RU2803586C2 - 3'-deamino-3'-(2"-pyrroline-1"-yl)-5-imino-13-deoxyantracyclins and production methods - Google Patents

Abstract

FIELD: medicine.

SUBSTANCE: present invention relates to a compound of the formula

,

where R₁ is H; R₂ and R₃ are OH; R₄ is O-C_1-6alkyl; R₅ is NH, and R₆ is 2-pyrrolino-daunozamine or 2-pyrrolino-epidaunozamine, which is used for suppression of fibrosarcoma cell growth. The invention also relates to a pharmaceutical composition and to a synergetic combined preparation containing this compound, as well as to a method for suppression of fibrosarcoma cell growth.

EFFECT: provision of an anthracycline compound containing 2-pyrrolino-group, which increases anticancer activity, while saving a good safety profile.

20 cl, 4 tbl, 4 ex

Description

Cross reference to related application

Priority is claimed to U.S. Patent Application No. 15/434,617, filed February 16, 2017, the entire contents of which are incorporated herein by reference.

Field of technology to which the invention relates

The present disclosure relates to 13-deoxyanthracyclines, and more particularly to 3'-deamino-3'-(2"-pyrrolin-1"-yl)-5-imino-13-deoxyanthracyclines, and to methods for their preparation and use. The present disclosure also relates to synergistic combinations of 2-pyrrolino-13-deoxyanthracyclines and 13-deoxyanthracyclines.

Prerequisites for creating the invention

Doxorubicin, an anthracycline antibiotic, is one of the most widely used and very potent anticancer agents. It has a broad spectrum of anticancer activity, being useful in the treatment of acute lymphoblastic leukemia, acute myeloblastic leukemia, Wilms tumor, neuroblastoma, soft tissue and bone sarcomas, breast carcinoma, ovarian carcinoma, transitional cell carcinoma of the bladder, thyroid carcinoma, gastric carcinoma, Hodgkin's, malignant lymphoma and bronchogenic carcinoma (instructions for use of doxorubicin, Pfizer Inc., New York, NY). To make wider use of the enormous tumoricidal potential associated with the structure of anthracycline antibiotics, thousands of synthetic derivatives have been described, including their analogs bound to various carrier macromolecules.

Nagy et al. (Proc. Natl. Acad. Sci. vol. 93, pp. 2464-2469, 1996) described that modification of doxorubicin (compound I ) to 3'-deamino-3'-(2"-pyrrolin-1"-yl) doxorubicin (compound II; 2-pyrrolinodoxorubicin) increased anticancer activity against human and murine cancer cells in vitro by 500-1000 times compared with doxorubicin.

Doxorubicin quickly penetrates into the cell nucleus and binds with high affinity to DNA through non-covalent intercalation between base pairs, leading to inhibition of the synthesis of biomacromolecules. It is generally accepted that the biological effects of doxorubicin are associated with its ability to act as an inhibitor of topoisomerase II (topoII), disrupting the religation step of this enzyme and forming a ternary doxorubicin-DNA-topoII cleavable complex. It turned out that 2-pyrrolino-doxorubicin acts in the same way (Stepankova et al., Biochemical Pharmacology 82: 227-235, 2011). There are only minor differences in the modifications of DNA by these anthracyclines and the resulting conformational changes in DNA. Similarly, the ability of 2-pyrrolino-doxorubicin DNA modifications to inhibit the catalytic activity of topoisomerase II is not significantly different from the ability of doxorubicin. (Stepankova et al., supra).

However, increasing the activity of doxorubicin by converting it to a 2-pyrroline compound made it nonspecific with regard to cytotoxicity. Consequently, 2-pyrrolinodoxorubicin is toxic or lethal at or below the doses required for anticancer effects (Jungwirth, A et al., International Journal of Oncology 10: 877-884, 1997; and Szepeshazi, K et al., Clinical Cancer Research, 7: 2854-2861, 2001). As a result, 2-pyrrolinodoxorubicin has not been useful as a drug for the treatment of cancer. Jungwirth et al., supra and Szepeshazi et al., supra proposed to modify 2-pyrrolinodoxorubicin by conjugating it at the 14-OH position with analogues of somatostatin or luteinizing hormone releasing factor. However, these modifications resulted in compounds that were specific for certain tumors, thereby limiting the broad spectrum of activity of the parent compound.

The essence of the invention

The present invention provides anthracycline compounds containing a 2-pyrrolino group that enhance broad-spectrum anticancer activity while maintaining the good safety profile and broad-spectrum characteristic of an anthracycline. In particular, the compounds of the present invention are represented by formula X :

Where:

R ₁ , R ₂ and R ₃ are each independently H or OH;

R ₄ is selected from the group consisting of H, OH, alkyl and O-alkyl;

R ₅ represents O or NH; And

R ₆ is 2-pyrrolino-daunosamine or 2-pyrrolino-epidaunosamine;

their pharmaceutically acceptable salts, their deuterated forms, their prodrugs, their isomers, their solvates and mixtures thereof.

Preferably, the compound is an anthracycline derivative selected from the group consisting of 2-pyrrolino-13-deoxydoxorubicin, 2-pyrrolino-13-deoxydaunorubicin, 2-pyrrolino-13-deoxyepirubicin, 2-pyrrolino-13-deoxycarminomycin and 2-pyrrolino-13-deoxycarminomycin. 13-deoxyidarubicin and their 5-imino (R ₅ = NH) analogues.

The present invention also relates to a pharmaceutical composition which includes a compound of formula X and a pharmaceutically acceptable carrier or excipient.

Another aspect of the present invention relates to a method for inhibiting cell growth, which includes administering to a mammal in need thereof a therapeutically effective amount of a compound of formulaXor composition comprising a compound of the formulaX.

The present invention also provides a therapeutically effective amount of a synergistic combination drug comprising a first compound and a second compound, wherein: the first compound is a 2-pyrrolino-13-deoxyanthracycline having formula X :

Where:

R ₁ , R ₂ and R ₃ are each independently H or OH;

R ₄ is selected from the group consisting of H, OH, alkyl and O-alkyl;

R ₅ represents O or NH; And

R ₆ is 2-pyrrolino-daunosamine or 2-pyrrolino-epidaunosamine;

its pharmaceutically acceptable salts, its prodrugs, its deuterated forms, its isomers, its solvates and mixtures thereof; And

the second compound is a 13-deoxyanthracycline, said 13-deoxyanthracycline having the formula:

Where

each of R ₁ , R ₂ and R ₃ is independently selected from the group consisting of H and OH;

R ₄ is selected from the group consisting of H, OH, alkyl and O-alkyl;

R ₅ is selected from the group consisting of O and NH; And

R ₆ represents a sugar component;

its pharmaceutically acceptable salts, its prodrugs, its deuterated forms, its isomers, its solvates and mixtures thereof;

And

wherein a therapeutically effective amount of the synergistic combination drug inhibits cell growth.

Preferably, the first compound is an anthracycline derivative selected from the group consisting of 2-pyrrolino-13-deoxydoxorubicin, 2-pyrrolino-13-deoxydaunorubicin, 2-pyrrolino-13-deoxyepirubicin, 2-pyrrolino-13-deoxycarminomycin and 2-pyrrolino-13-deoxycarminomycin. 13-deoxyidarubicin and their 5-imino analogues, and the second compound is an anthracycline derivative selected from the group consisting of 13-deoxydoxorubicin, 13-deoxydaunorubicin, 13-deoxyepirubicin, 13-deoxycarminomycin, 13-deoxyidarubicin, 13-deoxyannamycin and 13 - deoxyamrubicin and their 5-imino analogues.

