WO2015142367A1

WO2015142367A1 - Compositions and methods of treating liver cancer

Info

Publication number: WO2015142367A1
Application number: PCT/US2014/033675
Authority: WO
Inventors: Andrey Leonov; Roman Ivanov; Polina SHPIGOTSKAYA
Original assignee: Incuron LLC
Current assignee: Incuron LLC
Priority date: 2014-03-17
Filing date: 2014-04-10
Publication date: 2015-09-24
Anticipated expiration: 2016-09-17
Also published as: CN105611980A; EA201691846A1

Abstract

This invention relates to methods and compositions that are useful for the treatment of various cancers, including primary and secondary liver cancers. The cancers can be metastatic cancer in the liver, or cancers that have metastasized from liver. The invention relates to, in part, uses of quinacrine in the manufacture of a medicament for treatment-resistant liver cancer.

Description

CO POSITIONS AND METHODS OF TREATING OVER CANCER

CROSS-REFERENCE TO RELATED APPLICATIONS

This application claims the benefit of U.S. Provisional Patent Application No, 61/954,342, filed March 17, 2014, the entire contents of which are herein incorporated by reference.

FIELD OF THE INVENTION

This Invention relates to methods and compositions that are useful for the treatment of various cancers, including !lver cancer.

BACKGROUND

Liver cancer Is the fifth most common cancer ^'worldwide and the third most common cause of cancer-reiated death. Commonly Iiver cancers are treated via surgical resection or chemotherapy, chemoembolisation, or radiation therapy, if surgery is not an option. Surgical resection, however, is sometime impractical due to tumor size or location and co plicating Iiver diseases, such as advanced cirrhosis. Further, even if surgical resection is tenable, tumor recurrence/relapse complicates 70% of cases at 5 years post-resection. Liver cancer treatment is also compounded by the fact that chemofherapeuflc treatments tend to have reduced efficacy in Iiver cancer, possibly due to an increased ability of iiver cancer ceils to efflux chemotherapeutic agents.

There remains a need for agents and medicaments that can effectively treat iiver cancer,

SUMMARY OF THE INVENTION

Accordingly, the present invention provides for the use of qulnacrine in the manufacture of a medicament for the treatment of primary or secondary liver cancer that Is resistant to standard treatments and/or a method for treating primary or secondary liver cancer comprising administering an effective amount of quinacrine to a patient, wherein the liver cancer is resistant to standard treatment methods. In various embodiments, the Iiver cancer is resistant and/or poorly responsive to various chemotherapies, including, for example, any of doxorubicin, 5-f!uorouracii, or cisplatin. In various embodiments, the cancerous liver comprises hepatocytes which overexpress one or more multidrug-resistance pumps.

The invention also relates to various uses of quinacrine to supplement, for example as adjuvant or neoadjuvant care, or supplant standard treatments. For example, the present invention includes the use of quinacrine as a neoadjuvant that allows for a patient to undergo surgical resection or the use of quinacrine as an adjuvant in a patient that has undergone surgical resection. Also, the present invention includes the use of quinacrine as pailiative care in late stage liver cancer patients, especially in light of its relatively more palatable side effect profile, as compared to standard treatments.

Further, the present invention relates to the use of quinacrine in the manufacture of a medicament suitable for oral administration for the treatment of liver cancer, wherein the medicament comprises quinacrine in a dosage amount of from 300-400 mg. The invention also provides an oral dosage regimen comprising at least about 30 unit doses of from about 300-400 mg of quinacrine. The invention also provides methods of treating cancer in a liver cancer patient, comprising administering about 400 mg of quinacrine daily for about two weeks to achieve a steady state concentration and optionally then administering a maintenance dose of quinacrine. The maintenance dose in some emobodiments is from 300-400 mg, and in some embodiments, lower than about 400 mg of quinacrine, daily. Further the Invention provides for kits to allow a patient to easily comply with the treatment regimens described herein.

BRIEF DESCRIPTION OF THE FIGURES

Fig, 1 shows a graph of the percentage change of maximum diameter target lesions after the end of 8-week quinacrine therapy In comparison with baseline data. The label above each graphical bar corresponds to individual patient identification numbers.

DETAILED DESCRIPTION OF THE INVENTION

The present invention is based, in part, on the discovery that quinacrine may used in the manufacture of a medicament that is effective against treatment-resistant liver cancers. Further, the invention is based, in part, on the discovery that quinacrine is effective in treating primary or secondary liver cancers in human subject when orally administered In doses of about 400 mg daily and/or In a regime that is guided by the pharmacokinetic properties discovered herein.

Definitions

The following definitions are used in connection with the invention disclosed herein. Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood to one of skill in the art to which this invention belongs.

As used herein, "a," "an," or "the" can mean one or more than one.

Further, the term "about" when used in connection with a referenced numeric indication means the referenced numeric indication plus or minus up to 10% of that referenced numeric indication. For example, the language "about 50" covers the range of 45 to 55.

An "effective amount," when used in connection with medical uses is an amount that is effective for providing a measurable treatment, prevention, or reduction in the rate of pathogenesis of a disease of interest. As referred to herein, all compositional percentages are by weight of the total composition, unless otherwise specified. As used herein, the word 'Include," and Its variants, is intended to be non-limiting, such that recitation of items in a list Is not to the exclusion of other like Items that may also be useful in the compositions and methods of this technology. Similarly, the terms "can" and "may" and their variants are intended to be non- limiting, such that recitation that an embodiment can or may comprise certain elements or features does not exclude other embodiments of the present technology that do not contain those elements or features.

Although the open-ended term "comprising," as a synonym of terms such as including, containing, or having, is used herein to describe and claim the invention, the present invention, or embodiments thereof, may alternatively be described using alternative terms such as "consisting of" or "consisting essentially of."

As used herein, the words "preferred" and "preferably" refer to embodiments of the technology that afford certain benefits, under certain circumstances. However, other embodiments may also be preferred, under the same or other circumstances. Furthermore, the recitation of one or more preferred embodiments does not Imply that other embodiments are not useful, and Is not intended to exclude other embodiments from the scope of the technology.

Generally, for administering therapeutic agents (e.g. quinacrine) for therapeutic purposes, the therapeutic agents are given at a pharmacologically effective dose, I.e. within a therapeutic window as described herein. A "pharmacologically effective amount," "pharmacologically effective dose," "therapeutically effective amount," or "effective amount" refers to an amount sufficient to produce the desired physiological effect or amount capable of achieving the desired result, particularly for treating the disorder or disease. An effective amount as used herein would include an amount sufficient to, for example, delay the development of a symptom of the disorder or disease, alter the course of a symptom of the disorder or disease (e.g., slow the progression of a symptom of the disease), reduce or eliminate one or more symptoms or manifestations of the disorder or disease, and reverse a symptom of a disorder or disease. For example, administration of therapeutic agents to a patient suffering from cancer provides a therapeutic benefit not only when the underlying condition is eradicated or ameliorated, but also when the patient reports a decrease in the severity or duration of the symptoms associated with the disease, e.g., a decrease in tumor burden, a decrease in circulating tumor cells, an increase in progression free survival. Therapeutic benefit also includes halting or slowing the progression of the underlying disease or disorder, regardless of whether improvement Is realized. Such pharmaceutically effective amounts are limited by a drug's toxicity.

Effective amounts, toxicity, and therapeutic efficacy can sometimes be determined In cell cultures or experimental animals, e.g., for determining the LD50 (the dose lethal to about 50% of the population) and the ED50 (the dose therapeutically effective in about 50% of the population). This is crucial to determine whether a drug has a "therapeutic window," especially when applying the drug to an intractable condition. The dosage can vary depending upon the dosage form employed and the route of administration utilized. The dose ratio between toxic and therapeutic effects is the therapeutic index and can be expressed as the ratio LD50/ED50. In some embodiments, compositions and methods that exhibit large therapeutic indices are preierred. Although, for intractable conditions, such as treaiment-res!tant and/or unresectable liver cancer, therapeutic indices may be large. A therapeutically effective dose can be estimated initially from in vitro assays, including, for example, ceil culture assays. Also, a dose can be formulated in animal models to achieve a circulating plasma concentration range that includes the IC50 as determined in ceil culture, or in an appropriate animal model. Levels of the described compositions in plasma can be measured, for example, by high performance liquid chromatography. The effects of any particular dosage can be monitored by a suitable bloassay. The dosage can be determined by a physician and adjusted, as necessary, to suit observed effects of the treatment.

In some aspects, the present invention provides for a use of quinacrine in the manufacture of a medicament for the treatment of liver cancer that is resistant to standard treatment methods. In some aspects, the present invention provides a method for treating liver cancer that is resistant to standard treatment methods comprising administering an effective amount of quinacrine to a patient in need thereof.

In various embodiments, resistance to standard treatment is manifested through a lack of beneficial effect (e.g. failure to reduce tumor size and/or tumor load and/or stabilizing disease) and/or a recurrence of cancer. In various embodiments, resitatnce to standard treatment is apparent from the lack of response and/or inadequate response by a patient that has undergone standard treatment.

In some embodiments, the liver cancer is resistant to one or more of chemotherapy, chemoemboiization, radiation therapy, cryosurgery, radiofrequency ablation, or ethanol injection. In some embodiments, the liver cancer is resistant to any of the chemotherapies described herein as wither determined by a preior treatment of a patient or, in some membodiments, a surrogate biomarker. In some embodiments, the liver cancer responds poorly to chemotherapy and/or the liver cancer may experience only a transient response (such as that observed with, for example, doxorubicin or 5-fluorouraci! or cisplatin treatment). In some embodiments, the liver cancer is resistant to chemoemboiization, injection of chemotherapy agents and one or more blood vessel occluding agents directly into the arteries supplying blood to a tumor (e.g. the hepatic artery). In some embodiments, the liver cancer Is resistant to cryotherapy (e.g. using cold probes to freeze the tumor). In some embodiments, the tumor is resistant to radiofrequency ablation (e.g. ablation with the heat generated from high frequency alternating current) and/or ethanol injection, a.k.a, percutaneous ablation (e.g. using a needle to place alcohol directly into the tumor to kill it).

Surgical resection Is a common method of treating liver cancer; however resection is often untenable when a patient has underlying liver disease (Including, for example, cirrhosis and/or hepatitis and/or the other diseases described herein). In some embodiments, the liver cancer is unresectable and/or present In a subject that has coexisting medical condition that prevents surgery, selected from cirrhosis and hepatitis. In some embodiments, quinacrine is used in patients that have unresectable liver cancer because the cancer is spread throughout the liver and/or is close to the area where the liver meets the main arteries, veins, and bile ducts. In some embodiments, the liver cancer is present in a subject that Is not a candidate for transplantation. In some embodiments, the patient is not a candidate for a liver transplant If the patient has, for example, a single tumor of greater than about 5 cm across, tumors that have not been stable for at least about 6 months, and greater than about 5 small tumors, each one no larger than about 3 cm across.

In some aspects, the present invention provides for a method for treating cancer, comprising administering an effective amount of quinacrine to a patient in need thereof, wherein the cancer is a metastatic cancer that is of hepatic origin or a metastatic cancer that has metastasized to the liver from another organ.

In some aspects, the present invention provides for a method for treating liver cancer, comprising administering an effective amount of quinacrine to a patient in need thereof, wherein the patient is administered quinacrine at about 400 mg daily, or about 450 mg daily, or about 500 mg daily for about two weeks to achieve a steady state concentration. In various embodiments, the dose of quinacrine my be administered orally at least once, or twice, or thrice, or four, or five, or six, or seven, or eight, or nine, or ten, or eleven, or twelve times per day (e.g. four, 100 mg or eight, 50 mg oral administrations in the case of a daily 400 mg loading dose). In various embodiments, the presence and/or severity of side effects (as determined using any one of the methods described herein, e.g. blood tests and/or biopsies and/or examination by a medical practitioner) will direct a reduction in the loading dose. In various embodiments, the achievement of a steady state concentration is assessed using blood tests and/or liver biopsies. In some embodiments, the patient is administered a maintenance dose of quinacrine after achieving a steady state concentration of quinacrine. In some embodiments, the patient Is administered a maintenance dose of about 400 to about 50mg of quinacrine, such as, for example, about 350 mg, or about 300 mg, or about 250 mg, or about 200 mg, or about 150 mg, or about 100 mg of quinacrine. in various embodiments, the dose of quinacrine may be administered orally at least once, or twice, or thrice, or four, or five, or six, or seven, or eight, or nine, or ten, or eleven, or twelve times per day (e.g. one, 100 mg or two, 50 mg oral administrations in the case of a daily 100 mg maintenance dose). In various embodiments, the presence and/or severity of side effects (as determined using any one of the methods described herein, e.g. blood tests and/or biopsies and/or examination by a medical practitioner) will direct a reduction in the maintenance dose, In some embodiments, the patient is administered an additional agent that reduces side effects during the administration period to achieve a steady state concentration of quinacrine, ^'wherein the additional agent is one or more or an anti-emetic, loperamide, diphenoxylate, and atropine.

In various embodiments, the patient is administered the following treatment regimen: about 400 mg dally, or about 450 mg daily, or about 500 mg daily for about 10-40 days, such as, for example, about 10, or about 11 , or about 12, or about 13, or about 14, or about 15, or about 16, or about 17 days, or about 18 days, or about 19 days, or about 20 days, or about 1 week, or about 2 weeks, or about 4 weeks to achieve a steady state concentration of quinacrine and, optionally, Is administered at a maintenance dose of about 350 mg, or about 300 mg, or about 250 mg, or about 200 mg, or about 150 mg, or about 100 mg for a period of about 1 week, or about 2 weeks, or about 3 weeks, or about 4 weeks, or about 1 month, or about 8 weeks, or about 2 months, or about 3 months, or to about 6 months, in various embodiments, the dosing regiment is any of those described in Example 1 (e.g. Table 1) or Example 3 (e.g. the listed cohorts).

In various embodiments, the inventive regimen, or any of the uses or treatment described herein, may be supplemented with an additional agent to reduce side effects. In various embodiments, the side effect Is one or more of those found in Table 23. For instance an anti-emetic may be used to supplement quinacnne; including, for example, 5-HT3 blockers with or without dexamethasone. Also, an anti-diarrheal agent may be used, including, for example, loperamide, diphenoxylate, and atropine. In some embodiments, the inventive regimen, or any of the uses or treatment described herein, may be supplemented with one or more combination agents described herein (by way of non !imiting example, in the context of chemoemboiization).

In some aspects, the present invention provides for use of quinacrine in the manufacture of a medicament suitable for oral administration for the treatment of liver cancer, wherein the medicament comprises unit doses of quinacrine of an amount of from about 300-400 mg. In some embodiments, a single dose unit is contained in 1 or 2 tablets or capsules for oral administration, in some embodiments, the medicament is suitable for achieving a steady state drug concentration in a patient in about 10, or about 11 , or about 12, or about 13, or about 14, or about 15, or about 16, or about 17 days.

