RU2808427C2 - Methods of treating pancreas cancer using combination therapy including liposomal irinotecan - Google Patents

Abstract

FIELD: medicine; oncology.

SUBSTANCE: invention relates to a method of treating exocrine pancreatic cancer in a human patient who has been pretreated with gemcitabine, the said invention comprises intravenously administering to the patient an antitumor therapy consisting of the following: a) a dose of 60, 70 or 80 mg/m² of a liposomal irinotecan composition containing irinotecan liposomes in a diluted irinotecan injection formulation, wherein the diluted irinotecan injection formulation is prepared by diluting a volume of a liposomal irinotecan composition containing 5 mg/ml liposomal irinotecan in 500 ml of 5% dextrose injection, in combination with b) a dose of 200 mg/m² (l) form of leucovorin and a dose of 2,400 mg/m² of 5-fluorouracil, for the treatment of exocrine pancreatic cancer in a patient, where the liposomal composition of irinotecan contains a salt of irinotecan with sucrose octasulfate, encapsulated in irinotecan liposomes consisting of phosphatidylcholine, cholesterol and phosphatidylethanolamine modified with polyethylene glycol, and having a diameter of approximately 80–140 nm, wherein a diluted injectable formulation of irinotecan, leucovorin and 5-fluorouracil is administered to the patient sequentially, starting on the first day of a two-week cycle.

EFFECT: invention provides therapeutic synergy in combination therapy using liposomal irinotecan with 5-FU and leucovorin.

12 cl, 7 ex, 7 dwg, 14 tbl

Description

CROSS REFERENCE TO RELATED APPLICATIONS

This application claims priority to US Provisional Application No. 61/659211 (filed June 13, 2012) and US Provisional Application No. 61/784382 (filed March 14, 2013), each of which is incorporated herein by reference.

BACKGROUND OF THE ART

Despite advances in cancer treatment, there remains an urgent need to further improve treatment options to prolong patients' lives while maintaining quality of life, especially for advanced-stage cancers such as pancreatic cancer, which are often refractory or become resistant to current therapeutic treatments.

The incidence of pancreatic cancer has increased markedly over the past few decades. It is currently the fourth leading cause of cancer death in the United States. The high mortality rate from pancreatic cancer is due to the lack of effective treatments and the complete lack of reliable treatment options. Because of the location of the pancreas, pancreatic cancer is typically not diagnosed until the tumor is large enough to cause systemic symptoms. This, coupled with the lack of good screening tools and limited understanding of risk factors, means that patients tend to have advanced (often metastatic) cancer at the time of diagnosis. The prognosis for metastatic pancreatic cancer is poor and almost always fatal, with an overall five-year survival rate of less than 4%.

Chemotherapy using one or more of 5-fluorouracil (5-FU) and gemcitabine has been shown to prolong survival in pancreatic cancer. Combination therapy including folinic acid (leucovorin or levuloleucovorin), 5-fluorouracil and irinotecan (FOLFIRI), folinic acid, 5-fluorouracil, irinotecan and oxaliplatin (FOLFIRINOX) or, less commonly, a combination of folin acid, 5-fluorouracil and oxaliplatin (FOLFOX ) is also used to treat some types of pancreatic cancer. Irinotecan is 7-ethyl-10-[4-(1-piperidine)-1-piperidine]carbonyloxycapothecin, IUPAC name (S)-4,11-diethyl-3,4,12,14-tetrahydro-4-hydroxy -3,14-dioxo-1H-pyran-[3',4':6,7]-indolizine-[1,2-b]-quinolin-9-yl-[1,4'-bipiperidine]-1' -carboxylate. Irinotecan is a member of the topoisomerase I inhibitor class of drugs and is a semisynthetic, water-soluble analogue of the natural alkaloid camptothecin. Also known as CPT-11, irinotecan is currently marketed as an aqueous solution as Camptosar® (irinotecan hydrochloride injection). Topoisomerase I inhibitors, such as irinotecan, stop uncontrolled cell growth by inhibiting DNA unwinding, thereby preventing DNA replication.

Irinotecan has a complex pharmacology; activation, inactivation and elimination of this drug involve extensive metabolic transformations. Irinotecan is a prodrug that is converted by nonspecific carboxylesterases to a 100- to 1000-fold more active metabolite, SN-38. SN-38 is not recognized by P-glycoprotein, a drug transporter that plays an important role in acquired drug resistance by clearing certain drugs from the cell, so irinotecan is likely to be active in tumors that are resistant to other standard chemotherapeutic agents. SN-38 is excreted from the body through glucuronidation, for which major pharmacogenetic variability has been described, and excretion in bile. These properties of this drug substance contribute to the marked heterogeneity in the efficacy and toxicity of irinotecan observed in clinical settings. Irinotecan hydrochloride injection is approved in the United States for the treatment of metastatic colon or kidney cancer; This drug is also used to treat cancer of the colon, rectum, stomach, lungs, uterus, cervix and ovaries.

There are several approved treatment options for advanced or metastatic pancreatic cancer, especially those of exocrine origin. Gemcitabine monotherapy is the current standard of care for first-line treatment of advanced and metastatic pancreatic adenocarcinoma. In clinical studies, gemcitabine monotherapy consistently demonstrated a mean survival benefit of 5 to 6 months and 1-year survival of approximately 20%. The use of gemcitabine monotherapy is also approved as second-line treatment for patients previously treated but no longer responding to 5-fluorouracil; however, the median overall increase in life expectancy was 3.9 months.

Based on knowledge of the biology of pancreatic cancer, a number of targeting agents have been evaluated, but only erlotinib, a protein tyrosine kinase inhibitor that targets EGFR, has been approved for use as first-line therapy in advanced pancreatic cancer, and this approval has only been given for use in combination with gemcitabine. Coadministration of erlotinib with gemcitabine resulted in a statistically significant survival benefit and improved median survival (6.4 months vs. 5.9 months) and 1-year survival (24% vs. 17%) when compared with monotherapy using gemcitabine. Clinical studies evaluating other targeted agents, including studies of the antibodies bevacizumab and cetuximab, have been disappointingly negative. Thus, there is an urgent need for an improvement and an effective alternative to the current treatments for pancreatic cancer. The described invention satisfies this need and provides other advantages.

BRIEF DESCRIPTION OF THE INVENTION

Methods of treating pancreatic cancer in a patient (i.e., a human patient) are provided, comprising administering to the patient liposomal irinotecan (e.g., injection of a liposomal preparation of irinotecan octasulfate salt - sucrose, also called MM-398) alone or in combination with 5-fluorouracil ( 5-FU) and leucovorin (collectively referred to as 5-FU/LV) according to the specific clinical administration regimen. In addition, compositions adapted for use in such methods are provided.

In one aspect, there is provided a method of treating (e.g., effectively treating) pancreatic cancer in a patient, comprising: administering to the patient an effective amount of liposomal irinotecan, the method comprising at least one cycle, wherein the cycle is a period of 3 weeks, over of each cycle, liposomal irinotecan is administered on day 1 of the cycle at a dose of 120 mg/ ^m2 , except if the patient is homozygous for the UGT1A1*28 allele; in this case, liposomal irinotecan is administered on day 1 of cycle 1 at a dose of 80 mg/ ^m2 . In one embodiment, the dose of liposomal irinotecan administered to a patient homozygous for the UGT1A1*28 allele is increased after one in increments of 20 mg/m ² , but not to exceed 120 mg/m ² .

In yet another aspect, a method of treating pancreatic cancer in a patient includes coadministering to the patient an effective amount of liposomal irinotecan, 5-fluorouracil (5-FU), and leucovorin, the method comprising at least one administration cycle equal to a 2-week period, wherein for each cycle:

(a) liposomal irinotecan is administered to patients who are not homozygous for the UGT1A1*28 allele on day 1 of each cycle at a dose of 80 mg/ ^m2 , and to patients homozygous for the UGT1A1*28 allele on day 1 of cycle 1 at a dose of 60 mg/m ² and on day 1 of each subsequent cycle at a dose of 60 mg/ ^m2 to 80 mg/ ^m2 (for example, 60 mg/ ^m2 or 70 mg/ ^m2 or 80 mg/ ^m2 );

(b) 5-FU is administered at a dose of 2400 mg/m ² ; And

(c) leucovorin is administered at a dose of 200 mg/ ^m2 (l-form, or levoleucovorin) or 400 mg/ ^m2 (l+d-racemic form).

In one embodiment, the dose of liposomal irinotecan administered to a patient homozygous for the UGT1A1*28 allele is increased after one cycle to 80 mg/m ² . In one embodiment, in each cycle, liposomal irinotecan is administered before leucovorin and leucovorin is administered before 5-FU.

In yet another embodiment, liposomal irinotecan is administered intravenously over 90 minutes.

In yet another embodiment, 5-FU is administered intravenously over 46 hours.

In yet another embodiment, leucovorin is administered intravenously over 30 minutes.

In yet another embodiment, the patient is premedicated with dexamethasone and/or a 5-HT3 antagonist or other antiemetic prior to each administration of liposomal irinotecan.

In yet another embodiment, the pancreatic cancer is an exocrine pancreatic cancer selected from the group consisting of acinar cell carcinoma, adenocarcinoma, glandular squamous cell carcinoma, giant cell tumor, intraductal papillary mucinous neoplasm (IPMN), mucinous cystadenocarcinoma, pancreatoblastoma, serous cystadenocarcinoma and solid and pseudopapillary tumors.

In one embodiment, treatment of the patient results in a positive outcome, wherein the positive outcome is pathological complete response (pCR), complete response (CR), partial response (PR), or stable disease (SD). In yet another embodiment, combination therapy using liposomal irinotecan, 5-FU and leucovorin results in therapeutic synergy.

In yet another embodiment, liposomal irinotecan is formulated in an injectable liposomal form of irinotecan-sucrose octasulfate (MM-398). Irinotecan-Sucrose Octasulfate Injectable Liposomal Form may also be referred to as Irinotecan-HCl Injectable Liposomal Formulation because irinotecan-HCl is the active pharmaceutical ingredient used to load irinotecan into liposomes containing triethylammonium sucrose octasulfate to produce MM-398 liposomes. This nomenclature can be used even though the hydrochloride ion of irinotecan-HCl reacts with the triethylammonium ion of triethylammonium sucrose octasulfate to form triethylammonium chloride (triethylamine hydrochloride), leaving irinotecan-sucrose octasulfate as a pharmaceutical agent in the MM-398 liposomes. In yet another aspect, kits are provided for the treatment of pancreatic cancer in a patient, comprising a dose of liposomal irinotecan and instructions for use of the liposomal irinotecan, as described herein.

In yet another aspect, kits are provided for the treatment of pancreatic cancer in a patient, containing a dose of liposomal irinotecan, 5-fluorouracil (5-FU) and leucovorin, as well as instructions for use of liposomal irinotecan, 5-FU and leucovorin, as described herein.

In one embodiment, the kit covers the treatment of exocrine pancreatic cancer selected from the group consisting of acinar cell carcinoma, adenocarcinoma, glandular squamous cell carcinoma, giant cell tumor, intraductal papillary mucinous neoplasm (IPMN), mucinous cystadenocarcinoma, pancreatoblastoma, serous cystadenocarcinoma, and solid and pseudopapillary tumors.

In one embodiment, liposomal irinotecan is an injectable liposomal form of irinotecan-sucrose octasulfate (MM-398).

In yet another aspect, a liposomal irinotecan formulation is provided for coadministration with 5-fluorouracil (5-FU) and leucovorin for at least one cycle, wherein the cycle is a period of 2 weeks, the irinotecan formulation is a liposomal irinotecan formulation, and:

(a) liposomal irinotecan is administered to patients who are not homozygous for the UGT1A1*28 allele on day 1 of each cycle at a dose of 80 mg/ ^m2 , and to patients homozygous for the UGT1A1*28 allele on day 1 of cycle 1 at a dose of 60 mg/m ² and on day 1 of each subsequent cycle at a dose of 60 mg/ ^m2 or 80 mg/ ^m2 ;

(b) 5-FU is administered at a dose of 2400 mg/m ² ; And

(c) leucovorin is administered at a dose of 200 mg/ ^m2 (l-form, or levoleucovorin) or 400 mg/ ^m2 (l+d-racemic form).

In one embodiment, the dose of liposomal irinotecan administered to a patient homozygous for the UGT1A1*28 allele is increased after 1 cycle to 80 mg/m ² . In yet another embodiment, liposomal irinotecan is administered intravenously over 90 minutes.

In yet another embodiment, 5-FU is administered intravenously over 46 hours.

In yet another embodiment, leucovorin is administered intravenously over 30 minutes.

In yet another embodiment, the patient is premedicated with dexamethasone and/or a 5-HT3 antagonist or other antiemetic prior to each administration of liposomal irinotecan.

In yet another embodiment, the pancreatic cancer is an exocrine pancreatic cancer selected from the group consisting of acinar cell carcinoma, adenocarcinoma, glandular squamous cell carcinoma, giant cell tumor, intraductal papillary mucinous neoplasm (IPMN), mucinous cystadenocarcinoma, pancreatoblastoma, serous cystadenocarcinoma and solid and pseudopapillary tumors.

In yet another embodiment, the irinotecan liposomal formulation is an injectable liposomal form of irinotecan-sucrose octasulfate.

In yet another aspect, a method is provided for improving the results of chemotherapy by increasing tumor vascularization, comprising administering to a patient with a tumor an injection of a liposomal preparation of irinotecan octasulfate salt-sucrose in an amount effective to increase tumor vascularization, and simultaneously administering to the patient an effective amount of a non-irinotecan chemotherapeutic agent.

In yet another aspect, an injectable liposomal form of irinotecan-sucrose octasulfate is provided for co-administration to a patient with a tumor: 1) an injectable liposomal form of irinotecan-sucrose octasulfate in an amount effective to increase tumor vascularization, and 2) an effective amount of a non-irinotecan chemotherapeutic agent.

Further, the invention includes a method of treating exocrine pancreatic cancer, comprising intravenously administering antineoplastic therapy once every two weeks to a human patient having exocrine pancreatic cancer, wherein the antineoplastic therapy consists of:

a) administering a dose of 60-80 mg/m ² of the liposomal irinotecan composition in irinotecan dilute injectable formulation to a human patient in a single infusion over approximately 90 minutes, wherein the irinotecan dilute injectable formulation contains irinotecan liposomes in 500 ml of injectable fluid and a volume of 5 mg/ml liposomal irinotecan formulation is effective for delivering a dose of liposomal irinotecan, in combination with

b) administering a therapeutically effective amount of leucovorin and 5-fluorouracil,

for the treatment of exocrine pancreatic cancer in a patient, wherein irinotecan liposomes consist of phosphatidylcholine, cholesterol and polyethylene glycol modified phosphatidylethanolamine and have a diameter of approximately 80-140 nm.