In another aspect of the present invention, there is provided a pharmaceutical composition that includes the synergistic combination drug disclosed above and a pharmaceutically acceptable carrier or excipient.

The present invention also provides a method of inhibiting cell growth with a synergistic combination drug containing a first compound as defined above and a second compound as defined above, comprising administering to a mammal in need thereof a therapeutically effective amount of the synergistic combination drug, pharmaceutically acceptable salts thereof, deuterated forms thereof , its prodrugs, its isomers or its solvates.

A synergistic combination drug may be formulated and administered wherein the second compound is present in an amount that is less than a therapeutically effective amount thereof. It is also possible to formulate and administer a synergistic combination drug wherein the first compound is present in an amount that is less than a therapeutically effective amount thereof. As an example, when the second compound is present in an amount that is less than a therapeutically effective amount thereof, the weight ratio of the second compound to the first component can typically be from about 0.005 to about 0.10:1, and more typically from about 0.01 to about 0.10:1. Also, by way of example, when the first compound is present in an amount that is less than a therapeutically effective amount thereof, the weight ratio of the first compound to the second component can typically be from about 0.005 to about 0.10:1, and more typically from about 0.01 to about 0.10:1.

Other objects and advantages of the present invention will become apparent to those skilled in the art from the following detailed description, wherein only the preferred embodiments are shown and described merely by way of illustration of the best mode of implementation. As will be appreciated, the invention is capable of other and different embodiments, and certain details thereof may be modified in certain obvious respects without departing from the present disclosure. Accordingly, the description should be considered illustrative in nature and not limiting.

Detailed Description of the Invention

Although certain embodiments of the present invention are described in detail below, it should be understood that the invention is not limited in its application to the details of the compositions and combinations of compounds described in the accompanying examples and experiments, since the present invention may have other embodiments and may be carried out in various ways.

The 2-pyrrolino-13-deoxyanthracycline compounds used in accordance with the present invention have the following formula X :

wherein R ₁ , R ₂ and R ₃ are each independently H or OH; R ₄ is selected from the group consisting of H, OH, alkyl and O-alkyl; R ₅ represents O or NH; and R ₆ is 2-pyrrolino-daunosamine or 2-pyrrolino-epidaunosamine. More typical 2-pyrrolino-13-deoxyanthracyclines are 2-pyrrolino-13-deoxydoxorubicin, 2-pyrrolino-13-deoxydaunorubicin, 2-pyrrolino-13-deoxyepirubicin, 2-pyrrolino-13-deoxycarminomycin and 2-pyrrolino-13-deoxyidarubicin and their 5-imino analogues (R ₅ = NH).

Alkyl groups typically contain 1-6 carbon atoms and can be linear or branched. Examples of suitable alkyl groups include methyl, ethyl, isopropyl, n-propyl, butyl, n-butyl, pentyl and hexyl.

"Pharmaceutically acceptable salts" refer to derivatives of the disclosed compounds where the parent compound is modified by producing acid salts thereof. The compounds of the present invention form acid addition salts with a wide range of organic and inorganic acids and include physiologically acceptable salts that are often used in pharmaceutical chemistry. Such salts are also part of the present invention. Typical inorganic acids used to form such salts include hydrochloric, hydrobromic, hydroiodic, nitric, sulfuric, phosphoric, hypophosphoric, and the like. Salts formed from organic acids such as aliphatic mono- and dicarboxylic acids, phenyl-substituted alkonic acids, hydroxyalkanoic and hydroxyalkanedioic acids, aromatic acids, aliphatic and aromatic sulfonic acids can also be used. Such pharmaceutically acceptable salts thus include acetate, phenylacetate, trifluoroacetate, acrylate, ascorbate, benzoate, chlorobenzoate, dinitrobenzoate, hydroxybenzoate, methoxybenzoate, methyl benzoate, o-acetoxybenzoate, naphthalene-2-benzoate, bromide, isobutyrate, phenylbutyrate, β-hydroxybutyrate , butine-1,4-dioate, hexine-1,4-dioate, caprate, caprylate, chloride, cinnamate, citrate, formate, fumarate, glycolate, heptanoate, hippurate, lactate, malate, maleate, hydroxymaleate, malonate, mandelate, mesylate , nicotinate, isonicotinate, nitrate, oxalate, phthalate, teraphthalate, phosphate, monohydrogen phosphate, dihydrogen phosphate, metaphosphate, pyrophosphate, propiolate, propionate, phenylpropionate, salicylate, sebacate, succinate, suberate, sulfate, bisulfate, pyrosulfate, sulfite, bisulfite, sulfonate, benzenesulfonate , p-bromobenzenesulfonate, chlorobenzenesulfonate, ethanesulfonate, 2-hydroxyethanesulfonate, methanesulfonate, naphthalene-1-sulfonate, naphthalene-2-sulfonate, p-toluenesulfonate, xylenesulfonate, tartrate, etc.

Deuterated forms contain heavy hydrogen, including deuterium and/or tritium.

It should be understood that the compounds of the present invention refer to all optical isomers and stereoisomers at the various possible atoms in the molecule unless otherwise indicated.

The term “solvate” refers to a compound formed by the reaction of a solvent and a solute and includes hydrates. Solvates are usually crystalline solid adducts containing solvent molecules in a crystal structure in either stoichiometric or non-stoichiometric proportions.

Prodrugs of compounds containing various nitrogen-containing functional groups (amino) may include the following types of derivatives, where each R group independently may be hydrogen, substituted or unsubstituted alkyl, aryl, alkenyl, alkynyl, heterocyclic, alkylaryl, aralkyl, aralkenyl, an aralkynyl, cycloalkyl or cycloalkenyl group.

(a) Carboxamides, -NHC(O)R

(b) Carbamates, -NHC(O)OR

(c) (Acyloxy)alkylcarbamates, NHC(O)OROC(O)R

(d) Enamines, -NHCR(=CHCO2R) or -NHCR(=CHCONR2)

(e) Schiff base, -N=CR2.

The synthesis and purification of 13-deoxyanthracyclines are disclosed in US Patent Nos. 5,942,605; 5948896; and 7244829, which are incorporated herein by reference. A method for converting anthracyclines to 2-pyrrolino-anthracyclines is disclosed in Nagy et al., supra and US Pat. No. 6,184,374, which are incorporated herein by reference.

It has been found that the 13-deoxy forms of doxorubicin, daunorubicin, or other similar anthracyclines will not be metabolically converted to the cardiotoxic 13-dihydro forms and thus do not exhibit cumulative irreversible cardiotoxicity. In particular, see WO99/08687, US Patent Nos. 5,948,896 and 5,942,605 and PCT/US99/04704, the disclosure of which is incorporated herein by reference.

5-imino-13-deoxydoxorubicin was tested in Phase I clinical trials in patients with advanced tumors (Holstein, SA Investigative New Drugs. 2015, 33: 594-602). This compound has anticancer activity against various carcinomas, sarcomas and bladder cancer. In addition, the compound was well tolerated with relatively little anthracycline toxicity. Until now, it was not known whether such 13-deoxyanthracycline anticancer agents would have an enhanced anticancer effect when converted to 2-pyrrolino analogues, and whether they would simultaneously have an acceptable safety profile. As stated above, 2-pyrrolino-doxorubicin is very potent, but also toxic and lethal at or near the doses required for anticancer effectiveness.

2-Pyrrolino-13-deoxyanthracyclines of formula X demonstrate anticancer, antineoplastic and/or antineoplastic efficacy that is useful for all types of therapies for the treatment of cancer, neoplasms or tumors, including leukemia, melanoma, liver, breast, ovarian, prostate, gastric tumors , pancreas, lung, kidney, colon and central nervous system. The treatments of the present invention provide methods for inhibiting the growth of normal cells (such as psoriatic skin cells), cancers, tumors and neoplasms in mammals, including humans, using compounds of formula X.

