TWI888383B - Liquid pharmaceutical formulations of pth conjugates - Google Patents

Abstract

A liquid pharmaceutical formulation, wherein the pharmaceutical formulation comprises a PTH conjugate, a buffering agent, an isotonicity agent, a preservative and optionally an antioxidant and wherein the PTH conjugate comprises a PTH moiety that is covalently and reversibly conjugated to a water-soluble carrier moiety.

Description

Liquid pharmaceutical preparations of PTH conjugates

The present invention relates to a liquid pharmaceutical preparation comprising a PTH conjugate, a buffer, an isotonic agent, a preservative and optionally an antioxidant.

Hypoparathyroidism is a rare endocrine disorder characterized by low serum calcium levels and low (insufficient) levels of parathyroid hormone in the blood circulation, most often occurring in adults after thyroid surgery. Patients receiving immunotherapy targeting immune checkpoint molecules such as cytotoxic T lymphocyte antigen 4 (CTLA-4), programmed cell death protein 1 (PD-1) and its ligand PD-L1 may also develop hypoparathyroidism as an immune-related adverse event of the parathyroid gland. Standard treatments for hypoparathyroidism include activated vitamin D analogs and calcium supplements, which increase calcium and phosphate absorption and serum levels at the expense of abnormally increased urinary calcium excretion.

PTH (parathyroid hormone) is an endocrine hormone that is secreted from the parathyroid glands in response to decreased calcium levels. In 2015, Natpara (PTH (1-84)) was approved for once-daily subcutaneous injection as an adjunct to vitamin D and calcium in patients with hypoparathyroidism. Although this represents an important advance in the treatment of this disease, Natpara has not demonstrated the ability to reduce the incidence of hypercalcemia (elevated serum calcium levels), hypocalcemia (low serum calcium levels), or hypercalcuria (elevated urinary calcium) in treated patients compared to conventional therapies.

When administered subcutaneously, PTH is degraded by proteases and is therefore rapidly absorbed and metabolized. Therefore, there is an urgent need for improved PTH-based therapies for hypoparathyroidism.

Conjugating PTH with PEG by PEGylation is one approach to improve biostability. WO2003/064462A1 discloses a pharmaceutical formulation comprising a stable conjugate of PTH(1-34) and Cys-PTH(1-35) with a PEG derivative. Although the application also suggests that the PTH variant and the water-soluble polymer can be linked via a hydrolyzable bond obtained by the reaction of the aldehyde-PEG and the amine group on PTH, no details are provided on how such conjugates are obtained or stored prior to administration.

Range extension of increasing the in vivo half-life of PTH based on conjugation to a water-soluble carrier moiety (such as PEG) via a reversible prodrug linker group was explored in WO2017/148883A1, WO2018/060310A1, WO2018/060311A1 and WO2018/060312A1. However, no information is provided on liquid pharmaceutical formulations that allow stable storage of these reversible conjugates.

Pharmaceutical formulations of such PTH conjugates, in which a water-soluble carrier is bound to PTH via a reversible linkage, must provide sufficient PTH conjugate stability to avoid premature release of PTH during storage. If the reversible linkage between the carrier and PTH is degraded during storage, the concentration of the fast-acting drug will increase. This may lead to the administration of supraphysiological levels of PTH, posing a risk of overdose, which may cause hypercalcemia or osteopenia. Therefore, achieving sustained release of PTH will maintain calcium homeostasis and normal turnover, which is important for the treatment of hypoparathyroidism.

Additionally, any drug released during the storage period will undergo rapid renal clearance when administered to a patient, thus reducing the time over which a long-acting formulation can provide therapeutically relevant amounts of drug.

In addition, it is known that PTH or its variants, conjugates or derivatives may undergo degradation reactions during storage, which may lead to the formation of impurities/peptide damage in the corresponding preparations, such as:

˙Degradation products produced by oxidation of the methionine (Met/M) residue to methionine sulfoxide and methionine sulfoxide;

˙Degradation products produced by oxidation of tryptophan (Trp/T) residues to hydroxyindole-3-alanine, 5-hydroxytryptophan or by dioxidation to N-methylkynurenine and kynurenine;

˙Degradation products resulting from the isomerization of aspartic acid (Asp/D) or aspartic acid residues to isoaspartic acid or isoaspartic acid salts (e.g. via a succinimide intermediate);

˙Degradation products resulting from the cleavage of peptide bonds at aspartic acid or aspartic acid residues, such as C-terminal Asp truncated peptides;

˙Degradation products resulting from the deamination of the asparagine residue (Asn/N) to aspartic acid or aspartate salts and/or isoaspartic acid or isoaspartate salts (e.g. via a succinimide intermediate).

˙Degradation products resulting from peptide bond cleavage at the asparagine residue, such as C-terminal Asn or Asp truncated peptides;

˙Degradation products resulting from the deamination of glutamine residues (Gln/Q) to glutamate or isoglutamate (e.g. via a glutarimide intermediate); and

˙Aggregates produced by the aggregation of peptides.

Since the above-mentioned degradation products or aggregates that may be formed during storage may impair the biological activity of the PTH moiety, it is desirable to minimize their formation during storage. Moreover, the reversible connection between the PTH moiety and the water-soluble carrier makes the storage of liquid pharmaceutical formulations containing the PTH conjugate challenging.

Therefore, it is important to identify suitable liquid pharmaceutical formulations of PTH conjugates comprising PTH covalently linked to a water-soluble carrier via a reversible linking group, wherein the peptide will exhibit an acceptable impurity profile and limited premature release of PTH even after prolonged storage.

Therefore, the purpose of the present invention is to at least partially overcome the above-mentioned disadvantages.

This object is achieved by a liquid pharmaceutical formulation, wherein the liquid pharmaceutical formulation comprises a PTH conjugate, a buffer, an isotonic agent, a preservative and optionally an antioxidant, and wherein the PTH conjugate comprises a PTH moiety covalently and reversibly conjugated to a water-soluble carrier moiety.

It was surprisingly found that the liquid pharmaceutical preparation of the present invention can be stably stored for a long period of time. In addition, it was surprisingly found that in the liquid pharmaceutical preparation of the present invention, the aggregation of PTH complexes in the liquid pharmaceutical preparation was reduced.

Within the meaning of the present invention, these terms are used as follows.

As used herein, the term "PTH" refers to all PTH polypeptides from mammalian species, such as from humans and mammalian species, especially from humans and murine species, as well as variants, analogs, orthologs, homologs, and derivatives and fragments thereof, which are characterized by increasing serum calcium and renal phosphorus excretion and decreasing serum phosphorus and renal calcium excretion in certain embodiments.

The term "PTH" also refers to all PTHrP polypeptides that bind and activate the common PTH/PTHrP1 receptor. In certain embodiments, the term "PTH" refers to PTH polypeptides and variants, homologs, and derivatives thereof that exhibit substantially the same biological activity (i.e., increasing serum calcium and renal phosphorus excretion, and decreasing serum phosphorus and renal calcium excretion).

As used herein, the term "PTH polypeptide variant" refers to a polypeptide from the same species that is different from a reference PTH or PTHrP polypeptide. In certain embodiments, such a reference is a PTH polypeptide sequence. Typically, the differences are limited so that the reference amino acid sequence and the variant are generally very similar and, in many regions, identical. In certain embodiments, the PTH polypeptide variant is at least 70%, 80%, 90%, or 95% identical to the reference PTH or PTHrP polypeptide. By having a polypeptide that is "identical" to a query amino acid sequence by at least, for example, 95%, it is contemplated that the amino acid sequence of the subject polypeptide is identical to the query sequence, except that the subject polypeptide sequence may contain up to five amino acid changes per 100 amino acids in the query amino acid sequence. These changes in the reference sequence may occur at the amino (N-terminal) or carboxyl (C-terminal) position of the reference amino acid sequence or anywhere between these terminal positions, either individually interspersed in the residue of the reference sequence or as one or more contiguous groups interspersed in the reference sequence. The query sequence may be the entire amino acid sequence of the reference sequence or any fragment specified as described herein.

Such a PTH polypeptide variant may be a naturally occurring variant, such as a naturally occurring allelic variant encoded by one of several alternative forms of PTH or PTHrP occupying a given locus on a chromosome or organism, or an isoform encoded by a natural splice variant derived from a single primary transcript. Alternatively, the PTH polypeptide variant may be a variant that is not known to occur naturally and may be prepared by mutagenesis techniques known in the art.

It is known in the art that one or more amino acids can be deleted from the N-terminus or C-terminus of a biologically active polypeptide without substantially losing biological function. The term PTH polypeptide variant also encompasses such N- and/or C-terminal deletions.

It is also recognized by those of ordinary skill in the art that some amino acid sequences of PTH or PTHrP polypeptides can be altered without significantly affecting the structure or function of the polypeptide. Such mutants include deletions, insertions, inversions, duplications, and substitutions selected according to general rules known in the art so as to have little effect on activity. For example, Bowie et al. (1990), Science 247: 1306-1310, which is incorporated herein by reference in its entirety, provide guidance on how to make phenotypically silent amino acid substitutions, in which the authors note that there are two main approaches to studying the tolerance of amino acid sequences to changes.

PTH variants may also be peptides in which any one or more (up to all) residues susceptible to deamination or deamination-like reactions (e.g., isomerization) are intentionally converted to other residues to any extent before storage by deamination or deamination-like reactions, with the conversion rate of each converted residue being up to 100%. PTH variants may also be peptides in which any one or more (up to all) residues susceptible to oxidation are intentionally converted to other residues to any extent before storage, with the conversion rate of each converted residue being up to 100%.

In certain embodiments, the term "PTH" refers to the following polypeptide sequence: SEQ ID NO: 1 (PTH 1-84): SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRKKEDNVLVESHEKSLGEADKADVNVLTKAKSQ

SEQ ID NO: 2 (PTH 1-83): SSVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRKKEDNVLVESHEKSLGEADKADVNVLTKAKS

SEQ ID NO: 3 (PTH 1-82): SSVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRKKEDNVLVESHEKSLGEADKADVNVLTKAK

SEQ ID NO: 4 (PTH 1-81): SSVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRKKEDNVLVESHEKSLGEADKADVNVLTKA

SEQ ID NO: 5 (PTH 1-80): SSVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRKKEDNVLVESHEKSLGEADKADVNVLTK

SEQ ID NO: 6(PTH 1-79):SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRKKEDNVLVESHEKSLGEADKADVNVLT

SEQ ID NO: 7(PTH 1-78):SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRKKEDNVLVESHEKSLGEADKADVNVL

SEQ ID NO: 8 (PTH 1-77): SSVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRKKEDNVLVESHEKSLGEADKADVNV

SEQ ID NO: 9 (PTH 1-76): SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRKKEDNVLVESHEKSLGEADKADVN

SEQ ID NO: 10 (PTH 1-75): SSVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRKKEDNVLVESHEKSLGEADKADV

SEQ ID NO: 11 (PTH 1-74): SSVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRKKEDNVLVESHEKSLGEADKAD

SEQ ID NO: 12 (PTH 1-73): SSVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRKKEDNVLVESHEKSLGEADKA

SEQ ID NO: 13 (PTH 1-72): SSVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRKKEDNVLVESHEKSLGEADK

SEQ ID NO: 14 (PTH 1-71): SSVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRKKEDNVLVESHEKSLGEAD

SEQ ID NO: 15 (PTH 1-70): SSVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRKKEDNVLVESHEKSLGEA

SEQ ID NO: 16 (PTH 1-69): SSVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRKKEDNVLVESHEKSLGE

SEQ ID NO: 17(PTH 1-68): SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRKKEDNVLVESHEKSLG

SEQ ID NO: 18 (PTH 1-67): SSVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRKKEDNVLVESHEKSL

SEQ ID NO: 19 (PTH 1-66): SSVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRKKEDNVLVESHEKS

SEQ ID NO: 20 (PTH 1-65): SSVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRKKEDNVLVESHEK

SEQ ID NO: 21 (PTH 1-64): SSVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRKKEDNVLVESHE

SEQ ID NO: 22 (PTH 1-63): SSVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRKKEDNVLVESH

SEQ ID NO: 23(PTH 1-62):SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRKKEDNVLVES

SEQ ID NO: 24(PTH 1-61):SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRKKEDNVLVE

SEQ ID NO: 25(PTH 1-60): SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRKKEDNVLV

SEQ ID NO: 26 (PTH 1-59): SSVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRKKEDNVL

SEQ ID NO: 27(PTH 1-58):SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRKKEDNV

SEQ ID NO: 28(PTH 1-57):SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRKKEDN

SEQ ID NO: 29 (PTH 1-56): SSVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRKKED

SEQ ID NO: 30 (PTH 1-55): SSVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRKKE

SEQ ID NO: 31 (PTH 1-54): SSVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRKK

SEQ ID NO: 32 (PTH 1-53): SSVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRK

SEQ ID NO: 33 (PTH 1-52): SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPR

SEQ ID NO: 34 (PTH 1-51): SSVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRP

SEQ ID NO: 35 (PTH 1-50): SSVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQR

SEQ ID NO: 36 (PTH 1-49): SSVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQ

SEQ ID NO: 37 (PTH 1-48): SSVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGS

SEQ ID NO: 38 (PTH 1-47): SSVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAG

SEQ ID NO: 39 (PTH 1-46): SSVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDA

SEQ ID NO: 40 (PTH 1-45): SSVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRD

SEQ ID NO: 41 (PTH 1-44): SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPR

SEQ ID NO: 42 (PTH 1-43): SSVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAP

SEQ ID NO: 43 (PTH 1-42): SSVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLA

SEQ ID NO: 44 (PTH 1-41): SSVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPL

SEQ ID NO: 45 (PTH 1-40): SSVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAP

SEQ ID NO: 46 (PTH 1-39): SSVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGA

SEQ ID NO: 47 (PTH 1-38): SSVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALG

SEQ ID NO: 48 (PTH 1-37): SSVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVAL

SEQ ID NO: 49 (PTH 1-36): SSVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVA

SEQ ID NO: 50 (PTH 1-35): SSVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFV

SEQ ID NO: 51 (PTH 1-34): SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNF

SEQ ID NO: 52 (PTH 1-33): SSVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHN

SEQ ID NO: 53(PTH 1-32):SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVH

SEQ ID NO: 54 (PTH 1-31): SSVSEIQLMHNLGKHLNSMERVEWLRKKLQDV

SEQ ID NO: 55(PTH 1-30):SVSEIQLMHNLGKHLNSMERVEWLRKKLQD

SEQ ID NO: 56(PTH 1-29):SVSEIQLMHNLGKHLNSMERVEWLRKKLQ

SEQ ID NO: 57(PTH 1-28):SVSEIQLMHNLGKHLNSMERVEWLRKKL

SEQ ID NO: 58(PTH 1-27):SVSEIQLMHNLGKHLNSMERVEWLRKK

SEQ ID NO: 59 (PTH 1-26): SSVSEIQLMHNLGKHLNSMERVEWLRK

SEQ ID NO: 60(PTH 1-25):SVSEIQLMHNLGKHLNSMERVEWLR

SEQ ID NO: 61 (amidated PTH 1-84): SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRKKEDNVLVESHEKSLGEADKADVNVLTKAKSQ; wherein the C-terminus is amidated

SEQ ID NO: 62 (amidated PTH 1-83): SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRKKEDNVLVESHEKSLGEADKADVNVLTKAKS; wherein the C-terminus is amidated

SEQ ID NO: 63 (amidated PTH 1-82): SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRKKEDNVLVESHEKSLGEADKADVNVLTKAK; wherein the C-terminus is amidated

SEQ ID NO: 64 (amidated PTH 1-81): SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRKKEDNVLVESHEKSLGEADKADVNVLTKA; wherein the C-terminus is amidated

SEQ ID NO: 65 (amidated PTH 1-80): SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRKKEDNVLVESHEKSLGEADKADVNVLTK; wherein the C-terminus is amidated

SEQ ID NO: 66 (amidated PTH 1-79): SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRKKEDNVLVESHEKSLGEADKADVNVLT; wherein the C-terminus is amidated

SEQ ID NO: 67 (amidated PTH 1-78): SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRKKEDNVLVESHEKSLGEADKADVNVL; wherein the C-terminus is amidated

SEQ ID NO: 68 (amidated PTH 1-77): SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRKKEDNVLVESHEKSLGEADKADVNV; wherein the C-terminus is amidated

SEQ ID NO: 69 (amidated PTH 1-76): SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRKKEDNVLVESHEKSLGEADKADVN; wherein the C-terminus is amidated

SEQ ID NO: 70 (amidated PTH 1-75): SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRKKEDNVLVESHEKSLGEADKADV; wherein the C-terminus is amidated

SEQ ID NO: 71 (amidated PTH 1-74): SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRKKEDNVLVESHEKSLGEADKAD; wherein the C-terminus is amidated

SEQ ID NO: 72 (amidated PTH 1-73): SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRKKEDNVLVESHEKSLGEADKA; wherein the C-terminus is amidated

SEQ ID NO: 73 (amidated PTH 1-72): SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRKKEDNVLVESHEKSLGEADK; wherein the C-terminus is amidated

SEQ ID NO: 74 (amidated PTH 1-71): SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRKKEDNVLVESHEKSLGEAD; wherein the C-terminus is amidated

SEQ ID NO: 75 (amidated PTH 1-70): SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRKKEDNVLVESHEKSLGEA; wherein the C-terminus is amidated

SEQ ID NO: 76 (amidated PTH 1-69): SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRKKEDNVLVESHEKSLGE; wherein the C-terminus is amidated

SEQ ID NO: 77 (amidated PTH 1-68): SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRKKEDNVLVESHEKSLG; wherein the C-terminus is amidated

SEQ ID NO: 78 (amidated PTH 1-67): SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRKKEDNVLVESHEKSL; wherein the C-terminus is amidated

SEQ ID NO: 79 (amidated PTH 1-66): SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRKKEDNVLVESHEKS; wherein the C-terminus is amidated

SEQ ID NO: 80 (amidated PTH 1-65): SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRKKEDNVLVESHEK; wherein the C-terminus is amidated

SEQ ID NO: 81 (amidated PTH 1-64): SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRKKEDNVLVESHE; wherein the C-terminus is amidated

SEQ ID NO: 82 (amidated PTH 1-63): SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRKKEDNVLVESH; wherein the C-terminus is amidated

SEQ ID NO: 83 (amidated PTH 1-62): SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRKKEDNVLVES; wherein the C-terminus is amidated

SEQ ID NO: 84 (amidated PTH 1-61): SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRKKEDNVLVE; wherein the C-terminus is amidated

SEQ ID NO: 85 (amidated PTH 1-60): SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRKKEDNVLV; wherein the C-terminus is amidated

SEQ ID NO: 86 (amidated PTH 1-59): SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRKKEDNVL; wherein the C-terminus is amidated

SEQ ID NO: 87 (amidated PTH 1-58): SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRKKEDNV; wherein the C-terminus is amidated

SEQ ID NO: 88 (amidated PTH 1-57): SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRKKEDN; wherein the C-terminus is amidated

SEQ ID NO: 89 (amidated PTH 1-56): SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRKKE; wherein the C-terminus is amidated

SEQ ID NO: 90 (amidated PTH 1-55): SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRKKE; wherein the C-terminus is amidated

SEQ ID NO: 91 (amidated PTH 1-54): SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRKK; wherein the C-terminus is amidated

SEQ ID NO: 92 (amidated PTH 1-53): SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPRK; wherein the C-terminus is amidated

SEQ ID NO: 93 (amidated PTH 1-52): SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRPR; wherein the C-terminus is amidated

SEQ ID NO: 94 (amidated PTH 1-51): SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQRP; wherein the C-terminus is amidated

SEQ ID NO: 95 (amidated PTH 1-50): SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQR; wherein the C-terminus is amidated

SEQ ID NO: 96 (amidated PTH 1-49): SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGSQ; wherein the C-terminus is amidated

SEQ ID NO: 97 (amidated PTH 1-48): SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAGS; wherein the C-terminus is amidated

SEQ ID NO: 98 (amidated PTH 1-47): SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDAG; wherein the C-terminus is amidated

SEQ ID NO: 99 (amidated PTH 1-46): SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRDA; wherein the C-terminus is amidated

SEQ ID NO: 100 (amidated PTH 1-45): SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPRD; wherein the C-terminus is amidated

SEQ ID NO: 101 (amidated PTH 1-44): SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAPR; wherein the C-terminus is amidated

SEQ ID NO: 102 (amidated PTH 1-43): SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLAP; wherein the C-terminus is amidated

SEQ ID NO: 103 (amidated PTH 1-42): SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPLA; wherein the C-terminus is amidated

SEQ ID NO: 104 (amidated PTH 1-41): SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAPL; wherein the C-terminus is amidated

SEQ ID NO: 105 (amidated PTH 1-40): SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGAP; wherein the C-terminus is amidated

SEQ ID NO: 106 (amidated PTH 1-39): SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALGA; wherein the C-terminus is amidated

SEQ ID NO: 107 (amidated PTH 1-38): SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVALG; wherein the C-terminus is amidated

SEQ ID NO: 108 (amidated PTH 1-37): SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVAL; wherein the C-terminus is amidated

SEQ ID NO: 109 (amidated PTH 1-36): SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFVA; wherein the C-terminus is amidated

SEQ ID NO: 110 (amidated PTH 1-35): SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNFV; wherein the C-terminus is amidated

SEQ ID NO: 111 (amidated PTH 1-34): SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHNF; wherein the C-terminus is amidated

SEQ ID NO: 112 (amidated PTH 1-33): SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVHN; wherein the C-terminus is amidated

SEQ ID NO: 113 (amidated PTH 1-32): SVSEIQLMHNLGKHLNSMERVEWLRKKLQDVH; wherein the C-terminus is amidated

SEQ ID NO: 114 (amidated PTH 1-31): SVSEIQLMHNLGKHLNSMERVEWLRKKLQDV; wherein the C-terminus is amidated

SEQ ID NO: 115 (amidated PTH 1-30): SVSEIQLMHNLGKHLNSMERVEWLRKKLQD; wherein the C-terminus is amidated

SEQ ID NO: 116 (amidated PTH 1-29): SVSEIQLMHNLGKHLNSMERVEWLRKKLQ; wherein the C-terminus is amidated

SEQ ID NO: 117 (amidated PTH 1-28): SVSEIQLMHNLGKHLNSMERVEWLRKKL; wherein the C-terminus is amidated

SEQ ID NO: 118 (amidated PTH 1-27): SVSEIQLMHNLGKHLNSMERVEWLRKK; wherein the C-terminus is amidated

SEQ ID NO: 119 (amidated PTH 1-26): SVSEIQLMHNLGKHLNSMERVEWLRK; wherein the C-terminus is amidated

SEQ ID NO: 120 (amidated PTH 1-25): SVSEIQLMHNLGKHLNSMERVEWLR; wherein the C-terminus is amidated

SEQ ID NO: 121 (PTHrP):

The term PTH polypeptide also encompasses all PTH and PTHrP polypeptides encoded by PTH and PTHrP analogs, orthologs and/or species homologs. It is also recognized by those of ordinary skill in the art that PTHrP and PTHrP analogs bind to activate a common PTH/PTHrP1 receptor, and thus the term PTH polypeptide also encompasses all PTHrP analogs.

As used herein, the term "PTH analogs" refers to PTH and PTHrP of different and unrelated organisms, which perform the same function in each organism, but it does not originate from a common ancestral structure of the ancestor of the organism. Instead, similar PTH and PTHrP appeared separately and then evolved to perform the same or similar functions. In other words, similar PTH and PTHrP polypeptides are polypeptides with completely different amino acid sequences, but perform the same biological activity, namely, increasing serum calcium and renal phosphorus excretion, and reducing serum phosphorus and renal calcium excretion.

As used herein, the term "PTH orthologs" refers to PTH and PTHrP within two different species whose sequences are related to each other through a common homologous PTH or PTHrP in an ancestral species, but have evolved to diverge from each other.

As used herein, the term "PTH homologs" refers to PTH and PTHrP from different organisms that perform the same function in each organism and are derived from an ancestral structure common to the ancestors of the organisms. In other words, homologous PTH polypeptides are polypeptides with very similar amino acid sequences that perform the same biological activity, namely, increasing serum calcium and renal phosphorus excretion, and decreasing serum phosphorus and renal calcium excretion. In certain embodiments, a PTH polypeptide homolog may be defined as a polypeptide that exhibits at least 40%, 50%, 60%, 70%, 80%, 90%, or 95% identity to a reference PTH or PTHrP polypeptide.

Thus, the PTH polypeptide may be, for example: (i) wherein at least one amino acid residue is substituted with a conservative or non-conservative amino acid residue, in certain embodiments, a conservative amino acid residue, and such substituted amino acid residue may or may not be a residue encoded by the genetic code; and/or (ii) wherein at least one amino acid residue includes a substituent; and/or (iii) wherein the PTH polypeptide is fused to another compound, such as a compound that increases the half-life of the polypeptide (e.g., polyethylene glycol); and/or (iv) wherein additional amino acids are fused to the PTH polypeptide, such as an IgG Fc fusion region polypeptide or a leader sequence or secretory sequence, or a sequence for purifying the above forms of polypeptides or preprotein sequences.

As used herein, the term "PTH polypeptide fragment" refers to any polypeptide comprising a contiguous span of a portion of the amino acid sequence of a PTH or PTHrP polypeptide.

More specifically, a PTH polypeptide fragment comprises at least 6, such as at least 8, at least 10, or at least 17 consecutive amino acids of a PTH or PTHrP polypeptide. A PTH polypeptide fragment may be further described as a subgenus of a PTH or PTHrP polypeptide comprising at least 6 amino acids, wherein "at least 6" is defined as any integer between 6 and the integer representing the C-terminal amino acid of the PTH or PTHrP polypeptide. Also included are substances of PTH or PTHrP polypeptide fragments, as described above, having a length of at least 6 amino acids and further specified with respect to their N-terminal and C-terminal positions.

The term "PTH polypeptide fragment" also includes all PTH or PTHrP polypeptide fragments as separate substances, which are at least 6 amino acids in length and can be specifically specified by the N-terminal and C-terminal positions as described above. That is, on any given amino acid sequence of a PTH or PTHrP polypeptide, a fragment of at least 6 consecutive amino acid residues in length can occupy every combination of N-terminal and C-terminal positions.

The term "PTH" also includes polyamino acid conjugates having a sequence as described above, but having a backbone containing amide and non-amide bonds (such as ester bonds), such as depeptides. Depeptides are chains of amino acid residues, wherein the backbone contains both amide (peptide) and ester bonds. Thus, the term "side chain" as used herein refers to a portion attached to the α-carbon of the amino acid moiety, if the amino acid moiety is attached via an amine bond, as in a polypeptide, or to any carbon-containing portion of the backbone of the polyamino acid conjugate, such as in a depeptide.

In certain embodiments, the term "PTH" refers to a polypeptide having a backbone formed by amide (peptide) bonds.

Since the term PTH includes the above-mentioned variants, analogs, orthologs, homologs, derivatives and fragments of PTH and PTHrP, all references to specific positions within the reference sequence also include equivalent positions in variants, analogs, orthologs, homologs, derivatives and fragments of PTH or PTHrP portions, even if not specifically mentioned.

As used herein, the phrase "PTH conjugate, wherein the PTH portion" followed by the amount in mg/ml means that the liquid preparation contains the PTH conjugate, but for the corresponding amount, only the PTH portion is taken into account instead of the complete PTH conjugate, that is, the portion of the PTH conjugate other than the PTH portion, such as the water-soluble carrier portion, is not taken into account. The amount of the PTH portion in the PTH conjugate can be determined by quantitative amino acid analysis after the PTH conjugate is completely hydrolyzed under acidic conditions, or by any known analytical method that can quantify an unknown sample by comparison with a PTH conjugate with a known content of the PTH portion.

As used herein, the term "about" in combination with a numerical value is used to indicate a range from and including that numerical value plus and minus no more than 10% of the numerical value, in certain embodiments, no more than 8% of the numerical value, in certain embodiments, no more than 5% of the numerical value, and in certain embodiments, no more than 2% of the numerical value. For example, the phrase "about 200" is used to indicate a range from and including 200 +/- 10%, i.e., a range of 180-220. In certain embodiments, 200 +/- 8%, i.e., 184 to 216; in certain embodiments, 200 +/- 5%, i.e., 190-210. And in certain embodiments, 200 +/- 2%, i.e., 196-204. It should also be understood that a percentage stated as "about 20%" does not mean "20% +/- 10%", i.e., ranging from and including 10% to 30%, but "about 20%" means ranging from and including 18% to 22%, i.e., plus or minus 10% of the numerical value of 20.

As used herein, the term "antimicrobial agent" refers to a chemical substance that kills or inhibits the growth of microorganisms such as bacteria, fungi, yeast, protozoa and/or destroys viruses.

As used herein, the term "antiadsorbent" refers primarily to ionic or nonionic surfactant proteins or soluble polymers that are used to competitively coat or adsorb to the inner surface of a container containing a dosage form. The concentration and type of surfactant chosen depends on the effects to be avoided, but typically a monolayer of surfactant is formed at the interface just above the critical micelle concentration (CMC) value.

As used herein, the term "buffer" or "buffer" refers to a compound that maintains the pH in the desired range. Physiologically tolerated buffers are, for example, sodium phosphate, succinate, histidine, bicarbonate, citrate and acetate, sulfate, nitrate, chloride, pyruvate. Antacids such as Mg(OH) ₂ or ZnCO ₃ may also be used.