In a preferred embodiment, the polyethylene glycol modified phosphatidylethanolamine is present in an amount of approximately one molecule of polyethylene glycol per 200 molecules of phospholipids in irinotecan liposomes.

In one embodiment of the invention, irinotecan liposomes are monolamellar lipid bilayer vesicles encapsulating a salt of irinotecan with sucrose octasulfate.

The invention also applies to cases where exocrine pancreatic cancer has progressed during therapy with gemcitabine before administration of irinotecan liposomes.

Preferably, the therapeutically effective amount of leucovorin is 200 mg/m ² (l) form of leucovorin, optionally administered as 400 mg/m ² (l+d) racemic form of leucovorin, and the therapeutically effective amount of 5-fluorouracil is 2400 mg/m ² per dose .

In one embodiment, a diluted injectable formulation of irinotecan, leucovorin, and 5-fluorouracil is administered to a patient sequentially, beginning on the first day of a two-week cycle, resulting in the simultaneous presence of irinotecan, leucovorin, and 5-fluorouracil liposomes in the patient's blood.

Preferably, a diluted injectable formulation of irinotecan is administered to the patient prior to 5-fluorouracil.

Further, according to the invention:

a) the patient has been previously treated with gemcitabine;

b) leucovorin is administered as 400 mg/m ² (l+d) racemic form of leucovorin;

c) exocrine pancreatic cancer progressed during therapy with gemcitabine before administration of diluted injectable irinotecan;

d) irinotecan liposomes are monolamellar lipid bilayer vesicles encapsulating the irinotecan salt with sucrose octasulfate, and

e) polyethylene glycol modified phosphatidylethanolamine is present in an amount of approximately one molecule of polyethylene glycol per 200 molecules of phospholipids in irinotecan liposomes.

The invention may further include storing the diluted irinotecan injectable formulation at a temperature of approximately 2-8°C for no more than 24 hours prior to administration.

Preferably, irinotecan liposomes are monolamellar lipid bilayer vesicles that encapsulate an aqueous space containing irinotecan in a gelled or precipitated state as a sucrose octasulfate salt.

In a preferred embodiment of the invention, the injectable liquid is 5% dextrose injection solution.

The invention further includes a method of treating exocrine pancreatic cancer in a human patient who has been pretreated with gemcitabine, comprising intravenously administering to the patient an antineoplastic therapy consisting of:

a) a dose of 60-80 mg/m ² of a liposomal irinotecan composition containing irinotecan liposomes in a diluted irinotecan injection formulation, wherein the diluted injection formulation is prepared by diluting a volume of a liposomal irinotecan composition containing 5 mg/ml liposomal irinotecan in 500 ml of injectable liquid, in combinations with

b) a dose of 200 mg/m ² (l) form of leucovorin and a dose of 2400 mg/m ² 5-fluorouracil,

for the treatment of exocrine pancreatic cancer in a patient, wherein the irinotecan liposomal composition comprises a sucrose octasulfate salt of irinotecan encapsulated in irinotecan liposomes consisting of phosphatidylcholine, cholesterol and polyethylene glycol modified phosphatidylethanolamine and having a diameter of approximately 80-140 nm, wherein the diluted injectable formulation Irinotecan, leucovorin and 5-fluorouracil are administered to the patient sequentially, starting from the first day of the two-week cycle.

The invention may further include storing the diluted irinotecan injectable formulation at a temperature of approximately 2-8°C for no more than 24 hours prior to administration.

In a preferred embodiment, polyethylene glycol modified phosphatidylethanolamine is present in irinotecan liposomes in an amount of approximately one polyethylene glycol molecule for every 200 phospholipid molecules.

In one embodiment, the patient has previously been unsuccessfully treated with gemcitabine or has acquired resistance to gemcitabine prior to administration of the irinotecan diluted injectable formulation.

According to one embodiment of the invention, a dose of 200 mg/m ² (l) form of leucovorin is provided by administering a dose of 400 mg/m ² (l+d) form of leucovorin.

According to the invention, the dose of the liposomal irinotecan composition is 60 mg/ ^m2 , 70 mg/ ^m2 or 80 mg/ ^m2 , preferably the dose of the liposomal irinotecan composition is 80 mg/ ^m2 , and the human patient is not homozygous for the UGT1A1* allele 28.

In one embodiment, the liposomal irinotecan composition is administered as a diluted irinotecan injection formulation as a 90-minute infusion, leucovorin is administered after irinotecan for 30 minutes, and 5-fluorouracil is administered within 46 hours of leucovorin.

Preferably, irinotecan liposomes are monolamellar lipid bilayer vesicles that encapsulate an aqueous space containing irinotecan in a gelled or precipitated state as a sucrose octasulfate salt.

Preferably, the injectable liquid is 5% dextrose injection solution.

In addition, the invention relates to a method of treating pancreatic cancer in a human patient whose cancer has progressed after therapy with gemcitabine, comprising administering antitumor therapy to the human patient once every two weeks, wherein the antitumor therapy consists of intravenously administering to the patient human: a diluted injectable formulation of irinotecan providing a dose of 60, 70 or 80 mg/m ² liposomal irinotecan, in combination with a dose of 200 mg/m ² (l) form of leucovorin and a dose of 2400 mg/m ² 5-fluorouracil for the treatment of pancreatic cancer in a human patient, where

a) the liposomes in the diluted irinotecan injectable formulation have a diameter of approximately 80-140 nm, and consist of phosphatidylcholine, cholesterol and polyethylene glycol modified phosphatidylethanolamine; And

b) the diluted irinotecan injection formulation contains irinotecan liposomes in 500 ml of injectable liquid, stored at a temperature of approximately 2-8°C for no more than 24 hours before each administration.

According to the above method, a dose of 80 mg/ ^m2 liposomal irinotecan is administered to a human patient who is not homozygous for the UGT1A1*28 allele, and a dose of 200 mg/ ^m2 (l) form of leucovorin is provided by administering a dose of 400 mg/ ^m2 (l+ d) forms of leucovorin.

The method may further comprise administering an antiemetic to the patient prior to administering anticancer therapy; wherein liposomal irinotecan is administered as part of a diluted injectable irinotecan formulation as a single 90-minute infusion, a dose of leucovorin is administered after liposomal irinotecan for 30 minutes, and a dose of 5-fluorouracil is administered within 46 hours of leucovorin.

Preferably, irinotecan liposomes are monolamellar lipid bilayer vesicles that encapsulate an aqueous space containing irinotecan in a gelled or precipitated state as a sucrose octasulfate salt.

In this case, the injected liquid is a 5% dextrose injection solution.

The invention further includes a method for treating metastatic pancreatic adenocarcinoma after disease progression following gemcitabine therapy in a human patient who is not homozygous for the UGT1A1*28 allele, comprising the following steps, administered once every two weeks:

a) diluting a volume of 5 mg/ml of the irinotecan liposomal composition in 500 ml of injectable liquid to obtain a diluted irinotecan injectable formulation; then

b) optionally, storing the diluted irinotecan injection formulation at a temperature of approximately 2-8°C for no more than 24 hours prior to administration; then

c) administering antineoplastic therapy to a human patient, wherein the antineoplastic therapy consists of intravenously administering to the human patient: a diluted injectable formulation of irinotecan providing a dose of 80 mg/m ² liposomal irinotecan, in combination with a dose of 200 mg/m ² (l) form leucovorin and a dose of 2400 mg/m ² 5-fluorouracil, for the treatment of pancreatic cancer in a human patient;

wherein the liposomes in the irinotecan liposomal composition have a diameter of approximately 80-140 nm, and contain a monolamellar lipid bilayer vesicle that encapsulates an aqueous space containing irinotecan in a gelled or precipitated state as a sucrose octasulfate salt, wherein the monolamellar lipid bilayer consists of phosphatidylcholine, cholesterol and phosphatidylethanolamine modified with polyethylene glycol.

In a preferred embodiment, the injectable liquid is 5% dextrose injection solution.

Additionally, the invention provides an injectable liposomal form of irinotecan-sucrose octasulfate for co-administration to a patient with a tumor: 1) an amount of an injectable liposomal form of the irinotecan-sucrose octasulfate drug every two weeks effective to increase tumor vascularization, and 2) an effective amount of at least one anticancer drug agent that is not irinotecan.

BRIEF DESCRIPTION OF THE DRAWINGS

Figure 1 is a graph showing the antitumor activity of MM-398 in an orthotopic pancreatic tumor model expressing luciferase (L3.6pl).

Figure 2 is a graph showing the accumulation of SN-38 in a tumor after treatment with free irinotecan or liposomal irinotecan (MM-398).

Figure 3 is a graph showing the effect of MM-398 on carbonic anhydrase IX staining in the HT29 xenograft model.

Figure 4 shows the effect of MM-398 on low molecular weight Hoechst dye perfusion.

Figure 5 provides a summary of the pharmacokinetics of MM-398 in q3w (irinotecan, liposomal + free drug).

Figure 6 provides a summary of the pharmacokinetics of MM-398 in q3w.

Figure 7 is a schematic representation of the phase 3 study design.

DETAILED DESCRIPTION OF THE INVENTION

I. Definitions

As used herein, the term “subject” or “patient” is a person with cancer.

As used herein, “effective treatment” refers to treatment that produces a beneficial effect, such as improving at least one symptom of a disease or disorder. The positive effect may take the form of an improvement over baseline, i.e. improvements compared to the measurement or observation made before the start of treatment in accordance with the specified method. The beneficial effect may also take the form of stopping, slowing, delaying or stabilizing the harmful progression of a cancer marker. Effective treatment may relate to alleviation of at least one symptom of cancer. Such effective treatment may, for example, reduce a patient's pain, reduce the size and/or number of lesions, may reduce or prevent metastasis of a cancerous tumor, and/or may slow the growth of a cancerous tumor.

The term "effective amount" refers to the amount of an agent that provides the desired biological, therapeutic and/or prophylactic result. This result may be a reduction, improvement, alleviation, reduction, retardation and/or mitigation of one or more signs, symptoms or causes of a disease, or any other desired change in a biological system. In the context of cancer, an effective amount includes an amount sufficient to cause tumor shrinkage and/or reduce the rate of tumor growth (eg, suppress tumor growth) or prevent or delay other undesirable cell proliferation. In some embodiments, an effective amount is an amount sufficient to delay tumor development. In some embodiments, an effective amount is an amount sufficient to prevent or delay tumor recurrence. An effective amount may be administered in one or more doses. An effective amount of the drug or composition can: (i) reduce the number of cancer cells; (ii) reduce tumor size; (iii) inhibit, delay, slow down to some extent and stop the infiltration of cancer cells into peripheral organs; (iv) inhibit (ie, slow down to some extent and stop) tumor metastasis; (v) inhibit tumor growth; (vi) prevent or cause a delay in the onset and/or recurrence of a tumor; and/or (vii) relieve to some extent one or more symptoms associated with cancer.

The terms "combination therapy", "co-administration", "co-administered" or "simultaneous administration" (or these terms with minor variations) include the simultaneous administration of at least two therapeutic agents to a patient or their sequential administration during the period of time when the first one administered the therapeutic agent is still present in the patient upon administration of the second therapeutic agent administered.

The term “monotherapy” refers to the administration of a single drug to treat a disease or disorder in the absence of coadministration of any other therapeutic agent to treat the same disease or disorder.

"Dosage" refers to the parameters of administering a drug in specified quantities per unit of time (eg, per hour, day, week, month, etc.) to a patient. Such parameters include, for example, the size of each dose. Such parameters also include the configuration of each dose, which may be administered in one or more units, e.g., taken in a single administration, e.g., orally (e.g., one, two, three or more tablets, capsules, etc.) or by injection (eg, as a bolus). Dosage sizes may involve doses administered continuously (eg, as an intravenous infusion over several minutes or hours). Such parameters further include the frequency of administration of individual doses, which may vary over time.

"Dose" refers to the amount of drug taken in a single administration.

As used herein, “cancer” refers to a condition characterized by abnormal, unregulated, malignant cell growth. In one embodiment, the cancer is exocrine pancreatic cancer. In yet another embodiment, the exocrine pancreatic cancer is selected from the group consisting of acinar cell carcinoma, adenocarcinoma, glandular squamous cell carcinoma, giant cell tumor, intraductal papillary mucinous neoplasm (IPMN), mucinous cystadenocarcinoma, pancreatoblastoma, serous cystadenocarcinoma, and solid and pseudopapillary tumors .

The terms "resistant" and "resistant" refer to tumor cells that survive treatment with a therapeutic agent. Such cells may initially respond to the therapeutic agent, but subsequently demonstrate a decreased response during treatment, or fail to demonstrate an adequate response to the therapeutic agent in the sense that the cells continue to proliferate during treatment with the agent.

II. Injectable liposomal form of irinotecan - sucrose sulfate (MM-398; PEP02)

Herein, irinotecan is administered in a stable liposomal formulation in irinotecan-sucrose sulfate injectable liposomal form (otherwise referred to as "irinotecan-sucrose octasulfate injectable liposomal form" or "irinotecan sucrose sulfate injectable liposomal form"), which formulation is referred to herein as "MM-398 " (and also PEP02, see US 8147867). MM-398 can be provided as a sterile liquid for intravenous injection. The required amount of MM-398 can be diluted, for example, in 500 ml of 5% dextrose injection USP grade and administered by infusion over 90 minutes.

The MM-398 liposome is a monolamellar lipid bilayer vesicle with a diameter of approximately 80-140 nm, which encapsulates an aqueous space containing the irinotecan complex in a gel or precipitated state as a salt with sucrose octasulfate. The lipid membrane of the liposome consists of phosphatidylcholine, cholesterol and phosphatidylethanolamine modified with polyethylene glycol in an amount of approximately one molecule of polyethylene glycol (PEG) per 200 molecules of phospholipids.

