Methylenedioxybenzo [I] phenanthridine derivatives used to treat cancer

The invention provides compounds of formula I: wherein A, B, X, and Y have any of the values defined in the specification, as well as pharmaceutical compositions comprising such compounds, processes for preparing such compounds, and therapeutic methods for treating cancer and other topoisomerase mediated conditions.

BACKGROUND OF THE INVENTION

Topoisomerases are ubiquitous enzymes that participate in processes such as DNA replication, repair, transcription, and recombination as well as chromosome condensation and segregation. Topoisomerase I (TOP1) is the target of several antitumor agents based upon their ability to stabilize the enzyme-DNA cleavage complex, which results in DNA damage and ultimately cell death. Camptothecin (CPT) was the first compound identified as a TOP1-targeting agent (Hsaing, Y. H.; Hertsberg, R.; Hecht, S.; Liu, L. F. Camptothecin Induced Protein-Linked DNA Breaks Via Mammalian DNA Topoisomerase I,J. Biol. Chem.,1985, 260, 14873-14878). Two clinical TOP1-targeting agents, topotecan (Hycamtin®) and irinotecan (CPT-11/Camptosar®) have since been developed. The improved water-solubility of topotecan and irinotecan relative to CPT was critical to their development into the clinic. These agents have incorporated, within their structure, the core structure of camptothecin, which includes a δ-lactone. This lactone moiety is susceptible to hydrolysis and the resulting carboxylic acid has a high affinity for human serum albumin. In addition, it is known that both of these clinical agents are susceptible to transporter-mediated cellular efflux, which can limit intracellular accumulation and has been associated with multidrug resistance. Specifically overexpression of MDR1 (P-glycoprotein) and breast cancer resistance protein (BCRP) have been associated with resistance to these camptothecins.

Additional topoisomerase targeting agents with anticancer properties include those described by LaVoie et al. in U.S. Pat. No. 7,208,492. Particular compounds discussed include compound 206 and compound 216.

These are the compounds of formula II and formula I respectfully, as described in U.S. Pat. No. 7,208,492.

Despite these previous reports there is currently a need for additional agents that are useful for treating cancer. There is also a need for anticancer agents, particularly topoisomerase I targeting agents that have enhanced cytotoxicity or enhanced metabolic stability, prolonged half-lives or improved oral bioavailability in mammals, or for topoisomerase I targeting agents that are not substrates for an efflux transporter or that have a diminished ability to be removed from a cell by an efflux transporter.

SUMMARY OF THE INVENTION

The present invention provides compounds that show inhibitory activity against topoisomerase I and/or topoisomerase II and compounds that are effective cytotoxic agents against cancer cells including drug-resistant cancer cells. The compounds of the invention are based on a benzo[i]phenanthridine core with a carboxamide moiety comprising an alkyl group with a pendant amino group for which the methylene adjacent to the amino group is disubstituted. Applicant has discovered that disubstitution of this methylene adjacent to the amino group provides compounds with significantly enhanced cytotoxicity relative to the compounds for which the methylene is unsubstituted. Representative compounds of the invention were found not to be substrates of BCRP.

Accordingly there is provided a compound of the invention which is a compound of formula I:

one of A and B is —C(O)NH(CR6R7)nCR1R2NRaRband the other is H;

R1and R2are each independently (C1-C3) alkyl; or R1and R2together with the carbon to which they are attached form a 3-6 membered cycloalkyl;

Raand Rbare each independently H or (C1-C3) alkyl wherein (C1-C3) alkyl may be optionally substituted with aryl or heteroaryl; or Raand Rbtogether with the nitrogen to which they are attached form a piperazino, pyrrolidino or piperidino;

for each CR6R7; R6and R7are each independently H or CH3;

n is 1, 2, or 3;

X is —OCH3and Y is —OR3; or Y is —OCH3and X is OR3;

Rcand Rdare each independently H, aryl, heteroaryl, aryl(alkyl), heteroaryl(alkyl), or (C1-C6)alkyl; or Rcand Rdtogether with the nitrogen to which they are attached form a piperazino, pyrrolidino or piperidino;

or a salt or prodrug thereof.