The pharmaceutical composition of the present invention includes a compound or compounds of formula X, pharmaceutically acceptable salts thereof, deuterated forms thereof, prodrugs thereof, isomers thereof and/or solvates thereof in a pharmaceutically acceptable carrier or excipient. These pharmaceutical compositions of the present invention are useful in anticancer therapy and treatment of cell proliferative disorders. Pharmaceutically acceptable carriers or excipients are well known to those of ordinary skill in the art of formulating compounds into pharmaceutical compositions, combinations, mixtures and preparations. A pharmaceutically acceptable carrier refers to one or more compatible solid or liquid excipients, diluents or encapsulating agents that are suitable for administration to mammals, including humans. Pharmaceutical compositions, combinations, mixtures and preparations suitable for parenteral administration are prepared in sterile form, which may be a sterile solution or suspension in a suitable diluent or solvent.

The amount of active ingredient contained in the pharmaceutical composition can vary widely depending on many factors such as the route of administration and the carrier. In the present invention, the pharmaceutical composition may contain from about 0.1 to 1000 mg of 2-pyrrolino-13-deoxyanthracycline of formula X per ml or gram of pharmaceutical composition.

In the method of administration, 2-pyrrolino-13-deoxyanthracycline of formula X, pharmaceutically acceptable salts thereof, deuterated forms thereof, prodrugs thereof, isomers thereof and/or solvates thereof are administered to a patient in need thereof at a dose of about 0.1 mg/m ² body surface area to about 1000 mg/m ² body surface area, more typically from about 10 mg/m ² body surface area to about 500 mg/m ² body surface area. Doses of 2-pyrrolino-13-deoxyanthracyclines can be administered as frequently as needed. The actual route of administration will vary according to the particular dosage form, composition, combination, mixture or preparation, the particular type of cancer or cell proliferative disorder being treated, and the particular patient being treated.

The composition can be administered to a patient by any route that is medically acceptable, including oral, parenteral, topical or implantation. Oral administration includes administration of the composition in the form of tablets, capsules, lozenges, suspensions, solutions, emulsions, powders, syrups, and the like. The preferred route of administration is parenteral.

The actual route and order of administration may vary according to the particular pharmaceutical composition of the compound of formula X being used, the particular type of cancer being treated, the severity of the disease condition being treated, and the particular patient being treated. Dosage limits for administration of components may vary depending on the age, condition, gender and severity of the disease of the patient, and can be determined by one of ordinary skill in the art.

Examples of cancer that can be treated according to the present invention include breast cancer, bladder cancer, Karpi sarcoma, leukemia such as acute lymphoblastic leukemia and acute myeloblastic leukemia, Wilms tumor, neuroblastoma, lymphoma, soft tissue and bone sarcomas, cancer ovarian, bladder cancer, including transitional cell bladder cancer, thyroid cancer, gastric carcinoma, Hodgkin's disease and bronchogenic carcinoma.

To test whether 13-deoxyanthracycline could be converted to 2-pyrrolino-13-deoxyanthracycline, the method of Nagy et al., supra, was used to convert the amine in the sugar moiety to 3'-deamino-3'-(2''-pyrroline-1 ''-il) group. As an example, 5-imino-13-deoxydoxorubicinIII converted to 2-pyrrolino-5-imino-13-deoxydoxorubicinIV. CompoundIV tested in vitro And in vivo on anticancer activity and efficacy. Connection conversionIII into the connectionIV shown below:

Conversion of 13-deoxyanthracycline to 2-pyrrolino-13-deoxyanthracycline of formula X with improved activity and efficacy is shown, by way of example, in the following synthesis and biological tests, which are intended to illustrate and not limit the present invention.

Example 1

5-imino-13-deoxydoxorubicin HCl salt ( III , 62.4 mg, 0.1104 mmol) was dissolved in 10 ml of dimethylformamide (DMF) and 90 ml of dichloromethane in a flask. The flask was purged with nitrogen and then 4-iodobutyraldehyde (0.697 g, 3.535 mmol, 33 equivalents) in 10 ml of dichloromethane was added dropwise. N,N -Diisopropylethylamine (0.152 g, 205 μL, 1.178 mmol, 11 equivalents) was then added via syringe and the reaction flask was wrapped in foil and allowed to stir for 1.25 hours at room temperature. After completion of the reaction, as determined by high-performance liquid chromatography (HPLC), 20 ml of a 1% solution of trifluoroacetic acid (TFA) in methanol was added. The solution was concentrated on a rotary evaporator to ¼ of its original volume. Ether/hexane solution (1:5, 100 ml) was added and a dark purple precipitate formed. The solid was allowed to settle and the light purple stock solution was drained through the cannula. The remaining solid was washed with an additional volume of 1:5 ether/hexane and then dried overnight under high vacuum in an aluminum foil-wrapped flask. The yield of 2-pyrrolino-5-imino-13-deoxydoxorubicin IV was 61.0 mg (0.0878 mmol, 79.5%) as a purple powder that was 92% pure by HPLC.

Example 2

The growth inhibitory activities of 2-pyrrolino-5-imino-13-deoxydoxorubicin IV , 5-imino-13-deoxydoxorubicin III and doxorubicin I against several cancer cell lines were assessed in a fluorescence assay using resazurin. Growth inhibitory activity was measured as the inhibitory concentration of a compound required to inhibit cell growth by 50% (IC ₅₀ ). Cancer cells were seeded in 96-well plates (0.5-1×10 ⁴ cells/well) with appropriate media, incubated with the drug for 48 hours and then treated with resazurin (0.01% final concentration), a non-fluorescent blue dye, in 24 hours. Fluorescence (exc.560nm/em.590nm) was recorded using a BioTek SynergyHT multiwell plate fluorimeter. Healthy cells metabolize resazurin to resorufin, a fluorescent pink dye. Thus, fluorescence is an indicator of cell viability and is used to determine the effect of cytotoxic agents on cell cultures. The results of cytotoxic activity assays in five human cancer cell lines and one normal human cell line are presented in Table 1. Against all cancer cells, 2-pyrrolino-5-imino-13-deoxydoxorubicin IV was significantly more effective than the parent compound 5-imino- 13-deoxydoxorubicin III , with IC ₅₀ values that were 13-27 times lower than those of 5-imino-13-deoxydoxorubicin III . The activity of 2-pyrrolino-5-imino-13-deoxydoxorubicin IV was also higher than that of doxorubicin I , with IC ₅₀ values that were on average lower than those of doxorubicin I.

The compounds were also tested against a normal non-cancerous cell line, normal human fibroblasts (HADF). 2-pyrrolino-5-imino-13-deoxydoxorubicin IV had half the cytotoxicity (39.75/18.9) of doxorubicin I against HADF (normal) cells. The results of these studies are shown in Table 1.

The compounds were also tested against the doxorubicin-sensitive uterine fibrosarcoma cell line, MES-SA (Compound I ), and against the doxorubicin-resistant uterine fibrosarcoma cell line, MES-SA/MX2. These results are shown in Table 2.