As used herein, the term "C _1-4 alkyl" used alone or in combination refers to a straight or branched chain alkyl moiety having 1 to 4 carbon atoms. If present at the end of a molecule, examples of straight or branched chain C _1-4 alkyl are methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl and tert-butyl. When two parts of the molecule are connected through a C _1-4 alkyl group, then examples of such C _1-4 alkyl groups are -CH ₂ -, -CH ₂ -CH ₂ -, -CH(CH ₃ )-, -CH ₂ -CH ₂ -CH ₂ -, -CH(C ₂ H ₅ )- and -C(CH ₃ ) ₂ -. Each hydrogen of a C _1-4 alkyl carbon may be optionally substituted with a substituent as described above. Optionally, the C _1-4 alkyl group may be interrupted by one or more moieties as defined below.

As used herein, the term "C _1-6 alkyl" used alone or in combination refers to a straight or branched chain alkyl moiety having 1 to 6 carbon atoms. Examples of straight and branched chain C _1-6 alkyl groups are methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, tert-butyl, n-pentyl, 2-methylbutyl, 2,2-dimethylpropyl, n-hexyl, 2-methylpentyl, 3-methylpentyl, 2,2-dimethylbutyl, 2,3-dimethylbutyl and 3,3-dimethylpropyl if present at the molecular terminal. When the two parts of the molecule are linked through a C _1-6 alkyl group, then examples of such C _1-6 alkyl groups are -CH ₂ -, -CH ₂ -CH ₂ -, -CH(CH ₃ )-, -CH ₂ -CH ₂ -CH ₂ -, -CH(C ₂ H ₅ )-, and -C(CH ₃ ) ₂ -. Each hydrogen atom of the C _1-6 carbon may be optionally substituted with a substituent as described above. Optionally, the C _1-6 alkyl group may be interrupted by one or more moieties as defined below.

Thus, "C _1-10 alkyl", "C _1-20 alkyl" or "C _1-50 alkyl" refers to an alkyl chain having 1 to 10, 1 to 20, or 1 to 50 carbon atoms, respectively, wherein each hydrogen atom of the C _1-10 , C _1-20 or C _1-50 carbon may be optionally replaced by a substituent as defined above. Optionally, the C _1-10 , C _1-20 or C _1-50 alkyl may be interrupted by one or more moieties as defined below.

As used herein, the term " _C2-6 alkenyl" used alone or in combination refers to a straight or branched hydrocarbon moiety containing at least one carbon-carbon double bond and having 2 to 6 carbon atoms. If present at the end of the molecule, examples are -CH= _CH2 , -CH=CH- _CH3 , _-CH2- CH= _CH2 , -CH= _CHCH2 - _CH3 and -CH=CH-CH= _CH2 . When the two parts of the molecule are connected through the _C2-6 alkenyl group, then an example of such a _C2-6 alkenyl group is -CH=CH-. Each hydrogen atom of _{the C2-6} alkenyl moiety may be optionally replaced by a substituent as defined above. Optionally, the _C2-6 alkenyl group may be interrupted by one or more moieties as defined below.

Thus, the term " _C2-10 alkenyl", " _C2-20 alkenyl" or " _C2-50 alkenyl", used alone or in combination, refers to a straight or branched chain hydrocarbon moiety containing at least one carbon-carbon double bond and having 2 to 10, 2 to 20, or 2 to 50 carbon atoms. Each hydrogen atom of the _C2-10 alkenyl, _C2-20 alkenyl or _C2-50 alkenyl may be optionally replaced by a substituent as defined above. Optionally, the _C2-10 alkenyl, _C2-20 alkenyl or _C2-50 alkenyl may be interrupted by one or more moieties as defined below.

As used herein, the term "C _2-6 alkynyl", used alone or in combination, refers to a straight or branched hydrocarbon moiety containing at least one carbon-carbon triple bond and having 2 to 6 carbon atoms. If present at the end of the molecule, examples are -C≡CH, -CH ₂ -C≡CH, CH ₂ -CH ₂ -C≡CH and CH ₂ -C≡C-CH _3. When the two parts of the molecule are connected through the alkynyl group, then an example is -C≡C-. Each hydrogen atom of the _{C 2-6} alkynyl group may be optionally replaced by a substituent as defined above. Optionally, one or more double bonds may occur. Optionally, the C _2-6 alkynyl group may be interrupted by one or more moieties as defined below.

Thus, as used herein, the terms " _C2-10 alkynyl", " _C2-20 alkynyl" and " _C2-50 alkynyl", used alone or in combination, refer to a straight or branched chain hydrocarbon moiety comprising at least one carbon-carbon triple bond and having 2 to 10, 2 to 20, or 2 to 50 carbon atoms, respectively. Each hydrogen atom of the _C2-10 alkynyl, _C2-20 alkynyl or _C2-50 alkynyl may be optionally replaced by a substituent as defined above. Optionally, one or more double bonds may occur. Optionally, the _C2-10 alkynyl, _C2-20 alkynyl or _C2-50 alkynyl may be interrupted by one or more moieties as defined below.

As described above, _C1-4 alkyl, _C1-6 alkyl, _C1-10 alkyl, _C1-20 alkyl, _C1-50 alkyl, _C2-6 alkenyl, _C2-10 alkenyl, _C2-20 alkenyl, _C2-50 alkenyl, _C2-6 alkynyl, C2-10 alkynyl, _C2-20 alkynyl or _{C2-50 alkynyl} may in certain embodiments be optionally interrupted by one or more moieties selected from:

wherein the dashed line indicates attachment to the remainder of the moiety or reagent; and -R and -Ra ^are each independently selected from: -H, methyl, ethyl, propyl, butyl, pentyl, and hexyl.

As used herein, the term "C _3-10 cycloalkyl" refers to a cyclic alkyl chain having 3 to 10 carbon atoms, which may be saturated or unsaturated, for example, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cyclohexenyl, cycloheptyl, cyclooctyl, cyclononyl or cyclodecyl. Each hydrogen atom of the C _3-10 cycloalkyl carbon may be substituted with a substituent as described above. The term "C _3-10 cycloalkyl" also includes bridged bicyclics, such as norbornane or norbornene.

As used herein, the term "8- to 30-membered carbon polycyclic group" or "8- to 30-membered carbon polycyclic ring" refers to a cyclic moiety of two or more rings having 8 to 30 ring atoms, wherein two adjacent rings share at least one ring atom and may contain up to the maximum number of double bonds (fully, partially or unsaturated aromatic or non-aromatic rings). In certain embodiments, the 8- to 30-membered carbon polycyclic group refers to a cyclic moiety of two, three, four or five rings, and in certain embodiments, a cyclic moiety of two, three or four rings.

As used herein, the term "3- to 10-membered heterocyclic group" or "3- to 10-membered heterocycle" refers to a ring having 3, 4, 5, 6, 7, 8, 9 or 10 ring atoms, which may contain up to the maximum number of double bonds (fully, partially or unsaturated aromatic or non-aromatic rings), wherein at least one and up to 4 ring atoms are substituted by heteroatoms selected from sulfur (including -S(O)-, -S(O) _2- ), oxygen and nitrogen (including =N(O)-), and wherein the ring is linked to the rest of the molecule via a carbon or nitrogen atom. Examples of 3- to 10-membered heterocyclic rings include, but are not limited to, aziridine, ethylene oxide, ethylene sulfide, azide-doped acrylidine, ethylene oxide, thiodide-doped acrylcyclobutane, oxide-doped acrylcyclobutane, thiodide-doped acrylcyclobutane, furan, thiophene, pyrrole, pyrroline, imidazole, imidazoline, pyrazole, pyrazoline, oxazole, oxazoline, isoxazole, isoxazoline, thiazole, thiazoline, isothiazole, isothiazolinone. The invention also includes oxadiazole, thiadiazoline, tetrahydrofuran, tetrahydrothiophene, pyrrolidine, imidazolidine, pyrazolidine, oxazolidine, isoxazolidine, thiazolidine, isothiazolidine, thiadiazolidine, cyclobutane, pyran, dihydropyran, tetrahydropyran, imidazolidine, pyridine, oxazine, pyrazine, pyrimidine, piperazine, piperidine, morpholine, tetrazole, triazole, triazolidine, tetrazolidine, diazepine, azopine and homopiperazine. Each hydrogen atom of the 3- to 10-membered heterocyclic group or the 3- to 10-membered heterocyclic group may be substituted with a substituent as defined below.

As used herein, the term "8- to 11-membered heterobicyclic group" or "8- to 11-membered heterobicycle" refers to a heterocyclic moiety of two rings having 8 to 11 ring atoms, wherein at least one ring atom is shared by both rings and may contain up to the maximum number of double bonds (fully, partially or unsaturated aromatic or non-aromatic rings), wherein at least one and up to 6 ring atoms are substituted by heteroatoms selected from sulfur (including -S(O)-, -S(O) _2- ), oxygen and nitrogen (including =N(O)-), and wherein the ring is linked to the rest of the molecule via a carbon or nitrogen atom. Examples of 8- to 11-membered heterocycles are indole, dihydroindole, benzofuran, benzothiophene, benzoxazole, benzoisoxazole, benzothiazole, benzoisothiazole, benzimidazole, benzimidazolin, quinoline, quinazoline, dihydroquinazoline, quinoline, dihydroquinoline, tetrahydroquinoline, decahydroquinoline, isoquinoline, decahydroisoquinoline, tetrahydroisoquinoline, dihydroisoquinoline, benzazepine, purine and pteridine. The term 8- to 11-membered heterocyclic ring also includes a spirocyclic structure of two rings (such as 1,4-dioxa-8-azaspiro[4.5]decane) or a bridged heterocyclic ring (such as 8-azabicyclo[3.2.1]octane). Each hydrogen atom of the 8- to 11-membered heterobicyclic ring or the 8- to 11-membered heterobicyclic ring carbon may be substituted with a substituent as defined below.

Similarly, the term "8- to 30-membered heteropolycyclic group" or "8- to 30-membered heteropolycyclic ring" refers to a heterocyclic moiety of two or more rings having 8-30 ring atoms (in certain embodiments, three, four or five rings), wherein two adjacent rings share at least one ring atom and may contain up to the maximum number of double bonds (fully, partially or unsaturated aromatic or non-aromatic rings), wherein at least one and up to 10 ring atoms are substituted by heteroatoms selected from sulfur (including -S(O)-, -S(O) _2- ), oxygen and nitrogen (including =N(O)-), and wherein the ring is linked to the rest of the molecule through a carbon or nitrogen atom.

是指R^x和R^y形成以下結構：

It should be understood that the phrase "the R ^x /R ^y pair is linked together with the atoms to which they are attached to form a C _3-10 cycloalkyl or 3 to 10 membered heterocyclic group" refers to the following structures:

It means that R ^x and R ^y form the following structure:

wherein R is a C _3-10 cycloalkyl group or a 3-10 membered heterocyclic group.

It is also to be understood that the phrase "the R ^x /R ^y pair is linked together with the atoms to which they are attached to form Ring A" refers to the following structure:

It means that R ^x and R ^y form the following structure:

As used herein, the term "drug" as used herein refers to a substance used to treat, cure, prevent or diagnose a disease or to enhance physical or mental health. If a drug (such as PTH) is conjugated to another moiety, the portion of the resulting product derived from PTH is referred to as the "PTH moiety."

As used herein, the term "excipient" refers to a compound administered with a therapeutic agent, such as a buffer, an isotonic modifier, a preservative, a stabilizer, an anti-adsorption agent, an oxidation protectant or other adjuvant. However, in some cases, an excipient may have a dual or triple function. The term "excipient" may also refer to a diluent, adjuvant or vehicle administered with a therapeutic agent such as a drug or a drug conjugate. Such drug excipients may be sterile liquids such as water and oils (including those of petroleum, animal, plant or synthetic origin), including but not limited to: peanut oil, soybean oil, mineral oil, sesame oil, etc. When the drug formulation is administered orally, water is a preferred excipient. When the drug formulation is administered intravenously, salt and glucose aqueous solutions are preferred excipients. In certain embodiments, salt solutions and glucose aqueous solutions and glycerol solutions are used as liquid excipients for injectable solutions. Suitable drug excipients include: starch, glucose, lactose, sucrose, mannitol, trehalose, gelatin, malt, rice, flour, chalk, silica gel, sodium stearate, glyceryl monostearate, talc, sodium chloride, skimmed milk powder, glycerol, propylene, ethylene glycol, water, ethanol, etc. If necessary, the liquid pharmaceutical preparation may also contain a small amount of a wetting agent or emulsifier, a pH buffer, such as acetate, succinate, Tris (tris (hydroxymethyl) aminomethane), carbonate, phosphate, HEPES (4- (2-hydroxyethyl) -1- piperazineethanesulfonic acid), MES (2- (N-morpholino) ethanesulfonic acid), or may contain a detergent such as ^Tween® , poloxamer, poloxamine, CHAPS, ^Igepal® or an amino acid, such as glycine, lysine or histidine. These pharmaceutical preparations can be in the form of solutions, suspensions, emulsions, tablets, pills, capsules, powders, sustained-release preparations, and the like. The drug formulation can be formulated into a suppository with conventional binders and excipients such as triglycerides. Oral formulations may include standard excipients such as pharmaceutical grade mannitol, lactose, starch, magnesium stearate, sodium saccharin, cellulose, magnesium carbonate, etc. Such formulations will contain a therapeutically effective amount of the drug or drug portion and an appropriate amount of excipients to provide the patient with an appropriate administration form. The formulation should be suitable for the mode of administration.

As used herein, the term "formulation", "pharmaceutical formulation", "mixture" or "composition" refers to a formulation containing one or more active ingredients and one or more excipients, and any product directly or indirectly resulting from the combination, fusion or aggregation of any two or more of the ingredients in the formulation, or from the decomposition of one or more of the ingredients, or from other types of reactions or interactions of one or more of the ingredients. Therefore, the liquid pharmaceutical formulation of the present invention includes any formulation or composition prepared by mixing one or more PTH conjugates with pharmaceutically acceptable excipients such as buffers, isotonic agents, preservatives and optional antioxidants.

As used herein, the term "free form" of a drug refers to the unmodified, fully pharmacologically active form of the drug (e.g., after release from a conjugate).

As used herein, the term "functional group" refers to an atomic group that can react with other atomic groups. Functional groups include, but are not limited to, the following: carboxylic acid (-(C=O)OH), primary or secondary amine ( _-NH2 , -NH-), maleimide, thiol (-SH), sulfonic acid (-(O=S=O)OH), carbonate, carbamate (-O(C=O)N<), hydroxyl (-OH), aldehyde (-(C=O)H), ketone (-(C=O)-), hydrazine (>NN<), isocyanate, isothiocyanate, phosphoric acid (-O(P=O)OHOH), phosphonic acid (-O(P=O)OHH), halogenated acetyl, alkyl halide, acryl, aryl fluoride, hydroxyl amine, disulfide, sulfonamide, sulfuric acid, vinyl sulfide, vinyl ketone, diazoalkane, oxirane and aziridine.

As used herein, the term "halogen" refers to fluorine, chlorine, bromine or iodine. Generally, the preferred halogen is fluorine or chlorine.

As used herein, the term "interrupted" means that a moiety is inserted between two carbon atoms, or (if the insertion is at one of the ends of the moiety) between a carbon atom or a heteroatom and a hydrogen atom, and in certain embodiments, between a carbon atom and a heteroatom.

As used herein, the term "immune checkpoint inhibitor" refers to a compound that interferes with the function of a ligand or inhibits the binding of a ligand that is induced by a receptor expressed on the cell membrane, which inhibits inflammatory immune cell function after receptor activation. Such compounds can be, for example, biological agents such as antibodies, nanobodies, probodies, anticalins, or cyclic peptides or small molecule inhibitors.

As used herein, the term "isosmolar agent" refers to a chemical substance that minimizes pain, irritation, and tissue damage caused by cell damage due to osmotic pressure differences at the injection depot.

As used herein, the term "liquid pharmaceutical preparation" refers to a mixture comprising a water-soluble PTH conjugate and one or more solvents (such as water).

As used herein, the term "dried pharmaceutical preparation" or "dry pharmaceutical preparation" refers to providing the pharmaceutical preparation in a dry form. Suitable drying methods are spray drying and freeze drying, i.e., freeze drying. Such dry preparations containing PTH complexes have a maximum residual water content of 10% as determined using the Karl Fischer method; in some embodiments, less than 5%; in some embodiments, less than 2%. In some embodiments, the dry pharmaceutical preparation is dried by freeze drying.

As used herein, the term "moiety" refers to a portion of a molecule that lacks one or more atoms compared to the corresponding reagent. For example, if a reagent of the formula "H-X-H" reacts with another reagent and becomes part of the reaction product, the structure of the corresponding moiety of the reaction product is "H-X-" or "-X-", and each "-" represents a connection to another moiety. Thus, the drug moiety (such as the PTH moiety) is released from the conjugate as a drug (such as PTH).

可以連接到兩個部分，或可以作為

或作為

間斷一個部分。 It should be understood that if a sequence or chemical structure of a group of atoms is provided that connects two moieties or interrupts a moiety, the sequence or chemical structure can be connected to the two moieties in either direction unless otherwise expressly stated. For example, the moiety "-C(O)N(R ¹ )-" can be connected to two moieties or interrupt a moiety as "-C(O)N(R ¹ )-" or "-N(R ¹ )C(O)-". Similarly, the moiety

Can be connected to two parts, or can be used as

or as

Break off a section.

If the PTH moiety contains one or more acidic or alkaline groups, the liquid pharmaceutical preparation also contains its corresponding pharmaceutically or toxicologically acceptable salt, in particular its pharmaceutically utilizable salt. Thus, the PTH moiety containing one or more acidic groups can be present and used, for example, in the form of an alkali metal salt, an alkaline earth metal salt or an ammonium salt. More precise examples of such salts include sodium salts, potassium salts, calcium salts, magnesium salts or salts with ammonia or organic amines, such as ethylamine, ethanolamine, triethanolamine or amino acids, as well as other salts or amines known to those skilled in the art. PTH moieties containing one or more basic groups (i.e., groups that can be protonated) may be present and may be used in the form of addition salts thereof with inorganic or organic acids. Examples of suitable acids include: hydrogen chloride, hydrogen bromide, phosphoric acid, sulfuric acid, nitric acid, methanesulfonic acid, p-toluenesulfonic acid, naphthalene disulfonic acid, oxalic acid, acetic acid, tartaric acid, lactic acid, salicylic acid, benzoic acid, formic acid, propionic acid, pivalic acid, diethylacetic acid, malonic acid, succinic acid, pimelic acid, fumaric acid, maleic acid, apple acid, aminosulfonic acid, phenylpropionic acid, gluconic acid, ascorbic acid, nicotinic acid, citric acid, adipic acid, and other acids known to those of ordinary skill in the art. For those skilled in the art, other methods are known for converting basic groups into cations (e.g. alkylation of amine groups) to generate positively charged ammonium groups and suitable counterions of the salts. If the PTH moiety contains both acidic and basic groups, the pharmaceutical preparations of the invention include internal salts or betaines (amphoteric ions) in addition to the salt forms mentioned. The respective salts can be obtained by conventional methods known to those skilled in the art, for example by contacting these complexes with organic or inorganic acids or bases in solvents or dispersants, or by anion exchange or cation exchange with other salts. The preparations of the present invention also include all salts of PTH complexes, which are not directly suitable for use as drugs due to their low physiological compatibility, but can be used, for example, as intermediates in chemical reactions or for the preparation of pharmaceutically acceptable salts.

As used herein, the term "antioxidant" or "oxidation protectant" refers to a compound that inhibits peptide oxidation.

As used herein, the term "pH adjuster" refers to a compound used to adjust the pH of a liquid solution or formulation.

The term "pharmaceutically acceptable" refers to a substance that does not cause harm when administered to a patient and preferably refers to a substance that is approved for use in animals, preferably humans, by a regulatory agency such as EMA (Europe) and/or FDA (US) and/or any other national regulatory agency.

As used herein, the term "physiological conditions" refers to an aqueous buffer solution at pH 7.4 and 37°C.

As used herein, the term "polypeptide" refers to a chain of at least 2 and up to 50 amino acid monomer moieties connected by peptide (amide) bonds. For simplicity, for PTH drugs and PTH moieties only, sequences with more than 50 amino acids are also referred to as "polypeptides".

As used herein, the term "protein" refers to a chain of more than 50 amino acid monomer moieties linked by peptide bonds, wherein preferably no more than 12,000 amino acid monomer moieties are linked by peptide bonds, such as no more than 10,000 amino acid monomer moieties, no more than 8,000 amino acid monomer moieties, no more than 5,000 amino acid monomer moieties, or no more than 2,000 amino acid monomer moieties.

As used herein, the term "preservative" refers to compounds that have bacteriostatic and bactericidal properties by killing microorganisms such as bacteria and preventing the growth of such microorganisms.

As used herein, the term "polymer" refers to a molecule comprising repeating structural units (i.e., monomers) connected by chemical bonds in a linear, cyclic, branched, cross-linked or dendritic manner or a combination thereof, which may be synthetic or biologically derived or a combination of both. It should be understood that the polymer may also contain one or more other chemical groups and/or moieties, such as one or more functional groups. In some embodiments, the soluble polymer has a molecular weight of at least 0.5 kDa, for example, a molecular weight of at least 1 kDa, a molecular weight of at least 2 kDa, a molecular weight of at least 3 kDa, or a molecular weight of at least 5 kDa. If the polymer is soluble, its molecular weight is at most 1000 kDa, such as at most 750 kDa, such as at most 500 kDa, such as at most 300 kDa, such as at most 200 kDa, such as at most 100 kDa.

It should be understood that proteins or polypeptides are also polymers in which amino acids are repeated structural units, even though the side chains of each amino acid may be different.

As used herein, the term "polymer" or "polymer moiety" refers to an agent or moiety comprising one or more polymers or polymer moieties. A polymeric agent or moiety may also optionally comprise one or more other moieties, which in certain embodiments are selected from: C _1-50 alkyl, C _2-50 alkenyl, C _2-50 alkynyl, C _3-10 cycloalkyl, 3 to 10 membered heterocyclic, 8 to 11 membered heterobicyclic, phenyl, naphthyl, indenyl, indanyl and tetrahydronaphthyl; and a linking group selected from the following:

wherein the dashed line indicates attachment to the moiety or the remainder of the reagent, and -R and ^-Ra are each independently selected from: -H, methyl, ethyl, propyl, butyl, pentyl, and hexyl.

It is understood by those with ordinary knowledge in the art that the polymer products obtained by the polymerization reaction do not all have the same molecular weight, but rather exhibit a molecular weight distribution. Therefore, the molecular weight range, molecular weight, monomer number range, and monomer number range in the polymer used herein refer to the number average molecular weight and number average of the monomers, i.e., the arithmetic mean of the molecular weight of the polymer or polymer portion and the arithmetic mean of the number of monomers in the polymer or polymer portion.

Thus, in a polymer portion comprising "x" monomer units, any integer given to "x" corresponds to the arithmetic mean number of monomers. Any integer range given to "x" provides an integer range within which the arithmetic mean number of monomers lies. An integer given to "x" as "about x" means that the arithmetic mean number of monomers is within an integer range of x +/- 10%, in some embodiments, within an integer range of x +/- 8%, in some embodiments, within an integer range of x +/- 5%, and in some embodiments, within an integer range of x +/- 2%.

As used herein, the term "PEG-based" with respect to a moiety or agent means that the moiety or agent comprises PEG. In certain embodiments, the PEG-based moiety or agent comprises at least 10% (w/w) PEG, such as at least 20% (w/w) PEG, such as at least 30% (w/w) PEG, such as at least 40% (w/w) PEG, such as at least 50% (w/w), such as at least 60% (w/w) PEG, such as at least 70% (w/w) PEG, such as at least 80% (w/w) PEG, such as at least 90% (w/w) PEG, such as at least 95% (w/w) PEG. The remaining weight percent of the PEG-based moiety or agent is selected from the following moieties and other moieties of the linking group: ˙C _1-50 alkyl, C _2-50 alkenyl, C _2-50 alkynyl, C _3-10 cycloalkyl, 3 to 10 membered heterocyclic, 8 to 11 membered heterobicyclic, phenyl, naphthyl, indenyl, indanyl and tetrahydronaphthyl; and ˙a linking group selected from the following:

wherein the dashed line indicates attachment to the moiety or the remainder of the reagent, and -R and ^-Ra are each independently selected from: -H, methyl, ethyl, propyl, butyl, pentyl, and hexyl.

As used herein, the term "PEG-based comprising at least X% PEG" with respect to a moiety or agent means that the moiety or agent comprises at least X% (w/w) _ethylene glycol units ( _-CH2CH2O- ), wherein the ethylene glycol units may be arranged in blocks, alternatingly or may be randomly distributed in the moiety or agent. In certain embodiments, all of the ethylene glycol units of the moiety or agent are present in one block; In certain embodiments, the remaining weight percent of the PEG-based moiety or agent is selected from the following moieties and other moieties of linking groups: ˙C _1-50 alkyl, C _2-50 alkenyl, C _2-50 alkynyl, C _3-10 cycloalkyl, 3 to 10 membered heterocyclic, 8 to 11 membered heterobicyclic, phenyl, naphthyl, indenyl, indanyl, and tetrahydronaphthyl; and ˙a linking group selected from the following:

wherein the dashed line indicates attachment to the moiety or the remainder of the reagent, and -R and ^-Ra are each independently selected from: -H, methyl, ethyl, propyl, butyl, pentyl, and hexyl.

As used herein, the term "hyaluronic acid-based comprising at least X% hyaluronic acid" is used accordingly.

Those of ordinary skill in the art also recognize that the conjugates of the present invention are prodrugs. As used herein, the term "prodrug" refers to a drug moiety, such as a PTH moiety, reversibly and covalently conjugated to a water-soluble carrier (such as -Z) through a reversible linking group moiety. The prodrug releases the reversibly and covalently bound drug moiety in the form of its corresponding drug. In other words, the prodrug is a conjugate comprising a drug moiety (such as a PTH moiety) that is covalently and reversibly conjugated to a water-soluble carrier through a reversible linking group moiety, and wherein the conjugation of the carrier to the reversible linking group moiety is direct or through a spacer group. Such a prodrug or conjugate releases the previously conjugated drug moiety in the form of a free drug.

As used herein, the term "random coil" refers to a peptide or protein that, in certain embodiments, adopts/has/forms a conformation that substantially lacks defined secondary and tertiary structure as determined by circular dichroism in an aqueous buffer at ambient temperature and pH 7.4. In certain embodiments, the ambient temperature is about 20°C, i.e., between 18°C and 22°C, and in certain embodiments, the ambient temperature is 20°C.

As used herein, the term "reversible linkage" is a linkage that is cleavable under physiological conditions (in aqueous buffer at pH 7.4, 37°C) and in the absence of enzymes, and has a half-life of one hour to six months, such as one hour to four months, one hour to three months, one hour to two months, or one hour to one month. Thus, a stable linkage is a linkage that has a half-life of more than six months under physiological conditions (aqueous buffer at pH 7.4, 37°C).

As used herein, the term "reversibly linking moiety" is a moiety that is covalently linked to a drug moiety (e.g., a PTH moiety) through a reversible link and is also covalently linked to a water-soluble carrier (e.g., -Z), wherein the covalent link to the carrier is direct or through a spacer moiety such as -L ² -. In certain embodiments, the link between -Z and -L ² - is a stable link.

As used herein, the term "reagent" refers to a compound containing at least one functional group for reacting with a functional group of another compound or drug. It should be understood that drugs containing functional groups (such as primary amines, secondary amines, or hydroxyl functional groups) are also reagents.

As used herein, the term "spacer group" or "spacer moiety" refers to a moiety suitable for linking two moieties. Suitable spacer groups may be selected from: C _1-50 alkyl, C _2-50 alkenyl or C _2-50 alkynyl, which may be interrupted by one or more groups selected from: -NH-, -N(C _1-4 alkyl)-, -O-, -S-, -C(O)-, -C(O)NH-, -C(O)N(C _1-4 alkyl)-, -OC(O)-, -S(O)-, -S(O) ₂ -, 4 to 7 membered heterocyclic groups, phenyl and naphthyl.

As used herein, the term "substituted" refers to the replacement of one or more hydrogen atoms of a molecule or moiety with a different atom or group of atoms, which is referred to as "substitution".

