Treatment of dna damage related disorders

Abstract

The present invention provides methods and compositions for the treatment of DNA damage related disorders. One embodiment of the invention is a prophylactic method for the prevention of cancer using a chloroquine compound. Another embodiment is a method for the inhibition of side effects associated with chemotherapeutic and radiotherapeutic agents using chloroquine compounds.

Description

RELATED APPLICATIONS

[0001] This application claims the benefit of priority from U.S. patent application Ser. No. 10/351,733 filed Jan. 24, 2003 and Ser. No. 10/307,077, filed Nov. 27, 2002 which are incorporated herein by reference in their entirety.

BACKGROUND OF THE INVENTION

[0003] Cancer is now the second leading cause of death in the United States. Over 1 million new cases of cancer are expected to be diagnosed in 2003 and over 500,000 people are expected to die of cancer.

[0004] Cancer is typically treated with one or a combination of three types of therapies: surgery, radiation, and chemotherapy. Overall costs for cancer, including treatments, were approximately $170 billion dollars in 2002. The cancer treatments are not only expensive; they are ineffective most of the time and also have many side effects. Hence, there is a demand for more effective cancer prevention and treatment agents, as well as for the prevention and treatment of DNA damage related conditions.

SUMMARY OF THE INVENTION

[0005] The present invention provides compositions, methods, and kits for the treatment of DNA damage related disorders. In one embodiment, a chloroquine compound is administered for the prevention of DNA damage related disorders, like cancer. In another embodiment, the chloroquine compound does not prevent a localized skin cancer alone. In yet another embodiment, the cancers prevented by the chloroquine compounds are a localized skin cancer and a cancer that is not a localized skin cancer. Also described herein are methods of inhibiting the side effects of chemotherapeutic and/or radiotherapeutic agents using chloroquine compounds.

BRIEF DESCRIPTION OF THE FIGURES

[0006] FIG. 1 shows a Kaplan-Meier survival curve of C57/BL6 mice after exposure to 8 Gy total body irradiation (TBI). Half of the cohort received a dose of chloroquine (dashed line) by either IP injection (1.75 mg/kg or 3.5 mg/kg) or in their drinking water (3.5 mg/kg or 7 mg/kg) the day before the TBI. The one mouse which died in the chloroquine-treated group received 1.75 mg/kg by i.p. injection.

[0007] FIG. 2 shows that chloroquine treatment enhances survival after TBI by enhancing recovery of hematopoietic progenitor cells. Five mice received 3.5 mg/kg chloroquine (C) by i.p. injection 24 and 4 hours prior to TBI (bars with diagonal stripes). Five mice received no chloroquine (stippled bars). Fourteen days after irradiation, the cellularity (open bars) of hematopoietic tissues (spleen, thymus, bone marrow) was assessed by a blinded observer on a scale of 0-3 with 3 being normal cellularity. The bars represent the average cellularity of the tissues from the 5 mice in each group.

[0008] FIG. 3 shows a Kaplan-Meier survival curve of AT mice after exposure to 8 Gy TBI. Half of the cohort received a dose of 3.5 mg/kg chloroquine (CHL; dashed line) by i.p. injection 24 and 4 hours prior to the TBI.

[0009] FIG. 4 demonstrates that chloroquine treatment prevents the development of tumors in E.mu.-myc mice. After weaning, a cohort of transgenic mice expressing the c-myc oncogene were started on chloroquine (CHL) at 7.0 mg/kg in the drinking water ((+), solid line). Within 100 days, all of the mice with no drug in the water had died of leukemia, while none of the cohort of mice on drug had succumbed. The latter group of mice was then divided into two groups (timing of this event depicted by heavy arrow), one group of which was taken off of chloroquine ((-), dashed line) and the other group of which was started on i.p. injections of 3.5 mg/kg of chloroquine once a week. Within a month, all of the mice taken off of chloroquine had developed malignancies and all of the mice on the weekly i.p. injections remained tumor-free for months.

[0010] FIG. 5 illustrates that chloroquine treatment reduces the development of tumors in mice injected with the potent chemical carcinogen, 3-methylcholanthrene (3-MC). Chloroquine (CHL, 3.5 mg/kg) was given by i.p. injection 24 and 4 hours prior to 3-MC injection in 30 mice and 30 mice received the carcinogen with no chloroquine pretreatment. The percentage of animals remaining tumor-free is plotted. Statistical significance, log rank test P<0.0001.

[0011] FIG. 6 demonstrates that chloroquine treatment reduces the development of tumors in mice exposed to ionizing radiation in a protocol that induces thymic lymphomas. Chloroquine (CHL, 3.5 mg/kg) was given by i.p. injection 24 and 4 hours prior to irradiation in four successive weeks and animals were subsequently observed for the development of tumors. Statistical significance, log rank test P=0.0012.

DETAILED DESCRIPTION OF THE INVENTION

[0012] Chloroquine Compounds

[0013] The present invention provides methods, compositions, and kits for the prevention of DNA damage related disorders. Chloroquine compounds are useful in practicing the invention described herein. The term "chloroquine compounds" as used herein means chloroquine-like compounds, chloroquine and enantiomers, analogs, derivatives, metabolites, pharmaceutically acceptable salts, and mixtures thereof. Examples of chloroquine compounds include, but are not limited to, chloroquine phosphate, hydroxychloroquine, chloroquine diphosphate, chloroquine sulphate, hydroxychloroquine sulphate, and enantiomers, analogs, derivatives, metabolites, pharmaceutically acceptable salts, and mixtures thereof. The term "chloroquine-like compounds" as used herein means compounds that mimic chloroquine's biological and/or chemical properties.

[0014] In a specific embodiment, the invention is practiced with chloroquine. The chemical structure of chloroquine, N.sup.4-(7-Chloro-4-quinolinyl)-N.sup.1,N.sup.1-diethyl-1,4-pentanediamin- e or 7-chloro-4-(4-diethylamino-1-methylbutylamino)quinoline, is as follows: 1

[0015] Chloroquine (The Merck Index, p. 2220, 1996) is a synthetically manufactured drug containing a quinoline nucleus. Suitable synthesis techniques for chloroquine are well known in the art. For example see U.S. Pat. No. 2,233,970.

[0016] As mentioned above, the chloroquine compounds useful herein include chloroquine analogs and derivatives. A number of chloroquine analogs and derivatives are well known. For example, suitable compounds and methods for synthesizing the same are described in U.S. Pat. Nos. 6,417,177; 6,127,111; 5,639,737; 5,624,938; 5,736,557; 5,596,002; 5,948,791; 5,510,356; 2,653,940; 2,233,970; 5,668,149; 5,639,761; 4,431,807; and 4,421,920.

[0017] Examples of suitable chloroquine compounds include chloroquine phosphate; 7-chloro-4-(4-diethylamino-1-butylamino)quinoline (desmethylchloroquine); 7-hydroxy-4-(4-diethylamino-1-butylamino)quinolin- e; 7-chloro-4-(1-carboxy-4-diethylamino-1-butylamino)quinoline; 7-hydroxy-4-(1-carboxy-4-diethylamino-1-butylamino)quinoline; 7-chloro-4-(1-carboxy-4-diethylamino-1-methylbutylamino)quinoline; 7-hydroxy-4-(1-carboxy-4-diethylamino-1-methylbutylamino)quinoline; 7-chloro-4-(4-ethyl-(2-hydroxyethyl)-amino-1-methylbutylamino)quinoline (hydroxychloroquine); 7-hydroxy-4-(4-ethyl-(2-hydroxyethyl)-amino-1-methy- lbutylamino)quinoline; hydroxychloroquine phosphate; 7-chloro-4-(4-ethyl-(2-hydroxyethyl)-amino-1-butylamino)quinoline (desmethylhydroxychloroquine); 7-hydroxy-4-(4-ethyl-(2-hydroxyethyl)-amin- o-1-butylamino)quinoline; 7-chloro-4-(1-carboxy-4-ethyl-(2-hydroxyethyl)-a- mino-1-butylamino)quinoline; 7-hydroxy-4-(1-carboxy-4-ethyl-(2-hydroxyethy- l)-amino-1-butylamino)quinoline; 7-chloro-4-(1-carboxy-4-ethyl-(2-hydroxye- thyl)-amino-1-methylbutylamino)quinoline; 7-hydroxy-4-(1-carboxy-4-ethyl-(- 2-hydroxyethyl)-amino-1-methylbutylamino)quinoline; 8-[(4-aminopentyl)amino]-6-methoxydihydrochloride quinoline; 1-acetyl-1,2,3,4-tetrahydroquinoline; 8-[4-aminopentyl)amino]-6-methoxyqu- inoline dihydrochloride; 1-butyryl-1,2,3,4-tetrahydroquinoline; 7-chloro-2-(o-chlorostyryl)-4-[4-diethylamino-1-methylbutyl]aminoquinolin- e phosphate; 3-chloro-4-(4-hydroxy-.alpha.,.alpha.'-bis(2-methyl-1-pyrroli- dinyl)-2,5-xylidinoquinoline, 4-[(4-diethylamino)-1-methylbutyl)amino]-6-m- ethoxyquinoline; 3,4-dihydro-1 (2H)-quinolinecarboxyaldehyde; 1,1'-pentamethylenediquinoleinium diiodide; and 8-quinolinol sulfate, enantiomers thereof, as well as suitable pharmaceutical salts thereof.