The invention also provides a pharmaceutical composition comprising a compound of formula I, or a pharmaceutically acceptable salt or prodrug thereof, in combination with a pharmaceutically acceptable excipient, diluent or carrier.

The invention also provides a method for modulating topoisomerase activity in a mammal in need of such treatment comprising administering to the mammal (e.g. a human), a compound of formula I, or a pharmaceutically acceptable salt or prodrug thereof, effective to provide a topoisomerase modulating effect.

The invention also provides a method comprising inhibiting cancer (e.g. leukemia, non-small lung cancer, colon cancer, CNS cancer, melanoma, ovarian cancer, renal cancer, prostate cancer or breast cancer) cell growth by contacting said cancer cell in vitro or in vivo with an amount of a compound of formula I, or a pharmaceutically acceptable salt or prodrug thereof, effective to inhibit the growth of said cancer cell.

The invention also provides a method for treating cancer (e.g. leukemia, non-small lung cancer, colon cancer, CNS cancer, melanoma, ovarian cancer, renal cancer, prostate cancer or breast cancer) in a mammal (e.g. a human), comprising administering a compound of formula I, or a pharmaceutically acceptable salt or produg thereof, to the mammal.

The invention also provides a compound of formula I or a pharmaceutically acceptable salt or prodrug thereof for use in medical therapy (e.g. for use in treating cancer such as leukemia, non-small lung cancer, colon cancer, CNS cancer, melanoma, ovarian cancer, renal cancer, prostate cancer or breast cancer).

The invention also provides for the use of a compound of formula I or a pharmaceutically acceptable salt or produg thereof for the manufacture of a medicament useful for the treatment of cancer (e.g. leukemia, non-small lung cancer, colon cancer, CNS cancer, melanoma, ovarian cancer, renal cancer, prostate cancer or breast cancer) in a mammal (e.g. a human).

The invention provides a compound of formula I or a pharmaceutically acceptable salt or prodrug thereof for use in the prophylactic or therapeutic treatment of cancer (e.g. leukemia, non-small lung cancer, colon cancer, CNS cancer, melanoma, ovarian cancer, renal cancer, prostate cancer or breast cancer) in a mammal (e.g. a human).

The invention also provides processes and intermediates disclosed herein that are useful for preparing compounds of formula I or salts thereof.

DETAILED DESCRIPTION

The following definitions are used, unless otherwise described: halo is fluoro, chloro, bromo, or iodo. Alkyl, alkoxy, alkenyl, alkynyl, etc. denote both straight and branched groups; but reference to an individual radical such as propyl embraces only the straight chain radical, a branched chain isomer such as isopropyl being specifically referred to. Aryl denotes a phenyl radical or an ortho-fused bicyclic carbocyclic radical having about nine to ten ring atoms in which at least one ring is aromatic. Heteroaryl encompasses a radical of a monocyclic aromatic ring containing five or six ring atoms consisting of carbon and one to four heteroatoms each selected from the group consisting of non-peroxide oxygen, sulfur, and N(X) wherein X is absent or is H, O, (C1-C4)alkyl, phenyl or benzyl, as well as a radical of an ortho-fused bicyclic heterocycle of about eight to ten ring atoms comprising one to four heteroatoms each selected from the group consisting of non-peroxide oxygen, sulfur, and N(X).

Specific values listed below for radicals, substituents, and ranges, are for illustration only; they do not exclude other defined values or other values within defined ranges for the radicals and substituents.