In some aspects, the present invention provides for an ora! dosage regimen of from about 300-400 mg of quinacrine comprising at least about 30 unit doses, or about 60 unit doses, or about 90 unit doses, or about 120 unit doses, or about 150 unit doses, or about 180 unit doses. In some embodiments, the unit doses are for daily administration.

In some aspects, the present invention provides for kits that can simplify the administration of any agent described herein, including quinacrine. An illustrative kit of the invention comprises any composition described herein in unit dosage forms including loading unit doses and maintenance unit doses. In one embodiment, the unit dosage form is a container, a pill box (including one that Is divided by days) or a blister pack, containing any agent described herein and a pharmaceutically acceptable carrier, diluent, excipient, or vehicle. The kit can further comprise a label or printed Instructions instructing the use of any agent described herein. The kit can also further comprise one or more additional agent described herein. In one embodiment, the kit comprises a container containing an effective amount of a composition of the invention and an effective amount of another composition, such those described herein.

In some embodiments, the kit can provide a pill box or a blister pack that provides sufficient daily doses of quinacrine of about 400 mg, or about 450 mg, or about 500 mg. In some embodiments, the pill box or a blister pack contains about 10, or 1 1 , or 12, or 13, or 14, or 15, or 16, or 17 doses. In some embodiments, the kit further contains a pill box or a blister pack that provides sufficient daily maintenance doses of quinacrine (for example, about 350 mg, or 300 mg, or 250 mg, or 200 mg, or 150 mg, or 100 mg). Accordingly, some embodiments may provide for two dose levels of quinacrine for different periods of time. Administration of any quinacrine composition (and/or additional agents) described herein can, Independently, be one to four times daily or one to four times per month or one to six times per year or once every two, three, four or five years. Administration can be for the duration of one day or one month, two months, three months, six months, one year, two years, three years, and may even be for the life of the subject. Chronic, long-term administration of maintenance doses may be indicated in some cases. The dosage may be administered as a single dose or divided into multiple doses. In general, the desired dosage should be administered at set intervals for a prolonged period, usually at least over several weeks or months, although longer periods of administration of several months or years or more may be needed.

The dosage regimen utilizing any quinacrine composition (and/or additional agents) described herein can be selected in accordance with a variety of factors including type, species, age, weight, sex and medical condition of the subject; the severity of the condition to be treated: the route of administration; the renal or hepatic function of the subject; the pharmacogenomlc makeup of the individual; and the specific compound of the invention employed. Any quinacrine composition (and/or additional agents) described herein can be administered in a single daily dose, or the total daily dosage can be administered in divided doses of two, three or four times daily. The dosage of any quinacrine composition (and/or additional agents) described herein as well as the dosing schedule can depend on various parameters, including, but not limited to, the disease being treated, the subject's general health, and the administering physician's discretion. Any agent described herein, can be administered prior to (e.g., 5 minutes, 15 minutes, 30 minutes, 45 minutes, 1 hour, 2 hours, 4 hours, 6 hours, 12 hours, 24 hours, 48 hours, 72 hours, 96 hours, 1 week, 2 ^'weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 8 weeks, or 12 weeks before), concurrently with, or subsequent to (e.g., 5 minutes, 15 minutes, 30 minutes, 45 minutes, 1 hour, 2 hours, 4 hours, 6 hours, 12 hours, 24 hours, 48 hours, 72 hours, 96 hours, 1 ^'week, 2 weeks, 3 weeks, 4 ^'weeks, 5 weeks, 6 weeks, 8 ^'weeks, or 12 weeks after) the administration of an additional therapeutic agent, to a subject in need thereof. In various embodiments any agent described herein is administered 1 minute apart, 10 minutes apart, 30 minutes apart, less than 1 hour apart, 1 hour apart, 1 hour to 2 hours apart, 2 hours to 3 hours apart, 3 hours to 4 hours apart, 4 hours to 5 hours apart, 5 hours to 6 hours apart, 6 hours to 7 hours apart, 7 hours to 8 hours apart, 8 hours to 9 hours apart, 9 hours to 10 hours apart, 10 hours to 11 hours apart, 11 hours to 12 hours apart, no more than 24 hours apart or no more than 48 hours apart.

The amount of any quinacrine composition (and/or additional agents) described herein that is admixed with the carrier materials to produce a single dosage can vary depending upon the subject being treated and the particular mode of administration, in vitro or in vivo assays can be employed to help identify optimal dosage ranges.

In general, the doses that are useful are known to those In the art. For example, doses may be determined with reference Physicians' Desk Reference, 66th Edition, PDR Network; 2012 Edition (December 27, 2011), the contents of which are incorporated by reference in its entirety. In some embodiment, the present invention allows a patient to receive doses that exceed those determined with reference Physicians' Desk Reference. The dosage of any quinacrine composition (and/or additional agents) described herein can depend on several factors Including the severity of the condition, whether the condition is to be treated or prevented, and the age, ^'weight, and health of the subject to be treated. Additionally, pharmaeogenom!c (the effect of genotype on the pharmacokinetic, pharmacodynamic or efficacy profile of a therapeutic) information about a particular subject may affect dosage used. Furthermore, the exact Individual dosages can be adjusted somewhat depending on a variety of factors, including the specific combination of the agents being administered, the time of administration, the route of administration, the nature of the formulation, the rate of excretion, the particular disease being treated, the severity of the disorder, and the anatomical location of the disorder. Some variations in the dosage can be expected.

In various embodiments, the cancer described herein may be a primary cancer or a metastatic cancer. The primary cancer may be an area of cancer cells at an originating site that becomes clinically detectable, and may be a primary tumor. In contrast, the metastatic cancer may be the spread of a disease from one organ or part to another non-adjacent organ or part. The metastatic cancer may be caused by a cancer cell that acquires the ability to penetrate and infiltrate surrounding normal tissues in a local area, forming a new tumor, which may be a local metastasis. "Metastasis" refers to the spread of cancer from a primary site to other places In the body- Cancer cells can break away from a primary tumor, penetrate into lymphatic and blood vessels, circulate through the bloodstream, and grow in a distant focus (metastasize) in normal tissues elsewhere in the body. Metastasis can be local or distant. Metastasis is a sequential process, contingent on tumor ceils breaking off from the primary tumor, traveling through the bloodstream, and stopping at a distant site. At the new site, the cells establish a blood supply and can grow to form a life -threatening mass. Both stimulatory and inhibitory molecular pathways within the tumor cell regulate this behavior, and interactions between the tumor cell and host ceils in the distant site are also significant. Metastases may be detected through the sole or combined use of magnetic resonance imaging (MR!) scans, computed tomography (CT) scans, blood and platelet counts, liver function studies, chest X-rays and bone scans in addition to the monitoring of specific symptoms.

The cancer may also be caused by a cancer cell that acquires the ability to penetrate the walls of lymphatic and/or blood vessels, after which the cancer cell is able to circulate through the bloodstream (thereby being a circulating tumor cell) to other sites and tissues in the body. The cancer may be due to a process such as lymphatic or hematogeneous spread. The cancer may also be caused by a tumor ceil that comes to rest at another site, re-penetrates through the vessel or walls, continues to multiply, and eventually forms another clinically detectable tumor. The cancer may be this new tumor, which may be a metastatic (or secondary) tumor.

The cancer may be caused by tumor ceils that have metastasized, which may be a secondary or metastatic tumor. The cells of the tumor may be like those in the original tumor. As an example, if a breast cancer or colon cancer metastasizes to the liver, the secondary tumor, ^'while present in the liver, is made up of abnormal breast or colon cells, not of abnormal liver cells. The tumor in the liver may thus be a metastatic breast cancer or a metastatic colon cancer, not liver cancer. The cancer may have an origin from any tissue. The cancer may originate from melanoma, colon, breast, or prostate, and thus may be made up of cells that were originally skin, colon, breast, or prostate, respectively. The cancer may also be a hematological malignancy, which may be lymphoma.

In various embodiments, the cancer described herein is a liver cancer. In various embodiments, the iiver cancer described herein is primary Iiver cancer. In various embodiments, the primary Iiver cancer Is one of hepatocellular carcinoma (HCC), cholangiocarcinoma, angiosarcoma, and hepatoblastoma. In some embodiments, quinacrine is used in the manufacture of a medicament or in the treatment of a primary liver cancer in patient with cirrhosis. In various embodiments, the present invention includes treatment of primary liver cancers that are related to one or more of the following risk factors of liver cancer: cirrhosis, high alcohol consumption (including alcoholism), non alcoholic fatty Iiver disease, infection with hepatitis viruses, smoking, low immunity, family history, diabetes, gallbladder removal, radiation from X-rays or CT scans, high body weight, betel quid consumption, and aflatoxin consumption.

In some embodiments, the liver cancer Is a secondary Iiver cancer. In various embodiments, the secondary liver cancer is derived from one or more of the types of primary cancers that often metastasize to the liver, including, for example, colon, lung, stomach, pancreatic, breast cancers, biliary tract, esophageal, ovarian, prostate, kidney cancer, and melanoma. In some embodiments, the Invention relates to a method of treating cancer, comprising administering an effective amount of a quinacrine neoadjuvant therapy to a patient afflicted with a tumor likely to metastasize to the liver as enumerated above. In other embodiments, the secondary iiver cancer Is derived from one or more of a basal ceil carcinoma, biliary tract cancer: bladder cancer; bone cancer; brain and central nervous system cancer; breast cancer: cancer of the peritoneum; cervical cancer; choriocarcinoma; colon and rectum cancer; connective tissue cancer; cancer of the digestive system; endometrial cancer; esophageal cancer; eye cancer; cancer of the head and neck; gastric cancer (including gastrointestinal cancer); glioblastoma; hepatic carcinoma; hepatoma; intra-epitheiiai neoplasm; kidney or renal cancer; larynx cancer; leukemia; iiver cancer; lung cancer (e.g., small-cell lung cancer, non-small cell lung cancer, adenocarcinoma of the lung, and squamous carcinoma of the lung); melanoma; myeloma; neuroblastoma; oral cavity cancer (lip, tongue, mouth, and pharynx); ovarian cancer; pancreatic cancer; prostate cancer; retinoblastoma; rhabdomyosarcoma; rectal cancer; cancer of the respiratory system; salivary gland carcinoma; sarcoma; skin cancer; squamous cell cancer; stomach cancer; testicular cancer; thyroid cancer; uterine or endometrial cancer; cancer of the urinary system; vulva! cancer; lymphoma including Hodgkin's and non-Hodgkin's lymphoma, as well as B-ce!! lymphoma (Including low grade/foilicular non-Hodgkin's lymphoma (NHL); small lymphocytic (SL) NHL; Intermediate grade/follicular NHL; intermediate grade diffuse NHL; high grade immunob!astic NHL; high grade lymphoblastic NHL; high grade small non-cleaved cell NHL; bulky disease NHL; mantle cell lymphoma; A!DS-related lymphoma; and Waldenstrom's acroglobulinemia; chronic lymphocytic leukemia (CLL); acute lymphoblastic leukemia (ALL); Hairy cell leukemia; chronic myeloblasts leukemia; as well as other carcinomas and sarcomas; and post-transplant lymphoproliferative disorder (PTLD), as well as abnormal vascular proliferation associated with phakomatoses, edema (such as that associated with brain tumors), and Meigs' syndrome. In various embodiments, the liver cancer described herein has one or more of a liver cancer tissue marker, selected from, for example: GPC3; GPC3 + heat shock protein 70 + g!utamine synthetase; Telomerase; Proliferating cell nuclear antigen labeling Index; Ki-67; !B-1 E-eadherin, and β-caten!n. in various embodiments, the liver cancer described herein has one or more of a liver cancer serum marker selected from, for example: AFP; AFP-concanavalin A; AFP-LCA binding; HCC-speciflc AFP band on Isoelectric focusing (monosialylated AFP); AFP lectin-affinity subgroups (LCAreactive LCA-L3; eiythroagglutinatingphytohemagg!utiniri- E4 reactive AFP-P4 and P5); Circulating free AFP-lg compiexes; DCP/prothrombin produced by vitamin K absence or antagonism II; Soluble NH2 fragment of GPC-3, a heparin sulfate proteoglycan; Golgi protein 73; Iso-yGTP; Ferritin; Variant alkaline phosphatase; a 1 -Antitrypsin; a1-Acid glycoprotein; Osteopontin; Aldolase A; 5iprlmej- Nucleotide phosphodiesterase; CK18, CK19, TPA, TPS; Circulating free squamous cell carcinoma antigen-lg complexes; a-Fucosyi-transferase; a-L-fucosidase; Transforming growthfactor β1 ; Intercellular cell adhesion molecule 1 ; Anti-p53 antibody; !nterieukin 8; Interleukin 6; Insulin-like growth factor H; Telomerase or telomerase reverse transcriptase m NA; Vascular endothelial growth factor; Variant wild-type estrogen receptor; Vitamin B12-blndingprotein; Neurotensin; Free nucleic acids; Circulating cell-free serum DNA; Epigenetic abnormalities such as, for example, p16 hypermethylatlon; and Plasma proteasome.

In various embodiments, the liver cancer described herein has one or more of a liver cancer tumor ceil marker selected from, for example: circulating tumor cells in peripheral blood detected by RTPCR of AFP mRNA. In various embodiments, the liver cancer described herein has one or more of a liver cancer genetic marker, selected from, for example: plasma glutamate carboxy-peptidase phospholipases A2 G13 and G7 and other cDNA microarray-derived encoded proteins; Melanoma antigen gene 1 , 3; synovia! sarcoma on X chromosome 1 , 2, 4, 5; sarcoplasmic calcium-binding protein 1 ; New York esophageal squamous cell carcinoma 1 ; and Circulating methylated DNA (ras association domain family 1A).

In some embodiments, the liver cancer expresses a!pha-fetoprotein. Further details of markers that define, in some embodiments, the liver cancers of the present Invention are found at, for example, Sturgeon, et al. Use of Tumor Markers In Liver, Bladder, Cervical, and Gastric Cancers American Association for Clinical Chemistry, Inc. (2010), the contents of which are hereby incorporated by reference.

In various embodiments, the liver cancer described herein is classified as one or more of localized resectable, localized unresectable, advanced and recurrent.

Localized resectable liver cancer (some T1 or T2, NO, MO tumors) refers to an early stage cancer. Often the size of the tumor(s) is small and nearby blood vessels are not affected. Further, this type of cancer is often characterized by generally acceptable liver function and general health. In these embodiments, quinacrine may be used as the sole treatment to shrink small tumors and obviate the need for surgery (especially in, for example, patients that cannot easily undergo surgery, like the elderly). Or, in these embodiments, quinacrine may be used as an adjuvant or neoadjuvant therapy to compliment, for example, surgical resection and improve clinical outcome. Localized unresectable liver cancer (some T1 to T4, NO, MO tumors) refers to cancers that haven't yet spread, but that can't be removed safely by surgical resection for various reasons (e.g. a tumor is too large to be removed safely, a tumor is in a part of the liver that makes it hard to remove (such as very close to a large blood vessel), there are several tumors, and the rest of the liver is unhealthy (because of cirrhosis or other reasons)). These patients may be treated with a liver transplant if it is possible, in these embodiments, quinacrine may be used as a therapy that bridges the treatment gap to transplant (e.g. maintains patient health and/or suppresses tumor growth and/or metastasis until transplantation is possible). In some cases, quinacrine treatment may shrink the tumor(s) enough so that surgery (surgical resection or transplant) may become possible.

