Compositions for treating addiction, dependence, withdrawal, and / or opioid induced hyperalgesia

Abstract

A compound having a structure of formula (I), (II), (III), (IV), (V), (VI), or a pharmaceutically acceptable salt, a tautomer, or a solvate thereof for use in treating opioid addiction, withdrawal and / or dependence and / or loss of opioid induced analgesia and / or occurrence of opioid induced hyperalgesia in a subject in need thereof.

Description

COMPOSITIONS FOR TREATING ADDICTION, DEPENDENCE, WITHDRAWAL, AND / OR OPIOID INDUCED HYPERALGESIA RELATED APPLICATION

[0001] This application claims priority from U.S. Provisional Application No.63 / 607,836, filed December 8, 2023, the subject matter of which is incorporated herein by reference in its entirety. BACKGROUND

[0002] Substance addiction is a serious public health problem throughout the world. Heroin and other opioids, including prescription painkillers, are widely abused and account for a large percentage of illicit drug use. Opioid use is also linked to approximately 50% of violent crimes in the United States and costs the U.S. economy billions of dollars per year.

[0003] Acute withdrawal from drug dependence is characterized by dramatic and traumatic symptoms, including sweating, racing heart, palpitations, muscle tension, tightness in the chest, difficulty breathing, tremor, nausea, vomiting, diarrhea, grand mal seizures, heart attacks, strokes, hallucinations and delirium tremens (DTs). Once acute withdrawal symptoms have subsided, post-acute withdrawal syndrome can last for months or years. Post-acute withdrawal symptoms include fatigue, depression, lack of motivation, and increased pain sensitivity.

[0004] Numerous treatments have been developed in attempts to ameliorate acute and post-acute withdrawal symptoms. However, in most cases, treatment of withdrawal requires use of other addictive substances (e.g., morphine or methadone). Treatment also requires that the addict attend a clinic daily for an extended amount of time. Due to the severity and duration of withdrawal symptoms, opioid-addicted patients have a high rate of relapse. There is a significant need for effective, non-addictive treatment for acute and post-acute opioid withdrawal symptoms. SUMMARY

[0005] Embodiments described herein relate to compositions and methods of attenuating and / or treating opioid addiction, withdrawal and / or dependance in a subject in need thereof and / or preventing loss of opioid induced analgesia and occurrence of opioidinduced hyperalgesia, and particularly relates to the use of thiol-based compounds in compositions and methods of attenuating and / or treating opioid withdrawal and / or dependance and / or preventing loss of opioid induced analgesia and occurrence of opioid induced hyperalgesia.

[0006] In some embodiments, a compound can have a structure of formula (I):a pharmaceutically acceptable salt, a tautomer, or a solvate thereof; wherein: X is -OR2or -N(R3)2; X1is -SR4or -S(O)nOR5; R1is H, alkyl optionally substituted with one or more halogen, or -C(O)-alkyl optionally substituted with one or more halogen; R2, R5, and R6are each independently H or alkyl optionally substituted with one or more halogen; each R3is independently H or alkyl optionally substituted with one or more halogen; R4is -N=O or -OR6; and n is 1 or 2.

[0007] In some embodiments, R1is H, C1-C6alkyl optionally substituted with one or more halogen, or -C(O)-(C1-C6 alkyl) optionally substituted with one or more halogen. For example, R1is H, methyl, ethyl, propyl, butyl, -C(O)-methyl, -C(O)-ethyl, -C(O)-propyl, or - C(O)-butyl, each optionally substituted with one or more halogen.

[0008] In some R2and R6are each independently H or C1-C6 alkyl optionally substituted with one or more halogen. For example, R2and R6are each independently H, methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

[0009] In some embodiments, each R3is independently H or C1-C6alkyl optionally substituted with one or more halogen. For example, one of each R3is H and the other of R3is methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

[0010] In some embodiments, X is OR2, R1is H, C1-C6 alkyl optionally substituted with one or more halogen, or -C(O)-(C1-C6alkyl) optionally substituted with one or more halogen,and R2is H or C1-C6 alkyl optionally substituted with one or more halogen. For example, X is OR2, R1is H, methyl, ethyl, propyl, butyl, -C(O)-methyl, -C(O)-ethyl, -C(O)-propyl, or - C(O)-butyl, each optionally substituted with one or more halogen, and R2is H, methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

[0011] In other embodiments, X is N(R3)2; R1is H, C1-C6 alkyl optionally substituted with one or more halogen, or -C(O)-(C1-C6alkyl) optionally substituted with one or more halogen; and each R3is independently H or C1-C6 alkyl optionally substituted with one or more halogen. For example, X is N(R3)2; R1is H, methyl, ethyl, propyl, butyl, -C(O)-methyl, -C(O)-ethyl, -C(O)-propyl, or -C(O)-butyl, each optionally substituted with one or more halogen, and one R3is H and the other R3is methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

[0012] In some embodiments, X1is SR4and R4is -N=O.

[0013] In other embodiments, X1is SR4, R4is -OR6, and R6is H or C1-C6 alkyl optionally substituted with one or more halogen. For example, X1is SR4, R4is -OR6, and R6is H, methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

[0014] In some embodiments, X1is -S(O)nOR5and R5is H or C1-C6alkyl optionally substituted with one or more halogen. For example, X1is -S(O)nOR5and R5is H, methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

[0015] In some embodiments, the compound can have a structure of formula (II):a pharmaceutically acceptable salt, a tautomer, or a solvate thereof; wherein: X is -OR2or -N(R3)2; R1is H, alkyl optionally substituted with one or more halogen, or -C(O)-alkyl optionally substituted with one or more halogen; R2is independently H or alkyl optionally substituted with one or more halogen; each R3is independently H or alkyl optionally substituted with one or more halogen.

[0016] In some embodiments, R1is H, C1-C6 alkyl optionally substituted with one or more halogen, or -C(O)-(C1-C6alkyl) optionally substituted with one or more halogen. For example, R1is H, methyl, ethyl, propyl, butyl, -C(O)-methyl, -C(O)-ethyl, -C(O)-propyl, or - C(O)-butyl, each optionally substituted with one or more halogen.

[0017] In some embodiments, R2is H or C1-C6 alkyl optionally substituted with one or more halogen. For example, R2is H, methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

[0018] In some embodiments, each R3is independently H or C1-C6alkyl optionally substituted with one or more halogen. For example, one of each R3is H and the other of R3is methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

[0019] In some embodiments, the compound can have a structure of formula (III):a pharmaceutically acceptable salt, a tautomer, or a solvate thereof; wherein: X is -OR2or -N(R3)2; R1is H, alkyl optionally substituted with one or more halogen, or -C(O)-alkyl optionally substituted with one or more halogen; R2and R6are each independently H or alkyl optionally substituted with one or more halogen; and each R3is independently H or alkyl optionally substituted with one or more halogen.

[0020] In some embodiments, R1is H, C1-C6alkyl optionally substituted with one or more halogen, or -C(O)-(C1-C6 alkyl) optionally substituted with one or more halogen. For example, R1is H, methyl, ethyl, propyl, butyl, -C(O)-methyl, -C(O)-ethyl, -C(O)-propyl, or - C(O)-butyl, each optionally substituted with one or more halogen.

[0021] In some embodiments, R2and R6are each independently H or C1-C6alkyl optionally substituted with one or more halogen. For example, R2and R6are each independently H, methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

[0022] In some embodiments, each R3is independently H or C1-C6 alkyl optionally substituted with one or more halogen.

[0023] In some embodiments, where one of each R3is H, the other of R3is methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

[0024] In other embodiments, the compound can have a structure of formula (IV):a pharmaceutically acceptable salt, a tautomer, or a solvate thereof; wherein: X is -OR2or -N(R3)2; R1is H, alkyl optionally substituted with one or more halogen, or -C(O)-alkyl optionally substituted with one or more halogen; R2and R5are each independently H or alkyl optionally substituted with one or more halogen; and each R3is independently H or alkyl optionally substituted with one or more halogen.

[0025] In some embodiments, R1is H, C1-C6 alkyl optionally substituted with one or more halogen, or -C(O)-(C1-C6alkyl) optionally substituted with one or more halogen. For example, R1is H, methyl, ethyl, propyl, butyl, -C(O)-methyl, -C(O)-ethyl, -C(O)-propyl, or - C(O)-butyl, each optionally substituted with one or more halogen.

[0026] In some embodiments, R2and R5are each independently H or C1-C6 alkyl optionally substituted with one or more halogen. For example, R2and R5are each independently H, methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

[0027] In some embodiments, each R3is independently H or C1-C6alkyl optionally substituted with one or more halogen. For example, one of each R3is H and the other of R3is methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

[0028] In some embodiments, the compound can have a structure of formula (V):a pharmaceutically acceptable salt, a tautomer, or a solvate thereof; wherein: X is -OR2or -N(R3)2; R1is H, alkyl optionally substituted with one or more halogen, or -C(O)-alkyl optionally substituted with one or more halogen; R2and R5are each independently H or alkyl optionally substituted with one or more halogen; and each R3is independently H or alkyl optionally substituted with one or more halogen.