In certain embodiments, such one or more substituents are independently selected from: halogen, -CN, ^{-COORx1 , -ORx1 ,} -C(O) ^Rx1 , -C(O)N( ^Rx1Rx1a ), -S(O) _2N ( ^Rx1Rx1a ), -S(O)N( ^Rx1Rx1a ), -S(O) _2Rx1 , -S(O) ^Rx1 , -N( ^Rx1 ) ^S (O) _2N ^{( Rx1Rx1b} ), ^-SRx1 , -N( ^Rx1Rx1a ), _-NO2 , -OC(O) ^Rx1 , -N( ^Rx1 ) ^C (O) ^Rx1a , -N( ^Rx1 )S(O) _2Rx1a , -N( ^Rx1 )S(O) ^Rx1a , -N ^{( Rx1} ) ^{wherein -T0 , C1-50 alkyl} , _C2-50 alkenyl and _C2-50 alkynyl are optionally substituted ^{with one or more identical or different -Rx2, and wherein C1-50 alkyl, C2-50 alkenyl and C2-50 alkynyl are optionally interrupted by one or more groups selected from the following: -T0- ,} _{-C ( O} ^{) O-} _{, -O-} ^, -C( ^O )-, -C( _O )N(Rx3)-, -S(O) _2N ( ^Rx3 )-, _-S (O)N( ^Rx4 )- ^{; wherein -Rx1} , -Rx1a _, -Rx1b are independently _{selected from} ^: -H, -T0, C1-50 alkyl ^, C2-50 alkenyl and C2-50 alkynyl ^{; wherein -T0} , C1-50 ^{alkyl , C2-50} alkenyl and _C2-50 alkynyl are optionally substituted by one ^{or more} identical or _different -Rx2, and wherein C1-50 _alkyl , _C2-50 ^alkenyl and _C2-50 ^alkynyl are substituted by one _{or more} identical or _different ^-Rx2 . _2-50 alkynyl is optionally interrupted by one or more groups selected from the following groups: ^-T0- , -C(O)O-, -O-, -C(O)-, -C(O)N( ^Rx3 )-, -S(O) _2N ( ^Rx3 )-, -S(O)N( ^Rx3 )-, -S(O) _2- , -S(O)-, -N( ^Rx3 )S(O) _2N ( ^Rx3a )-, -S-, -N( ^Rx3 )-, ^-OC (ORx3)( ^Rx3a )-, -N( ^Rx3 )C(O)N( ^Rx3a )- and -OC(O)N( ^Rx3 )-; each ^T0 is independently selected from: phenyl, naphthyl, indenyl, indanyl, tetrahydronaphthyl, C _3-10 membered cycloalkyl, 3- to 10-membered heterocycloalkyl and 8- to 11-membered heterobicycloalkyl. wherein each ^T0 is independently optionally substituted by one or more identical or different ^-Rx2 ; each ^-Rx2 is independently selected from: halogen, -CN, oxo (=O), ^-COORx4 , ^-ORx4 , -C(O) ^Rx4 , -C(O)N( ^Rx4Rx4a ), -S( ^O ) _2N ( ^Rx4Rx4a ), -S(O)N( ^{Rx4Rx4a )} , -S(O) _2Rx4 , -S(O) ^Rx4 , -N( ^Rx4 ) ^S ( ^{O)2N(Rx4aRx4b), -SRx4, -N(Rx4Rx4a} ₎ ^{, -NO2 ,} _-OC (O) ^Rx4 , -N( ^Rx4 )C(O) ^Rx4a , -N( ^Rx4 ) ^S ( ^O ₎ ^2Rx4a , -N(R ^x4 )S(O)R ^x4a , -N(R ^x4 )C(O)OR ^x4a , -N(R ^x4 )C(O)N(R ^x4a R ^x4b ), -OC(O)N(R ^x4 R ^x4a ) and C _1-6 alkyl; wherein the C _1-6 alkyl is optionally substituted by one or more the same or different halogens; each -R ^x3 , -R ^x3a , -R ^x4 , -R ^x4a , -R ^x4b is independently selected from: -H and C _1-6 alkyl; wherein the C _1-6 alkyl is optionally substituted by one or more the same or different halogens.

In certain embodiments, one or more substituents are independently selected from: halogen, -CN, -COORx1, ^-ORx1 , -C(O) ^Rx1 , -C(O) ^N ( ^Rx1Rx1a ), -S(O) _2N ^{( Rx1Rx1a} ), -S(O)N ^{( Rx1Rx1a} ), -S(O) _2Rx1 , -S(O) ^Rx1 , -N( ^Rx1 ) ^S (O) ^2N ( ^Rx1aRx1b ), ^-SRx1 , -N( ^Rx1Rx1a ) _{, -NO2} , -OC(O) ^Rx1 , -N( ^{Rx1 )} C(O) ^Rx1a , -N( ^Rx1 )S(O) ^2Rx1a , -N( ^{Rx1 )} S(O ₎ ^Rx1a , -N( ^Rx1) )C(O) ^ORx1a , -N( ^Rx1 ) ^C (O)N(Rx1aRx1b), -OC(O)N( ^{Rx1Rx1a )} , ^-T0 , ^C1-10 alkyl, _C2-10 alkenyl and _C2-10 alkynyl; wherein ^-T0 , _C1-10 alkyl, _C2-10 alkenyl and _C2-10 alkynyl are optionally substituted by one or more identical or different ^-Rx2 , and wherein _{C1-10 alkyl} , _C2-10 alkenyl and _C2-10 alkynyl are optionally interrupted by one or more groups selected from the following: ^-T0- , -C(O)O-, -O-, -C(O)-, -C(O)N( ^Rx3 )-, -S(O) _2N ( ^Rx3 )-, -S(O)N( ^Rx3) - )-, -S(O) ₂ -, -S(O)-, -N(R ^x3 )S(O) ₂ N(R ^x3a )-, -S-, -N(R ^x3 )-, -OC(OR ^x3 )(R ^x3a )-, -N(R ^x3 )C(O)N(R ^x3a )-, and -OC(O)N(R ^x3 )-; each -R ^x1 , -R ^x1a , -R ^x1b , -R ^x3 , -R ^x3a is independently selected from: -H, halogen, C _1-6 alkyl, C _2-6 alkenyl, and C _2-6 alkynyl; each T ⁰ is independently selected from: phenyl, naphthyl, indenyl, indanyl, tetrahydronaphthyl (tetralinyl), C _3-10 membered cycloalkyl, 3 to 10 membered heterocyclic group and 8 to 11 membered heterobicyclic group; wherein each T ⁰ is independently optionally substituted by one or more identical or different -R ^x2 ; each -R ^x2 is independently selected from: halogen, -CN, oxo (=O), -COOR ^x4 , -OR ^x4 , -C(O)R ^x4 , -C(O)N(R ^x4 R ^x4a ), -S(O) ₂ N(R ^x4 R ^x4a ), -S(O)N(R ^x4 R ^x4a ), -S(O) ₂ R ^x4 , -S(O)R ^x4 , -N(R ^c4 )S(O) ₂ N(R ^x4a R ^x4b ), -SR ^x4 , -N(R ^x4 R ^x4a ), -NO ₂ , -OC(O)R ^x4 wherein the C 1-6 ^{alkyl is optionally} substituted with _one or ^more of the same or different halogens; each -R ^x4 , -R ^x4a , -R ^x4b is independently selected from: ^-H , halogen, C _{1-6 alkyl, C 2-6} ^alkenyl _{and C} ^2-6 _alkynyl ^{; in certain} embodiments, one or more substituents are independently selected from: halogen ^, -CN, ^{-COOR x1 ,} _-OR ^x1 , -C(O)R ^x1 , -C(O)N(R ^x1 R ^x1a ), -S(O) ₂ N(R ^x1 R ^x1a ), -S(O)N(R ^x1 R ^x1a ), -S(O) ₂ R ^x1 , -S(O)R ^x1 , -N(R ^x1 )S(O) ₂ N(R ^x1a R ^x1b ), -SR ^x1 , -N(R ^x1 R ^x1a ), -NO ₂ , -OC(O)R ^x1 , -N(R ^x1 )C(O)R ^x1a , -N(R ^x1 )S(O) ₂ R ^x1a , -N(R ^x1 )S(O)R ^x1a , -N(R ^x1 )C(O)OR ^x1a , -N(R ^x1 )C(O)N(R ^x1a R ^x1b wherein ^-T0 , _C1-6 alkyl, _C2-6 alkenyl and _C2-6 alkynyl are optionally substituted with one or more identical or ^{different -Rx2 ,} and wherein _C1-6 alkyl, _C2-6 alkenyl and _C2-6 alkynyl are optionally interrupted by one or more groups selected from the following _: ^{-T0- ,} -C( _O )O-, -O-, -C(O ₎ -, -C(O)N(Rx3)-, -S(O)2N( ^Rx3 )-, -S(O)N( ^Rx3 )-, -S(O) _{2- ,} -S(O)-, -N( ^Rx3 ) ^S (O) _2N ( ^Rx3a )-, -S-, -N(R ^x3 )-, -OC(OR ^x3 )(R ^x3a )-, -N(R ^x3 )C(O)N(R ^x3a )- and -OC(O)N(R ^x3 )-; each -R ^x1 , -R ^x1a , -R ^x1b , -R ^x3 , -R ^x3a is independently selected from: -H, halogen, C _1-6 alkyl, C _2-6 alkenyl and C _2-6 alkynyl; each T ⁰ is independently selected from: phenyl, naphthyl, indenyl, indanyl, tetrahydronaphthyl, C _3-10 cycloalkyl, 3 to 10 membered heterocyclic group and 8 to 11 membered heterobicyclic group; wherein each T ⁰ is independently optionally substituted by one or more identical or different -R ^x2 .

In certain embodiments, up to 6 hydrogen atoms of the optionally substituted molecule are independently substituted by substituents, for example, 5 hydrogen atoms are independently substituted by substituents, 4 hydrogen atoms are independently substituted by substituents, 3 hydrogen atoms are independently substituted by substituents, 2 hydrogen atoms are independently substituted by substituents, or 1 hydrogen atom is substituted by a substituent.

As used herein, the terms "stable" and "stability" with respect to a pharmaceutical formulation mean that after a storage period, for example after one month, two months, four months, six months, eight months, twelve months, eighteen months, twenty-four months, thirty-six months, in particular after a specified storage period, the pharmaceutical formulation contains less than 5% of the drug in free form and less than 20%, for example less than 10%, for example less than 5%, of impurities, for example impurities resulting from oxidation of methionine or tryptophan, isomerization of aspartate or aspartate salts, cleavage of peptide bonds on aspartate, aspartate salts or asparagine, deamination of asparagine or glutamine and aggregation of peptides. Impurities can be quantified by RP-HPLC or SEC based on their individual peak areas in the chromatogram relative to the total peak area of all PTH conjugate-related peaks, and after the PTH fraction is released from the PTH conjugate, the impurities in the PTH fraction of the PTH conjugate can be determined.

As used herein, the term "stabilizer" refers to a compound used to stabilize a drug conjugate. Stabilization can be achieved by enhancing peptide stabilization or by directly conjugating the excipient to the drug conjugate.

As used herein, the term "surfactant" refers to a wetting agent that reduces the surface tension of a liquid.

As used herein, the term "sealed container" means a container that is closed in an airtight manner, thereby not allowing gas exchange between the inside and the outside and maintaining the sterility of the contents.

As used herein, the term "therapeutically effective amount" refers to an amount sufficient to cure, alleviate or partially prevent the clinical manifestations of a given disease and its complications. The effective amount for each purpose will depend on the severity of the disease or injury, as well as the weight and general condition of the subject. It will be understood that the appropriate dosage can be determined using routine experiments by constructing a matrix of values and testing different points in the matrix, which are all ordinary skills of trained physicians. Within the scope of the present invention, a therapeutically effective amount involves an amount intended to achieve a therapeutic effect over an extended period of time, i.e., at least one day, such as two days, such as three days, such as four days, such as 5 days, such as 6 days, such as 1 week, or such as 2 weeks.

As used herein, the term "traceless linking group" refers to a reversible linking group that releases the drug in free form upon cleavage.

As used herein, the term "unit dose" refers to the amount of drug administered to a patient in a single dose.

As used herein, the term "water-soluble" with respect to a water-soluble carrier means that when such a carrier is a part of a PTH complex, at least 1 g of the PTH complex containing such a water-soluble carrier can be dissolved in one liter of water at 20°C to form a uniform solution.

Generally speaking, the term "comprising" also covers "consisting of".

In certain embodiments, the PTH portion of the PTH conjugate has a sequence of SEQ ID NO: 47, SEQ ID NO: 48, SEQ ID NO: 49, SEQ ID NO: 50, SEQ ID NO: 51, SEQ ID NO: 52, SEQ ID NO: 53, SEQ ID NO: 54, SEQ ID NO: 55, SEQ ID NO: 107, SEQ ID NO: 108, SEQ ID NO: 109, SEQ ID NO: 110, SEQ ID NO: 111, SEQ ID NO: 112, SEQ ID NO: 113, SEQ ID NO: 114, or SEQ ID NO: 115. In certain embodiments, the PTH portion has a sequence of SEQ ID NO: 50, SEQ ID NO: 51, SEQ ID NO: 52, SEQ ID NO: 110, SEQ ID NO: 111, or SEQ ID NO: 112. In certain embodiments, the PTH portion has a sequence of SEQ ID NO: 50. In some embodiments, the PTH portion has a sequence of SEQ ID NO: 52. In some embodiments, the PTH portion has a sequence of SEQ ID NO: 110. In some embodiments, the PTH portion has a sequence of SEQ ID NO: 111. In some embodiments, the PTH portion has a sequence of SEQ ID NO: 112. In some embodiments, the PTH portion has a sequence of SEQ ID NO: 51.

In certain embodiments, a water-soluble carrier moiety is defined as variable -Z, which is described in more detail elsewhere herein.

The liquid pharmaceutical preparation of the present invention contains a buffer. The buffer can be selected from: succinic acid, citric acid, lactic acid, acetic acid, glutamic acid, fumaric acid, aspartic acid, glutaric acid, phosphoric acid, histidine, gluconic acid, tartaric acid, apple acid and mixtures thereof. It is clear to those skilled in the art that the corresponding conjugate base or salt of the buffer, such as succinate, citrate, lactate, acetate, glutamate, fumarate, aspartic acid, glutarate, phosphate, gluconate, tartrate, apple acid and mixtures thereof are also included.

In some embodiments, the buffer is succinic acid. In some embodiments, the buffer is citric acid. In some embodiments, the buffer is lactic acid. In some embodiments, the buffer is acetic acid. In some embodiments, the buffer is glutamate. In some embodiments, the buffer is fumaric acid. In some embodiments, the buffer is aspartic acid. In some embodiments, the buffer is glutaric acid. In some embodiments, the buffer is phosphoric acid. In some embodiments, the buffer is histidine. In some embodiments, the buffer is gluconic acid. In some embodiments, the buffer is tartaric acid. In some embodiments, the buffer is malic acid.

In some embodiments, the concentration of the buffer is 0.25 to 24 mg/ml. In some embodiments, the concentration of the buffer is 0.6 to 6.0 mg/ml. In some embodiments, the concentration of the buffer is 1.0 to 1.4 mg/ml. In some embodiments, the concentration of the buffer is about 1.18 mg/ml.

In order to maintain a certain pH or pH range, the liquid pharmaceutical formulation contains a buffer. The buffer maintains the pH of the liquid pharmaceutical formulation within the desired range. In certain embodiments, the pH of the liquid pharmaceutical formulation is not higher than 6 because the reversible linkage within the PTH complex may be unstable under alkaline conditions.

In some embodiments, the pH of the liquid pharmaceutical formulation is about pH 3.0 to about pH 6.0. In some embodiments, the pH of the liquid pharmaceutical formulation is about pH 3.5 to about pH 5.0. In some embodiments, the pH of the liquid pharmaceutical formulation is about pH 3.7 to about pH 4.3. In some embodiments, the pH of the liquid pharmaceutical formulation is 4.0.

The liquid pharmaceutical preparation of the present invention comprises an isotonic agent. The isotonic agent can be selected from: mannitol, trehalose, sucrose, raffinose, gelatin, lactose, calcium hydrogen phosphate, sorbitol, xylitol, glycine, histidine, ethanol, hydroxyethyl starch, potassium chloride, sodium chloride, glucose, dextran, polysucrose ( ^Ficoll® ), propylene glycol and mixtures thereof.

In certain embodiments, the isotonic agent is selected from: mannitol, trehalose, sucrose, raffinose, gelatin, lactose, calcium hydrogen phosphate, sorbitol, xylitol, glycine, histidine, ethanol, hydroxyethyl starch, potassium chloride, sodium chloride, glucose, dextran, propylene glycol and mixtures thereof.

In some embodiments, the isosmotic agent is mannitol. In some embodiments, the isosmotic agent is trehalose. In some embodiments, the isosmotic agent is sucrose. In some embodiments, the isosmotic agent is raffinose. In some embodiments, the isosmotic agent is gelatin. In some embodiments, the isosmotic agent is lactose. In some embodiments, the isosmotic agent is calcium hydrogen phosphate. In some embodiments, the isosmotic agent is sorbitol. In some embodiments, the isosmotic agent is xylitol. In some embodiments, the isosmotic agent is glycine. In some embodiments, the isosmotic agent is histidine. In some embodiments, the isosmotic agent is ethanol. In some embodiments, the isosmotic agent is hydroxyethyl starch. In some embodiments, the isosmotic agent is potassium chloride. In some embodiments, the isosmotic agent is sodium chloride. In some embodiments, the isosmotic agent is glucose. In some embodiments, the isosmotic agent is dextran. In some embodiments, the isosmotic agent is polysucrose. In some embodiments, the isosmotic agent is propylene glycol.

As defined herein, the term "trehalose" is intended to cover all salts and hydration states of trehalose, such as anhydrous trehalose or trehalose dihydrate. In certain embodiments, the term "trehalose" refers to anhydrous trehalose. In certain embodiments, the term "trehalose" refers to trehalose dihydrate.

As defined herein, the term "mannitol" is intended to cover D-mannitol and L-mannitol and mixtures thereof. In certain embodiments, the term "mannitol" refers to L-mannitol. In certain embodiments, the term "mannitol" refers to D-mannitol. In certain embodiments, the term "mannitol" refers to a mixture of L-mannitol and D-mannitol.

In some embodiments, the concentration of the isosmotic agent is 10 to 200 mg/ml. In some embodiments, the concentration of the isosmotic agent is 30 to 60 mg/ml. In some embodiments, the concentration of the isosmotic agent is 36 to 48 mg/ml. In some embodiments, the concentration of the isosmotic agent is about 41.7 mg/ml.

The liquid pharmaceutical preparation of the present invention contains a preservative. The preservative can be selected from: m-cresol, benzyl alcohol, benzoic acid, phenol, methyl p-hydroxybenzoate, ethyl p-hydroxybenzoate, propyl p-hydroxybenzoate, butyl p-hydroxybenzoate, potassium sorbate, chlorobutanol, benzyl alcohol, phenylmercuric nitrate, thimerosal, sorbic acid, potassium sorbate, chlorocresol, benzalkonium chloride, 2-ethoxyethanol, chlorhexidine, chlorobutanol, phenylethanol, phenylmercuric acetate and mixtures thereof.

In some embodiments, the preservative is m-cresol. In some embodiments, the preservative is benzyl alcohol. In some embodiments, the preservative is benzoic acid. In some embodiments, the preservative is phenol. In some embodiments, the preservative is methyl parahydroxybenzoate. In some embodiments, the preservative is ethyl parahydroxybenzoate. In some embodiments, the preservative is propyl parahydroxybenzoate. In some embodiments, the preservative is butyl parahydroxybenzoate. In some embodiments, the preservative is potassium sorbate. In some embodiments, the preservative is chlorocresol. In some embodiments, The preservative is benzyl alcohol. In some embodiments, the preservative is phenylmercuric nitrate. In some embodiments, the preservative is thimerosal. In some embodiments, the preservative is sorbic acid. In some embodiments, the preservative is potassium sorbate. In some embodiments, the preservative is chlorocresol. In some embodiments, the preservative is benzalkonium chloride. In some embodiments, the preservative is 2-ethoxyethanol. In some embodiments, the preservative is chlorhexidine. In some embodiments, the preservative is chlorobutanol. In some embodiments, the preservative is phenylethyl alcohol. In some embodiments, the preservative is phenylmercuric acetate.

In some embodiments, the concentration of the preservative is 1 to 10 mg/ml. In some embodiments, the concentration of the preservative is 1.5 to 3.5 mg/ml. In some embodiments, the concentration of the preservative is 2 to 3 mg/ml. In some embodiments, the concentration of the preservative is about 2.5 mg/ml.

The liquid pharmaceutical formulation of the present invention may further comprise a pH adjuster. In certain embodiments, the pH adjuster is an acid. Examples of acids may be selected from: hydrochloric acid, phosphoric acid, carbonic acid, nitric acid, and mixtures thereof.

In some embodiments, the pH adjuster is hydrochloric acid. In some embodiments, the pH adjuster is phosphoric acid. In some embodiments, the pH adjuster is carbonic acid. In some embodiments, the pH adjuster is nitric acid.

In certain embodiments, the pH adjuster is a base. Examples of bases may be selected from: Tris (tris(hydroxymethyl)aminomethane), potassium hydroxide, lysine, sodium hydroxide, and mixtures thereof.

In some embodiments, the pH adjuster is Tris. In some embodiments, the pH adjuster is potassium hydroxide. In some embodiments, the pH adjuster is lysine. In some embodiments, the pH adjuster is sodium hydroxide.

In some embodiments, the pH adjuster is a mixture of at least one base and at least one acid. In some embodiments, the pH adjuster is a mixture of one base and one acid. In some embodiments, the pH adjuster is a mixture of sodium hydroxide and hydrochloric acid.

In certain embodiments, the concentration of the pH adjuster or mixture of pH adjusters is 0.01 to 5 mg/ml. In certain embodiments, the concentration of the pH adjuster or mixture of pH adjusters is 0.04 to 2.5 mg/ml. In certain embodiments, the concentration of the pH adjuster or mixture of pH adjusters is 0.08 to 1.25 mg/ml. In certain embodiments, the concentration of the pH adjuster or mixture of pH adjusters is about 0.13 mg/ml. It should be understood that in the case of a mixture of pH adjusters, the concentration provided refers to the total concentration of all pH adjusters.

The liquid pharmaceutical preparation of the present invention optionally contains an antioxidant. Examples of antioxidants can be selected from: methionine, butylhydroxytoluene, butylhydroxyanisole, tocopherol, propyl gallate, ascorbic acid, sodium bisulfite, ethylenediaminetetraacetic acid (EDTA), cysteine, glutathione, monothioglycerol, poly(ethyleneimine), vitamin E, tetrahydropyrimidine, morin and mixtures thereof.

In some embodiments, the antioxidant is methionine. In some embodiments, the antioxidant is ascorbic acid. In some embodiments, the antioxidant is butylated hydroxytoluene. In some embodiments, the antioxidant is butylated hydroxyanisole. In some embodiments, the antioxidant is tocopherol. In some embodiments, the antioxidant is propyl gallate. In some embodiments, the antioxidant is sodium bisulfite. In some embodiments, the antioxidant is monothioglycerol. In some embodiments, the antioxidant is EDTA. In some embodiments, the antioxidant is cysteine. In some embodiments, the antioxidant is glutathione. In some embodiments, the antioxidant is polyethyleneimine. In some embodiments, the antioxidant is vitamin E. In some embodiments, the antioxidant is tetrahydropyrimidine. In some embodiments, the antioxidant is morin.

As defined herein, the term "methionine" is intended to encompass D-methionine and L-methionine and mixtures thereof. In certain embodiments, the term "methionine" refers to L-methionine. In certain embodiments, the term "methionine" refers to D-methionine. In certain embodiments, the term "methionine" refers to a mixture of D-methionine or L-methionine.

As defined herein, the term "EDTA" is intended to cover all forms of EDTA known in the art, such as EDTA salts, including EDTA metal salts, such as EDTA disodium salt, EDTA dipotassium salt, EDTA calcium salt, EDTA dimagnesium salt, or mixtures thereof. In certain embodiments, EDTA refers to EDTA disodium salt. In certain embodiments, the term "EDTA" refers to EDTA dicalcium salt. In certain embodiments, the term "EDTA" refers to anhydrous EDTA.

In certain embodiments, the molar ratio of the antioxidant to the PTH portion is from about 0.1:1 to about 100:1. In certain embodiments, the molar ratio of the antioxidant to the PTH portion is from about 0.1:1 to about 70:1. In certain embodiments, the molar ratio of the antioxidant to the PTH portion is from about 0.1:1 to about 15:1. In certain embodiments, the molar ratio of the antioxidant to the PTH portion is from about 1:1 to about 10:1. In certain embodiments, the molar ratio of the antioxidant to the PTH portion is from about 3:1 to about 7:1.

In certain embodiments, the liquid pharmaceutical formulation of the present invention does not contain an antioxidant.

The liquid pharmaceutical preparation of the present invention contains a PTH conjugate.

In certain embodiments, the liquid pharmaceutical preparation comprises a PTH conjugate, wherein the PTH portion is present at a concentration of 0.05 to 5.0 mg/ml. In certain embodiments, the liquid pharmaceutical preparation comprises a PTH conjugate, wherein the PTH portion is present at a concentration of 0.1 to 5.0 mg/ml. In certain embodiments, the liquid pharmaceutical preparation comprises a PTH conjugate, wherein the PTH portion is present at a concentration of 0.1 to 1.5 mg/ml. In certain embodiments, the liquid pharmaceutical preparation comprises a PTH conjugate, wherein the PTH portion is present at a concentration of 0.25 to 0.35 mg/ml. In certain embodiments, the liquid pharmaceutical preparation comprises a PTH conjugate, wherein the PTH portion is present at a concentration of about 0.3 mg/ml. It should be understood that the concentrations provided above refer to the amount of the PTH portion, not the entire PTH complex.

In certain embodiments, the PTH conjugate has the structure of Formula (Ia) or (Ib):

其中-D是PTH部分；-L¹-是通過PTH的官能團與PTH部分-D連接的可逆連接基團部分；-L²-是單化學鍵或間隔基團部分；-Z是水溶性載體部分；x是選自1、2、3、4、5、6、7、8、9、10、11、12、13、14、15和16的整數；並且y是選自1、2、3、4和5的整數。

wherein -D is a PTH moiety; -L ¹ - is a reversible linker moiety linked to the PTH moiety -D via a functional group of PTH; -L ² - is a single chemical bond or a spacer moiety; -Z is a water-soluble carrier moiety; x is an integer selected from 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15 and 16; and y is an integer selected from 1, 2, 3, 4 and 5.

In certain embodiments, -D is covalently and reversibly linked to -L ¹ -.

In some embodiments, x of formula (Ia) is an integer selected from 1, 2, 3, 4, 6 and 8. In some embodiments, x of formula (Ia) is an integer selected from 1, 2, 4 and 6. In some embodiments, x of formula (Ia) is an integer selected from 1, 4 and 6. In some embodiments, x of formula (Ia) is 1.

In some embodiments, y of formula (Ib) is an integer selected from 2, 3, 4 and 5. In some embodiments, y of formula (Ib) is an integer selected from 2, 3 and 4. In some embodiments, y of formula (Ib) is an integer selected from 2 and 3.

In some embodiments, y of formula (Ib) is an integer selected from 1, 2, and 3. In some embodiments, y of formula (Ib) is 1. In some embodiments, y of formula (Ib) is 2.

In certain embodiments, the PTH conjugate has formula (Ia) where x=1.

In certain embodiments, -D has a sequence of SEQ ID NO: 47, SEQ ID NO: 48, SEQ ID NO: 49, SEQ ID NO: 50, SEQ ID NO: 51, SEQ ID NO: 52, SEQ ID NO: 53, SEQ ID NO: 54, SEQ ID NO: 55, SEQ ID NO: 107, SEQ ID NO: 108, SEQ ID NO: 109, SEQ ID NO: 110, SEQ ID NO: 111, SEQ ID NO: 112, SEQ ID NO: 113, SEQ ID NO: 114, or SEQ ID NO: 115.

In certain embodiments, -D has a sequence of SEQ ID NO: 50, SEQ ID NO: 51, SEQ ID NO: 52, SEQ ID NO: 110, SEQ ID NO: 111, or SEQ ID NO: 112.

In some embodiments, -D has a sequence of SEQ ID NO: 50. In some embodiments, -D has a sequence of SEQ ID NO: 52. In some embodiments, -D has a sequence of SEQ ID NO: 110. In some embodiments, -D has a sequence of SEQ ID NO: 111. In some embodiments, -D has a sequence of SEQ ID NO: 112. In some embodiments, -D has a sequence of SEQ ID NO: 51.

The -L ¹ -moiety is conjugated to a functional group of the side chain of the amino acid residue of -D, the N-terminal amine functional group or the C-terminal carboxyl functional group of -D, or a nitrogen atom in the backbone polypeptide of -D. The attachment of -D to the N-terminus or C-terminus can be directly through the corresponding amine or carboxyl functional group, respectively, or indirectly, wherein the spacer moiety is first conjugated to the amine or carboxyl functional group to which the spacer moiety -L ¹ - is conjugated.

In certain embodiments, the amino acid residue of PTH conjugated to -L ¹ - comprises a functional group selected from the group consisting of carboxylic acid, primary and secondary amines, maleimide, thiol, sulfonic acid, carbonate, carbamate, hydroxyl, aldehyde, ketone, hydrazine, isocyanate, isothiocyanate, phosphoric acid, phosphonic acid, halogenated acetyl, halogenated alkane, acryl, aryl fluoride, hydroxylamine, sulfate, disulfide, vinyl sulfide, vinyl ketone, diazoalkane, oxirane, guanidine, and aziridine. In certain embodiments, the amino acid residue of PTH conjugated to -L ¹ - comprises a functional group selected from the group consisting of hydroxyl, primary and secondary amines, and guanidine. In certain embodiments, the amino acid residue of PTH conjugated to -L ¹ - comprises a primary or secondary amine functional group. In certain embodiments, the amino acid residue of PTH conjugated to -L ¹ - comprises a primary amine functional group.

If the -L ¹ - moiety is conjugated to a functional group of a side chain of an amino acid residue of PTH, the amino acid residue is selected from: a proteinogenic amino acid residue and a non-proteinogenic amino acid residue.