Specifically, (C1-C3)alkyl can be methyl, ethyl, propyl or isopropyl; (C1-C6)alkyl can be methyl, ethyl, propyl, isopropyl, butyl, iso-butyl, sec-butyl, pentyl, 3-pentyl, or hexyl; (C3-C6)cycloalkyl can be cyclopropyl, cyclobutyl, cyclopentyl, or cyclohexyl. As used herein the term “aryl(C1-C6) alkyl” refers to a (C1-C6) alkyl radical in which one or more of the hydrogen atoms of the (C1-C6) alkyl radical is replaced with an aryl radical. As used herein the term “heteroaryl(C1-C6) alkyl” refers to a (C1-C6) alkyl radical in which one or more of the hydrogen atoms of the (C1-C6) alkyl radical is replaced with a heteroaryl radical.

A specific group of compounds of formula I are compounds wherein A is —C(O)NH(CR6R7)nCR1R2NRaRband B is H.

A specific group of compounds of formula I are compounds wherein B is —C(O)NH(CR6R7)nCR1R2NRaRband A is H.

A specific value for n is 1 or 2.

A specific value for n is 1.

A specific value for CR6R7is CH2.

A specific value for R1is (C1-C3)alkyl.

A specific value for R2is (C1-C3)alkyl.

A specific group of compounds of formula I are compounds wherein R1and R2are each independently (C1-C3)alkyl.

A specific value for R1is methyl.

A specific value for R2is methyl.

A specific group of compounds of formula I are compounds wherein R1and R2are each methyl.

A specific value for Rais H or (C1-C3)alkyl wherein (C1-C3) alkyl may be optionally substituted with aryl or heteroaryl.

A specific value for Rbis H or (C1-C3)alkyl wherein (C1-C3) alkyl may be optionally substituted with aryl or heteroaryl.

A specific group of compounds of formula I are compounds wherein Raand Rbare each independently H or (C1-C3)alkyl.

A specific group of compounds of formula I are compounds wherein Raand Rbare each independently (C1-C3)alkyl.

A specific group of compounds of formula I are compounds wherein Raand Rbare each methyl.

A specific group of compounds of formula I are compounds wherein X is —OCH3and Y is —OR3.

A specific group of compounds of formula I are compounds wherein Y is —OCH3and X is —OR3.

A specific value for R3is —C(O)R4, —C(O)OR5or —C(O)NRcRd.

A specific value for R3is H.

A specific value for R3is CH3.

A specific value for R4is (C1-C6)alkyl.

A specific value for R5is (C1-C6)alkyl.

A specific value for Rcis H or (C1-C6)alkyl.

A specific value for Rdis H or (C1-C6)alkyl.

A specific compound of formula I is the compound 2,3-dimethoxy-N-(2-(dimethylamino)-2-methylpropyl)-8,9-methylenedioxybenzo[i]phenanthridine-12-carboxamide; or N-(2-(dibenzylamino)-2-methylpropyl)-2,3-dimethoxy-8,9-methylenedioxybenzo[i]phenanthridine-12-carboxamide; or N-(2-amino-2-methylpropyl)-2,3-dimethoxy-8,9-methylenedioxybenzo[i]phenanthridine-12-carboxamide; or 2,3-dimethoxy-8,9-methylenedioxybenzo[i]phenanthridine-11-carboxylic acid 2-(dimethylamino)-2-methylpropylamide; or 2,3-dimethoxy-N-(3-(dimethylamino)-3-methylbutyl)-8,9-methylenedioxybenzo[i]phenanthridine-12-carboxamide; or 2,3-dimethoxy-N-(4-(dimethylamino)-4-methylpentyl)-8,9-methylenedioxybenzo[i]phenanthridine-12-carboxamide; 2,3-dimethoxy-8,9-methylenedioxybenzo[i]phenanthridine-11-carboxylic acid 3-(dimethylamino)-3-methylbutylamide; 2,3-dimethoxy-8,9-methylenedioxybenzo[i]phenanthridine-11-carboxylic acid 4-(dimethylamino)-4-methylpentylamide or a salt or prodrug thereof.

The term “prodrug” as used herein refers to any compound that when administered to a biological system generates the drug substance, i.e. active ingredient, as a result of spontaneous chemical reaction(s), enzyme catalyzed chemical reaction(s), photolysis, and/or metabolic chemical reaction(s) or by some other process. A prodrug is thus a modified (e.g. covalently modified) analog or latent form of a therapeutically-active compound. A prodrug may also be an active metabolite or therapeutically-active compound itself.