[0029] In some embodiments, R1is H, C1-C6 alkyl optionally substituted with one or more halogen, or -C(O)-(C1-C6 alkyl) optionally substituted with one or more halogen. For example, R1is H, methyl, ethyl, propyl, butyl, -C(O)-methyl, -C(O)-ethyl, -C(O)-propyl, or - C(O)-butyl, each optionally substituted with one or more halogen.

[0030] In some embodiments, R2and R5are each independently H or C1-C6 alkyl optionally substituted with one or more halogen. For example, R2and R5are each independently H, methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

[0031] In some embodiments, each R3is independently H or C1-C6alkyl optionally substituted with one or more halogen. For example, one of each R3is H and the other of R3is methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

[0032] Other embodiments, described herein relate to a pharmaceutical composition comprising the compounds described herein.

[0033] Still other embodiments described herein relate to a pharmaceutical composition for use in treating addiction, withdrawal and / or dependence in a subject in need thereof that includes a compound having a structure of formulas:adduct, a pharmaceutically acceptable salt, a tautomer, or a solvate thereof; wherein:X is -OR2or -N(R3)2; X1is -SR4or -S(O)nOR5; X2is -OR9or -N(R10)2; X3is -OR11or -N(R12)2; R1, R7, and R8are each independently H, alkyl optionally substituted with one or more halogen, or -C(O)-alkyl optionally substituted with one or more halogen; R2, R5, R6, R9and R11are each independently H or alkyl optionally substituted with one or more halogen; each R3, R10, and R12are independently H or alkyl optionally substituted with one or more halogen; R4is H, -N=O or -OR6; and n is 1 or 2.

[0034] In some embodiments, X is not -OR2if R1is H.

[0035] In other embodiments, X3is not -OR11if X2is -OR9and R7and R8are H.

[0036] In some embodiments, the adduct of the compound of formulas I or VI is biologically active and includes at least one of an albumin adduct, a glucose adduct, an L- cysteine adduct, an L-glutathione adduct, or a D-cysteine adduct.

[0037] In some embodiments, R1, R7, and R8are each independently H, C1-C6 alkyl optionally substituted with one or more halogen, or -C(O)-(C1-C6alkyl) optionally substituted with one or more halogen. For example, R1, R7, and R8are each independently H, methyl, ethyl, propyl, butyl, -C(O)-methyl, -C(O)-ethyl, -C(O)-propyl, or -C(O)-butyl, each optionally substituted with one or more halogen.

[0038] In some embodiments, R2, R5, R6, R9and R11are each independently H or C1-C6alkyl optionally substituted with one or more halogen. For example, R2, R5, R6, R9and R11are each independently H, methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

[0039] In some embodiments, each R3, R10, and R12is independently H or C1-C6 alkyl optionally substituted with one or more halogen.

[0040] In some embodiments, one of each R3, R10, and R12is H and the other of R3, R6, and R9is methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

[0041] In some embodiments, X is -OR2, R1is H, C1-C6 alkyl optionally substituted with one or more halogen, or -C(O)-(C1-C6alkyl) optionally substituted with one or more halogen, and R2is H or C1-C6 alkyl optionally substituted with one or more halogen. For example, X is -OR2, R1is H, methyl, ethyl, propyl, butyl, -C(O)-methyl, -C(O)-ethyl, -C(O)- propyl, or -C(O)-butyl, each optionally substituted with one or more halogen, and R2is H, methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

[0042] In some embodiments, X is -N(R3)2; R1is H, C1-C6 alkyl optionally substituted with one or more halogen, or -C(O)-(C1-C6alkyl) optionally substituted with one or more halogen; and each R3is independently H or C1-C6 alkyl optionally substituted with one or more halogen. For example, X is -N(R3)2; R1is H, methyl, ethyl, propyl, butyl, -C(O)- methyl, -C(O)-ethyl, -C(O)-propyl, or -C(O)-butyl, each optionally substituted with one or more halogen, and one R3is H and the other R3is methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

[0043] In some embodiments, X1is -SR4and R4is -N=O.

[0044] In other embodiments, X1is -SR4and R4is H.

[0045] In some embodiments, X1is -SR4, R4is -OR6, and R6is H or C1-C6alkyl optionally substituted with one or more halogen. For example, X1is -SR4, R4is -OR6, and R6is H, methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

[0046] In some embodiments, X1is -S(O)nOR5and R5is H or C1-C6alkyl optionally substituted with one or more halogen. For example, X1is -S(O)nOR5and R5is H, methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

[0047] In some embodiments, X2is -OR9, R7is H, C1-C6 alkyl optionally substituted with one or more halogen, or -C(O)-(C1-C6 alkyl) optionally substituted with one or more halogen, R9is H or C1-C6alkyl optionally substituted with one or more halogen; X3is OR11, R8is H, C1-C6 alkyl optionally substituted with one or more halogen, or -C(O)-(C1-C6 alkyl) optionally substituted with one or more halogen, and R11is H or C1-C6alkyl optionally substituted with one or more halogen. For example, X2is OR9, R7is H, methyl, ethyl, propyl, butyl, -C(O)-methyl, -C(O)-ethyl, -C(O)-propyl, or -C(O)-butyl, each optionally substituted with one or more halogen, R9is H, methyl, ethyl, or propyl, or butyl, each optionally substituted with one ormore halogen, X3is OR11; R8is H, methyl, ethyl, propyl, butyl, -C(O)-methyl, -C(O)-ethyl, - C(O)-propyl, or -C(O)-butyl, each optionally substituted with one or more halogen, and R11is H, methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

[0048] In other embodiments, X2is N(R10)2, R7is H, C1-C6alkyl optionally substituted with one or more halogen, or -C(O)-(C1-C6 alkyl) optionally substituted with one or more halogen, each R10is independently H or C1-C6alkyl optionally substituted with one or more halogen, X3is N(R12)2, R8is H, C1-C6 alkyl optionally substituted with one or more halogen, or -C(O)-(C1-C6alkyl) optionally substituted with one or more halogen, and each R12is independently H or C1-C6 alkyl optionally substituted with one or more halogen. For example, X2is N(R10)2, R7is H, methyl, ethyl, propyl, butyl, -C(O)-methyl, -C(O)-ethyl, -C(O)-propyl, or -C(O)-butyl, each optionally substituted with one or more halogen, one R10is H and the other R10is methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen, X3is N(R12)2; R8is H, methyl, ethyl, propyl, butyl, -C(O)-methyl, - C(O)-ethyl, -C(O)-propyl, or -C(O)-butyl, each optionally substituted with one or more halogen, and one R12is H and the other R12is methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

[0049] In some embodiments, the compound is not cysteine, cystine, a cysteine alkylester, or cystine dialkylester.

[0050] In some embodiments, the compound is not N-acetylcysteine.

[0051] In some embodiment, the pharmaceutical composition further includes a pharmaceutically acceptable carrier or excipient.

[0052] In some embodiments, the subject has an addiction, withdrawal and / or dependence from at least one of alcohol, amphetamine, cocaine, methamphetamine, nicotine, or opioid.

[0053] In some embodiments, the subject has an opioid withdrawal, dependence, and / or addiction.

[0054] In some embodiments, the subject has neonatal opioid withdrawal syndrome.

[0055] In some embodiments, the subject has an opioid use disorder (OUD).

[0056] In some embodiments, the opioid comprises at least one of alfentanil, buprenorphine, butorphanol, carfentanil, codeine, diamorphine, dextromoramide, dezocine, dihydrocodeine, fentanyl, hydrocodone, hydromorphone, levorphanol, meperidine, meptazinol, methadone, morphine, nalbuphine, nalorphine, opium, oxycodone,oxymorphone, pentazocine, propoxyphene, remifentanil, sufentanil, tapentadol, and tramadol, and pharmaceutically acceptable salts thereof. For example, the opioid is carfentanil, fentanyl, remifentanil, or sufentanil.

[0057] In some embodiments, the subject has been administered an opioid antagonist and the composition inhibits opioid antagonist withdrawal. The opioid antagonist can include, for example, naloxone, an oxymorphol analog of naloxone, a naloxone salt, or a naloxone dihydrate.

[0058] In some embodiments, the composition can be administered concurrently with opioid administration and / or up to about 10 minutes, up to about 20 minutes, up to about 30 minutes, up to about 40 minutes, up to about 50 minutes, up to about 60 minutes, up to about 70 minutes, up to about 80 minutes, up to about 90 minutes, up to about 100 minutes, up to about 110 minutes, or up to about 120 minutes before or after initiation of opioid administration.

[0059] Other embodiments described herein relate to a pharmaceutical composition for use in preventing loss of opioid induced analgesia and occurrence opioid induced hyperalgesia in a subject in need thereof that includes any of the compounds described herein.