This stable liposomal formulation of irinotecan has several properties that may provide an improved therapeutic index. Controlled and sustained release improves the activity of the specified drug depending on the dosage schedule by increasing the duration of exposure of the drug to the tumor tissue; this property allows it to be present in a higher percentage of cells during the S phase of the cell cycle, when DNA unwinding is required as a precursor to DNA replication. The pharmacokinetics of prolonged exposure to the bloodstream and the high intravascular retention of drug in liposomes may contribute to the enhanced permeability and retention (EPR) effect. EPR allows liposomes to accumulate in areas such as malignant tumors where the normal integrity of blood vessels (particularly capillaries) is compromised, resulting in particles such as liposomes leaking out of the capillary lumen. EPR may thus facilitate site-specific delivery of liposomes to solid tumors. The EPR of MM-398 may lead to a subsequent depot effect in which liposomes accumulate in tumor associated macrophages (TAMs), which metabolize irinotecan, locally converting it to the significantly more toxic SN-38. This local bioactivation is thought to result in decreased drug exposure to potential sites of toxicity and increased exposure to cancer cells within the tumor.

Pharmacogenetics of irinotecan glucuronidation

The enzyme produced by the UGT1A1 gene, UDP-glucuronosyltransferase 1, is responsible for the metabolism of bilirubin and also mediates glucuronidation of SN-38, which is the first step in the predominant metabolic clearance pathway of this active metabolite of irinotecan. In addition to its antitumor activity, SN-38 is also responsible for the high toxicity sometimes associated with irinotecan therapy. Thus, glucuronidation of SN-38 to the inactive form, SN-38 glucuronide, is an important step in the regulation of irinotecan toxicity.

A mutational polymorphism of the UGT1A1 gene promoter has been described, in which there is a variable number of thymine-adenine (ta) repeats. It was found that promoters containing seven thymine adenine (ta) repeats (in the UGT1A1*28 allele) were less active than wild-type promoters containing six repeats, resulting in decreased expression of UDP-glucuronyltransferase 1. Patients carrying two defective UGT1A1 alleles show reduced glucuronidation of SN-38. In some cases, individuals homozygous for the UGT1A1*28 allele (referred to as UGT1A1 7/7 genotype individuals, since both alleles are UGT1A1*28 alleles containing 7 ta repeats, as opposed to the UGT1A1 6/6 genotype (wild type) have been reported type) in which both alleles contain 6 ta repeats) and characterized by unstable increases in serum bilirubin (eg, patients with Gilbert's syndrome) may be at greater risk of toxicity when receiving standard doses of irinotecan. This suggests an association between homozygosity for the UGT1A1*28 allele, bilirubin levels and irinotecan toxicity.

The metabolic transformation of MM-398 to SN-38 (eg, in plasma) involves two critical steps: (1) release of irinotecan from liposomes and (2) conversion of free irinotecan to SN-38. Without taking into account theoretical aspects, it is believed that once irinotecan leaves the liposomes, it undergoes catabolism through the same metabolic pathways as regular (free) irinotecan. Therefore, such human genetic polymorphisms, which are predictive of toxicity and efficacy, can be considered similar for irinotecan and MM-398. However, due to the lower tissue distribution, reduced clearance, higher systemic exposure, and longer half-life of SN-38 in MM-398 compared to free irinotecan, defective genetic polymorphisms may show a greater association with severe adverse events and /or efficiency.

Patients with reduced UGT1A1 activity

It has been shown that individuals homozygous for the UGT1A1*28 allele (UGT1A1 7/7 genotype) are characterized by an increased risk of neutropenia after initiation of treatment with irinotecan. According to the prescribing information for irinotecan (Camptosar®), in a study of 66 patients receiving irinotecan monotherapy (350 mg/m ² times every 3 weeks), the incidence of grade 4 neutropenia in patients homozygous for the UGT1A1*28 allele was 50 %, and in patients heterozygous for this allele (UGT1A1 6/7 genotype) the incidence was 12.5%. Importantly, grade 4 neutropenia was not observed in patients homozygous for the wild-type allele (UGT1A1 6/6 genotype). Other studies have described a lower prevalence of life-threatening neutropenia. For this reason, in patients recruited into the phase 3 study described in the Examples section of this document and homozygous for the UGT1A1*28 allele (UGT1A1 7/7 genotype), treatment with MM-398 was initiated at a lower dose than that of patients with one (eg, UGT1A1 6/7) or two (UGT1A1 6/6) wild-type alleles.

Additional genotypic modifiers of irinotecan metabolism

Although the UGT1A1*28 allele is relatively common in Caucasians (estimated at 10%), its prevalence varies among other ethnic groups. In addition, additional UGT1A1 genotypes have been found with higher prevalence, for example in Mongoloid populations; this may have important implications for irinotecan metabolism in these populations. For example, the UGT1A1*6 allele is more common among Mongoloids. This allele is not associated with ta repeats, but with the Gly71Arg mutation, which reduces enzyme activity. Previous and ongoing studies of MM-398 collected pharmacogenetic data in recruited patients. In the study entitled PEP0203, a clear relationship of genetic polymorphisms of the UGT1A family and DPYD (dihydropyrimidine dehydrogenase, an enzyme associated with 5-FU catabolism) with the pharmacokinetic parameters and toxicity of MM-398 was not established due to the small sample size of the subjects evaluated. However, it was noted that patients with the combined UGT1A1*6/*28 polymorphism had an increased dose-normalized SN-38 AUC and underwent DLT.

III. 5-fluorouracil (5-FU) and leucovorin

5-fluorouracil is a pyrimidine antagonist that affects the biosynthesis of nucleic acids. The deoxyribonucleotide of this drug inhibits thymidylate synthetase, thereby inhibiting the formation of thymidylic acid from deoxyuridylic acid and interfering with DNA synthesis. It also interferes with RNA synthesis.

Leucovorin (also called folinic acid) acts as a biochemical cofactor for 1-carbon transfer reactions in the synthesis of purines and pyrimidines. Leucovorin does not require the enzyme dihydrofolate reductase (DHFR) to convert to tetrahydrofolic acid. Leucovorin inhibits the effects of methotrexate and other DHFR antagonists. Leucovorin may enhance the cytotoxic effects of fluorinated pyrimidines (i.e. fluorouracil and floxuridine). Once 5-FU is activated in the cell, it is accompanied by a folate cofactor and inhibits the enzyme thymidylate synthetase, thereby inhibiting pyrimidine synthesis. Leucovorin increases the folate pool, thereby enhancing the binding of folate cofactor and active 5-FU to thymidylate synthetase.

Leucovorin has dextro- and levo-isomers, only the latter of which is suitable for pharmacological purposes. Therefore, the use of the bioactive levo-isomer (“levoleucovorin”) has also been approved by the FDA for the treatment of cancer. The dosage of levoleucovorin is usually half that of a racemic mixture containing dextro-(d) and levo-(l) isomers.

FU and leucovorin should be handled and stored in accordance with the country-specific package insert.

IV. Introduction

Liposomal irinotecan is administered intravenously, alone or in combination with 5-fluorouracil (5-FU) and/or leucovorin. In one embodiment, liposomal irinotecan is administered prior to 5-FU and leucovorin. In yet another embodiment, leucovorin is administered before 5-FU. In yet another embodiment, liposomal irinotecan is administered intravenously over 90 minutes. In yet another embodiment, 5-FU is administered intravenously over 46 hours. In yet another embodiment, leucovorin is administered intravenously over 30 minutes. In various embodiments, the liposomal irinotecan is MM-398.

V. Patient populations

In one embodiment, a patient receiving treatment using the methods and compositions described herein exhibits evidence of recurrent or recurrent pancreatic cancer following primary chemotherapy.

In yet another embodiment, the patient has a history of unsuccessful treatment of the underlying or recurrent disease with at least one platinum-based chemotherapy regimen, for example, a chemotherapy regimen comprising carboplatin, cisplatin, or another organic platinum compound.

In a further embodiment, the patient has previously received treatment with gemcitabine without success or has acquired resistance to gemcitabine.

In one embodiment, a resistant or resistant tumor is a tumor in which the treatment-free interval for patients with the tumor is less than 6 months (eg, due to cancer recurrence) or in which the tumor progresses during a course of treatment.

In yet another embodiment, the pancreatic cancer in the patients being treated is an advanced pancreatic cancer, which is a pancreatic tumor exhibiting distant metastasis and/or peripancreatic tumor proliferation.

The compositions and methods described herein can be used to treat all types of pancreatic cancer, including pancreatic cancer that is refractory or refractory to other anticancer treatments.

VI. Combination therapy

In one embodiment, liposomal irinotecan is administered to patients with pancreatic cancer along with 5-fluorouracil (5-FU) and leucovorin according to a specific clinical dosing regimen, such as that described herein. In one embodiment, the liposomal irinotecan is MM-398.

As used herein, additional or combined administration (co-administration) includes simultaneous administration of compounds in the same or different dosage forms, or separate administration of compounds (eg, sequential administration). For example, liposomal irinotecan can be administered simultaneously with 5-FU and leucovorin. Alternatively, liposomal irinotecan can be administered in combination with 5-FU and leucovorin, wherein liposomal irinotecan, 5-FU and leucovorin are formulated for separate administration and are administered simultaneously or sequentially. For example, liposomal irinotecan may be administered first, followed (eg, immediately afterwards) by 5-FU and leucovorin. Such parallel or sequential administration has the advantage that liposomal irinotecan, 5-FU and leucovorin are simultaneously present in the body of patients receiving treatment. In a specific embodiment, liposomal irinotecan is administered prior to 5-FU and leucovorin. In yet another specific embodiment, leucovorin is administered before 5-FU.

In yet another embodiment, liposomal irinotecan, 5-FU and leucovorin are formulated for intravenous administration. In a specific embodiment, the patient is administered an effective amount of each of the following: liposomal irinotecan, 5-fluorouracil (5-FU), and leucovorin, the treatment comprising at least one cycle, where the cycle is a period of 2 weeks, and for each cycle: (a) liposomal irinotecan is administered on day 1 of the cycle at a dose of 80 mg/ ^m2 , unless the patient is homozygous for the UGT1A1*28 allele; in this case, liposomal irinotecan is administered on day 1 of cycle 1 at a dose of 60 mg/ ^m2 ; (b) 5-FU is administered at a dose of 2400 mg/m ² ; and (c) leucovorin is administered at a dose of 200 mg/m ² (l-form) or 400 mg/m ² (l+d-racemic form). In a specific embodiment, the dose of liposomal irinotecan administered to a patient homozygous for the UGT1A1*28 allele is increased after one cycle to 80 mg/m ² .

In one embodiment, liposomal irinotecan may be administered at a high dose initially and the dose reduced over time. In yet another embodiment, liposomal irinotecan is initially administered at a low dose and the dose is increased over time. In one embodiment, liposomal irinotecan is administered as monotherapy.

In yet another embodiment, the dose of 5-FU is varied over time. For example, 5-FU can be administered at a high dose initially and the dose reduced over time. In yet another embodiment, 5-FU is initially administered at a low dose and the dose is increased over time.

In yet another embodiment, the dose of leucovorin is varied over time. For example, leucovorin can be given at a high dose initially and the dose reduced over time. In yet another embodiment, leovorin is administered at a low dose initially and the dose is increased over time.

VII. Treatment protocols

Suitable treatment protocols include, for example, protocols where the patient is administered an effective amount of liposomal irinotecan, the treatment comprising at least one cycle equal to a 3-week period, and in each cycle liposomal irinotecan is administered on the first day of the cycle at a dose of 120 mg/m ² , except if the patient is homozygous for the UGT1A1*28 allele; in this case, liposomal irinotecan is administered on day 1 of cycle 1 at a dose of 80 mg/ ^m2 . In one embodiment, the dose of liposomal irinotecan administered to a patient homozygous for the UGT1A1*28 allele is increased after one in increments of 20 mg/m ² , but not to exceed 120 mg/m ² .

In yet another embodiment, the treatment protocol includes administering to a patient an effective amount of each of the following: liposomal irinotecan, 5-fluorouracil (5-FU), and leucovorin, the treatment comprising at least one cycle, wherein a cycle is a period of 2 weeks, and for each cycle: (a) liposomal irinotecan is administered on day 1 of the cycle at a dose of 80 mg/ ^m2 , unless the patient is homozygous for the UGT1A1*28 allele; in this case, liposomal irinotecan is administered on day 1 of cycle 1 at a dose of 60 mg/ ^m2 ; (b) 5-FU is administered at a dose of 2400 mg/m ² ; and (c) leucovorin is administered at a dose of 200 mg/m ² (l-form) or 400 mg/m ² (l+d-racemic form). In a specific embodiment, the dose of liposomal irinotecan administered to a patient homozygous for the UGT1A1*28 allele is increased after one cycle to 80 mg/m ² .

VIII. results

The present invention provides methods for treating pancreatic cancer in a patient, comprising administering to the patient liposomal irinotecan (MM-398) alone or in combination with 5-fluorouracil (5-FU) and leucovorin, according to a specific clinical dosage regimen.

Preferably, when combined therapy using liposomal irinotecan with 5-FU and leucovorin, therapeutic synergy is observed.

“Therapeutic synergy” refers to the phenomenon in which treating patients with a combination of therapeutic agents results in therapeutically superior results compared to those achieved by each individual component of the combination given at the optimal dose (T. H. Corbett et al., 1982, Cancer Treatment Reports, 66, 1187). In this context, a therapeutically improved outcome is one in which patients: a) exhibit a lower incidence of adverse events while receiving a therapeutic benefit that is equal to or greater than the effect obtained when each of the individual components of the combination is administered as monotherapy at the same dose as and in combination, or b) do not exhibit dose-limiting toxicity while producing a therapeutic benefit greater than that obtained when each individual component of the combination is administered at the same doses as in the combination(s). In xenograft models, when the combination is administered at its maximum tolerated dose, with each component present at a dose typically not exceeding its own maximum tolerated dose, therapeutic synergy is observed if the combination achieves a reduction in tumor growth greater than the best reduction value. tumor growth with the introduction of individual components.

Thus, when combined, the components of such combinations have additive or superadditive effects in suppressing pancreatic tumor growth compared to monotherapy with liposome-encapsulated irinotecan alone or treatment with chemotherapeutic agent(s) in the absence of liposomal irinotecan. The term “additive” means a result that is greater (for example, in terms of the degree of reduction in tumor mitotic index or tumor growth or the degree of reduction in tumor size or the frequency and/or duration of symptom-free or symptom-reducing periods) than the best individual result achieved by monotherapy using each individual component, and the term "superadditive" is used to denote a result greater than the sum of such individual results. In one embodiment, the additive effect is measured as slowing or stopping the growth of a pancreatic tumor. The additive effect can also be measured as, for example, a decrease in pancreatic tumor size, a decrease in tumor mitotic index, a decrease in the number of metastases over time, an increase in overall response rate, and an increase in median or overall survival.