By way of example a prodrug may generate the active inhibitory compound during metabolism, systemically, inside a cell, by hydrolysis, enzymatic cleavage, or by some other process (Bundgaard, Hans, “Design and Application of Prodrugs” inA Textbook of Drug Design and Development(1991), P. Krogsgaard-Larsen and H. Bundgaard, Eds. Harwood Academic Publishers, pp. 113-191; Tranoyl-Opalinski, I., Fernandes, A., Thomas, M., Gesson, J.-P., and Papot, S., Anti-Cancer Agents in Med. Chem., 8 (2008) 618-637). Enzymes which are capable of an enzymatic activation mechanism with the prodrug compounds of the invention include, but are not limited to nitroreductase, proteases (e.g. serine proteases such as prostate specific antigen (PSA), amidases, esterases, microbial enzymes, phospholipases, cholinesterases, and phosphases.

Certain compounds of formula I can function as prodrugs for other compounds of formula I and are thus embodiments of the invention. For example, a compound of formula I wherein R3is —C(O)R4, —C(O)OR5, or —C(O)NRcRdcan function as a prodrug for a corresponding compound of formula I wherein R3is hydrogen.

Particularly useful prodrugs are those that are linked through a phenolic functional group. Accordingly, in one embodiment the invention provides a prodrug comprising a compound of formula I that releases a phenol of a compound of formula I. In another embodiment the invention provides prodrugs that comprise a targeting moiety (e.g. an antibody).

Processes for preparing compounds of formula I are provided as further embodiments of the invention and are illustrated by the following procedures in which the meanings of the generic radicals are as given above unless otherwise qualified.

Representative compounds of the invention were prepared as illustrated below in Schemes 1-6.

A compound of formula I wherein B is —C(O)NH(CR6R7)nCR1R2NRaRband A is hydrogen can be prepared by converting a corresponding acid of formula 2:

to the compound of formula I, for example by coupling the acid of formula 2 with an amine to provide a compound of formula I. Thus, the intermediate acid of formula 2 is useful for preparing a compound of formula I.

A compound of formula I wherein A is —C(O)NH(CR6R7)nCR1R2NRaRband B is hydrogen can be prepared by converting a corresponding acid of formula 3:

to the compound of formula I, for example by coupling the acid of formula 3 with an amine to provide a compound of formula I. Thus, the intermediate acid of formula 3 is useful for preparing a compound of formula I.

Accordingly, the invention provides a method:a) for preparing a compound of formula I wherein B is —C(O)NH(CR6R7)nCR1R2NRaRbcomprising treating a compound of formula 2 with an appropriate amine (e.g. H2N(CR6R7)nCR1R2NRaRb) to provide the compound of formula I.b) for preparing a compound of formula I wherein A is —C(O)NH(CR6R7)nCR1R2NRaRbcomprising treating a compound of formula 3 with an appropriate amine (e.g. H2N(CR6R7)nCR1R2NRaRb) to provide the compound of formula I.c) for preparing a compound of formula I comprising deprotecting a corresponding compound bearing one or more protecting groups to provide the compound of formula I.d) for preparing a salt of a compound of formula I comprising treating a compound of formula I with an acid (e.g. an organic acid or inorganic acid) or base (e.g. an alkali base or alkaline base) to provide the salt of the compound of formula I.

In cases where compounds are sufficiently basic or acidic, a salt of a compound of formula I can be useful as an intermediate for isolating or purifying a compound of formula I. Additionally, administration of a compound of formula I as a pharmaceutically acceptable acid or base salt may be appropriate. Examples of pharmaceutically acceptable salts are organic acid addition salts formed with acids which form a physiological acceptable anion, for example, tosylate, methanesulfonate, acetate, citrate, malonate, tartrate, succinate, benzoate, ascorbate, α-ketoglutarate, and α-glycerophosphate. Suitable inorganic salts may also be formed, including hydrochloride, sulfate, nitrate, bicarbonate, and carbonate salts.

