Compositions, systems and methods for gene expression noise drug screening and uses thereof

Abstract

HIV-1's ability to enter a transcriptionally dormant state and establish a reservoir of latently infected cells is considered the major barrier to eradicating the virus from infected patients. Stochastic noise (i.e. fluctuations) in an HIV-1 transcriptional positive-feedback loop is one mechanism that enables HIV-1 to establish latency. Here, Applicants demonstrate that small-molecule modulation of noise in HIV-1 gene expression radically perturbs HIV-1 latency.

What is claimed is: 
     
         1 . A pharmaceutical composition for reactivating a latent Human Immunodeficiency Virus Type-1 (“HIV-1”) reservoir in a mammalian cell having an integrated HIV-1 genome, the composition comprising:
 (i) a variability enhancer in an amount sufficient to modulate variability of expression of a gene promoter without substantially changing the mean expression level of the gene promoter; 
 (ii) an activator of latent HIV-1 gene expression in an amount effective to activate latent HIV-1 gene expression; and 
 (iii) a pharmaceutically acceptable carrier. 
 
     
     
         2 . The pharmaceutical according to  claim 1 , wherein the variability enhancer and the activator of latent HIV-1 gene expression synergize to reactivate the latent HIV-1 reservoir. 
     
     
         3 . The pharmaceutical according to  claim 1 , wherein upon reactivation of the latent HIV-1 reservoir, latent HIV-1 is rendered sensitive to killing by an immunotoxin. 
     
     
         4 . The pharmaceutical composition according to  claim 1 , wherein the variability enhancer is selected from the group consisting of thiamylal sodium, Estramustine, griseofulvin, telmisartan, docetaxel, riboflavin, pantothenic acid(d) na salt, mercaptopurine, pemetrexed, ethinyl estradiol, irinotecan hydrochloride, dutasteride, felbinac, vincristine sulfate, thiram, bezafibrate, Indomethacin, mebendazole, ouabain, sulfaquinoxaline sodium, oxybutynin chloride, oxyphencyclimine hydrochloride, saxagliptin, phenylmercuric acetate, troclosene potassium, artemisinin, cytarabine, thioguanine, hydroquinone, acetophenazine maleate, hexylresorcinol, oxytetracycline, cefadroxil, tolnaftate, phenylbutyric acid, atorvastatin calcium, carboplatin, 5-azacytidine, levodopa, lamivudine, sotalol hydrochloride, clavulanate lithium, digoxin, sodium monofluorophosphate, dexpanthenol, rabeprazole sodium, cetirizine hydrochloride, trilostane, modafinil, thiabendazole, adapalene, sulfameter, Fluoxetine hydrochloride, trimethobenzamide hydrochloride, Metoclopramide hydrochloride, guanethidine sulfate, trichlormethine, sucralfate, valganciclovir hydrochloride, bemotrizinol, parachlorophenol, (±)-Verapamil hydrochloride, oxidopamine hydrochloride, fomepizole hydrochloride, Xylazine hydrochloride, diperodon hydrochloride, zomepirac sodium, closantel, fenbendazole, finasteride, Procainamide hydrochloride, celecoxib, colistin sulfate, imipenem, cefoxitin sodium, azithromycin, clidinium bromide, Tolazamide, chlorpheniramine maleate, colesevalam hydrochloride, Pargyline hydrochloride, hydralazine hydrochloride, anastrozole, vinorelbine, bleomycin, dasatinib, busulfan, vorinostat, selamectin, idoxuridine, carvedilol, (±)-Isoproterenol hydrochloride, racephedrine hydrochloride, vardenafil hydrochloride, guaiacol, clobetasol propionate, mycophenolic acid, fludrocortisone acetate, zaleplon, nitroglycerin, riboflavin 5-phosphate sodium, acetriazoic acid, digitoxin, carnitine hydrochloride, albendazole, taurine, Taxol, mitomycin c, clofarabine, arginine hydrochloride, tetramizole hydrochloride, thiostrepton, lonidamine, melphalan, fludarabine phosphate, methotrexate(+/−), rifaximin, Arecoline hydrobromide, zalcitabine, erythrosine sodium, nikethamide, oxfendazole, editol, doxapram hydrochloride, aminacrine, dobutamine hydrochloride, rutin, single stereoisomers, mixtures of stereoisomers, pharmaceutically acceptable salts, prodrugs and functional derivatives thereof. 
     
     
         5 . The pharmaceutical composition according to  claim 1 , wherein the activator of latent HIV-1 gene expression is selected from the group consisting of a histone deacetylase (HDAC) inhibitor, a Protein Kinase C (PKC) agonist, a methylation inhibitor, a bromodomain inhibitor, and an anti-cancer drug. 
     
     
         6 . The pharmaceutical composition according to  claim 5 , wherein, the HDAC inhibitor is selected from the group consisting of 12-deoxyphorbol 13-phenylacetate (DPP), Suberoylannilide Hydroxamic Acid (SAHA), MS-275, valproic acid (VPA), trichostatin A (TSA), single stereoisomers, mixtures of stereoisomers, pharmaceutically acceptable salts, and prodrugs thereof. 
     
     
         7 . The pharmaceutical composition according to  claim 5 , wherein the PKC agonist is selected from the group consisting of tumor necrosis factor alpha (TNFα), prostratin, phorbol 12-myristate 13-acetate (PMA), bryostatin, single stereoisomers, mixtures of stereoisomers, pharmaceutically acceptable salts, and prodrugs thereof. 
     
     
         8 . The pharmaceutical composition according to  claim 5 , wherein the methylation inhibitor is selected from the group consisting of azacytidine, fludarabine, adenosine, single stereoisomers, mixtures of stereoisomers, pharmaceutically acceptable salts, and prodrugs thereof. 
     