Advanced liver cancer (Includes all N1 or M1 tumors) refers to cancers that have spread outside the liver (either to the lymph nodes or to other organs). Because these cancers are widespread, they cannot be treated with surgery, if your liver is functioning well enough (e.g. Chiid-Pugh class A or B), quinacrine alone or as a combination therapy {e.g. ^'with the targeted therapy soraferiib) may help control the growth of the cancer for a time and may extend life.

Recurrent liver cancer refers to cancer that after treatment. Recurrence can be local (in or near the same place it started) or distant (spread to organs such as the lungs or bone). Treatment of liver cancer that returns after initial therapy depends on many factors, Including where It comes back, the type of initial treatment, and how well the liver is functioning. Patients with localized resectable disease that recurs in the liver might be eligible for further surgery or local treatments like ablation or embolization. If the cancer is widespread, targeted therapy (e.g. soraferiib) or chemotherapy may be options. In all of these scenarios, quinacrine may be used to compliment or supplant treatment plans.

In various embodiments, the liver cancer described herein is classified with the AJCC (TN ) staging system. Stages are labeled using Roman numerals I through IV (1-4). Some stages are further sub-divided Into A and B or even C. For the most part, the lower the number, the less the cancer has spread. A higher number, such as stage IV (4), means a more advanced cancer. The staging systems for most types of cancer depend only on the extent of the cancer, but most patients with liver cancer have damage to the rest of their liver along with their cancer. This means that the liver might not be ^'working as well as it should, which also affects treatment options and the outlook for the patient. Quinacrine finds uses across this spectrum of stages (e.g. IA, or IB, or IC, or IIA, or lib, or NIC, or IIIA, or IIIB, or IliC, or IVA, or IVB, or IVC, including one or more of IX, TO, Tls, T1 , T2, T3, T4, NX, NO, N1 , N2, N3, MX, MO, M1 and any grades of 1 , or 2, or 3, or 4, or 5). For example, quinacrine is useful to provide liver cancer cure or attenuation In lower stages or it may be used as a palliative treatment in higher stages. In ail stages, quinacrine may be used as an adjuvant or neoadjuvant.

In various embodiments, the invention relates to uses of quinacrine in the manufacture of a medicament in the treatment of various other cancers besides liver, including primary cancers. In various embodiments, the invention relates to methods of treating of various other cancers besides liver, including primary cancers. For example, the cancers for treatment may be one or more of those described in Table 3 and Table 8.

In various embodiments, the liver cancer is present in a liver having cells that overexpress one or more muitidrug-resistance pumps, selected from, for example, the multidrug-resistance pump Is one or more of the P- glycoprotein and the multidrug resistance-associated protein (MRP). In various embodiments, quinacrine's ability to direct to the liver may offset hepatic ability to efflux it from liver ceils.

In various embodiments, the treatment as described herein refers to, for example, reducing liver tumor load, reducing liver tumor volume, affecting a biomarker level (e.g. decreasing AFP levels), and affecting a clinical marker of hepatic function (e.g. reducing a clinical marker of impaired hepatic function, such as elevated aminotransferase levels, elevated plasma ammonia levels, changes in one or more of alanine aminotransferase (ALT), aspartate aminotransferase (AST), alkaline phosphatase, 5' nucleotidase, a gamma-glutamyl transpeptidase (GGT), and bilirubin).

In some embodiments, the therapeutic effect of quinacrine may be evaluated using a blood test or biopsy to measure the aforementioned treatment effects (including, for example, immunometric assays). In some embodiments, the therapeutic effect of quinacrine may be evaluated using, for example, hepatic imaging (e.g. one or more of ultrasounds, CT, MR, and PET, PET/CT and may involve various Imaging agents (e.g. gadolinium chelates)). Techniques for imaging may be found in Ros and Morteie, Hepatic Imaging. An overview Clin Liver Dis. 2002 Feb;6(1 ): 1-16, the contents of which are hereby incorporated by reference in their entirety.

In some embodiments, the present invention provides for methods of using quinacrine as palliative care, including, for example, in patients with late stage liver cancer.

In some embodiments, the present invention provides for methods of using quinacrine as adjuvant and/or neoadjuvant or uses of quinacrine in the manufacture of a medicament suitable for the treatment of liver cancer as adjuvant and/or neoadjuvant. In one embodiment, the quinacrine is useful to increase the effectiveness of chemotherapies that provide little to no benefit to the patient. In another embodiment, quinacrine is useful to increase the effectiveness of chemotherapies that fail to provide long term beneficial liver cancer effects, including, for example, doxorubicin or 5-fluorouracil or cisplatin. In one embodiment, a patient receives quinacrine to make other resection more tenable or effective. For example, quinacrine can be used as a neoadjuvant to make a patient suitable for resection (e.g. by reducing the size and/or number of tumors) or quinacrine can be used as an adjuvant to improve the likelihood that resection will be successful (e.g. by reducing the likelihood of recurrence). In some embodiments, quinacrine can be used as a therapy that prepares a patient for transplantation.

In various embodiments, the present invention relates to uses or methods of treatment of a patient that is afflicted with a liver disease selected from, for example, Alpha 1 Anii-Trypsin Deficiency, Autoimmune Hepatitis, Biliary Atresia, Cirrhosis, Cystic Disease of the Liver, Fatty Liver Disease, Galactosemia, Gallstones, Gilbert's Syndrome, Hemochromatosis, Liver Cancer, Liver disease In pregnancy, Neonatal Hepatitis, Primary Biliary Cirrhosis, Primary Sclerosing Cholangitis, Porphyria, Reye's Syndrome, Sarcoidosis, Toxic Hepatitis, Type 1 Glycogen Storage Disease, Tyrosinemia, Viral Hepatitis A, B, C, and Wilson Disease. In various embodiments, any of these diseases make the patient less suitable for transplant or resection and qulnacrine may thus be used as an adjuvant or neoadjuvant.

In some embodiments, quinacrine is used with one or more combination agents, including but not limited to sorafenib, TRC105, Erlotinlb, Bevacizumab, doxorubicin, 5-fluorouracil, and cisplatin. in some embodiments, quinacrine is part of a chemoteherapeutic cocktail comprising any of the combination agents described herein. In some embodiments, quinacrine Is part of a chemoteherapeutic cocktail comprising any of the combination agents described herein that is used for chemoemboiizatlon. In various embodiments, there is provided a method of treating liver cancer In a patient undergoing treatment with one or more combination agents described herein comprising administering an effective amount of quinacrine (e.g. a liver cancer patient undergoing treatment with one or more of sorafenib, TRC105, doxorubicin, 5-fluorouracil, and cisplatin).

Further examples of combination agents are also provided and Include, but are not limited to, alkylating agents such as thiotepa and CYTOXAN cyc!osphosphamide; alkyl sulfonates such as busulfan, improsulfan and piposulfan; aziridlnes such as benzodopa, carboquone, meturedopa, and uredopa; ethylenimlnes and methylamelamines including a!tretamine, triethyienemeiamine, trietyienephosphoramide, triethiylenethiophosphoramide and trimethyiolomelamlne; acetogenins (e.g., bullatacin and buiiatacinone); a camptothecin (including the synthetic analogue topotecan); bryostatin; caily statin; CC-1065 (including its adozelesin, carzelesin and bizelesin synthetic analogues); cryptophycins (e.g., cryptophycin 1 and cryptophycin 8); dolastatin; duoearmycln (including the synthetic analogues, KW-2189 and CB 1-TM1 ); eleutherobln; pancratistatin; a sarcodictyin; sponglstatin; nitrogen mustards such as chlorambucil, chiornaphazine, choiophosphamide, estramusiine, ifosfamide, mechloreiham!ne, mechloreihamine oxide hydrochloride, melphaian, novembichin, phenesterine, prednimustine, trofosfamide, uracil mustard; nitrosureas such as carmustine, chlorozotocin, fotemustine, lomustine, nimustine, and ranimnustine; antibiotics such as the enediyne antibiotics (e.g., ca!icheamicin, especially calicheamicin gamma!i and calicheamicin omegaii (see, e.g. , Agnew, Chem. Intl. Ed. Engl., 33: 183-186 (1994)); dynem!c!n, including dynemicin A; bisphosphonates, such as clodronate; an esperamicin; as well as neocarzinostatin chromophore and related chromoprotein enediyne antibiotic chromophores), ac!acinomysins, actinomycin, authramycin, azaserlne, bleomycins, cactinomydn, carabicin, caminomycin, carzinophilin, chromomycinis, dactinomycin, daunorubicin, detorubicin, 6-diazo-5-oxo-L- norleucine, AD IA YCIN doxorubicin (including morpholino- doxorubicin, cyanomorpholino-doxorubicin, 2- pyrrolino-doxorubicin and deoxy doxorubicin), epirubicin, esorubicin, idarubicin, marcellomycin, mitomycins such as mitomycin C, mycophenolic acid, nogalamycin, olivomycins, peplomycin, potfiromycin, puromycin, quelamycin, rodorubicin, streptonigrin, streptozocin, tubercidin, ubenimex, zinostatin, zorublcin; anti-metabolites such as methotrexate and 5-fluorouracil (5-FU); folic acid analogues such as denopterin, methotrexate, pteropterin, trimetrexate; purine analogs such as fludarabine, 6-mercaptopurine, thiamiprine, thioguanine; pyrimidine analogs such as ancitabine, azacitidine, 6-azauridine, carmofur, cytarabine, dideoxyuridine, doxlf!uridine, enocitabine, floxuridine; androgens such as calusterone, dromostanolone propionate, epitiostanol, mepitiostane, testolactone; anti-adrenals such as minogiuteihimide, mitotane, triiostane; folic acid replenisher such as fro!inic acid: aceglatone; aidophosphamide glycoside; aminolevulinic acid: eni!uracil; amsacrine; bestrabucii; bisanirene; edatraxate; def of amine; demecolcine; diaziquone; eiformlthine; eiiiptlnium acetate; an epothilone; etoglucid; gaiiium nitrate; hydroxyurea; lentinan; lonidainine; maytansinoids such as maytansine and arisamitocins; mitoguazone; mitoxantrone; mopidanmol; nitraerine; pentostatin; phenamet; pirarubicln; losoxantrorie; podophylllnic acid; 2-ethylhydrazide; procarbazine; PSK polysaccharide complex (JHS Natural Products, Eugene, Oreg.); razoxane; rhizoxin; sizofuran; spirogermanium; tenuazonic acid; trlaziquone; 2,2',2"- trichlorotriethyiamine; trichothecenes {e.g., T-2 toxin, verracurin A, roridin A and anguidine); urethan; vindesine; dacarbazine; mannomustine; mitobronitoi; mitolactoi; pipobroman; gacytosine; arablnoside ("Ara-C"); cyclophosphamide; thiotepa; taxoids, e.g., TAXOL paciitaxei (Bristol-Myers Squibb Oncology, Princeton, N.J.), ABRAXANE Cremophor-free, albumin-engineered nanopartic!e formulation of paciitaxei (American Pharmaceutical Partners, Schaumberg, 11 1.), and TAXOTERE doxetaxe! (Rhone-Pou!enc Rorer, Antony, France); chlorambucil; GEMZAR gemcitabine; 6-thloguanine; mercaptopurlne; methotrexate; platinum analogs such as cisp!atin, oxa!ipiatln and carbopiatln; vinblastine; platinum; etoposide (VP-16); ifosfamide; mitoxantrone; vincristine; NAVELBiNE. vinoreibine; novantrone; teniposlde; edatrexate; daunomycin; amine-pterin; xeloda; ibandronate; irinotecan (Camptosar, CPT-11) (including the treatment regimen of irlnotecan with 5-FU and leucovorin); topoisomerase inhibitor RFS 2000; difiuoromethy!ornithine (D FO); retinoids such as retlnoic acid; capecitabine; combretastatin; leucovorin (LV); oxaliplaiin, including the oxa!iplatln treatment regimen (FOLFOX); lapatinib (Tykerb); inhibitors of PKC-a, Raf, H-Ras, EGFR (e.g., erlotinib (Tarceva)) and VEGF-A that reduce cell proliferation and pharmaceutically acceptable salts, acids or derivatives of any of the above. In addition, the methods of treatment can further include the use of radiation. In addition, the methods of treatment can further include the use of photodynamic therapy.

In some embodiments, the quinacrine compositions (and/or additional agents) described herein, include derivatives that are modified, i.e., by the covalent attachment of any type of molecule to the composition such that covalent attachment does not prevent the activity of the composition. For example, but not by way of limitation, derivatives include composition that have been modified by, inter alia, glycosylation, lipidation, acetyiation, pegylation, phosphorylation, amidation, derivatization by known protecting/blocking groups, proteolytic cleavage, linkage to a cellular iigand or other protein, etc. Any of numerous chemical modifications can be carried out by known techniques, including, but not limited to specific chemical cleavage, acetyiation, formyiation, metabolic synthesis of turicamycin, etc. Additionally, the derivative can contain one or more non- classical amino acids.

Quinacrine compositions (and/or additional agents) described herein can possess a sufficiently basic functional group, which can react with an inorganic or organic acid, or a carboxyl group, which can react with an Inorganic or organic base, to form a pharmaceutically acceptable salt. A pharmaceutically acceptable acid addition salt is formed from a pharmaceutically acceptable acid, as is well known In the art. Such salts include the pharmaceutically acceptable salts listed in, for example, Journal of Pharmaceutical Science, 66, 2-19 (1977) and The Handbook of Pharmaceutical Salts; Properties, Selection, and Use. P. H. S!ahl and C. G. Wermuth (eds.), Verlag, Zurich (Switzerland) 2002, which are hereby incorporated by reference in their entirety.

Pharmaceutically acceptable salts include, by way of non-limiting example, sulfate, citrate, acetate, oxalate, chloride, bromide, iodide, nitrate, bisu!fate, phosphate, acid phosphate, isonicotinate, lactate, salicylate, acid citrate, tartrate, oieate, {annate, pantothenate, bltartrate, ascorbaie, succinate, maleate, gentisinate, fumarate, gluconate, glucaronate, saccharate, formate, benzoate, g!utamate, methanesulfonate, ethanesuiionate, benzenesulfonate, p-toluenesulfonate, camphorsulfonate, pamoate, phenyiacetate, trifluoroacetate, acryiate, chlorobenzoate, dinitrobenzoate, hydroxybenzoate, methoxybenzoate, methyibenzoate, o-acetoxybenzoate, naphthalene-2-benzoate, isobutyrate, phenylbutyrate, o-hydroxybutyrate, butyne-1 ,4-dicarboxylate, hexyne-1 ,4- dicarboxyiate, caprate, capryiate, cinnamate, glycoiiate, heptanoate, hippurate, malate, hydroxymaleate, malonate, mandeiate, mesylate, nicotinate, phthalate, teraphthalate, propiolate, propionate, phenylpropionate, sebacate, suberate, p-bromobenzenesuifonate, chlorobenzenesuifonate, ethylsuifonate, 2-hydroxyethylsulfonate, methylsulfonate, naphihaiene-1 -sulfonate, naphthalene-2 -sulfonate, naphthalene-1 ,5-sulfonate, xylenesulfonate, and tartarate salts.