Table 1
Antiproliferative activity (IC ₅₀ ) of Compound IV , Compound III and Compound I against human cancer cell lines and a normal cell line determined using a resazurin assay. IC ₅₀ values are the mean (±SEM) of 3 separate assays, each performed in triplicate. Cell line Cell type Drug, IC ₅₀ values (µM) Compound IV Compound III Compound I SW-872 Liposarcoma 0.57 ± 0.23 7.38 ± 0.45 33.7 ± 7.5 R.D. Rhabdomyosarcoma 0.44 ± 0.13 11.89 ± 3.99 0.43 ± 0.15 GCT Histiocytoma 0.175 ± 0.015 3.37 ± 0.35 0.279 ± 0.036 HeLa Cervical adenocarcinoma 0.467 ± 0.104 9.82 ± 0.93 0.605 ± 0.095 T47D Ductal carcinoma of the breast 0.817 ± 0.561 18 ± 13.7 10.3 ± 7.8 Average value 0.49 10.09 9.06 HADF Human mature dermal fibroblasts 39.75 58.9 18.9

Compound IV was shown to be on average 20 (10.09/0.49) times more effective at inhibiting cell growth of cancer cells than Compound III, and 18 (9.06/0.49) times more effective than Compound III. than compound I. Compound IV was on average 81 (39.75/0.49) times more active against cancer cells than against normal non-cancerous cell lines. Compound III is on average only 5.8 (58.9/10.09) times more active against cancer cells than against normal cells, and compound I is only 2 (18.9/9.06) times more active. Thus, the selectivity of compound IV for cancer cells compared to normal cells was shown to be 14 (81/5.8) times greater than compound III and 40 (81/2) times greater than compound I . This result was unexpected and is in apparent contrast to what was reported for compound II (2-pyrrolinodoxorubicin), which had little or no selectivity for cancer cells compared to normal cells. Even more surprising and unpredictable was that the selectivity of Compound IV for cancer cells over normal cells was shown to be 40 times greater than Compound I (doxorubicin). Thus, the formation of the 2-pyrrolino analogue of 5-imino-13-deoxydoxorubicin ( III ) produced a more effective and safer anthracycline compared to the formation of the 2-pyrrolino analogue of doxorubicin ( I ), which produced a highly active toxin II without any therapeutic benefit.

table 2
Antiproliferative activity (IC ₅₀ ) of Compound IV , Compound III and Compound I against human cancer cell lines sensitive and resistant to doxorubicin. IC ₅₀ values are the mean (±SEM) of 3 separate assays, each performed in triplicate. Cell line Cell type Drug, IC ₅₀ values (µM) Compound IV Compound III Compound I MES-SA Uterine fibrosarcoma (sensitive) 0.76 ± 0.16 10.5 ± 1.0 0.55 ± 0.08 MES-SA/MX2 Uterine fibrosarcoma (resistant) 0.66 ± 0.003 90 7.8 ± 1.8

Compound IV is more potent than Compound III and is approximately equal in potency to Compound I in a doxorubicin-sensitive fibrosarcoma strain. In the resistant fibrosarcoma strain, Compound I was 14 (7.8/0.55) times less potent than in the sensitive fibrosarcoma strain, as expected. On the other hand, Compound IV is approximately as effective in a resistant strain of fibrosarcoma as in a sensitive strain of fibrosarcoma. Thus, Compound IV is 12 times more effective than Compound I in an anthracycline-resistant cell line, indicating that when cancer is treated, cancer tissue will not become resistant to Compound IV . This distinctive property is a significant improvement in compound IV compared to compounds III and I.

Example 3

To evaluate in vivo drug activity on primary tumor growth in a sarcoma model, an experiment was performed using commercially available human fibrosarcoma HT1080 cells engineered to express luciferase (HT1080-luc; PerkinElmer). The inhibitory activity of Compound III and Compound IV against HT1080-luc was measured in vitro by the methods described in Example 2. The IC ₅₀ ± SEM (μM) value for Compound IV was 3.51 ± 1.79 and for Compound III was 10.0 ± 6.1. HT1080-luc tumors were stimulated in 6-week-old female SCID mice (Harlan) by orthotopic subcutaneous injection of 1 x 10 ⁶ cells into each flank. After tumors had been established for 6 days, mice were randomized to one of three treatment groups and treated three times a week with an intraperitoneal injection of Compound IV or Compound III , 2.4 mg/kg in saline buffered saline (PBS), or a vehicle injection (PBS). . The results are shown in Table 3. Based on in vivo bioluminescence imaging results, tumor progression was reduced in a statistically significant manner at day 20 in mice treated with Compound IV compared to mice treated with PBS alone. Tumor progression was not significantly reduced by Compound III . At day 20, the reduction in tumor weight was 53% with Compound IV , whereas it was only 36% with Compound III at the same given dose. These results indicate increased efficacy of Compound IV compared to Compound III at a given dose and are consistent with the in vitro results presented above. Compound IV was well tolerated in mice without deaths or overt signs of toxicity such as ptosis, lordosis, lethargy, and weight loss. This is in contrast to 2-pyrrolinodoxorubicin, which can be lethal at the effective dose range.

Table 3
Average tumor weight at day 20. An asterisk (*) indicates a significant decrease compared to PBS-treated controls (p < 0.05). Values are means + standard error of the mean. Treatment: Average tumor weight (g) S.E.M. PBS 2.18 ±0.48 Compound III (2.4 mg/kg) 1.40 ±0.30 Compound IV (2.4 mg/kg) 1.03* ±0.21

The anti-cell proliferative therapeutic effects of 2-pyrrolino-13-deoxyanthracyclines of formula X were significantly enhanced when combined with 13-deoxyanthracyclines of formula Y. This enhanced therapeutic effect is the result of an unexpected synergy between 2-pyrrolino-13-deoxyanthracyclines and 13-deoxyanthracyclines. The 13-deoxyanthracycline compounds used in accordance with the present invention have the following formula Y :

wherein R ₁ , R ₂ and R ₃ are each independently H or OH; R ₄ is selected from the group consisting of H, OH, alkyl and O-alkyl; R ₅ represents O or NH; and R ₆ is a sugar containing a primary amino group or not containing an amino group; their pharmaceutically acceptable salts, their deuterated forms, their prodrugs, their isomers and/or their solvates. More typical 13-deoxyanthracyclines are 13-deoxydoxorubicin, 13-deoxydaunorubicin, 13-deoxyepirubicin, 13-deoxycarminomycin, 13-deoxyidarubicin, 13-deoxyannamycin and 13-deoxyamrubicin and their 5-imino analogues ( _R5 = NH).

The alkyl group in the above formula Y is the same as disclosed above.

In addition, the treatment of the present invention using a combination of 2-pyrrolino-13-deoxyanthracyclines and 13-deoxyanthracyclines demonstrates anticancer, antitumor and/or antineoplastic efficacy, which is useful for all types of therapy for the treatment of cell proliferative disorders such as inflammation, psoriasis, cancer, neoplasms or tumors, including leukemia, melanoma, tumors of the liver, breast, ovary, prostate, stomach, pancreas, lung, kidney, colon and central nervous system. The treatment of the present invention provides a method of inhibiting the growth of cancers, tumors, neoplasms and normal cells in mammals, including humans.

A typical composition, combination, mixture or preparation comprising the components of the present disclosure is a compound of formula X , a pharmaceutically acceptable salt thereof, a deuterated form thereof, a prodrug thereof, an isomer thereof and/or a solvate thereof in combination with a compound of formula Y ; a pharmaceutically acceptable salt thereof, a deuterated form thereof, a prodrug thereof, an isomer thereof and/or a solvate thereof. A more typical composition, combination, mixture or preparation is 2-pyrrolino-13-deoxyanthracycline selected from the group consisting of 2-pyrrolino-13-deoxydoxorubicin, 2-pyrrolino-13-deoxydaunorubicin, 2-pyrrolino-13-deoxyepirubicin, 2 -pyrrolino-13-deoxycarminomycin and 2-pyrrolino-13-deoxyidarubicin and their 5-imino analogues; a pharmaceutically acceptable salt thereof, a deuterated form thereof, a prodrug thereof, an isomer thereof and/or a solvate thereof, and a 13-deoxyanthracycline selected from the group consisting of 13-deoxydoxorubicin, 13-deoxydaunorubicin, 13-deoxyepirubicin, 13-deoxyidarubicin, 13-deoxyannamycin , 13-deoxycarminomycin, 13-deoxyamrubicin and their 5-imino analogues; a pharmaceutically acceptable salt thereof, a deuterated form thereof, a prodrug thereof, an isomer thereof and/or a solvate thereof.