In general, however, a suitable dose will be in the range of from about 0.1 to about 100 mg/kg, e.g., from about 0.5 to about 75 mg/kg of body weight per day, such as 1 to about 50 mg per kilogram body weight of the recipient per day, preferably in the range of 1 to 20 mg/kg/day.

The compound is conveniently formulated in unit dosage form; for example, containing 0.5 to 100 mg, conveniently 1 to 75 mg, most conveniently, 0.5 to 25 mg of active ingredient per unit dosage form. In one embodiment, the invention provides a composition comprising a compound of the invention formulated in such a unit dosage form.

The ability of a compound of the invention to effect topoisomerase I or II mediated DNA cleavage can be determined using pharmacological models that are well known in the art, for example, using a model like Test A described below.

Human topoisomerase I was expressed inEscherichia coliand isolated as a recombinant fusion protein using a T7 expression system as described previously (31). Plasmid YepG was also purified by the alkali lysis method followed by phenol deproteination and CsCl/ethidium isopycnic centrifugation method as described (32). The 3′ end labeling of the plasmid was accomplished by digestion with a restriction enzyme followed by end filling with Klenow polymerase as previously described (33). The cleavage assays were performed as previously reported (34,35). The drug and the DNA in presence of topoisomerase I was incubated for 30 min at room temperature. The reactions were terminated by the addition of 5 μL of 5% SDS and 1 mg/mL protein kinase K with an additional 1 h of incubation at 37° C. Samples were then alkali denatured by the addition of NaOH, EDTA, sucrose, and bromophenol blue to final concentrations of 75 mM, 2.5%, and 0.05 mg/mL, respectively, prior to loading onto a neutral agarose gel. After development of the gels, typically 24-h exposure was used to obtain autoradiograms outlining the extent of DNA fragmentation. Topoisomerase I-mediated DNA cleavage values are reported as Relative Effective Concentration (REC). The REC value reflects the concentrations relative to camptothecin, whose value is arbitrarily assumed as 0.2, that is able to produce the same 10% cleavage on the plasmid DNA in the presence of human topoisomerase I. Results for the assay for representative compounds of the invention of formula I along with comparison compounds are shown in Table 1. The data demonstrate that representative compounds of the invention target topoisomerase I.

A similar assay can be used to evaluate the ability of a compound of the invention to effect topoisomerase II mediated DNA cleavage by replacing the human topoisomerase I used in Test A with a suitable topoisomerase II.

The cytotoxic effects of a compound of the invention can be determined using pharmacological models that are well known in the art, for example, using a model like Test B described below.

Test B. Cytotoxic Assays (Cancer Cell Lines and Efflux Transporter Cell Lines)

The cytotoxicity was determined using the MTT-microtiter plate tetrazolinium cytotoxicity assay (MTA). The human lymphoblast RPMI 8402 and its camptothecin-resistant variant cell line, CPT-K5 was provided by Dr. Toshiwo Andoh (Aichi Cancer Center Research Institute, Nagoya, Japan). (36) The P388 mouse leukemia cell line and its CPT-resistant TOP1-deficient variant P388/CPT45 (37) were obtained from Michael R. Mattern and Randal K. Johnson (GlaxoSmithKline, King of Prussia, Pa.). The KB3-1 cell line and its multidrug-resistant variant KBV-1 (38) were obtained from K. V. Chin (The Cancer Institute of New Jersey, New Brunswick, N.J.). The KBH5.0 cell line as noted previously (21) was derived from KB3-1 by stepwise selection against Hoechst 33342. The cytotoxicity assay was performed using 96-well microtiter plates. Cells were grown in suspension at 37° C. in 5% CO2and maintained by regular passage in RPMI medium supplemented with 10% heat inactivated fetal bovine serum, L-glutamine (2 mM), penicillin (100 U/mL), and Streptomycin (0.1 mg/mL). For determination of IC50, cells were exposed continuously for FOUR days to varying concentrations of drug, and MTT assays were performed at the end of the fourth day. Each assay was performed with a control that did not contain any drug. All assays were performed at least twice in six replicate wells.