[0060] In some embodiments, the composition is formulated for systemic administration, continuous intravenous infusion, oral administration, intranasal administration, and / or inhalation.

[0061] Still other embodiments relate to a pharmaceutical composition that includes at least one opioid and an amount of a compound described herein effective to reduce opioid addiction, withdrawal, and / or dependence and / or loss of opioid induced analgesia and / or occurrence opioid induced hyperalgesia.

[0062] In some embodiments, the composition includes a pharmaceutically acceptable carrier or excipient.

[0063] In some embodiments, the opioid comprises at least one of alfentanil, buprenorphine, butorphanol, carfentanil, codeine, diamorphine, dextromoramide, dezocine, dihydrocodeine, fentanyl, hydrocodone, hydromorphone, levorphanol, meperidine, meptazinol, methadone, morphine, nalbuphine, nalorphine, opium, oxycodone, oxymorphone, pentazocine, propoxyphene, remifentanil, sufentanil, tapentadol, and tramadol, and pharmaceutically acceptable salts thereof.BRIEF DESCRIPTION OF THE DRAWINGS

[0064] Fig.1 illustrates withdrawal behaviors elicited by a bolus injection of naloxone HCl (1.5 mg / kg, IP) in rats treated for 36h with a subcutaneous depot of morphine (150 mg / kg) along with continuous infusion of vehicle (saline, 20 μL / h, IV) or L-cysteine (20.8 μmol / kg / h, IV) or L-cysteine ethylester (L-CYSee, 20.8 μmol / kg / h, IV). Withdrawal Signs: Jumps - all 4 paws off the floor; WDS, wet-dog shakes; Rears - rearing on hind legs; FPL - episodes of fore-paw licking; Circles, a 360orotation; Writhes, fully body contortion; Sneezes, abrupt expulsion of air. The data are presented as mean ± SEM. There were 9 rats in each group. ANOVA statistics: Jumps: F2,24 = 18.8, P = 0.00001; WDS: F2,24 = 29.9, P < 0.00001; Rears: F2,24= 16.2, P = 0.00004; FPL: F2,24= 27.0, P < 0.00001; Circles: F2,24= 24.1, P < 0.00001; Writhes: F2,24 = 15.4, P < 0.00005; Sneezes: F2,24 = 0.09, P = 0.92. *P < 0.05, significant responses.†P < 0.05, L-CYSee or L-cysteine versus vehicle.

[0065] Fig.2 illustrates incidence of apneas (> 1.5 sec between breaths) and increases in mean arterial blood pressure (MAP, mmHg) and heart rate (HR, beats / min) elicited by a bolus injection of naloxone HCl (1.5 mg / kg, IV) in rats treated for 36h with a subcutaneous depot of morphine (150 mg / kg) along with continuous infusion of vehicle (saline, 20 μL / h, IV), L-cysteine (20.8 μmol / kg / h, IV) or L-cysteine ethyl ester (L-CYSee, 20.8 μmol / kg / h, IV). The data are presented as mean ± SEM. There were 9 rats in each group. ANOVA statistics: MAP: F2,24 = 28.5, P < 0.00001; HR: F2,24 = 47.8, P < 0.00001; Apneas: F2,24 = 37.2, P < 0.00001. *P < 0.05, significant responses.†P < 0.05, L-CYSee versus vehicle or L- cysteine.

[0066] Figs.3(A-B) illustrate arithmetic changes in body temperature (Panel A) and body weights (Panel B) elicited by a bolus injection of naloxone HCl (1.5 mg / kg, IP) in rats treated for 24h with a subcutaneous depot of morphine (100 mg / kg) or for 48h with a subcutaneous depot of morphine (150 mg / kg) with or without continuous infusion of L- cysteine (20.8 μmol / kg / h, IV) or L-CYSee (20.8 μmol / kg / h, IV). The data are presented as mean ± SEM. There were 9 rats in each group. ANOVA statistics: Body Temperature: 36h: F2,24 = 51.4, P < 0.00001; 48h: F2,24 = 50.1, P < 0.00001; Body Weight 36h: F2,24 = 22.8, < 0.00001; 48h: F2,24= 18.6, P < 0.00001. *P < 0.05, significant responses. †P < 0.05, L-CYSee or L-cysteine versus vehicle.

[0067] Fig.4 illustrates withdrawal behaviors elicited by a bolus injection of naloxone HCl (1.5 mg / kg, IP) in rats treated for 48h with a subcutaneous depot of morphine (150 mg / kg) along with continuous infusion of vehicle (saline, 20 μL / h, IV) or L-cysteine (20.8 μmol / kg / h, IV) or L-cysteine ethylester (L-CYSee, 20.8 μmol / kg / h, IV) at began after 36 of morphine administration. Withdrawal Signs: Jumps - all 4 paws off the floor; WDS, wet-dog shakes; Rears - rearing on hind legs; FPL - episodes of fore-paw licking; Circles, a 360orotation; Writhes, fully body contortion; Sneezes, abrupt expulsion of air. The data are presented as mean ± SEM. There were 9 rats in each group. ANOVA statistics: Jumps: F2,24= 48.8, P < 0.00001; WDS: F2,24 = 33.9, P < 0.00001; Rears: F2,24 = 38.7, < 0.00001; FPL: F2,24 = 42.3, P < 0.00001; Circles: F2,24 = 29.4, P < 0.00001; Writhes: F2,24 = 34.0, P < 0.00005; Sneezes: F2,24 = 0.11, P = 0.89. *P < 0.05, significant responses.†P < 0.05, L- CYSee or L-cysteine versus vehicle.

[0068] Fig.5 illustrates incidence of apneas (> 1.5 sec between breaths) and transient increases in mean arterial blood pressure (MAP, mmHg) and heart rate (HR, beats / min) elicited by a bolus injection of naloxone HCl (1.5 mg / kg, IV) in rats treated for 48h with a subcutaneous depot of morphine (150 mg / kg) along with continuous infusion of vehicle (saline, 20 μL / h, IV), L-cysteine (20.8 μmol / kg / h, IV) or L-cysteine ethyl ester (L-CYSee, 20.8 μmol / kg / h, IV) that began at 12h of morphine administration. The data are presented as mean ± SEM. There were 9 rats in each group. ANOVA statistics: MAP: F2,24 = 33.3, P < 0.00001; HR: F2,24 = 41.8, P < 0.00001; Apneas: F2,24 = 69.9, P < 0.00001. *P < 0.05, significant responses.†P < 0.05, L-CYSee versus vehicle or L-cysteine.

[0069] Figs.6(A-D) illustrate responses elicited by injection of naloxone HCl (1.5 mg / kg, IV) in rats that had received 5 injections of fentanyl (125 µg / kg, IV) plus 5 co- injections of vehicle, N-acetyl-L-cysteine (L-NAC, 500 µmol / kg, IV) or L-NAC methyl ester (L-NACme, 500 µmol / kg, IV). Panel A: Behavioral responses – jumps, wet-dog shakes (WDS), rears, fore-paw licking (FPL), circles, writhes, sneezes. Panel B: Cardiorespiratory responses, mean arterial blood pressure (MAP), heart rate and apneas. Panel C: Body temperature. Panel D: Body weights Data are shown as mean ± SEM. There were 9 rats in each group. *P < 0.05, significant response.†P < 0.05, L-NAC or L-NACme versus vehicle. P < 0.05, L-NACme versus L-NAC.

[0070] Figs.7(A-D) illustrate responses elicited by injection of naloxone HCl (1.5 mg / kg, IV) in rats that had received 10 injections of fentanyl (125 µg / kg, IV) plus 10 co- injections of vehicle, N-acetyl-L-cysteine (L-NAC, 500 µmol / kg, IV) or L-NAC methyl ester (L-NACme, 500 µmol / kg, IV). Panel A: Behavioral responses – jumps, wet-dog shakes (WDS), rears, fore-paw licking (FPL), circles, writhes, sneezes. Panel B: Cardiorespiratory responses, mean arterial blood pressure (MAP), heart rate and apneas. Panel C: Body temperature. Panel D: Body weights. Data are shown as mean ± SEM. There were 9 rats in each group. *P < 0.05, significant response.†P < 0.05, L-NAC or L-NACme versus vehicle. P < 0.05, L-NACme versus L-NAC.

[0071] Figs.8(A-D) illustrate responses elicited by injection of naloxone HCl (1.5 mg / kg, IV) in rats that had received 10 injections of fentanyl (125 µg / kg, IV) plus 5 co- injections of vehicle, N-acetyl-L-cysteine (L-NAC, 500 µmol / kg, IV) or L-NAC methyl ester (L-NACme, 500 µmol / kg, IV) beginning at fentanyl injection 6. Panel A: Behavioral responses – jumps, wet-dog shakes (WDS), rears, fore-paw licking (FPL), circles, writhes, sneezes. Panel B: Cardiorespiratory responses, mean arterial blood pressure (MAP), heart rate and apneas. Panel C: Body temperature. Panel D: Body weights. Data are shown as mean ± SEM. There were 9 rats in each group. *P < 0.05, significant response.†P < 0.05, L-NAC or L-NACme versus vehicle. P < 0.05, L-NACme versus L-NAC.