One non-limiting example of a possible quantitative indicator of the effectiveness of therapeutic treatment is the calculation of the tenth logarithm of cell killing, determined by the following formula:

lg cell destruction = TC(days)/3.32×Td

where TC is cell growth inhibition, i.e. the average time (in days) for tumors in the treated group (T) and tumors in the control group (C) to reach a given value (for example, 1 g, or 10 ml), and Td is the time (in days), required to double tumor volume in control animals. When using this indicator, a product is considered active if the cell killing lg is greater than or equal to 0.7, and very active if the cell killing lg exceeds 2.8. When applied to a combination used at its maximum tolerated dose, with each component present at a dose typically not exceeding its own maximum tolerated dose, therapeutic synergy is observed if the 10 lg cell kill exceeds the best 10 lg cell kill of the individual components. . Typically, the 10 lg cell kill for the combination exceeds the best value of 10 lg cell kill when administering the individual component of the combination by at least 0.1 lg cell kill, at least 0.5 lg cell kill, or at least 1.0 lg cell destruction.

Response to treatment may include:

Pathological complete response (pCR): absence of invasive breast and lymph node cancer after primary systemic treatment.

complete response (CR): disappearance of all target lesions. Any pathological lymph nodes (target or non-target) reduced in short axis by <10 mm;

partial response (PR): at least 30% reduction in the sum of target lesion sizes compared to the initial sum of diameters;

stable disease (SD): absence of both a significant decrease in size consistent with partial response and a significant increase consistent with progressive disease compared with the smallest sum of diameters recorded during the study; or

wherein non-CR/non-PD means persistence of one or more non-target lesions and/or maintenance of tumor marker levels above normal.

Progressive disease (PD) refers to an increase in the sum of the target lesion sizes by at least 20% compared with the smallest sum recorded during the study (including the original sum if it is the smallest during the study). In addition to the relative increase of 20%, there must be an absolute increase in the amount of 5 mm. The appearance of one or more new lesions is also considered progression.

In a typical outcome, patients treated in accordance with the methods described herein may experience improvement in at least one of the symptoms of pancreatic cancer.

In one embodiment, a patient receiving such treatment experiences pCR, CR, PR, or SD.

In yet another embodiment, a patient receiving such treatment experiences a reduction in tumor size and/or a reduction in growth rate, i.e. suppression of tumor growth. In yet another embodiment, unwanted cell proliferation is reduced or inhibited. In yet another embodiment, one or more of the following may occur: the number of cancer cells may decrease; tumor size may decrease; Inhibition, delay, slowing or stopping of the penetration of cancer cells into peripheral organs may occur; tumor metastasis may be slowed down or inhibited; tumor growth may be inhibited; prevention or delay of tumor recurrence can be achieved; Some relief of one or more cancer-related symptoms may occur.

In other embodiments, such improvement is measured by a reduction in the number and/or size of measurable tumor lesions. Measurable lesions are defined as lesions that can be accurately measured in at least one dimension (with maximum diameter recorded) as >10 mm as measured by computed tomography (using a CT slice no thicker than 5 mm), 10 mm as measured with calipers on clinical examination or >20 mm on chest x-ray. As part of the enhancement, it is also possible to measure the size of non-target lesions, such as abnormal lymph nodes. In one embodiment, lesions can be measured on chest radiographs or CT or MRI films.

In other embodiments, cytology and histology may be used to assess response to treatment. When differentiating response or stable disease from progressive disease, cytologic confirmation of neoplastic origin of any discharge that appears or worsens during treatment should be considered if measurable tumors meet criteria for response or stable disease (discharge may be a side effect of treatment).

In some embodiments, administration of an effective amount of liposomal irinotecan, 5-FU, and leucovorin according to any of the methods provided herein results in at least one therapeutic effect selected from the group consisting of reduction in breast tumor size, reduction in the number of metastatic sites occurring over time, complete remission, partial remission, stable disease, increasing rate of overall response, or pathological completed response. In some embodiments, the proposed treatments result in a comparable degree of clinical benefit (CBR=CR+PR+SD ≥6 months) better than that achieved by the same combination of anticancer agents administered without concomitant administration of MM-398. In other embodiments, the improvement in the rate of clinical benefit is approximately 20%, 20%, 30%, 40%, 50%, 60%, 70%, 80% or more compared to the same combinations of anticancer agents administered without concomitant administration of MM -398.

The following examples are illustrative and should in no way be construed as limiting the scope of the present description; Various modifications and equivalent variations to what has been described will become apparent to those skilled in the art upon reading the present disclosure.

EXAMPLES

Example 1: Activity of MM-398 in an orthotopic pancreatic tumor model expressing luciferase (L3.6pl)

The antitumor activity of MM-398 was assessed in the orthotopic model of pancreatic cancer (L3.6pl), a highly hypoxic preclinical tumor model. Approximately 2.5 x 10-5 L3.6pl pancreatic tumor cells were implanted by direct injection into the pancreas. Tumor burden/score was monitored over time using bioluminescence imaging (BLI). MM-398 and free irinotecan were administered at a dose of 20 mg/kg/week for three weeks. As shown in Figure 1, MM-398 (liposomal CPT11) had significant antitumor activity compared with control (HBS) and free CPT11.

Example 2: Accumulation of SN-38 in Tumors Following Treatment with Free Irinotecan or Liposomal Irinotecan (MM-398)

It was hypothesized that the antitumor activity observed in an orthotopic model of pancreatic cancer was due to the local action of macrophages to convert irinotecan to the more active SN-38. To test this assumption, human colon cancer cells (HT-29) were injected subcutaneously into SCID mice, and after tumors reached a size of 1000 mm ³ , 40 mg/kg free irinotecan or MM-398 was administered intravenously. Mice with tumors were sacrificed at different time points, tumors were removed from animals in both groups, and SN-38 concentrations were measured.

As shown in Figure 2, a 20-fold increase in AUCSN-38 was observed in the tumor for MM-398 compared to free irinotecan. The duration of exposure allowed for prolonged exposure of slowly proliferating cancer cells to the active metabolite throughout the cell cycle. In addition, it has been suggested that this activity is also due to a decrease in hypoxia within the tumor and a subsequent suppressive effect on angiogenesis, metastasis and immunosuppressive conditions in the tumor.

Example 3: Effect of MM-398 on Carbonic Anhydrase IX Staining in the HT29 Xenograft Model

To test the effect of MM-398 on reducing hypoxia markers, experiments were performed on a human colon cancer cell model (HT-29). Specifically, HT-29 cells were injected subcutaneously into athymic nude mice and on day 13, PBS control or 1.25, 2.5, 5, 10, or 20 mg/kg MM-398 were administered intravenously. MM-398 was administered once a week for 4 weeks at the prescribed doses. Tumors from both groups of animals (n=5) were removed 24 hours after the last dose. Frozen tumor sections were used for carbonic anhydrase IX (CAIX) immunohistochemical staining. Quantification of CAIX staining was performed using Definiens® software (Definiens AG, Munich).

As shown in Figure 3, MM-398 reduced the levels of hypoxia markers. Specifically, the graphs in Figure 3 show the percentage of cells with medium (middle third) or high (top third) intensity of CAIX staining. Representative samples from each group are shown, along with the group mean (mean +/- standard deviation). Treatment with MM-398 altered the tumor microenvironment, reducing the percentage of intermediate- and high-CAIX positive cells in a dose-dependent manner. Since hypoxia is a hallmark of persistent and aggressive disease; Reducing hypoxia is expected to increase the sensitivity of tumor cells to chemotherapy.

Example 4: MM-398 Increases Hoechst Dye Perfusion

In addition to altering the sensitivity of tumor cells to chemotherapy by modifying the tumor microenvironment, decreased hypoxia may indicate improved tumor vascularization, which may facilitate delivery of small molecule therapeutics. Treatment with MM-398 resulted in increased microvascular density 6 days after treatment as measured by CD31 (endothelial cell-platelet adhesion molecule) staining in the HT29 xenograft study. To further evaluate the effect of MM-398 on low molecular weight tumor vascularization, a Hoechst 33342 dye perfusion experiment was performed. Specifically, a primary pancreatic tumor was grown in a NOD-SCID mouse and a single dose of MM-398 (20 mg/kg) was administered. After 24 hours, Hoechst 33342 dye was injected, and the animal was sacrificed 20 minutes later. As shown in Figure 4, the increase in staining intensity in treated mice was statistically significant, p<0.001. These data indicate that MM-398 altered the tumor microenvironment, increasing tumor susceptibility to agents such as 5-FU/LV by reducing tumor hypoxia and increasing small molecule perfusion.

Example 5: Pharmacokinetics of MM-398 in the human body (Phase I)

The pharmacokinetic profile of MM-398 when used as monotherapy was studied in a phase I clinical study (PEP0201) in patients at dose levels of 60, 120 or 180 mg/ ^m2 and in a phase II clinical study in patients with gastric cancer (PEP0206) at 120 mg. / ^m2 . These studies measured plasma levels of total irinotecan, SN-38, and encapsulated irinotecan.

Peak serum total irinotecan concentrations ( _Cmax ) for 120 mg/m ² MM-398 were in the range of 48-79 μg/ml, which was approximately 50 times higher than for 125 mg/m ² free irinotecan. The half-life of total irinotecan (t _1/2 ) for MM-398 ranged from 21 to 48 hours, which was approximately 2-3 times higher than for 125 mg/m ² free irinotecan. Overall, one-week exposure to total irinotecan (AUC 0-T) at a dose of 120 mg/ ^m2 MM-398 ranged from 1200-3000 (mcg*h/mL), which was approximately 50-100 times higher than for 300 mg/m ² free irinotecan. In contrast, SN38 C _max levels at 120 mg/m ² MM-398 ranged from 9 to 17 ng/ml, which was approximately 50% less than at 125 mg/m ² free irinotecan. Overall, one-week SN38 exposures (AUC 0-T) ranged from 474 to 997 ng/mL and were only 1-2 times higher than 300 mg/m ² free irinotecan. For both SN38 and total irinotecan, the AUC increased less than proportionally to the MM-398 dose. The PK parameters of encapsulated irinotecan were almost identical to those of total irinotecan, indicating that most of the irinotecan remained encapsulated in liposomes during circulation. PK parameters of MM-398 were not significantly changed in combination with 5-FU/LV. Figures 5 and 6 provide summary information on the PK results of previous studies of MM-398.

Example 6: Phase 1 Dose Escalation Study

A regimen using a combination of fluorouracil, leucovorin and MM-398 was studied in a phase 1 study of solid tumors in 16 subjects, of which 5 were patients with pancreatic cancer. Objective tumor response rate, duration of response, and disease control rate were the outcome measures of efficacy in this study. Among 15 patients in whom efficacy could be assessed, PR was confirmed in 2 (13.3%), SD was observed in 9 (60.0%), and PD was observed in 4 (26.7%). The overall disease control rate was 73.3%. A partial response was observed in one patient with gastric cancer (at the 80 mg/m ² dose level) and one patient with breast cancer (at the 100 mg/m ² dose level) with response durations of 142 and 76 days, respectively. Among 6 patients who received an MTD dose of 80 mg/ ^m2 , 1 PR, 4 SD and 1 PD were observed. The tumor response rate and disease control rate were 16.7% and 83.3%, respectively. The major DLTs were grade 3 diarrhea, leukopenia, neutropenia, and febrile neutropenia. The MTD for MM-398 was 80 mg/ ^m2 .

In the phase 1 dose-escalation study of MM-398 in combination with 5-FU/LV in advanced solid tumors (PEP0203), a total of 401 AEs were reported in 16 treated subjects (safety study population), of which 74 (18.4%) were CTC category 3 or higher. Among all AEs, according to the investigators, 231 AEs (57.6%) were related to treatment. The most common treatment-related AEs included nausea (81.3%), diarrhea (75.0%), vomiting (68.8%), fatigue (43.8%), mucositis (43.8%), leukopenia (37.5%), neutropenia (37.5%), weight loss (37.5%), anemia (31.3%) and alopecia (31.3%). Acute cholinergic diarrhea has been observed rarely. Table 1 presents the incidence of treatment-emergent adverse events by maximum CTC category and causality (incidence ≥ 20%) in the PEP0203 study. Table 2 shows the incidence of category 3 or higher treatment-emergent adverse events observed in 5 patients with pancreatic cancer treated in the PEP0203 study.

Table 1
Incidence of treatment-emergent adverse events by maximum CTC category and causality (incidence ≥20%) in study PEP0203 Organ system class
Preferred term Total
(N=16) Severity (category) ¹ Causality ² I II III IV Yes No Blood and lymphatic system disorders Anemia 7 (43.8%) 3 2 2 0 5 2 Leukopenia 6 (37.5%) 0 3 2 1 6 0 Neutropenia 6 (37.5%) 0 2 3 1 6 0 Gastrointestinal disorders Abdominal pain 7 (43.8%) 3 2 2 0 3 4 Constipation 6 (37.5%) 3 3 0 0 0 6 Diarrhea 12 (75.0%) 3 4 5 0 12 0 Nausea 13 (81.3%) 6 6 1 0 13 0 Vomit 12 (75.0%) 3 8 1 0 eleven 1 General and administration site disorders Fatigue 8 (50.0%) 4 3 1 0 7 1 Inflammation of the mucous membrane 7 (43.8%) 4 3 0 0 7 0 Fever 7 (43.8%) 3 4 0 0 2 5 Infections and infestations Infection 6 (37.5%) 0 3 3 0 2 4 Laboratory and instrumental data Increased ALT 5 (31.3%) 3 2 0 0 4 1 Increased AST 4 (25.0%) 3 1 0 0 1 3 Weight loss 8 (50.0%) 4 4 0 0 6 2 Metabolic and nutritional disorders Anorexia 4 (25.0%) 1 2 1 0 3 1 Hypoalbuminemia 4 (25.0%) 0 3 1 0 0 4 Hypocalcemia 5 (31.3%) 1 4 0 0 0 5 Hypokalemia 8 (50.0%) 2 0 5 1 2 6 Hyponatremia 4 (25.0%) 2 0 0 2 0 4 Nervous system disorders Dizziness 4 (25.0%) 4 0 0 0 1 3 Mental disorders Insomnia 4 (25.0%) 4 0 0 0 1 3 Disorders of the respiratory system, chest and mediastinal organs Cough 5 (31.3%) 3 1 1 0 0 5 Disorders of the skin and subcutaneous tissues Alopecia 5 (31.3%) 5 0 0 0 5 0

¹ : Severity classification uses the maximum grade ever recorded for each subject reporting that adverse event.