In certain embodiments, -L ¹ - is conjugated to a functional group pendant to a non-proteinogenic amino acid residue of PTH. It should be understood that such a non-proteinogenic amino acid is not found in the sequence of native PTH or fragments thereof and can only be found in variants, analogs, orthologs, homologs and derivatives of PTH.

In certain embodiments, -L ¹ - is linked to a functional group of the side chain of the proteogenic amino acid residue of PTH. In certain embodiments, the amino acid is selected from: histidine, lysine, tryptophan, serine, threonine, tyrosine, aspartic acid, glutamic acid and arginine. In certain embodiments, the amino acid is selected from: lysine, aspartic acid, arginine and serine. In certain embodiments, the amino acid is selected from: lysine, arginine and serine.

In certain embodiments, -L ¹ - is conjugated to a functional group of the side chain of histidine of PTH. In certain embodiments, -L ¹ - is conjugated to a functional group of the side chain of lysine of PTH. In certain embodiments, -L ¹ - is conjugated to a functional group of the side chain of tryptophan of PTH. In certain embodiments, -L ¹ - is conjugated to a functional group of the side chain of serine of PTH. In certain embodiments, -L ¹ - is conjugated to a functional group of the side chain of threonine of PTH. In certain embodiments, -L ¹ - is conjugated to a functional group of the side chain of tyrosine of PTH. In certain embodiments, -L ¹ -is conjugated to a functional group of the side chain of aspartic acid of PTH. In certain embodiments, -L ¹ -is conjugated to a functional group of the side chain of glutamic acid of PTH. In certain embodiments, -L ¹ -is conjugated to a functional group of the side chain of arginine of PTH.

It should be understood that not every PTH moiety may contain all of these amino acid residues.

In certain embodiments, -L ¹ - is directly conjugated to the N-terminal amine function of PTH through the corresponding amine function, or indirectly conjugated to the N-terminal amine function of PTH, wherein the spacer moiety is first conjugated to the amine function to which the spacer moiety -L ¹ - is conjugated. In certain embodiments, -L ^{1 - is directly conjugated to the N-terminal amine function of PTH, such as PTH 1-34, i.e., PTH having the sequence of SEQ ID NO: 51. The N-terminal attachment of -L 1 -} is advantageous, i.e., attachment of -L ¹ - to the N-terminal of PTH, because it was found that this attachment site protects the N-terminal end that is critical for the activity of PTH. The major metabolite formed from the N-terminally linked PTH conjugate with -L ¹ - is PTH(1-33), the 33 N-terminal amino acids of PTH, which is known to be active.

In certain embodiments, -L ¹ - is directly conjugated to the C-terminal functional group of PTH via the corresponding carboxyl functional group, or is indirectly conjugated to the C-terminal functional group of PTH, wherein the spacer moiety is first conjugated to the carboxyl functional group conjugated to the spacer group -L ¹ -.

In certain embodiments, -L ¹ - is directly conjugated to the N-terminal amine functionality of PTH.

The -L ¹ - moiety can be linked to -D via any type of linking group, as long as it is reversible. In certain embodiments, -L ¹ - is linked to -D via a linking group selected from the group consisting of amides, esters, carbamates, acetals, aldehydes, imines, oximes, hydrazones, disulfides, and acylguanidines. In certain embodiments, -L ¹ - is linked to -D via a linking group selected from the group consisting of amides, esters, carbamates, and acylguanidines. It will be appreciated that some of these linking groups are themselves irreversible, but adjacent groups included in -L ¹ - render these linking groups reversible.

In certain embodiments, -L ¹ - is linked to -D via an ester linking group. In certain embodiments, -L ¹ - is linked to -D via a carbamate linking group. In certain embodiments, -L ¹ - is linked to -D via an acylguanidine linking group. In certain embodiments, -L ¹ - is linked to -D via an amide linking group.

The -L ¹ - moiety is a reversible linking group from which the drug, i.e. PTH, is released in its free form, which means that -L ¹ - is a traceless linking group. Suitable reversible linking groups are known in the art, for example, the reversible linking group moieties disclosed in WO 2005/099768 A2, WO 2006/136586 A2, WO 2011/089216 A1 and WO 2013/024053 A1, which are incorporated herein by reference.

In certain embodiments, -L ¹ - is a reversible linking group as described in WO 2011/012722 A1, WO 2011/089214 A1, WO 2011/089215 A1, WO 2013/024052 A1, and WO 2013/160340 A1, which are incorporated herein by reference.

The -L ¹ -moiety is disclosed in WO 2009/095479 A2. Therefore, in certain embodiments, the -L ¹ -moiety has a structure of formula (II):

wherein the dashed line indicates attachment to the nitrogen, hydroxyl ^{or thiol of -D, wherein -D is a PTH moiety; -X- is -C(R4R4a )-; -N(R4)-; -O-; -C(R4R4a )-C(R5R5a ) - ; -C ( R5R5a} )-C ^{( R4R4a} )-; -C( ^R4R4a )-N ^{( R6} )-; -N( ^R6 ) ^-C ( ^R4R4a )-; -C( ^R4R4a )-O-; -OC ^{( R4R4a} )- ^; or -C( ^R7R7a )-; ^X1 is C; or S(O); ^-X2- is -C( ^R8R8a )-; ^or -C( ^{R8R8a ) -} )-C(R ⁹ R ^9a )-; =X ³ is =O; =S; or =N-CN; -R ¹ , -R ^1a , -R ² , -R ^2a , -R ⁴ , -R ^4a , -R ⁵ , -R ^5a , -R ⁶ , -R ⁸ , -R ^8a , -R ⁹ , -R ^9a are independently selected from: -H and C _1-6 alkyl; -R ³ and -R ^3a are independently selected from: -H and C _1-6 alkyl, in the case where one or both of -R ³ and -R ^3a are not -H, they are connected to the N to which they are connected through an sp ³ -hybridized carbon atom; -R ⁷ is -N(R ¹⁰ R ^10a ) or -NR ¹⁰ -(C=O)-R ¹¹ ; -R ^7a , -R ¹⁰ , -R ^10a , and -R ¹¹ are independently -H or C _1-6 alkyl.

Optionally, one or more of the pairs -R ^1a / -R ^4a , -R ^1a / -R ^5a , -R ^1a / -R ^7a , -R ^4a / -R ^5a , -R ^8a / -R ^9a form a chemical bond; Optionally, one or more of the pairs -R ¹ / -R ^1a , -R ² / -R ^2a , -R ⁴ / -R ^4a , -R ⁵ / -R ^5a , -R ⁸ / -R ^8a , -R ⁹ / -R ^9a are linked together with the atoms to which they are linked to form a C _3-10 cycloalkyl group; or a 3 to 10 membered heterocyclic group; Optionally, -R ¹ / -R ⁴ , -R ¹ / -R ⁵ , -R ¹ / -R ⁶ , -R ¹ / -R One or more of the pairs ^{-R 7a} , -R ⁴ /-R ⁵ , -R ⁴ /-R ⁶ , -R ⁸ /-R ⁹ , -R ² /-R ³ and the atoms to which they are attached are linked together to form ring A; optionally, R ³ /R ^3a and the nitrogen atom to which they are attached are linked together to form a 3- to 10-membered heterocyclic ring; A is selected from: phenyl, naphthyl, indenyl, indanyl, tetrahydronaphthyl, C _3-10 cycloalkyl, 3- to 10-membered heterocyclic ring and 8- to 11-membered heterobicyclic ring; wherein -L ¹ - is substituted by -L ² -Z, and wherein -L ¹ - is optionally further substituted, provided that the hydrogen marked with an asterisk in formula (II) is not substituted by -L ² -Z or a substituent; wherein -L ² - is a single chemical bond or a spacer group; and -Z is a water soluble carrier.

In certain embodiments, -L ¹ - of formula (II) is substituted with one -L ² -Z moiety.

In certain embodiments, -L ¹ - of formula (II) is not further substituted.

其中虛線表示與-L¹-的其餘部分相連；該環包含3-10個原子且包含至少一個氮的原子；和R^#和R^##表示sp³-雜化的碳原子。 It should be understood that if R ³ /-R ^3a of formula (II) is linked together with the nitrogen atom to which they are attached to form a 3- to 10-membered heterocyclic ring, only a 3- to 10-membered heterocyclic ring can be formed in which the atom directly linked to the nitrogen is an sp ³ -hybridized carbon atom. In other words, the 3- to 10-membered heterocyclic ring formed by R ³ /-R ^3a together with the nitrogen atom to which they are attached has the following structure:

wherein the dashed line indicates attachment to the remainder of -L ¹ -; the ring contains 3-10 atoms and contains at least one nitrogen atom; and R ^# and R ^## represent sp ³ -hybridized carbon atoms.

It should also be understood that the 3- to 10-membered heterocyclic ring may be further substituted.

Exemplary embodiments of suitable 3- to 10-membered heterocyclic rings formed by R ³ /-R ^3a of formula (II) together with the nitrogen atom to which they are attached are as follows:

wherein the dashed line indicates attachment to the rest of the molecule; and -R is selected from: H and C _1-6 alkyl.

部分的氮仍然是伯胺、仲胺或叔胺的一部分，即-R³和-R^3a彼此獨立地為H，或通過sp³-雜化碳原子與-N<連接。 -L ¹ - in formula (II) may be optionally further substituted. Generally, any substituent may be used as long as it does not affect the cleavage principle, i.e. the hydrogen marked with an asterisk in formula (II) is not substituted, and formula (II)

Part of the nitrogen is still part of a primary, secondary or tertiary amine, ie -R ³ and -R ^3a are independently H or connected to -N< via an sp ³ -hybridized carbon atom.

In certain embodiments, -R ¹ or -R ^1a of formula (II) is substituted with -L ² -Z. In certain embodiments, -R ² or -R ^2a of formula (II) is substituted with -L ² -Z. In certain embodiments, -R ³ or -R ^3a of formula (II) is substituted with -L ² -Z. In certain embodiments, -R ⁴ of formula (II) is substituted with -L ² -Z. In certain embodiments, -R ⁵ or -R ^5a of formula (II) is substituted with -L ² -Z. In certain embodiments, -R ⁶ of formula (II) is substituted with -L ² -Z. In certain embodiments, -R ⁷ or -R ^7a of formula (II) is substituted with -L ² -Z. In certain embodiments, -R ⁸ or -R ^8a of formula (II) is substituted with -L ² -Z. In certain embodiments, -R ⁹ or -R ^9a of formula (II) is substituted with -L ² -Z. In certain embodiments, -R ¹⁰ of formula (II) is substituted with -L ² -Z. In certain embodiments, -R ¹¹ of formula (II) is substituted with -L ² -Z.

In certain embodiments, X of formula (II) is selected from: -C(R ⁴ R ^4a )-, -N(R ⁴ )-, and -C(R ⁷ R ^7a )-.

In certain embodiments, X of formula (II) is -C(R ⁴ R ^4a )-.

In certain embodiments, X of formula (II) is -C(R ⁷ R ^7a )-.

In certain embodiments, -R ⁷ of formula (II) is -NR ¹⁰ -(C=O)-R ¹¹ .

In certain embodiments, ^-R7a of formula (II) is selected from: -H, methyl and ethyl. In certain embodiments, ^-R7a of formula (II) is -H.

In certain embodiments, -R ¹⁰ of formula (II) is selected from: -H, methyl and ethyl. In certain embodiments, -R ¹⁰ of formula (II) is methyl.

In certain embodiments, -R ¹¹ of formula (II) is selected from: -H, methyl and ethyl. In certain embodiments, -R ¹¹ of formula (II) is -H.

In certain embodiments, -R ¹¹ of formula (II) is substituted with -L ² -Z.

In certain embodiments, -X- of formula (II) is -N(R ⁴ )-.

In certain embodiments, -R ⁴ of formula (II) is selected from: -H, methyl, and ethyl.

In certain embodiments, -R ⁴ of formula (II) is -H.

In certain embodiments, ^X1 of formula (II) is C.

In certain embodiments, =X ³ of formula (II) is =0.

In certain embodiments, -X ² - of formula (II) is -C(R ⁸ R ^8a )-.

In certain embodiments, -R ⁸ and -R ^8a of formula (II) are independently selected from: -H, methyl and ethyl. In certain embodiments, at least one of -R ⁸ and -R ^8a of formula (II) is -H. In certain embodiments, both -R ⁸ and -R ^8a of formula (II) are -H.

In certain embodiments, -R ¹ and -R ^1a of formula (II) are independently selected from: -H, methyl and ethyl.

In certain embodiments, at least one of -R ¹ and -R ^1a of formula (II) is -H.

In certain embodiments, -R ¹ and -R ^1a of formula (II) are both -H.

In certain embodiments, at least one of -R ¹ and -R ^1a of formula (II) is methyl.

In certain embodiments, -R ¹ and -R ^1a of formula (II) are both methyl.

In certain embodiments, -R ² and -R ^2a of formula (II) are independently selected from: -H, methyl and ethyl. In certain embodiments, at least one of -R ² and -R ^2a of formula (II) is -H. In certain embodiments, -R ² and -R ^2a of formula (II) are both -H.

In certain embodiments, -R ³ and -R ^3a of formula (II) are independently selected from: -H, methyl, ethyl, propyl and butyl.

In certain embodiments, at least one of -R ³ and -R ^3a of formula (II) is methyl.

In certain embodiments, -R ³ of formula (II) is methyl, and -R ^3a of formula (II) is -H.

In certain embodiments, -R ³ and -R ^3a of formula (II) are both -H.

In certain embodiments, -D is linked to -L ¹ - through the nitrogen by forming an amide bond.

其中虛線表示通過形成醯胺鍵與作為PTH部分的-D的氮連接。 In certain embodiments, the -L ¹ - moiety has the structure of Formula (IIa-i):

The dotted line indicates the connection to the nitrogen of -D as the PTH part through the formation of an amide bond.

As defined in formula (II), using ^-R1 , ^-R1a , ^-R2 , ^-R2a , ^-R3 , ^-R3a , ^-R7 , ^-R7a and ^-X2- ; and wherein ^-L1- is substituted with ^-L2 -Z, and wherein ^-L1- is optionally further substituted, with the proviso that the hydrogen marked with an asterisk in formula (IIa-i) is not substituted with -L2 ^- Z or a substituent.

In certain embodiments, -L ¹ - of formula (IIa-i) is substituted with one -L ² -Z moiety.

In certain embodiments, the -L ¹ - moiety of formula (IIa-i) is not further substituted.

In certain embodiments, -R ¹ and -R ^1a of formula (IIa-i) are independently selected from: -H, methyl and ethyl. In certain embodiments, at least one of -R ¹ and -R ^1a of formula (IIa-i) is -H. In certain embodiments, -R ¹ and -R ^1a of formula (IIa-i) are both -H. In certain embodiments, -R ⁷ of formula (IIa-i) is -NR ¹⁰ -(C=O)-R ¹¹ .

In certain embodiments, R ^7a of formula (IIa-i) is selected from: -H, methyl and ethyl. In certain embodiments, R ^7a of formula (IIa-i) is -H.

In certain embodiments, -R ¹⁰ of formula (IIa-i) is selected from: -H, methyl and ethyl. In certain embodiments, -R ¹⁰ of formula (IIa-i) is methyl.

In certain embodiments, -R ¹¹ of formula (IIa-i) is selected from: -H, methyl and ethyl. In certain embodiments, -R ¹¹ of formula (IIa-i) is -H.

In certain embodiments, -R ¹¹ of formula (IIa-i) is substituted with -L ² -Z.

In certain embodiments, -X ² - of formula (IIa-i) is -C(R ⁸ R ^8a )-.

In certain embodiments, -R ⁸ and -R ^8a of formula (IIa-i) are independently selected from: -H, methyl and ethyl. In certain embodiments, at least one of -R ⁸ or -R ^8a of formula (IIa-i) is -H. In certain embodiments, -R ⁸ and -R ^8a of formula (IIa-i) are both -H.

In certain embodiments, -R ² and -R ^2a of formula (IIa-i) are independently selected from: -H, methyl and ethyl. In certain embodiments, at least one of -R ² or -R ^2a of formula (IIa-i) is -H. In certain embodiments, -R ² and -R ^2a of formula (IIa-i) are both -H.

In certain embodiments, -R ³ and -R ^3a of formula (IIa-i) are independently selected from: -H, methyl, ethyl, propyl and butyl. In certain embodiments, at least one of -R ³ or -R ^3a of formula (IIa-i) is methyl.

In certain embodiments, -R ³ of formula (IIa-i) is -H, and -R ^3a of formula (IIa-i) is methyl.

其中虛線表示通過形成醯胺鍵與作為PTH部分的-D的氮連接；如式(II)所定義，使用-R³、-R^3a、-R¹⁰、-R¹¹和-X²-；其中-L¹-被-L²-Z取代，並且其中-L¹-任選地被進一步取代，條件是式(IIa-ii)中標有星號的氫不被-L²-Z或取代基取代。 In certain embodiments, the -L ¹ - moiety has the structure of Formula (IIa-ii):

wherein the dashed line indicates linkage to the nitrogen of -D as the PTH moiety via formation of an amide bond; as defined in formula (II), using -R ³ , -R ^3a , -R ¹⁰ , -R ¹¹ and -X ² -; wherein -L ¹ - is substituted with -L ² -Z, and wherein -L ¹ - is optionally further substituted, provided that the hydrogen marked with an asterisk in formula (IIa-ii) is not substituted with -L ² -Z or a substituent.

In certain embodiments, -L ¹ - of formula (IIa-ii) is substituted with one -L ² -Z moiety.

In certain embodiments, the -L ¹ - moiety of formula (IIa-ii) is not further substituted.

In certain embodiments, -X ² - of formula (IIa-ii) is -C(R ⁸ R ^8a )-.

In certain embodiments, -R ⁸ and -R ^8a of formula (IIa-ii) are independently selected from: -H, methyl and ethyl. In certain embodiments, at least one of -R ⁸ or -R ^8a of formula (IIa-ii) is -H. In certain embodiments, -R ⁸ and -R ^8a of formula (IIa-ii) are both -H.

In certain embodiments, -R ³ and -R ^3a of formula (IIa-ii) are independently selected from: -H, methyl, ethyl, propyl and butyl. In certain embodiments, at least one of -R ³ or -R ^3a of formula (IIa-ii) is methyl.

In certain embodiments, -R ³ of formula (IIa-ii) is -H, and -R ^3a of formula (IIa-ii) is methyl.

In certain embodiments, -R ¹⁰ of formula (IIa-i) is selected from: -H, methyl and ethyl. In certain embodiments, -R ¹⁰ of formula (IIa-ii) is methyl.

In certain embodiments, -R ¹¹ of formula (IIa-ii) is selected from: -H, methyl and ethyl. In certain embodiments, -R ¹¹ of formula (IIa-i) is -H.

In certain embodiments, -R ¹¹ of formula (IIa-ii) is substituted with -L ² -Z.

其中虛線表示通過形成醯胺鍵與作為PTH部分的-D的氮連接；標有星號的虛線表示與-L²-連接；如式(II)所定義，使用-R³、-R^3a、-R¹⁰和-X²-；以及其中-L¹-被-進一步取代，條件是式(IIa-ii’)中標有星號的氫不被取代基取代。 In certain embodiments, the -L ¹ - moiety has the structure of Formula (IIa-ii'):

wherein the dashed line indicates linkage to the nitrogen of -D as the PTH moiety via formation of an amide bond; the dashed line marked with an asterisk indicates linkage to -L ² -; as defined in formula (II), using -R ³ , -R ^3a , -R ¹⁰ and -X ² -; and wherein -L ¹ - is further substituted with -, provided that the hydrogen marked with an asterisk in formula (IIa-ii') is not substituted with a substituent.

In certain embodiments, the -L ¹ - moiety of formula (IIa-ii') is not further substituted.

In certain embodiments, -X ² - in formula (IIa-ii') is -C(R ⁸ R ^8a )-.

In certain embodiments, -R ⁸ and -R ^8a of formula (IIa-ii') are independently selected from: -H, methyl and ethyl. In certain embodiments, at least one of -R ⁸ or -R ^8a of formula (IIa-ii') is -H. In certain embodiments, -R ⁸ and -R ^8a of formula (IIa-ii') are both -H.

In certain embodiments, -R ³ and -R ^3a of formula (IIa-ii') are independently selected from: -H, methyl, ethyl, propyl and butyl. In certain embodiments, at least one of -R ³ or -R ^3a of formula (IIa-ii') is methyl.

In certain embodiments, -R ³ of formula (IIa-ii') is -H, and -R ^3a of formula (IIa-ii') is methyl.

In certain embodiments, -R ¹⁰ of formula (IIa-i) is selected from: -H, methyl, and ethyl. In certain embodiments, -R ¹⁰ of formula (IIa-ii') is methyl.

其中虛線表示通過形成醯胺鍵與作為PTH部分的-D的氮連接；其中-L¹-被-L²-Z取代，並且其中-L¹-任選地被進一步取代，條件是式(IIa-iii)中標有星號的氫不被-L²-Z或-L²-Z’或取代基取代。 In certain embodiments, the -L ¹ - moiety has the structure of Formula (IIa-iii):

wherein the dashed line indicates linkage to the nitrogen of -D as the PTH moiety via formation of an amide bond; wherein -L ¹ - is substituted by -L ² -Z, and wherein -L ¹ - is optionally further substituted, provided that the hydrogen marked with an asterisk in formula (IIa-iii) is not substituted by -L ² -Z or -L ² -Z' or a substituent.

In certain embodiments, -L ¹ - of formula (IIa-iii) is substituted with one -L ² -Z moiety.

In certain embodiments, the -L ¹ - moiety of formula (IIa-iii) is not further substituted.

其中虛線表示通過形成醯胺鍵與作為PTH部分的-D的氮連接；標有星號的虛線表示與-L²-連接； 其中-L¹-被-進一步取代，條件是式(IIa-iii’)中標有星號的氫不被取代基取代。 In certain embodiments, the -L ¹ - moiety has the structure of Formula (IIa-iii'):

The dashed line indicates the connection to the nitrogen of -D as the PTH part via the formation of an amide bond; the dashed line marked with an asterisk indicates the connection to -L ² -; wherein -L ¹ - is further substituted with -, provided that the hydrogen marked with an asterisk in formula (IIa-iii') is not replaced by a substituent.

In certain embodiments, the -L ¹ - moiety of formula (IIa-iii') is not further substituted.

其中虛線表示通過形成醯胺鍵與作為PTH部分的-D的氮連接；如式(II)所定義，使用-R¹、-R^1a、-R²、-R^2a、-R³、-R^3a、-R⁴和-X²；其中-L¹-被-L²-Z取代，並且其中-L¹-任選地被進一步取代，條件是式(IIb-i)中標有星號的氫不被-L²-Z或取代基取代。 In certain embodiments, the -L ¹ - moiety has the structure of Formula (IIb-i):

wherein the dashed line indicates linkage to the nitrogen of -D as the PTH moiety via formation of an amide bond; as defined in formula (II), using ^-R1 , ^-R1a , ^-R2 , ^-R2a , ^-R3 , ^-R3a , ^-R4 and ^-X2 ; wherein ^-L1- is substituted with ^-L2 -Z, and wherein ^-L1- is optionally further substituted, provided that the hydrogen marked with an asterisk in formula (IIb-i) is not substituted with -L2 ^- Z or a substituent.

In certain embodiments, -L ¹ - of formula (IIb-i) is substituted with one -L ² -Z moiety.

In certain embodiments, the -L ¹ - moiety of formula (IIb-i) is not further substituted.

In certain embodiments, -R ¹ and -R ^1a of formula (IIb-i) are independently selected from: -H, methyl and ethyl. In certain embodiments, at least one of -R ¹ or -R ^1a of formula (IIb-i) is methyl. In certain embodiments, -R ¹ and -R ^1a of formula (IIb-i) are both methyl.

In certain embodiments, -R ⁴ of formula (IIb-i) is selected from: -H, methyl and ethyl. In certain embodiments, -R ⁴ of formula (IIb-i) is -H.

In certain embodiments, -X ² - of formula (IIb-i) is -C(R ⁸ R ^8a )-.

In certain embodiments, -R ⁸ and -R ^8a of formula (IIb-i) are independently selected from: -H, methyl and ethyl. In certain embodiments, at least one of -R ⁸ and -R ^8a of formula (IIb-i) is -H. In certain embodiments, -R ⁸ and -R ^8a of formula (IIb-i) are both -H.

In certain embodiments, -R ² and -R ^2a of formula (IIb-i) are independently selected from: -H, methyl and ethyl. In certain embodiments, at least one of -R ² or -R ^2a of formula (IIb-i) is -H. In certain embodiments, -R ² and -R ^2a of formula (IIb-i) are both -H.

In certain embodiments, -R ³ and -R ^3a of formula (IIb-i) are independently selected from: -H, methyl, ethyl, propyl and butyl. In certain embodiments, at least one of -R ³ or -R ^3a of formula (IIb-i) is -H. In certain embodiments, -R ³ and -R ^3a of formula (IIb-i) are both -H.

其中虛線表示通過形成醯胺鍵與作為PTH部分的-D的氮連接；如式(II)所定義，使用-R²、-R^2a、-R³、-R^3a和-X²；其中-L¹-被-L²-Z取代，並且其中-L¹-任選地被進一步取代，條件是式(IIb-ii)中標有星號的氫不被-L²-Z或取代基取代。 In certain embodiments, the -L ¹ - moiety has the structure of Formula (IIb-ii):

wherein the dashed line indicates linkage to the nitrogen of -D as the PTH moiety via formation of an amide bond; as defined in formula (II), using -R ² , -R ^2a , -R ³ , -R ^3a and -X ² ; wherein -L ¹ - is substituted with -L ² -Z, and wherein -L ¹ - is optionally further substituted, provided that the hydrogen marked with an asterisk in formula (IIb-ii) is not substituted with -L ² -Z or a substituent.

In certain embodiments, -L ¹ - of formula (IIb-ii) is substituted with one -L ² -Z moiety.

In certain embodiments, the -L ¹ - moiety of formula (IIb-ii) is not further substituted.

In certain embodiments, -X ² - of formula (IIb-ii) is -C(R ⁸ R ^8a )-.

In certain embodiments, -R ⁸ and -R ^8a of formula (IIb-i) are independently selected from: -H, methyl and ethyl. In certain embodiments, at least one of -R ⁸ or -R ^8a of formula (IIb-i) is -H. In certain embodiments, -R ⁸ and -R ^8a of formula (IIb-ii) are both -H. In certain embodiments, -R ² and -R ^2a of formula (IIb-ii) are independently selected from: -H, methyl and ethyl. In certain embodiments, at least one of -R ² or -R ^2a of formula (IIb-ii) is -H. In certain embodiments, -R ² and -R ^2a of formula (IIb-ii) are both -H.

In certain embodiments, -R ³ and -R ^3a of formula (IIb-ii) are independently selected from: -H, methyl, ethyl, propyl and butyl. In certain embodiments, at least one of -R ³ or -R ^3a of formula (IIb-ii) is -H. In certain embodiments, -R ³ and -R ^3a of formula (IIb-ii) are both -H.

其中虛線表示通過形成醯胺鍵與作為PTH部分的-D的氮連接；標有星號的虛線表示與-L²-連接；如式(II)所定義，使用-R²、-R^2a、-R^3a和-X²；以及其中-L¹-被-L²-Z取代，並且其中-L¹-任選地被進一步取代，條件是式(IIb-ii’)中標有星號的氫不被-L²-Z或取代基取代。 In certain embodiments, the -L ¹ - moiety has the structure of Formula (IIb-ii'):

wherein the dashed line indicates linkage to the nitrogen of -D as the PTH moiety via formation of an amide bond; the dashed line marked with an asterisk indicates linkage to -L ² -; as defined in formula (II), using -R ² , -R ^2a , -R ^3a and -X ² ; and wherein -L ¹ - is substituted with -L ² -Z, and wherein -L ¹ - is optionally further substituted, provided that the hydrogen marked with an asterisk in formula (IIb-ii') is not substituted with -L ² -Z or a substituent.

In certain embodiments, the -L ¹ - moiety of formula (IIb-ii') is not further substituted.

In certain embodiments, -X ² - of formula (IIb-ii') is -C(R ⁸ R ^8a )-.

In certain embodiments, -R ⁸ and -R ^8a of formula (IIb-ii') are independently selected from: -H, methyl and ethyl. In certain embodiments, at least one of -R ⁸ or -R ^8a of formula (IIb-ii') is -H. In certain embodiments, -R ⁸ and -R ^8a of formula (IIb-ii') are both -H.

In certain embodiments, -R ² and -R ^2a of formula (IIb-ii') are independently selected from: -H, methyl and ethyl. In certain embodiments, at least one of -R ² or -R ^2a of formula (IIb-ii') is -H. In certain embodiments, -R ² and -R ^2a of formula (IIb-ii') are both -H.

In certain embodiments, -R ^3a of formula (IIb-ii') is independently selected from: -H, methyl, ethyl, propyl and butyl. In certain embodiments, -R ^3a of formula (IIb-ii') is -H.

其中虛線表示通過形成醯胺鍵與作為PTH部分的-D的氮連接；其中-L¹-被-L²-Z取代，並且其中-L¹-任選地被進一步取代，條件是式(IIb-ii)中標有星號的氫不被-L²-Z或取代基取代。 In certain embodiments, the -L ¹ - moiety has the structure of Formula (IIb-iii):

wherein the dashed line indicates linkage to the nitrogen of -D as the PTH moiety via amide bond formation; wherein -L ¹ - is substituted by -L ² -Z, and wherein -L ¹ - is optionally further substituted, provided that the hydrogen marked with an asterisk in formula (IIb-ii) is not substituted by -L ² -Z or a substituent.