[0072] Fig.9 illustrates chemical structures of L-NAC and analogues.

[0073] Figs.10(A-D) illustrate responses elicited by the injection of naloxone HCl (1.5 mg / kg, IV) in rats that had received 5 injections of fentanyl (125 µg / kg, IV) plus 5 co- injections of vehicle, D-cysteine (250 µmol / kg, IV), D-cysteine ethyl ester (D-CYSee, 250 µmol / kg, IV), or D-cysteine ethyl amide (D-CYSea, 100 µmol / kg, IV). Panel A: Behavioral responses – jumps, wet-dog shakes (WDS), rears, fore-paw licking (FPL), circles, writhes, sneezes. Panel B: Cardiorespiratory responses, mean arterial blood pressure (MAP), heart rate and apneas. Panel C: Body temperature. Panel D: Body weights Data are shown as mean ± SEM. There were 9 rats in each group. ANOVA statistics: Panel A: Jumps (F3,32 = 28.0, P < 0.0001); WDS (F3,32 = 22.2, P < 0.0001); Rears (F3,32 = 47.3, P < 0.0001); FPL (F3,32 = 24.3, P < 0.0001); Circles (F3,32= 20.5, P = 0.0009); Writhes (F3,32= 20.0, P < 0.0001); Sneezes (F3,32 = 4.34, P = 0.011). Panel B: MAP (F3,32 = 31.7, P < 0.0001); Heart rate (F3,32 =38.2, P < 0.0001); Apneas (F3,32 = 32.7, P < 0.0001). Panel C: Body temperature (F3,32 = 23.5, P < 0.0001). Panel D: Body weights (F3,32 = 18.1, P < 0.0001). *P < 0.05, significant response.†P < 0.05, D-CYSea or D-CYSee versus vehicle. P < 0.05, D-CYSea versus D- CYSee.

[0074] Figs.11(A-D) illustrates responses elicited by the injection of naloxone HCl (1.5 mg / kg, IV) in rats that had received 10 injections of fentanyl (125 µg / kg, IV) plus 10 co- injections of vehicle, D-cysteine (250 µmol / kg, IV), D-cysteine ethyl ester (D-CYSee, 250 µmol / kg, IV), or D-cysteine ethyl amide (D-CYSea, 100 µmol / kg, IV). Panel A: Behavioral responses – jumps, wet-dog shakes (WDS), rears, fore-paw licking (FPL), circles, writhes, sneezes. Panel B: Cardiorespiratory responses, mean arterial blood pressure (MAP), heart rate and apneas. Panel C: Body temperature. Panel D: Body weights. Data are shown as mean ± SEM. There were 9 rats in each group. ANOVA statistics: Panel A: Jumps (F3,32 = 48.5, P < 0.0001); WDS (F3,32= 38.7, P < 0.0001); Rears (F3,32= 34.7, P < 0.0001); FPL (F3,32= 42.8, P < 0.0001); Circles (F3,32= 19.3, P = 0.0009); Writhes (F3,32= 30.2, P < 0.0001); Sneezes (F3,32 = 6.65, P = 0.001). Panel B: MAP (F3,32 = 37.6, P < 0.0001); Heart rate (F3,32 = 88.4, P < 0.0001); Apneas (F3,32= 25.8, P < 0.0001). Panel C: Body temperature (F3,32= 48.4, P < 0.0001). Panel D: Body weights (F3,32= 18.5, P < 0.0001).*P < 0.05, significant response.†P < 0.05, D-CYSea or D-CYSee versus vehicle. P < 0.05, D-CYSea versus D- CYSee.

[0075] Figs.12(A-D) illustrate responses elicited by the injection of naloxone HCl (1.5 mg / kg, IV) in rats that had received 10 injections of fentanyl (125 µg / kg, IV) plus 5 co- injections of vehicle, D-cysteine (250 µmol / kg, IV), D-cysteine ethyl ester (D-CYSee, 250 µmol / kg, IV), or D-cysteine ethyl amide (D-CYSea, 100 µmol / kg, IV) beginning at fentanyl injection 6. Panel A: Behavioral responses – jumps, wet-dog shakes (WDS), rears, fore-paw licking (FPL), circles, writhes, sneezes. Panel B: Cardiorespiratory responses, mean arterial blood pressure (MAP), heart rate and apneas. Panel C: Body temperature. Panel D: Body weights. Data are shown as mean ± SEM. There were 9 rats in each group. ANOVA statistics: Panel A: Jumps (F2,24 = 36.2, P < 0.0001); WDS (F2,24 = 46.8, P < 0.0001); Rears (F2,24 = 35.8, P < 0.0001); FPL (F2,24 = 29.8, P < 0.0001); Circles (F2,24 = 32.2, P = 0.0009); Writhes (F2,24= 33.1, P < 0.0001); Sneezes (F2,24= 8.6, P = < 0.0003). Panel B: MAP (F2,24=31.9, P < 0.0001); Heart rate (F2,24 = 45.3, P < 0.0001); Apneas (F2,24 = 52.7, P < 0.0001). Panel C: Body temperature (F2,24 = 86.6, P < 0.0001). Panel D: Body weights (F2,24 = 22.4, P < 0.0001). *P < 0.05, significant response.†P < 0.05, D-CYSea or D-CYSee versus vehicle. P < 0.05, D-CYSea versus D-CYSee.

[0076] Fig.13 illustrates withdrawal behaviors elicited by a bolus injection of naloxone HCl (1.5 mg / kg, IP) in rats treated for 36h with a subcutaneous depot of morphine (150 mg / kg) along with continuous infusion of vehicle (saline, 20 μL / h, IV) or D-cysteine (20.8 μmol / kg / h, IV) or D-cysteine ethyl ester (D-CYSee, 20.8 μmol / kg / h, IV). Withdrawal Signs: Jumps - all 4 paws off the floor; WDS, wet-dog shakes; Rears - rearing on hind legs; FPL - episodes of fore-paw licking; Circles, a 360orotation; Writhes, fully body contortion; Sneezes, abrupt expulsion of air. The data are presented as mean ± SEM (9 rats per group). *P < 0.05, significant responses.†P < 0.05, D-CYSee or D-cysteine versus vehicle.

[0077] Fig.14 illustrates the incidence of apneas (> 1.5 sec) and transient increases in mean arterial blood pressure (MAP, mmHg) and heart rate (HR, beats / min) elicited by a bolus injection of naloxone HCl (1.5 mg / kg, IV) in rats treated for 36h with a subcutaneous depot of morphine (150 mg / kg) along with continuous infusion of vehicle (saline, 20 μL / h, IV), D-cysteine (20.8 μmol / kg / h, IV) or D-cysteine ethyl ester (D-CYSee, 20.8 μmol / kg / h, IV). The data are presented as mean ± SEM (9 rats per group). *P < 0.05, significant responses.†P < 0.05, D-CYSee versus vehicle or D-cysteine.

[0078] Figs.15(A-B) illustrate arithmetic changes in body temperature (Panel A) and body weights (Panel B) elicited by a bolus injection of naloxone HCl (1.5 mg / kg, IP) in rats treated for 24h with a subcutaneous depot of morphine (100 mg / kg) or for 48h with a subcutaneous depot of morphine (150 mg / kg) with or without continuous infusion of D- cysteine (20.8 μmol / kg / h, IV) or D-cysteine ethyl ester (D-CYSee, 20.8 μmol / kg / h, IV). The data are presented as mean ± SEM (9 rats per group). *P < 0.05, significant responses. †P < 0.05, D-CYSee or D-cysteine versus vehicle.

[0079] Fig.16 illustrates withdrawal behaviors elicited by a bolus injection of naloxone HCl (1.5 mg / kg, IP) in rats treated for 48h with a subcutaneous depot of morphine (150 mg / kg) along with continuous infusion of vehicle (saline, 20 μL / h, IV) or D-cysteine (20.8 μmol / kg / h, IV) or D-cysteine ethylester (D-CYSee, 20.8 μmol / kg / h, IV) at began after 36 of morphine administration. Withdrawal Signs: Jumps - all 4 paws off the floor; WDS, wet-dogshakes; Rears - rearing on hind legs; FPL - episodes of fore-paw licking; Circles, a 360orotation; Writhes, fully body contortion; Sneezes, abrupt expulsion of air. The data are presented as mean ± SEM (9 rats per group). *P < 0.05, significant responses.†P < 0.05, D- CYSee or D-cysteine versus vehicle.