² : Defined as a subject who has ever experienced an AE related to the study drug or not

table 2
Incidence of category 3 or higher treatment-emergent adverse events in pancreatic cancer patients in study PEP0203 Total number N=5 60 mg/m ² N=1 80 mg/m ² N=3 120 mg/m ² N=1 Main class of organ system Preferred term n (%) n (%) n (%) n (%) - Any major organ system class - Total 3 (60.0) 0 2 (66.7) 1 (100.0) Infections and infestations - Total 3 (60.0) 0 2 (66.7) 1 (100.0) Viral hepatitis 1 (20.0) 0 1 (33.3) 0 Infection 1 (20.0) 0 0 1 (100.0) Pneumonia 1 (20.0) 0 1 (33.3) 0 Septic shock 1 (20.0) 0 1 (33.3) 0 Blood and lymphatic system disorders - Total 2 (40.0) 0 1 (33.3) 1 (100.0) Lymphopenia 1 (20.0) 0 0 1 (100.0) Neutropenia 1 (20.0) 0 1 (33.3) 0 Leukocyte disorders 1 (20.0) 0 0 1 (100.0) Gastrointestinal disorders - Total 2 (40.0) 0 1 (33.3) 1 (100.0) Diarrhea 2 (40.0) 0 1 (33.3) 1 (100.0) Abdominal pain 1 (20.0) 0 0 1 (100.0) Gastrointestinal bleeding 1 (20.0) 0 1 (33.3) 0 Laboratory and instrumental data - Total 2 (40.0) 0 1 (33.3) 1 (100.0) Increased bilirubin in the blood 1 (20.0) 0 1 (33.3) 0 Increased lipase 1 (20.0) 0 0 1 (100.0) Decreased neutrophil count 1 (20.0) 0 0 1 (100.0) Decreased white blood cell count 1 (20.0) 0 0 1 (100.0) Metabolic and nutritional disorders - Total 2 (40.0) 0 1 (33.3) 1 (100.0) Hypoalbuminemia 1 (20.0) 0 1 (33.3) 0 Hypokalemia 1 (20.0) 0 1 (33.3) 0 Hyponatremia 1 (20.0) 0 0 1 (100.0) Hypophosphatemia 1 (20.0) 0 0 1 (100.0) Disorders of the respiratory system, chest and mediastinal organs - Total 2 (40.0) 0 1 (33.3) 1 (100.0) Dyspnea 1 (20.0) 0 0 1 (100.0) Pleural effusion 1 (20.0) 0 1 (33.3) 0 General and administration site disorders - Total 1 (20.0) 0 0 1 (100.0) Death 1 (20.0) 0 0 1 (100.0)

Example 7: Phase 3 Study

Promising efficacy and safety data from the phase 1 study (described above) warranted further investigation of the combination of MM-398, 5-FU and leucovorin in a phase 3 study.

A. Goals

The primary objective of the phase 3 study was to compare overall survival after treatment with MM-398 with or without 5-FU and leucovorin compared with 5-FU and leucovorin in patients with metastatic pancreatic cancer progressing on therapy with use of gemcitabine. Additional goals included:

compare time-to-event endpoints between intervention and control groups (i.e., progression-free survival (PFS) and time to treatment failure (TTF));

- compare the objective response rate (ORR) in the treatment groups;

- compare the response of the tumor marker CA 19-9 in the treatment groups;

- compare the clinical response rate (CBR) in the treatment groups;

- evaluate patient-reported outcomes (PROs) in treatment groups using the European Organization for Research and Treatment of Cancer (EORTC) Quality of Life Core Questionnaire (EORTC-QLQ-C30);

- compare the safety profile and adverse events in the treatment groups; And

- determine the pharmacokinetic properties of MM-398 when used as monotherapy and in combination with 5-FU and leucovorin.

The key objective of the present study was to examine biomarkers associated with toxicity and treatment efficacy with MM-398 and MM-398 in combination with 5-FU and leucovorin.

B. Study design

This study is an open-label, randomized, phase 3 trial of MM-398 with or without 5-FU and leucovorin versus 5-fluorouracil (5-FU) and leucovorin (also known as folinic acid) in three groups of patients with metastatic cancer. pancreas that progressed during previous therapy with gemcitabine.

This global study will enroll approximately 405 eligible patients under protocol version 2 or later. All patients will take part in a screening period of up to 28 days, during which their suitability will be assessed and the UGT1A1*28 allele will be screened. Eligible patients will be randomized in a 1:1:1 ratio to one of the following treatment groups:

Group A (experimental group): MM-398 MM 398, 120 mg/ ^m2 IV over 90 minutes every 3 weeks. Patients homozygous for the UGT1A1*28 allele will receive the first cycle of treatment at a reduced dose of 80 mg/ ^m2 . If the patient does not experience the toxic effect of the drug after the first dose of MM-398, then from cycle 2 onwards you can increase the dose in increments of 20 mg/ ^m2 to a maximum value of 120 mg/ ^m2 . Group B (control group):
5-FU and leucovorin 5-FU 2000 mg/ ^m2 IV over 24 hours (+/-30 minutes) was administered weekly for 4 weeks (on days 1, 8, 15 and 22) followed by 2 weeks of rest as part of a 6-week period cycle.
Levoleucovorin at a dose of 200 mg/ ^m2 or leucovorin (l+d-racemic mixture) at a dose of 400 mg/ ^m2 was administered intravenously over 30 minutes once a week for 4 weeks (on days 1, 8, 15 and 22 ) followed by 2 weeks of rest as part of a 6-week cycle. Group C (experimental group): MM-398, 5-FU and leucovorin MM-398 80 mg/ ^m2 IV over 90 minutes once every 2 weeks. Patients homozygous for the UGT1A1*28 allele and randomized to group C will receive the first cycle of treatment at a reduced dose of 60 mg/m ² . If the patient does not experience the toxic effect of the drug after the first dose of MM-398, then from cycle 2 onwards the dose can be increased to 80 mg/ ^m2 .
5-FU 2400 mg/ ^m2 IV over 46 hours, every 2 weeks.
Levoleucovorin at a dose of 200 mg/ ^m2 or l+d-racemic mixture at a dose of 400 mg/ ^m2 IV for 30 minutes once every 2 weeks.
MM-398 should be administered before 5-FU and leucovorin; Leucovorin should always be given before 5-FU. Other drugs should not be administered in combination if MM-398 or 5-FU/leucovorin must be stopped.

Patients will be uniformly randomized into treatment groups using a centralized interactive Internet-based system (IWRS). Randomization is stratified based on the following prognostic factors:

- baseline albumin level (≥4.0 g/dL versus <4.0 g/dL);

- KPS (70 and 80 versus ≥90);

- ethnicity (Caucasians vs. East Asians vs. everyone else).

The drug will be administered in cycles. Patients will be treated until there are signs of disease progression (radiological or clinical deterioration), intolerable toxicity, or other reasons to discontinue the study. Tumor response will be assessed using the RECIST guidelines (Eisenhauer, E.A., et al., “New response evaluation criteria in solid tumors: Revised RECIST guideline (version 1.1). European Journal of Cancer, 2009. 45:pp. 228-247) every 6 weeks or sooner if disease progression is evident based on clinical signs and symptoms. Tumor measurement images will be collected and stored for all patients throughout the study. However, all treatment decisions will be based on the assessment of disease status by the local radiologist or study director. If independent analysis of ORR and/or PFS is required, independent image review can be performed.

A follow-up visit is required 30 days after stopping treatment. Thereafter, all patients will be followed up monthly for overall survival (by telephone or study site visit) until death or study completion, whichever occurs first. Patients who discontinue study treatment for reasons unrelated to objective disease progression (including patients who discontinue early from the study due to worsening symptoms) should continue to be monitored every 6 weeks during follow-up for radiological progression.

All patients will be asked to complete a pain and analgesic diary throughout their participation in the study, in which the patient will document pain severity ratings and daily analgesic consumption. Patient responses will be used to assess clinical benefit along with other parameters. All patients will also be required to complete the EORTC-QLQ-C30 questionnaire to assess quality of life.

To address the objectives of this study, all sites will be required to participate in an additional translational study (TR) protocol (MM-398-07-03-01.TR), unless prohibited by local regulations. Participation in this study will be optional for patients; they will be required to provide separate consent for the translational study.

The primary analysis of overall survival (OS) will be performed after the death of at least 305 patients enrolled in the study under protocol version 2 or later. Patients receiving study treatment during the main overall survival analysis will continue to receive treatment until one of the discontinuation criteria is met. During this study, an independent Data Monitoring Board (DSMB) will regularly review the safety data. Figure 7 shows the study design.

C. Patient selection and early termination of participation

This global study will enroll approximately 405 patients using protocol version 2 or later. To be included in the study, patients must have:

1. Histologically or cytologically confirmed exocrine adenocarcinoma of the pancreas.

2. Documented metastatic tumor; Disease status may or may not be measurable according to RECIST guidelines ver. 1.1.

3. Documented disease progression during previous use of gemcitabine or a gemcitabine-containing drug for locally advanced or metastatic cancer. Examples of acceptable drugs include, but are not limited to:

- monotherapy with gemcitabine;

- any gemcitabine-based treatment regimen, with or without gemcitabine-based maintenance treatment;

- monotherapy with gemcitabine, to which a platinum-based agent, fluoropyrimidine or erlotinib was subsequently added;

- gemcitabine, administered in combination with an adjuvant in case of disease relapse within 6 months after completion of adjuvant therapy.

4. Karnofsky General Condition Index (KPS)>70.

5. Adequate bone marrow reserves, confirmed by the following parameters:

- ANC>1500 cells/μl without the use of hematopoietic growth factors;

- platelet count >100,000 cells/μl; And

- hemoglobin >9 g/dl (for patients with hemoglobin levels below 9 g/dl, blood transfusion is allowed).

6. Adequate liver function, confirmed by the following parameters:

- total serum bilirubin within the normal range for organization (in case of obstruction of the bile ducts, their drainage is allowed);

- albumin level ≥3.0 g/dl;

- level of aspartate aminotransferase (AST) and alanine aminotransferase (ALT)≤2.5×VPN (in the presence of liver metastases, a level of ≤5×VPN is acceptable)

7. Adequate renal function, confirmed by a serum creatinine level ≤1.5×ULN.

8. Normal ECG or ECG without clinically significant abnormalities.

9. Recover from the effects of previous surgery, radiation therapy, or other cancer treatment.

10. At least 18 years of age.

11. Able to understand and sign the informed consent (or have a legal representative who is able to do so).

Patients must meet all inclusion criteria listed above and not meet any of the following exclusion criteria:

1. Active metastases to the central nervous system (manifested by clinical symptoms, cerebral edema, need for steroids, or progressive disease).

2. Clinically significant gastrointestinal disorders, including liver disorders, bleeding, inflammation, obstruction or grade 1 diarrhea.

3. History of any other malignant neoplasm in the last 5 years; subjects with a previous history of carcinoma in situ or basal or squamous cell carcinoma of the skin were eligible to participate. Subjects with other malignancies were eligible to participate if they had shown no evidence of disease in the past 5 years.

4. Severe arterial thromboembolism (myocardial infarction, unstable angina, stroke) less than 6 months before inclusion.

5. NYHA class III or IV congestive heart failure, ventricular arrhythmias, or uncontrolled blood pressure.

6. Active infection or unexplained fever >38.5°C during screening visits or on the first day of scheduled dosing (patients with tumor fever may be eligible for enrollment at the discretion of the investigator), which in the opinion of the investigator may jeopardize the patient's participation in the study or influences the outcome of the study

7. Known hypersensitivity to any of the components of MM-398, other liposomal products, fluoropyrimidines or leucovorin.

8. The investigational drug administered for 4 weeks or less than 5 times the half-life of the investigational agent prior to the day of the first scheduled drug dose in the study, whichever is longer.

9. Other medical or social conditions that, in the opinion of the investigator, may interfere with the patient's ability to sign informed consent, cooperate and participate in the study, or interfere with the interpretation of the results.

10. Pregnancy or breastfeeding; Women of childbearing potential must have a negative pregnancy test based on a urine or serum test at the time of inclusion in the study. Men and women of childbearing potential must agree to use a reliable method of birth control during the study and for up to 3 months after the last dose of study drug.

The admission criteria must be strictly adhered to. Discontinuation of the study drug is provided for patients in the following cases:

- the patient has signs of disease progression based on RECIST v1.1 criteria;

- the patient experiences a worsening of symptoms;

- the patient experiences an intolerable toxicity or adverse event that requires:

third dose reduction;

stopping treatment for more than 21 days from the start of the next cycle if, in the opinion of the investigator, the patient does not benefit from the study drug;

- the patient, in the opinion of the study director, does not adhere to the procedures according to the study;

- the patient or the patient's physician asks the patient to refrain from taking the study drug;

- the investigator or sponsor, for any reason, taking into account the rights, safety and welfare of patients and in accordance with ICH/GCP guidelines and local law, terminates the study or the patient's participation in the study.

If a patient is lost to follow-up or withholds study drug, an attempt should be made to contact the patient to determine the reason for discontinuation. If a patient is lost to follow-up, at least 3 documented attempts to contact the patient, including one by certified mail, should be made before the patient is considered lost to follow-up. If a patient discontinues study drug for reasons unrelated to objective progression, continued radiographic assessment of the patient's disease every 6 weeks until objective progression is detected.

All patients who discontinue study drug should remain for follow-up as required by the protocol. The only circumstance in which a patient should not have study endpoints monitored is if the patient's consent is withdrawn. Withdrawal of consent must be the patient's decision and must indicate not only the patient's desire to discontinue study drug and follow-up visits, but also that the investigator may not make any further efforts to contact the patient, including attempts to determine the patient's survival status.

D. Method of assigning patients to treatment groups

Once all screening tests are completed and UGT1A1*28 test results are available, patients should be randomized using a computerized interactive Internet-based system (IWRS) in a 1:1:1 ratio to one of the following treatment groups:

- group A (experimental group): MM-398;

- group B (control group): 5-FU and leucovorin;

- group C (experimental group): MM-398, 5-FU and leucovorin.