The components of the composition, combination, mixture or preparation can be administered to a patient simultaneously, separately, sequentially or one after the other. The components can be administered to a patient by any suitable route that is medically acceptable, including oral, parenteral, topical or implantation. Oral administration includes the administration of components of compositions, combinations, mixtures or preparations in the form of tablets, capsules, lozenges, suspensions, solutions, emulsions, powders, syrups and the like. The preferred route of administration is parenteral.

The actual route and order of administration of the components may vary according to the particular 2-pyrrolino-13-deoxyanthracycline pharmaceutical composition of formula X and the particular 13-deoxyanthracycline pharmaceutical composition of formula Y being used, the particular cancer or cell proliferative disorder being treated, the severity of the disease state , to be treated, and the specific patient being treated. Dosage limits for administration of components may vary depending on the age, condition, gender and severity of the disease of the patient, and can be determined by one of ordinary skill in the art.

The pharmaceutical composition of the present invention includes 2-pyrrolino-13-deoxyanthracycline of formula X; a pharmaceutically acceptable salt thereof, a deuterated form thereof, a prodrug thereof, an isomer thereof and/or a solvate thereof mixed together with a 13-deoxy anthracycline of formula Y; a pharmaceutically acceptable salt thereof, a deuterated form thereof, a prodrug thereof, an isomer thereof and/or a solvate thereof in a pharmaceutically acceptable carrier or excipient. These pharmaceutical compositions of the present invention are useful for anticancer therapy and treatment of cell proliferative disorders.

Pharmaceutically acceptable carriers or excipients are well known to those of ordinary skill in the art of formulating compounds into pharmaceutical compositions, combinations, mixtures and preparations. A pharmaceutically acceptable carrier refers to one or more compatible solid or liquid excipients, diluents or encapsulating agents that are suitable for administration to mammals, including humans. Pharmaceutical compositions, combinations, mixtures and preparations suitable for parenteral administration are provided in sterile form, which may be a sterile solution or suspension in a suitable diluent or solvent.

The amount of active ingredient contained in the pharmaceutical composition can vary widely depending on a variety of factors such as the route of administration and the carrier. In the present invention, the pharmaceutical composition may contain from about 0.1 to 1000 mg of 2-pyrrolino-13-deoxyanthracycline of formula X , a pharmaceutically acceptable salt thereof, a deuterated form thereof, a prodrug thereof, an isomer thereof and/or a solvate thereof, and from about 0.1 up to 1000 mg of 13-deoxyanthracycline of formula Y , a pharmaceutically acceptable salt thereof, a deuterated form thereof, a prodrug thereof, an isomer thereof and/or a solvate thereof.

In the method of the present invention, 2-pyrrolino-13-deoxyanthracycline of formula X , a pharmaceutically acceptable salt thereof, a deuterated form thereof, a prodrug thereof, an isomer thereof and/or a solvate thereof is administered to a patient in need thereof at a dose of about 0.1 mg/day. m ² body surface area to about 1000 mg/m ² body surface area, more typically from about 10 mg/m ² body surface area to about 500 mg/m ² body surface area, and more typically by parenteral administration. The 13-deoxy anthracycline of formula Y , a pharmaceutically acceptable salt thereof, a deuterated form thereof, a prodrug thereof, an isomer thereof and/or a solvate thereof of the present invention is administered to a patient in need thereof at a dose of from about 0.1 mg/m ² to about 1000 mg/m ² , more typically from about 10 mg/m ² to about 500 mg/m ² , and more typically by parenteral route of administration. 2-Pyrrolino-13-deoxyanthracycline and 13-deoxyanthracycline can be administered together in one composition, combination, mixture or preparation, or they can be administered separately in any order, sequentially in any order, or one after the other in any order. If they are not administered together, the second compound is typically administered within 72 hours of administration of the first compound.

The anticancer/antiproliferative therapeutic effects of 2-pyrrolino-13-deoxyanthracycline are significantly enhanced by 13-deoxyanthracycline without increasing toxicity. Similarly, the anticancer/antiproliferative therapeutic effects of 13-deoxyanthracycline are significantly enhanced by 2-pyrrolino-13-deoxyanthracycline without increasing toxicity. This lack of increase in toxicity may be partially attributed to the synergism between the two types of compounds. Doses of 2-pyrrolino-13-deoxyanthracycline and 13-deoxyanthracycline can be administered as frequently as needed. The actual route and order of administration will vary according to the particular dosage form, composition, combination, mixture or preparation, the particular type of cancer being treated and the particular patient being treated.

The enhanced anti-cell proliferative effect of the combination of 2-pyrrolino-13-deoxyanthracycline of formula X with 13-deoxyanthracycline of formula Y of the present invention is shown, by way of example, in the following biological test, which is intended to illustrate and not limit the present invention.

Example 4

The fibrosarcoma cell line, HT1080, was used to measure IC ₅₀ values and IC ₁₀ values (the inhibitory concentration of a compound required to inhibit cell growth by 10%) of Compound IV and Compound III as described in Example 2. The IC ₅₀ concentration represents the therapeutic amount of the compound . IC ₁₀ represents the non-therapeutic sub-effective amount of the compound. Cells were passaged and seeded at a density of 5000 cells/well in 96-well plates and the plates were incubated for 24 hours. After incubation, the medium was replaced and the IC ₅₀ and IC ₁₀ values of Compound IV and Compound III were measured after 48 hours of incubation. IC ₅₀ values were then measured for compound IV in the presence of compound III at IC ₁₀ concentration and for compound III in the presence of compound IV at IC ₁₀ concentration after 48 hours of incubation. After incubation, 20 μM 0.1% AlamarBlue in 1xPBS was added to each well. The plates were then incubated for an additional 24 hours. The plates were read on a microplate reader at a/c. 485/528.

Table 4
IC ₅₀ and IC ₁₀ values (micromolar) of compound IV and compound III , IC ₅₀ of compound IV in the presence of IC ₁₀ of compound III, and IC ₅₀ of compound III in the presence of IC ₁₀ of compound IV , in HT1080 cells. Compound IV Compound III Compound IV plus IC ₁₀ III Compound III plus IC ₁₀ IV IC ₅₀ 12.53 63.7 1.01 1.78 IC ₁₀ 0.84 5.34 - -

Compound IV is 12-fold (12.5/1.01) more potent in the presence of non-therapeutic amounts of Compound III, and Compound III is 35-fold (63.7/1.78) more potent in the presence of non-therapeutic amounts of Compound IV . These results clearly indicate that compounds IV and III are synergistic with each other. This result is unexpected since both compounds are anthracyclines and are expected to act by the same mechanism and therefore should only be additive in combination. That is, one of ordinary skill in the art would expect that IC ₁₀ concentrations of one compound would have little or no effect on the activity of another compound. Thus, the combination of 2-pyrrolino-13-deoxyanthracyclines of formula X with 13-deoxyanthracyclines of formula Y provides an improved composition with increased activity and effectiveness compared to the compounds used alone.