[0080] Fig.17 illustrates incidence of apneas (> 1.5 sec) and transient repetitive increases in mean arterial blood pressure (MAP, mmHg) and heart rate (HR, beats / min) elicited by a bolus injection of naloxone HCl (1.5 mg / kg, IV) in rats treated for 48h with a subcutaneous depot of morphine (150 mg / kg) along with continuous infusion of vehicle (saline, 20 μL / h, IV), D-cysteine (20.8 μmol / kg / h, IV) or D-cysteine ethyl ester (D-CYSee, 20.8 μmol / kg / h, IV) that began at 12h of morphine administration. The data are presented as mean ± SEM (9 rats per group). *P < 0.05, significant responses.†P < 0.05, D-CYSee versus vehicle or D-cysteine.

[0081] Figs.18(A-E) illustrate the changes in tail-flick latencies expressed as maximal possible effect (MPE, %) elicited by co-injections 1-10 of fentanyl (125 µg / kg, IV) + vehicle or fentanyl (125 µg / kg, IV) + naloxone HCl (NLX, 5 mg / kg, IV). The data are presented as mean ± SEM. There were 9 rats in each group. *P < 0.05, significant response.†P < 0.05, NLX versus vehicle.

[0082] Figs.19(A-E) illustrate the changes in tail-flick latencies expressed as maximal possible effect (MPE, %) elicited by co-injections 6-10 of fentanyl (125 µg / kg, IV) + vehicle or fentanyl (125 µg / kg, IV) + naloxone HCl (NLX, 5 mg / kg, IV). The data are presented as mean ± SEM. There were 9 rats in each group. *P < 0.05, significant response.†P < 0.05, NLX versus vehicle.

[0083] Figs.20(A-C) illustrate total (cumulative) changes in tail-flick latencies for analgesia or hyperalgesia expressed as the sum of the maximal possible effect (MPE, %) elicited by co-injections 1-10 of fentanyl (125 µg / kg, IV) + vehicle or fentanyl (125 µg / kg, IV) + naloxone HCl (NLX, 5 mg / kg, IV). C: Changes in Pre-values for tail-flick latencies expressed as arithmetic difference from Pre-injection 1. The data are presented as mean ± SEM. There were 9 rats in each group. *P < 0.05, significant response.†P < 0.05, NLX versus vehicle.

[0084] Figs.21(A-E) illustrate the changes in tail-flick latencies expressed as maximal possible effect (MPE, %) elicited by co-injections 1-5 of fentanyl (125 µg / kg, IV) + vehicleor fentanyl (125 µg / kg, IV) + N-acetyl-L-cysteine (L-NAC, 500 µmol / kg, IV) or fentanyl (125 µg / kg, IV) + L-NAC methyl ester (L-NACme, 500 µmol / kg, IV). The data are presented as mean ± SEM. There were 9 rats in each group. *P < 0.05, significant response.†P < 0.05, L-NAC or L-NACme versus vehicle. P < 0.05, L-NACme versus L-NAC.

[0085] Figs.22(A-E) illustrate the changes in tail-flick latencies expressed as maximal possible effect (MPE, %) elicited by co-injections 6-10 of fentanyl (125 µg / kg, IV) + vehicle or fentanyl (125 µg / kg, IV) + N-acetyl-L-cysteine (L-NAC, 500 µmol / kg, IV) or fentanyl (125 µg / kg, IV) + L-NAC methyl ester (L-NACme, 500 µmol / kg, IV). The data are presented as mean ± SEM. There were 9 rats in each group. *P < 0.05, significant response.†P < 0.05, L-NAC or L-NACme versus vehicle. P < 0.05, L-NACme versus L-NAC.

[0086] Figs.23(A-B) illustrate the total (cumulative) changes in tail-flick latencies for analgesia (Panel A) and for hyperalgesia (Panel B) expressed as the sum of the maximal possible effect (MPE, %) values elicited by co-injections 1-10 of fentanyl (125 µg / kg, IV) + vehicle or fentanyl (125 µg / kg, IV) + N-acetyl-L-cysteine (L-NAC, 500 µmol / kg, IV) or fentanyl (125 µg / kg, IV) + L-NAC methyl ester (L-NACme, 500 µmol / kg, IV). The data are s mean ± SEM. There were 9 rats in each group. *P < 0.05, significant response.†P < 0.05, L-NAC or L-NACme versus vehicle. P < 0.05, L-NACme versus L-NAC.

[0087] Figs.24(A-B) illustrate (A) Baseline tail-flick latencies prior to each set of co- injections of fentanyl (125 µg / kg, IV) + vehicle or fentanyl (125 µg / kg, IV) + N-acetyl-L- cysteine (L-NAC, 500 µmol / kg, IV) or fentanyl (125 µg / kg, IV) + L-NAC methyl ester (L- NACme, 500 µmol / kg, IV). B: Changes in Pre-values for tail-flick latencies expressed as arithmetic difference from Pre-injection 1. The data are presented as mean ± SEM. There were 9 rats in each group. *P < 0.05, significant response.†P < 0.05, L-NAC or L-NACme versus vehicle. P < 0.05, L-NACme versus L-NAC.

[0088] Fig.25 are chemical structures of L-NAC and analogues.

[0089] Figs.26(A-H) illustrate the changes in tail-flick latencies expressed as maximal possible effect (MPE, %) elicited by co-injections 1-10 of vehicle + vehicle or vehicle + N- acetyl-L-cysteine (L-NAC, 500 µmol / kg, IV) or vehicle + L-NAC methyl ester (L-NACme, 500 µmol / kg, IV). The data are presented as mean ± SEM. There were 9 rats in each group.There were no differences from Pre values for ant administration protocol (P > 0.05, for all responses).

[0090] Figs.27(A-B) illustrate baseline tail-flick latencies prior to each set of co- injections of fentanyl (125 µg / kg, IV) + vehicle or fentanyl (125 µg / kg, IV) + naloxone HCl (NLX, 5 mg / kg, IV). Panel B: Changes in Pre-values for tail-flick latencies expressed as arithmetic difference from Pre-injection 1. The data are presented as mean ± SEM. There were 9 rats in each group. There were no changes in baseline values from the initial set of co-injections (P > 0.05, for all responses).

[0091] Fig.28 illustrates Co-administration of L-NAC prevents acquisition of fentanyl- induced reward seeking in rats. Treatment with L-NAC (100 mg / kg, i.p.) 30 min or 120 min prior to fentanyl conditioning (5 µg / kg, s.c.) prevents acquisition of fentanyl conditioned place preference compared to saline treatment. *p <0.01. DETAILED DESCRIPTION

[0092] For convenience, certain terms employed in the specification, examples, and appended claims are collected here. Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this application belongs.

[0093] The articles "a" and "an" are used herein to refer to one or to more than one (i.e., to at least one) of the grammatical object of the article. By way of example, "an element" means one element or more than one element.

[0094] The terms "comprise," "comprising," "include," "including," "have," and "having" are used in the inclusive, open sense, meaning that additional elements may be included. The terms "such as", "e.g.," as used herein are non-limiting and are for illustrative purposes only. "Including" and "including but not limited to" are used interchangeably.

[0095] The term "or" as used herein should be understood to mean "and / or", unless the context clearly indicates otherwise.

[0096] The term "about" or "approximately" as used herein refers to a quantity, level, value, number, frequency, percentage, dimension, size, amount, weight or length that varies by as much as 15%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2% or 1% to a reference quantity, level, value, number, frequency, percentage, dimension, size, amount, weight or length. In one embodiment, the term "about" or "approximately" refers a range of quantity, level, value,number, frequency, percentage, dimension, size, amount, weight or length ± 15%, ± 10%, ± 9%, ± 8%, ± 7%, ± 6%, ± 5%, ± 4%, ± 3%, ± 2%, or ± 1% about a reference quantity, level, value, number, frequency, percentage, dimension, size, amount, weight or length.

[0097] It will be noted that the structure of some of the compounds of the application include asymmetric (chiral) carbon or sulfur atoms. It is to be understood accordingly that the isomers arising from such asymmetry are included herein, unless indicated otherwise. Such isomers can be obtained in substantially pure form by classical separation techniques and by stereochemically controlled synthesis. The compounds of this application may exist in stereoisomeric form, therefore, can be produced as individual stereoisomers or as mixtures.

[0098] The term "isomerism" refers to compounds that have identical molecular formulae but that differ in the nature or the sequence of bonding of their atoms or in the arrangement of their atoms in space. Isomers that differ in the arrangement of their atoms in space are termed "stereoisomers". Stereoisomers that are not mirror images of one another are termed "diastereoisomers", and stereoisomers that are non-superimposable mirror images are termed "enantiomers", or sometimes optical isomers. A carbon atom bonded to four nonidentical substituents is termed a "chiral center" whereas a sulfur bound to three or four different substituents, e.g., sulfoxides or sulfinimides, is likewise termed a “chiral center”.