Randomization must occur within 7 days before the planned drug intake. Randomization is stratified based on the following prognostic factors:

- baseline albumin level (≥ 4.0 g/dL versus <4.0 g/dL);

- KPS (70 and 80 versus ≥90);

- ethnicity (Caucasian/East Asian/all others).

E. Description of MM-398

MM-398 is irinotecan (also known as CPT-11) encapsulated in a liposomal drug delivery system. It will be supplied in sterile, single-use vials containing 9.5 ml of MM-398 at a concentration of 5 mg/ml. Vials will contain an additional 0.5 ml to facilitate extraction of the specified amount from each 10 ml vial.

MM-398 must be stored in the refrigerator at a temperature of 2-8°C, protected from light. No protection from light is required during infusion. MM-398 should not be frozen. Responsible persons should check the contents of the vial for particulate matter before and after withdrawing the drug from the vial into the syringe.

MM-398 must be diluted before administration. The diluted solution is physically and chemically stable for 6 hours at room temperature (15-30°C), but should preferably be stored at refrigerator temperature (2-8°C) away from light. The diluted solution should not be frozen. Due to the potential for microbial contamination during dilution, it is advisable to use the diluted solution within 24 hours when stored in a refrigerator (2-8°C) and within 6 hours when stored at room temperature (15-30°C).

Twenty vials of MM-398 will be packed in a cardboard container. Individual vials, as well as outer cartons, will be labeled in accordance with local regulatory requirements.

MM-398 will be administered as follows. All patients will be initially screened for the UGT1A1*28 allele.

Group A - Patients without the homozygous UGT1A1*28 allele will receive MM-398 at a dose of 120 mg/ ^m2 .
- Patients homozygous for UGT1A1*28 will receive the first cycle of treatment at a reduced dose of 80 mg/ ^m2 . If the patient does not experience the toxic effect of the drug after the first dose of MM-398, then from cycle 2 onwards you can increase the dose in increments of 20 mg/ ^m2 to a maximum value of 120 mg/ ^m2 . Group C - Patients without the homozygous UGT1A1*28 allele will receive MM-398 at a dose of 80 mg/ ^m2 .
- Patients homozygous for the UGT1A1*28 allele and randomized to group C will receive the first cycle of treatment at a reduced dose of 60 mg/m ² . If the patient does not experience the toxic effect of the drug after the first dose of MM-398, then from cycle 2 onwards the dose can be increased to 80 mg/ ^m2 .
- MM-398 should be administered before 5-FU and leucovorin.

In Arm A, MM-398 will be administered by IV infusion over 90 minutes on the first day of each 3-week cycle at the study site. In Group C, MM-398 will be administered by IV infusion over 90 minutes during the first cycle; the infusion time can be reduced to 60 minutes, starting from cycle 2, if no acute reaction to the infusion occurred during cycle 1. The cycle duration is 3 weeks for group A and 2 weeks for group C. The first day of cycle 1 is a fixed day; subsequent doses should be administered on the first day of each cycle +/-3 days.

Before administration, the appropriate dose of MM-398 must be diluted with 5% dextrose injection (D5W) to a final volume of 500 mL. Be careful not to use built-in filters or solvents other than D5W. MM-398 can be administered using standard PVC bags and intravenous tubing.

The actual dose of MM-398 administered will be determined by calculating the patient's body surface area at the beginning of each cycle. For ease of administration, a +/-5% variation in the calculated total dose will be allowed. Because MM-398 vials are single-use, center staff should not store the unused portion of the vial for future use; any unused portion of the product should be discarded.

Before MM-398 infusion, all patients should receive standard doses of dexamethasone and a 5-HT3 antagonist or other antiemetic in accordance with standard institutional practice for irinotecan administration. For patients who have experienced acute cholinergic symptoms in previous cycles, atropine can be prescribed prophylactically.

F. Description of 5-FU and Leucovorin

5-fluorouracil is a pyrimidine antagonist that affects the biosynthesis of nucleic acids. The deoxyribonucleotide of this drug inhibits thymidylate synthetase, thereby inhibiting the formation of thymidylic acid from deoxyuridylic acid and interfering with DNA synthesis. It also interferes with RNA synthesis.

Leucovorin acts as a biochemical cofactor for 1-carbon transfer reactions in the synthesis of purines and pyrimidines. Leucovorin does not require the enzyme dihydrofolate reductase (DHFR) to convert to tetrahydrofolic acid. Leucovorin inhibits the effects of methotrexate and other DHFR antagonists. Leucovorin may enhance the cytotoxic effects of fluorinated pyrimidines (i.e. fluorouracil and floxuridine). Once 5-FU is activated in the cell, it is accompanied by a folate cofactor and inhibits the enzyme thymidylate synthetase, thereby inhibiting pyrimidine synthesis. Leucovorin increases the folate pool, thereby enhancing the binding of folate cofactor and active 5-FU to thymidylate synthetase.

FU and leucovorin should be handled and stored in accordance with the country-specific package insert. Commercially available 5-FU and leucovorin will be provided to all patients in the study randomized to arms B and C.

5-FU and leucovorin will be administered as follows.

Group B - 5-FU will be administered at a dose of 2000 mg/ ^m2 as an infusion over 24 hours (+/-30 minutes) every week for 4 weeks (on days 1, 8, 15 and 22) followed by 2 weeks of rest within a 6 week cycle.
- Leucovorin will be administered at a dose of 200 mg/ ^m2 (l-form) or 400 mg/ ^m2 (l+d-racemic form) as an IV infusion over 30 minutes once a week for 4 weeks (on days 1, 8, 15 and 22) followed by 2 weeks of rest as part of a 6-week cycle Group C - 5-FU will be administered at a dose of 2400 mg/ ^m2 as an infusion over 46 hours (+/-60 minutes) every 2 weeks.
- Leucovorin will be administered at a dose of 200 mg/ ^m2 (l-form) or 400 mg/ ^m2 (l+d-racemic form) as an infusion over 30 minutes every 2 weeks

Leucovorin should be reconstituted according to the package insert instructions or standard departmental guidelines for leucovorin dissolution. Leucovorin should be administered before the 5-FU infusion.

The actual dose of 5-FU and leucovorin administered will be determined by calculating the patient's body surface area prior to each cycle. For ease of administration, a +/-5% variation in the calculated total dose will be allowed.

After cycle 1, there will be a +/-3 day window for starting each new cycle, and a +/-1 day window for infusions on days 8, 15 and 22.

All patients should receive standard doses of dexamethasone, prochlorperazine, or other equivalent antiemetic prior to infusion of 5-FU and leucovorin according to standard institutional practice for administering 5-FU.

G. Important considerations for treatment with MM-398

Data from previous studies of MM-398 showed no unexpected toxic effects compared to the extensively studied active ingredient irinotecan. Warnings and precautions for the use of irinotecan and procedures for treating these toxic effects are listed below.

Diarrhea

Irinotecan can cause early and late forms of diarrhea, which appear to be due to different mechanisms. Early diarrhea (occurring during or shortly after irinotecan infusion) is cholinergic in nature. It is usually temporary and rarely severe. It may be accompanied by symptoms of rhinitis, increased salivation, miosis, lacrimation, sweating, flushing and increased intestinal motility, which can cause cramping abdominal pain. For patients who experienced early cholinergic symptoms during a previous cycle of MM-398, atropine will be administered prophylactically at the discretion of the investigator.

Late diarrhea (usually occurring more than 24 hours after irinotecan administration) can be life-threatening as it can be prolonged and lead to dehydration, electrolyte imbalance, or sepsis. Late-onset diarrhea should be promptly treated with loperamide, and if diarrhea persists after loperamide, octreotide should be considered. Loss of fluid and electrolytes associated with persistent or severe diarrhea can lead to life-threatening dehydration, kidney failure, and electrolyte imbalance and contribute to cardiovascular disease. There is an increased risk of infectious complications that can lead to sepsis in patients with chemotherapy-induced neutropenia. Patients with diarrhea should be closely monitored with fluid and electrolyte replacement if dehydration begins and antibiotic support if ileus, fever, or severe neutropenia develops.

Neutropenia

Deaths due to sepsis following severe neutropenia have been reported in patients receiving irinotecan. Neutropenic complications should be promptly treated with antibiotic support. G-CSF may be used to treat neutropenia at the discretion of the investigator. Patients who have experienced grade 3 or 4 neutropenia with prior anticancer therapy require careful monitoring and treatment.

Hypersensitivity

Hypersensitivity reactions, including severe anaphylactic or anaphylactoid reactions, have been observed. If hypersensitivity occurs, immediately stop taking the suspected drug and initiate invasive therapy.

Colitis/intestinal obstruction

Cases of colitis complicated by ulceration, bleeding, intestinal obstruction and infection have been observed. Patients experiencing intestinal obstruction should receive prompt antibiotic support.

Thromboembolism

Thromboembolic events have been reported in patients receiving a treatment regimen that included irinotecan; the specific causes of these phenomena were not determined.

Pregnancy

Irinotecan is classified as safety category D for use during pregnancy. Women of childbearing potential should be advised to avoid pregnancy during treatment with irinotecan. If pregnancy is reported, treatment should be discontinued. The patient should be withdrawn from the study and the pregnancy monitored until the outcome is known.

Caring for the IV Site

Hemorrhages should be avoided; it is necessary to monitor for signs of inflammation in the infusion area. If bleeding occurs, it is recommended to flush the area with sterile saline and apply ice.

Patients at special risk

In clinical studies with a weekly dosing schedule of irinotecan, it was noted that patients with moderately elevated baseline serum total bilirubin levels (1.0-2.0 mg/dL) were significantly more likely to experience grade 3 or 4 neutropenia during the first cycle than patients with bilirubin levels less than 1.0 mg/dL (50.0% [19/38] vs. 17.7% [47/226]; p<0.001). Patients with abnormalities of bilirubin glucuronidation, such as Gilbert's syndrome, may also be at increased risk of myelosuppression when treated with irinotecan.

Acute reactions associated with infusion

Acute infusion-related reactions characterized by flushing, shortness of breath, facial swelling, headache, chills, back pain, chest or pharyngeal tightness, and hypotension have been reported in a small number of patients receiving liposomal formulations. In most patients, these reactions resolved within 24 hours after stopping the infusion. In some patients, these reactions resolved when the infusion rate was reduced. Most patients who experienced reactions associated with acute infusion of liposomal drugs were able to tolerate further infusions without complications.

Other possible toxic effects

MM-398, a new liposomal formulation of irinotecan, is different from unencapsulated irinotecan and therefore there is the potential for toxic effects not caused by free irinotecan. Signs and symptoms of drug toxicity should be closely monitored in all patients, especially during initiation of treatment.

H. Requirements for dose changes

Dosing may be withheld for up to 3 weeks from the scheduled date to allow the patient to recover from toxic effects associated with the study drug. If recovery from a toxic effect requires more than 3 weeks, the patient's participation in the study should be discontinued, provided that the patient does not benefit from the study drug; otherwise, the investigator and the sponsor or sponsor's representative should discuss the possibility of continued patient participation based on the risks and benefits of continuation.

If a patient's dose is reduced during a study due to toxicity, the dose should remain reduced throughout the study; repeated increase in dose to the previous level is not allowed. If a patient requires a dose reduction for a third time due to persistence of an adverse event after two dose reductions, the patient should be discontinued from the study.

Infusion reactions should be monitored. Infusion reactions will be assessed according to the National Cancer Institute CTCAE (version 4.0) definition for allergic/infusion reactions and anaphylaxis below:

Grade 1: Temporary flushing or rash, drug-induced fever <38°C (<100.4°F); intervention is not indicated Grade 2: Intervention or interruption of infusion is indicated; prompt response to symptomatic treatment (eg, antihistamines, NSAIDs, narcotics); Prophylactic medications are indicated for <24 hours Grade 3: Symptomatic bronchospasm with or without urticaria; parenteral intervention is indicated; allergic edema/angioedema; hypotension Grade 4: Life-threatening consequences; urgent intervention indicated

When treating infusion reactions, the study site guidelines or the following guidelines should be used.

1st degree
- Reduce infusion rate by 50%
- Monitor the patient's condition every 15 minutes for deterioration 2nd degree
- Stop infusion
- Administer 50 mg diphenhydramine hydrochloride IV, 650 mg acetaminophen po, give oxygen
- Resume infusion at 50% rate after infusion response resolves
- Monitor the patient's condition every 15 minutes for deterioration
- Before all subsequent infusions, administer 25-50 mg diphenhydramine hydrochloride IV 3rd degree
- Stop the infusion and disconnect the infusion tubing from the patient
- Administer diphenhydramine hydrochloride 50 mg IV, dexamethasone 10 mg IV, bronchodilators for bronchospasm, and other medications or oxygen as needed
- Further treatment using MM-398 is not allowed 4th degree
- Stop the infusion and disconnect the infusion tubing from the patient
- Administer epinephrine, bronchodilators, or oxygen as indicated for bronchospasm
- Administer 50 mg diphenhydramine hydrochloride IV, 10 mg dexamethasone IV
- Consider hospitalization for observation
- Further treatment using MM-398 is not allowed

For patients who experience grade 1 or 2 infusion reactions, subsequent infusions may be administered at a reduced rate (over 120 minutes) at the discretion of the investigator.

For patients who experience a grade 1 or 2 infusion reaction a second time, dexamethasone 10 mg IV should be administered. All subsequent infusions should be preceded by 50 mg diphenhydramine hydrochloride IV, 10 mg dexamethasone IV, and 650 mg acetaminophen po.

I. Changing the dose of MM-398 for hematological toxicity

Before starting a new cycle of therapy, patients should have the following indicators:

- ACN>1500/mm ³ ;

- platelet count >100,000/ ^mm3 .

Treatment should be delayed to allow sufficient time for recovery and recovery; Treatment should be carried out in accordance with the guidelines given in the tables below. For febrile neutropenia, the ANC should be restored to ≥1500/mm ³ and the patient should recover from infection.