[0018] Additional suitable chloroquine derivatives include aminoquinoline derivatives and their pharmaceutically acceptable salts such as those described in U.S. Pat. Nos. 5,948,791 and 5,596,002. Suitable examples include (S)-N.sub.2-(7-Chloro-quinolin-4-yl)-N.sub.1,N.sub.1-dimethyl-pro- pane-1,2-diamine; (R)-N.sub.2-(7-chloro-quinolin-4-yl)-N.sub.1,N.sub.1-dim- ethyl-propane-1,2-diamine; N.sub.1-(7-chloro-quinolin-4-yl)-2,N.sub.2,N.su- b.2-trimethyl-propane-1,2-diamine; N.sub.3-(7-chloro-quinolin-4-yl)-N.sub.- 1,N.sub.1-diethyl-propane-1,3-diamine; (RS)-(7-chloro-quinolin-4-yl)-(1-me- thyl-piperidin-3-yl)-amine; (RS)-(7-chloro-quinolin-4-yl)-(1-methyl-pyrrol- idin-3-yl)-amine; (RS)-N.sub.2-(7-Chloro-quinolin-4-yl)-N.sub.1,N.sub.1-di- methyl-propane-1,2-diamine; (RS)-N.sub.2-(7-chloro-quinolin-4-yl)-N.sub.1,- N.sub.1-diethyl-propane-1,2-diamine; (S)-N.sub.2-(7-chloro-quinolin-4-yl)-- N.sub.1,N.sub.1-diethyl-propane-1,2-diamine; (R)-N.sub.2-(7-chloro-quinoli- n-4-yl)-N.sub.1,N.sub.1-diethyl-propane-1,2-diamine; (RS)-7-chloro-quinolin-4-yl)-(1-methyl-2-pyrrolidin-1-yl-ethyl)-amine; N.sub.2-(7-chloro-quinolin-4-yl)-N.sub.1,N.sub.1-dimethyl-ethane-1,2-diam- ine; N.sub.2-(7-chloro-quinolin-4-yl)-N.sub.1,N.sub.1-diethyl-ethane-1,2-d- iamine; N.sub.3-(7-chloro-quinolin-4-yl)-N.sub.1,N.sub.1-dimethyl-propane-- 1,3-diamine; (R)-N.sub.1-(7-chloro-quinolin-4-yl)-N.sub.2,N.sub.2-dimethyl- -propane-1,2-diamine; (S)-N.sub.1-(7-chloro-quinoline-4-yl)-N.sub.2,N.sub.- 2-dimethyl-propane-1,2-diamine; (RS)-(7-chloro-quinolin-4-yl)-(1-methyl-py- rrolidin-2-yl-methyl)-amine; N.sub.1-(7-Chloro-quinolin-4-yl)-N.sub.2-(3-c- hloro-benzyl)-2-methyl-propane-1,2-diamine; N.sub.1-(7-chloro-quinolin-4-y- l)-N.sub.2-(benzyl)-2-methyl-propane-1,2-diamine; N.sub.1-(7-chloro-quinol- in-4-yl)-N.sub.2-(2-hydroxy-3-methoxy-benzyl)-2-methyl-propane-1,2-diamine- ; N.sub.1-(7-chloro-quinolin-4-yl)-N.sub.2-(2-hydroxy-5-methoxy-benzyl)-2-- methyl-propane-1,2-diamine; and N.sub.1-(7-chloro-quinolin-4-yl)-N.sub.2-(- 4-hydroxy-3-methoxy-benzyl)-2-methyl-propane-1,2-diamine; (1S,2S)-N.sub.1-(7-chloro-quinolin-4-yl)-N.sub.2-(benzyl)-cyclohexane-1,2- -diamine; (1S,2S)-N.sub.1-(7-chloro-quinolin-4-yl)-N.sub.2-(4-chlorobenzyl- )-cyclohexane-1,2-diamine; (1S,2S)-N.sub.1-(7-chloro-quinolin-4-yl)-N.sub.- 2-(4-dimethylamino-benzyl)-cyclohexane-1,2-diamine; cis-N.sub.1-(7-chloro-quinolin-4-yl)-N.sub.4-(4-dimethylamino-benzyl)-cyc- lohexane-1,4-diamine; cis-N.sub.1-(7-chloro-quinolin-4-yl)-N.sub.4-(benzyl- )-cyclohexane-1,4-diamine; cis-N.sub.1-(7-chloro-quinolin-4-yl)-N.sub.4-(3- -chloro-benzyl)-cyclohexane-1,4-diamine; cis-N.sub.1-(7-chloro-quinolin-4-- yl)-N.sub.4-(2-hydroxy-4-methoxy-benzyl)-cyclohexane-1,4-diamine; cis-N.sub.1-(7-chloro-quinolin-4-yl)-N.sub.4-(3,5-dimethoxy-benzyl)-cyclo- hexane-1,4-diamine; cis-N.sub.1-(7-chloro-quinolin-4-yl)-N.sub.4-(4-methyl- sulphanyl-benzyl)-cyclohexane-1,4-diamine; cis-N.sub.1-(7-chloro-quinolin-- 4-yl)-N.sub.4-(4-diethylamino-benzyl)-cyclohexane-1,4-diamine; cis-N.sub.1-(7-chloro-quinolin-4-yl)-N.sub.4-(biphenyl-4-yl)methyl-cycloh- exane-1,4-diamine; trans-N.sub.1-(7-chloro-quinolin-4-yl)-N.sub.4-[2-(3,5-- dimethoxy-phenyl)-ethyl]-cyclohexane-1,4-diamine; cis-N.sub.1-(7-chloro-qu- inolin-4-yl)-N.sub.4-(4-methoxy-benzyl)-cyclohexane-1,4-diamine; trans-N.sub.1-(7-chloro-quinolin-4-yl)-N.sub.4-(4-dimethylamino-benzyl)-c- yclohexane-1,4-diamine; and trans-N.sub.1-(7-chloro-quinolin-4-yl)-N.sub.4- -(2,6-difluoro-benzyl)-cyclohexane-1,4-diamine.

[0019] Chloroquine compounds such as chloroquine may exhibit the phenomena of tautomerism, conformational isomerism, geometric isomerism, and/or optical isomerism. The invention covers any tautomeric, conformational isomeric, optical isomeric and/or geometric isomeric forms of the chloroquine compounds, as well as mixtures of these various different forms.

[0020] Chloroquine and hydroxychloroquine are generally racemic mixtures of (-)- and (+)-enantiomers. The (-)-enantiomers are also known as (R)-enantiomers (physical rotation) and 1-enantiomers (optical rotation). The (+)-enantiomers are also known as (S)-enantiomers (physical rotation) and r-enantiomers (optical rotation). The metabolism of the (+)- and the (-)-enantiomers of chloroquine are described in Augustijins and Verbeke (1993) Clin. Pharmacokin. 24(3):259-69; Augustijins, et al. (1999) Eur. J. Drug Metabol. Pharmacokin. 24(1):105-8; DuCharme and Farinotti (1996) Clin. Pharmacokin. 31(4):257-74; Ducharme, et al. (1995) Br. J. Clin. Pharmacol. 40(2):127-33. Preferably, the (-)-enantiomer of chloroquine is used. The enantiomers of chloroquine and hydroxychloroquine may be prepared by procedures known to the art.

[0021] The chloroquine compounds may metabolize to produce active metabolites. The used of active metabolites is also within the scope of the present invention.

[0022] Not intending to be limited by one mechanism, it is believed that the chloroquine compounds and chloroquine-like compounds act by enhancing the activity of Ataxia-Telangiectasia Mutated (ATM) kinase. The agonistic properties of chloroquine on ATM kinase have been demonstrated. Hence, it is intended herein that chloroquine-like compounds include compounds that are agonists of ATM kinase. Agonists of ATM kinase include compounds that promote the dissociation of ATM into active monomers and/or compounds that promote phosphorylation of a serine corresponding to the residue 1981 of ATM kinase of SEQ ID NO:1.