Experimental results from Test B for representative compounds of the invention which are compounds of formula I and comparator compounds are shown in the Table 2 below. These results demonstrate that compounds of the invention can function as cytotoxic agents against tumor cell lines. Accordingly compounds of the invention of formula I may be useful as therapeutic agents for the treatment of cancer (e.g. leukemia, non-small lung cancer, colon cancer, CNS cancer, melanoma, ovarian cancer, renal cancer, prostate cancer or breast cancer) and to treat tumors that are that are resistant to other chemotherapeutic agents.

Additionally, compounds of the invention may be useful as pharmacological tools for the further investigation of topoisomerase I function.

The ability of a compound of the invention to be actively transported can be determined using pharmacological models that are well known in the art, for example, using a model like the test described below.

The cytotoxicity of the representative compounds of the invention were also tested on cell line KB3-1 (parent cell line), KBV-1 (a variant that overexpresses efflux transporter MDR1) and KBH5.0 (a variant that overexpresses BCRP). The data is tabulated in Table 3. Differences in the relative cytotoxicity between the parent and variant cell lines may be indicative of a compound that is a substrate for an efflux transporter. These data suggest that all of the compounds tested may be substrates to varying degrees for MDR1 and that compounds 31 and 34 are not substrates for BCRP. Accordingly, compounds of the invention of formula I may be useful to treat tumors that are resistant to other anticancer agents, especially anticancer agents that are susceptible to efflux by BCRP (e.g. anthracyclines, mitoxantrone, topotecan, irinotecan, bisanthrone, doxorubicin, daunorubicin, and epirubin.

The in vivo antitumor activity of a compound of the invention can be determined using pharmacological models that are well known in the art, for example, using a model like Test C described below.

Test C. Human Tumor Xenograft Assay

Bioassays were performed using female NCR/NU NU mice of approximately 9 weeks of age as obtained from Taconic Farms, Inc. (Germantown, N.Y., USA). Mice were housed 4 per cage in laminar flow HEPA filtered microisolator caging (Allentown Caging Equipment Co., Allentown, N.J., USA). Mice were fed Purina autoclavable breeder chow #5021 and given drinking water, purified by reverse-osmosis, ad libitum. Five days after arrival within the animal facility, the mice were inoculated on the right flank with 1.5×106MDA-MB-435 tumor cells in 0.1 mL of RPMI 1640 Media by sc injection (25 gauge needle×⅝″). The MDA-MB-435 cells were grown in 75 cm2flasks using RPMI 1640 Media and 10% fetal bovine serum. Tumors were of sufficient size at 19-20 days after inoculation. Tumor-bearing mice were evenly matched in each experimental group based on tumor volume. Tumor volume was calculated by measuring the tumor with a microcaliper. The length (l) is the maximum two dimensional distance of the tumor and the width (w) is the maximum distance perpendicular to this length measured in mm. Tumor volume was calculated using the formula (l*w2)/2. Every mouse in this study was weighed individually on a daily basis. Dose adjustments for each experimental group, as indicated in Table 4, were made throughout the study based upon the effect or lack of an effect of treatment on average body weights. Tumor volume was determined for each individual mouse every other day. Compound 31, the α,α-dimethyl analog of 206, was better tolerated and significantly more effective than 206 as an antitumor agent as indicated in Table 4.