In certain embodiments, -L ¹ - of formula (IIb-iii) is substituted with one -L ² -Z moiety.

In certain embodiments, the -L ¹ - moiety of formula (IIb-iii) is not further substituted.

其中虛線表示通過形成醯胺鍵與作為PTH部分的-D的氮連接；標有星號的虛線表示與-L²-連接；以及其中-L¹-被-L²-Z取代，並且其中-L¹-任選地被進一步取代，條件是式(IIb-ii’)中標有星號的氫不被-L²-Z或取代基取代。 In certain embodiments, the -L ¹ - moiety has the structure of Formula (IIb-iii'):

wherein the dashed line indicates linkage to the nitrogen of -D as the PTH moiety via formation of an amide bond; the dashed line marked with an asterisk indicates linkage to -L ² -; and wherein -L ¹ - is substituted with -L ² -Z, and wherein -L ¹ - is optionally further substituted, provided that the asterisked hydrogen in formula (IIb-ii') is not substituted with -L ² -Z or a substituent.

In certain embodiments, the -L ¹ - moiety of formula (IIb-iii') is not further substituted.

其中 虛線表示分別通過形成醯胺鍵或酯鍵與作為PTH部分的-D的伯胺或仲胺或羥基連接；-R¹、-R^1a、-R²、-R^2a、-R³和-R^3a獨立地選自：-H、-C(R⁸R^8aR^8b)、-C(=O)R⁸、-C≡N、-C(=NR⁸)R^8a、-CR⁸(=CR^8aR^8b)、-C≡CR⁸和-T；-R⁴、-R⁵和-R^5a彼此獨立地選自：-H、-C(R⁹R^9aR^9b)和-T；a1和a2彼此獨立為0或1；每個-R⁶、-R^6a、-R⁷、-R^7a、-R⁸、-R^8a、-R^8b、-R⁹、-R^9a、-R^9b彼此獨立地選自：-H、鹵素、-CN、-COOR¹⁰、-OR¹⁰、-C(O)R¹⁰、-C(O)N(R¹⁰R^10a)、-S(O)₂N(R¹⁰R^10a)、-S(O)N(R¹⁰R^10a)、-S(O)₂R¹⁰、-S(O)R¹⁰、-N(R¹⁰)S(O)₂N(R^10aR^10b)、-SR¹⁰、-N(R¹⁰R^10a)、-NO₂、-OC(O)R¹⁰、-N(R¹⁰)C(O)R^10a、-N(R¹⁰)S(O)₂R^10a、-N(R¹⁰)S(O)R^10a、-N(R¹⁰)C(O)OR^10a、-N(R¹⁰)C(O)N(R^10aR^10b)、-OC(O)N(R¹⁰R^10a)、-T、C_1-20烷基、C_2-20烯基和C_2-20炔基；其中T、C_1-20烷基、C_2-20烯基和C_2-20炔基任選地被一個或多個相同或不同的-R¹¹取代，並且其中C_1-20烷基、C_2-20烯基和C_2-20炔基任選地被一個或多個選自以下的基團間斷：-T-、-C(O)O-、-O-、-C(O)-、-C(O)N(R¹²)-、-S(O)₂N(R¹²)-、-S(O)N(R¹²)-、-S(O)₂-、-S(O)-、-N(R¹²)S(O)₂N(R^12a)-、-S-、-N(R¹²)-、-OC(OR¹²)(R^12a)-、-N(R¹²)C(O)N(R^12a)-和-OC(O)N(R¹²)-；每個-R¹⁰、-R^10a、-R^10b獨立地選自：-H、-T、C_1-20烷基、C_2-20烯基和C_2-20炔基；其中T、C_1-20烷基、C_2-20烯基和C_2-20炔基任選地被一個或多個相同或不同的R¹¹取代，並且其中C_1-20烷基、C_2-20烯基和C_2-20炔基任選地被一個或多個選自以下的基團間斷：-T-、-C(O)O-、-O-、-C(O)-、-C(O)N(R¹²)-、-S(O)₂N(R¹²)-、-S(O)N(R¹²)-、-S(O)₂-、-S(O)-、-N(R¹²)S(O)₂N(R^12a)-、-S-、-N(R¹²)-、-OC(OR¹²)(R^12a)-、-N(R¹²)C(O)N(R^12a)-和-OC(O)N(R¹²)-； 每個T彼此獨立地選自：苯基、萘基、茚基、茚滿基、四氫萘基、C_3-10環烷基、3至10元雜環基和8至11元雜二環基；其中每個T獨立地任選被一個或多個相同或不同的-R¹¹取代；每個-R¹¹彼此獨立地選自：鹵素、-CN、氧代(=O)、-COOR¹³、-OR¹³、-C(O)R¹³、-C(O)N(R¹³R^13a)、-S(O)₂N(R¹³R^13a)、-S(O)N(R¹³R^13a)、-S(O)₂R¹³、-S(O)R¹³、-N(R¹³)S(O)₂N(R^13aR^13b)、-SR¹³、-N(R¹³R^13a)、-NO₂、-OC(O)R¹³、-N(R¹³)C(O)R^13a、-N(R¹³)S(O)₂R^13a、-N(R¹³)S(O)R^13a、-N(R¹³)C(O)OR^13a、-N(R¹³)C(O)N(R^13aR^13b)、-OC(O)N(R¹³R^13a)和C_1-6烷基；其中C_1-6烷基任選地被一個或多個相同或不同的鹵素取代；每個-R¹²、-R^12a、-R¹³、-R^13a、-R^13b獨立地選自：-H和C_1-6烷基。其中C_1-6烷基任選地被一個或多個相同或不同的鹵素取代；任選地，-R¹/-R^1a、-R²/-R^2a、-R³/-R^3a、-R⁶/-R^6a、-R⁷/-R^7a對中的一個或多個與它們所連接的原子連接在一起，形成C_3-10環烷基或3-10元雜環基；任選地，-R¹/-R²、-R¹/-R³、-R¹/-R⁴、-R¹/-R⁵、-R¹/-R⁶、-R¹/-R⁷、-R²/-R³、-R²/-R⁴、-R²/-R⁵、-R²/-R⁶、-R²/-R⁷、-R³/-R⁴、-R³/-R⁵、-R³/-R⁶、-R³/-R⁷、-R⁴/-R⁵、-R⁴/-R⁶、-R⁴/-R⁷、-R⁵/-R⁶、-R⁵/-R⁷、-R⁶/-R⁷對中的一個或多個與它們所連接的原子連接在一起形成環A；A選自：苯基、萘基、茚基、茚滿基、四氫萘基、C_3-10環烷基、3至10元雜環基和8至11元雜雙環基；其中-L¹-被-L²-Z取代，並且其中-L¹-任選地被進一步取代；其中-L²-是單化學鍵或間隔基團；和-Z是水溶性載體。 Another -L ¹ -part is disclosed in WO2016020373A1, which is incorporated herein by reference in its entirety. Thus, in certain embodiments, the -L ¹ -part has a structure of formula (III):

wherein the dotted line indicates connection with the primary or secondary amine or hydroxyl group of -D as the PTH part by forming an amide bond or an ester bond, respectively; -R ¹ , -R ^1a , -R ² , -R ^2a , -R ³ and -R ^3a are independently selected from: -H, -C(R ⁸ R ^8a R ^8b ), -C(=O)R ⁸ , -C≡N, -C(=NR ⁸ )R ^8a , -CR ⁸ (=CR ^8a R ^8b ), -C≡CR ⁸ and -T; -R ⁴ , -R ⁵ and -R ^5a are independently selected from: -H, -C(R ⁹ R ^9a R ^9b ) and -T; a1 and a2 are independently 0 or 1; each of -R ⁶ , -R ^6a , -R ^{7 10a} ), -S( ^O)2R10 , -S( ^O)R10 , -N ^(R10) S( ^{O )2N(} R10R10a), -SR10, ^-N (R10R10a ⁾ , ^-NO2 , -OC(O) ^R10 , -N(R10R10a), -S(O) ^2R10 , -S(O) _R10 , -N( ^R10 )S(O)2N( ^R10aR10b ), ^{-SR10 ,} -N( ^R10R10a ) ^, -NO2, -OC(O) _R10 , -N(R10R10a), -S(O) ^2R10 , -S( ^O ) _R10 , -N( ^R10 )S(O) ^2N ( ^R10aR10b ), ^-SR10 , _-N ( ^R10R10a ), -NO2, -OC(O) ^R10 , -N( ^R10 )C(O)R ^10a , -N(R ¹⁰ )S(O) ₂ R ^10a , -N(R ¹⁰ )S(O)R ^10a , -N(R ¹⁰ )C(O)OR ^10a , -N(R ¹⁰ )C(O)N(R 10a R 10b ), -OC(O)N(R ¹⁰ R ^10a ), -T, C _1-20 alkyl, C 2-20 alkenyl, and C _2-20 alkynyl; wherein T, C _1-20 alkyl, C 2-20 alkenyl, and C _2-20 alkynyl are optionally substituted by one or more identical or ^different -R ¹¹ , and wherein C 1-20 alkyl, C _2-20 alkenyl, and C _2-20 alkynyl are optionally substituted by one or more identical or different -R ¹¹ , and wherein C _1-20 alkyl, C _2-20 alkenyl, and C _2-20 The alkynyl group is optionally interrupted by one or more groups selected from the group consisting of -T-, -C(O)O-, -O-, -C(O)-, -C(O)N(R ¹² )-, -S(O) ₂ N(R ¹² )-, -S(O)N(R ¹² )-, -S(O) ₂ -, -S(O)-, -N(R ¹² )S(O) ₂ N(R ^12a )-, -S-, -N(R ¹² )-, -OC(OR ¹² )(R ^12a )-, -N(R ¹² )C(O)N(R ^12a )-, and -OC(O)N(R ¹² )-; each of -R ¹⁰ , -R ^10a , -R ^10b is independently selected from: -H, -T, C _1-20 alkyl, C _2-20 alkenyl and C _2-20 alkynyl; wherein T, C _1-20 alkyl, C _2-20 alkenyl and C _2-20 alkynyl are optionally substituted by one or more identical or different R ¹¹ , and wherein C _1-20 alkyl, C _2-20 alkenyl and C _2-20 alkynyl are optionally interrupted by one or more groups selected from the following: -T-, -C(O)O-, -O-, -C(O)-, -C(O)N(R ¹² )-, -S(O) ₂ N(R ¹² )-, -S(O)N(R ¹² )-, -S(O) ₂ -, -S(O)-, -N(R ¹² )S(O) ₂ N(R ^12a wherein the present invention comprises: -S-, -N(R ¹² )-, -OC(OR ¹² )(R ^12a )-, -N(R ¹² )C(O)N(R ^12a )- and -OC(O)N(R ¹² )-; each T is independently selected from phenyl, naphthyl, indenyl, indanyl, tetrahydronaphthyl, C _3-10 cycloalkyl, 3 to 10 membered heterocyclic group and 8 to 11 membered heterobicyclic group; wherein each T is independently optionally substituted with one or more identical or different -R ¹¹ ; each -R ¹¹ is independently selected from halogen, -CN, oxo (=O), -COOR ¹³ , -OR ¹³ , -C(O)R ¹³ , -C(O)N(R ^{13R 13a} ), -S(O) ₂ N(R ¹³ R ^13a ), -S(O)N(R ¹³ R ^13a ), -S(O) ₂ R ¹³ , -S(O)R ¹³ , -N(R ¹³ )S(O) ₂ N(R ^13a R ^13b ), -SR ¹³ , -N(R ¹³ R ^13a ), -NO ₂ , -OC(O)R ¹³ , -N(R ¹³ )C(O)R ^13a , -N(R ¹³ )S(O) ₂ R ^13a , -N(R ¹³ )S(O)R ^13a , -N(R ¹³ )C(O)OR ^13a , -N(R ¹³ )C(O)N(R ^13a R ^13b ), -OC(O)N(R ¹³ R ^13a ) and C _1-6 alkyl; wherein the C _1-6 alkyl is optionally substituted with one or more halogens which are the same or different; each -R ¹² , -R ^12a , -R ¹³ , -R ^13a , -R ^13b is independently selected from: -H and C _1-6 alkyl. wherein the C _1-6 alkyl is optionally substituted by one or more identical or different halogens; optionally, one or more of the pairs -R ¹ / -R ^1a , -R ² / -R ^2a , -R ³ / -R ^3a , -R ⁶ / -R ^6a , -R ⁷ / -R ^7a are linked together with the atoms to which they are linked to form a C _3-10 cycloalkyl or a 3-10 membered heterocyclic group; optionally, -R ¹ / -R ² , -R ¹ / -R ³ , -R ¹ / -R ⁴ , -R ¹ / -R ⁵ , -R ¹ / -R ⁶ , -R ¹ / -R ⁷ , -R ² / -R ³ , -R ² / -R ⁴ , -R ² / -R ⁵ , -R ² / -R ⁶ -R ² /-R ⁷ , -R ³ /-R ⁴ , -R ³ /-R ⁵ , -R ³ /-R ⁶ , -R ³ /-R ⁷ , -R ⁴ /-R ⁵ , -R ⁴ /-R ⁶ , -R ⁴ /-R ⁷ , -R ⁵ /-R ⁶ , -R ⁵ /-R ⁷ , -R ⁶ /-R ⁷ are linked together with the atoms to which they are linked to form ring A; A is selected from: phenyl, naphthyl, indenyl, indanyl, tetrahydronaphthyl, C _3-10 cycloalkyl, 3 to 10 membered heterocyclic group and 8 to 11 membered heterobicyclic group; wherein -L ¹ - is substituted by -L 2 -Z, and wherein -L 1 - is substituted by -L ² -Z, and wherein -L ¹ - is optionally further substituted; wherein -L ² - is a single chemical bond or a spacer group; and -Z is a water-soluble carrier.

In certain embodiments, the optional other substituents of -L ¹ - in formula (III) are as described above.

In certain embodiments, -L ¹ - of formula (III) is substituted with one -L ² -Z moiety.

In certain embodiments, -L ¹ - of formula (III) is not further substituted.

Other embodiments for -L ¹ - are disclosed in EP1536334B1, WO2009/009712A1, WO2008/034122A1, WO2009/143412A2, WO2011/082368A2 and US8618124B2, the entire contents of which are incorporated herein by reference.

其中虛線表示與作為PTH部分的-D連接，其中通過選自OH、SH和NH₂的-D的官能團連接。 Other embodiments for -L ¹ - are disclosed in US8946405B2 and US8754190B2, the entire contents of which are incorporated herein by reference. Therefore, the -L ¹ - moiety has a structure of formula (IV):

Wherein the dotted line indicates the connection to -D as the PTH part, wherein the connection is through a functional group of -D selected from OH, SH and _NH2 .

m is 0 or 1; at least one or both of -R ¹ and -R ² are independently selected from: -CN, -NO ₂ , optionally substituted aryl, optionally substituted heteroaryl, optionally substituted alkenyl, optionally substituted alkynyl, -C(O)R ³ , -S(O)R ³ , -S(O) ₂ R ³ and -SR ⁴ ; one or only one of -R ¹ and -R ² is selected from: -H, optionally substituted alkyl, optionally substituted arylalkyl and optionally substituted heteroarylalkyl; -R ³ is selected from: -H, optionally substituted alkyl, optionally substituted aryl, optionally substituted arylalkyl, optionally substituted heteroaryl, optionally substituted heteroarylalkyl, -OR ⁹ and -N(R ⁹ ) ₂ ; -R ^-R4 is selected from: optionally substituted alkyl, optionally substituted aryl, optionally substituted arylalkyl, optionally substituted heteroaryl and optionally substituted heteroarylalkyl; each ^-R5 is independently selected from: -H, optionally substituted alkyl, optionally substituted alkenylalkyl, optionally substituted alkynylalkyl, optionally substituted aryl, optionally substituted arylalkyl, optionally substituted heteroaryl and optionally substituted heteroarylalkyl; ^-R9 is selected from: -H and optionally substituted alkyl; -Y- is absent, and -X- is O or S; or -Y- is -N(Q)CH2, and -X- is O; Q is selected from: optionally substituted alkyl, optionally substituted aryl, optionally substituted arylalkyl, optionally substituted heteroaryl and optionally substituted heteroarylalkyl; optionally, -R1 and -R2 are selected from: -H and optionally substituted alkyl; -Y- is absent, and -X- is O or S; or -Y- is -N( ^Q)CH2 _, and -X- is O; Q is selected from: optionally substituted alkyl, optionally substituted aryl, optionally substituted arylalkyl, optionally substituted heteroaryl and optionally substituted heteroarylalkyl; optionally, wherein -L ¹ - is substituted by -L ² -Z, and wherein -L ¹ - is optionally further substituted; wherein -L ² - is a single chemical bond or a ^spacer group; and ^-Z is a water-soluble carrier.

In the context of formula (IV) only, the terms used have the following meanings: As used herein, the term "alkyl" includes straight chain, branched chain or cyclic saturated alkyl groups of 1 to 8 carbon atoms, or in some embodiments, 1 to 6, or 1 to 4 carbon atoms.

The term "alkoxy" includes alkyl groups bonded to oxygen, including methoxy, ethoxy, isopropoxy, cyclopropoxy, cyclobutoxy, etc.

The term "alkenyl" includes non-aromatic unsaturated hydrocarbons having a carbon-carbon double bond.

The term "alkynyl" includes non-aromatic unsaturated hydrocarbons having a carbon-carbon triple bond.

The term "aryl" includes aromatic hydrocarbon groups of 6 to 18 carbons, and in certain embodiments 6 to 10 carbons, including groups such as phenyl, naphthyl, and anthracenyl. The term "heteroaryl" includes aromatic rings containing 3 to 15 carbons and at least one N, O, or S atom, preferably 3 to 7 carbons and at least one N, O, or S atom, including groups such as pyrrolyl, pyridyl, pyrimidyl, imidazolyl, oxazolyl, isoxazolyl, thiazolyl, isothiazolyl, quinolyl, indolyl, indenyl, etc.

In some cases, the alkenyl, alkynyl, aryl or heteroaryl moiety may be attached to the rest of the molecule through an alkylene bond. In those cases, the substituent will be referred to as alkenylalkyl, alkynylalkyl, arylalkyl or heteroarylalkyl, indicating that the alkylene moiety is between the alkenyl, alkynyl, aryl or heteroaryl moiety and the molecule to which the alkenyl, alkynyl, aryl or heteroaryl is attached.

The term "halogen" includes bromine, fluorine, chlorine and iodine.

The term "heterocyclic" refers to a 4- to 8-membered aromatic or non-aromatic ring containing 3 to 7 carbon atoms and at least one N, O or S atom. Examples are piperidinyl, piperazinyl, tetrahydropyranyl, pyrrolidine and tetrahydrofuranyl, as well as the exemplary groups provided above for the term "heteroaryl".

When the ring system is optionally substituted, suitable substituents are selected from: alkyl, alkenyl, alkynyl and additional rings, each optionally further substituted. Optional substituents on any group including the above groups include: halogen, nitro, cyano, -OR, -SR, _-NR2 , -OCOR, -NRCOR, -COOR, _-CONR2 , -SOR, _-SO2R , _-SONR2 , _-SO2NR2 , wherein each R is independently alkyl, alkenyl, alkynyl, aryl or heteroaryl, or two R groups together with the atoms to which they are attached _form a ring.

In certain embodiments, -L ¹ - of formula (IV) is substituted with one -L ² -Z moiety.

其中虛線表示與作為PTH部分的-D連接，其中通過-D的胺官能團連接。 Another embodiment for -L ¹ - is disclosed in WO 2013/036857 A1, which is incorporated herein by reference in its entirety. Thus, the -L ¹ - moiety has a structure of formula (V):

The dotted line indicates the connection to -D as the PTH part, wherein the connection is through the amine functional group of -D.

^-R1 is selected from: optionally substituted _C1 - _C6 linear, branched or cyclic alkyl; optionally substituted aryl; optionally substituted heteroaryl; _alkoxy ; and ^-NR52 ; ^-R2 is selected from: -H; optionally substituted _C1 - _C6 alkyl; optionally substituted aryl; optionally substituted heteroaryl; ^-R3 is selected from: -H; optionally substituted _C1 -C6 alkyl; optionally substituted aryl; optionally substituted heteroaryl; ^-R4 is selected from: -H; optionally substituted _C1 - _C6 alkyl; optionally substituted aryl; optionally substituted heteroaryl; each ^-R5 is independently selected from: -H; optionally substituted _C1 - _C6 alkyl; optionally substituted aryl; optionally substituted heteroaryl; or _two -R wherein -L ¹ - is substituted with -L ² -Z, and wherein -L ¹ - is optionally further substituted; wherein -L ² - is a single chemical bond or a spacer group; and -Z is ^a water-soluble carrier.

In the context of formula (V) only, the terms used have the following meanings: "alkyl", "alkenyl" and "alkynyl" include straight chain, branched or cyclic hydrocarbon groups of 1 to 8 carbons, or 1 to 6 carbons, or 1 to 4 carbons, wherein alkyl is a saturated hydrocarbon, alkenyl includes one or more carbon-carbon double bonds, and alkynyl includes one or more carbon-carbon triple bonds. Unless otherwise specified, they contain 1 to 6 carbons.

"Aryl" includes aromatic hydrocarbon groups of 6 to 18 carbons, preferably 6 to 10 carbons, including groups such as phenyl, naphthyl and anthracene.

"Heteroaryl" includes aromatic rings containing 3 to 15 carbon atoms and at least one N, O or S atom. In certain embodiments, it contains 3 to 7 carbon atoms and at least one N, O or S atom, including groups such as pyrrolyl, pyridyl, pyrimidyl, imidazolyl, oxazolyl, isoxazolyl, thiazolyl, isothiazolyl, quinolyl, indolyl, indenyl, etc.

The term "substituted" refers to an alkyl, alkenyl, alkynyl, aryl or heteroaryl group containing one or more substituents replacing one or more hydrogen atoms. The substituents can generally be selected from: halogen, including F, Cl, Br and I; lower alkyl, including linear, branched and cyclic; lower halogenated alkyl, including fluoroalkyl, chloroalkyl, bromoalkyl and iodoalkyl; OH; lower alkoxy, including linear, branched and cyclic; SH; lower alkylthio, including linear, branched and cyclic; amine, alkylamine, dialkylamine, silyl (including alkylsilyl, alkoxysilyl and arylsilyl); nitro; cyano; carbonyl; carboxylic acid, carboxylic acid ester, carboxylic acid acyl; Amine, aminocarbonyl; aminoacyl; carbamate; urea; thiocarbamate; thiourea; ketone; sulfonate; sulfonamide; aryl, including phenyl, naphthyl and anthracenyl; heteroaryl, including 5-membered heteroaryl (including pyrrole, imidazole, furan, thiophene, oxazole, thiazole, isoxazole, isothiazole, thiadiazole, triazole, oxadiazole and tetrazole), 6-membered heteroaryl (including pyridine, pyrimidine, pyrazine) and fused heteroaryl (including benzofuran, benzothiophene, benzoxazole, benzimidazole, indole, benzothiazole, benzoisoxazole and benzoisothiazole).

In certain embodiments, -L ¹ - of formula (V) is substituted with one -L ² -Z moiety.

其中虛線表示與作為PTH部分的-D連接，其中通過-D的胺官能團連接。 Another embodiment of -L ¹ - is disclosed in US7585837B2, which is incorporated herein by reference in its entirety. Therefore, the -L ¹ - moiety has the structure of formula (VI):

The dotted line indicates the connection to -D as the PTH part, wherein the connection is through the amine functional group of -D.

^R1 and ^R2 are independently selected from: hydrogen, alkyl, alkoxy, alkoxyalkyl, aryl, alkaryl, aralkyl, halogen, nitro, _-SO3H , _-SO2NHR5 , amine, ammonium, _carboxyl , _PO3H2 and _OPO3H2 ; ^R3 , ^{R4 and R5} are independently selected from: hydrogen, alkyl and _aryl ; wherein ^-L1- is substituted by ^-L2 -Z, and wherein ^-L1- is optionally further substituted; wherein ^-L2- is a single chemical bond or a spacer group; and -Z is a water-soluble carrier.

Suitable substituents of formula (VI) are: alkyl (e.g. C _1-6 alkyl), alkenyl (e.g. C _2-6 alkenyl), alkynyl (e.g. C _2-6 alkynyl), aryl (e.g. phenyl), heteroalkyl, heteroalkenyl, heteroalkynyl, heteroaryl (e.g. aromatic 4 to 7 membered heterocyclic ring) or halogen moiety.

In the context of formula (VI) only, the terms used have the following meanings: The terms "alkyl", "alkoxy", "alkoxyalkyl", "aryl", "alkaryl" and "aralkyl" refer to alkyl groups of 1 to 8 carbon atoms, in certain embodiments alkyl groups of 1 to 4 carbon atoms, such as methyl, ethyl, propyl, isopropyl and butyl, and aryl groups of 6 to 10 carbon atoms, such as phenyl and naphthyl. The term "halogen" includes bromine, fluorine, chlorine and iodine.

In certain embodiments, -L ¹ - of formula (VI) is substituted with one -L ² -Z moiety.

其中虛線表示與作為PTH部分的-D的連接，其中通過-D的胺官能團連接。 Another embodiment of -L ¹ - is disclosed in WO 2002/089789 A1, which is incorporated herein by reference in its entirety. Therefore, in certain embodiments, -L ¹ - has a structure of formula (VII):

The dotted line indicates the connection to -D as the PTH part, wherein the connection is through the amine function of -D.

L ₁ is a bifunctional linking group; Y ₁ and Y ₂ are independently O, S or NR ⁷ ; R ² , R ³ , R ⁴ , R ⁵ , R ⁶ and R ⁷ are independently selected from: hydrogen, C _1-6 alkyl, C _3-12 branched alkyl, C _3-8 cycloalkyl, C _1-6 substituted alkyl, C _3-8 substituted cycloalkyl, aryl, substituted aryl, aralkyl, C _1-6 heteroalkyl, substituted C _1-6 heteroalkyl, C _1-6 alkoxy, phenoxy and C _1-6 heteroalkoxy; Ar is a moiety that forms a polysubstituted aromatic hydrocarbon or a polysubstituted heterocyclic group when included in formula (VII); X is a chemical bond or a moiety that is actively transported to target cells, a hydrophobic moiety or a combination thereof, and y is 0 or 1; wherein -L ¹ - is substituted with -L ² -Z, and wherein -L ¹ - is optionally further substituted; wherein -L ² - is a single chemical bond or a spacer group; and -Z is a water soluble carrier.

In the context of formula (VII) only, the terms used have the following meanings: The term "alkyl" is understood to include, for example, straight chain, branched, substituted C _1-12 alkyl, including alkoxy, C _3-8 cycloalkyl or substituted cycloalkyl, etc.

The term "substituted" is understood to include the addition or replacement of one or more atoms contained in a functional group or compound with one or more different atoms. Substituted alkyl includes carboxyalkyl, aminoalkyl, dialkylamino, hydroxyalkyl and alkylalkyl; substituted cycloalkyl includes moieties such as 4-chlorocyclohexyl; aryl includes moieties such as naphthyl; substituted aryl includes moieties such as 3-bromo-phenyl; aralkyl includes moieties such as toluyl; heteroalkyl includes moieties such as ethylthiophene; substituted heteroalkyl includes moieties such as 3-methoxythiophene; alkoxy includes moieties such as methoxy; phenoxy includes moieties such as 3-nitrophenoxy. Halogen is understood to include fluorine, chlorine, iodine and bromine.

In certain embodiments, -L ¹ - of formula (VII) is substituted with one -L ² -Z moiety.

其中標有星號的虛線表示通過形成醯胺鍵與作為PTH部分的-D的氮連接。 In certain embodiments, -L ¹ - comprises a substructure of formula (VIII):

The dashed line marked with an asterisk indicates the connection to the nitrogen of -D as the PTH part through the formation of an amide bond.

The unlabeled dashed line indicates linkage to the remainder of -L ¹ -; and wherein -L ¹ - is substituted with -L ² -Z, and wherein -L ¹ - is optionally further substituted; wherein -L ² - is a single chemical bond or a spacer group; and -Z is a water soluble carrier.

In certain embodiments, -L ¹ - of formula (VIII) is substituted with one -L ² -Z moiety.

In certain embodiments, -L ¹ - of Formula (VIII) is not further substituted.

其中標有星號的虛線表示通過形成胺基甲酸酯鍵與作為PTH部分的-D的氮連接；未標記的虛線表示與-L¹-的其餘部分連接；和其中-L¹-被-L²-Z取代，並且其中-L¹-任選地被進一步取代；其中-L²-是單化學鍵或間隔基團；和-Z是水溶性載體。 In certain embodiments, -L ¹ - comprises a substructure of formula (IX):

wherein the dashed line marked with an asterisk indicates attachment to the nitrogen of -D as the PTH moiety via formation of a carbamate bond; the unmarked dashed line indicates attachment to the remainder of -L ¹ -; and wherein -L ¹ - is substituted with -L ² -Z, and wherein -L ¹ - is optionally further substituted; wherein -L ² - is a single chemical bond or a spacer group; and -Z is a water soluble carrier.