Table
Changing the dose of MM-398 with different numbers of neutrophils ANC: cells/mm ³ (worst grade according to CTCAE) Changing the dose of MM-398 for the next cycle ^a Group A: Patients not homozygous for UGT1A1*28 Group A: Patients homozygous for UGT1A1*28 ^d
Group C: Patients not homozygous for UGT1A1*28 Group C: Patients homozygous for UGT1A1*28 ^d ≥1000-1999 (grade 1 or 2) 100% of previous dose 100% of previous dose 100% of previous dose < 1000
(grade 3/4) or febrile neutropenia Reduce dose by 20 mg/ ^m2 to a minimum dose of 80 mg/ ^{m2 b} Reduce dose to 60 mg/ ^m2 for the first case and to 50 mg/ ^m2 for the second case ^c,d Reduce dose to 50 mg/ ^m2 for the first case and to 40 mg/ ^m2 for the second case ^e,d

^a All dose changes should be based on worst prior toxicity.

^b Patients requiring further dose reduction below 80 mg/ ^m2 should be excluded from the study.

^c Patients requiring further dose reduction below 50 mg/ ^m2 should be excluded from the study.

^d For patients homozygous for UGT1A1*28 whose dose is increased, the dose should be reduced according to the guidance for patients not homozygous for UGT1A1*28.

^e Patients requiring further dose reduction below 40 mg/ ^m2 should be excluded from the study.

Table
Changing the dose of MM-398 for other types of hematological toxicity Worst CTCAE toxicity rating Changing the MM-398 dose for the next cycle ^a Group A: Patients not homozygous for UGT1A1*28 Group A: Patients homozygous for UGT1A1*28 ^d
Group C: Patients not homozygous for UGT1A1*28 Group C: Patients homozygous for UGT1A1*28 ^d ≤2 degree 100% of previous dose 100% of previous dose 100% of previous dose 3/4 degree Reduce dose by 20 mg/ ^m2 to a minimum dose of 80 mg/ ^{m2 b} Reduce dose to 60 mg/ ^m2 for the first case and to 50 mg/ ^m2 for the second case ^c,d Reduce dose to 50 mg/ ^m2 for the first case and to 40 mg/ ^m2 for the second case ^e,d

^a All dose changes should be based on worst prior toxicity.

^b Patients requiring further dose reduction below 80 mg/ ^m2 should be excluded from the study.

^c Patients requiring further dose reduction below 50 mg/ ^m2 should be excluded from the study.

^d For patients homozygous for UGT1A1*28 whose dose is increased, the dose should be reduced according to the guidance for patients not homozygous for UGT1A1*28.

^e Patients requiring further dose reduction below 40 mg/ ^m2 should be excluded from the study.

J. MM-398 Dose Modification for Nonhematologic Toxicity

Treatment should be delayed until diarrhea is corrected to grade ≤1, and for other non-hematologic grade 3 or 4 toxicities until they are corrected to grade 1 or baseline. Guidance for dose adjustments of MM-398 for drug-induced diarrhea and other grade 3 or 4 nonhematologic toxicities is provided below. In case of infusion reactions, proceed as described above.

Table
Changing the dose of MM-398 for diarrhea Worst CTCAE toxicity rating Changing the MM-398 dose for the next cycle ^a Group A: Patients not homozygous for UGT1A1*28 Group A: Patients homozygous for UGT1A1*28 ^d
Group C: Patients not homozygous for UGT1A1*28 Group C: Patients homozygous for UGT1A1*28 ^d Grade 1 or 2 (stool 2-3 times a day more often than before treatment or stool 4-6 times a day more often than before treatment) 100% of previous dose 100% of previous dose 100% of previous dose Grade 3 (stool 7-9 times a day more often than before treatment) or grade 4 (stool >10 times a day more often than before treatment) Reduce dose by 20 mg/ ^m2 to a minimum dose of 80 mg/ ^{m2 b} Reduce dose to 60 mg/ ^m2 for the first case and to 50 mg/ ^m2 for the second case ^c,d Reduce dose to 50 mg/ ^m2 for the first case and to 40 mg/ ^m2 for the second case ^e,d

^a All dose changes should be based on worst prior toxicity.

^b Patients requiring further dose reduction below 80 mg/ ^m2 should be excluded from the study.

^c Patients requiring further dose reduction below 50 mg/ ^m2 should be excluded from the study.

^d For patients homozygous for UGT1A1*28 whose dose is increased, the dose should be reduced according to the guidance for patients not homozygous for UGT1A1*28.

^e Patients requiring further dose reduction below 40 mg/ ^m2 should be excluded from the study.

Table
Change in dose of MM-398 for non-hematological toxicity not associated with diarrhea, asthenia and anorexia grade 3 ^d Worst CTCAE toxicity rating Changing the MM-398 dose for the next cycle ^a Group A: Patients not homozygous for UGT1A1*28 Group A: Patients homozygous for UGT1A1*28 ^e
Group C: Patients not homozygous for UGT1A1*28 Group C: Patients homozygous for UGT1A1*28 ^e 1st or 2nd degree 100% of previous dose 100% of previous dose 100% of previous dose Grade 3 or 4 (except nausea and vomiting) Reduce dose by 20 mg/ ^m2 to a minimum dose of 80 mg/ ^{m2 b} Reduce dose to 60 mg/ ^m2 for the first case and to 50 mg/ ^m2 for the second case ^c,e Reduce dose to 50 mg/ ^m2 for the first case and to 40 mg/ ^m2 for the second case ^f,e Grade 3 or 4 nausea and/or vomiting despite antiemetics Optimize antiemetic therapy AND reduce dose by 20 mg/ ^m2 to a minimum dose of 80 mg/ ^{m2 b} Optimize antiemetic therapy and reduce the dose to 60 mg/ ^m2 ; if the patient is already receiving 60 mg/ ^m2 , reduce the dose to 50 mg/ ^{m2 c,e} Optimize antiemetic therapy and reduce the dose to 50 mg/ ^m2 ; if patient is already receiving 50 mg/ ^m2 , reduce dose to 40 mg/ ^{m2 f,e}

^a All dose changes should be based on worst prior toxicity.

^b Patients requiring further dose reduction below 80 mg/ ^m2 should be excluded from the study.

^c Patients requiring further dose reduction below 50 mg/ ^m2 should be excluded from the study.

^d For asthenia and anorexia grade 3, no dose change is required.

^e For patients homozygous for UGT1A1*28 whose dose is increased, the dose should be reduced according to the guidance for patients not homozygous for UGT1A1*28.

^f Patients requiring further dose reduction below 40 mg/ ^m2 should be excluded from the study.

K. Dose modification of 5-FU and leucovorin (groups B and C)

Below are recommendations for changing your 5-FU dose. No leucovorin dose adjustment is required due to toxicity. Leucovorin must be administered immediately before each dose of 5-FU; therefore, if the 5-FU dose is delayed, the leucovorin dose should also be delayed. In the event that a patient experiences an infusion reaction, the departmental guidelines or guidelines for the management of infusion reactions with MM-398 should be used.

L. 5-FU Dose Adjustment for Hematologic Toxicity

Before administering the next dose in a cycle or starting a new treatment cycle, patients should have the following:

- ACHN≥1500/mm3

- leukocytes ≥3500/mm ³ ;

- platelet count ≥75,000/mm ³ (according to the European summary of product characteristics for 5-FU, platelets must recover to a level ≥100,000/mm ³ before starting treatment).

Treatment should be delayed to allow sufficient time for recovery and recovery; Treatment should be carried out in accordance with the guidelines given in the table below. The length of the cycles is fixed at 6 weeks, and if the patient is unable to receive a dose of D8, D15 or D22 due to toxicity, that dose will be considered missed.

Table
5-FU Dose Adjustment for Hematologic Toxicity
(group B and C) ACN (cells/mm ³ ) Platelets
(cells/mm ³ ) Dose of 5-FU in D8, D15, D22 ^a Dose of 5-FU for next cycle ^a ≥1000 And ≥50000 100% of previous dose 100% of previous dose 500-999 Or <50000-25000 Suspend; after elimination, reduce dose by 25% ^b Reduce dose by 25% ^b <500 or febrile neutropenia Or <25,000 or thrombocytopenia with bleeding Suspend reception; after elimination, reduce dose by 25% ^b Reduce dose by 25% ^b

^a All dose changes should be based on worst prior toxicity.

^b Patients requiring more than 2 dose reductions should be excluded from the study.

M. 5-FU Dose Adjustment for Nonhematologic Toxicity

Treatment should be delayed until all grade 3 or 4 non-hematologic toxicities have corrected to grade 1 or baseline. Guidelines for dose adjustments of 5-FU due to toxicity are provided below. The length of the cycles is fixed at 6 weeks, and if the patient is unable to receive a dose of D8, D15 or D22 due to toxicity, that dose will be considered missed.

Table
Change in dose of 5-FU for non-hematological toxicity not associated with asthenia and anorexia grade 3 c (group B and C) Worst CTCAE toxicity rating Dose of 5-FU in D8, D15, D22 ^a Dose of 5-FU for next cycle ^a 1st or 2nd degree 100% of previous dose, excluding grade 2 hand-foot syndrome, grade 2 cardiac toxicity, or any grade neurocerebral toxicity 100% of previous dose, excluding grade 2 hand-foot syndrome, grade 2 cardiac toxicity, or any grade neurocerebral toxicity Hand-foot syndrome 2nd degree Reduce dose by 25% ^b Reduce dose by 25% ^b Neurocerebral toxicity of any grade or ≥ cardiac toxicity grade 2 Stop treatment Stop treatment 3rd or 4th degree Suspend; after elimination, reduce the dose by 25% ^b , with the exception of grade 3 or 4 hand-foot syndrome Reduce dose by 25% ^b except for grade 3 or 4 hand-foot syndrome Hand-foot syndrome, grade 3 or 4 Stop treatment Stop treatment

^a All dose changes should be based on worst prior toxicity.

^b Patients requiring more than 2 dose reductions should be excluded from the study.

^c For asthenia and anorexia grade 3, no dose change is required.

N. Other toxic effects requiring special attention

For the 5-FU and MM-398 treated groups, QTc prolongation secondary to electrolyte imbalance caused by diarrhea should be treated with appropriate electrolyte saturation. Once the underlying abnormality and ECG abnormalities have been corrected, treatment may be resumed, subject to close monitoring and appropriate dosage adjustments for diarrhea as described above.

O. Concomitant therapy

All concurrent medical conditions and complications of the underlying malignancy should be treated at the discretion of the investigator in accordance with acceptable local standards of care. Patients should receive analgesics, antiemetics, antibiotics, antipyretics, and blood products as needed. Although therapy with anticoagulants such as warfarin is allowed, careful monitoring of coagulation parameters is necessary to avoid complications of possible drug-drug interactions. All concomitant medications, including blood transfusions, should be recorded on the appropriate individual registration card.

Treatment guidelines for some diseases are discussed below; however, departmental guidelines for treating these conditions can also be used. Concomitant medications that require special attention are discussed below.

Antiemetics

Dexamethasone and a 5-HT3 blocker (for example, ondansetron or granisetron) can be administered to all patients as part of drug preparation in the absence of individual contraindications. Antiemetics will also be prescribed as clinically indicated during the study period.

Colony-stimulating factors

The use of granulocyte colony-stimulating factor (G-CSF) is acceptable for the treatment of patients with neutropenia or febrile neutropenia; Prophylactic use of G-CSF is limited to patients who have had at least one episode of grade 3 or 4 neutropenia or febrile neutropenia while receiving the study drug or documented grade 3 or 4 neutropenia or febrile neutropenia while receiving antineoplastic drugs in the past.

Treatment of diarrhea

Acute diarrhea and cramping abdominal pain developing during or within 24 hours after administration of MM-398 may be part of the cholinergic syndrome. This syndrome should be treated with atropine. For patients experiencing cholinergic symptoms during the study, the need for prophylactic or therapeutic administration of atropine should be considered.

Diarrhea can be debilitating and, in rare cases, potentially life-threatening. The guidelines developed by the ASCO panel for the treatment of chemotherapy-induced diarrhea are summarized below.

Table
Recommendations for the treatment of diarrhea caused by chemotherapy Clinical manifestation Intervention Diarrhea of any degree Oral loperamide (2 mg every 2 hours for irinotecan-induced diarrhea; 2 mg every 4 hours for 5-FU-induced diarrhea): continue until diarrhea is free for ≥12 hours Diarrhea persists when taking loperamide for >24 hours Oral fluoroquinolone ×7 days Diarrhea persists when taking loperamide for >48 hours Stop taking loperamide; hospitalize the patient; administer intravenous fluids ANC<500 cells/µl, regardless of fever or diarrhea Oral fluoroquinolone (continue until neutropenia resolves) Fever with persistent diarrhea, even in the absence of neutropenia Oral fluoroquinolone (continue until fever and diarrhea resolve)

The synthetic octapeptide octreotide has been shown to be effective in the control of diarrhea caused by fluoropyrimidine chemotherapy when administered in increasing doses by continuous infusion or subcutaneous injection. Octreotide can be administered in doses ranging from 100 mcg twice daily to 500 mcg three times daily, with a maximum tolerated dose of 2000 mcg three times daily over a 5-day course. Patients should be advised to drink plenty of fluids throughout treatment.

Other treatments

Symptomatic treatment of other toxic effects should be carried out according to institutional guidelines. It is possible to prevent alopecia with a cooling cap or stomatitis with ice-cold mouth rinses.

P. Prohibited Therapeutics

The instructions for use of irinotecan indicate that the following drugs interact with irinotecan: St. John's wort, CYP3A4-inducing anticonvulsants (phenytoin, phenobarbital, carbamazepine), ketoconazole, itraconazole, troleandomycin, erythromycin, diltiazem and verapamil. Treatment with these and other agents that interact with irinotecan should be avoided if possible. Because 5-FU interacts with warfarin, caution should be exercised if coadministration is necessary. For other drug-drug interactions, see the country-specific package inserts for 5-FU and leucovorin.

The following therapeutic agents are not permitted during the study:

- other antitumor therapeutic agents, including cytotoxic agents, targeted agents, hormonal therapeutic agents or other antibodies;

- potentially curative radiation therapy; the use of palliative radiation therapy is allowed; And

- the use of any other study drugs is not allowed.

Q. Laboratory Procedures

General blood analysis

A complete blood count (CBC) will be performed on a local basis and should include an assessment of the white blood cell (WBC) count, leukocyte count, hemoglobin, hematocrit and platelet count.

Serum biochemical analysis

The serum biochemical panel will be performed centrally. In addition, biochemical testing can be performed on a local basis and results obtained from the local laboratory can be used for enrollment and treatment decisions when results from the central laboratory are not available. If local laboratory results are used for enrollment, then local laboratory results should be used in all subsequent treatment decisions. Serum biochemical analysis should include determination of electrolytes (sodium, potassium, chloride and bicarbonate), BUN, serum creatinine, glucose, direct and total bilirubin, AST, ALT, alkaline phosphatase, LDH, uric acid, total protein, albumin, calcium, magnesium and phosphate.