The term "pharmaceutically acceptable salt" also refers to a salt of the compositions of the present invention having an acidic functional group, such as a carboxylic acid functional group, and a base. Suitable bases include, but are not limited to, hydroxides of alkali metals such as sodium, potassium, and lithium; hydroxides of alkaline earth metal such as calcium and magnesium; hydroxides of other metals, such as aluminum and zinc; ammonia, and organic amines, such as unsubstituted or hydroxy-substituted mono-, di-, or tri-alkyiamines, dicyclohexylamine; tributyi amine; pyridine; N-methyi, N-ethylamine; diethyiamine; triethylamine; mono-, bis-, or tris-(2-OH-lower alkyiamines), such as mono-; bis-, or tris-(2-hydroxyethyi)amine, 2-hydroxy-tert-butylamine, or tris-(hydroxymethy!)methy!amine, N,N-di-lower alky!-N-(hydroxyl-lower alkyl)-amines, such as N,N-dimethyl-N-(2- hydroxyethyl)amine or tri-(2-hydroxyethyi)amine; N-methyi-D-glucamine; and amino acids such as arginine, lysine, and the like. In some embodiments, the compositions described herein are in the form of a pharmaceutically acceptable salt.

Further, any quinacrine compositions (and/or additional agents) described herein can be administered to a subject as a component of a composition that comprises a pharmaceutically acceptable carrier or vehicle. Such compositions can optionally comprise a suitable amount of a pharmaceutically acceptable excipient so as to provide the form for proper administration.

Pharmaceutical exclpients can be liquids, such as water and oils, including those of petroleum, animal, vegetable, or synthetic origin, such as peanut oil, soybean oil, mineral oil, sesame oil and the like. The pharmaceutical exclpients can be, for example, saline, gum acacia, gelatin, starch paste, talc, keratin, colloidal silica, urea and the like, in addition, auxiliary, stabilizing, thickening, lubricating, and coloring agents can be used. In one embodiment, the pharmaceutically acceptable exclpients are sterile when administered to a subject. Water Is a useful excipient when any agent described herein is administered intravenously. Saline solutions and aqueous dextrose and glycerol solutions can also be employed as liquid excipients, specifically for injectable soiutions. Suitable pharmaceutical excipients also include starch, glucose, lactose, sucrose, gelatin, malt, rice, flour, chalk, silica gel, sodium stearate, glycerol monostearate, talc, sodium chloride, dried skim milk, glycerol, propylene, glycol, water, ethanol and the like. Any agent described herein, if desired, can also comprise minor amounts of wetting or emulsifying agents, or pH buffering agents.

The present Invention Includes the described quinacrine related compositions (and/or additional agents) in various formulations. Any quinacrine composition (and/or additional agents) described herein can take the form of solutions, suspensions, emulsion, drops, tablets, pills, pellets, capsules, capsules containing liquids, powders, sustained-release formulations, suppositories, emulsions, aerosols, sprays, suspensions, or any other form suitable for use. In one embodiment, the composition is in the form of a capsule (see, e.g., U.S. Patent No. 5,898,155, the contents of which are hereby Incorporated herein by reference). Other examples of suitable pharmaceutical excipients are described in Remington's Pharmaceutical Sciences 1447-1676 (Alfonso R. Gennaro eds., 19th ed. 1995), the contents of which are hereby incorporated herein by reference.

Where necessary, the quinacrine compositions (and/or additional agents) can also include a soiubilizing agent. Also, the agents can be delivered with a suitable vehicle or delivery device as known in the art. Combination therapies outlined herein can be co-delivered In a single delivery vehicle or delivery device. Compositions for administration can optionaiiy include a local anesthetic such as, for example, lignocaine to lessen pain at the site of the injection,

The formulations comprising the quinacrine compositions (and/or additional agents) of the present invention may conveniently be presented in unit dosage forms and may be prepared by any of the methods well known in the art of pharmacy. Such methods generally include the step of bringing the therapeutic agents into association with a carrier, which constitutes one or more accessory ingredients. Typically, the formulations are prepared by uniformly and intimately bringing the therapeutic agent into association with a liquid carrier, a finely divided solid carrier, or both, and then, if necessary, shaping the product into dosage forms of the desired formulation (e.g., wet or dry granulation, powder blends, etc., followed by tabieting using conventional methods known in the art)

In one embodiment, any quinacrine composition (and/or additional agents) described herein is formulated in accordance with routine procedures as a composition adapted for a mode of administration described herein.

Routes of administration include, for example: oral, intradermal, intramuscular, intraperitoneal, intravenous, subcutaneous, intranasal, epidural, sublingual, intranasal, intracerebral, intravaginal, transdermal, recta!ly, by inhalation, or topically, particularly to the ears, nose, eyes, or skin. In some embodiments, the administering is effected orally or by parenteral Injection. The mode of administration can be left to the discretion of the practitioner, and depends in-part upon the site of the medical condition. In most instances, administration results in the release of any agent described herein into the bloodstream. Any quinacrine composition (and/or additional agents) described herein can be administered orally. Such quinacrine compositions (and/or additional agents) can also be administered by any other convenient route, for example, by Intravenous Infusion or bolus Injection, by absorption through epithelial or mucocutaneous linings (e.g., oral mucosa, rectal and intestinal mucosa, etc.) and can be administered together ^'with another biologically active agent. Administration can be systemic or local. Various delivery systems are known, e.g., encapsulation in liposomes, microparticles, microcapsules, capsules, efc, and can be used to administer.

In specific embodiments, It may be desirable to administer locally to the area In need of treatment.

In one embodiment, any quinacrine composition (and/or additional agents) described herein is formulated in accordance with routine procedures as a composition adapted for oral administration to humans. Compositions for oral delivery can be in the form of tablets, lozenges, aqueous or oi!y suspensions, granules, powders, emulsions, capsules, syrups, or elixirs, for example. Orally administered compositions can comprise one or more agents, for example, sweetening agents such as fructose, aspartame or saccharin; flavoring agents such as peppermint, oil of wintergreen, or cherry; coloring agents; and preserving agents, to provide a pharmaceutically palatable preparation. Moreover, where in tablet or pill form, the compositions can be coated to delay disintegration and absorption in the gastrointestinal tract thereby providing a sustained action over an extended period of time. Selectively permeable membranes surrounding an osmotically active driving any quinacrine composition (and/or additional agents) described herein are also suitable for orally administered compositions. In these latter platforms, fluid from the environment surrounding the capsule Is imbibed by the driving compound, ^'which swells to displace the agent or agent composition through an aperture. These delivery platforms can provide an essentially zero order delivery profile as opposed to the spiked profiles of immediate release formulations. A time-delay material such as glycerol monostearate or glycerol stearate can also be useful. Oral compositions can include standard excipienis such as mannitoi, lactose, starch, magnesium stearate, sodium saccharin, cellulose, and magnesium carbonate. In one embodiment, the excipients are of pharmaceutical grade- Suspensions, in addition to the active compounds, may contain suspending agents such as, for example, eihoxy!ated Isosteary! alcohols, poiyoxyethyiene sorbitol and sorbltan esters, microcrysta!iine cellulose, aluminum metahydroxide, bentonite, agar-agar, tragacanth, etc.. and mixtures thereof.

Dosage forms suitable for parenteral administration (e.g. intravenous, intramuscular, intraperitoneal, subcutaneous and intra-articular injection and infusion) Include, for example, solutions, suspensions, dispersions, emulsions, and the like. They may also be manufactured In the form of sterile solid compositions (e.g. iyophilized composition), which can be dissolved or suspended In sterile injectable medium immediately before use. They may contain, for example, suspending or dispersing agents known in the art.

In another embodiment, delivery can be In a vesicle, in particular a liposome (see Langer, 1990, Science 249: 1527-1533; Treat ef a/., in Liposomes in the Therapy of infectious Disease and Cancer, Lopez- Berestein and Fidler (eds.), Liss, New York, pp. 353-365 (1989), Any quinacrine composition (and/or additional agents) described herein can be administered by coniroiied- reiease or sustained-release means or by delivery devices that are well known to those of ordinary skill In the art. Examples Include, but are not limited to, those described In U.S. Patent Nos. 3,845,770; 3,916,899; 3,536,809; 3,598,123; 4,008,719; 5,674,533; 5,059,595; 5,591 ,767; 5,120,548; 5,073,543; 5,639,476; 5,354,556; and 5,733,556, each of which Is incorporated herein by reference in its entirety. Such dosage forms can be useful for providing controlled- or sustained-release of one or more active ingredients using, for example, hydropropylmethyi cellulose, other polymer matrices, gels, permeable membranes, osmotic systems, multilayer coatings, microparticles, liposomes, microspheres, or a combination thereof to provide the desired release profile in varying proportions. Suitable controlled- or sustained-release formulations known to those skilled in the art, including those described herein, can be readily selected for use with the active ingredients of the agents described herein. The invention thus provides single unit dosage forms suitable for oral administration such as, but not limited to, tablets, capsules, geicaps, and caplets that are adapted for controlled- or sustained-release.

In some embodiments, the quinacrine composition is formulated and/or conjugated to increase the bioavailability and decrease a clearance of the drug. For example, quinacrine may be conjugated to a polymer, such as, for example, polyethylene glycol) (PEG), polysiaiic acid, giycolic acid, polyamino acid polymers (e.g. poiygiutamic acid (PGA), N-(2-hydroxypropyl)methacrylamide copolymer (HP A), and hybrid modified PEG polymers), and Hydroxyethyi starch (HES). Also, quinacrine can be incorporated into drug delivery systems that slow or prevent clearance, such as, for example, polymeric micelles, hydrogels, microparticles, self-diffusion systems, biodegradable polymers, cellulose derivatives, porous membranes, and dendrimers. Also, quinacrine can be incorpotrated into lipid delivery systems, such as, for example, liposomes, solid lipid nanopartic!es, oily suspensions, submicron lipid emulsions, lipid implants, lipid microbubbies, inverse lipid micelles, cochiiar liposomes, and lipid microtubules, and lipid microcylinders.

Controlled- or sustained-release of an active ingredient can be stimulated by various conditions, including but not limited to, changes in pH, changes In temperature, stimulation by an appropriate wavelength of light, concentration or availability of enzymes, concentration or availability of water, or other physiological conditions or compounds.

In another embodiment, polymeric materials can be used (see Medical Applications of Controlled Release, Langer and Wise (eds.), CRC Pres., Boca Raton, Florida (1974); Controlled Drug Bioavailability, Drug Product Design and Performance, Smolen and Ball (eds.), Wiley, New York (1984); Ranger and Peppas, 1983, J. Macromoi. Sci. Rev. Macromoi. Chem. 23:61 ; see also Levy ef a/., 1985, Science 228: 190; During ei a/., 1989, Ann. Neurol. 25:351 ; Howard ef a/. , 1989, j. Neurosurg. 71 :105).

In another embodiment, a controlled-release system can be placed in proximity of the target area to be treated, thus requiring only a fraction of the systemic dose (see, e.g., Goodson, in Medical Applications of Controlled Release, supra, vol. 2, pp. 115-138 (1984)). Other controlled-release systems discussed in the review by Langer, 1990, Science 249: 1527-1533) may be used. In some embodiments, the subject and/or animal is a mammal, e.g., a human, mouse, rat, guinea pig, dog, cat, horse, cow, pig, rabbit, sheep, or non-human primate, such as a monkey, chimpanzee, or baboon. In other embodiments, the subject and/or animal is a non-mammal, such, for example, a zebrafish. In some embodiments, the subject and/or animal may comprise fluorescent!y-tagged cells (with e.g. GFP). In some embodiments, the subject and/or animal is a transgenic animal comprising a fluorescent ceil.

In some embodiments, the subject and/or animal is a human. In some embodiments, the human Is a pediatric human. In other embodiments, the human is an adult human, in other embodiments, the human is a geriatric human. In other embodiments, the human may be referred to as a patient.

In certain embodiments, the human has an age in a range of from about 1 to about 5 years old, from about 5 to about 10 years old, from about 10 to about 15 years old, from about 15 to about 20 years old, from about 20 to about 25 years old, from about 25 to about 30 years old, from about 30 to about 35 years old, from about 35 to about 40 years old, from about 40 to about 45 years old, from about 45 to about 50 years old, from about 50 to about 55 years old, from about 55 to about 60 years old, from about 60 to about 65 years old, from about 65 to about 70 years old, from about 70 to about 75 years old, from about 75 to about 80 years old, from about 80 to about 85 years old, from about 85 to about 90 years old, from about 90 to about 95 years old or from about 95 to about 100 years old.

In specific embodiments, the human has an age in a range of at least 60 years old, or at least 65 years old, or at least 70 years old, or at least 75 years old, or at least 80 years old, or at least 85 years old. Primary liver cancers are particularly prevalent among the elderly. Accordingly, the present invention provides for methods and uses of quinacrine in the treatment of liver cancers in the elderly. In an illustrative embodiment, the use of quinacrine provides an elderly patient that cannot withstand and/or Is resistant to standard treatment, including surgical resection. In another embodiment, an elderly patient, including one that cannot withstand surgical resection is provided quinacrine as an adjuvant or a neoadjuvant treatment alongside one or more of chemotherapy, chemoembolization, radiation therapy, cryosurgery, radiofrequency ablation, or ethanol injection. In another illustrative embodiment, the use of quinacrine provides an elderly patient with palliative care when afflicted with a late stage liver cancer.

In other embodiments, the subject is a non-human animal, and therefore the invention pertains to veterinary use. In a specific embodiment, the non-human animal Is a household pet. In another specific embodiment, the non- human animal is a livestock animal.

This invention is further illustrated by the following non-limiting examples.

EXAMPLES

A muiticenter, open-label, dose escalation, Phase 1 b safety and toierabiiitv study of quinacrine in patients with liver tumors of different etiology was designed and conducted. EXAMPLE 1 : STUDY PLAN (DESCRIPTION OF GENERAL STUDY DESIGN AND PLAN)

After signing the informed consent, patients received screening clinical examinations. The compliance to inclusion/exclusion criteria was confirmed prior to the study therapy.

Study subjects were successively enrolled to 7 dose cohorts. The dose was increased in each next cohort. First, patients took a loading dose. During the next study ^'weeks, beginning from Week 2, the dose ^'was reduced and patients took maintenance dose up to the study completion (Table 1 ). Patients in cohort 5 took the same dose for all 8 weeks of treatment.