[0023] Use of Chloroquine Compounds

[0024] In one aspect, the invention provides methods of treating an animal subject, including a human. The term "animal subject" as used herein includes humans as well as other mammals. The methods described herein generally involve the administration of effective amounts of chloroquine compounds and/or chloroquine like compounds for the treatment of DNA damage related disorders. The term "DNA damage related disorders" include, but are not limited to, cancer, aging, disorders caused by damage to DNA due to exposure to carcinogens, toxins, free radicals, like oxygen radical, or DNA damaging radiations like ionizing radiation and UV radiation. The chloroquine compounds are also useful for prevention of tissue injury resulting from ischemia, such as that which occurs following myocardial infarction or stroke. The effects of the chloroquine compounds used in the methods described herein include systemic, local, and topical effects. It is preferred that the effects of the chloroquine compounds in the methods described herein are systemic.

[0025] In one embodiment, the chloroquine compounds are used as prophylactics to prevent DNA damage related disorders. The chloroquine compounds are useful in the prevention of cancers caused by toxins, carcinogens, DNA damaging radiations, and/or genetic mutations. For example, chloroquine compounds are useful in the prevention of cancers caused by exposure to toxins and carcinogens like aromatic hydrocarbons, cigarette smoke, acetyl amino fluorine, MTBE, etc. Also, chloroquine compounds are useful in prevention of preventing cancers caused by DNA damaging radiations like UV and ionizing radiation. The ionizing radiations includes both natural and therapeutic radiation exposures. Examples of ionizing radiations are X-rays for diagnostics and radiation therapy used for tumors.

[0026] The prophylactic uses for cancer described herein are not envisioned to encompass the prevention solely of localized skin carcinomas like basal cell epithelioma and squamous cell carcinoma, skin carcinomas, Burkitt's lymphoma, or skin pathologies caused by harmful radiation. When used in patients with actinic keratosis, it is envisioned the chloroquine compounds do not solely inhibit basal cell epithelioma and squamous cell carcinoma. In one embodiment, the chloroquine compounds are used to prevent a localized skin carcinoma and at least one cancer that is not a localized skin carcinoma. Examples of cancers that are not localized skin carcinomas include, but are not limited to, melanomas, lymphomas, prostate cancer, breast cancer, colon cancer, lung cancer, retinoblastoma, neuroblastoma, sarcomas, and ovarian cancer.

[0027] In a preferred embodiment, chloroquine compounds are used in the prevention of one or more of the following cancers--melanomas, prostate cancer, breast cancer, colon cancer, lung cancer, non-Hodgkins lymphoma, retinoblastoma, neuroblastoma, sarcomas, and ovarian cancer.

[0028] The chloroquine compounds can be used to prevent secondary cancers, i.e., cancers that are caused by radiation therapy and chemotherapy used to treat the primary cancer. In one embodiment, the chloroquine compounds are used to prevent the occurrence of breast cancer in patients receiving radiation therapy for non-Hodgkin's lymphoma. Also, in these patients the chloroquine compounds can be used to inhibit the cellular damage caused by the radiation therapy to normal cells and enhance the repair process of the normal cells. The chloroquine compounds are also suitable for prevention of the reoccurrence of cancers in patients who have had prior incidences of cancer.

[0029] In one embodiment, the chloroquine compounds are administered to decrease or prevent the side-effects of radiation therapy used to treat cancer. The chloroquine compounds can be administered prior to, during, or after treatment with radiation. In this embodiment, the beneficial effect of the chloroquine compounds is contemplated to be not solely limited to a beneficial effect on pathological skin conditions like skin carcinomas and dermatoses. The use of chloroquine compounds in combination with radiation therapy is contemplated to protect the normal cells and inhibit the cellular damage caused by the radiation therapy to normal cells and enhance the repair process of the normal cells.

[0030] In one embodiment, the chloroquine compounds are used in immunosuppressed patients, like transplant patients. In immunosuppressed patients, the chloroquine compounds can be used to prevent cancers. The chloroquine compounds can be used to prevent Epstein Barr virus induced lymphoproliferative syndrome.

[0031] In another embodiment, chloroquine compounds are used as prophylactics to inhibit side effects of frequent exposure to X-rays in athletes. This method would also be useful for other patient populations that are frequently exposed to DNA damaging radiations, such as X-ray technicians, police officers, astronauts, and the like. It is known that exposure to X-rays causes DNA damage. Administration of chloroquine compounds is contemplated to inhibit the side-effects of frequent exposure to DNA damaging radiations, including inhibit the damage to cells due to damage to DNA.

[0032] The present invention also provides methods for preventing DNA damage, inhibiting the effects of DNA damage, and stimulating cellular response to DNA damage by administering an effective amount of chloroquine compounds. Not intending to be limited by one mechanism of action, it is contemplated that cellular responses are enhanced by an agonistic activity on ATM kinase by priming the cell to respond to agents which cause DNA damage. Further details on ATM kinase are provided in U.S. Ser. No. ______, attorney docket number SJ-0042, entitled "ATM Kinase Compositions and Methods," filed on the same day as the present invention, which is incorporated by reference herein in its entirety.

[0033] The prophylactic benefits of chloroquine compounds can be obtained by administering in advance of exposure to the DNA damaging agent to provide the enhancing effect in one embodiment. The amount of time prior to the exposure to the DNA damaging agent that the chloroquine compound is administered can vary from days, hours, to minutes. Also, the chloroquine compounds can be administered during exposure to the DNA damaging agent or after such exposure. In one embodiment, the effective amount of a chloroquine compound is an amount which reduces DNA damage, reduces DNA mutation or increases survival of cells exposed to a DNA damaging agent when compared to cells exposed to the same DNA damaging agent and not receiving a chloroquine compound.

[0034] The prophylactic use of chloroquine includes the prevention of tissue injury resulting from ischemia, such as that which occurs following myocardial infarction or stroke. While not intending to be limited to one mechanism of action, it is believed that the chloroquine compounds prevent cellular death due to oxidative damage during reperfusion and as such can ameliorate tissue injury resulting from ischemic injury.

[0035] In one embodiment, chloroquine compounds are used in the treatment of DNA damage related disorders. The chloroquine compounds are used preferably in combination with chemotherapeutic or radiotherapeutic agents to prevent the side-effects associated with the chemotherapeutic agents. It is known that chloroquine compounds can inhibit multiple drug resistance. Hence, it is not intended that the methods described herein produce a beneficial effect on multiple drug resistance alone. In a preferred embodiment, the beneficial effects of chloroquine compounds, when used in combination with chemotherapeutic agents, are due to modulation of ATM kinase activity. It is contemplated that the chloroquine compounds protect the normal cells and inhibit the cellular damage caused by the radiation therapy to normal cells and enhance the repair process of the normal cells.

[0036] In one embodiment, the chloroquine compounds are used to treat and/or prevent disorders caused by oxidative damage. The chloroquine compounds can be administered with anti-oxidants, like vitamin B12, to stimulate the cellular response to DNA damage and promote the repair of the cells exposed to the oxidative agents.

[0037] Therapeutic and Prophylactic Benefits

[0038] In one embodiment, the chloroquine compounds are used as prophylactic agents. For prophylactic benefit, the chloroquine compound may be administered to a patient at risk of developing a DNA damage related disorder like cancer or to a patient reporting one or more of the physiological symptoms of a DNA damage related disorder, even though a diagnosis of such disorder may not have been made. A prophylactic benefit is achieved when a disorder is prevented from afflicting a patient. This prevention can include the affliction of the patient with a milder form of the disorder or the appearance of fewer or no symptoms of the disorder being prevented or the absence of the disorder in the patient being treated.

[0039] In addition to a prophylactic benefit, the chloroquine compounds can be used for their therapeutic benefits. In one embodiment, the chloroquine compounds are used to treat DNA damage related disorders. In a preferred embodiment, the beneficial effect of the chloroquine compounds is not due to an inhibition of multiple drug resistance. The term "treating" as used herein includes achieving a therapeutic benefit and/or a prophylactic benefit. By therapeutic benefit is meant eradication or amelioration of the underlying disorder being treated. For example, in a cancer patient, therapeutic benefit includes eradication or amelioration of the underlying cancer. Also, a therapeutic benefit is achieved with the eradication or amelioration of one or more of the physiological symptoms associated with the underlying disorder such that an improvement is observed in the patient, notwithstanding that the patient may still be afflicted with the underlying disorder. For example, administration of a chloroquine compound to a patient suffering from cancer provides therapeutic benefit not only when the patient's tumor marker level is decreased, but also when an improvement is observed in the patient with respect to other complications that accompany the cancer like pain and psychiatric disorders.

[0040] Effective Amount

[0041] A physician or veterinarian having ordinary skill in the art may readily determine and prescribe the effective amount of the chloroquine compound required in the methods described herein. Pharmaceutical compositions suitable for use in the present invention include compositions wherein the chloroquine compound and other optional active ingredients are present in an effective amount. The effective amounts include doses that partially or completely achieve the desired therapeutic, prophylactic, and/or biological effect. The actual amount effective for a particular application will depend on the condition being treated and the route of administration. Determination of an effective amount is well within the capabilities of those skilled in the art, especially in light of the disclosure herein.