CA 19-9

CA 19-9 levels will be measured centrally for all patients.

Pregnancy test

All women of childbearing potential should undergo a pregnancy test using serum or urine.

Allele UGT1A1*28

Whole blood samples will be collected from all patients at baseline and sent to a central laboratory to verify UGT1A1*28 allele status. Local laboratory results can be used if central laboratory results are not available at the time of randomization.

Pharmacokinetic assessments

PK analysis will be carried out centrally. Plasma samples for PK assessment will be collected during the first cycle from all patients randomized in this study at the following time points:

- group A: immediately before the infusion, during the infusion (80 to 90 minutes after the start of the infusion), between 2 and a half and four hours after the start of the infusion and in C1D8;

- group B: one sample at the end of the 5-FU infusion (C1D2);

- Group C: immediately before the MM-398 infusion, during the MM-398 infusion (80 to 90 minutes after the start of the infusion), between 2 and a half and four hours after the start of the MM-398 infusion, at the end of the 5-FU infusion and at C1D8.

In addition, a sample for PK assessment will be collected during the first cycle, any time between 8 and 72 hours after MM-398 administration from patients randomized to groups A and C who provide additional consent for this sample to be collected.

R. Assessment of pain and analgesic consumption

Patients will be provided with pain and analgesic consumption diaries to record daily pain intensity on a visual analogue scale and document daily analgesic consumption.

S.EORTC-QLQ-C30

Quality of life will be assessed using the EORTC-QLQ-C30 instrument. The EORTC-QLQ-C30 is a reliable and objective measure of quality of life in cancer patients in cross-national clinical trials. It includes nine multi-item scales: five functional scales (physical, role, cognitive, emotional and social); three symptom scales (fatigue, pain, and nausea and vomiting); and a scale of general health and quality of life. In addition, it includes several single-item symptom measures.

Patients are required to complete the EORTC-QLQ-C30 questionnaire at the time points specified in the assessment schedule. On days when the patient is scheduled to receive study drug, assessments should be completed prior to initiation of study drug administration. The questionnaire should only be completed by patients for whom validated translations of the EORTC-QLQ-C30 questionnaire are available.

T. Overall survival/follow-up after study

Overall survival data will be collected after the patient completes the 30-day follow-up visit, every month (+/-1 week) after the date of the 30-day follow-up visit. Data collected after discontinuation should include: date of disease progression (if not already documented; if the patient discontinues study drug early for reasons unrelated to objective disease progression, the patient should continue to undergo tumor assessment every 6 weeks until new anticancer therapy is initiated or disease progression); documentation of cancer treatment received by the patient, including dates of systemic therapy, radiation therapy, or surgery after discontinuation of study participation; and date of death. All patients must be enrolled until death or study closure, whichever occurs first.

U. Determination of severity and association of adverse events

The category of each adverse event will be determined in accordance with NCI CTCAE ver. 4.0, located at http://ctep.cАЧНer.gov/reporting/ctc.html. Events not listed in the CTCAE will be designated as mild, moderate, severe, life-threatening, or fatal, corresponding to NCI CTCAE categories 1, 2, 3, 4, and 5, respectively, and the following definitions:

- mild: a phenomenon that does not lead to incapacity or disability and can be eliminated without intervention;

- moderate: a phenomenon that does not lead to disability or disability, but requires intervention;

- severe: a phenomenon leading to temporary incapacity or disability and requiring intervention;

- life-threatening: an event in which the patient is at risk of death during the event

- fatal: a phenomenon that leads to the death of the patient

The investigator should attempt to determine whether there is a reasonable likelihood that the adverse event is related to the study drug. This relationship should be described as "related" or "unrelated".

V. Analysis of overall survival

Overall survival (OS) is the primary endpoint of the study. Overall survival is defined as the time from the date of randomization of patients to the date of death or the date the subject was last known to be alive. For each patient who is not known to have died at the time of data collection completion, overall survival will be revised based on the analysis at the time of last contact before data collection was completed.

The primary analysis of the study will include two pairwise comparisons of study drug survival in the ITT population using the unstratified log-rank test. Testing will be carried out according to the Bonferroni-Holm procedure, which tightly controls the error rate adjusted for the effect of multiple comparisons at the 0.05 level (two-sided) [25]:

Reject :SA(t)=SB(t), i.e. monotherapy with MM-398 has no effect compared with control if the log-rank p value for this test is at least 0.025, or if the log-rank p value for this test is less than 0.05 and the log rank p value for the comparison of groups B and C less than 0.025.

Reject :SC(t)=SB(t), i.e. combination therapy with MM-398 has no effect compared with control if the log-rank p value for this test is at least 0.025, or if the log-rank p value for this test is less than 0.05 and the log rank p value for the comparison of groups A and B less than 0.025.

For each treatment group, Kaplan-Meier analysis will be performed to obtain nonparametric estimates of the survival function and median survival time. The corresponding 95% confidence intervals will be calculated using the log-log method. Cox proportional hazards modeling will be used to estimate hazard ratios and associated 95% confidence intervals.

For overall survival in the ITT population (except as noted), the following additional sensitivity analyzes will be performed to assess the robustness of the primary analysis results:

log-rank comparison of treatments in the PP population;

stratified log-rank analyzes using randomization stratification factors [with hazard ratio parameters estimated from stratified Cox modeling];

Wilcoxon treatment comparison;

Cox regression model with stepwise selection (entry p < 0.25, residual p < 0.15) of model parameters, where treatment and prognostic factors (noted below) are candidates for inclusion;

univariate analysis to evaluate potential independent prognostic factors using Cox regression;

Subgroup analyzes to test for differences in treatment effects across different segments of the study population.

Repeat all analyzes (primary and sensitivity analyses) using only patients enrolled under protocol version 2 (and later)

Prognostic factors tested include: baseline KPS, baseline albumin, ethnicity, geographic location, disease stage at diagnosis, baseline tumor location, number of prior chemotherapy courses, prior radiation therapy, prior surgery, time since last treatment, best response to previous treatment, baseline CA 19-9, gender and age.

W. Additional Efficacy Analyzes

Progression-free survival

PFS is defined as the number of months from the date of randomization to the date of death or progression, whichever occurs first (according to RECIST 1.1). If neither death nor progression is observed during the study, PFS data will be revised to the time of the last reliable tumor assessment.

Comparisons of PFS between treatment groups will be performed using the paired unstratified log-rank test. PFS curves will be assessed using Kaplan-Meier estimates. The Cox proportional hazards model will produce hazard ratio estimates and associated 95% confidence intervals. In addition, stratified analyzes using randomization stratification factors will be performed. Treatment effects adjusted for stratification variables and other prognostic covariates will be examined. In addition, various methods for selecting and imputing missing data can be used to perform PFS sensitivity analyses. The sensitivity analysis methodology will be fully described in the statistical analysis plan.

Analyzes will be performed on the ITT, PP and EP populations.

Time until no treatment effect is declared

Time to treatment failure is defined as the time from randomization to disease progression, death, or early discontinuation due to toxicity. Kaplan-Meier analyzes will be performed for time to treatment failure as indicated for progression-free survival analyses.

Analyzes will be performed on the ITT, PP and EP populations.

Frequency of objective reactions

Tumor scores associated with ORR will be determined using RECIST v1.1. If a sponsor requires an independent review of radiologic assessments to support a new use of a drug or for any other reason, the response status of all patients may be reviewed by an independent panel of clinicians and the sponsor or its designee. In the event of a discrepancy between the independent panel's assessment and the investigator's assessment, the independent panel's assessment will take precedence.

The objective response rate (ORR) for each treatment group will be calculated by adding the number of patients with the best overall response confirmed CR or PR according to RECIST. ORR represents the best response recorded from randomization to progression or end of study. The number and percentage of patients experiencing an objective response (confirmed by CR+PR) at the time of analysis will be reported and the 95% confidence interval for the proportion calculated. Comparisons of objective response rates between treatment groups will be performed using pairwise Fisher's exact test. Analyzes will be performed on the ITT, PP and EP populations.

Tumor Marker Response Analysis

Serum CA 19-9 levels will be assessed for 7 days before treatment (baseline) and every 6 weeks thereafter. Tumor marker CA19-9 response will be assessed by changes in serum CA19-9 levels. Response is defined as at least one 50% decrease in CA 19-9 from baseline during the treatment period. Tumor marker response rate calculations will only be performed for patients with elevated baseline CA 19-9 levels (>30 U/mL).

Analysis of results according to patient reports

The EORTC-QLQ-C30 questionnaires will be analyzed in accordance with the EORTC guidelines [22].

Security Analysis

Treatment-emergent adverse events will be presented by treatment group, patient, NCI CTCAE category, and MedDRA system organ class (SOC). Separate lists will be provided for common adverse events, serious adverse events, study drug-related adverse events, and grade 3 and 4 adverse events. Laboratory data will be reported by treatment group and visit. Abnormal laboratory values will be assessed according to NCI CTCAE category whenever possible. The QTc estimate will be performed based on the Friederici's formula correction method. CTCAE criteria will be applied to QTcF (ie category 3 = QTc>500 ms). All safety analyzes will be performed according to treatment group, treatment cycle, and week, as appropriate. Overall safety will also be assessed based on category over multiple cycles, SOC, and exposure. In addition, the safety analysis will include comparisons between treatment groups for all patients in the population to assess safety:

- required number of blood transfusions;

- proportion of patients requiring G-CSF;

- adverse events leading to a delay or change in dose.

Pharmacokinetic analysis

Pharmacokinetic data will be collected for all patients randomized to either arm receiving MM-398. Plasma concentration-time data will be analyzed using population pharmacokinetic methods. Pharmacokinetic parameters will be assessed by nonlinear mixed effects modeling using NONMEM® version 7, level 1.0 (ICON Development Solutions, Dublin, Ireland). PK parameters will include _Cmax , _Tmax , plasma AUC (area under the concentration curve), clearance, volume of distribution and terminal half-life. The influence of patient-specific factors (age, race, sex, body weight, liver and kidney function tests, ECOG value, etc.) on pharmacokinetic parameters will be assessed. Population-based PK/PD techniques will be used to assess the relationship between drug exposure and efficacy and/or toxicity parameters (eg, neutropenia, diarrhea). Additional exploratory analysis of PK samples may be performed to clarify safety, efficacy, or PK issues related to MM-398 that arise during the study. 5-FU concentration levels will be summarized descriptively.

Notes

Although the present invention has been described in connection with specific embodiments, it is understood that further changes thereto are possible, and the present application is intended to cover any changes, uses or modifications of the present invention generally consistent with the spirit of the invention and including such departures from the present description , which arise within the limits of known or generally accepted practice within the field of technology to which the invention relates, and may apply to the essential functions set forth herein. The disclosure of any U.S., international or other patents or patent applications or publications referenced herein is incorporated herein by reference in its entirety.

Claims (19)

1. A method of treating exocrine pancreatic cancer in a human patient who has been previously treated with gemcitabine, comprising intravenously administering to the patient an antitumor therapy consisting of: a) a dose of 60, 70 or 80 mg/m ² of a liposomal irinotecan composition containing irinotecan liposomes in a diluted irinotecan injection formulation, wherein the diluted irinotecan injection formulation is prepared by diluting a volume of a liposomal irinotecan composition containing 5 mg/ml liposomal irinotecan in 500 ml 5 % dextrose solution for injection, in combination with b) a dose of 200 mg/m ² (l) form of leucovorin and a dose of 2400 mg/m ² 5-fluorouracil, for the treatment of exocrine pancreatic cancer in a patient, wherein the irinotecan liposomal composition comprises a sucrose octasulfate salt of irinotecan encapsulated in irinotecan liposomes consisting of phosphatidylcholine, cholesterol and polyethylene glycol modified phosphatidylethanolamine and having a diameter of approximately 80-140 nm, wherein the diluted injectable formulation Irinotecan, leucovorin and 5-fluorouracil are administered to the patient sequentially, starting from the first day of the two-week cycle. 2. The method of claim 1, wherein the polyethylene glycol modified phosphatidylethanolamine is present in an amount of approximately one polyethylene glycol (PEG) molecule per 200 phospholipid molecules in the irinotecan liposomes. 3. The method according to claim 1, where the irinotecan liposomes are monolamellar lipid bilayer vesicles encapsulating the irinotecan salt with sucrose octasulfate. 4. The method according to claim 1, in which exocrine pancreatic cancer has progressed during therapy with gemcitabine before administration of irinotecan liposomes. 5. The method according to claim 1, in which 200 mg/m ² (l) form of leucovorin is introduced as 400 mg/m ² (l+d) racemic form of leucovorin. 6. The method of claim 1, wherein a diluted injectable formulation of irinotecan is administered to the patient prior to 5-fluorouracil. 7. The method according to claim 6, in which: a) leucovorin is administered as 400 mg/m ² (l+d) racemic form of leucovorin; b) exocrine pancreatic cancer progressed during therapy with gemcitabine before administration of diluted injectable irinotecan; c) irinotecan liposomes are monolamellar lipid bilayer vesicles encapsulating the irinotecan salt with sucrose octasulfate, and d) polyethylene glycol modified phosphatidylethanolamine is present in an amount of approximately one polyethylene glycol molecule per 200 phospholipid molecules in irinotecan liposomes. 8. The method of claim 1, further comprising storing the diluted irinotecan injectable formulation at a temperature of approximately 2-8°C for no more than 24 hours prior to administration. 9. The method of claim 1, wherein irinotecan liposomes are monolamellar lipid bilayer vesicles that encapsulate an aqueous space containing irinotecan in a gelled or precipitated state as a sucrose octasulfate salt. 10. The method of claim 1, wherein the patient has previously been unsuccessfully treated with gemcitabine or has acquired resistance to gemcitabine prior to administration of the diluted irinotecan injectable formulation. 11. The method of claim 1, wherein the dose of the liposomal irinotecan composition is 80 mg/m ² and the human patient is not homozygous for the UGT1A1*28 allele. 12. The method of claim 11, wherein the liposomal irinotecan composition is administered as a diluted irinotecan injection formulation as a 90-minute infusion, leucovorin is administered after irinotecan for 30 minutes, and 5-fluorouracil is administered within 46 hours after leucovorin.