Table 1. Dosing regimen of quinacrine

At least 3 patients were enrolled to each cohort. If no patient experienced a dose limiting toxicity (DLT) after 4 weeks of treatment, then the next 3 patients were enrolled at the next cohort. If any patient prematurely discontinued the study therapy from Day 1 to Week 4 for reasons not related to the study therapy, then a new patient was enrolled to the same dose cohort for 3 patients to take the study therapy for at least 4 ^'weeks. Identification numbers of subjects withdrawn from the study were not re-assigned. If at least 1 patient of the cohort experienced DLT, 3 additional patients took the same dose of the study product, if none of the additional 3 patients experienced a DLT (1/6), then dose escalation was continued in the next cohort. If 1 or more of the additional 3 patients experienced a DLT (2/6), then dose escalation was ceased. The previous dose was considered the maximum tolerated dose and was used for the phase I! clinical study. If 2 of the original 3 patients in any dose cohort experienced a DLT, dose escalation was ceased, and the previous dose was considered the maximum tolerated dose.

The decision on possible dose escalation In the next cohort was made by the Data Safety Monitoring Board after 4 weeks of treatment. Review of the results of clinical and laboratory tests ^'were made for these 4 ^'weeks. The decision on maximum tolerated dose was made depending on the nature and degree of toxicity. Dose limiting toxicity was defined by any of the following adverse events: grade 4 neutropenia that lasted longer than 7 days, febrile neutropenia (increased body temperature 38^C'C and ANC < 1000 cl/μϋ, grade 4 thrombocytopenia or grade 3 thrombocytopenia with bleeding, QTcF of >500 msec, and any other drug-related adverse reactions of Grade 3 or 4 toxicity except: nausea or vomiting that responded to standard anti-emetic treatment and non-hematoiogical grade 3 laboratory AEs that were asymptomatic and reversible (returned to baseline or to grade 1 during less than 14 days of quinacrine interruption). Adverse events were graded according to the toxicity criteria of CTCAE 4.0.

The total study period was 2 years and 9 months. Patient enrollment lasted for approximately 2.5 years. The period of study participation for a patient was up to 13 weeks including 1 screening week, 8 weeks of treatment, and 4 weeks of follow-up after treatment discontinuation. 2 patients received extra courses of quinacrine therapy after 8 weeks of treatment when the patients had stable disease. After 8 weeks of treatment with quinacrine, treatment efficacy ^'was assessed in accordance with the RECIST 1.1. Criteria.

The decision to continue quinacrine therapy In study subjects that had not progressed was made individually for each patient. Treatment was continued for patients with stable disease and partial or complete response. Such patients received several extra cycles of the study therapy in the same maintenance dose that was taken in the dose cohort to which the patient was enrolled. Patients with disease progression and patients that had to discontinue quinacrine due to AE or refusal from participation In the study were foilowed-up with for 4 weeks after the study therapy.

Table 2: Study flow chart

*After 8 ^'weeks of quinacrine therapy, efficacy was assessed in accordance with the RECIST 1.1 criteria, a decision to continue quinacrine therapy in patients that had not progressed was made individually.

** Visit Windows ± 1 Day were allowed during the treatment.

1. The procedure was made within 28 days prior to the first dose of quinacrine;

2. Vital signs included: BP, heart rate, respiratory rate, temperature, weight, and height (only on screening);

3. Hematology included: hemoglobin, hematocrit, VVBC differential, RBC, and platelet count;

4- Clinical chemistry included: sodium, potassium, chloride, biearbonate/COj/urea, creatinine, glucose, total bilirubin, ALP, ALT, AST, total protein, albumin, calcium, phosphorus, and magnesium;

5. Urinalysis included: coior, clarity, specific gravity, determination of hemoglobin, bilirubin, glucose, and ketones were made with dipsticks, microscopic examination was made with dipstick determinations 2+ and higher;

6. PK sampling ^'was made on study Day 1 prior to the first dose and30 minutes, 1 , 2, 4, 8, and 24 hours post-dose in patients that gave consent for the procedure;

7. PK sampling was made pre-dose on Day 15 (Week 2), Day 29 (Week 4), and Day 43 (Week 6);

8. Core-needie biopsy of the liver tumor and healthy tissue was made on study Week 4 in patients that gave consent for the procedure. Quinacrine concentration was also determined in the biopsy sample;

9. Serum tumor markers were determined at the same time points as the CT scan. Specific markers were determined for each tumor: for example, aifa- fetoprotein (AFP) for hepatocellular carcinoma, CEA and Ca-19.9 for stomach cancer, colon cancer, cho!angiocarcino as, and pancreatic cancer. If serum levels of the tumor markers were normal prior to treatment, tumor marker levels were not re-determined†

10. Protrombin time (PT);

11. At Week12, clinical chemistry included: creatinine, glucose, total bilirubin, ALP, ALT, AST, total protein, and albumin;

12. After Day 29 (Week 4) - assessment of quinacrine safety/to era bl ! i ty for each patient;

13. The Amendment to the informed Consent was signed on the visit by patients that were recommended an extra course of the study therapy.

14 Quinacrine was prescribed in the same maintenance dose if the patient was recommended an extra course of the study therapy.

Screening visits were made no earlier than one week prior to administration of the study product on study day 1. No study-related procedures were performed prior to obtaining written informed consent. The maximum screening period was 7 days. To determine patients' compliance to necessary selection criteria, the clinical assessment Included the following: demographic data (including birth date, sex, age, and race), medical history (Including information on chronic infections such as HIV, hepatitis C, hepatitis B, and concomitant medications), vital signs (blood pressure, respiration rate, pulse, temperature, height, and weight), assessment of ECOG status, 12-!ead EGG at rest, CT-scan of the liver (made as per institutional standards), laboratory safety tests, tumor marker test, blood coagulation test, andpregnancv test (for women). If a patient met all inclusion criteria and did not have any exclusion criteria, he/she was initiated into the study therapy.

Day 1: on study day 1. the following examinations and procedures were made: physical examination, vital signs (blood pressure, respiration rate, pulse, temperature, and weight), assessment of ECOG status, administration of the study product, assessment of adverse events after administration of the study product, assessment of changes in concomitant medications, PK blood sampling, and patient training in diary completion and diary dispensing.

Day 8 (Week 1), Day 15 (Week 2), Day 22 (Week 3), Day 29 (Week 4), and Day 43 (Week 6): on the above mentioned days, the following examinations and procedures were made: physical examination, vital signs (blood pressure, respiration rate, pulse, temperature, and weight), assessment of ECOG status, laboratory safety tests, 12-lead ECG at rest, blood coagulation test, assessment of adverse events, assessment of changes in concomitant medications, PK blood sampling (Day 15 (W2), Day 29 (W4), and Day 43 (W6) (the test was made only in patients that gave consent for the procedure)), liver biopsy to determine quinacrine concentrate in biopsy sample (made at Week 4 in patients that gave consent for the procedure), and dispensing of patient diary.

Day 57 (Week 8) - end-of study/premature discontinuation visit: all procedures were made after 8 weeks of quinacrine therapy on the visit of Week 8. Visit Windows ± 1 Day were allowed during treatment. All procedures for treatment discontinuation were also made if quinacrine therapy was terminated due to adverse event or patient's refusal from further participation from the study. The following examinations and procedures were made: physical examination, vital signs (blood pressure, respiration rate, pulse, temperature, and weight), assessment of ECOG status, CT-scan of liver (made in patients that completed the 8-week drug therapy), laboratory safety tests, tumor markers, assessment of adverse events, assessment of changes in concomitant medications, assessment of RECIST 1.1 criteria or adoption of the decision on possible continuation of the study therapy (visit procedures as described In Procedures and visits in extra cycles of study therapy) or patient's follow-up in the observation period of the protocol, and dispensing of the patient diary. Note that the conclusion on stable tumor process in patients and the necessity to continue treatment was signed by the study doctor and approved by the National study coordinator of the protocol. Follow-up period: 14 days (± 3 days) after the last dose of IMP - telephone call: the following examination and procedures were made: assessment of ECOG status, assessment of adverse events, and assessment of changes In concomitant medications.

Follow-up period: 28 days (± 3 days) after the last dose of IMP: the following examinations and procedures were made: physical examination, vital signs (blood pressure, respiration rate, pulse, temperature, and weight), assessment of ECOG status, laboratory safety tests (urinalysis was not made), assessment of adverse events, and assessment of changes in concomitant medications

Premature study termination: the study therapy was discontinued if Degree 3 AE per toxicity criteria CTCAE 4.0 occurred which, on the investigators opinion, was treatment-related or symptoms of dose-limiting toxicity developed (see Description of General Study Design and Plan). The study therapy was discontinued and the patient had all of the visit procedures of Week 8. The patient was fo!!owed-up with for 4 weeks after treatment discontinuation. All study withdrawals were fully documented in source documents and CRF. If Degree 3 AEs per CTCAE 4.0 toxicity criteria occurred that were not treatment-related, on the investigator's opinion, the study drug was not discontinued If it was deemed unnecessary. Any patient was withdrawn from the study If Degree 4 AE per CTCAE 4.0 criteria occurred, regardless of the relationship between the event and the study product. Any patient was also withdrawn from the study if: AE was Inconsistent with planned examinations and procedures, the disease progressed, concomitant disease or serious aggravation of the disease was inconsistent with the study participation, the protocol deviated, (Including treatment non-compliance), and for administrative reasons. Patients ^'were withdrawn from the study at any moment if the Investigator considered that it was in the patient's best interest. If any patient did not come to the clinic on the next visit, the patient was contacted by phone to ensure that it was not due to AE, if the study therapy was prematurely discontinued, the principal reason of such discontinuation was recorded in source documents and the appropriate section of CRF. Aii best efforts were made to complete and obtain ail test results as fully as possible.

Pharmacokinetic assessment: aii patients had a pharmacokinetic assessment. Aii patients enrolled to the protocol gave consent for participation in the study and blood samples were taken in time points specified in the study protocol. 2 patients - 0101 and 0307 - gave consent for liver biopsy, 2 tissue samples - native and tumor - ^'were taken to determine product concentration after 4 ^'weeks of quinacrine therapy. Patients had food restrictions, namely: to abstain from grapefruit juice, ginger, coffee, cocoa, tea, and chocolate one day prior to the visit of the clinic and on the day of the pharmacokinetic assessment.

Procedures and visits in extra cycles of study therapy: the number of extra treatment cycles was determined by the Principal Investigator and depended on stable disease values that were assessed every 8 weeks. The decision to continue the treatment was made by the Principal Investigator. Within one cycle, any patient came to the site every 4 weeks to receive: an extra cycie of therapy 1 (Week 12 and Week 16), an extra cycle of therapy 2 (Week 20 and Week 24), and an extra cycle of therapy N (Week n and Week n+4), where n corresponded to Week 4 from the onset of the relevant N cycle. Week 12, 20. 28, n of extra treatment cycles: the following examinations and procedures were made: physical examination, vital signs (biood pressure, respiration rate, pulse, temperature, and weight), assessment of ECOG status, laboratory safety tests (including hematology, biood biochemistry (including alanine transaminase (ALT), alkaline phosphatase, bilirubin (total), creatinine, glucose, and total protein), and urinalysis), assessment of adverse events, assessment of changes in concomitant medications, and dispensing of the patient diary.

Week 16, 24, 32 (n+4) of extra treatment cycles: The following examinations and procedures were made: physical examination, vital signs (blood pressure, respiration rate, pulse, temperature, and weight), assessment of ECOG status, CT-scart of the liver, laboratory safety tests (including hematology and blood biochemistry (Including alanine transaminase (ALT), alkaline phosphatase, bilirubin (total), creatinine, glucose, and total protein)), urinalysis, tumor markers, assessment of adverse events, assessment of changes in concomitant medications, assessment of the RECIST 1.1 criterion and adoption of the decision on additional cycle of the treatment, and dispensing of the patient diary. Patients that continued the study were included to the safety analysis in accordance with safety criteria and efficacy analysis in accordance with efficacy criteria of the study protocol, with regards to extra cycles of the study therapy. The study therapy with quinacrlne was performed until signs of tumor progression or dose-limiting progression occurred, after which the patient was withdrawn from the study in accordance with the exclusion criteria in the study protocol or patient's decision to discontinue the study. After the last dose of the study product, follow-up visits were made in accordance with the Protocol (see Follow- up period: 14 days (+/- 3 days) after the last dose of IMP^■■ telephone call and Follow-up period: 28 days (+/- 3 days after the last dose of IMP).

Selection of Study Population: 36 patients were screened to the study, and 32 of them were enrolled to 7 dose cohorts with gradual dose escalation of the study product. At most, 6 patients were enrolled to each dose cohort (see Description of General Study Design and Plan). In cohort a7 (the last cohort with maximum possible dose approved in the study), 2 cases of dose-limiting toxicity ^'were reported due to what next dose escalation was not required. The sponsor company made a decision to consider the dose as maximum tolerated dose (MTD) in cohort N°5 (400 mg/day) and to stop next patient enrollment. The Drug Safety Monitoring Board was timely informed about the sponsors decision and did not object that patient enrolment was stopped since the primary study objective was achieved.

inclusion criteria: 1. Patients with liver metastases of solid tumors of epithelial origin or hepatocellular carcinoma for which standard therapy had failed or did not exist; 2, ECOG PS 0 - 1 and life expectancy > 12 weeks; 3. Men and ^'women in the age >18 and < 70 years; 4. Adequate organ function as defined by the following criteria: absolute neutrophil count (ANC) > 1.500 cl/pL, platelet count > 100.000 ci/pL, hemoglobin > 9.0 g/di,_serum creatinine < 1.5 x ULN, total serum bilirubin < 2.0 x ULN, and ALT and AST < 5.0 x ULN; 5. Patients with hepatocellular carcinoma Cniid-Pugh class A; 6. Patients agreed to use effective forms of contraception in the study; 7. Patients were fully recovered from any prior surgery, radiotherapy, local, or systemic therapy, except for therapy-induced alopecia or anemia if inclusion criterion 4 was met; and 8. Patients provided a written informed consent form for participation in the study. Exclusion criteria: Palienls wilh acute infection within 3 days of the first dose of quinacrine; patients with other malignancies within the preceding 5 years except for non-melanoma skin cancers and cervical intra-epitheiia! neoplasia; known hypersensitivity to quinacrine; patients with untreated CNS metastases; patients with treated metastases not taking corticosteroids at the study and neurologicaiiy stable for at least 2 months participated in the trial; for women of childbearing potential - pregnant, lactating or refusing to use an effective form of contraception in the study; NYHA class 3 or 4, myocardial infarction or acute coronary syndrome within the preceding 6 months, diabetes mellitus or COPD that required hospitalization within the preceding 6 months; patients with QTcF of >470 msec (females) or >450 msec (males) on EGG; Patients that were HIV positive.; acute relapsing chronic B or C hepatitis; Patients that had organ transplants; patients that had psoriasis or porphyria; history of psychosis and other neuroses; History of deficiency of giucose-6-phosphaie dehydrogenase; any other condition, including alcohol and drug abuse thai, in the opinion of the investigator, prevented patient's participation in the study or increased risks in the study; and participation in another clinical study at least 1 month prior to the first dose of quinacrine.