[0042] The effective amount for use in humans can be determined from animal models. For example, a dose for humans can be formulated to achieve circulating and/or gastrointestinal concentrations that have been found to be effective in animals.

[0043] In one embodiment, the effective amount can include the dose ranges, modes of administration, formulations, etc., that have been recommended or approved by any of the various regulatory or advisory organizations in the medical or pharmaceutical arts (eg, FDA, AMA) or by the manufacturer or supplier. Effective amounts of chloroquine can be found, for example, in the Physicians Desk Reference.

[0044] The daily dosage range of chloroquine, in one embodiment, can vary between about 0.1 mg/kg to about 2 gm/kg body weight. The daily dose of a chloroquine compound may be less than about 2 gm/kg, less than about 1.5 gm/kg, or less than about 1 gm/kg. In one embodiment, the daily dose of a chloroquine coumpound is more than about 0.5 mg/kg, more than about 500 mg/kg, or more than about 1 gm/kg. Preferred daily dosage ranges of a chloroquine compound are about 0.5 mg/kg to about 50 mg/kg or about 1.0 mg/kg to about 10 mg/kg body weight. Preferred doses of chloroquine diphosphate are about 3.5 mg/kg and 7.0 mg/kg.

[0045] In one embodiment, the effective amount of chloroquine is administered every other week, once a week, more than once a week, or once a day. The dose of chloroquine can be administered once or more than once a day. In yet another embodiment, the effective amount of a chloroquine compound is an amount that produces the intended beneficial effects but does not produce the side-effects associated with chloroquine compounds, like retinoblastoma.

[0046] In one embodiment, the invention provides a kit comprising a chloroquine compound packaged in association with instructions teaching a method of using the compound according to one or more of the above-described methods. The kit can contain the chloroquine compound packaged in unit dosage form.

[0047] Routes of Administration and Formulation

[0048] The compounds useful in the present invention, or pharmaceutically acceptable salts thereof, can be delivered to the patient using a wide variety of routes or modes of administration. Suitable routes of administration include, but are not limited to, inhalation, transdermal, oral, rectal, transmucosal, intestinal and parenteral administration, including intramuscular, subcutaneous and intravenous injections.

[0049] The formulations useful herein can administer the chloroquine compounds topically or systemically. In one embodiment, the formulation of chloroquine compound is administered systemically. In another embodiment, the formulation of chloroquine compound has a systemic effect if administered either topically or systemically.

[0050] The term "pharmaceutically acceptable salt" means those salts which retain the biological effectiveness and properties of the compounds used in the present invention, and which are not biologically or otherwise undesirable. Such salts include salts with inorganic or organic acids, such as hydrochloric acid, hydrobromic acid, phosphoric acid, nitric acid, sulfuric acid, methanesulfonic acid, p-toluenesulfonic acid, acetic acid, fumaric acid, succinic acid, lactic acid, mandelic acid, malic acid, citric acid, tartaric acid or maleic acid. In addition, if the compounds used in the present invention contain a carboxy group or other acidic group, it may be converted into a pharmaceutically acceptable addition salt with inorganic or organic bases. Examples of suitable bases include sodium hydroxide, potassium hydroxide, ammonia, cyclohexylamine, dicyclohexyl-amine, ethanolamine, diethanolamine and triethanolamine.

[0051] If necessary, the compounds and useful herein may be administered in combination with other therapeutic agents. The choice of therapeutic agents that can be co-administered with the compounds of the invention will depend, in part, on the condition being treated.

[0052] Agents used in accordance with the methods of the invention may be conveniently administered in a pharmaceutical composition containing the active compound in combination with a suitable carrier. Such pharmaceutical compositions may be prepared by methods and contain carriers which are well-known in the art. A generally recognized compendium of such methods and ingredients is Remington: The Science and Practice of Pharmacy, Alfonso R. Gennaro, editor, 20th ed. Lippingcott Williams & Wilkins: Philadelphia, Pa., 2000. A pharmaceutically-acceptabl- e carrier, composition or vehicle, such as a liquid or solid filler, diluent, excipient, or solvent encapsulating material, is involved in carrying or transporting the subject compound from one organ, or portion of the body, to another organ, or portion of the body. Each carrier must be acceptable in the sense of being compatible with the other ingredients of the formulation and not injurious to the patient.

[0053] Examples of materials which may serve as pharmaceutically-acceptabl- e carriers include sugars, such as lactose, glucose and sucrose; starches, such as corn starch and potato starch; cellulose, and its derivatives, such as sodium carboxymethyl cellulose, ethyl cellulose and cellulose acetate; powdered tragacanth; malt; gelatin; talc; excipients, such as cocoa butter and suppository waxes; oils, such as peanut oil, cottonseed oil, safflower oil, sesame oil, olive oil, corn oil and soybean oil; lycols, such as propylene glycol; polyols, such as glycerin, sorbitol, mannitol and polyethylene glycol; esters, such as ethyl oleate and ethyl laurate; agar; buffering agents, such as magnesium hydroxide and aluminum hydroxide; alginic acid; pyrogen-free water; isotonic saline; Ringer's solution; ethyl alcohol; pH buffered solutions; polyesters, polycarbonates and/or polyanhydrides; and other non-toxic compatible substances employed in harmaceutical formulations. Wetting agents, emulsifiers and lubricants, such as sodium lauryl sulfate and magnesium stearate, as well as coloring agents, release agents, coating agents, sweetening, flavoring and perfuming agents, preservatives and antioxidants can also be present in the compositions.

[0054] Agents of use in the invention may be administered parenterally (for example, by intravenous, intraperitoneal, subcutaneous or intramuscular injection), topically (including buccal and sublingual), orally, intranasally, intravaginally, or rectally, with oral administration being particularly preferred.

[0055] For oral therapeutic administration, the composition may be combined with one or more carriers and used in the form of ingestible tablets, buccal tablets, troches, capsules, elixirs, suspensions, syrups, wafers, chewing gums, foods and the like. Also, for oral consumption the active ingredient may be dissolved or suspended in water or other edible oral solutions. Such compositions and preparations should contain at least 0.1% of active compound. The percentage of the compositions and preparations may, of course, be varied and may conveniently be between about 0.1 to about 100% of the weight of a given unit dosage form. The amount of active agent in such therapeutically useful compositions is such that an effective dosage level will be obtained.

[0056] The tablets, troches, pills, capsules, and the like may also contain the following: binders such as gum tragacanth, acacia, corn starch or gelatin; excipients such as dicalcium phosphate; a disintegrating agent such as corn starch, potato starch, alginic acid and the like; a lubricant such as magnesium stearate; and a sweetening agent such as sucrose, fructose, lactose or aspartame or a flavoring agent such as peppermint, oil of wintergreen, or cherry flavoring. The above listing is merely representative and one skilled in the art could envision other binders, excipients, sweetening agents and the like. When the unit dosage form is a capsule, it may contain, in addition to materials of the above type, a liquid carrier, such as a vegetable oil or a polyethylene glycol. Various other materials may be present as coatings or to otherwise modify the physical form of the solid unit dosage form. For instance, tablets, pills, or capsules may be coated with gelatin, wax, shellac or sugar and the like.

[0057] For administration orally, the compounds may be formulated as a sustained release preparation. Numerous techniques for formulating sustained release preparations are described in the following references--U.S. Pat. Nos. 4,891,223; 6,004,582; 5,397,574; 5,419,917; 5,458,005; 5,458,887; 5,458,888; 5,472,708; 6,106,862; 6,103,263; 6,099,862; 6,099,859; 6,096,340; 6,077,541; 5,916,595; 5,837,379; 5,834,023; 5,885,616; 5,456,921; 5,603,956; 5,512,297; 5,399,362; 5,399,359; 5,399,358; 5,725,883; 5,773,025; 6,110,498; 5,952,004; 5,912,013; 5,897,876; 5,824,638; 5,464,633; 5,422,123; and 4,839,177; and WO 98/47491. These references are hereby incorporated herein by reference in their entireties. In a preferred embodiment, the sustained release formulation utilized has an enteric coating.

[0058] For administration by inhalation, the active compound(s) may be conveniently delivered in the form of an aerosol spray presentation from pressurized packs or a nebulizer, with the use of a suitable propellant, e.g., dichlorodifluoromethane, trichlorofluoromethane, dichlorotetrafluoroethane, carbon dioxide or other suitable gas. In the case of a pressurized aerosol the dosage unit may be determined by providing a valve to deliver a metered amount. Capsules and cartridges of e.g. gelatin for use in an inhaler or insufflator may be formulated containing a powder mix of the compound and a suitable powder base such as lactose or starch.

[0059] A syrup or elixir may contain the active agent, sucrose or fructose as a sweetening agent, methyl and propylparabens as preservatives, a dye and flavoring such as cherry or orange flavor. Of course, any material used in preparing any unit dosage form should be pharmaceutically acceptable and substantially non-toxic in the amounts employed. In addition, the active components may be incorporated into sustained-release preparations and devices including, but not limited to, those relying on osmotic pressures to obtain a desired release profile. Once daily formulations for each of the active components are specifically included.

