Apoptosis inhibitor formulations for prevention of hearing loss

Abstract

A formulation for sustained release of an apoptosis inhibitor in the inner ear to protect from hearing loss, especially due to exposure to chemotherapy with drugs such as cisplatin. The formulation can be injected through a small gauge needle into the inner ear, where it gels to form a sustained release depot for controlled delivery of drug over a few days. In the preferred embodiment, the formulation includes a thermoresponsive sol-gel polymer such as POLOXAMER 407 and an apoptosis inhibitory agent, preferably an inhibitor of apoptotic protease activating factor-1 (APAF-1), in an effective amount to prevent hearing loss, for example, due to the administration of platinum-based chemotherapeutic agents. As demonstrated by the examples, the hydrogel provides sustained release of an apoptosis inhibitory agent, LPT99, an anti-apoptosis agent that inhibits apoptotic protease activating factor-1 (APAF-1), as well as safety and efficacy in in vitro and in vivo models.

We claim: 
     
         1 . A sustained release formulation delivering an effective amount of an apoptotic inhibitory agent for a period of at least three days for prevention or treatment of a condition, disease or disorder resulting in apoptosis, the formulation comprising a solution or suspension of the apoptotic inhibitory agent in a polymer transitioning from a liquid state at room temperature which can be injected through a 23-30 gauge needle to a gel state at body temperature. 
     
     
         2 . The formulation of  claim 1  wherein the apoptotic inhibitor agent inhibits a protein in an apoptosis pathway. 
     
     
         3 . The formulation of  claim 2  wherein the apoptotic inhibitory agent inhibits apoptotic protease activating factor-1 (Apaf-1) or reduces cyt-c release. 
     
     
         4 . The formulation of  claim 1 , wherein the condition is hearing loss associated with exposure to ototoxic agents such as chemotherapeutics, exposure to loud noise, aging, Meniere's Disease, sudden sensorineural hearing loss, autoimmune inner ear disease or a combination thereof. 
     
     
         5 . The formulation of  claim 1  wherein the polymer enhances solubility of the apoptotic inhibitory agent at least about 30-fold, 100-fold, 300-fold, or 1000-fold, compared to a corresponding formulation lacking the polymer or to water. 
     
     
         6 . The formulation of  claim 5  wherein the apoptotic inhibitory agent is stable in solution with the polymer at room temperature for a period of at least three months. 
     
     
         7 . The formulation of  claim 1 , wherein the apoptosis inhibitory agent is hydrophobic and/or a small molecule. 
     
     
         8 . The formulation of  claim 1 , wherein the apoptosis inhibitory agent inhibits apoptosis of sensorineural cells, such as cochlear hair cells from an insult that causes loss of a portion of hair cells, neurons, or stria vascularis cells or maintains the tight junctions between the cells. 
     
     
         9 . The formulation of  claim 1 , wherein the apoptosis inhibitory agent contains a 1,4-piperazine-2,5-dione moiety or a pharmaceutically acceptable salt thereof. 
     
     
         10 . The formulation of  claim 1 , wherein the apoptosis inhibitory agent is a compound of Formula I or a pharmaceutically acceptable salt thereof: 
       
         
           
           
               
               
           
         
         wherein: 
         R 1  and R 2  are independently hydrogen, C 1-5  alkyl, C 2-5  alkenyl, (CH 2 ) 0-3 -cycloalkyl, —(CH 2 ) 1-3 -heterocycle, —(CH 2 ) 0-3 -aryl, —(CH 2 ) 0-3 -heteroaryl, —(CH 2 ) 1-2 —CH(aryl) 2 , —(CH 2 ) 1-2 —CH(aryl)(heteroaryl), or —(CH 2 ) 1-2 —CH(heteroaryl) 2 ; 
         R 3  is selected from hydrogen, C 1-5  alkyl, C2-5 alkenyl, (CH 2 ) 0-3 -cycloalkyl, —(CH 2 ) 1-3 -heterocycle, —(CH 2 ) 1-3 -aryl, —(CH 2 ) 1-3 -heteroaryl, —(CH 2 ) 1-3 —CONR 5 R 6 , —(CH 2 ) 1-2 —CH(aryl) 2 , —(CH 2 ) 1-2 —CH(aryl)(heteroaryl), and —(CH 2 ) 1-2 —CH(heteroaryl) 2 ; 
         R 4  is hydrogen, —C 1-5  alkyl, —(CHR 7 ) 1-3 —CO—NR 5 R 6 , —(CHR 7 ) 1-3 —CO—OR 5 , —(CH 2 ) 1-3 —NR 5 R 6 , —(CH 2 ) 1-3 —CO[NCHR 7 CO] m NH 2 , —(CH 2 ) 1-3 —CO[NCHR 7 CO] m OR 5 ; 
         n is 1 or 2; m is 1, 2, or 3; 
         R 5  and R 6  are independently from hydrogen, —C 1-5  alkyl, or —(CH 2 ) 0-3 -aryl; 
         each R 7  is independently hydrogen, —C 1-5  alkyl, —(CH 2 ) 1-3 -aryl, or —(CH 2 ) 1-3 -heteroaryl; 
         wherein the C 1-5  alkyl, C 2-5  alkenyl, cycloalkyl and heterocycle groups are optionally substituted with one or more substituents selected independently from halogen, OR 5 , OCF 3 , SH, SR 5 , NR 5 R 6 , NHCOR 5 , COOH, COOR 5 , OCOR 5 , aryl, and heteroaryl; 
         wherein the aryl and heteroaryl groups are optionally substituted with one or more substituents selected independently from halogen, CF 3 , OR 5 , OCF 3 , SH, SR 5 , NH 2 , NHCOR 5 ; NO 2 , CN, COR 5 , COOR 5 , OCOR 5 , CONR 5 R 6 , —(CH 2 ) 0-3 NR 5 R 6 , SO 2 NH 2 , NHSO 2 CH 3 , C 1-5  alkyl, aryl and heteroaryl; 
         wherein the heterocycle and heteroaryl groups are optionally substituted on a secondary nitrogen atom with C 1-5  alkyl, cycloalkyl, or —(CH 2 ) 0-3 -aryl; and 
         on the condition that (i) when R 2  is 2-(4-fluorophenyl)ethyl, R 4  is —CH 2 —CO—NH 2  and n is 1; (ii) if R 1  is 2-(4-fluorophenyl)ethyl, R 3  is not 2-(4-methoxyphenyl)ethyl, 2-(2-pyridyl)ethyl or 2-(2,4-dichlorophenyl)ethyl; and (iii) if R 1  is 2-(2,4-dichlorophenyl)ethyl, R 3  is not 2-(4-methoxyphenyl)ethyl, or 2-(2-pyridyl)ethyl. 
       