A synergistic interaction occurs when the pharmacological effect of administering a combination of two drugs is greater than the additive effects of the two drugs administered separately. For example, assume that the dose of the first drug to produce a 30% response is 100 units, and the dose of the second drug to produce a 30% response is 10 units. If 100 units of the first drug plus 10 units of the second drug provide a 60% response, then there is an additive effect between the two drugs. However, if 100 units of the first drug plus 10 units of the second drug provide a 90% response, then there is a synergistic or superadditive effect between the two drugs. If the compounds interacted in an additive manner, the IC ₅₀ of compound IV would be expected to decrease from 12.53 μM to 10 μM in the presence of compound III at the IC ₁₀ concentration. However, the IC ₅₀ of compound IV decreased to 1 μM. Likewise, the IC ₅₀ of compound III can be expected to decrease from 63.37 μM to 50 μM in the presence of compound IV at an IC ₁₀ concentration. However, the IC ₅₀ of compound III decreased to 1.78 μM. Thus, the 2-pyrrolino-13-deoxyanthracyclines of formula X and the 13-deoxyanthracyclines of formula Y of the present invention act in a potent manner or synergistically with each other to provide highly active and effective compositions, combinations, mixtures or preparations for inhibiting the growth of cancer cells.

In accordance with the present invention, the compounds of the present invention can be used alone or in a suitable association, and can also be used in combination with pharmaceutically acceptable carriers and other pharmaceutically active compounds. The active agent may be present in the pharmaceutical composition in any suitable amount.

Pharmaceutically acceptable carriers described herein, for example, carriers, adjuvants, excipients or diluents, are well known to those skilled in the art. Generally, the pharmaceutically acceptable carrier is chemically inert to the active compounds and has no harmful side effects or toxicity under conditions of use. Pharmaceutically acceptable carriers may include polymers and polymer matrices.

The choice of carrier will be determined in part by the particular method used to administer the composition. Accordingly, there is a wide variety of suitable dosage forms for the pharmaceutical composition of the present invention. The following dosage forms for oral, aerosol, parenteral, subcutaneous, intravenous, intraarterial, intramuscular, intraperitoneal, intrathecal, rectal, and vaginal administration are illustrative only and are not in any way limiting.

Dosage forms suitable for oral administration may include (a) liquid solutions, such as an effective amount of the compound dissolved in diluents such as water, saline, or orange juice; (b) capsules, sachets, tablets, lozenges and lozenges, each containing a predetermined amount of the active ingredient in the form of solid particles or granules; (c) powders; (d) suspension in a suitable liquid; and (e) suitable emulsions. Liquid compositions may include diluents such as water, cyclodextrin, dimethyl sulfoxide and alcohols such as ethanol, benzyl alcohol, propylene glycol, glycerin and polyethylene alcohols including polyethylene glycol, with or without the addition of a pharmaceutically acceptable surfactant, suspending agent or emulsifier. The capsules may be conventional hard or soft shell gelatin capsules containing, for example, surfactants, lubricants and excipients such as lactose, sucrose, calcium phosphate and corn starch. Tablet forms may include one or more of the following: lactose, sucrose, mannitol, corn starch, potato starch, alginic acid, microcrystalline cellulose, gum acacia, gelatin, guar gum, colloidal silicon dioxide, croscarmellose sodium, talc, magnesium stearate, stearate calcium, zinc stearate, stearic acid and other excipients, coloring agents, diluents, buffering agents, disintegrants, humectants, preservatives, flavoring agents and pharmacologically compatible carriers. Lozenge forms may include the active ingredient in a flavoring agent, usually sucrose and gum acacia or tragacanth, as well as lozenges containing the active ingredient in an inert base such as gelatin and glycerin, or sucrose and gum acacia, emulsions and gels containing in addition to the active ingredient such carriers as are known in the art.

The compounds, alone or in combination with other suitable components, can be formulated into aerosol compositions for administration by inhalation. These aerosol compositions can be placed in suitable pressurized propellants such as dichlorodifluoromethane, propane and nitrogen. They may also be formulated as pharmaceutical compositions for preparations under normal pressure, such as in a nebulizer or nebulizer.

Compositions suitable for parenteral administration include aqueous and non-aqueous isotonic sterile injectable solutions, which may contain antioxidants, buffers, bacteriostats and solutes that render the composition isotonic with the blood of the intended recipient, as well as aqueous and non-aqueous sterile suspensions, which may include suspending agents. , solubilizers, thickeners, stabilizers and preservatives. The compound can be administered in a physiologically acceptable diluent in a pharmaceutical carrier, such as a sterile liquid or mixture of liquids, including water, saline, aqueous dextrose and solutions of appropriate sugars, alcohol such as ethanol, isopropanol or hexadecyl alcohol, glycols such as propylene glycol or polyethylene glycol such as poly(ethylene glycol) 400, glycerol ketals such as 2,2-dimethyl-1,3-dioxolane-4-methanol, ethers, oil, fatty acid, fatty acid ester or glyceride or acetylated fatty acid glyceride, with or without the addition of a pharmaceutically acceptable surfactant such as soap or detergent, a suspending agent such as pectin, carbomers, methylcellulose, hydroxypropylmethylcellulose or carboxymethylcellulose, or emulsifiers and other pharmaceutical adjuvants.

Oils that can be used in parenteral compositions include petroleum, animal, vegetable or synthetic oils. Specific examples of oils include peanut, soybean, sesame, cottonseed, corn, olive, petrolatum and mineral oil. Suitable fatty acids for use in parenteral compositions include oleic acid, stearic acid and isostearic acid. Ethyl oleate and isopropyl myristate are examples of suitable fatty acid esters. Suitable soaps for use in parenteral compositions include fatty acid alkali metal salts, ammonium and triethanolamine, and suitable detergents include (a) cationic detergents such as, for example, dimethyldialkylammonium halides and alkylpyridinium halides, (b) anionic detergents such as, for example , alkyl, aryl and olefin sulfonates, alkyl, olefin, ether and monoglyceride sulfates and sulfosuccinates, (c) nonionic detergents such as, for example, fatty amine oxides, fatty acid alkanolamides and polyoxyethylene-polypropylene copolymers, (d) amphoteric detergents, such as, for example, alkyl-β-aminopropionates and quaternary ammonium salts of 2-alkylimidazoline, and (e) mixtures thereof.

Parenteral compositions typically contain from about 0.5% wt. up to about 25% wt. active ingredient in solution. Suitable preservatives and buffers may be used in such compositions. To minimize or eliminate injection site irritation, such compositions may contain one or more nonionic surfactants having a hydrophilic-lipophilic balance (HLB) of about 12 to about 17. The amount of surfactant in such compositions ranges from about 5% up to about 15% by weight. Suitable surfactants include polyethylene sorbitan fatty acid esters such as sorbitan monooleate, and high molecular weight ethylene oxide hydrophobic base adducts formed by condensation of propylene oxide with propylene glycol.

Pharmaceutically acceptable excipients are also well known to those skilled in the art. The choice of excipient will be determined in part by the particular compound as well as the particular method used to administer the composition. Accordingly, there is a wide variety of suitable formulations of the pharmaceutical composition of the present invention. The following methods and excipients are merely illustrative and not limiting in any way. Pharmaceutically acceptable excipients preferably do not interfere with the action of the active ingredients and do not cause adverse side effects. Suitable carriers and excipients include solvents such as water, alcohol and propylene glycol, solid absorbents and diluents, surfactants, a suspending agent, tableting binders, lubricants, fragrances and coloring agents.