Study Patients: Patient Allocation in general, 38 patients were screened to the study (there were 4 screening failures) and 32 took at least one dose of the study product. The main reasons for screening failures were: high ALT level (0201), age > 70 years (0202), recent participation In another clinical study (0302), and high bilirubin level (0801). 13 of 32 patients (41 %) completed all visits in accordance with the study protocol, had 8 weeks quinacrine therapy, and complied between 80 to 120% within all 8 weeks of treatment. 19 patients (59%) were ^•withdrawn from the study: 11 patients - due to disease progression, 7 patients - due to adverse events, 1 patient - due to refusal to follow the protocol requirements. All study subjects (32 patients) were Included to safety and pharmacokinetic assessments, while all patients that completed the study (13 patients) were Included to the efficacy analysis. The age of the patients that completed all screening procedures varied from 25 to 71 years, ^•while the mean age was 56 years. All patients ^'were European. The majority of patients (23 patients - 72%) were female. Summary tables of the patients enrolled to the study are provided below.

Table 3. Summary data of 32 patients enrolled to study Q102LT/R1 and taking quinaerine therapy

* PD - disease progression; SD- stable disease; PR- partial response

Exclusion of patients with missing data from ITT-popuiation 13 patients completed the study therapy, there were 4 screening failures: 0201 and 0202 - based on Inclusion criterion s3: "Men and women in the age >18 and < 60 years" (in accordance with the Study Protocol, version Ns 1 ). Patient 0303 did not comply with exclusion criterion N°15: 'Participation in another clinical study prior 1 month". Patient 0801 did not comply with inclusion criterion Ns4: "Total bilirubin < 2.0 x ULN". 19 patients were prematurely withdrawn from the study (11 patients - due to disease progression, 7 patients - due to adverse events, and 1 patient - due to patient's refusal to follow the protocol requirements).

The study was set up to determine, inter alia, the maximum tolerated dose (MTD) (including in liver tumors of different etiology). Also, the safety profile of quinacrine, determination of the pharmacokinetic parameters of quinacrine, and determination of the possible efficacy of quinacrine therapy in patients with liver tumors of different etiology based on the RECIST 1.1 criteria was undertaken.

EXAMPLE 2; STATISTICAL METHODS SPECIFIED BY THE PROTOCOL AND DETERMINATION OF SAMPLE SIZE

Statistical and analytical plans With regards to illustrative study goals (e.g. determination of dose-limiting toxicity, assessment of safety, to!erability, and pharmacokinetics), the main analysis instruments were descriptive statistics and graphical representations. Numeric data were summarized and described with the following parameters: number of observations, number of missing observations, minimum, maximum, mean arithmetic, standard deviation, median, mode, 25 percentile, and 75 percentile. To assess qualitative data, the number of cases and percentages were provided. The last measurements/examinations made prior to the first dose of the study product (visit on Day 1 ), were considered as baseline data. Changes from the baseline were calculated as the difference of results of measurements/examinations on visit X from the results of measurements/examinations on Visit on Day 1. The statistical methodology was detailed in the Statistical Analysis Plan. The statistical analysis was made In the SAS system (version 9.3, SAS Institute Inc., Gary, NC, USA).

Determination of sample size 36 patients were screened in the study , 32 of them were enrolled to the study , and 13 patients completed the study taking 8-week study therapy. Safety was assessed in all study subjects, while possible efficacy was assessed In patients that completed the study (see below).

Analysis Data Sets Two populations were analyzed In the study: intention-to-treat population of patients (ITT): Intention-to treat-population of patients (ITT) consisted of all patients enrolled to the study that took at least one dose of the study product. Safety population: Safety population consisted of all patients enrolled to the study that took at least one dose of the study product. The study ITT-population and safety population coincided.

EXAMPLE 3; TREATMENT

Quinacrine (capsules 50 mg) was taken per os. The dosing regimen depended on the dose that was prescribed to the patient depending on the study cohort.

Cohort 1

Day 1-7 of loading dose - 100 mg (2 capsules) three times a day.

Day 8-56 of maintenance dose - 100 mg (2 capsules) once daily in the morning.

Cohort 2

Day 1-7 of loading dose - 100 mg (2 capsules) three times a day.

Day 8-56 of maintenance dose - 100 mg (2 capsules) in the morning and 100 mg (2 capsules) in 4 hours.

If DLT occurred in the first cohort, the dose for the second cohort was reduced in 2 times: loading dose 150 mg a day and maintenance dose 50 mg a day.

Cohort 3

Day 1-7 of loading dose - 150 mg (3 capsules) in the morning, 100 mg (2 capsules) In the afternoon, and 100 mg (2 capsules) in the evening.

Day 8-56 of maintenance dose - 150 mg (3 capsules) in the morning and 100 mg (2 capsules) in 4 hours.

Cohort 4

Day 1-7 of loading dose - 150 mg (3 capsules) in the morning, 150 mg (3 capsules) in the afternoon, and 100 mg (2 capsules) in the evening.

Day 8-56 of maintenance dose - 150 mg (3 capsules) in the morning and 150 mg (3 capsules) in 4 hours.

Cohort 5

Day 8-56 of maintenance dose - 150 mg (3 capsules) in the morning, 150 mg (3 capsules) In the afternoon, and 100 mg (2 capsules) in the evening.

Cohort 6

Day 1-7 of loading dose - 200 mg (4 capsules) in the morning, 200 mg (4 capsules) in the afternoon, and 100 mg (2 capsules) in the evening.

Day 8-56 of maintenance dose - 200 mg (4 capsules) in the morning, 150 mg (3 capsules) in the afternoon, and 100 mg (2 capsules) in the evening.

Cohort 7

Day 1-7 of loading dose - 250 mg (5 capsules) in the morning, 250 mg (5 capsules in the afternoon, and 100 mg (2 capsules) in the evening.

Day 8-56 of maintenance dose - 200 mg (4 capsules) in the morning, 200 mg (4 capsules) in the afternoon, and 100 mg (2 capsules) in the evening.

Previous and concomitant therapy All drug products except for those prohibited by the protocol which were considered necessary for patient's well-being and did not Interfere with the study product were prescribed at the Investigator's discretion. The Investigator recorded all drug products taken by patients in the study from the date of informed consent to the appropriate section of CRF. As it was a partially outpatient study, emphasis was made on patient surveys on any products which they took on their own. Patients were informed that they may take drug products for management of concomitant diseases. Dose and dosing frequency were constant throughout the study period. During the study, patients were not allowed to use any of the following products In combination with the study drug: gold-based drugs (auranofin, aurothiogiucose, sodium aurothiomaiate, and solganal), tranquillizers and antipsychotic agents (thioridazine, pimozide, and melieril), and other drug products that are metabolized in the human body with cytochrome P450 (atomoxetlne, codeine, enabiex, fesoterodine, timolol, aricept, and tolterodine). During the treatment, patients abstained from aicohoi, as exposed by quinacrine, aldehyde dehydrogenase ^'was inhibited, and alcohol degradation was suppressed on the stage of acetaidehyde formation which resulted in intoxication symptoms (shiver, asphyxia, palpitations, fear, face erythema, and increased temperature).

Compliance Study subjects were instructed to bring back their bottles with the study product empty and or partially filled) for the entire treatment period. All bottles were collected. The results of tablet accountability were recorded in CRF. The compliance of study therapy was determined with calculation of the remaining tablets. The compliance was assessed in the range of 80 to 120%. 4 out of 32 study subjects did not comply with the range.

EXAMPLE 4: EFFICACY AND SAFETY PARAMETERS (DETERMINATION OF EFFICACY AND SAFETY PARAMETERS AND SCHE E)

Measurements of efficacy parameters Possible efficacy was assessed after 8 weeks of quinacrlne therapy in accordance with the RECIST1.1 criteria.

Measurements of safety parameters. Ail patients that took at least one dose of quinacrine were included in the safety analysis. The safety of patients was constantly assessed in the study by the following parameters: Adverse events and serious adverse events (AE and SAE): all information on adverse events was collected after obtaining informed consent. Their frequency., severity, and relationship with the study product ^'were detailed and analyzed in the Efficacy Analysis section, as well as compared between treatment groups (dose cohorts). The frequency and incidence of individual AEs was also reported.

Changes in the routine laboratory tests ^'were assessed (predominantly, hematology and clinical chemistry parameters), Moreover, laboratory results were tabulated in comparison with normal laboratory values and the tables and distribution plots were provided in the study statistical report. Clinical changes of laboratory values were assessed on their relationship with adverse events or, if asymptomatic, determined as independent AEs. Such changes were detailed and analyzed as described herein.

Table 5. Laboratory safety parameters.

Clinical biochemistry ~ Alanine transaminase (ALT); Aspargine transaminase (AST); Alkaline

phosphatase (ALP); Albumin; Bilirubin (total); BUN Hydrocarbonate/C02 Creatinine; Glucose; Sodium; Potassium; Chlorides; Calcium; Magnesium; Total protein; Non-organic phosphorus.

Abbreviated clinical biochemistry Alanine transaminase (ALT); Aspargine transaminase (AST); Alkaline phosphatase; Albumin; Bilirubin (total); Creatinine; Glucose; Total protein;

Hematology Hemoglobin; Hematocrit; RBC count; Platelet count; WBG count and differential (neurophils, eosinophils, basophiles, lymphocytes, monocytes)

Urinalysis Color; Clarity; Specific gravity; Hemoglobin measurement with dipstick; Glucose measurement with dipstick; Ketone measurement with dipstick; Bilirubin measurement with dipstick Microscopic

examination ^'was made if dipstick determinations ^'were ++ and higher.

Biood coagulation test Protrombin time;

Pregnancy test Dipstick pregnancy test

Assay of tumor markers in blood serum Aipha-fetoprotein (for HCC); CEA and Ca-19.9 (for stomach, colon cancer, cholangiocarcinoma, and pancreatic cancer); Other tumor markers specific for the type of tumor

Reporting of vital signs, ital signs included: measurements of BP, HR, RR, temperature, weight, and height (only on screening).

ECG assessment ECG data were assessed on screening, visit on Day 8, Day 15, Day 22, Day 29, and Day 43.

Adverse events (AE) The working definition of adverse event was: any untoward medical occurrence in a patient or a clinical study subject administered a pharmaceutical product that does not necessarily have a causal relationship with this treatment. An AE, therefore, was any unfavorable and unintended sign (e.g., a clinically abnormal laboratory finding), symptom, or disease temporally associated with the use of the study drug, whether or not considered related to the study drug. All study subjects had at least one adverse reaction. 27 of 31 patients reported adverse reactions considered by the investigator as treatment-related: dermatitis (29,0%), nausea (29.0%), weakness (19.4%), anorexia (22.6%), lymphopenia (12.9%), weight loss (19.4%), hyperglycemia (19,4%), decreased hemoglobin (16.1 %), and dizziness (12.9%). Most adverse reactions had toxicity of Degree 1-2. Adverse events of Degree 1 were reported in 12 patients (38,7%), Degree 2 in 13 patients (41.9%), and Degree 3 in 6 patients (19.4%), There were no adverse events of Degree 4, All adverse reactions reported in the study, ^'without wishing to be bound by theory, have previously been listed for quinacrine. The results of the study showed that the product quinacrine had an acceptable safety profile in patients with androgen-independent metastatic prostate cancer in dose 300 mg/day (100 mg 3 times a day) for the first week, ^•with subsequent switch to maintenance dose of 100 mg/day. Based on the assessment of adverse reactions, laboratory tests, EGG, vital signs, and results of physical examination, there were no adverse reactions of high toxicity.

Serious adverse events (SAE) Any untoward medical occurrence meeting definition of AE which at any dose: resulted in death, was life-threatening, resulted in persistent or significant disability or incapacity, required hospitalization or prolongation of existing hospitalization, and was a congenita! anomaly or birth defect in a child or fetus of patient faking the study product prior to conception or during pregnancy. Medical and scientific judgment was exercised in deciding whether other conditions were also considered serious, such as important medical events that were not immediately life threatening (did not result in death or hospitalization) but jeopardized the subject's safety or required intervention to prevent one of the other outcomes listed in the definition above. These ^'were also considered serious. Definition of "life threatening" - An AE was life threatening if the subject was at immediate risk of death from the event as it occurred; i.e. It does not include a reaction that if it had occurred in a more serious form it might have caused death. Death: despite the fact, considering the study population, death was an expected event, and it was repotted as any SAE. Hospitalization: any hospitalization was reported as SAE, except for: therapeutic or surgical procedures scheduled per protocol, routine health examinations that required hospitalization to assess baseline status/trends (for example, for instrumental test methods), admission for therapeutic/surgical indications other than treatment of the pathological condition (scheduled prior to the study; relevant documents were required), or admission due to other life circumstances that are not related with health conditions and did not require therapy/surgical intervention (for example, homelessness, inadequate economic status, interruption of care, family circumstances, and administrative reasons). Identification, registration, and reporting of adverse events and concurrent diseases. During the study, study doctors foiiowed-up adverse events. All adverse events reported from the data of signed Informed consent were detailed In source documents and transferred to CRF regardless of the Investigator's judgment on relationship with the study product. To assess maximum intensity of each adverse event, the Investigators used the following 3-scale score: Mild: an AE that was easily tolerated by the subject not interfering with everyday activities, Moderate: an AE that was sufficiently discomforting and interfered ^'with normal everyday activities, or Severe: an AE that was incapacitating and prevented normal ever/day activities.

The Investigator assessed the relationship between the event and study product as: Unrelated: Clearly and definitely associated with external factors and did not meet criteria of unlikely or probable relationship, or Related: definite tempera! relationship to the study product (associated with a clinical condition or external factors, or other indicated treatment methods i.e. there was a clear temporary relationship between therapy discontinuation or dose decrease or Improvement (it re-occurred with rechallenge and It had a known pattern of response to the study drug)).

Seven serious adverse events were reported in the study, see detailed information in the Pharmacokinetics section.

Efficacy was assessed after 8 weeks of qu!nacrine therapy in accordance with the RECIST1.1 criteria. On the visit of Day 57, the status of the present target and non-target lesions, as well as overall treatment effect was assessed.

Pharmacokinetic parameters and pharmacokinetic assessment. PK was assessed by serum concentrations of quinacrine in serum of every subject taking the study product. The data is provided below. The comprehensive pharmacokinetic assessment of individual serum concentrations of quinacrine in study subjects is provided in the individual pharmacokinetic report of the study. Vials with serum samples for pharmacokinetic assessment were kept in the study site at -200°C. Once accumulated, they were sent in a thermal container in dry ice for testing in the laboratory of Institution of the Russian Academy of Medical Sciences "Mental Health Research Center RAMS", Russia. Along with serum samples that were used to determine serum quinacrine concentrations, biopsy materials of 2 patients - 0101 and 0307 - were provided to the laboratory to assess product distribution in liver tissues after 4 weeks of the study therapy, in each case, biopsy material was taken from liver tumor tissue and intact region of liver.