[0099] The term "chiral isomer" refers to a compound with at least one chiral center. It has two enantiomeric forms of opposite chirality and may exist either as an individual enantiomer or as a mixture of enantiomers. A mixture containing equal amounts of individual enantiomeric forms of opposite chirality is termed a "racemic mixture". A compound that has more than one chiral center has 2n-1 enantiomeric pairs, where n is the number of chiral centers. Compounds with more than one chiral center may exist as either an individual diastereomer or as a mixture of diastereomers, termed a "diastereomeric mixture". When one chiral center is present, a stereoisomer may be characterized by the absolute configuration (R or S) of that chiral center. Alternatively, when one or more chiral centers are present, a stereoisomer may be characterized as (+) or (-). Absolute configuration refers to the arrangement in space of the substituents attached to the chiral center. The substituents attached to the chiral center under consideration are ranked in accordance with the Sequence Rule of Cahn, Ingold and Prelog. (Cahn et al, Angew. Chem. Inter. Edit.1966, 5, 385; errata 511; Cahn et al., Angew. Chem.1966, 78, 413; Cahn and Ingold, J Chem. Soc.1951 (London), 612; Cahn et al., Experientia 1956, 12, 81; Cahn, J., Chem. Educ.1964, 41, 116).

[0100] The term "geometric isomers" refers to diastereomers that owe their existence to hindered rotation about double bonds. These configurations are differentiated in their names by the prefixes cis and trans, or Z and E, which indicate that the groups are on the same or opposite side of the double bond in the molecule according to the Cahn-Ingold-Prelog rules. Further, the structures and other compounds discussed in this application include all atropic isomers thereof.

[0101] The term "atropic isomers" refers to a type of stereoisomer in which the atoms of two isomers are arranged differently in space. Atropic isomers owe their existence to a restricted rotation caused by hindrance of rotation of large groups about a central bond. Such atropic isomers typically exist as a mixture, however as a result of recent advances in chromatography techniques, it has been possible to separate mixtures of two atropic isomers in select cases.

[0102] The terms "addiction" and "dependence" are used interchangeably to refer to the patient's inability to stop using the opioid or opioid-like drug, even when it would be in his / her best interest to stop. The Diagnostic and Statistical Manual of Mental Disorders Text Revision (DSMIV-TR) criteria for dependency include: Dependence or significant impairment or distress, as manifested by 3 or more of the following during a 12 month period: 1. Tolerance or markedly increased amounts of the substance to achieve intoxication or desired effect or markedly diminished effect with continued use of the same amount of substance. 2. Withdrawal symptoms or the use of certain substances to avoid withdrawal symptoms. 3. Use of a substance in larger amounts or over a longer period than was intended. 4. Persistent desire or unsuccessful efforts to cut down or control substance use. 5. Involvement in chronic behavior to obtain the substance, use the substance, or recover from its effects. 6. Reduction or abandonment of social, occupational or recreational activities because of substance use. 7. Use of substances even though there is a persistent or recurrent physical or psychological problem that is likely to have been caused or exacerbated by the substance.

[0103] "Drug withdrawal" refers to a group of symptoms that occur upon the abrupt discontinuation or sudden decrease in intake of medications or recreational drugs. Consequently, "opioid withdrawal" refers to the group of symptoms that occur upon the dramatic reduction, abrupt discontinuation or decrease in intake of opioids or opiates. Withdrawal symptoms may also start between doses. Withdrawal symptoms from opioidsinclude but are not limited to anxiety, depression, sweating, vomiting, and diarrhea, muscle cramping, agitation, insomnia, yawning dilated pupils, goose bumps, abdominal cramping, runny nose and increased tearing, for example.

[0104] The term "opioid" refers to naturally-occurring opiates and synthetic or semi- synthetic opioids that have psychoactive effects. Non-limiting examples include acetyl- alpha-methylphentanyl, acetylmethadol, alfentanil, allylprodine, alphacetylmethadol, alphamethadol, alpha-methylfentanyl, alpha-methylthiofentanyl, alphaprodine, anileridine, benzylmorphine, benzethidine, betacetylmethadol, beta-hydroxyfentanyl, beta-hydroxy-3- methylfentanyl, betameprodine, betacetylmethadol, beta-hydroxyfentanyl, beta-hydroxy-3- methylfentanyl, betameprodine, betamethadol, betaprodine, bezitramide, buprenorphine, butorphanol, carfentanil, clonitazene, codeine, desomorphine, dextromoramide, dextropropoxyphene, dezocine, diampromide, diamorphone, diethylthiambutene, dihydrocodeine, dihydroetorphine, dihydromorphine, dimenoxadol, dimepheptanol, dimethyl- thiambutene, dioxaphetyl butyrate, diphenoxylate, difenoxin, dipipanone, eptazocine, ethoheptazine, ethylmethylthiambutene, ethylmorphine, etonitazene, etorphine, etoxeridine, fentanyl, furethidine, heroin, hydrocodone, hydromorphone, hydroxypethidine, isomethadone, ketobemidone, levo-alphacetylmethadol, levomethorphan, levorphanol, levophenacylmorphan, levomoranude, lofentanil, loperamide, laudanum, meperidine, meptazinol, metazocine, methadone, 3-methylfentanyl, 3-methylthiofentanyl, metopon, morphine, morpheridine, MPPP (1-methyl-4-phenyl-4-propionoxypiperidine), myrophine, narceine, nicomorphine, noracymethadol, norlevorphanol, normethadone, nalorphine, nalbuphene, normorphine, norpipanone, opium, oxycodone, oxymorphone, papaveretum, para-fluorofentanyl, paregoric, PEPAP (1-(-2-phenethyl)-4-phenyl-4-acetoxypiperidine), pentazocine, phenadoxone, phenampromide, phenomorphan, phenazocine, phenoperidine, piminodine, piritramide, propheptazine, promedol, properidine, propiram, propoxyphene, racemoramide, racemethorphan, racemorphan, remifentanil, sufentanil, tapentadol, thebaine, thiofentanyl, tilidine, tramadol, trimeperidine, mixtures of any of the foregoing, salts of any of the foregoing, derivatives of any of the foregoing, and the like. The term opioids also encompasses opioid intermediates, including 4-cyano-2-dimethylamino-4,4-diphenyl butane, 2-methyl-3-morpholino-1,1-diphenylpropane-carboxylic acid, 4-cyano-1-methyl-4- phenylpiperidine, ethyl-4-phenylpiperidine-4-carboxylate, and 1-methyl-4-phenylpiperidine- 4-carboxylic acid.

[0105] Neonatal abstinence syndrome (NAS) is a complex of signs and symptoms in the postnatal period associated with the sudden withdrawal of maternally transferred opioids. The main manifestations include increased muscle tone, autonomic instability, irritability, poor sucking reflex, and impaired weight increase.

[0106] The phrases "parenteral administration" and "administered parenterally" are art- recognized terms, and include modes of administration other than enteral and topical administration, such as injections, and include, without limitation, intravenous, intramuscular, intrapleural, intravascular, intrapericardial, intraarterial, intrathecal, intracapsular, intraorbital, intracardiac, intradermal, intraperitoneal, transtracheal, subcutaneous, subcuticular, intra-articular, subcapsular, subarachnoid, intraspinal and intrastemal injection and infusion.

[0107] The terms “treatment," "treating," and "treat" refer to acting upon a disease, disorder, or condition with an agent to reduce or ameliorate harmful or any other undesired effects of the disease, disorder, or condition and / or its symptoms. "Treatment," as used herein, covers the treatment of a human patient, and includes: (a) reducing the risk of occurrence of the condition in a patient determined to be predisposed to the condition but not yet diagnosed as having the condition, (b) impeding the development of the condition, and / or (c) relieving the condition, i.e., causing regression of the condition and / or relieving one or more symptoms of the condition. "Treating" or "treatment of" a condition or patient refers to taking steps to obtain beneficial or desired results, including clinical results such as the reduction of symptoms. For purposes of this invention, beneficial or desired clinical results include, but are not limited to: treating opioid or opioid-like drug addiction; treating, preventing, and / or attenuating acute withdrawal symptoms; treating, preventing, and / or attenuating long-term (post-acute) withdrawal symptoms; and preventing relapse of opioid or opioid-like drug use.

[0108] The term "preventing" is art-recognized and includes stopping a disease, disorder or condition from occurring in a subject, which may be predisposed to the disease, disorder and / or condition but has not yet been diagnosed as having it. Preventing a condition related to a disease includes stopping the condition from occurring after the disease has been diagnosed but before the condition has been diagnosed.

[0109] The term "pharmaceutical composition" refers to a formulation containing the disclosed compounds in a form suitable for administration to a subject. In someembodiments, the pharmaceutical composition is in bulk or in unit dosage form. The unit dosage form is any of a variety of forms, including, for example, a capsule, an IV bag, a tablet, a single pump on an aerosol inhaler, or a vial. The quantity of active ingredient (e.g., a formulation of the disclosed compound or salts thereof) in a unit dose of composition is an effective amount and is varied according to the particular treatment involved. One skilled in the art will appreciate that it is sometimes necessary to make routine variations to the dosage depending on the age and condition of the patient. The dosage will also depend on the route of administration. A variety of routes are contemplated, including oral, pulmonary, rectal, parenteral, transdermal, subcutaneous, intravenous, intramuscular, intraperitoneal, intranasal, inhalational, and the like. Dosage forms for the topical or transdermal administration of a compound described herein includes powders, sprays, ointments, pastes, creams, lotions, gels, solutions, patches, nebulized compounds, and inhalants. In some embodiments, the compound or active ingredient is mixed under sterile conditions with a pharmaceutically acceptable carrier, and with any preservatives, buffers, or propellants that are required.