[0060] The compounds may also be formulated in rectal compositions such as suppositories or retention enemas, e.g., containing conventional suppository bases such as cocoa butter or other glycerides.

[0061] In addition to the formulations described previously, the compounds may also be formulated as a depot preparation. Such long acting formulations may be administered by implantation or transcutaneous delivery (for example subcutaneously or intramuscularly), intramuscular injection or a transdermal patch. Thus, for example, the compounds may be formulated with suitable polymeric or hydrophobic materials (for example as an emulsion in an acceptable oil) or ion exchange resins, or as sparingly soluble derivatives, for example, as a sparingly soluble salt.

[0062] The selected dosage level will depend upon a variety of factors including the activity of the particular compound of the present invention employed, the route of administration, the time of administration, the rate of excretion or metabolism of the particular compound being employed, the duration of the treatment, other drugs, compounds and/or materials used in combination with the particular compound employed, the age, sex, weight, condition, general health and prior medical history of the patient being treated, and like factors well-known in the medical arts.

[0063] The invention is described in greater detail by the following non-limiting examples.

EXAMPLES

Example 1

Radioprotection Assay

[0064] HeLa cells were treated with 2 .mu.g/ml of chloroquine for one hour, washed for one hour, and irradiated at 2 or 6 Gy. Subsequently, 1000 cells were plated and assessed for colony formation. Table 1 shows that exposure to chloroquine prior to irradiation increased cell survival by 30%.

1TABLE 1 Treatment Average Number of Colonies* Std. Dev. 2 Gy 444 19.5 Chloroquine + 2 Gy 580 21.2 6 Gy 94.6 10.6 Chloroquine + 6 Gy 129 8.6 *Averages were from five individual samples.

[0065] To test the possibility that chloroquine activation of ATM may cause radioprotection, C57/BL6 mice were exposed to 8 Gy IR, a dose which kills approximately 80% of the mice at around two weeks. Death appears to result from hematopoietic toxicities. The day before total body irradiation (TBI), mice were either given an i.p. injection of chloroquine or chloroquine was added to the drinking water (5 mice--i.p. 1.75 mg/kg chloroquine; 5 mice--i.p. 3.5 mg/kg chloroquine; 5 mice--1.75 mg/kg chloroquine in drinking water; 5 mice--3.5 mg/kg chloroquine in drinking water). FIG. 1 shows a Kaplan-Meier survival curve indicating that a dose of chloroquine prior to the TBI provided significant protection from death. The experiment was reproduced numerous times and analyses of tissues indicated that the protective effect was due to enhanced recovery of hematopoietic cells (bone marrow, spleen, thymus) following irradiation (FIG. 2). Injection of chloroquine prior to the TBI had no effect on the survival of mice lacking ATM genes (FIG. 3), thus indicating that radioprotection may be dependent on ATM.

Example 2

Cancer Prevention

[0066] Transgenic mice expressing the c-myc oncogene under the control of the immunoglobulin enhancer (i.e., E.mu.-myc mice) develop B-cell lymphomas and leukemias with relatively short latencies. Chloroquine was added to the drinking water of a cohort of E.mu.-myc mice and the mice were observed for the development of B-cell malignancies. FIG. 4 demonstrates that 100% of the control transgenic mice developed malignancies within 100 days of birth while 0% of the transgenic mice on chloroquine developed tumors. After .about.120 days, half of the cohort of chloroquine-treated mice were taken off of chloroquine and the other half were switched to receiving a dose of chloroquine by i.p. injection once a week. Within .about.30 days, all of the transgenic mice taken off of the chloroquine had developed tumors while none of the mice receiving weekly i.p. injections developed cancer. At .about.10 months of age, these mice on weekly chloroquine remained cancer-free and appeared healthy and normal.

[0067] The carcinogen 3-methylcholanthrene (3-MC) induces soft tissue sarcomas if injected into muscle and skin carcinomas if applied to the skin (Smart et al., 1986; Noguchi et al., 1996; Horak et al., 1984). This model system has been used to demonstrate that superinduction of p53 after DNA damage (e.g., in a mouse carrying an extra copy of chromosomal DNA containing the p53 gene) protects mice from the development of cancers induced by chemical carcinogen treatments (Garcia-Cao et al., 2002). Therefore, it was determined whether the protective effect observed in these studies could likewise be achieved by biochemically enhancing p53 induction. As demonstrated herein, ATM kinase activation by chloroquine did not induce strand breaks or induce phosphorylation of substrates that normally get phosphorylated by ATM at the sites of DNA breaks, however, it did lead to induction and phosphorylation of p53 protein. Thus, chloroquine pre-treatment may prevent/reduce tumor development resulting from 3-MC injections. Accordingly, doses of 3.5 mg/kg of chloroquine were given by i.p. injection 24 and 4 hours prior to 3-MC injection in 30 mice. Results are shown in FIG. 5. The occurrence of these tumors was readily apparent by visual inspection and confirmed by histologic assessment.

[0068] Multiple exposures to non-lethal doses of ionizing radiation can induce thymic lymphomas in C57BL/6 mice (Boniver et al., 1990). Using a classical, tumor-inducing protocol (Kaplan and Brown, 1952), which consists of four weekly whole-body exposures of 1.75 Gy each, the effect of chloroquine administration on thymic lymphoma formation was examined. Chloroquine (3.5 mg/kg) was administered to 4-week old female C57BL/6 mice by i.p. injection 24 hours and 4 hours prior to each of the four doses of radiation described in the protocol. According to the protocol, tumors were expected to appear within 4-6 months after the last dose of irradiation in 90% of control (untreated) mice. FIG. 6 shows the results of this analysis.

[0069] All publications and patent applications mentioned in this specification are herein incorporated by reference to the same extent as if each individual publication or patent application was specifically and individually indicated to be incorporated by reference.

[0070] It will be apparent to one of ordinary skill in the art that many changes and modifications can be made thereto without departing from the spirit or scope of the appended claims.