The compositions may be presented in sealed single-dose or multi-dose containers, such as ampoules and vials, and may be stored in a freeze-dried (lyophilized) state, requiring only the addition of a sterile liquid excipient, such as water for injection, immediately prior to use. Extemporaneous injection solutions and suspensions can be prepared from sterile powders, granules and tablets. The requirements for effective pharmaceutical carriers for injectable compositions are well known to those skilled in the art. See Pharmaceutics and Pharmacy Practice , J.B. Lippincott Co., Philadelphia, PA, Banker and Chalmers, Eds., 238-250 (1982) and ASHP Handbook on Injectable Drugs , Toissel, ^4th ed., 622-630 (1986).

Compositions suitable for topical use include lozenges comprising the active ingredient in a flavoring agent, typically sucrose and gum acacia or tragacanth; lozenges comprising the active ingredient in an inert base such as gelatin and glycerin, or sucrose and acacia; and mouthwashes comprising the active ingredient in a suitable liquid carrier; as well as creams, emulsions and gels containing, in addition to the active ingredient, such carriers as are known in the art.

In addition, compositions suitable for rectal administration can be presented in the form of suppositories by mixing with various bases, such as emulsifying bases or water-soluble bases. Compositions suitable for vaginal administration may be presented in the form of pessaries, tampons, creams, gels, pastes, foams or sprays containing, in addition to the active ingredient, such carriers as are known in the art as suitable.

Those skilled in the art will appreciate that suitable methods for exogenously administering a compound of the present invention to an animal are available, and while multiple routes of administration may be used to administer a particular compound, a particular route may provide a more immediate and more effective response than another route.

Exemplary embodiments of the present invention include:

Embodiment A. The compound represented by the formula:

Where:

R ₁ , R ₂ and R ₃ are each independently H or OH;

R ₄ is selected from the group consisting of H, OH, alkyl and O-alkyl;

R ₅ represents O or NH; And

R ₆ is 2-pyrrolino-daunosamine or 2-pyrrolino-epidaunosamine;

its pharmaceutically acceptable salts, its deuterated form, its prodrug, its isomer, its solvate and mixtures thereof.

Embodiment B. A compound of Embodiment A, wherein said compound is an anthracycline derivative selected from the group consisting of 2-pyrrolino-13-deoxydoxorubicin, 2-pyrrolino-13-deoxydaunorubicin, 2-pyrrolino-13-deoxyepirubicin, 2-pyrrolino -13-deoxycarminomycin and 2-pyrrolino-13-deoxyidarubicin and their 5-imino analogues.

Embodiment C. A pharmaceutical composition comprising a compound of Embodiment A or B and a pharmaceutically acceptable carrier or excipient.

Embodiment D. A method of inhibiting cell growth, comprising administering to a mammal in need thereof a therapeutically effective amount of a compound comprising:

2-pyrrolino-13-deoxyanthracycline having the formula:

Where:

R ₁ , R ₂ and R ₃ are each independently H or OH;

R ₄ is selected from the group consisting of H, OH, alkyl and O-alkyl;

R ₅ represents O or NH; And

R ₆ is 2-pyrrolino-daunosamine or 2-pyrrolino-epidaunosamine,

its pharmaceutically acceptable salt, its deuterated form, its prodrug, its isomer, its solvate and mixtures thereof.

Embodiment E. The method of embodiment D, wherein said compound is an anthracycline derivative selected from the group consisting of 2-pyrrolino-13-deoxydoxorubicin, 2-pyrrolino-13-deoxydaunorubicin, 2-pyrrolino-13-deoxyepirubicin, 2-pyrrolino -13-deoxycarminomycin and 2-pyrrolino-13-deoxyidarubicin and their 5-imino analogues.

Embodiment F. A therapeutically effective amount of a synergistic combination drug comprising a first compound and a second compound, wherein: the first compound is a 2-pyrrolino-13-deoxyanthracycline having the formula:

Where:

R ₁ , R ₂ and R ₃ are each independently H or OH;

R ₄ is selected from the group consisting of H, OH, alkyl and O-alkyl;

R ₅ represents O or NH; And

R ₆ is 2-pyrrolino-daunosamine or 2-pyrrolino-epidaunosamine; a pharmaceutically acceptable salt thereof, a deuterated form thereof, a prodrug thereof, an isomer thereof, a solvate thereof and mixtures thereof; And

the second compound is a 13-deoxyanthracycline having the formula:

Where

each of R ₁ , R ₂ and R ₃ is independently selected from the group consisting of H and OH;

R ₄ is selected from the group consisting of H, OH, alkyl and O-alkyl;

R ₅ is selected from the group consisting of O and NH; And

R ₆ represents a sugar component; a pharmaceutically acceptable salt thereof, a deuterated form thereof, a prodrug thereof, an isomer thereof, a solvate thereof and mixtures thereof; And

wherein said therapeutically effective amount inhibits cell growth.

Embodiment G. The synergistic combination drug of embodiment F, wherein said first compound is an anthracycline derivative selected from the group consisting of 2-pyrrolino-13-deoxydoxorubicin, 2-pyrrolino-13-deoxydaunorubicin, 2-pyrrolino-13-deoxyepirubicin, 2-pyrrolino-13-deoxycarminomycin and 2-pyrrolino-13-deoxyidarubicin and their 5-imino analogues, and said second compound is an anthracycline derivative selected from the group consisting of 13-deoxydoxorubicin, 13-deoxydaunorubicin, 13-deoxyepirubicin, 13 -deoxycarminomycin, 13-deoxyidarubicin, 13-deoxyannamycin and 13-deoxyamrubicin and their 5-imino analogues.

Embodiment H. A pharmaceutical composition comprising a synergistic combination drug of Embodiment F or G and a pharmaceutically acceptable carrier or excipient.

Embodiment I. A method of inhibiting cell growth using a synergistic combination drug containing a first compound and a second compound, comprising administering to a mammal in need thereof a therapeutically effective amount of said synergistic combination drug, wherein said first compound is 2-pyrrolino-13-deoxyanthracycline , having the formula:

Where:

R ₁ , R ₂ and R ₃ are each independently H or OH;

R ₄ is selected from the group consisting of H, OH, alkyl and O-alkyl;

R ₅ represents O or NH; And

R ₆ is 2-pyrrolino-daunosamine or 2-pyrrolino-epidaunosamine; its pharmaceutically acceptable salt, its deuterated forms, its prodrug, its isomer, its solvate and mixtures thereof; And

said second compound is a 13-deoxyanthracycline having the formula:

Where

each of R ₁ , R ₂ and R ₃ is independently selected from the group consisting of H and OH;

R ₄ is selected from the group consisting of H, OH, alkyl and O-alkyl;

R ₅ is selected from the group consisting of O and NH; And

R ₆ represents a sugar component; a pharmaceutically acceptable salt thereof, a deuterated form thereof, a prodrug thereof, an isomer thereof, a solvate thereof and mixtures thereof; And

wherein said therapeutically effective amount inhibits cell growth.

Embodiment J. The method of embodiment I, wherein said first compound is an anthracycline derivative selected from the group consisting of 2-pyrrolino-13-deoxydoxorubicin, 2-pyrrolino-13-deoxydaunorubicin, 2-pyrrolino-13-deoxyepirubicin, 2- pyrrolino-13-deoxycarminomycin and 2-pyrrolino-13-deoxyidarubicin and their 5-imino analogues, and the second compound is an anthracycline derivative selected from the group consisting of 13-deoxydoxorubicin, 13-deoxydaunorubicin, 13-deoxyepirubicin, 13-deoxycarminomycin, 13-deoxyidarubicin, 13-deoxyannamycin and 13-deoxyamrubicin and their 5-imino analogues.