In certain embodiments, -L ¹ - of formula (IX) is substituted with one -L ² -Z moiety.

In certain embodiments, -L ¹ - of Formula (IX) is not further substituted.

In certain embodiments, -L ² - is a chemical bond or a spacer group moiety.

In certain embodiments, -L ² - is a chemical bond.

In certain embodiments, -L ² - is a spacer group moiety.

When ^-L2- is not a single chemical bond, ^-L2- is selected from: -T-, -C(O)O-, -O-, -C(O)-, -C(O)N( ^Ry1 )-, -S(O) _2N ( ^Ry1 )-, -S(O)N( ^Ry1 )-, -S(O) _2- , -S(O)-, -N( ^Ry1 )S(O) _2N ( ^Ry1a )-, -S-, -N( ^Ry1 )-, -OC(ORy1)( ^Ry1a )-, -N( ^Ry1 )C(O)N( ^Ry1a )-, -OC(O)N( ^Ry1 )-, _C1-50 alkyl, _C2-50 alkenyl and _C2-50 alkynyl ^; wherein T, C wherein the C _1-50 alkyl, C _2-50 alkenyl and C _2-50 alkynyl are optionally substituted by one or more identical or different -R ^y2 groups, and wherein the C _1-50 alkyl, C _2-50 alkenyl and C _2-50 alkynyl are optionally interrupted by one or more groups selected from the following groups: -T-, -C(O)O-, -O-, -C(O)-, -C(O)N(R ^y3 )-, -S(O) ₂ N(R ^y3 )-, -S(O)N(R ^y3 )-, -S(O) ₂ -, -S(O)-, -N(R ^y3 )S(O) ₂ N(R ^y3a )-, -S-, -N(R ^y3 )-, -OC(OR ^y3 )(R ^y3a )-, -N(R ^y3 )C(O)N(R ^y3a )- and -OC(O)N(R ^y3 )-.

-R ^y1 and -R ^y1a are independently selected from: -H, -T, C _1-50 alkyl, C _2-50 alkenyl and C _2-50 alkynyl; wherein -T, C _1-50 alkyl, C _2-50 alkenyl and C _2-50 alkynyl are optionally substituted by one or more identical or different -R ^y2 , and wherein C _1-50 alkyl, C _2-50 alkenyl and C _2-50 alkynyl are optionally interrupted by one or more groups selected from the following: -T-, -C(O)O-, -O-, -C(O)-, -C(O)N(R ^y4 )-, -S(O) ₂ N(R ^y4 )-, -S(O)N(R ^y4 )-, -S(O) ₂ -, -S(O)-, -N(R ^y4 )S(O) ₂ N(R ^y4a )-, -S-, -N(R ^y4 )-, -OC(OR ^y4 )(R ^y4a )-, -N(R ^y4 )C(O)N(R ^y4a )- and -OC(O)N(R ^y4 )-; each T is independently selected from: phenyl, naphthyl, indenyl, indanyl, tetrahydronaphthyl, C _3-10 cycloalkyl, 3 to 10 membered heterocyclic group and 8 to 11 membered heterobicyclic group, 8 to 30 membered carbon polycyclic group and 8 to 30 membered heteropolycyclic group; wherein each T is independently optionally substituted by one or more identical or different -R ^y2 ; each -R ^y2 is independently selected from: halogen, -CN, oxo (=O), -COOR ^y5 , -OR ^y5 , -C(O)R ^y5 , -C(O)N(R ^y5 R ^y5a ), -S(O) ₂ N(R ^y5 R ^y5a ), -S(O)N(R ^y5 R ^y5a ), -S(O) ₂ R ^y5 , -S(O)R ^y5 , -N(R ^y5 )S(O) ₂ N(R ^y5a R ^y5b ), -SR ^y5 , -N(R ^y5 R ^y5a ), -NO ₂ , -OC(O)R ^y5 , -N(R ^y5 )C(O)R ^y5a , -N(R ^y5 )S(O) ₂ R ^y5a , -N(R ^y5 )S(O)R ^y5a , -N(R ^y5 )C(O)OR ^y5a , -N(R ^y5 )C(O)N(R ^y5a R ^y5b ), -OC(O)N(R ^y5 R ^y5a ) and C _1-6 alkyl; wherein the C _1-6 alkyl is optionally substituted by one or more the same or different halogens; and each -R ^y3 , -R ^y3a , -R ^y4 , -R ^y4a , -R ^y5 , -R ^y5a and -R ^y5b is independently selected from: -H and C _1-6 alkyl, wherein the C _1-6 alkyl is optionally substituted by one or more the same or different halogens.

When ^-L2- is not a single bond, in certain embodiments ^-L2- is selected from: -T-, -C(O)O-, -O-, -C(O)-, -C(O)N( ^Ry1 )-, -S(O) _2N ( ^Ry1 )-, -S(O)N( ^Ry1 )-, -S(O) _2- , -S(O)-, -N(Ry1)S(O) _2N ( ^Ry1a )-, -S-, -N( ^Ry1 )-, -OC( ^ORy1 )( ^Ry1a )-, -N( ^Ry1 )C(O)N( ^Ry1a )-, -OC(O)N( ^Ry1 )-, _C1-20 alkyl, _C2-20 alkenyl and _C2-20 alkynyl ^; wherein T, C wherein the C _1-20 alkyl, C _2-20 alkenyl and C _2-20 alkynyl are optionally substituted by one or more identical or different -R ^y2 groups, and wherein the C _1-20 alkyl, C _2-20 alkenyl and C _2-20 alkynyl are optionally interrupted by one or more groups selected from the following groups: -T-, -C(O)O-, -O-, -C(O)-, -C(O)N(R ^y3 )-, -S(O) ₂ N(R ^y3 )-, -S(O)N(R ^y3 )-, -S(O) ₂ -, -S(O)-, -N(R ^y3 )S(O) ₂ N(R ^y3a )-, -S-, -N(R ^y3 )-, -OC(OR ^y3 )(R ^y3a )-, -N(R ^y3 )C(O)N(R ^y3a )- and -OC(O)N(R ^y3 )-.

-R ^y1 and -R ^y1a are independently selected from: -H, -T, C _1-10 alkyl, C _2-10 alkenyl and C _2-10 alkynyl; wherein -T, C _1-10 alkyl, C _2-10 alkenyl and C _2-10 alkynyl are optionally substituted by one or more identical or different -R ^y2 , and wherein C _1-10 alkyl, C _2-10 alkenyl and C _2-10 alkynyl are optionally interrupted by one or more groups selected from the following: -T-, -C(O)O-, -O-, -C(O)-, -C(O)N(R ^y4 )-, -S(O) ₂ N(R ^y4 )-, -S(O)N(R ^y4 )-, -S(O) ₂ -, -S(O)-, -N(R ^y4 )S(O) ₂ N(R ^y4a )-, -S-, -N(R ^y4 )-, -OC(OR ^y4 )(R ^y4a )-, -N(R ^y4 )C(O)N(R ^y4a )- and -OC(O)N(R ^y4 )-; each T is independently selected from: phenyl, naphthyl, indenyl, indanyl, tetrahydronaphthyl, C _3-10 cycloalkyl, 3 to 10 membered heterocyclic group and 8 to 11 membered heterobicyclic group, 8 to 30 membered carbon polycyclic group and 8 to 30 membered heteropolycyclic group; wherein each T is independently optionally substituted by one or more identical or different -R ^y2 ; -R ^y2 is independently selected from: halogen, -CN, oxo (=O), -COOR ^y5 , -OR ^y5 , -C(O)R ^y5 , -C(O)N(R ^y5 R ^y5a ), -S(O) ₂ N(R ^y5 R ^y5a ), -S(O)N(R ^y5 R ^y5a ), -S(O) ₂ R ^y5 , -S(O)R ^y5 , -N(R ^y5 )S(O) ₂ N(R ^y5a R ^y5b ), -SR ^y5 , -N(R ^y5 R ^y5a ), -NO ₂ , -OC(O)R ^y5 , -N(R ^y5 )C(O)R ^y5a , -N(R ^y5 )S(O) ₂ R ^y5a , -N(R ^y5 )S(O)R ^y5a , -N(R ^y5 )C(O)OR ^y5a ,-N(R wherein the C _1-6 alkyl is optionally substituted by one or more the same or different halogens ^{; and each -R y3 , -R y3a , -R y4} , -R ^y4a , -R ^y5 , -R ^y5a and -R ^y5b is independently selected from: -H and C _1-6 alkyl, wherein _the C _1-6 alkyl is optionally substituted by one or more the same or different halogens ^.

When ^-L2- is not a single chemical bond, ^-L2- is selected from: -T-, -C(O)O-, -O-, -C(O)-, -C(O)N( ^Ry1 )-, -S(O) _2N ( ^Ry1 )-, -S(O)N( ^Ry1 )-, -S(O) _2- , -S(O)-, -N( ^Ry1 )S(O) _2N ( ^Ry1a )-, -S-, -N( ^Ry1 )-, -OC(ORy1)( ^Ry1a )-, -N( ^Ry1 )C(O)N( ^Ry1a )-, -OC(O)N( ^Ry1 )-, _C1-50 alkyl, _C2-50 alkenyl and _C2-50 alkynyl ^; wherein T, C wherein the C _1-50 alkyl, C _2-50 alkenyl and C _2-50 alkynyl are optionally substituted by one or more identical or different -R ^y2 groups, and wherein the C _1-50 alkyl, C _2-50 alkenyl and C _2-50 alkynyl are optionally interrupted by one or more groups selected from the following groups: -T-, -C(O)O-, -O-, -C(O)-, -C(O)N(R ^y3 )-, -S(O) ₂ N(R ^y3 )-, -S(O)N(R ^y3 )-, -S(O) ₂ -, -S(O)-, -N(R ^y3 )S(O) ₂ N(R ^y3a )-, -S-, -N(R ^y3 )-, -OC(OR ^y3 )(R ^y3a )-, -N(R ^y3 )C(O)N(R ^y3a )- and -OC(O)N(R ^y3 )-; -R ^y1 and -R ^y1a are independently selected from: -H, -T, C _1-10 alkyl, C _2-10 alkenyl and C _2-10 alkynyl; each T is independently selected from: phenyl, naphthyl, indenyl, indanyl, tetrahydronaphthyl, C _3-10 cycloalkyl, 3 to 10 membered heterocyclic group and 8 to 11 membered heterobicyclic group, 8 to 30 membered carbon polycyclic group and 8 to 30 membered heteropolycyclic group; each -R ^y2 is independently selected from: halogen and C _1-6 alkyl; and each -R ^y3 , -R ^y3a , -R ^y4 , -R ^y4a , -R ^y5 , -R ^y5a and -R ^y5b are independently selected from: -H and C _1-6 alkyl, wherein the C _1-6 alkyl is optionally substituted with one or more halogens which may be the same or different.

In certain embodiments, -L ² - is a C _1-20 alkyl chain, which is optionally interrupted by one or more groups independently selected from -O-, -T- and -C(O)N(R ^y1 ); and the C _1-20 alkyl chain is optionally substituted by one or more groups independently selected from -OH, -T and -C(O)N(R ^y6 R ^y6a ); wherein -R ^y1 , -R ^y6 , -R ^y6a are independently selected from: H and C _1-4 alkyl, and wherein T is selected from: phenyl, naphthyl, indenyl, indanyl, tetrahydronaphthyl, C The invention also includes _{a 3-10-} membered cycloalkyl group, a 3-10-membered heterocyclic group and an 8-11-membered heterobicyclic group, an 8-30-membered carbon polycyclic group and an 8-30-membered heteropolycyclic group.

In certain embodiments, -L ² - has a chain length of 1 to 20 atoms.

As used herein, the term "chain length" with respect to the -L ² - moiety refers to the number of atoms of -L ² - present in the shortest connection between -L ¹ - and Z.

其中標有星號的虛線表示與-L¹-連接；未標記的虛線表示與-Z的連接；n選自0、1、2、3、4、5、6、7、8、9、10、11、12、13、14、15、16、17和18；和其中式(i)的部分任選地被進一步取代。 In certain embodiments, -L ² - has a structure of formula (i):

wherein a dashed line marked with an asterisk indicates linkage to -L ¹ -; an unmarked dashed line indicates linkage to -Z; n is selected from 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17 and 18; and wherein the moiety of formula (i) is optionally further substituted.

In some embodiments, n of formula (i) is selected from 3, 4, 5, 6, 7, 8 and 9. In some embodiments, n of formula (i) is 4, 5, 6 or 7. In some embodiments, n of formula (i) is 4. In some embodiments, n of formula (i) is 5. In some embodiments, n of formula (i) is 6.

其中未標記的虛線表示通過形成醯胺鍵與作為PTH部分的-D的氮連接；標有星號的虛線表示與-Z的連接。 In certain embodiments, the -L ¹ -L ² - moiety is selected from:

The unmarked dashed line indicates the connection to the nitrogen of -D as the PTH part through the formation of an amide bond; the dashed line marked with an asterisk indicates the connection to -Z.

其中未標記的虛線表示通過形成醯胺鍵與作為PTH部分的-D的氮連接；標有星號的虛線表示與-Z的連接。 In certain embodiments, the -L ¹ -L ² - moiety is selected from:

The unmarked dashed line indicates the connection to the nitrogen of -D as the PTH part through the formation of an amide bond; the dashed line marked with an asterisk indicates the connection to -Z.

In certain embodiments, the -L ¹ -L ² - moiety has the structure of Formula (IIca-ii).

In certain embodiments, the -L ¹ -L ² - moiety has the structure of Formula (IIcb-iii).

In certain embodiments, the PTH conjugate has a structure of formula (Ia), wherein x=1.

The carrier -Z comprises a C _8-24 alkyl group or a polymer. In some embodiments, -Z comprises a polymer. In some embodiments, -Z comprises a polymer selected from the following: 2-methacryl-oxyethylphosphatidylcholine, polyacrylic acid, polyacrylate, polyacrylamide, polyalkoxy polymer, polyamide, polyamidoamine, polyamino acid, polyanhydride, polyaspartamide, polybutyric acid, polyglycolic acid, polybutylene terephthalate, polycaprolactone, polycarbonate, polycyanoacrylate, polydimethacrylamide, polyester, polyethylene, polyethylene glycol, polyethylene oxide, polyethyl phosphate, polyethyl oxazoline, polyglycolic acid, polyhydroxyethyl acrylate, polyhydroxyethyl-oxazoline, polyhydroxymethacrylate, polyhydroxypropylmethacrylamide, polyhydroxypropylmethacrylate, polyhydroxypropyloxazoline, polyhydroxypropyloxazoline, polyhydroxyethyloxazoline, polyhydroxyethyloxazoline, polyhydroxyethyloxazoline, polyhydroxyethyloxazoline, polyhydroxyethyloxazoline, polyhydroxyethyloxazoline, polyhydroxyethyloxazoline, polyhydroxyethyloxazoline, polyhydroxyethyloxazoline, polyhydroxyethyloxazoline, polyhydroxyethyloxazoline, polyhydroxyethyloxazoline, polyhydroxyethyloxazoline, polyhydroxyethyloxazoline, polyhydroxypropyl ... oxazoline, polyimino carbonate, polylactic acid, polylactic acid hydroxyacetic acid copolymer, polymethacrylamide, polymethacrylate, polymethyloxazoline, polyorganophosphazene, polyorthoester, polyoxazoline, polypropylene glycol, polysiloxane, polyurethane, polyvinyl alcohol, polyvinylamine, polyvinyl methyl ether, polyvinyl pyrrolidone, silicone, cellulose, carboxymethyl cellulose, hydroxypropyl methyl cellulose, chitin, chitosan, dextran, dextrin, gelatin, hyaluronic acid and its derivatives, functionalized hyaluronic acid, mannan, pectin, rhamnogalacturonic acid, starch, hydroxyalkyl starch, hydroxyethyl starch and other carbohydrate-based polymers, xylan and its copolymers.

In some embodiments, -Z has a molecular weight of 5-200 kDa. In some embodiments, -Z has a molecular weight of 8 to 100 kDa. In some embodiments, -Z has a molecular weight of 10 to 80 kDa. In some embodiments, -Z has a molecular weight of 12 to 60 kDa. In some embodiments, -Z has a molecular weight of 15 to 40 kDa. In some embodiments, -Z has a molecular weight of about 20 kDa. In some embodiments, -Z has a molecular weight of about 40 kDa.

In some embodiments, such water-soluble carrier-Z comprises a protein. In some embodiments, the protein is selected from the carboxyl terminal polypeptide of human gonadotropin as described in US 2012/0035101A1, which is incorporated herein by reference. Albumin; XTEN sequence as described in WO2011/123813 A2, which is incorporated herein by reference. Proline/alanine random coil sequence as described in WO2011/144756A1, which is incorporated herein by reference. Proline/alanine/serine random coil sequence as described in WO2008/155134A1 and WO2013/024049A1, both of which are incorporated herein by reference; and Fc fusion protein.

In some embodiments, -Z is polysarcosine. In some embodiments, -Z comprises poly-N-methylglycine. In some embodiments, -Z comprises a random coiled protein portion.

In some embodiments, -Z comprises one random curli protein portion. In some embodiments, -Z comprises two random curli protein portions. In some embodiments, -Z comprises three random curli protein portions. In some embodiments, -Z comprises four random curli protein portions. In some embodiments, -Z comprises five random curli protein portions. In some embodiments, -Z comprises six random curli protein portions. In some embodiments, -Z comprises seven random curli protein portions. In some embodiments, -Z comprises eight random curli protein portions.

In some embodiments, the random curly protein portion comprises at least 25 amino acid residues and at most 2000 amino acids. In some embodiments, the random curly protein portion comprises at least 30 amino acid residues and at most 1500 amino acid residues. In some embodiments, the random curly protein portion comprises at least 50 amino acid residues and at most 500 amino acid residues.

In some embodiments, -Z comprises a random curli protein portion, wherein at least 80%, in some embodiments at least 85%, in some embodiments at least 90%, in some embodiments at least 95%, in some embodiments at least 98%, and in some embodiments at least 99% of the total number of amino acids forming the random curli protein portion are selected from alanine and proline. In some embodiments, at least 10% but less than 75%, in some embodiments less than 65% of the total number of amino acid residues in such random curli protein portion are proline residues. In some embodiments, such random curli protein portion is as described in WO2011/144756A1, which is incorporated herein by reference in its entirety.

In certain embodiments, -Z comprises at least one portion selected from the group consisting of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, SEQ ID NO: 10, SEQ ID NO: 11, SEQ ID NO: 12, SEQ ID NO: 13, SEQ ID NO: 14, SEQ ID NO: 15, SEQ ID NO: 16, SEQ ID NO: 17, SEQ ID NO: 51 and SEQ ID NO: 61 disclosed in WO2011/144756, which is incorporated herein by reference. The portion comprising such aranine- and proline-containing random coiled protein is referred to as "PA" or "PA portion".

Therefore, -Z includes the PA part.

In some embodiments, -Z comprises a random curli protein portion, wherein at least 80%, in some embodiments at least 85%, in some embodiments at least 90%, in some embodiments at least 95%, in some embodiments at least 98%, and in some embodiments at least 99% of the total number of amino acids forming the random curli protein portion are selected from alanine, serine and proline. In some embodiments, at least 4% but less than 40% of the total number of amino acid residues in such random curli protein portion are proline residues. In some embodiments, such a random curli protein portion is as described in WO2008/155134A1, which is incorporated herein by reference in its entirety. In certain embodiments, -Z comprises at least one portion selected from SEQ ID NO: 2, SEQ ID NO: 4, SEQ ID NO: 6, SEQ ID NO: 8, SEQ ID NO: 10, SEQ ID NO: 12, SEQ ID NO: 14, SEQ ID NO: 16, SEQ ID NO: 18, SEQ ID NO: 20, SEQ ID NO: 22, SEQ ID NO: 24, SEQ ID NO: 26, SEQ ID NO: 28, SEQ ID NO: 30, SEQ ID NO: 32, SEQ ID NO: 34, SEQ ID NO: 36, SEQ ID NO: 40, SEQ ID NO: 42, SEQ ID NO: 44, SEQ ID NO: 46, SEQ ID NO: 50, SEQ ID NO: 52, SEQ ID NO: 54 and SEQ ID NO: 56 disclosed in WO2008/155134A1, which is incorporated herein by reference. The portion containing this random coiled protein portion containing alanine, serine and proline is called "PAS" or "PAS portion".

Therefore, -Z includes the PAS part.

In some embodiments, -Z comprises a random curli protein portion, wherein at least 80%, in some embodiments at least 85%, in some embodiments at least 90%, in some embodiments at least 95%, in some embodiments at least 98%, and in some embodiments at least 99% of the total number of amino acids forming the random curli protein portion are selected from alanine, glycine, and proline. The portion comprising such a random curli protein portion containing alanine, glycine, and proline is referred to as a "PAG" or a "PAG portion".

Therefore, -Z includes the PAG part.

In some embodiments, -Z comprises a random curli protein portion, wherein at least 80%, in some embodiments at least 85%, in some embodiments at least 90%, in some embodiments at least 95%, in some embodiments at least 98%, and in some embodiments at least 99% of the total number of amino acids forming the random curli protein portion are selected from proline and glycine. The portion comprising such a random curli protein portion containing proline and glycine will be referred to as "PG" or "PG portion".

In certain embodiments, such a PG portion comprises a portion of formula (a-0) [(Gly) _p -Pro-(Gly) _q ] _r (a-0); wherein p is selected from 0, 1, 2, 3, 4 and 5; q is selected from 0, 1, 2, 3, 4 and 5; r is an integer ranging from 10 to 1000 and including 10 and 1000; provided that at least one of p and q is at least 1; in certain embodiments, p of formula (a-0) is selected from 1, 2 and 3.

In certain embodiments, q of formula (a-0) is selected from 0, 1 and 2.

In certain embodiments, the PG portion comprises the sequence of SEQ ID NO: 122: GGPGGPGPGGPGPGGPGPGGPG.

In certain embodiments, the PG portion comprises the sequence of SEQ ID NO:97 of the formula (a-0-a): (GGPGGPGPGGPGGPGPGGPG) _v (a-0-a), wherein v is an integer ranging from 1 to 50 and including 1 and 50.

Therefore, -Z includes the PG part.

In some embodiments, Z comprises a random curli protein portion, wherein at least 80%, in some embodiments at least 85%, in some embodiments at least 90%, in some embodiments at least 95%, in some embodiments at least 98%, and in some embodiments at least 99% of the total number of amino acids forming the random curli protein portion are selected from alanine, glycine, serine, threonine, glutamate and proline. In some embodiments, such a random curli protein portion is as described in WO2010/091122A1, which is incorporated herein by reference. In certain embodiments, -Z comprises at least one moiety selected from the group consisting of SEQ ID NO: 182, SEQ ID NO: 183, SEQ ID NO: 184; SEQ ID NO: 185, SEQ ID NO: 186, SEQ ID NO: 187, SEQ ID NO: 188, SEQ ID NO: 189, SEQ ID NO: 190, SEQ ID NO: 191, SEQ ID NO: 192, SEQ ID NO: 193, SEQ ID NO: 194, SEQ ID NO: 195, SEQ ID NO: 196, SEQ ID NO: 197, SEQ ID NO: 198, SEQ ID NO: 199, SEQ ID NO: 200, SEQ ID NO: 201, SEQ ID NO: 202, SEQ ID NO: 203, SEQ ID NO: 204, SEQ ID NO: 205, SEQ ID NO: 206, SEQ ID NO: 207, SEQ ID NO: 208 NO: 208, SEQ ID NO: 209, SEQ ID NO: 210, SEQ ID NO: 211, SEQ ID NO: 212, SEQ ID NO: 213, SEQ ID NO: 214, SEQ ID NO: 215, SEQ ID NO: 216, SEQ ID NO: 217, SEQ ID NO: 218, SEQ ID NO: 219, SEQ ID NO: 220, SEQ ID NO NO: 221, SEQ ID NO: 759, SEQ ID NO: 760, SEQ ID NO: 761, SEQ ID NO: 762, SEQ ID NO: 763, SEQ ID NO: 764, SEQ ID NO: 765, SEQ ID NO: 766, SEQ ID NO: 767, SEQ ID NO: 768, SEQ ID NO: 769, SEQ ID NO: 770, SEQ ID NO: 771, SEQ ID NO: 772, SEQ ID NO: 773, SEQ ID NO: 774, SEQ ID NO: 775, SEQ ID NO: 776, SEQ ID NO: 777, SEQ ID NO: 778, SEQ ID NO: 779, SEQ ID NO: 1715, SEQ ID NO: 1716, SEQ ID NO: 1718, SEQ ID NO: 1719, SEQ ID NO: 1720, SEQ ID NO: 1721 and SEQ ID NO: 1722, which are incorporated herein by reference. Portions comprising such random coiled protein portions containing alanine, glycine, serine, threonine, glutamate and proline are referred to as "XTEN" or "XTEN portions".

Therefore, -Z contains the XTEN portion.

In some embodiments, -Z comprises a fatty acid derivative. In some embodiments, the fatty acid derivative is those disclosed in WO2005/027978A2 and WO2014/060512A1, both of which are incorporated herein by reference.

In certain embodiments, -Z is a hyaluronic acid-based polymer.

In some embodiments, -Z is a carrier disclosed in WO2012/02047A1, which is incorporated herein by reference. In some embodiments, -Z is a carrier disclosed in WO2013/024048A1, which is incorporated herein by reference.

In certain embodiments, -Z is a PEG-based polymer, such as a linear, branched, or multi-armed PEG-based polymer.

In certain embodiments, -Z is a linear PEG-based polymer.

In some embodiments, -Z is a multi-arm PEG-based polymer. In some embodiments, -Z is a multi-arm PEG-based polymer having at least 4 PEG-based arms.

In certain embodiments, the polymer-Z based on multi-arm PEG is connected to a plurality of -L ² -L ¹ -D parts, wherein in certain embodiments, each -L ² -L ¹ -D part is connected to the end of the arm. In certain embodiments, the polymer-Z based on multi-arm PEG is connected to 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15 or 16 -L ² -L ¹ -D parts. In certain embodiments, the polymer-Z based on multi-arm PEG is connected to 2, 3, 4, 6 or 8 -L ² -L ¹ -D parts. In certain embodiments, the polymer-Z based on multi-arm PEG is connected to 2, 4 or 6 -L ² -L ¹ -D parts. In certain embodiments, such a multi-arm PEG-based polymer-Z is linked to 4 or 6 -L ² -L ¹ -D moieties, and in certain embodiments, such a multi-arm PEG-based polymer-Z is linked to 4 -L ² -L ¹ -D moieties.

In certain embodiments, such multi-arm PEG-based polymer-Z is a multi-arm PEG derivative, for example, as described in detail in the product list of JenKem Technology in the United States (accessible by downloading from http://www.jenkemusa.com/Pages/PEGProducts.aspx on December 18, 2014), such as a 4-arm PEG derivative, specifically a 4-arm PEG derivative containing a pentaerythritol core, an 8-arm PEG derivative containing a hexaglycerol core, and an 8-arm PEG derivative containing a tripentaerythritol core. In certain embodiments, the water-soluble PEG-based carrier-Z comprises a portion selected from the following: a 4-arm PEG amine containing a pentaerythritol core:

n is 20 to 500; 8-arm PEG amine containing a hexaglycerol core:

n is 20 to 500; and R = hexaglycerol or tripentaerythritol core structure; and a 6-arm PEG amine containing a sorbitol or dipentaerythritol core:

n is 20 to 500; and R = a sorbitol or dipentaerythritol core; and wherein the dashed line indicates the attachment to the remainder of the PTH conjugate.

In some embodiments, -Z is a branched PEG-based polymer. In some embodiments, -Z is a branched PEG-based polymer with one, two, three, four, five, or six branch points. In some embodiments, Z is a branched PEG-based polymer with one, two, or three branch points. In some embodiments, Z- is a branched PEG-based polymer with one branch point. In some embodiments, -Z is a branched PEG-based polymer with two branch points. In some embodiments, -Z is a branched PEG-based polymer with three branch points.

In some embodiments, the branch point is selected from -N<, -CH< and >C<.

In certain embodiments, the -Z moiety based on the branched PEG has a molecular weight of at least 10 kDa.

In some embodiments, the molecular weight of this branch-Z portion is from 10kDa to 500kDa, including 10kDa and 500kDa. In some embodiments, the molecular weight of this branch-Z portion is from 10kDa to 250kDa, including 10kDa and 250kDa. In some embodiments, the molecular weight of this branch-Z portion is from 10kDa to 150kDa, including 10kDa and 150kDa. In some embodiments, the molecular weight of this branch-Z portion is from 12kDa to 100kDa, including 12kDa and 100kDa. In some embodiments, the molecular weight of this branch-Z portion is from 15kDa to 80kDa, including 15kDa and 80kDa.

In certain embodiments, the molecular weight of this branch-Z part is 10kDa to 80kDa, and includes 10kDa and 80kDa. In certain embodiments, the molecular weight is about 10kDa. In certain embodiments, the molecular weight of this branch-Z part is about 20kDa. In certain embodiments, the molecular weight of this branch-Z part is about 30kDa. In certain embodiments, the molecular weight of this branch-Z part is about 40kDa. In certain embodiments, the molecular weight of this branch-Z part is about 50kDa. In certain embodiments, the molecular weight of this branch-Z part is about 60kDa. In certain embodiments, the molecular weight of this branch-Z part is about 70kDa. In certain embodiments, the molecular weight of this branch-Z part is about 80kDa. In certain embodiments, the molecular weight of this branched-Z moiety is about 40 kDa.