EXAMPLE 5: EFFICACY ANALYSIS

Summary information with the total number of patients, patients prematurely withdrawn from the study (due to AE and progression) in absolute figures, % of patients thai completed 8 weeks of treatment in absolute figures, % effect assessment (overall and target lesions) in absolute figures, and % in individual cohorts and in iota! is provided in table 6.

Summary information with total number of patients, patients prematurely withdrawn from the study (due to AE and progression) in absolute figures, % of patients which completed 8 weeks of treatment in absolute figures, % effect assessment (overall and target lesions) in absolute figures, and % in groups per diagnosis (% of total number of patients in the group ^'with one diagnosis) is provided in table 7. The efficacy analysis was made after 8 weeks of quinacrine therapy in accordance with the RECIST1.1 criteria. The graphic presentation of % changes of maximum diameter target lesion after 8 weeks of quinacrine therapy in comparison with baseline data is provided in Figure 1. Table 6 is a summary table with total number of patients, patients prematurely withdrawn (based on AE and progression) In absolute figures, % completed 8 weeks of treatment in absolute figures, % assessment of effect (total and per target lesions) in absolute figures, and % in individual cohorts and overall.

Table 6

Cohort Cohort Cohort Cohort Cohort Cohort Cohort Total B1 mi N93 Ns6 mi

abs %¹ abs abs %¹ abs %¹ abs ¹ abs %¹ abs ¹ abs ²

Number of patients

6 18.7 4 12.5 6 18.7 3 9.4 7 21.9 3 9.4 9.4 32 100

Prematurely

withdrawn for the

reason:

- AE 2 33.3 0 0 16.7 1 33.3 1 14.3 0 0 2 66.7 7 21.9

- Progression 2 33.3 2 50 33.3 0 0 3 42.9 1 33.3 1 33.3 11 34.4

- Patient's refusal 0 0 1 25 0 0 0 0 0 0 0 0 0 0 1 3.1

Completed 8

2 33 1 25 3 50 2 67 3 43 2 67 0 0 13 40.6 weeks of treatment

Overall response:

- CR 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0

- PR 0 0 0 0 0 0 0 0 1 33.3 0 0 0 0 1 7.7

- SD 1 50 o 0 1 33.3 0 o 0 0 2 100 0 0 4 30.8

- PD 1 50 1 100 2 66.7 2 100 2 66.7 0 0 0 0 8 61.5

- NE 0 0 0 0 0 0 0 0 0 0 o 0 o 0 0 0

Status of current

target lesions:

- CR 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0

- PR 0 0 0 0 0 0 0 0 1 33.3 0 0 0 0 1 7.7 - SD 2 100 0 0 2 66,7 0 0 0 0 2 100 0 0 6 46.2

- PD 0 0 1 100 1 33.3 2 100 2 66.7 0 0 0 0 6 46.2

- NE 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0

- NA 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0

^* CR = Complete Response NA = Not Applicable PR = Partial Response PD = Progressive Disease

SD = Stable Disease NE = Inevaiuabie

% calculated from the total number of patients in the cohort; % calculated from the total number of patients (32 patients - for calcuiation of patients withdrawn and competed the study therapy, 13 patients - for calculation of overall response of patients and status of target lesions)

Table 7 Summary table with total number of patients, patients prematurely withdrawn (based on AE and progression) in absolute figures. % completed 8 weeks of treatment in absolute figures. % assessment of effect (total and per target lesions) in absolute figures, and % in groups by diagnoses (% of total number of atients in the group with one diagnosis)

CR = Complete Response; NA = Not Applicable; PR = Partial Response; PD = Progressive Disease; SD =

Stable Disease; HE - Inevaluable

% calculated from the total number of patients in the cohort; % calculated from the total number of patients (32 patients - for calculation of patients withdrawn and competed the study therapy, 13 patients - for calculation of overall response of patients and status of target lesions)

Status of target and non-target lesions after 8 weeks of study therapy After 8 weeks of quinacrine therapy 6 (46%) patients had target lesion status assessed as disease progression, 6 patients (46%) - stable disease and 1 patient (8%) - partial response. Stable disease per target lesion status was shown in 2 patients in cohort 1 (100%), 2 patients in cohort 3 (67%), and 2 patients in cohort 6 (100%).

Non-target lesion after 8 weeks of study therapy was assessed as disease progression in 5 patients (39%), partial response/stable disease - 4 patients (31 %), and not applicable - 4 patients (31 %). Of the 6 patients that target lesion status was assessed as stable disease, 3 patients had non-target lesion status assessed as incomplete response/stable disease (1 patient in cohort 3 and 2 patients in cohort 6). 1 patient had non-target lesion status as not applicable, and the remaining 2 patients had non-target lesion status as disease progression. New lesions were observed in 7 patients (54%), two of which had target lesion status as stable disease.

Overall treatment response after 8 weeks of study therapy. After 8 weeks of study therapy, overall treatment response was assessed in 8 patients (62%) as disease progression: 1 patient (50%) in cohort 1 , 1 patient (100%) in cohort 2, 2 patients (67%) in cohort 3, 2 patients (100%) in cohort 4, and 2 patients (67%) in cohort 5. 4 patients (31 %) had overall treatment response assessed as stable disease, while 1 patient (8%) had overall treatment response assessed as partial response. None of the 4 patients with overall response assessed as stable disease experienced any SAEs. The patient whose overall response was assessed as partial response had 1 SAE related to the disease (hydrothorax).

Post-hoc analysis. The mean percentage change in sum of maximum diameters of target lesion at Week 8 in comparison with the results on Screening visit (baseline data) was an Increase of 21 % (range: -46 to 98%). Two patients had a decrease in sum of maximum diameters from baseline to Week 8: patient 0306 in cohort 5 and patient 0117 in cohort 6. Patient 0306 had target lesion status as partial response, non-target lesion status as partial response/stable disease and overall response as partial response. Patient 0117 had target lesion status as stable disease, non-target lesion status as partial response/stable disease and overall response as stable disease. The maximum serum concentrations of the study product in both patients ^'were observed at Week 6: 219 ng/ml for patient 0308 and 196 ng/mi for patient 0117.

Analysis of Response Heterogeneity in Patients Taking Study Therapy the data were provided on clinical response In 13 patients that completed quinacrine therapy in the 1 b phase clinical study. The study design aimed to find maximum tolerated dose of the product and did not suggest the presence of a control group of patients taking placebo. The assessment of possible efficacy of quinacrine therapy based on the RECIST 1.1 criteria allowed identification of the difference In response to the 8-week therapy in patients ^'with one of the diagnoses in comparison with other patients of the sample, as the number of measurements remained acceptable for statistical analysis.

As tested parameters, the following were used: percentage change in diameter of target lesion in comparison to the screening diameter prior to the study {% target lesions change) and target lesion response to the study therapy (target lesions response) and overall response In scores: 0 - disease progression (PD), 1 - stable disease (SD), 2 - partial response (PR), and 3 - complete response.

Table 8. Baseline test data - response to study therapy in 13 patients of 1b Phase clinical study,

Target treatment

Patient Ms Diagnosis (brief) % Target change Overall response effect

01-01 Hepatocellular carcinoma 13,3 SD SD

01-04 Hepatocellular carcinoma 2.4 SD PD

01-06 Stomach cancer 44.7 PD PD

01-09 Colon cancer 15.4 SD PD

01-1 1 Colon cancer 36,5 PD PD

03-05 Breast cancer 16.8 SD SD

06-02 Colon cancer 95.5 PD PD

06-03 Colon cancer 26.1 PD PD

01-13 Small Intestine cancer 30,4 PD PD

03-06 Breast cancer -46.3 PR PR 06-04 Colon cancer 35.7 PD PD

Salivary gland cancer

01-16 8.4 SD SD

(head/neck)

01-17 Colon cancer -1.9 SD SD

Table 9. Comparison of response to study therapy in individual groups of patients and remaining sample based on Student's T-test.

Table 10. Comparison of response to study therapy in combined group of patients and remaining sample based on Student's T-test.

Patients with gastrointestinal cancer {HI = 8 patients) in comparison with other patients {U2 = 5 patients)

Mean 1 Mean 2 t-value P N1 H2

% Target change 35.3 -1.08 2.32655 0.040112 8 5 Target treatment effect 0.25 \ .Δ -3.64430 0.003859 8 5

Overall response 0.13 1.0 -3.00223 0.012032 8 5 i he differences show that patients with Gi cancer had disease progression, and patients with hepatic carcinoma, breast cancer, and salivary gland carcinoma had stable disease during the study.

The data on the structure of patients enrolled to the study that completed the treatment, with regards to the diagnosis and results of response assessment on treatment, are provided in the tables below (Tables 11-15):

Table 11, Structure of patients enrolled to the study with various tumor types

% caiculated from the total number of patients in the cohort, 2 - % calculated from the totai number of patients (32 patients)

Table 12, Structure of patients with various tumor types that completed treatment course

% calculated from the total number of patients in the cohort, 2 -% calculated from the total number of patients (13 patients)

Table 13. Analysis of total response in patients with various tumor types that completed 8 weeks of treatment in accordance with the RECIST criteria

Table 14. Assessment of target lesion response in patients with various tumor types that completed 8 weeks of treatment in accordance with the RECIST criteria

Table 15. Mean response yalues of target lesions in groups with various tumor types

Assessment of Treatment Compliance the estimated treatment compliance was in the range of 80-120% for a!! patients enrolled to the study except for 5 events reported in 4 patients: 1 patient on premature study withdrawal/termination visit in cohort 3, 2 patients on premature study withdrawal/termination visit in cohort 5, and 1 patient on premature study withdrawal/termination visit at Week 3.

Pharmacokinetic Assessment Materials and methods Blood samples were taken from each patient to determine quinacrine concentration prior to drug administration (day 1) and on 9 time points (total - 10 time points):day 1 - 1 hour prior to administration of the study drug, day 1 - 30 min after administration of the study drug, day 1 - 1 hour after administration of the study drug, day 1 - 2 hours after administration of the study drug, day 1 - 4 hours after administration of the study drug, day 1 - 8 hours after administration of the study drug, day 2 - 24 hours after administration of the study drug, day 15 - prior to administration of the study drug, day 29 - prior to administration of the study drug, and day 43 - prior to administration of the study drug. Biopsy samples of 2 patients (0101 and 0307} were taken after 4 weeks of quinacrine therapy from two regions (tumor tissue and normal tissue) in order to determine quinacrine concentration. The assay of quinacrine in serum samples was conducted with mass-spectrometry as part of analysis of the biological samples obtained from 32 patients with liver tumors of different etiology that took Quinacrine In study Q102LT/R1. Detection was made with triple quadrupole mass-spectrometer Agilent 6410. Homogenized samples of liver tissue were analyzed with the method of high-efficiency liquid chromatography - tandem mass-spectrometry.

The results showed that steady-state plasma concentration of quinacrine was achieved already on day 15 after the dosing, without wishing to be bound by theory, which may be associated with high loading doses on the first day of the therapy. Table 16. Steady-state corfcentratiors of quinacrine after multiple-day study therapy in various cohorts (ng/ml).

The linearity of product pharmacokinetics was established in the range of tested doses which was shown as invariance of concentration/dose ratio. The value was 0.44±0.08 (ng/n g)x10 ^s and did not differ between study cohorts (Cohort Ne7 was not considered due to too few measurements).

Table 17. Pharmacokinetic parameters of quinacrine after multiple day study therapy in various cohorts

Quinacrine was characterized by intensive elimination; total clearance values averaged to 23 ml/{min*kg) which, without wishing to be bound by theory, was comparable to renal blood flow in humans - 21 mi/(min*kg).

The results of liver biopsy in two patients are provided in the tables below:

Table 18. Patient 0101 , study therapy - 4 weeks

Mass of tissue Plasma concentration,

Sample sample, mg Extract volume, mi Result, pg/g ng/ml

10^* 2 0.5 55

10.4

1 H^** 5 0.5 69 Table 19, Patient 0307, study therapy - 4 weeks

* - tumor sample ** - normal tissue sample

Without wishing to be bound by theory, the following conclusions were made: there was a high variability of individual drug concentrations in the blood (variation coefficient of concentration values was >100%), a steady- state concentration of quinacrine was achieved already on day 15 day after the dosing, the linearity of product pharmacokinetics was established In the range of tested maintenance doses (100-450 mg/day), the product clearance of quinacrine (characterized by intensive elimination) was comparable to such physiological value as renal blood flow rate, and the hepatic product concentration was higher than blood concentration in many times.

Of the 32 patients taking quinacrine, 19 patients prematurely discontinued the study: 11 patients due to disease progression, 7 patients due to adverse events (4 patients - DLT, 3 patients - other adverse events that were associated with disease progression and were not treatment-related), and 1 patient due to refusal to follow the protocol requirements.

Anticancer efficacy of 8 weeks of treatment with quinacrine was assessed in 13 patients. One patient with breast cancer (8%) had partial response, four patients (31 %) - stable disease (1 patient each of breast cancer, hepatocellular carcinoma, salivary gland cancer, and rectal cancer), and eight patients (62%) had disease progression (5 patients with colon cancer and 1 patient each with stomach cancer, hepatocellular carcinoma, and small intestine cancer).

Target treatment response that was predominantly localized in liver, was more marked: partial response - 1 patient (8%), stable disease - 6 patients (46%), and disease progression - 6 patients (46%).

The period of ciinica! effect was not assessed; however, one patient with hepatocellular carcinoma had stable disease for long period with quinacrine therapy (7.5 months).

Despite high product concentration in the liver, quinacrine had an anticancer effect on liver metastases of solid tumors and hepatocellular carcinoma. The exception was patients with liver metastases of breast cancer that had a clinical effect of quinacrine therapy. Both patients had multiple previous courses of chemotherapy and hormonal therapy, and had liver metastases.

EXAMPLE 8: SAFETY ANALYSIS

Extent of Exposure The study product was provided to all sites prior to the study initiation by the sponsor company as bottles of quinacrine with 50 mg each (60 capsules). 32 patients took at least 1 dose of the study product, 13 of them completed quinacrine therapy (8 weeks of treatment with quinacrine). Adverse Events reporting began at the start of screening procedures (the screening period lasted up to 4 weeks). Despite the fact that the patients did not take study therapy in the period, several procedures on patient selection ^'were made. Safety population included data of aii patients that took at least 1 dose of the study product.

Each of the 32 patients had at least 1 AE. A total of 246 AEs were reported In the study, 135 of them reported in 29 patients (91 %) were assessed as treatment-related. In general, the proportion of patients that reported treatment-related AEs was comparable between groups of patients.

During the study, 7 patients (22%) reported 7 serious adverse events (SAE) not related with the study product: Cohort 1 : 4 patients (67%): 0102, 0104, 0303 (disease progression), and 0103 (jaundice cholestatic), Cohort 3: 1 patient (17%): 0108 (cholestasis), and Cohort 5: 2 patients (29%): 0306 (hydrothorax) and 0307 (ascitis).