[0110] The term "flash dose" refers to compound formulations that are rapidly dispersing dosage forms.

[0111] The term "immediate release" is defined as a release of compound from a dosage form in a relatively brief period of time, generally up to about 60 minutes. The term "modified release" is defined to include delayed release, extended release, and pulsed release. The term "pulsed release" is defined as a series of releases of drug from a dosage form. The term "sustained release" or "extended release" is defined as continuous release of a compound from a dosage form over a prolonged period.

[0112] The phrase "pharmaceutically acceptable" is art-recognized. In certain embodiments, the term includes compositions, polymers and other materials and / or dosage forms which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of human beings and animals without excessive toxicity, irritation, allergic response, or other problem or complication, commensurate with a reasonable benefit / risk ratio.

[0113] The phrase "pharmaceutically acceptable carrier" is art-recognized, and includes, for example, pharmaceutically acceptable materials, compositions or vehicles, such as a liquid or solid filler, diluent, excipient, solvent or encapsulating material, involved in carrying or transporting any subject composition from one organ, or portion of the body, toanother organ, or portion of the body. Each carrier must be "acceptable" in the sense of being compatible with the other ingredients of a subject composition and not injurious to the patient. In certain embodiments, a pharmaceutically acceptable carrier is non-pyrogenic. Some examples of materials, which may serve as pharmaceutically acceptable carriers include: (1) sugars, such as lactose, glucose and sucrose; (2) starches, such as corn starch and potato starch; (3) cellulose, and its derivatives, such as sodium carboxymethyl cellulose, ethyl cellulose and cellulose acetate; (4) powdered tragacanth; (5) malt; (6) gelatin; (7) talc; (8) excipients, such as cocoa butter and suppository waxes; (9) oils, such as peanut oil, cottonseed oil, sunflower oil, sesame oil, olive oil, corn oil and soybean oil; (10) glycols, such as propylene glycol; (11) polyols, such as glycerin, sorbitol, mannitol and polyethylene glycol; (12) esters, such as ethyl oleate and ethyl laurate; (13) agar; (14) buffering agents, such as magnesium hydroxide and aluminum hydroxide; (15) alginic acid; (16) pyrogen-free water; (17) isotonic saline; (18) Ringer's solution; (19) ethyl alcohol; (20) phosphate buffer solutions; and (21) other non-toxic compatible substances employed in pharmaceutical formulations.

[0114] The compounds of the application are capable of further forming salts. All of these forms are also contemplated herein.

[0115] "Pharmaceutically acceptable salt" of a compound means a salt that is pharmaceutically acceptable and that possesses the desired pharmacological activity of the parent compound. For example, the salt can be an acid addition salt. One embodiment of an acid addition salt is a hydrochloride salt. The pharmaceutically acceptable salts can be synthesized from a parent compound that contains a basic or acidic moiety by conventional chemical methods. Generally, such salts can be prepared by reacting the free acid or base forms of these compounds with a stoichiometric amount of the appropriate base or acid in water or in an organic solvent, or in a mixture of the two; generally, non-aqueous media like ether, ethyl acetate, ethanol, isopropanol, or acetonitrile being preferred. Lists of salts are found in Remington's Pharmaceutical Sciences, 18th ed. (Mack Publishing Company, 1990).

[0116] The compounds described herein can also be prepared as esters, for example pharmaceutically acceptable esters. For example, a carboxylic acid function group in a compound can be converted to its corresponding ester, e.g., a methyl, ethyl, or other ester. Also, an alcohol group in a compound can be converted to its corresponding ester, e.g., an acetate, propionate, or other ester.

[0117] The compounds described herein can also be prepared as prodrugs, for example pharmaceutically acceptable prodrugs. The terms "pro-drug" and "prodrug" are used interchangeably herein and refer to any compound, which releases an active parent drug in vivo. Since prodrugs are known to enhance numerous desirable qualities of pharmaceuticals (e.g., solubility, bioavailability, manufacturing, etc.) the compounds can be delivered in prodrug form. Thus, the compounds described herein are intended to cover prodrugs of the presently claimed compounds, methods of delivering the same and compositions containing the same. "Prodrugs" are intended to include any covalently bonded carriers that release an active parent drug in vivo when such prodrug is administered to a subject. Prodrugs are prepared by modifying functional groups present in the compound in such a way that the modifications are cleaved, either in routine manipulation or in vivo, to the parent compound. Prodrugs include compounds wherein a hydroxy, amino, sulfhydryl, carboxy, or carbonyl group is bonded to any group that may be cleaved in vivo to form a free hydroxyl, free amino, free sulfhydryl, free carboxy or free carbonyl group, respectively. Prodrugs can also include a precursor (forerunner) of a compound described herein that undergoes chemical conversion by metabolic processes before becoming an active or more active pharmacological agent or active compound described herein.

[0118] Examples of prodrugs include, but are not limited to, esters (e.g., acetate, dialkylaminoacetates, formates, phosphates, sulfates, and benzoate derivatives) and carbamates (e.g., N,N-dimethylaminocarbonyl) of hydroxy functional groups, ester groups (e.g., ethyl esters, morpholinoethanol esters) of carboxyl functional groups, N-acyl derivatives (e.g., N-acetyl) N-Mannich bases, Schiff bases and enaminones of amino functional groups, oximes, acetals, ketals and enol esters of ketone and aldehyde functional groups in compounds, and the like, as well as sulfides that are oxidized to form sulfoxides or sulfones..

[0119] The term "protecting group" refers to a grouping of atoms that when attached to a reactive group in a molecule masks, reduces or prevents that reactivity. Examples of protecting groups can be found in Green and Wuts, Protective Groups in Organic Chemistry, (Wiley, 2.sup.nd ed.1991); Harrison and Harrison et al., Compendium of Synthetic Organic Methods, Vols.1-8 (John Wiley and Sons, 1971-1996); and Kocienski, Protecting Groups, (Verlag, 3rded.2003).

[0120] The term "amine protecting group" is intended to mean a functional group that converts an amine, amide, or other nitrogen-containing moiety into a different chemical group that is substantially inert to the conditions of a particular chemical reaction. Amine protecting groups are preferably removed easily and selectively in good yield under conditions that do not affect other functional groups of the molecule. Examples of amine protecting groups include, but are not limited to, formyl, acetyl, benzyl, t-butyldimethylsilyl, t-butyldiphenylsilyl, t-butyloxycarbonyl (Boc), p-methoxybenzyl, methoxymethyl, tosyl, trifluoroacetyl, trimethylsilyl (TMS), fluorenyl-methyloxycarbonyl, 2-trimethylsilyl- ethyoxycarbonyl, 1-methyl-1-(4-biphenylyl) ethoxycarbonyl, allyloxycarbonyl, benzyloxycarbonyl (CBZ), 2-trimethylsilyl-ethanesulfonyl (SES), trityl and substituted trityl groups, 9-fluorenylmethyloxycarbonyl (FMOC), nitro-veratryloxycarbonyl (NVOC), and the like. Those of skill in the art can identify other suitable amine protecting groups.

[0121] Representative hydroxy protecting groups include those where the hydroxy group is either acylated or alkylated such as benzyl, and trityl ethers as well as alkyl ethers, tetrahydropyranyl ethers, trialkylsilyl ethers and allyl ethers.

[0122] Additionally, the salts of the compounds described herein, can exist in either hydrated or unhydrated (the anhydrous) form or as solvates with other solvent molecules. Nonlimiting examples of hydrates include monohydrates, dihydrates, etc. Nonlimiting examples of solvates include ethanol solvates, acetone solvates, etc.

[0123] The term "solvates" means solvent addition forms that contain either stoichiometric or non-stoichiometric amounts of solvent. Some compounds have a tendency to trap a fixed molar ratio of solvent molecules in the crystalline solid state, thus forming a solvate. If the solvent is water the solvate formed is a hydrate, when the solvent is alcohol, the solvate formed is an alcoholate. Hydrates are formed by the combination of one or more molecules of water with one of the substances in which the water retains its molecular state as H2O, such combination being able to form one or more hydrate.