Sequence CWU 1

1

1 1 3056 PRT Homo sapiens 1 Met Ser Leu Val Leu Asn Asp Leu Leu Ile Cys Cys Arg Gln Leu Glu 1 5 10 15 His Asp Arg Ala Thr Glu Arg Lys Lys Glu Val Glu Lys Phe Lys Arg 20 25 30 Leu Ile Arg Asp Pro Glu Thr Ile Lys His Leu Asp Arg His Ser Asp 35 40 45 Ser Lys Gln Gly Lys Tyr Leu Asn Trp Asp Ala Val Phe Arg Phe Leu 50 55 60 Gln Lys Tyr Ile Gln Lys Glu Thr Glu Cys Leu Arg Ile Ala Lys Pro 65 70 75 80 Asn Val Ser Ala Ser Thr Gln Ala Ser Arg Gln Lys Lys Met Gln Glu 85 90 95 Ile Ser Ser Leu Val Lys Tyr Phe Ile Lys Cys Ala Asn Arg Arg Ala 100 105 110 Pro Arg Leu Lys Cys Gln Glu Leu Leu Asn Tyr Ile Met Asp Thr Val 115 120 125 Lys Asp Ser Ser Asn Gly Ala Ile Tyr Gly Ala Asp Cys Ser Asn Ile 130 135 140 Leu Leu Lys Asp Ile Leu Ser Val Arg Lys Tyr Trp Cys Glu Ile Ser 145 150 155 160 Gln Gln Gln Trp Leu Glu Leu Phe Ser Val Tyr Phe Arg Leu Tyr Leu 165 170 175 Lys Pro Ser Gln Asp Val His Arg Val Leu Val Ala Arg Ile Ile His 180 185 190 Ala Val Thr Lys Gly Cys Cys Ser Gln Thr Asp Gly Leu Asn Ser Lys 195 200 205 Phe Leu Asp Phe Phe Ser Lys Ala Ile Gln Cys Ala Arg Gln Glu Lys 210 215 220 Ser Ser Ser Gly Leu Asn His Ile Leu Ala Ala Leu Thr Ile Phe Leu 225 230 235 240 Lys Thr Leu Ala Val Asn Phe Arg Ile Arg Val Cys Glu Leu Gly Asp 245 250 255 Glu Ile Leu Pro Thr Leu Leu Tyr Ile Trp Thr Gln His Arg Leu Asn 260 265 270 Asp Ser Leu Lys Glu Val Ile Ile Glu Leu Phe Gln Leu Gln Ile Tyr 275 280 285 Ile His His Pro Lys Gly Ala Lys Thr Gln Glu Lys Gly Ala Tyr Glu 290 295 300 Ser Thr Lys Trp Arg Ser Ile Leu Tyr Asn Leu Tyr Asp Leu Leu Val 305 310 315 320 Asn Glu Ile Ser His Ile Gly Ser Arg Gly Lys Tyr Ser Ser Gly Phe 325 330 335 Arg Asn Ile Ala Val Lys Glu Asn Leu Ile Glu Leu Met Ala Asp Ile 340 345 350 Cys His Gln Val Phe Asn Glu Asp Thr Arg Ser Leu Glu Ile Ser Gln 355 360 365 Ser Tyr Thr Thr Thr Gln Arg Glu Ser Ser Asp Tyr Ser Val Pro Cys 370 375 380 Lys Arg Lys Lys Ile Glu Leu Gly Trp Glu Val Ile Lys Asp His Leu 385 390 395 400 Gln Lys Ser Gln Asn Asp Phe Asp Leu Val Pro Trp Leu Gln Ile Ala 405 410 415 Thr Gln Leu Ile Ser Lys Tyr Pro Ala Ser Leu Pro Asn Cys Glu Leu 420 425 430 Ser Pro Leu Leu Met Ile Leu Ser Gln Leu Leu Pro Gln Gln Arg His 435 440 445 Gly Glu Arg Thr Pro Tyr Val Leu Arg Cys Leu Thr Glu Val Ala Leu 450 455 460 Cys Gln Asp Lys Arg Ser Asn Leu Glu Ser Ser Gln Lys Ser Asp Leu 465 470 475 480 Leu Lys Leu Trp Asn Lys Ile Trp Cys Ile Thr Phe Arg Gly Ile Ser 485 490 495 Ser Glu Gln Ile Gln Ala Glu Asn Phe Gly Leu Leu Gly Ala Ile Ile 500 505 510 Gln Gly Ser Leu Val Glu Val Asp Arg Glu Phe Trp Lys Leu Phe Thr 515 520 525 Gly Ser Ala Cys Arg Pro Ser Cys Pro Ala Val Cys Cys Leu Thr Leu 530 535 540 Ala Leu Thr Thr Ser Ile Val Pro Gly Ala Val Lys Met Gly Ile Glu 545 550 555 560 Gln Asn Met Cys Glu Val Asn Arg Ser Phe Ser Leu Lys Glu Ser Ile 565 570 575 Met Lys Trp Leu Leu Phe Tyr Gln Leu Glu Gly Asp Leu Glu Asn Ser 580 585 590 Thr Glu Val Pro Pro Ile Leu His Ser Asn Phe Pro His Leu Val Leu 595 600 605 Glu Lys Ile Leu Val Ser Leu Thr Met Lys Asn Cys Lys Ala Ala Met 610 615 620 Asn Phe Phe Gln Ser Val Pro Glu Cys Glu His His Gln Lys Asp Lys 625 630 635 640 Glu Glu Leu Ser Phe Ser Glu Val Glu Glu Leu Phe Leu Gln Thr Thr 645 650 655 Phe Asp Lys Met Asp Phe Leu Thr Ile Val Arg Glu Cys Gly Ile Glu 660 665 670 Lys His Gln Ser Ser Ile Gly Phe Ser Val His Gln Asn Leu Lys Glu 675 680 685 Ser Leu Asp Arg Cys Leu Leu Gly Leu Ser Glu Gln Leu Leu Asn Asn 690 695 700 Tyr Ser Ser Glu Ile Thr Asn Ser Glu Thr Leu Val Arg Cys Ser Arg 705 710 715 720 Leu Leu Val Gly Val Leu Gly Cys Tyr Cys Tyr Met Gly Val Ile Ala 725 730 735 Glu Glu Glu Ala Tyr Lys Ser Glu Leu Phe Gln Lys Ala Asn Ser Leu 740 745 750 Met Gln Cys Ala Gly Glu Ser Ile Thr Leu Phe Lys Asn Lys Thr Asn 755 760 765 Glu Glu Phe Arg Ile Gly Ser Leu Arg Asn Met Met Gln Leu Cys Thr 770 775 780 Arg Cys Leu Ser Asn Cys Thr Lys Lys Ser Pro Asn Lys Ile Ala Ser 785 790 795 800 Gly Phe Phe Leu Arg Leu Leu Thr Ser Lys Leu Met Asn Asp Ile Ala 805 810 815 Asp Ile Cys Lys Ser Leu Ala Ser Phe Ile Lys Lys Pro Phe Asp Arg 820 825 830 Gly Glu Val Glu Ser Met Glu Asp Asp Thr Asn Gly Asn Leu Met Glu 835 840 845 Val Glu Asp Gln Ser Ser Met Asn Leu Phe Asn Asp Tyr Pro Asp Ser 850 855 860 Ser Val Ser Asp Ala Asn Glu Pro Gly Glu Ser Gln Ser Thr Ile Gly 865 870 875 880 Ala Ile Asn Pro Leu Ala Glu Glu Tyr Leu Ser Lys Gln Asp Leu Leu 885 890 895 Phe Leu Asp Met Leu Lys Phe Leu Cys Leu Cys Val Thr Thr Ala Gln 900 905 910 Thr Asn Thr Val Ser Phe Arg Ala Ala Asp Ile Arg Arg Lys Leu Leu 915 920 925 Met Leu Ile Asp Ser Ser Thr Leu Glu Pro Thr Lys Ser Leu His Leu 930 935 940 His Met Tyr Leu Met Leu Leu Lys Glu Leu Pro Gly Glu Glu Tyr Pro 945 950 955 960 Leu Pro Met Glu Asp Val Leu Glu Leu Leu Lys Pro Leu Ser Asn Val 965 970 975 Cys Ser Leu Tyr Arg Arg Asp Gln Asp Val Cys Lys Thr Ile Leu Asn 980 985 990 His Val Leu His Val Val Lys Asn Leu Gly Gln Ser Asn Met Asp Ser 995 1000 1005 Glu Asn Thr Arg Asp Ala Gln Gly Gln Phe Leu Thr Val Ile Gly Ala 1010 1015 1020 Phe Trp His Leu Thr Lys Glu Arg Lys Tyr Ile Phe Ser Val Arg Met 1025 1030 1035 1040 Ala Leu Val Asn Cys Leu Lys Thr Leu Leu Glu Ala Asp Pro Tyr Ser 1045 1050 1055 Lys Trp Ala Ile Leu Asn Val Met Gly Lys Asp Phe Pro Val Asn Glu 1060 1065 1070 Val Phe Thr Gln Phe Leu Ala Asp Asn His His Gln Val Arg Met Leu 1075 1080 1085 Ala Ala Glu Ser Ile Asn Arg Leu Phe Gln Asp Thr Lys Gly Asp Ser 1090 1095 1100 Ser Arg Leu Leu Lys Ala Leu Pro Leu Lys Leu Gln Gln Thr Ala Phe 1105 1110 1115 1120 Glu Asn Ala Tyr Leu Lys Ala Gln Glu Gly Met Arg Glu Met Ser His 1125 1130 1135 Ser Ala Glu Asn Pro Glu Thr Leu Asp Glu Ile Tyr Asn Arg Lys Ser 1140 1145 1150 Val Leu Leu Thr Leu Ile Ala Val Val Leu Ser Cys Ser Pro Ile Cys 1155 1160 1165 Glu Lys Gln Ala Leu Phe Ala Leu Cys Lys Ser Val Lys Glu Asn Gly 1170 1175 1180 Leu Glu Pro His Leu Val Lys Lys Val Leu Glu Lys Val Ser Glu Thr 1185 1190 1195 1200 Phe Gly Tyr Arg Arg Leu Glu Asp Phe Met Ala Ser His Leu Asp Tyr 1205 1210 1215 Leu Val Leu Glu Trp Leu Asn Leu Gln Asp Thr Glu Tyr Asn Leu Ser 1220 1225 1230 Ser Phe Pro Phe Ile Leu Leu Asn Tyr Thr Asn Ile Glu Asp Phe Tyr 1235 1240 1245 Arg Ser Cys Tyr Lys Val Leu Ile Pro His Leu Val Ile Arg Ser His 1250 1255 1260 Phe Asp Glu Val Lys Ser Ile Ala Asn Gln Ile Gln Glu Asp Trp Lys 1265 1270 1275 1280 Ser Leu Leu Thr Asp Cys Phe Pro Lys Ile Leu Val Asn Ile Leu Pro 1285 1290 1295 Tyr Phe Ala Tyr Glu Gly Thr Arg Asp Ser Gly Met Ala Gln Gln Arg 1300 1305 1310 Glu Thr Ala Thr Lys Val Tyr Asp Met Leu Lys Ser Glu Asn Leu Leu 1315 1320 1325 Gly Lys Gln Ile Asp His Leu Phe Ile Ser Asn Leu Pro Glu Ile Val 1330 1335 1340 Val Glu Leu Leu Met Thr Leu His Glu Pro Ala Asn Ser Ser Ala Ser 1345 1350 1355 1360 Gln Ser Thr Asp Leu Cys Asp Phe Ser Gly Asp Leu Asp Pro Ala Pro 1365 1370 1375 Asn Pro Pro His Phe Pro Ser His Val Ile Lys Ala Thr Phe Ala Tyr 1380 1385 1390 Ile Ser Asn Cys His Lys Thr Lys Leu Lys Ser Ile Leu Glu Ile Leu 1395 1400 1405 Ser Lys Ser Pro Asp Ser Tyr Gln Lys Ile Leu Leu Ala Ile Cys Glu 1410 1415 1420 Gln Ala Ala Glu Thr Asn Asn Val Tyr Lys Lys His Arg Ile Leu Lys 1425 1430 1435 1440 Ile Tyr His Leu Phe Val Ser Leu Leu Leu Lys Asp Ile Lys Ser Gly 1445 1450 1455 Leu Gly Gly Ala Trp Ala Phe Val Leu Arg Asp Val Ile Tyr Thr Leu 1460 1465 1470 Ile His Tyr Ile Asn Gln Arg Pro Ser Cys Ile Met Asp Val Ser Leu 1475 1480 1485 Arg Ser Phe Ser Leu Cys Cys Asp Leu Leu Ser Gln Val Cys Gln Thr 1490 1495 1500 Ala Val Thr Tyr Cys Lys Asp Ala Leu Glu Asn His Leu His Val Ile 1505 1510 1515 1520 Val Gly Thr Leu Ile Pro Leu Val Tyr Glu Gln Val Glu Val Gln Lys 1525 1530 1535 Gln Val Leu Asp Leu Leu Lys Tyr Leu Val Ile Asp Asn Lys Asp Asn 1540 1545 1550 Glu Asn Leu Tyr Ile Thr Ile Lys Leu Leu Asp Pro Phe Pro Asp His 1555 1560 1565 Val Val Phe Lys Asp Leu Arg Ile Thr Gln Gln Lys Ile Lys Tyr Ser 1570 1575 1580 Arg Gly Pro Phe Ser Leu Leu Glu Glu Ile Asn His Phe Leu Ser Val 1585 1590 1595 1600 Ser Val Tyr Asp Ala Leu Pro Leu Thr Arg Leu Glu Gly Leu Lys Asp 1605 1610 1615 Leu Arg Arg Gln Leu Glu Leu His Lys Asp Gln Met Val Asp Ile Met 1620 1625 1630 Arg Ala Ser Gln Asp Asn Pro Gln Asp Gly Ile Met Val Lys Leu Val 1635 1640 1645 Val Asn Leu Leu Gln Leu Ser Lys Met Ala Ile Asn His Thr Gly Glu 1650 1655 1660 Lys Glu Val Leu Glu Ala Val Gly Ser Cys Leu Gly Glu Val Gly Pro 1665 1670 1675 1680 Ile Asp Phe Ser Thr Ile Ala Ile Gln His Ser Lys Asp Ala Ser Tyr 1685 1690 1695 Thr Lys Ala Leu Lys Leu Phe Glu Asp Lys Glu Leu Gln Trp Thr Phe 1700 1705 1710 Ile Met Leu Thr Tyr Leu Asn Asn Thr Leu Val Glu Asp Cys Val Lys 1715 1720 1725 Val Arg Ser Ala Ala Val Thr Cys Leu Lys Asn Ile Leu Ala Thr Lys 1730 1735 1740 Thr Gly His Ser Phe Trp Glu Ile Tyr Lys Met Thr Thr Asp Pro Met 1745 1750 1755 1760 Leu Ala Tyr Leu Gln Pro Phe Arg Thr Ser Arg Lys Lys Phe Leu Glu 1765 1770 1775 Val Pro Arg Phe Asp Lys Glu Asn Pro Phe Glu Gly Leu Asp Asp Ile 1780 1785 1790 Asn Leu Trp Ile Pro Leu Ser Glu Asn His Asp Ile Trp Ile Lys Thr 1795 1800 1805 Leu Thr Cys Ala Phe Leu Asp Ser Gly Gly Thr Lys Cys Glu Ile Leu 1810 1815 1820 Gln Leu Leu Lys Pro Met Cys Glu Val Lys Thr Asp Phe Cys Gln Thr 1825 1830 1835 1840 Val Leu Pro Tyr Leu Ile His Asp Ile Leu Leu Gln Asp Thr Asn Glu 1845 1850 1855 Ser Trp Arg Asn Leu Leu Ser Thr His Val Gln Gly Phe Phe Thr Ser 1860 1865 1870 Cys Leu Arg His Phe Ser Gln Thr Ser Arg Ser Thr Thr Pro Ala Asn 1875 1880 1885 Leu Asp Ser Glu Ser Glu His Phe Phe Arg Cys Cys Leu Asp Lys Lys 1890 1895 1900 Ser Gln Arg Thr Met Leu Ala Val Val Asp Tyr Met Arg Arg Gln Lys 1905 1910 1915 1920 Arg Pro Ser Ser Gly Thr Ile Phe Asn Asp Ala Phe Trp Leu Asp Leu 1925 1930 1935 Asn Tyr Leu Glu Val Ala Lys Val Ala Gln Ser Cys Ala Ala His Phe 1940 1945 1950 Thr Ala Leu Leu Tyr Ala Glu Ile Tyr Ala Asp Lys Lys Ser Met Asp 1955 1960 1965 Asp Gln Glu Lys Arg Ser Leu Ala Phe Glu Glu Gly Ser Gln Ser Thr 1970 1975 1980 Thr Ile Ser Ser Leu Ser Glu Lys Ser Lys Glu Glu Thr Gly Ile Ser 1985 1990 1995 2000 Leu Gln Asp Leu Leu Leu Glu Ile Tyr Arg Ser Ile Gly Glu Pro Asp 2005 2010 2015 Ser Leu Tyr Gly Cys Gly Gly Gly Lys Met Leu Gln Pro Ile Thr Arg 2020 2025 2030 Leu Arg Thr Tyr Glu His Glu Ala Met Trp Gly Lys Ala Leu Val Thr 2035 2040 2045 Tyr Asp Leu Glu Thr Ala Ile Pro Ser Ser Thr Arg Gln Ala Gly Ile 2050 2055 2060 Ile Gln Ala Leu Gln Asn Leu Gly Leu Cys His Ile Leu Ser Val Tyr 2065 2070 2075 2080 Leu Lys Gly Leu Asp Tyr Glu Asn Lys Asp Trp Cys Pro Glu Leu Glu 2085 2090 2095 Glu Leu His Tyr Gln Ala Ala Trp Arg Asn Met Gln Trp Asp His Cys 2100 2105 2110 Thr Ser Val Ser Lys Glu Val Glu Gly Thr Ser Tyr His Glu Ser Leu 2115 2120 2125 Tyr Asn Ala Leu Gln Ser Leu Arg Asp Arg Glu Phe Ser Thr Phe Tyr 2130 2135 2140 Glu Ser Leu Lys Tyr Ala Arg Val Lys Glu Val Glu Glu Met Cys Lys 2145 2150 2155 2160 Arg Ser Leu Glu Ser Val Tyr Ser Leu Tyr Pro Thr Leu Ser Arg Leu 2165 2170 2175 Gln Ala Ile Gly Glu Leu Glu Ser Ile Gly Glu Leu Phe Ser Arg Ser 2180 2185 2190 Val Thr His Arg Gln Leu Ser Glu Val Tyr Ile Lys Trp Gln Lys His 2195 2200 2205 Ser Gln Leu Leu Lys Asp Ser Asp Phe Ser Phe Gln Glu Pro Ile Met 2210 2215 2220 Ala Leu Arg Thr Val Ile Leu Glu Ile Leu Met Glu Lys Glu Met Asp 2225 2230 2235 2240 Asn Ser Gln Arg Glu Cys Ile Lys Asp Ile Leu Thr Lys His Leu Val 2245 2250 2255 Glu Leu Ser Ile Leu Ala Arg Thr Phe Lys Asn Thr Gln Leu Pro Glu 2260 2265 2270 Arg Ala Ile Phe Gln Ile Lys Gln Tyr Asn Ser Val Ser Cys Gly Val 2275 2280 2285 Ser Glu Trp Gln Leu Glu Glu Ala Gln Val Phe Trp Ala Lys Lys Glu 2290 2295 2300 Gln Ser Leu Ala Leu Ser Ile Leu Lys Gln Met Ile Lys Lys Leu Asp 2305 2310 2315 2320 Ala Ser Cys Ala Ala Asn Asn Pro Ser Leu Lys Leu Thr Tyr Thr Glu 2325 2330 2335 Cys Leu Arg Val Cys Gly Asn Trp Leu Ala Glu Thr Cys Leu Glu Asn 2340 2345 2350 Pro Ala Val Ile Met Gln Thr Tyr Leu Glu Lys Ala Val Glu Val Ala 2355 2360 2365 Gly Asn Tyr Asp Gly Glu Ser Ser Asp Glu Leu Arg Asn Gly Lys Met 2370 2375 2380 Lys Ala Phe Leu Ser Leu Ala Arg Phe Ser Asp Thr Gln Tyr Gln Arg 2385 2390 2395 2400 Ile Glu Asn Tyr Met Lys Ser Ser Glu Phe Glu Asn Lys Gln Ala Leu 2405 2410 2415 Leu Lys Arg Ala Lys Glu Glu Val