Embodiment K. A synergistic combination drug of embodiment F or G, wherein said second compound is present in an amount that is less than a therapeutically effective amount thereof.

Embodiment L. The synergistic combination drug of Embodiment K, wherein the weight ratio of the second compound to the first component is from about 0.005 to about 0.10:1.

Embodiment M. The synergistic combination drug of Embodiment K, wherein the weight ratio of the second compound to the first component is from about 0.01 to about 0.10:1.

Embodiment N. A synergistic combination drug of embodiment F or G, wherein the first compound is present in an amount that is less than a therapeutically effective amount thereof.

Embodiment O. The synergistic combination formulation of Embodiment N, wherein the weight ratio of the first compound to the second component is from about 0.005 to about 0.10:1.

Embodiment P. The synergistic combination drug of Embodiment N, wherein the weight ratio of the first compound to the second component is from about 0.01 to about 0.10:1.

Embodiment Q. The method of Embodiment I or J, wherein said second compound is administered in an amount that is less than a therapeutically effective amount thereof.

Embodiment R. The method of embodiment Q, wherein the weight ratio of the second compound to the first component is from about 0.005 to about 0.10:1.

Embodiment S. The method of embodiment Q, wherein the weight ratio of the second compound to the first component is from about 0.01 to about 0.10:1.

Embodiment T. The method of Embodiment I or J, wherein said second compound is administered in an amount that is less than a therapeutically effective amount thereof.

Embodiment U. The method of embodiment T, wherein the weight ratio of the first compound to the second component is from about 0.005 to about 0.10:1.

Embodiment V. The method of Embodiment Q, wherein the weight ratio of the first compound to the second component is from about 0.01 to about 0.10:1.

The term “including” (and its grammatical variations) in the context of this application is used in the inclusive sense of “comprising” or “including”, and not in the exclusive sense of “consisting only of”. The terms defined by the articles “a”, “an” and “the”, as used herein, are to be understood to cover both the plural and singular unless otherwise indicated.

The above description illustrates and describes the present invention. Moreover, the disclosure presents and describes only the preferred embodiments, but, as stated above, it is to be understood that the preferred embodiments may be formed in various other combinations, modifications and conditions, and may be changed or modified within the scope of the inventive concepts presented in this application, in accordance with the above instructions and/or skill or knowledge in the relevant field of technology. The embodiments described above are also intended to explain the best methods known to the applicant and to enable others skilled in the art to use the invention in such or other embodiments and with various modifications necessary for its particular applications or uses. Accordingly, the description is not intended to limit the invention to the form disclosed herein. It is also intended that the appended claims be construed to include alternative embodiments.

All publications and patent applications cited herein are incorporated herein by reference and for any and all purposes as if each individual publication or patent application had been specifically and individually identified as being incorporated by reference. In the event of any inconsistency between this disclosure and any publications or patent applications incorporated by reference herein, the present disclosure controls.

Claims (62)

1. The compound represented by the formula where R ₁ represents H; R ₂ and R ₃ represent OH; R ₄ represents O-C _1-6 alkyl; R ₅ represents NH; And R ₆ is 2-pyrrolino-daunosamine or 2-pyrrolino-epidaunosamine. 2. A compound according to claim 1, wherein said compound is a 5-imino analogue of a 2-pyrrolino-13-deoxyepirubicin derivative. 3. A pharmaceutical composition for inhibiting the growth of fibrosarcoma cells, containing an effective amount of a compound according to claim 1 and a pharmaceutically acceptable carrier or excipient. 4. A pharmaceutical composition for inhibiting the growth of fibrosarcoma cells, containing an effective amount of a compound according to claim 2 and a pharmaceutically acceptable carrier or excipient. 5. A method of inhibiting the growth of fibrosarcoma cells, comprising administering to a mammal in need thereof a therapeutically effective amount of a compound comprising: 2-pyrrolino-13-deoxyanthracycline, wherein said 2-pyrrolino-13-deoxyanthracycline is a compound of the formula where R ₁ represents H; R ₂ and R ₃ represent OH; R ₄ represents O-C _1-6 alkyl; R ₅ represents NH; And R ₆ is 2-pyrrolino-daunosamine or 2-pyrrolino-epidaunosamine. 6. The method according to claim 5, where the specified compound is a 5-imino analogue of a 2-pyrrolino-13-deoxyepirubicin derivative. 7. A synergistic combination drug in a therapeutically effective amount for inhibiting the growth of fibrosarcoma cells, comprising a first compound and a second compound, wherein the first compound is 2-pyrrolino-13-deoxyanthracycline having the formula , where R ₁ represents H; R ₂ and R ₃ represent OH; R ₄ represents O-C _1-6 alkyl; R ₅ represents NH; And R ₆ is 2-pyrrolino-daunosamine or 2-pyrrolino-epidaunosamine; And the second compound is a 13-deoxyanthracycline having the formula , where R ₁ represents H; R ₂ and R ₃ represent OH; R ₄ represents O-C _1-6 alkyl; R ₅ represents NH; And R ₆ represents ; And wherein said therapeutically effective amount inhibits cell growth. 8. The synergistic combination drug according to claim 7, wherein said first compound is a 5-imino analogue of a 2-pyrrolino-13-deoxyepirubicin derivative, and said second compound is a 5-imino analogue of a 13-deoxyepirubicin derivative. 9. A pharmaceutical composition for inhibiting the growth of fibrosarcoma cells, containing an effective amount of the synergistic combination drug according to claim 7 and a pharmaceutically acceptable carrier or excipient. 10. A pharmaceutical composition for inhibiting the growth of fibrosarcoma cells, containing an effective amount of the synergistic combination drug according to claim 8 and a pharmaceutically acceptable carrier or excipient. 11. A method of inhibiting the growth of fibrosarcoma cells using a synergistic combination drug containing a first compound and a second compound, comprising administering to a mammal in need thereof a therapeutically effective amount of said synergistic combination drug, wherein said first compound is 2-pyrrolino-13-deoxyanthracycline, having a formula , where R ₁ represents H; R ₂ and R ₃ represent OH; R ₄ represents O-C _1-6 alkyl; R ₅ represents NH; And R ₆ is 2-pyrrolino-daunosamine or 2-pyrrolino-epidaunosamine; And said second compound is a 13-deoxyanthracycline having the formula Where R ₁ represents H; R ₂ and R ₃ represent OH; R ₄ represents O-C _1-6 alkyl; R ₅ represents NH; And R ₆ represents ; And wherein said therapeutically effective amount inhibits cell growth. 12. The method of claim 11, wherein said first compound is a 5-imino analogue of a 2-pyrrolino-13-deoxyepirubicin derivative, and the second compound is a 5-imino analogue of a 13-deoxyepirubicin derivative. 13. The synergistic combination drug of claim 7, wherein said second compound is present in an amount that is less than a therapeutically effective amount thereof. 14. The synergistic combination drug according to claim 13, where the mass ratio of the second compound to the first compound is from 0.005 to 0.10:1. 15. The synergistic combination drug of claim 7, wherein said first compound is present in an amount that is less than a therapeutically effective amount thereof. 16. The synergistic combination drug according to claim 15, where the mass ratio of the first compound to the second compound is from 0.005 to 0.10:1. 17. The method of claim 11, wherein said second compound is administered in an amount that is less than a therapeutically effective amount thereof. 18. The method according to claim 17, where the mass ratio of the second compound to the first compound is from 0.005 to 0.10:1. 19. The method of claim 11, wherein said second compound is administered in an amount that is less than a therapeutically effective amount thereof. 20. The method according to claim 19, where the mass ratio of the first compound to the second compound is from 0.005 to 0.10:1.