Topoisomerase inhibitors are also known to possess antibacterial, antifungal, antiprotozoal, antihelmetic, and antiviral activity. Accordingly, the topoisomerase inhibitors of the invention may also be useful as antibacterial, antifungal, antipsoritic (psoriasis) antiprotozoal, antihelmetic, or antiviral agents. In particular, compounds of the invention that demonstrate little or no activity as mammalian topoisomerase I poisons, because of the possibility of similar molecular mechanism of action, could be highly active and selective antibacterial, antifungal, antiprotozoal, antihelmetic, or antiviral agents. Thus, certain compounds of the invention may be particularly useful as systemic antibacterial, antifungal, antiprotozoal, antihelmetic, or antiviral agents in mammals. The invention also provides the use of a compound of the invention for the manufacture of a medicament useful for producing an antibacterial, antifungal, antiprotozoal, antihelmetic, or antiviral effect in a mammal.

As used herein, the term “solid mammalian tumors” include cancers of the head and neck, lung, mesothelioma, mediastinum, esophagus, stomach, pancreas, hepatobiliary system, small intestine, colon, rectum, anus, kidney, ureter, bladder, prostate, urethra, penis, testis, gynecological organs, ovarian, breast, endocrine system, skin central nervous system; sarcomas of the soft tissue and bone; and melanoma of cutaneous and intraocular origin. The term “hematological malignancies” includes childhood leukemia and lymphomas, Hodgkin's disease, lymphomas of lymphocytic and cutaneous origin, acute and chronic leukemia, plasma cell neoplasm and cancers associated with AIDS. The preferred mammalian species for treatment are humans and domesticated animals.

EXAMPLES

Synthesis of Compound 31

The required amine, 2-dimethylamino-2-methylpropylamine (7), was prepared as follows.

A solution of KCN (13 g, 200 mmol) in 100 mL water was added to a stirred, cooled suspension of dimethylamine hydrochloride (16.3 g, 200 mmol) and acetone (6.96 g, 120 mmol). The mixture was stirred overnight at room temperature and then extracted with ether (50 mL×3). The organic layer was dried over Na2SO4and then concentrated under vacuum to provide product 9.32 g in 92% yield as a colorless, water-like liquid.1H NMR (CDCl3) δ 1.42 (s, 6H), 2.36 (s, 6H);13C NMR (CDCl3) δ 26.8, 40.8, 57.2, 119.7.

To a suspension of LAH (3.8 g, 100 mmol) in 150 mL ether was added a solution of 6 (5.6 g, 50 mmol) in ether (12 mL) dropwise at −5° C. The reaction was stirred at room temperature for 5 hours and then cooled down to −5° C. 4 mL Water, 4 mL 15% NaOH and 12 mL water were added sequentially. The resulting mixture was filtered and filtrate was extracted with water, brine and dried over Na2SO4. The organic extract was concentrated under vacuum and then distilled to afford a colorless water-like liquid 5.3 g in 91% yield. bp 145-147° C.;1H NMR (CDCl3) δ 0.95 (s, 6H), 1.38 (s, 2H), 2.20 (s, 6H), 2.56 (s, 2H);13C NMR (CDCl3) δ 19.2, 37.5, 49.9, 55.8.

Synthesis of Compound 32

The required amine, 2-dibenzylamino-2-methyl-propylamine (28), was prepared as follows.

To a solution of 2-amino-2-methylpropan-1-ol (5.1 mL, 53.3 mmol) in acetone and water (4:1, 100 mL) were added benzyl bromide and potassium carbonate (14.74 g, 106.6 mmol). The resulting reaction mixture was heated to reflux for 40 h. The reaction mixture was evaporated and partitioned in dichloromethane and water. The organic layer was then washed with brine (100 mL), dried (Mg2SO4), and evaporated, yielding 15 g of 13 in 98% yield as a light yellow solid;1H NMR (CDCl3) δ 1.13 (s, 6H), 3.02 (s, 1H), 3.47 (s, 2H), 3.76 (s, 4H), 7.17-7.28 (m, 10H).