Gly Leu Leu Arg Glu His Lys Ile 2420 2425 2430 Gln Thr Asn Arg Tyr Thr Val Lys Val Gln Arg Glu Leu Glu Leu Asp 2435 2440 2445 Glu Leu Ala Leu Arg Ala Leu Lys Glu Asp Arg Lys Arg Phe Leu Cys 2450 2455 2460 Lys Ala Val Glu Asn Tyr Ile Asn Cys Leu Leu Ser Gly Glu Glu His 2465 2470 2475 2480 Asp Met Trp Val Phe Arg Leu Cys Ser Leu Trp Leu Glu Asn Ser Gly 2485 2490 2495 Val Ser Glu Val Asn Gly Met Met Lys Arg Asp Gly Met Lys Ile Pro 2500 2505 2510 Thr Tyr Lys Phe Leu Pro Leu Met Tyr Gln Leu Ala Ala Arg Met Gly 2515 2520 2525 Thr Lys Met Met Gly Gly Leu Gly Phe His Glu Val Leu Asn Asn Leu 2530 2535 2540 Ile Ser Arg Ile Ser Met Asp His Pro His His Thr Leu Phe Ile Ile 2545 2550 2555 2560 Leu Ala Leu Ala Asn Ala Asn Arg Asp Glu Phe Leu Thr Lys Pro Glu 2565 2570 2575 Val Ala Arg Arg Ser Arg Ile Thr Lys Asn Val Pro Lys Gln Ser Ser 2580 2585 2590 Gln Leu Asp Glu Asp Arg Thr Glu Ala Ala Asn Arg Ile Ile Cys Thr 2595 2600 2605 Ile Arg Ser Arg Arg Pro Gln Met Val Arg Ser Val Glu Ala Leu Cys 2610 2615 2620 Asp Ala Tyr Ile Ile Leu Ala Asn Leu Asp Ala Thr Gln Trp Lys Thr 2625 2630 2635 2640 Gln Arg Lys Gly Ile Asn Ile Pro Ala Asp Gln Pro Ile Thr Lys Leu 2645 2650 2655 Lys Asn Leu Glu Asp Val Val Val Pro Thr Met Glu Ile Lys Val Asp 2660 2665 2670 His Thr Gly Glu Tyr Gly Asn Leu Val Thr Ile Gln Ser Phe Lys Ala 2675 2680 2685 Glu Phe Arg Leu Ala Gly Gly Val Asn Leu Pro Lys Ile Ile Asp Cys 2690 2695 2700 Val Gly Ser Asp Gly Lys Glu Arg Arg Gln Leu Val Lys Gly Arg Asp 2705 2710 2715 2720 Asp Leu Arg Gln Asp Ala Val Met Gln Gln Val Phe Gln Met Cys Asn 2725 2730 2735 Thr Leu Leu Gln Arg Asn Thr Glu Thr Arg Lys Arg Lys Leu Thr Ile 2740 2745 2750 Cys Thr Tyr Lys Val Val Pro Leu Ser Gln Arg Ser Gly Val Leu Glu 2755 2760 2765 Trp Cys Thr Gly Thr Val Pro Ile Gly Glu Phe Leu Val Asn Asn Glu 2770 2775 2780 Asp Gly Ala His Lys Arg Tyr Arg Pro Asn Asp Phe Ser Ala Phe Gln 2785 2790 2795 2800 Cys Gln Lys Lys Met Met Glu Val Gln Lys Lys Ser Phe Glu Glu Lys 2805 2810 2815 Tyr Glu Val Phe Met Asp Val Cys Gln Asn Phe Gln Pro Val Phe Arg 2820 2825 2830 Tyr Phe Cys Met Glu Lys Phe Leu Asp Pro Ala Ile Trp Phe Glu Lys 2835 2840 2845 Arg Leu Ala Tyr Thr Arg Ser Val Ala Thr Ser Ser Ile Val Gly Tyr 2850 2855 2860 Ile Leu Gly Leu Gly Asp Arg His Val Gln Asn Ile Leu Ile Asn Glu 2865 2870 2875 2880 Gln Ser Ala Glu Leu Val His Ile Asp Leu Gly Val Ala Phe Glu Gln 2885 2890 2895 Gly Lys Ile Leu Pro Thr Pro Glu Thr Val Pro Phe Arg Leu Thr Arg 2900 2905 2910 Asp Ile Val Asp Gly Met Gly Ile Thr Gly Val Glu Gly Val Phe Arg 2915 2920 2925 Arg Cys Cys Glu Lys Thr Met Glu Val Met Arg Asn Ser Gln Glu Thr 2930 2935 2940 Leu Leu Thr Ile Val Glu Val Leu Leu Tyr Asp Pro Leu Phe Asp Trp 2945 2950 2955 2960 Thr Met Asn Pro Leu Lys Ala Leu Tyr Leu Gln Gln Arg Pro Glu Asp 2965 2970 2975 Glu Thr Glu Leu His Pro Thr Leu Asn Ala Asp Asp Gln Glu Cys Lys 2980 2985 2990 Arg Asn Leu Ser Asp Ile Asp Gln Ser Phe Asp Lys Val Ala Glu Arg 2995 3000 3005 Val Leu Met Arg Leu Gln Glu Lys Leu Lys Gly Val Glu Glu Gly Thr 3010 3015 3020 Val Leu Ser Val Gly Gly Gln Val Asn Leu Leu Ile Gln Gln Ala Ile 3025 3030 3035 3040 Asp Pro Lys Asn Leu Ser Arg Leu Phe Pro Gly Trp Lys Ala Trp Val 3045 3050 3055