     
         9 . The pharmaceutical composition according to  claim 5 , wherein the bromodomain inhibitor is JQ1, GSK1210151A, or PFI-1, single stereoisomers, mixtures of stereoisomers, pharmaceutically acceptable salts, and prodrugs thereof. 
     
     
         10 . The pharmaceutical composition according to  claim 5 , wherein the anticancer drug is cytarabine, single stereoisomers, mixtures of stereoisomers, pharmaceutically acceptable salts, and prodrugs thereof. 
     
     
         11 . The pharmaceutical composition according to  claim 1 , wherein the gene promoter is a promoter involved in establishing or maintaining HIV-1 latency. 
     
     
         12 . The pharmaceutical composition according to  claim 11 , wherein the gene promoter is an HIV-1 long terminal repeat promoter. 
     
     
         13 . The pharmaceutical composition according to  claim 1 , further comprising:
 (iv) a HAART compound.   
     
     
         14 . The pharmaceutical composition according to  claim 13 , wherein the HAART compound comprises a nucleoside reverse transcriptase inhibitor, a non-nucleoside reverse transcriptase inhibitor or a protease inhibitor. 
     
     
         15 . The pharmaceutical composition according to  claim 14 , wherein the nucleoside reverse transcriptase inhibitor is selected from the group consisting of ZIDOVUDINE® (AZT), STAVUDINE® (d4T), and LAMIVUDINE® (3TC). 
     
     
         16 . The pharmaceutical composition according to  claim 14 , wherein the non-nucleoside reverse transcriptase inhibitor is NEVIRAPINE® (BI-RG-587). 
     
     
         17 . The pharmaceutical composition according to  claim 14 , wherein the nucleoside reverse transcriptase inhibitor is selected from the group consisting of INDINAVIR® (MK-639), and SAQINAVIR® (Ro-31-8959), and NELFINAVIR® (AG-1343). 
     
     
         18 . The pharmaceutical composition according to  claim 1 , further comprising:
 (iv) an immunotoxin.   
     
     
         19 . A kit for reactivating a latent Human Immunodeficiency Virus Type 1 (“HIV-1”) reservoir in a mammalian cell having an integrated HIV-1 genome, the kit comprising:
 (i) a first container containing a variability enhancer in an amount sufficient to modulate variability of expression of a gene promoter without substantially changing the mean expression level of the gene promoter; 
 (ii) a second container containing an activator of latent HIV-1 gene expression in an amount effective to activate latent HIV-1 gene expression; and 
 (iii) an instruction for using (i) and (ii) for reactivating the latent HIV-1 reservoir in the mammalian cell. 
 
     
     
         20 . The kit according to  claim 19 , wherein the variability enhancer is selected from the group consisting thiamylal sodium, Estramustine, griseofulvin, telmisartan, docetaxel, riboflavin, pantothenic acid(d) na salt, mercaptopurine, pemetrexed, ethinyl estradiol, irinotecan hydrochloride, dutasteride, felbinac, vincristine sulfate, thiram, bezafibrate, Indomethacin, mebendazole, ouabain, sulfaquinoxaline sodium, oxybutynin chloride, oxyphencyclimine hydrochloride, saxagliptin, phenylmercuric acetate, troclosene potassium, artemisinin, cytarabine, thioguanine, hydroquinone, acetophenazine maleate, hexylresorcinol, oxytetracycline, cefadroxil, tolnaftate, phenylbutyric acid, atorvastatin calcium, carboplatin, 5-azacytidine, levodopa, lamivudine, sotalol hydrochloride, clavulanate lithium, digoxin, sodium monofluorophosphate, dexpanthenol, rabeprazole sodium, cetirizine hydrochloride, trilostane, modafinil, thiabendazole, adapalene, sulfameter, Fluoxetine hydrochloride, trimethobenzamide hydrochloride, Metoclopramide hydrochloride, guanethidine sulfate, trichlormethine, sucralfate, valganciclovir hydrochloride, bemotrizinol, parachlorophenol, (±)-Verapamil hydrochloride, oxidopamine hydrochloride, fomepizole hydrochloride, Xylazine hydrochloride, diperodon hydrochloride, zomepirac sodium, closantel, fenbendazole, finasteride, Procainamide hydrochloride, celecoxib, colistin sulfate, imipenem, cefoxitin sodium, azithromycin, clidinium bromide, Tolazamide, chlorpheniramine maleate, colesevalam hydrochloride, Pargyline hydrochloride, hydralazine hydrochloride, anastrozole, vinorelbine, bleomycin, dasatinib, busulfan, vorinostat, selamectin, idoxuridine, carvedilol, (±)-Isoproterenol hydrochloride, racephedrine hydrochloride, vardenafil hydrochloride, guaiacol, clobetasol propionate, mycophenolic acid, fludrocortisone acetate, zaleplon, nitroglycerin, riboflavin 5-phosphate sodium, acetriazoic acid, digitoxin, carnitine hydrochloride, albendazole, taurine, Taxol, mitomycin c, clofarabine, arginine hydrochloride, tetramizole hydrochloride, thiostrepton, lonidamine, melphalan, fludarabine phosphate, methotrexate(+/−), rifaximin, Arecoline hydrobromide, zalcitabine, erythrosine sodium, nikethamide, oxfendazole, editol, doxapram hydrochloride, aminacrine, dobutamine hydrochloride, rutin, single stereoisomers, mixtures of stereoisomers, pharmaceutically acceptable salts, prodrugs and functional derivatives thereof.