In some embodiments, -Z contains a portion

In certain embodiments, -Z comprises an amide bond.

其中虛線表示與-L2-或-Z的其餘部分連接；BP^a是選自-N<，-CR<和>C<的分支點；-R選自：-H和C_1-6烷基；如果BP^a為-N<或-CR<，則a為0；如果BP^a為>C<，則n為1。 In certain embodiments, -Z comprises a moiety of formula (a):

Wherein the dotted line indicates connection to the rest of -L2- or -Z; BP ^a is a branch point selected from -N<, -CR< and >C<; -R is selected from: -H and C _1-6 alkyl; if BP ^a is -N< or -CR<, a is 0; if BP ^a is >C<, n is 1.

^-Sa- , ^-Sa'- , -Sa ^" - and -Sa ^''' are independently a bond or selected from: _C1-50 alkyl, _C2-50 alkenyl and _C2-50 alkynyl; wherein _C1-50 alkyl, _C2-50 alkenyl and _C2-50 alkynyl are optionally substituted by one or more identical or different ^-R1 , and wherein _C1-50 alkyl, _C2-50 alkenyl and _C2-50 alkynyl are optionally interrupted by one or more groups selected from the following: -T-, -C(O)O-, -O-, -C(O)-, -C(O)N( ^R2 )-, -S(O) _2N ( ^R2 )-, -S(O)N( ^R2 )-, -S(O) _2- , -S(O)-, -N( ^R2 )S(O) ₂ N(R ^2a )-, -S-, -N(R ² )-, -OC(OR ² )(R ^2a )-, -N(R ² )C(O)N(R ^2a )- and -OC(O)N(R ² )-; each -T- is independently selected from: phenyl, naphthyl, indenyl, indanyl, tetrahydronaphthyl, C _3-10 cycloalkyl, 3 to 10 membered heterocyclic group and 8 to 11 membered heterobicyclic group, 8 to 30 membered carbon polycyclic group and 8 to 30 membered heteropolycyclic group; wherein each -T- is independently optionally substituted by one or more identical or different -R ¹ ; each -R ¹ is independently selected from: halogen, -CN, oxo (=O), -COOR ³ , -OR ³ , -C(O)R ³ , -C(O)N(R ³ R ^3a ), -S(O) ₂ N(R ³ R ^3a ), -S(O)N(R ³ R ^3a ), -S(O) ₂ R ³ , -S(O)R ³ , -N(R ³ )S(O) ₂ N(R ^3a R ^3b ), -SR ³ , -N(R ³ R ^3a ), -NO ₂ , -OC(O)R ³ , -N(R ³ )C(O)R ^3a , -N(R ³ )S(O) ₂ R ^3a , -N(R ³ )S(O)R ^3a , -N(R ³ )C(O)OR ^3a , -N(R ³ )C(O)N(R ^3a R ^3b ), -OC(O)N(R ³ R ^3a ) and C _1-6 alkyl; wherein the C _1-6 alkyl is optionally substituted by one or more halogens which are the same or different; each of -R ² , -R ^2a , -R ³ , -R ^3a and -R ^3b is independently selected from: -H and C _1-6 alkyl, wherein the C _1-6 alkyl is optionally substituted by one or more halogens which are the same or different; and -P ^a' , -P ^a" and -P ^a''' are independently polymer moieties.

In certain embodiments, BP ^a of formula (a) is -N<. In certain embodiments, BP ^a of formula (a) is >C<. In certain embodiments, BP ^a of formula (a) is -CR<. In certain embodiments, -R is -H. Therefore, a of formula (a) is 0.

In certain embodiments, -S ^a - of formula (a) is a chemical bond.

In certain embodiments, -S ^a - of formula (a) is selected from: C _1-10 alkyl, C _2-10 alkenyl and C _2-10 alkynyl, wherein C _1-10 alkyl, C _2-10 alkenyl and C _2-10 alkynyl are optionally interrupted by one or more chemical groups selected from the following: -T-, -C(O)O-, -O-, -C(O)-, -C(O)N(R ⁴ )-, -S(O) ₂ N(R ⁴ )-, -S(O)N(R ⁴ )-, -S(O) ₂ -, -S(O)-, -N(R ⁴ )S(O) ₂ N(R ^4a )-, -S-, -N(R ⁴ )-, -OC(OR ⁴ )(R ^4a )-, -N(R ⁴ )C(O)N(R 4 wherein ^-T- is a 3- to ¹⁰ -membered heterocyclic group; and -R ⁴ and -R ^4a are independently selected from: -H, methyl, ethyl, propyl and butyl.

In certain embodiments, -S ^a - of formula (a) is C _1-10 alkyl interrupted by one or more chemical groups selected from -T-, -C(O)N(R ⁴ )- and -O-.

In certain embodiments, -S ^a'- of formula (a) is a chemical bond.

In certain embodiments, -S ^a' of formula (a) is selected from: C _1-10 alkyl, C _2-10 alkenyl and C _2-10 alkynyl, wherein C _1-10 alkyl, C _2-10 alkenyl and C _2-10 alkynyl are optionally interrupted by one or more chemical groups selected from the following: -C(O)O-, -O-, -C(O)-, -C(O)N(R ⁴ )-, -S(O) ₂ N(R ⁴ )-, -S(O)N(R ⁴ )-, -S(O) ₂ -, -S(O)-, -N(R ⁴ )S(O) ₂ N(R ^4a )-, -S-, -N(R ⁴ )-, -OC(OR ⁴ )(R ^4a )-, -N(R ⁴ )C(O)N(R ^4a )- and -OC(O)N(R ⁴ )-; wherein -R ⁴ and -R ^4a are independently selected from: -H, methyl, ethyl, propyl and butyl. In certain embodiments, -S ^a'- of formula (a) is selected from: methyl, ethyl, propyl, butyl, which is optionally interrupted by one or more chemical groups selected from -O-, -C(O)- and -C(O)N(R ⁴ )-.

In certain embodiments, -Sa ^" - of formula (a) is a chemical bond.

In certain embodiments, -S ^a" - of formula (a) is selected from: C _1-10 alkyl, C _2-10 alkenyl and C _2-10 alkynyl, wherein C _1-10 alkyl, C _2-10 alkenyl and C _2-10 alkynyl are optionally interrupted by one or more chemical groups selected from the following: -C(O)O-, -O-, -C(O)-, -C(O)N(R ⁴ )-, -S(O) ₂ N(R ⁴ )-, -S(O)N(R ⁴ )-, -S(O) ₂ -, -S(O)-, -N(R ⁴ )S(O) ₂ N(R ^4a )-, -S-, -N(R ⁴ )-, -OC(OR ⁴ )(R ^4a )-, -N(R ⁴ )C(O)N(R ^4a )- and -OC(O)N(R ⁴ )-; wherein -R ⁴ and -R ^4a are independently selected from: -H, methyl, ethyl, propyl and butyl. In certain embodiments, -S ^a ″- of formula (a) is selected from: methyl, ethyl, propyl, butyl, which is optionally interrupted by one or more chemical groups selected from -O-, -C(O)- and -C(O)N(R ⁴ )-.

In certain embodiments, Sa ^''' of formula (a) is a chemical bond.

In certain embodiments, Sa ^''' of formula (a) is selected from: _C1-10 alkyl, _C2-10 alkenyl and _C2-10 alkynyl, wherein _C1-10 alkyl, _C2-10 alkenyl and _C2-10 alkynyl are optionally interrupted by one or more chemical groups selected from the following: -C(O)O-, -O-, -C(O)-, -C(O)N( ^R4 )-, -S(O) _2N ( ^R4 )-, -S(O)N( ^R4 )-, -S(O) _2- , -S(O)-, -N( ^R4 )S(O) _2N ( ^R4a )-, -S-, -N( ^R4 )-, -OC( ^OR4 )( ^R4a )-, -N( ^R4 )C(O)N( ^R4a) )- and -OC(O)N(R ⁴ )-; wherein -R ⁴ and -R ^4a are independently selected from: -H, methyl, ethyl, propyl and butyl. In certain embodiments, Sa ^''' of formula (a) is selected from: methyl, ethyl, propyl, butyl, which is optionally interrupted by one or more chemical groups selected from -O-, -C(O)- and -C(O)N(R ⁴ )-.

In certain embodiments, -Pa ^' , -Pa ^" and -Pa ^''' of formula (a) independently comprise a polymer selected from the group consisting of 2-methacryloyl-oxyethylphosphatidylcholine, polyacrylic acid, polyacrylate, polyacrylamide, polyalkoxy polymer, polyamide, polyamidoamine, polyamino acid, polyanhydride, polyaspartamide, polybutyric acid, polyglycolic acid, polybutylene terephthalate, polycaprolactone, polycarbonate, polycyanoacrylate, polydimethacrylamide, polyester, polyethylene, polyethylene glycol, polyethylene oxide, polyethyl phosphate, Polyethyloxazoline, polyglycolic acid, polyhydroxyethyl acrylate, polyhydroxyethyl-oxazoline, polyhydroxymethacrylate, polyhydroxypropylmethacrylamide, polyhydroxypropylmethacrylate, polyhydroxypropyloxazoline, polyiminocarbonate, polylactic acid, polylactic acid-hydroxyacetic acid copolymer, polymethacrylamide, polymethacrylate, polymethyloxazoline, polyorganophosphazene, polyorthoester, polyoxazoline, polypropylene glycol, polysiloxane, polyurethane Esters, polyvinyl alcohol, polyvinylamine, polyvinyl methyl ether, polyvinyl pyrrolidone, silicone, cellulose, carboxymethyl cellulose, hydroxypropyl methyl cellulose, chitin, chitosan, dextran, dextrin, gelatin, hyaluronic acid and its derivatives, functionalized hyaluronic acid, mannan, pectin, rhamnogalacturonic acid, starch, hydroxyalkyl starch, hydroxyethyl starch and other carbohydrate-based polymers, xylan and its copolymers.

In certain embodiments, -Pa ^' , -Pa ^" and -Pa ^" of formula (a) independently comprise a PEG-based moiety. In certain embodiments, Pa', Pa' and Pa''' of formula (a) independently comprise a PEG-based moiety comprising at least 20% PEG, in certain embodiments at least 30% PEG, in certain embodiments at least 40% PEG, in certain embodiments at least 50% PEG, in certain embodiments at least 60% PEG, in certain embodiments at least 70% PEG, in certain embodiments at least 80% PEG, and in certain embodiments at least 90% PEG.

In certain embodiments, -Pa ^' , -Pa ^" and -Pa ^" of formula (a) independently have a molecular weight ranging from 5kDa to 50kDa and including 5kDa and 50kDa, and in certain embodiments, have a molecular weight ranging from 5kDa to 40kDa and including 5kDa and 40kDa. In certain embodiments, it is in the range of 7.5kDa to 35kDa and including 7.5kDa and 35kDa, in certain embodiments, it is in the range of 7.5kDa to 30kDa and including 7.5kDa and 30kDa, and in certain embodiments, it is in the range of 10kDa to 30kDa and including 10kDa and 30kDa.

In certain embodiments, the molecular weight of -Pa ^' , -Pa ^" and -Pa ^" of formula (a) is about 5 kDa. In certain embodiments, the molecular weight of -Pa ^' , -Pa ^" and -Pa ^" of formula (a) is about 7.5 kDa. In certain embodiments, the molecular weight of -Pa ^' , -Pa ^" and -Pa ^" of formula (a) is about 10 kDa. In certain embodiments, the molecular weight of -Pa ^' , -Pa ^" and -Pa ^" of formula (a) is about 12.5 kDa. In another embodiment, the molecular weight of -Pa ^' , -Pa ^" and -Pa ^" of formula (a) is about 15 kDa. In certain embodiments, the molecular weight of -Pa ^' , -Pa ^" and -Pa ^" of formula (a) is about 20 kDa.

In some embodiments, -Z comprises one portion of formula (a). In some embodiments, -Z comprises two portions of formula (a). In some embodiments, -Z comprises three portions of formula (a). In some embodiments, -Z is a portion of formula (a).

其中虛線表示與-L²-或-Z的其餘部分連接；m和p彼此獨立地為150至1000範圍並包括150和1000的整數；在某些實施方案中，為150至500範圍並包括150和500的整數；在某些實施方案中，為200至500範圍並包括200和500的整數。在某些實施方案中，為400至500範圍並包括400和500的整數。 In certain embodiments, -Z comprises a moiety of formula (b):

wherein the dashed line indicates connection to the rest of -L ² - or -Z; m and p are independently integers ranging from 150 to 1000, including integers between 150 and 1000; in certain embodiments, ranging from 150 to 500, including integers between 150 and 500; in certain embodiments, ranging from 200 to 500, including integers between 200 and 500. In certain embodiments, ranging from 400 to 500, including integers between 400 and 500.

In some embodiments, m and p of formula (b) are the same integer. In some embodiments, m and p of formula (b) are about 450.

In certain embodiments, -Z is a moiety of formula (b).

In some embodiments, the total mass of the PTH conjugate is at least 10 kDa, such as at least 12 kDa, such as at least 15 kDa, such as at least 20 kDa, or such as at least 30 kDa. In some embodiments, the total mass of the PTH conjugate is at most 250 kDa, such as at most 200 kDa, 180 kDa, 150 kDa or 100 kDa.

其中未標記的虛線表示通過形成醯胺鍵而與作為PTH部分的-D的氮連接；標有星號的虛線表示與

部分連接；其中m和p彼此獨立地為400至500範圍並包括400和500的整數。 In certain embodiments, the PTH conjugate has the structure of Formula (IIe-i):

The unmarked dashed lines indicate that the nitrogen of -D as the PTH part is connected by forming an amide bond; the dashed lines marked with an asterisk indicate that the nitrogen of -D as the PTH part is connected by forming an amide bond.

Partial connection; wherein m and p are independently integers ranging from 400 to 500 and including 400 and 500.

In certain embodiments, -D is linked to the PTH conjugate of formula (IIe-i) via the N-terminal amine functional group of the PTH moiety.

其中未標記的虛線表示通過形成醯胺鍵而與作為PTH部分的-D的氮連接；標有星號的虛線表示與

部分連接；其中m和p彼此獨立地為400至500範圍並包括400和500的整數。 In certain embodiments, the PTH conjugate has the structure of Formula (IIf-i):

The unmarked dashed lines indicate that the nitrogen of -D as the PTH part is connected by forming an amide bond; the dashed lines marked with an asterisk indicate that the nitrogen of -D as the PTH part is connected by forming an amide bond.

Partial connection; wherein m and p are independently integers ranging from 400 to 500 and including 400 and 500.

In certain embodiments, -D is linked to the PTH conjugate of formula (IIf-i) via the N-terminal amine functional group of the PTH moiety.

In certain embodiments, the liquid drug formulation of the present invention may contain one or more other excipients, such as stabilizers, anti-adsorption agents, viscosity regulators, and antibiotics. In certain embodiments, one excipient may have multiple functions, such as dual or triple functions.

In certain embodiments, the liquid pharmaceutical preparation of the present invention may further comprise a stabilizer, such as a stabilizer selected from the following: alanine; arginine; aspartic acid; glycine; histidine; lysine; proline; sugars such as glucose, sucrose and trehalose; polyols such as glycerol and sorbitol; salts such as potassium phosphate and sodium sulfate; chelating agents such as EDTA and hexaphosphate; ligands such as divalent metal ions; other salts or organic molecules such as phenol derivatives; oligomers or polymers such as cyclodextrin, dextran, dendrimer, PEG, PVP, protamine and HAS. It is understood that sugars can have more than one function, such as acting as an isotonic agent and a stabilizer.

In certain embodiments, the liquid pharmaceutical formulation of the present invention may further comprise an anti-adsorption agent, such as an anti-adsorption agent selected from the following: mainly ionic or non-ionic surfactants or other proteins or soluble polymers, which are used to competitively coat or adsorb to the inner surface of the formulation or formulation container, such as poloxamer (Pluronic F-68), PEG dodecyl ether (Brij 35), polysorbate 20 and 80, dextran, polyethylene glycol, PEG-polyhistidine, BSA, HSA and gelatin. The concentration and type of the excipient selected depends on the effect to be avoided, but generally a monolayer of surfactant is formed at the interface above the CMC value.

Surprisingly, it was found that in the liquid pharmaceutical formulation of the present invention, no oxidation on the methionine residue of the PTH moiety was observed for at least 6 months, thereby eliminating the need for antioxidants. Therefore, in certain embodiments, the liquid pharmaceutical formulation does not contain an antioxidant.

In certain embodiments, the liquid pharmaceutical preparation of the present invention comprises a PTH conjugate, succinic acid, mannitol, m-cresol, and optionally an antioxidant.

In certain embodiments, the liquid pharmaceutical preparation comprises: a PTH conjugate, wherein the PTH portion is 0.05-5.0 mg/ml

And the pH is 3.0 to 6.0.

In certain embodiments, the liquid pharmaceutical preparation comprises: a PTH conjugate, wherein the PTH portion is 0.05-5.0 mg/ml

And the pH is 3.5 to 5.0.

In certain embodiments, the liquid pharmaceutical preparation comprises: a PTH conjugate, wherein the PTH portion is 0.05-5.0 mg/ml

And the pH is 3.7 to 4.3.

In certain embodiments, the liquid pharmaceutical preparation comprises: a PTH conjugate, wherein the PTH portion is 0.1-5.0 mg/ml

And the pH is 3.0 to 6.0.

In certain embodiments, the liquid pharmaceutical preparation comprises: a PTH conjugate, wherein the PTH portion is 0.1-5.0 mg/ml

And the pH is 3.5 to 5.0.

In certain embodiments, the liquid pharmaceutical preparation comprises: a PTH conjugate, wherein the PTH portion is 0.1-5.0 mg/ml

And the pH is 3.7 to 4.3.

In certain embodiments, the liquid pharmaceutical preparation comprises: PTH conjugate, wherein the PTH portion is 0.1-1.5 mg/ml

And the pH is 3.0 to 6.0.

In certain embodiments, the liquid pharmaceutical preparation comprises: a PTH conjugate, wherein the PTH portion is 0.1-1.5 mg/ml

And the pH is 3.5 to 5.0.

In certain embodiments, the liquid pharmaceutical preparation comprises: a PTH conjugate, wherein the PTH portion is 0.1-1.5 mg/ml

And the pH is 3.7 to 4.3.

In certain embodiments, the liquid pharmaceutical preparation comprises: a PTH conjugate, wherein the PTH portion is 0.25-0.35 mg/ml

Meta-cresol 2.0-3.0mg/ml

And the pH is 3.0 to 6.0.

In certain embodiments, the liquid pharmaceutical preparation comprises: a PTH conjugate, wherein the PTH portion is 0.25-0.35 mg/ml

And the pH is 3.5 to 5.0.

In certain embodiments, the liquid pharmaceutical preparation comprises: a PTH conjugate, wherein the PTH portion is 0.25-0.35 mg/ml

And the pH is 3.7 to 4.3.

In certain embodiments, the liquid pharmaceutical preparation comprises: a PTH conjugate, wherein the PTH portion is approximately 0.15 mg/ml

And the pH is 3.0 to 6.0.

In certain embodiments, the liquid pharmaceutical preparation comprises: a PTH conjugate, wherein the PTH portion is 0.15 mg/ml

And the pH is 3.0 to 6.0.

In certain embodiments, the liquid pharmaceutical formulation of the present invention comprises a PTH conjugate comprising about 0.3 mg/ml PTH fraction, about 1.18 mg/ml succinic acid, about 41.7 mg/ml D-mannitol, about 2.5 mg/ml m-cresol, wherein the pH is about 4.0.

In certain embodiments, the liquid pharmaceutical preparation of the present invention comprises a PTH conjugate comprising 0.3 mg/ml PTH fraction, 1.2 mg/ml succinic acid, 42 mg/ml D-mannitol, 3 mg/ml m-cresol, wherein the pH is 4.

In certain embodiments, the liquid pharmaceutical preparation of the present invention comprises a PTH conjugate comprising 0.3 mg/ml PTH fraction, 1.18 mg/ml succinic acid, 41.7 mg/ml D-mannitol, 2.5 mg/ml m-cresol, wherein the pH is 4.0.

It is recognized by those skilled in the art that the liquid pharmaceutical preparation of the present invention may contain a pH adjuster.

In certain embodiments, the liquid pharmaceutical preparation of the present invention comprises a PTH conjugate comprising about 0.3 mg/ml PTH fraction, about 1.18 mg/ml succinic acid, about 41.7 mg/ml D-mannitol, about 2.5 mg/ml m-cresol, about 3.5 mg/ml 1.0N sodium hydroxide, and a pH of about 4.0.

In certain embodiments, the liquid pharmaceutical preparation of the present invention comprises a PTH conjugate, which comprises 0.3 mg/ml PTH fraction, 1.2 mg/ml succinic acid, 42 mg/ml D-mannitol, 3 mg/ml m-cresol, 4 mg/ml 1.0N sodium hydroxide, and has a pH of 4.

In certain embodiments, the liquid pharmaceutical preparation of the present invention comprises a PTH conjugate comprising 0.3 mg/ml PTH fraction, 1.18 mg/ml succinic acid, 41.7 mg/ml D-mannitol, 2.5 mg/ml m-cresol, 3.5 mg/ml 1.0N sodium hydroxide, and a pH of 4.0.

The liquid pharmaceutical formulation as described above is stable for at least 6 months, such as at least 7 months, such as at least 8 months, such as at least 9 months, such as at least 10 months, such as at least 11 months, such as at least 12 months. In certain embodiments, the liquid pharmaceutical formulation is stable for at least 14 months, such as at least 16 months, such as at least 18 months, such as at least 20 months, such as at least 22 months, such as lasting at least 24 months, such as at least 36 months.

In certain embodiments, the temperature of the liquid pharmaceutical preparation as described above is stored at -80°C to 25°C, such as -20°C to 25°C, such as -15°C to 25°C, such as -10°C to 25°C, such as -5°C to 25°C, such as 0°C to 25°C, such as 2°C to 8°C. In certain embodiments, the liquid pharmaceutical preparation is stored at 2°C. In certain embodiments, the liquid pharmaceutical preparation is stored at 4°C. In certain embodiments, the liquid pharmaceutical preparation is stored at 5°C. In certain embodiments, the liquid pharmaceutical preparation is stored at 8°C. In certain embodiments, the liquid pharmaceutical preparation is stored at 10°C. In certain embodiments, the liquid pharmaceutical preparation is stored at 16°C. In certain embodiments, the liquid pharmaceutical preparation is stored at 20°C. In some embodiments, the liquid pharmaceutical formulation is stored at 25°C. In some embodiments, the liquid pharmaceutical formulation is stored at 30°C. In some embodiments, the liquid pharmaceutical formulation is stored at 40°C.

In certain embodiments, the liquid pharmaceutical formulation is stable for at least 36 months when stored at 2°C to 8°C. In certain embodiments, the liquid pharmaceutical formulation is stable for at least 36 months when stored at 2°C. In certain embodiments, the liquid pharmaceutical formulation is stable for at least 6 months when stored at 5°C. In certain embodiments, the liquid pharmaceutical formulation is stable for at least 2 weeks when stored at 30°C.

The applicant surprisingly found that in the liquid pharmaceutical formulation of the present invention, the reversible connection between PTH and the water-soluble carrier is stable, thereby eliminating the need for freeze-drying and reconstitution from the freeze-dried material. However, if desired, the liquid pharmaceutical formulation of the present invention can be dried, such as by freeze-drying to form a dry, such as a freeze-dried pharmaceutical formulation.

In certain embodiments, the method of preparing a liquid pharmaceutical preparation of the present invention comprises the following steps: (i) mixing a PTH complex with at least a buffer, an isotonic agent, a preservative, and optionally an antioxidant; (ii) adjusting the pH of the mixture of step (i); (iii) optionally, filtering the mixture from step (ii); (iv) transferring an amount of the mixture of step (ii) or (iii) equivalent to the desired dose into a container; (v) sealing the container; and wherein the order of steps (ii) and (iii) can be optionally reversed.

In certain embodiments, the PTH complex in step (i) is mixed with a buffer, an isotonic agent, a preservative, and optionally an antioxidant.

In certain embodiments, the method of preparing a liquid pharmaceutical preparation of the present invention comprises the following steps: (i) mixing a PTH complex with at least succinic acid, mannitol, m-cresol and optionally an antioxidant; (ii) adjusting the pH of the mixture of step (i); (iii) optionally filtering the mixture from step (ii); (iv) transferring an amount of the mixture of step (ii) or (iii) equivalent to the desired dose into a container; (v) sealing the container; and wherein the order of steps (ii) and (iii) can be optionally reversed.

In certain embodiments, steps (ii) and (iii) are not reversed.

In certain embodiments, the PTH complex in step (i) is mixed with succinic acid, mannitol, m-cresol and optionally an antioxidant.

In certain embodiments, the method of preparing a liquid pharmaceutical preparation of the present invention comprises the following steps:

(i) mixing a PTH conjugate with at least succinic acid, mannitol, m-cresol and optionally an antioxidant to produce a formulation comprising: a PTH conjugate wherein the PTH portion is 0.05-5.0 mg/ml

(ii) The pH of the mixture of step (i) was adjusted to pH 3.0 to pH 6.0 using NaOH and HCl.

(iii) optionally, filtering the mixture from step (ii);

(iv) transferring an amount of the mixture of step (ii) or (iii) equivalent to the required dosage into a container;

(v) sealing the container; and wherein the order of steps (ii) and (iii) may optionally be reversed.

In certain embodiments, steps (ii) and (iii) are not reversed.

In certain embodiments, the PTH conjugate in step (i) is mixed with succinic acid, mannitol and m-cresol to produce a formulation comprising: a PTH conjugate, wherein the PTH portion is 0.05-5.0 mg/ml

In some embodiments, in step (ii), the pH is adjusted to pH 3.5 to pH 5.0. In some embodiments, in step (ii), the pH is adjusted to pH 3.7 to pH 4.3.

In certain embodiments, the method of preparing a liquid pharmaceutical preparation of the present invention comprises the following steps:

(i) mixing a PTH conjugate with at least succinic acid, mannitol, m-cresol and optionally an antioxidant to produce a formulation comprising: a PTH conjugate wherein the PTH portion is 0.10-5.0 mg/ml

(ii) The pH of the mixture of step (i) was adjusted to pH 3.0 to pH 6.0 using NaOH and HCl.

(iii) optionally, filtering the mixture from step (ii);

(iv) transferring an amount of the mixture of step (ii) or (iii) equivalent to the required dosage into a container;

(v) sealing the container; and wherein the order of steps (ii) and (iii) may optionally be reversed.

In certain embodiments, steps (ii) and (iii) are not reversed.

In certain embodiments, the PTH conjugate in step (i) is mixed with succinic acid, mannitol and m-cresol to produce a formulation comprising: a PTH conjugate, wherein the PTH portion is 0.10-5.0 mg/ml

In some embodiments, in step (ii), the pH is adjusted to pH 3.5 to pH 5.0. In some embodiments, in step (ii), the pH is adjusted to pH 3.7 to pH 4.3.

In certain embodiments, the method of preparing a liquid pharmaceutical formulation of the present invention comprises the following steps: (i) mixing a PTH conjugate with at least succinic acid, mannitol, m-cresol and an optional antioxidant to produce a formulation comprising the following components: PTH conjugate, wherein the PTH portion is 0.10-1.5 mg/ml

(ii) adjusting the pH of the mixture of step (i) to pH 3.0 to pH 6.0 with NaOH and HCl; (iii) optionally, filtering the mixture from step (ii); (iv) transferring an amount of the mixture of step (ii) or (iii) equivalent to the required dose into a container; (v) sealing the container; and wherein the order of steps (ii) and (iii) can be optionally reversed.

In certain embodiments, steps (ii) and (iii) are not reversed.

In certain embodiments, the PTH conjugate in step (i) is mixed with succinic acid, mannitol and m-cresol to produce a formulation comprising: a PTH conjugate, wherein the PTH portion is 0.10-1.5 mg/ml

In some embodiments, in step (ii), the pH is adjusted to pH 3.5 to pH 5.0. In some embodiments, in step (ii), the pH is adjusted to pH 3.7 to pH 4.3.

In certain embodiments, the method of preparing a liquid pharmaceutical formulation of the present invention comprises the following steps: (i) mixing a PTH conjugate with at least succinic acid, mannitol, m-cresol and an optional antioxidant to produce a formulation comprising the following components: a PTH conjugate, wherein the PTH portion is 0.25-0.35 mg/ml

Succinic acid 1.0-1.4mg/ml

(ii) adjusting the pH of the mixture of step (i) to pH 3.0 to pH 6.0 with NaOH and HCl; (iii) optionally, filtering the mixture from step (ii); (iv) transferring an amount of the mixture of step (ii) or (iii) equivalent to the required dose into a container; (v) sealing the container; and wherein the order of steps (ii) and (iii) can be optionally reversed.

In certain embodiments, steps (ii) and (iii) are not reversed.

In certain embodiments, the PTH conjugate in step (i) is mixed with succinic acid, mannitol and m-cresol to produce a formulation comprising: a PTH conjugate, wherein the PTH portion is 0.25-0.35 mg/ml

In some embodiments, in step (ii), the pH is adjusted to pH 3.5 to pH 5.0. In some embodiments, in step (ii), the pH is adjusted to pH 3.7 to pH 4.3.