Claims

1. A method of prophylactic treatment for a DNA damage related disorder comprising administering to an animal subject in need thereof an effective amount of a chloroquine compound, with the proviso that the chloroquine compound does not prevent a localized skin carcinoma alone.

2. The method of claim 1 wherein the chloroquine compound prevents a localized skin carcinoma and at least one cancer that is not a localized skin carcinoma.

3. The method of claim 1 wherein the DNA damage related disorder is aging.

4. The method of claim 1 wherein the DNA damage related disorder is cancer.

5. The method of claim 1 wherein the DNA damage related disorder is caused by radiation, toxins, carcinogens, natural processes or exposure to agents which cause oxidative DNA damage.

6. A method of inhibiting side-effects of radiation therapy comprising administering to an animal subject in need thereof an effective amount of a chloroquine compound.

7. A method of inhibiting development of secondary cancers comprising administering to an animal subject in need thereof an effective amount of a chloroquine compound.

8. A method of inhibiting side-effects of chemotherapy comprising administering to an animal subject in need thereof an effective amount of a chloroquine compound.

9. A prophylactic method of prevention of X-ray related DNA damage comprising administering to an athlete in need thereof an effective amount of a chloroquine compound.

10. The method of claims 1, 6, 7, 8, or 9 wherein the chloroquine compound is selected from the group consisting of chloroquine, chloroquine phosphate, hydroxychloroquine, chloroquine diphosphate, chloroquine sulphate, hydroxychloroquine sulphate, or enantiomers, derivatives, analogs, metabolites, pharmaceutically acceptable salts, and mixtures thereof.

11. The method of claim 10 wherein the compound is chloroquine, chloroquine phosphate or chloroquine diphosphate.

12. The method of claims 1, 6, 7, 8, or 9 wherein chloroquine compound has a systemic effect.

13. The method of claims 1, 6, 7, 8, or 9 wherein the amount of the compound administered is at least about 0.1 mg/kg/day.

14. The method of claims 1, 6, 7, 8, or 9 wherein the amount of the compound administered is up to about 10 mg/kg/day.

15. The method of claims 1, 6, 7, 8, or 9 wherein the amount of the compound administered is more than about 0.1 mg/kg/day.

16 The method of claims 1, 6, 7, 8, or 9 wherein the amount of the compound administered is more than about 1.0 mg/kg/day.

17. The method of claims 1, 6, 7, 8, or 9 wherein the amount of the compound administered is less than about 50 mg/kg/day.

18. The method of claims 1, 6, 7, 8, or 9 wherein the amount of the compound administered is less than about 10 mg/kg/day.

19. The method of claims 1, 6, 7, 8, or 9 wherein the chloroquine compound is administered more than once a week.

20. The method claims 1, 6, 7, 8, or 9 wherein the chloroquine compound is administered daily.

21. The method of claims 1, 6, 7, 8, or 9 wherein the chloroquine compound is formulated in a sustained release formulation.