To a solution of triphenylphosphine (1.96 g, 7.5 mmol), phthalimide (1.1 g, 7.5 mmol), and 2-dibenzylamino-2-methylpropan-1-ol (1.35 g, 5.0 mmol) in anhydrous THF was added DEAD (1.3 g, 7.5 mmol) in THF dropwise as not to allow the reaction to exceed room temperature. The reaction was stirred at room temperature for 2 hours under a nitrogen atmosphere. The reaction mixture was concentrated under reduced pressure and the residue subjected to flash column chromatography using 20-30% CHCl3in hexanes to provide 1.25 g (62.5% yield) of 27.

To a solution of 2-(dibenzylamino)-2-methylpropyl)isoindoline-1,3-dione (1.25 g. 3.14 mmol) in absolute ethanol (6 ml) and benzene (4 ml) was added 0.54 ml of acetic acid (9.42 mmol) followed by 50% aqueous hydrazine (0.46 ml, 9.42 mmol) and the mixture stirred at reflux for 8 hours. The resulting solid residue was filtered and the filtrate concentrated under reduced pressure. The residue was dissolved in EtOAc and extracted twice with 1.0 N HCl. The aqueous layer was separated, made basic with 5% NaOH, and extracted with EtOAc. The organic layer was concentrated under reduce pressure and purified by flash column chromatography using a gradient from 1-2% MeOH in CHCl3to provide 474 mg (56.4% yield) of 28.

Synthesis of Compound 33

Synthesis of Compound 34

Synthesis of Compound 42

The required amine, 2-dimethylamino-2-methylbutylamine (40) was prepared as follows.

To a solution of 37 (1 g, 5.4 mmol) in DMF (10 mL) was added potassium phthalimide (2 g, 10.8 mmol) and the resulting reaction mixture was heated up to 70° C. for 24 hours. The reaction mixture was quenched by 1 mL water and concentrated to dryness. The residue was dissolved in chloroform and washed with water, extracted with 2 N HCl and then basified. The precipitate was again extracted by chloroform (100 mL×2). The concentrate of organic solution finally afforded a yellow oil 700 mg, in 50% yield. The yellow oil (700 mg, 2.7 mmol) was heated with hydrazine (0.8 mL, 16.2 mmol) in ethanol (100 ml) to 60° C. for 18 hours. The reaction mixture was cooled, filtered and the filtrate was concentrated carefully to give a colorless oil 200 mg, in 57% yield.1H NMR (CDCl3) δ 1.01 (s, 6H), 1.47 (s, br, 1H), 1.56 (t, 2H, J=8.0), 2.22 (s, 6H), 2.76 (t, 2H, J=8.0);13C NMR (CDCl3) δ 21.9, 37.1, 37.6, 42.5, 54.4. HRMS calcd for C7H18N2H: 131.1543. found 131.1549.

Synthesis of Compound 43

The required amine, 2-dimethylamino-2-methylpentylamine (41) was prepared as follows.

To a solution of KCN (1.55 g, 23.8 mmol) in acetonitrile was added 18-crown-6 (229 mg, and then compound 37 (1.3 g, 8.1 mmol) was added. The reaction mixture was heated to reflux overnight and concentrated. The residue was partitioned in chloroform and water and the organic solvent was concentrated to give a yellow oil 400 mg, in 38% yield. To a suspension of LAH (380 mg, 10 mmol) in 150 mL ether was added a solution of previously obtained oil (350 mg, 2.7 mmol) in ether (2 mL) dropwise at −5° C. The reaction was stirred at room temperature for 5 hours and then cooled down to −5° C. 0.4 mL water, 0.4 mL 15% NaOH and 1.2 mL water were added sequentially. The resulting mixture was filtered and filtrate was extracted with water, brine and dried over Na2SO4. The organic extract was concentrated under vacuum and then distilled to afford a colorless water-like liquid 320 mg in 82% yield.1H NMR (CDCl3) δ 1.00 (s, 6H), 1.40 (m, 2H), 1.46 (m, 2H), 2.20 (s, 6H), 2.68 (m, 2H).

Synthesis of Compound 44

Synthesis of Compound 45

The following illustrate representative pharmaceutical dosage forms, containing a compound of formula I (‘Compound X’), for therapeutic or prophylactic use in humans.