     
     
         11 . The formulation of  claim 10 , wherein the apoptosis inhibitory agent is a compound having the structure: 
       
         
           
           
               
               
           
         
       
     
     
         12 . The formulation of  claim 1  comprising a therapeutically effective amount of between about 10 mM and about 1000 mM, between about 100 and about 800 mM, or between about 125 and about 500 mM of the apoptosis inhibitory agent (e.g. a compound containing a 1,4-piperazine-2,5-dione moiety) to produce a concentration of between about 125 μg/mL and about 500 mg/ml, and a concentration of the apoptosis inhibitory agent of between about 0.0031% w/w and about 1.5% w/w of the composition, of a 1,4-piperazine-2,5-dione moiety, or pharmaceutically acceptable salt thereof, dissolved in a formulation comprising between about 10% to about 30% weight by weight of a poly(ethylene oxide)-poly(propylene oxide) triblock copolymer having a formula A-B-A or B-A-B, wherein, A is poly(ethylene oxide) and B is poly(propylene oxide). 
     
     
         13 . The formulation of  claim 12 , wherein the 1,4-piperazine-2,5-dione compound is 2-(4-(2,4-dichlorophenethyl)-3,6-dioxo-1-(2-(thiophen-2-yl)ethyl)piperazin-2-yl)-N-(2-(5-methoxy-1H-indol-3-yl)ethyl)acetamide. 
     
     
         14 . The formulation of  claim 1 , wherein the gel state provides sustained release of the apoptosis inhibitory agent for a period of between at least about three days and 30 days. 
     
     
         15 . The formulation of  claim 1 , wherein the synthetic polymer is a non-ionic, amphiphilic polymer. 
     
     
         16 . The formulation of  claim 15 , wherein the synthetic polymer is selected from the group consisting of synthetic polymers such as N-isopropylacrylamide (NiPAAM) polymers, poly(ethylene oxide)-b-poly(propylene oxide)-b-poly(ethylene oxide) (PEO-PPO-PEO), poly(ethylene glycol) (PEG)-biodegradable polyester copolymers, block copolymers of ethylene oxide and propylene oxide); and tetrafunctional block copolymer derived from sequential addition of propylene oxide and ethylene oxide to ethylenediamine. 
     
     
         17 . The formulation of  claim 16 , wherein the synthetic polymer enhances solubility of the agent between at least about 30-fold, 100-fold, 300-fold, or 1000-fold compared to a corresponding formulation lacking the synthetic polymer or water. 
     
     
         18 . The formulation of  claim 1 , having a pH between about 6.8 and about 7.7, preferably 7.2, and optionally having an osmolality between about 240 mOsmol/kg and about 350 mOsmol/kg, preferably about 280 mOsmol/kg. 
     
     
         19 . A method of treating a condition, disease or disorder associated with apoptosis or cell death, comprising:
 administering into the inner ear of a person in need or at risk of the condition, disease or disorder a sustained release formulation delivering an effective amount of an apoptotic inhibitory agent for a period of at least three days for treatment of a condition, disease or disorder resulting in apoptosis, the formulation comprising a solution or suspension of the apoptotic inhibitory agent in a polymer transitioning from a liquid state at room temperature which can be injected through a 23 gauge or higher needle to a gel state at body temperature.   
     
     
         20 . A method of making a sustained release formulation delivering an effective amount of an apoptotic inhibitory agent for a period of at least three days for treatment of a condition, disease or disorder resulting in apoptosis, by mixing the apoptotic inhibitory agent in a formulation comprising an effective amount of a polymer transitioning from a liquid state at room temperature to form a liquid which can be injected through a 23 gauge needle into the ear to form a stable hydrogel matrix at body temperature.