In certain embodiments, the method of preparing a liquid pharmaceutical formulation of the present invention comprises the following steps: (i) mixing a PTH conjugate with at least succinic acid, mannitol, m-cresol and an optional antioxidant to produce a formulation comprising: a PTH conjugate, wherein the PTH portion is about 0.15 mg/ml

(ii) adjusting the pH of the mixture of step (i) to pH 3.0 to pH 6.0 with NaOH and HCl; (iii) optionally, filtering the mixture from step (ii); (iv) transferring an amount of the mixture of step (ii) or (iii) equivalent to the required dose into a container; (v) sealing the container; and wherein the order of steps (ii) and (iii) can be optionally reversed.

In certain embodiments, steps (ii) and (iii) are not reversed.

In certain embodiments, the PTH conjugate in step (i) is mixed with succinic acid, mannitol and m-cresol to produce a formulation comprising: a PTH conjugate wherein the PTH portion is approximately 0.15 mg/ml

In some embodiments, in step (ii), the pH is adjusted to pH 3.5 to pH 5.0. In some embodiments, in step (ii), the pH is adjusted to pH 3.7 to pH 4.3.

In certain embodiments, the method of preparing a liquid pharmaceutical preparation of the present invention comprises the following steps: (i) mixing a PTH conjugate with at least succinic acid, mannitol, m-cresol and an optional antioxidant to produce a preparation comprising the following components: a PTH conjugate, wherein the PTH portion is 0.30 mg/ml

(ii) adjusting the pH of the mixture of step (i) to about pH 4 with NaOH and HCl; (iii) optionally, filtering the mixture from step (ii); (iv) transferring an amount of the mixture of step (ii) or (iii) equivalent to the required dose into a container; (v) sealing the container; and wherein the order of steps (ii) and (iii) can be optionally reversed.

In certain embodiments, steps (ii) and (iii) are not reversed.

In certain embodiments, the PTH conjugate in step (i) is mixed with succinic acid, mannitol and m-cresol to produce a formulation comprising: a PTH conjugate, wherein the PTH portion is 0.30 mg/ml

In certain embodiments, the pH of the formulation of step (i) is adjusted to pH 4.

Another aspect of the present invention relates to a container containing the liquid pharmaceutical formulation of the present invention.

In certain embodiments, the container can be selected from: a vial; a syringe, such as a dual-chamber syringe; an ampoule and a cartridge, such as a dual-chamber cartridge.

In some embodiments, the cartridge is used with a pen injector, such as an autoinjector.

In certain embodiments, the liquid pharmaceutical formulation of the present invention is provided in a unit dose, meaning that a container containing the liquid pharmaceutical formulation contains one therapeutic dose.

In certain embodiments, the liquid pharmaceutical formulation contains multiple doses, meaning that the container containing the liquid pharmaceutical formulation contains more than one therapeutic dose.

In certain embodiments, the multi-dose liquid pharmaceutical preparation comprises at least 2 doses, such as at least 4 doses, such as at least 6 doses, such as at least 8 doses, such as at least 10 doses, such as at least 12 doses; in certain embodiments, such as at least 14 doses of PTH conjugates.

In certain embodiments, the multi-dose liquid pharmaceutical formulation comprises at least 2, 4, 6, 8, 10, 12, or 14 doses of a PTH conjugate.

Therefore, in another aspect of the invention, the liquid pharmaceutical formulation is provided as a multi-dose formulation.

Another aspect of the invention is a liquid pharmaceutical formulation of the invention for use as a medicament.

In another aspect, the present invention relates to a liquid pharmaceutical formulation of the present invention for treating, controlling, delaying or preventing one or more diseases that can be treated, controlled, delayed or prevented by PTH.

In certain embodiments, the present invention relates to a liquid pharmaceutical formulation of the present invention for treating one or more diseases that can be treated with PTH.

Another aspect of the present invention is a method for treating, controlling, delaying or preventing one or more diseases that can be treated with PTH in a patient, the method comprising administering to the patient a therapeutically effective amount of a PTH liquid pharmaceutical preparation of the present invention.

In certain embodiments, the one or more diseases that can be treated, controlled, delayed or prevented with PTH are selected from: hypoparathyroidism, hyperphosphatemia, osteoporosis, fracture repair, osteomalacia, osteomalacia and osteoporosis in patients with hypophosphatasia, steroid-induced osteoporosis, osteoporosis in men, arthritis, osteoarthritis, osteogenesis imperfecta, fibrodysplasia, rheumatoid arthritis, Paget's disease, humoral hypercalcemia associated with malignant tumors, osteopenia, periodontal disease, fractures, alopecia, chemotherapy-induced alopecia and thrombocytopenia.

In certain embodiments, the one or more diseases that can be treated, controlled, delayed or prevented with PTH are selected from: hypoparathyroidism, hyperphosphatemia, osteoporosis, fracture repair, osteomalacia, osteomalacia and osteoporosis in patients with hypophosphatasia, steroid-induced osteoporosis, male osteoporosis, Arthritis, osteoarthritis, osteogenesis imperfecta, fibrodysplasia, rheumatoid arthritis, Paget's disease, humoral hypercalcemia associated with malignant tumors, osteopenia, periodontal disease, fractures, hair loss, chemotherapy-induced hair loss and thrombocytopenia, chronic periodontitis, mandibular osteonecrosis, and poor fracture healing caused by ALPL gene mutations.

In certain embodiments, the disease is hypoparathyroidism.

In certain embodiments, the disease is hypoparathyroidism induced by treatment with an immune checkpoint inhibitor.

In certain embodiments, the disease is targeted by CTLA-4 (cytotoxic T lymphocyte-associated protein 4), PD-1 (programmed cell death protein 1), PD-L1 (programmed death ligand 1), PD-L2 (programmed death ligand 2), KIR (killer cell immunoglobulin-like receptor), B7-H3, B7-H4, BTLA (B and T lymphocyte detoxifying agent), LAG3 (lymphocyte activation gene 3), TIM-3 (T cell immunoglobulin and mucin domain-containing protein-3), Treatment of induced parathyroid dysfunction with immune checkpoint inhibitors of VISTA (V-domain Ig inhibitor of T cell activation), ILT2/LILRB1 (Ig-like transcript 2/leukocyte Ig-like receptor 1), ILT3/LILRB4 (Ig-like transcript 3/leukocyte Ig-like receptor 4), ILT4/LILRB2 (Ig-like transcript 4/leukocyte Ig-like receptor 2), TIGIT (T cell immune receptor with Ig and ITIM domains), NKG2A, PVRIG or a combination thereof.

In certain embodiments, the disease is hypoparathyroidism induced by treatment with immune checkpoint inhibitors targeting CTLA-4, PD-1, PD-L1, or a combination thereof.

In certain embodiments, the disease is induced parathyroid hypofunction treated with an immune checkpoint inhibitor targeting CTLA-4, such as ipilimumab, tremelimumab, MK-1308, FPT155, PRS010, BMS-986249, BPI-002, CBT509, JS007, ONC392, TE1254, IBI310, BR02001, CG0161, KN044, PBI5D3H5, BCD145, ADU1604, AGEN1884, AGEN1181, CS1002, or CP675206.

In certain embodiments, the disease is induced parathyroid hypofunction treated with an immune checkpoint inhibitor targeting PD-1, such as pembrolizumab, nivolumab, pidilizumab, AMP-224, BMS-936559, cemiplimab, or PDR001.

In certain embodiments, the disease is induced parathyroidism treated with an immune checkpoint inhibitor targeting PD-1, such as MDX-1105, MEDI4736, atezolizumab, avelumab, BMS-936559, or durvalumab.

In certain embodiments, the disease is hypoparathyroidism induced by treatment with an immune checkpoint inhibitor targeting PD-L2.

In certain embodiments, the disease is induced hypoparathyroidism treated with an immune checkpoint inhibitor targeting KIR, such as lirilumab (IPH2102) or IPH2101.

In certain embodiments, the disease is induced hypoparathyroidism treated with an immune checkpoint inhibitor targeting B7-H3, such as MGA271.

In certain embodiments, the disease is induced hypoparathyroidism treated with an immune checkpoint inhibitor targeting B7-H4, such as FPA150.

In certain embodiments, the disease is BTLA-targeted immune checkpoint inhibitor therapy-induced hypoparathyroidism.

In certain embodiments, the disease is induced parathyroid dysfunction treated with an immune checkpoint inhibitor targeting LAG3, such as IMP321 (eftilagimod alpha), relatlimab, MK-4280, AVA017, BI754111, ENUM006, GSK2831781, INCAGN2385, LAG3Ig, LAG525, REGN3767, Sym016, Sym022, TSR033, TSR075, or XmAb22841.

In certain embodiments, the disease is induced hypoparathyroidism treated with an immune checkpoint inhibitor targeting TIM-3, such as LY3321367, MBG453, or TSR-022.

In certain embodiments, the disease is induced parathyroid hypofunction treated with an immune checkpoint inhibitor targeting VISTA, such as JNJ-61610588.

In certain embodiments, the disease is hypoparathyroidism induced by treatment with an immune checkpoint inhibitor targeting ILT2/LILRB1.

In certain embodiments, the disease is hypoparathyroidism induced by treatment with immune checkpoint inhibitors targeting ILT3/LILRB4.

In certain embodiments, the disease is induced hypoparathyroidism treated with an immune checkpoint inhibitor targeting ILT3/LILRB2, such as MK-4830.

In certain embodiments, the disease is induced parathyroid dysfunction treated with an immune checkpoint inhibitor targeting TIGIT, such as MK-7684, PTZ-201, RG6058, or COM902.

In certain embodiments, the disease is induced hypoparathyroidism treated with an immune checkpoint inhibitor targeting NKG2A, such as IPH-2201.

In certain embodiments, the disease is induced parathyroid hypofunction treated with an immune checkpoint inhibitor targeting PVRIG, such as COM701.

In certain embodiments, the disease is hypoparathyroidism induced by treatment with immune checkpoint inhibitors targeting PD-1 and CTLA-4.

In certain embodiments, the disease is immune checkpoint inhibitory treatment with nivolumab-induced hypoparathyroidism.

In certain embodiments, the disease is hypoparathyroidism induced by immune checkpoint inhibitor treatment with ipilimumab.

In certain embodiments, the disease is immune checkpoint inhibitory treatment with pembrolizumab induced hypoparathyroidism.

In certain embodiments, the disease is immune checkpoint inhibitor-induced hypoparathyroidism treated with a combination of nivolumab and ipilimumab.

In certain embodiments, the disease is rheumatoid arthritis induced by treatment with an immune checkpoint inhibitor.

In certain embodiments, the disease is rheumatoid arthritis induced by treatment with an immune checkpoint inhibitor targeting PD-1 or PD-L1.

In certain embodiments, the disease is rheumatoid arthritis induced by treatment with immune checkpoint inhibitors targeting PD-1.

In certain embodiments, the disease is rheumatoid arthritis induced by treatment with an immune checkpoint inhibitor targeting PD-L1.

In certain embodiments, the disease is rheumatoid arthritis induced by immune checkpoint inhibitor treatment with nivolumab.

In certain embodiments, the disease is rheumatoid arthritis induced by immune checkpoint inhibitor treatment with pembrolizumab.

In certain embodiments, the disease is rheumatoid arthritis induced by immune checkpoint inhibitor treatment with a combination of nivolumab and ipilimumab.

In certain embodiments, the disease is rheumatoid arthritis that relapses after treatment with an immune checkpoint inhibitor.

The liquid pharmaceutical preparations of the present invention can be administered, for example, topically, enterally or parenterally, as well as by methods of external application, injection or infusion, including intra-articular, peri-articular, intradermal, subcutaneous, intramuscular, intravenous, intraosseous, intraperitoneal, intrathecal, intracapsular, intraorbital, intravitreal, intratympanic, intravesical, intracardiac, transtracheal, subcutaneous, subcapsular, subarachnoid, intraspinal, intracardiac, intracardiac, intravenous, intravenous, intraperitoneal, intrathecal, intracapsular, intraorbital, intravitreal, intratympanic, intravesical, intracardiac, intratracheal, subcutaneous, subcapsular, subarachnoid, intraspinal, intracardiac ... Intraventricular, intrasternal injection, infusion, intranasal, oral, pulmonary and transdermal administration, direct delivery to the brain via an implanted device allowing delivery of the present or similar formulations to brain tissue or cerebral fluid (e.g., ventricular storage sacs), direct intraventricular injection or infusion, injection or infusion into the brain or areas associated with the brain, injection into the subchoroidal space, retro-orbital injection and intraocular instillation, preferably by subcutaneous injection.

In certain embodiments, the liquid pharmaceutical formulation of the present invention is administered by subcutaneous injection.

In certain embodiments, the liquid pharmaceutical formulations of the present invention are administered by subcutaneous injection using a syringe and needle or a pen injector such as an autoinjector.

In certain embodiments, the liquid pharmaceutical formulation of the present invention is administered by subcutaneous injection using a syringe and needle.

In certain embodiments, the liquid pharmaceutical formulation of the present invention is administered by subcutaneous injection using a pen injector.

In certain embodiments, the liquid pharmaceutical formulation of the present invention is administered by subcutaneous injection using an automatic injector.

In certain embodiments, the time period between two consecutive subcutaneous administrations, i.e., the administration interval, is at least every 12 hours, 24 hours, 36 hours, 48 hours, 60 hours, 72 hours, 84 hours, 96 hours, 108 hours, 120 hours, 132 hours, 144 hours, 156 hours, one week, two weeks, three weeks, or four weeks.

In some embodiments, the time period between two consecutive subcutaneous administrations is 12 hours. In some embodiments, the time period between two consecutive subcutaneous administrations is 24 hours. In some embodiments, the time period between two consecutive subcutaneous administrations is 48 hours. In some embodiments, the time period between two consecutive subcutaneous administrations is 72 hours. In some embodiments, the time period between two consecutive subcutaneous administrations is 96 hours. In some embodiments, the time period between two consecutive subcutaneous administrations is 120 hours. In some embodiments, the time period between two consecutive subcutaneous administrations is 144 hours. In some embodiments, the time period between two consecutive subcutaneous administrations is one week.

Embodiment

Materials and methods

Unless otherwise stated, all materials are commercially available.

To prepare the formulation, a formulation excipient stock solution was prepared. All excipients were mixed and filled to 90% of the final volume with water, and then the calculated amount of NaOH solution was spiked into the solution after the addition of compound 1 to achieve the desired pH value of the final formulation. Subsequently, the appropriate amount of compound 1 was added and dissolved. After adjusting the pH, the volume was adjusted to the final volume in a volumetric flask. Before filling, the formulation was filtered through a 0.2μm PVDF filter.

Visual inspection: Cartridges were inspected for visible or invisible particles with gentle, manual, radial agitation for 5 seconds against a white background and for 5 seconds against a black background according to the European Pharmacopoeia (8th edition; monograph 2.9.20). The inspection was performed independently by two trained inspectors. For classification of observed visible particles, a score based on the "Deutscher Arzneimittel-Codex" (DAC 2006) was used.

pH: The pH of the preparation was measured using a calibrated pH meter at room temperature using a standard ionic strength electrode.

Osmotic pressure: The osmotic pressure of a sample is measured by the freezing point depression method.

Microflow Imaging: Microflow imaging measurements were performed on the MFI-5200 particle analyzer system.

Purity of compound 1 was determined by HP-SEC: The sample was analyzed on a 10x300 mm column consisting of cross-linked agarose and dextran graded for fractionating globular proteins with a molecular weight of 10,000 to 60,0000 g/mol. The detection wavelength was 215 nm.

RP-HPLC was used to detect free PTH and determine the purity and content of compound 1 : using a C18 2.1x100mm column with a pore size of 130Å and a particle size of 1.7μm; detecting at a wavelength of 215nm; and determining the content by measuring a reference sample.

RP-HPLC analysis after in vitro release: Release of PTH from compound 1 was performed at pH 10.1 and 5°C to minimize degradation of the peptide. After incubation at 5°C for 76 hours, release was terminated by the addition of acetic acid. The resulting samples were analyzed by RP-HPLC on a C18 2.1x100mm column with a pore size of 130Å and a particle size of 1.7μm; detection was performed at a wavelength of 215nm.

Example 1 : Synthesis of Compound 1

Compound 1 was synthesized according to the method of compound 18 described in WO2017/148883A1.

Example 2: Stability test of formulation containing compound 1

The effect of the pH of liquid drug formulations on the amount of PTH (1-34) released from Compound 1 was evaluated. To this end, four different formulations (F1, F2, F3 and F4) containing Compound 1 were prepared (Table 1). Each formulation contained 0.4 mg PTH (1-34) / mL.

The formulations were loaded into cartridges and incubated for 3 months in incubators set at 5°C, 25°C/60%RH, and 40°C/75%RH. After 2 weeks, 1 month, and 3 months, one cartridge of each formulation and storage condition was removed and analyzed. Table 2 shows the amount of free PTH released over time from F1, F2, F3, and F4 as detected by RP-HPLC. It was observed that higher amounts of free PTH were released in liquid drug formulations with higher pH values.

n.d.=below the limit of quantification

Next, the purity of PTH(1-34) in F1, F2, F3 and F4 was evaluated. For this purpose, the release of PTH from compound 1 was induced at pH 10.1 and 5°C. After 76 hours of incubation at 5°C, the release was terminated by the addition of acetic acid. The obtained samples were subjected to RP-HPLC analysis. Table 3 shows that the percentage of deamidated peptide species was more significant for formulations with higher pH values, while the percentage of truncated peptide species, namely PTH(1-30), was more significant in formulations with lower pH values. Overall, the highest peptide purity was observed over time for formulation F1 for samples incubated at 40°C/75%RH.

Example 3: Stability of formulations containing compound 1 to oxidation

The effects of antioxidants and oxygen in the headspace (HS) on the stability of compound 1 and free PTH were investigated. Four different formulations (F5, F6, F7, and F8) containing compound 1 at pH 4.0 were prepared with different concentrations of antioxidants and volumes of HS (Table 4). Each formulation contained 0.4 mg PTH (1-34)/mL.

The formulations were loaded into cartridges with various headspaces (no HS, 50 μL HS, or 200 μL HS). The cartridges were incubated in an incubator set at 5°C and 25°C/60%RH for up to 6 months. After 2 weeks, 1 month, 3 months, and 6 months, one cartridge per formulation and storage condition was removed and analyzed by RP-HPLC.

Table 5 shows that for the formulation stored at 5°C, no free PTH was released over a period of 6 months. For the formulation stored at 25°C, about 0.9% free PTH was detected after 3 months.

n.d.=below the limit of quantification

After induced release of PTH from compound 1 (pH 10.1 and 5°C), the resulting mixture was subjected to RP-HPLC analysis. As shown in Table 6, the amount of oxidative substances detected did not change for all formulations. Changes in headspace volume and antioxidant concentration had no significant effect on the levels of oxidative substances.

Example 4: Effect of preservative concentration on the stability of compound 1

The effect of preservative concentration and preservative type on the stability of compound 1 and peptides was studied. To achieve this goal, four different formulations (F9, F10, F11 and F12) containing compound 1 at pH 4.0 were prepared. Each formulation contained 0.4 mg PTH (1-34) / mL. The formulations were filled in cartridges and cultured in an incubator set at 5 ° C and 40 ° C / 75% RH for up to 6 months. After 1 month, 3 months and 6 months, one cartridge of each formulation and storage condition was removed from the respective incubator and analyzed.

RP-HPLC analysis showed that all formulations stored at 5°C for 6 months contained no free PTH. For formulations stored at 40°C, the free PTH content increased to 5.0% and 5.5% after three months, respectively. Table 8 shows that no significant differences were found for the formulations tested.

n.d.=below the limit of quantification

After release of PTH from Compound 1 (pH 10.1 and 5°C), the resulting solution was subjected to RP-HPLC analysis. As shown in Table 9, RP-HPLC analysis did not show a decrease in peptide purity for all formulations stored at 5°C. After 6 months of storage at 5°C, deamidation, truncation levels (i.e., PTH (1-30) amount and oxidation) remained unchanged. Formulations stored at 40°C for 3 months showed a decrease in peptide purity from approximately 90% at t0 to 56-58% after 3 months. Deamidation increased from 2% to approximately 17%, and PTH (1-30) increased from 0.1-0.2% to 10%. The oxidation level did not change, except for a slight increase in formulation F9. No substantial differences were observed between the formulations analyzed. Therefore, the concentration or type of preservative had no significant effect on the stability of compound 1 and the purity of the peptide.

Example 5: Stability test of formulation containing compound 1

The effects of the concentration of compound 1 and the excipient as well as the effect of pH were evaluated. To this end, 19 formulations containing compound 1 were prepared at pH 3.5-4.5 (F13-F31, see Table 10). Each formulation contained 0.2-0.8 mg PTH(1-34)/mL

The formulations were filled into cartridges and incubated for up to 6 months in incubators set at 5°C, 25°C/60%RH, 30°C/65%RH, and 40°C/75%RH. After 1 month, 3 months, and 6 months, one cartridge per formulation and storage condition was removed from the respective incubator and analyzed. Visual inspection results showed that for all liquid drug formulations F13 to F31, the samples were clear, colorless, and particle-free during the stability study (6 months). Only a few visible particles were detected sporadically. In addition, no visible aggregates were formed under the selected conditions.

The pH values of liquid drug formulations F13 to F31 were monitored throughout 24 months when stored at 5°C, throughout 6 months when stored at 25°C and 30°C, and throughout 3 months when stored at 40°C; and were observed to remain within the specification range of ±0.1 of the target value. No formulation differences were observed.

Next, different osmotic pressure values were measured for liquid drug formulations F13 to F31 due to the different amounts of excipients in the formulations. No changes in osmotic pressure values were detected for all formulations after 24 months at 5°C, 6 months at 25°C and 30°C, and 3 months at 40°C.

Regarding the microfluidic imaging (MFI) results, very low particle concentrations were observed for all formulations at T0, as well as in the stability studies (when the formulations were stored at 5°C for up to 24 months, when the formulations were stored at 25°C and 30°C for up to 6 months, and when the formulations were stored at 40°C for up to 3 months). Only sporadic silicone oil droplet-like particles were found.

Likewise, photostability studies were conducted by light exposure testing, where a liquid formulation containing 0.3 mg PTH(1-34)/mL, 1.18 mg/mL succinic acid, 41.7 mg/mL mannitol, 2.5 mg/mL m-cresol at pH 4.0 was exposed to 71798 lux hours of light, plus 8 wh/ ^m2 of UV exposure. No differences were observed between the exposed formulation and the reference sample.

According to HP-SEC analysis, the fraction of high molecular weight substances remained constant within 24 months when the formulation was stored at 5°C and within 6 months of the stability study when the formulation was stored at 25°C, 30°C and 40°C; the constant amount of HMW substances indicated that there was no aggregate formation within 24 months when the formulation was stored at 5°C and within 6 months when the formulation was stored at 25°C, 30°C and 40°C.

RP-HPLC analysis showed that the content of compound 1 did not change when the formulations were stored at 5°C for up to 24 months (Table 11). The content of some formulations decreased slightly when stored at 25°C for 6 months. This trend was more obvious after storage at higher temperatures, as shown in Table 12. Formulations with higher pH values showed a more obvious decrease in the content of compound 1 than formulations with lower pH values.

The purity of compound 1 decreased only slightly when the formulations were stored at 5°C for up to 24 months (Table 13). For all formulations, the decrease in purity was more pronounced at higher temperatures (Table 14). Formulations with higher pH values were more affected than those with lower pH.

No free PTH was detected in formulations stored at 5°C for 24 months. A slight increase in free PTH was observed over time for formulations stored at higher temperatures.

n.d.=below the limit of quantification

n.d.=below the limit of quantification

Formulations with higher pH values showed a stronger increase in free PTH species than formulations with lower pH values, but no substantial differences were observed between the formulations analyzed.

n.d.=below the limit of quantification

n.d.=低於定量限

nd = below the limit of quantitation

RP-HPLC after release of PTH (pH 10.1 and 5°C) showed no significant change in the purity of free PTH for formulations stored at 5°C for up to 24 months. At higher temperatures, a decrease in purity with time and temperature was found. The lower pH formulation was characterized by a slightly lower purity than the higher pH formulation.

As shown in Tables 21 and 22, no significant increase in oxidative species was found for any formulation or storage condition after PTH release. Only a small increase in one oxidative species was found in all formulations. This increase was slightly significant at high temperature.

RP-HPLC analysis showed that PTH(1-30) increased with time and temperature for all formulations. The formulation with lower pH was characterized by slightly higher amounts of PTH(1-30) compared to the formulation with higher pH.

Tables 24 and 25 show that the amount of deamide substances in all formulations increases with time and temperature. For formulations stored at 5°C, there is no significant change in the amount of deamide substances after 24 months of storage.

Abbreviation

CHAPS-3-[(3-cholesterolamidopropyl) dimethylammonium]-1-propane sulfonate

EDTA-ethylenediaminetetraacetic acid

HEPES-4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid

LC-MS-Liquid Chromatography Coupled Mass Spectrometry

M-Month

n.d.-Cannot be determined or below limit of quantification (LOQ)

pH-hydrogen ion potential

PTH-parathyroid hormone

RH-Relative Humidity

RP-HPLC-Reversed Phase High Performance Liquid Chromatography

SEC-Size Exclusion Chromatography

sec-seconds

TFA-trifluoroacetic acid

TRIS-Tris(hydroxymethyl)aminomethane

UPLC-Ultra-Performance Liquid Chromatography

W-week

Claims (13)

A liquid pharmaceutical preparation, wherein the pharmaceutical preparation comprises a PTH conjugate, a buffer, an isotonic agent and a preservative, and wherein the PTH conjugate comprises a PTH portion covalently and reversibly conjugated to a water-soluble carrier, wherein the pharmaceutical preparation comprises: the PTH conjugate, wherein the PTH portion 0.3 ± 0.03 mg/ml, succinic acid 1.18 ± 0.118 mg/ml, D-mannitol 41.7 ± 4.17 mg/ml, m-cresol 2.5 ± 0.25 mg/ml; and wherein the pH is 4 ± 0.4, wherein the PTH conjugate has the structure of formula (Ia): (Ia) wherein -D is the PTH moiety; the ‑L ¹ ‑L ² ‑moieties are selected from: (IIcb-i), (IIcb-ii) and (IIcb-iii); wherein the unlabeled dashed line indicates the connection to the nitrogen of -D through the formation of an amide bond; and the dashed line marked with an asterisk indicates the connection to -Z; -Z is a water-soluble carrier portion and comprises a portion of formula (b): (b), wherein the dashed line indicates connection with ‑L ² ‑; and m and p are independently integers ranging from 150 to 1000 and including 150 and 1000; and x is 1. A liquid pharmaceutical preparation as described in claim 1, wherein the pharmaceutical preparation further comprises a pH adjuster. A liquid pharmaceutical preparation as described in claim 1 or 2, wherein the pharmaceutical preparation comprises 0.3 mg/ml of the PTH fraction, 1.18 mg/ml of the succinic acid, 41.7 mg/ml of the D-mannitol, and 2.5 mg/ml of the m-cresol, and wherein the pH is 4.0. The liquid pharmaceutical preparation according to claim 1 or 2, wherein -L ¹ - is directly conjugated to the N-terminal amine functional group of -D. A liquid pharmaceutical formulation as described in claim 1 or 2, wherein -D has a sequence of SEQ ID NO:47, SEQ ID NO:48, SEQ ID NO:49, SEQ ID NO:50, SEQ ID NO:51, SEQ ID NO:52, SEQ ID NO:53, SEQ ID NO:54, SEQ ID NO:55, SEQ ID NO:107, SEQ ID NO:108, SEQ ID NO:109, SEQ ID NO:110, SEQ ID NO:111, SEQ ID NO:112, SEQ ID NO:113, SEQ ID NO:114, or SEQ ID NO:115. A liquid pharmaceutical formulation as described in claim 1 or 2, wherein -D has a sequence of SEQ ID NO:51. A liquid pharmaceutical formulation as described in claim 1 or 2, wherein m and p are independently integers ranging from 400 to 500 and including 400 and 500. The liquid pharmaceutical preparation of claim 1 or 2, wherein the PTH conjugate has the structure of formula (IIf-i): (IIf-i), wherein the unlabeled dashed line indicates the connection to the nitrogen of the N-terminal amine functional group of -D through the formation of an amide bond; and the dashed line marked with an asterisk indicates the connection to Partial connection; wherein m and p are independently integers ranging from 400 to 500 and including 400 and 500. A method for producing a liquid pharmaceutical preparation as described in any one of claims 1 to 8, wherein the method comprises the following steps: (i) mixing the PTH complex with at least the buffer, the isotonic agent, the preservative and optionally the antioxidant; (ii) adjusting the pH of the mixture of step (i); (iii) optionally filtering the mixture of step (ii); (iv) transferring an amount of the mixture of step (ii) or (iii) equivalent to the required dose into a container; (v) sealing the container; and wherein the order of steps (ii) and (iii) can be optionally reversed. A production method as described in claim 9, wherein steps (ii) and (iii) are not reversed. The liquid pharmaceutical preparation according to claim 1 or 2, which is used as a medicine. The liquid pharmaceutical preparation according to claim 1 or 2, which is used for treating hypoparathyroidism. A use of the liquid pharmaceutical preparation as described in any one of claims 1 to 8 in the preparation of a medicament for treating hypoparathyroidism.