2 patients had AEs assessed as life-threatening: patient 0102 (Cohort 1) ^■■ asthenia, patient 0108 (Cohort 2) - cholestasis. The adverse events ^'were not treatment-related; subsequently, the patients progressed.

7 patients (22%) had AEs resulting in premature study withdrawal, 4 of them ^'were assessed as DLTs, and the other 3 AEs were reported in patients with disease progression (non-treatment related): Cohort 1 : 2 patients (33%): 0103 (increased bilirubin) and 0303 (paraneoplastic syndrome, iliofemoral deep vein thrombosis of lower extremities), Cohort 3: 1 patient (17%): 0304 (abdominal pain upper - DLT), Cohort 4: 1 patient (33%): 0601 (acute renal dysfunction), Cohort 5: 1 patient (14%): 0112 (diarrhea - DLT), and Cohort 7: 2 patients (67%): 0120 and 0308 (ALT increased - DLT). 3 patients (9%) had AEs resulting In death: Cohort 1 : 2 patients (33%): 0102 and 0104 (disease progression), and Cohort 3: 1 patient (17%): 0108 (cholestasis).

Summary information of all AEs that were reported in the reporting period is provided in Tables 20 and 21 ,

Table 20. Types of AEs reported in the study in various dose cohorts of patients reviewed for safety assessment.

1- % calculated from the total number of patients in the cohort

2- % calculated from the iota! number of patients (32 patients)

Table 21. Severity of AEs reported in the study in various dose cohorts of patients reviewed for safety assessment.

% calculated from the total number of patients In the cohort % calculated from the total number of patients (32 patients)

Assessment of adverse events The majority of reported AEs were of mild or moderate severity - 206 of 246 AEs. 18 patients (56%) had at !east 1 severe AE (4 patients (67%) in cohort 1 , 1 patient (25%) in cohort 2, 2 patients (33%) in cohort 3, 2 patients (67%) in cohort 4, 5 patients (71 %) in cohort 5, patient (33%) in cohort 6, and 3 patients (100%) in cohort 7).

The most common severe AEs were asthenia (5 of 18 patients), abdominal pain upper (4 of 18 patients), ALT increased (4 of 18 patients), AST increased (3 of 18 patients), increased bilirubin (3 of 18 patients), and disease progression (2 of 18 patients). A!! severe AEs were reported as Isolated cases. Ail cohorts had similar incidence of AEs (on average, 6 - 9 AEs per patient).

The most common adverse events were: Yellow skin (26 patients (81 %)): in cohort 1 - 6 patients (100%), in cohort 2 - 3 patients (75%), In cohort 3 - 6 patients (100%), in cohort 4 - 3 patients (100%), in cohort 5 - 5 patients (71 %), in cohort 6 - 1 patient (33%), and in cohort 7 - 2 patients (67%), Asthenia (18 patients (56%)): in cohort 1 - 4 patients (67%), in cohort 2 - 2 patients (50%), in cohort 3 - 2 patients (33%), in cohort 4 - 1 patients (33%), in cohort 5 - 6 patients (86%), in cohort 6 - 1 patient (33%), and in cohort 7 - 2 patients (67%), Abdominal pain upper (12 patients (38%)): in cohort 1 - 4 patients (67%), in cohort 3 - 2 patients (33%), in cohort 4 - 1 patients (33%), in cohort 5 - 3 patients (43%), and In cohort 6 - 2 patients (67%), and Nausea (10 patients (31 %)): in cohort 1 - 2 patients (33%), i cohort 2 - 1 patient (25%), in cohort 3 - 1 patient (17%), in cohort 4 - 1 patient (33%), in cohort 5 - 4 patients (57%), and in cohort 7 - 1 patient (33%).

The most common treatment-related AEs were: Yellow skin (24 patients (75%)), Nausea (9 patients (28%)), Abdominal pain upper (8 patients (25%)), Diarrhea (7 patients (22%)), ALT increased (7 patients (22%)), AST increased (5 patients (16%)), and Asthenia (4 patients (13%)).

Table 22, Summary fable of most frequently reported AEs

Table 23. Summary table of most common treatment-related AEs

% calculated from the iota! number of patients in the cohort 2- % calculated from the total number of patients (32 patients)

Table 24, Table of individual most common AEs

Table 25, Table of irtdividuai most common treatment-related AEs

Grade 2 0 0 0 0 1 16.7 0 0 14 3 0 0 0 0 2 6.3

Grade 3 0 0 0 0 1 16.7 0 0 0 0 0 0 2 66.7 3 9.4

Grade 4 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0

Grade 5 0 o o 0 0 0 0 0 0 0 0 0 o o 0 0

Asthenia Grade 1 0 o o 0 2 33.3 0 0 14.3 0 0 33.3 4 12.5

Grade 2 0 o 0 0 1 16.7 0 0 14.3 0 0 o 0 2 6,3

Grade 3 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0

Grade 4 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0

Grade 5 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0

% calculated from the total number of patients In the cohort 2- % calculated from the total number of patients (32 patients)

Ail other AEs were reported in less than 10% of patients. Of all the laboratory abnormalities, the most common abnormalities were AST increased (10 events) and ALT increased (10 events), in general, 15 patients reported 48 ciinicai laboratory abnormalities. On investigators' opinion, without ^'wishing to be bound by theory, there were no treatment-related SAEs,

Product quinacrine showed a relatively low toxicity. Treatment-related serious adverse events, life threatening events, and deaths were not reported, except for 4 DLTs.

Laboratory Ciinicai Analysis The following main laboratory safety parameters were constantly assessed by the investigators: hematology, clinical chemistry, blood coagulation values, results of genera! urinalysis, and ECG. The tests were conducted in local laboratories of the study sites. The test results were recorded in patient CRFs. Summary information on all clinical laboratory abnormalities recorded in the study Is provided In Table 26.

Table 28. Clinical laboratory abnormalities

In total, 15 patients reported 48 clinical laboratory abnormalities. Physical examination. During the reporting period, each patient reported clinical abnormal results of at least one physical examination. On screening visit, 6 patients had clinical abnormal results of physical examination, while there ^'were no abnormalities on visit at Day 1.

The number of patients with clinical abnormal results of physical examination on each visit was comparable between the visits at Week 1 - Week 6 (11-15 patients (34-47%)). Of the 30 patients who had physical examination on premature study discontinuation/end-of-study visit, clinical abnormalities occurred in 24 patients (75%) (such were absent on screening visit, Day 1).

In conclusion, without wishing to be bound by theory, the clinical studies showed that quinacrlne had a rather low toxicity in the range of dose levels 1-8 (from dosing by the scheme: loading dose - 300 mg/day, maintenance - 100 mg, and inclusively till dosing: loading dose 500 mg/day, maintenance - 450 mg/day). Oral route of administration and its selective hepatic accumulation are associated with gastrointestinal toxicity, which prevailed in dinicai picture. The drug-related gastrointestinal toxicity events were nausea (28% patients), abdominal pain upper in liver projection (25%), diarrhea (22%), ALT increased (22%), and AST increased (16%). Poor tendency of Increased incidence and severity of diarrhea and increased liver enzymes was shown with increased drug dose (Table 23). Other most common treatment-related adverse effects were skin pigmentation (75%) and asthenia (13%). The two latter complications had similar incidence and intensity in all cohorts.

During the study, 7 patients had serious adverse events (SAE) and 2 patients had adverse events (AE) with severity assessed as life threatening. Without wishing to be bound by theory, the adverse events were associated with disease progression and not treatment-related. 7 patients had adverse events leading to premature study withdrawal, 4 of them were treatment-related and assessed as DLTs (abdominal pain upper, diarrhea, and ALT increased In 2 patients), and the 3 other adverse events were related to disease progression.

During treatment, 3 patients died due to disease progression. No deaths were reported which were related to drug toxicity. Without wishing to be bound by theory, quinacrine did not affect cardiac activity, function of bone- marrow hemopoiesis, and other laboratory values. In general, treatment-related adverse events had similar incidence in all cohorts, and no serious and life-threatening adverse events related to treatment were registered, except for 4 DLTs.

EQUIVALENTS

While the invention has been described in connection ^'with specific embodiments thereof, it will be understood that it is capable of further modifications and this application is intended to cover any variations, uses, or adaptations of the invention following, in general, the principles of the invention and including such departures from the present disclosure as come within known or customary practice within the art to which the invention pertains and as may be applied to the essentia! features hereinbefore set forth and as follows in the scope of the appended claims. Those skilled in the ari will recognize, or be able to ascertain, using no more than routine experimentation, numerous equivalents to the specific embodiments described speciiically herein. Such equivalents are Intended to be encompassed in the scope of the following claims. INCORPORATION BY REFERENCE

All patents and publications referenced herein are hereby incorporated by reference in their entireties.

The publications discussed herein are provided solely for their disclosure prior to the filing date of the present application. Nothing herein is to be construed as an admission that the present invention Is not entitled to antedate such publication by virtue of prior Invention.

As used herein, all headings are simply for organization and are not intended to limit the disclosure in any manner. The content of any individual section may be equally applicable to ail sections.

Claims

CLAIMS What is claimed is:

1. Use of quinacrine in the manufacture of a medicament for the treatment of liver cancer that is resistant to standard treatment.

2. Use of quinacrine In the manufacture of a medicament for the treatment of a metastatic cancer that is of hepatic origin or has metastasized to the liver from another organ.

3. The use of claim 1 , wherein the liver cancer is resistant to one or more of chemotherapy, chemoemboiization, radiation therapy, cryosurgery, radiofrequency ablation, or ethanoi Injection.

4. The use of any one of claims 1-3, wherein the liver cancer is unresectable and/or present in a subject that has coexisting medical condition that prevents surgery, selected from cirrhosis and hepatitis.

5. The use of any one of claims 1-4, wherein the liver cancer is present in a liver having cells that overexpress one or more muitidrug-resistance pumps.

6. The use of claim 5, ^'wherein the muitidrug-resistance pump is one or more of the P-giycoprotein and the multidrug resistance-associated protein (MRP).

7. The use of any one of claims 1-6, ^'wherein the treatment comprises one or more of reducing liver tumor load, reducing liver tumor volume, and stabilizing disease,

8. The use of any one of claims 1-6, wherein the treatment comprises palliative care.

9. The use of any one of claims 1-8, wherein the liver cancer Is undergoing sorafenib and/or TRC105 treatment.

10. The use of any one of claims 1-9, wherein the liver cancer is a primaiy liver cancer.

11. The use of claim 10, wherein the primary liver cancer is one of hepatocellular carcinoma (HCC), choiangiocarcinoma, angiosarcoma, and hepatoblastoma.

12. The use of any one of claims 1 -9, wherein the liver cancer is a secondary liver cancer,

13. The use of claim 12, wherein the secondary liver cancer is one or more of: colon, lung, stomach, pancreatic, breast cancers, biliary tract, esophageal, ovarian, prostate, kidney cancer, and melanoma.

14. The use of any one of claims 1-9, wherein the liver cancer is classified as one or more of localized resectable, localized unresectable, and advanced.

15. The use of claims 1 -14, wherein the liver cancer expresses one or more of tissue marker, serum marker, tumor cell marker, and a genetic marker.

16. The use of any one of claims 15, wherein the liver cancer expresses alpha-fetoprotein.

17. A method for treating liver cancer, comprising administering an effective amount of quinacnne to a patient in need thereof, wherein the liver cancer is resistant to standard treatment methods.

18. A method for treating cancer, comprising administering an effective amount of quinacrine to a patient in need thereof, wherein the cancer is a metastatic cancer that is of hepatic origin or a metastatic cancer that has metastasized to the liver from another organ.

19. A method for treating liver cancer, comprising administering an effective amount of quinacrine to a patient in need thereof, wherein the patient Is administered quinacrine at about 400 mg daily for about two weeks to achieve a steady state concentration.

20. The method of claim 19, wherein the patient is administered a maintenance dose of quinac ine after achieving a steady state concentration of quinacrine.

21. The method of claim 20, wherein the maintenance dose is about 350 mg, or 300 mg, or 250 mg, or 200 mg, or 150 mg, or 100 mg of quinacrine,

22. The method of any one of claims 17-21 , wherein the patient Is administered an additional agent that reduces side effects during the administration period to achieve a steady state concentration of quinacrine, wherein the additional agent Is one or more or an anti-emetic, loperamide, diphenoxylate, and atropine,

23. The method of any one of claims 17-22, wherein the liver cancer Is resistant to one or more of chemotherapy, chemoembolization, radiation therapy, cryosurgery, radiofrequency ablation, or ethanoi injection.

24. The method of any one of claims 17-23, wherein the liver cancer is unresectable and/or present In a subject that has coexisting medical condition that prevents surgery, selected from cirrhosis and hepatitis.

25. The method of any one of claims 17-24, wherein the patient's hepatocytes overexpress one or more muitidrug-resistance pumps.

26. The method of claim 25, wherein the muitidrug-resistance pump is one or more of the P-giycoprotein and multidrug resistance-associated protein (MRP),

27. The method of any one of claims 17-26, wherein the treatment comprises one or more of reducing liver tumor load and reducing liver tumor volume,

28. The method of any one of claims 17-26, wherein the treatment comprises palliative care.

29. The method of any one of claims 17-28, wherein the patient is undergoing sorafenlb and/or TRC105 treatment.

30. The method of any one of claims 17-29, wherein the liver cancer is a primary liver cancer.

31. The method of claim 30, wherein the primary liver cancer is one of hepatocellular carcinoma (HCC), choiangiocarcinoma, angiosarcoma, and hepatoblastoma.

32. The method of any one of claims 1 -28, wherein the liver cancer is a secondary liver cancer.

33. The method of claim 32, wherein the secondary liver cancer Is one or more of: coion, lung, stomach, pancreatic, breast cancers, bi!iary tract, esophageal, ovarian, prostate, kidney cancer, and melanoma.

34. The method of any one of claims 17-33, wherein the liver cancer expresses one or more of tissue marker, serum marker, tumor cell marker, and a genetic marker.

35. The method of any one of claims 34, wherein the liver cancer expresses aipha-fetoprotein.

36. The use of any one of claims 17-35, wherein the liver cancer is classified as one or more of localized resectable, localized unresectable, and advanced.

37. A method of treating cancer, comprising administering an effective amount of a quinacrine neoadjuvant therapy to a patient afflicted with a tumor iikely to metastasize to the liver.

38. The method of c!aim 37, wherein the tumor likely to metastasize to the liver is one or more of: colon, lung, stomach, pancreatic, breast cancers, biliary tract, esophageal, ovarian, prostate, kidney cancer, and melanoma.

39. Use of quinacrine In the manufacture of a medicament suitable for oral administration for the treatment of liver cancer, wherein the medicament comprises quinacrine in a dosage amount of from 300-400 mg.

40. An oral dosage regimen of from about 300-400 mg of quinacrine comprising at least about 30 unit