[0124] The compounds, salts and prodrugs described herein can exist in several tautomeric forms, including the enol and imine form, and the keto and enamine form and geometric isomers and mixtures thereof. Tautomers exist as mixtures of a tautomeric set in solution. In solid form, usually one tautomer predominates. Even though one tautomer may be described, the present application includes all tautomers of the present compounds. A tautomer is one of two or more structural isomers that exist in equilibrium and are readilyconverted from one isomeric form to another. This reaction results in the formal migration of a hydrogen atom accompanied by a switch of adjacent conjugated double bonds. In solutions where tautomerization is possible, a chemical equilibrium of the tautomers will be reached. The exact ratio of the tautomers depends on several factors, including temperature, solvent, and pH. The concept of tautomers that are interconvertable by tautomerizations is called tautomerism.

[0125] Of the various types of tautomerism that are possible, two are commonly observed. In keto-enol tautomerism a simultaneous shift of electrons and a hydrogen atom occurs.

[0126] Tautomerizations can be catalyzed by: Base: 1. deprotonation; 2. formation of a delocalized anion (e.g., an enolate); 3. protonation at a different position of the anion; Acid: 1. protonation; 2. formation of a delocalized cation; 3. deprotonation at a different position adjacent to the cation.

[0127] A "patient," "subject," or "host" to be treated by the compounds or methods described herein may mean either a human or non-human animal, such as a mammal, a fish, a bird, a reptile, or an amphibian. Thus, the subject of the herein disclosed methods can be a human, non-human primate, horse, pig, rabbit, dog, sheep, goat, cow, cat, guinea pig or rodent. The term does not denote a particular age or sex. Thus, adult and newborn subjects, as well as fetuses, whether male or female, are intended to be covered. In one aspect, the subject is a mammal. A patient refers to a subject afflicted with a disease or disorder.

[0128] The terms "prophylactic” or “therapeutic" treatment is art-recognized and includes administration to the host of one or more of the subject compounds. If it is administered prior to clinical manifestation of the unwanted condition (e.g., disease or other unwanted state of the host animal) then the treatment is prophylactic, i.e., it protects the host against developing the unwanted condition, whereas if it is administered after manifestation of the unwanted condition, the treatment is therapeutic (i.e., it is intended to diminish, ameliorate, or stabilize the existing unwanted condition or side effects thereof).

[0129] The terms "therapeutic agent", "drug", "medicament", “active ingredient”, and "bioactive substance" are art-recognized and include molecules and other agents that are biologically, physiologically, or pharmacologically active substances that act locally or systemically in a patient or subject to treat a disease or condition. The terms include without limitation pharmaceutically acceptable salts thereof and prodrugs. Such agents may beacidic, basic, or salts; they may be neutral molecules, polar molecules, or molecular complexes capable of hydrogen bonding; they may be prodrugs in the form of ethers, esters, amides and the like that are biologically activated when administered into a patient or subject.

[0130] The phrase "therapeutically effective amount" or “pharmaceutically effective amount” is an art-recognized term. In certain embodiments, the term refers to an amount of a therapeutic agent that produces some desired effect at a reasonable benefit / risk ratio applicable to any medical treatment. In certain embodiments, the term refers to that amount necessary or sufficient to eliminate, reduce or maintain a target of a particular therapeutic regimen. The effective amount may vary depending on such factors as the disease or condition being treated, the particular targeted constructs being administered, the size of the subject or the severity of the disease or condition. One of ordinary skill in the art may empirically determine the effective amount of a particular compound without necessitating undue experimentation. In certain embodiments, a therapeutically effective amount of a therapeutic agent for in vivo use will likely depend on a number of factors, including: the rate of release of an agent from a polymer matrix, which will depend in part on the chemical and physical characteristics of the polymer; the identity of the agent; the mode and method of administration; and any other materials incorporated in the polymer matrix in addition to the agent.

[0131] With respect to any chemical compounds, the present application is intended to include all isotopes of atoms occurring in the present compounds. Isotopes include those atoms having the same atomic number but different mass numbers. By way of general example and without limitation, isotopes of hydrogen include tritium and deuterium, and isotopes of carbon include C-13 and C-14.

[0132] When a bond to a substituent is shown to cross a bond connecting two atoms in a ring, then such substituent can be bonded to any atom in the ring. When a substituent is listed without indicating the atom via which such substituent is bonded to the rest of the compound of a given formula, then such substituent can be bonded via any atom in such substituent. Combinations of substituents and / or variables are permissible, but only if such combinations result in stable compounds.

[0133] When an atom or a chemical moiety is followed by a subscripted numeric range (e.g., C1-6), it is meant to encompass each number within the range as well as all intermediateranges. For example, "C1-6 alkyl" is meant to include alkyl groups with 1, 2, 3, 4, 5, 6, 1-6, 1- 5, 1-4, 1-3, 1-2, 2-6, 2-5, 2-4, 2-3, 3-6, 3-5, 3-4, 4-6, 4-5, and 5-6 carbons.

[0134] The term "alkyl" is intended to include both branched (e.g., isopropyl, tert-butyl, isobutyl), straight-chain e.g., methyl, ethyl, propyl, butyl, pentyl, hexyl, heptyl, octyl, nonyl, decyl), and cycloalkyl (e.g., alicyclic) groups (e.g., cyclopropyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl), alkyl substituted cycloalkyl groups, and cycloalkyl substituted alkyl groups. Such aliphatic hydrocarbon groups have a specified number of carbon atoms. For example, C1-6alkyl is intended to include C1, C2, C3, C4, C5, and C6alkyl groups. As used herein, "lower alkyl" refers to alkyl groups having from 1 to 6 carbon atoms in the backbone of the carbon chain. "Alkyl" further includes alkyl groups that have oxygen, nitrogen, sulfur or phosphorous atoms replacing one or more hydrocarbon backbone carbon atoms. In certain embodiments, a straight chain or branched chain alkyl has six or fewer carbon atoms in its backbone (e.g., C1-C6 for straight chain, C3-C6 for branched chain), for example four or fewer. Likewise, certain cycloalkyls have from three to eight carbon atoms in their ring structure, such as five or six carbons in the ring structure.

[0135] The term "substituted alkyls" refers to alkyl moieties having substituents replacing a hydrogen on one or more carbons of the hydrocarbon backbone. Such substituents can include, for example, alkyl, alkenyl, alkynyl, halogen, hydroxyl, alkylcarbonyloxy, arylcarbonyloxy, alkoxycarbonyloxy, aryloxycarbonyloxy, carboxylate, alkylcarbonyl, arylcarbonyl, alkoxycarbonyl, aminocarbonyl, alkylaminocarbonyl, dialkylaminocarbonyl, alkylthiocarbonyl, alkoxyl, phosphate, phosphonato, phosphinato, cyano, amino (including alkylamino, dialkylamino, arylamino, diarylamino, and alkylarylamino), acylamino (including alkylcarbonylamino, arylcarbonylamino, carbamoyl and ureido), amidino, imino, sulfhydryl, alkylthio, arylthio, thiocarboxylate, sulfates, alkylsulfinyl, sulfonato, sulfamoyl, sulfonamido, nitro, trifluoromethyl, cyano, azido, heterocyclyl, alkylaryl, or an aromatic or heteroaromatic moiety. Cycloalkyls can be further substituted, e.g., with the substituents described above. An "alkylaryl" or an "aralkyl" moiety is an alkyl substituted with an aryl (e.g., phenylmethyl (benzyl)). If not otherwise indicated, the terms "alkyl" and "lower alkyl" include linear, branched, cyclic, unsubstituted, substituted, and / or heteroatom-containing alkyl or lower alkyl, respectively.

[0136] The term "alkenyl" refers to a linear, branched or cyclic hydrocarbon group of 2 to about 24 carbon atoms containing at least one double bond, such as ethenyl, n-propenyl,isopropenyl, n-butenyl, isobutenyl, octenyl, decenyl, tetradecenyl, hexadecenyl, eicosenyl, tetracosenyl, cyclopentenyl, cyclohexenyl, cyclooctenyl, and the like. Generally, although again not necessarily, alkenyl groups can contain 2 to about 18 carbon atoms, and more particularly 2 to 12 carbon atoms. The term "lower alkenyl" refers to an alkenyl group of 2 to 6 carbon atoms, and the specific term "cycloalkenyl" intends a cyclic alkenyl group, preferably having 5 to 8 carbon atoms. The term "substituted alkenyl" refers to alkenyl substituted with one or more substituent groups, and the terms "heteroatom-containing alkenyl" and "heteroalkenyl" refer to alkenyl or heterocycloalkenyl (e.g., heterocylcohexenyl) in which at least one carbon atom is replaced with a heteroatom. If not otherwise indicated, the terms "alkenyl" and "lower alkenyl" include linear, branched, cyclic, unsubstituted, substituted, and / or heteroatom-containing alkenyl and lower alkenyl, respectively.

[0137] The term "alkynyl" refers to a linear or branched hydrocarbon group of 2 to 24 carbon atoms containing at least one triple bond, such as ethynyl, n-propynyl, and the like. Generally, although again not necessarily, alkynyl groups can contain 2 to about 18 carbon atoms, and more particularly can contain 2 to 12 carbon atoms. The term "lower alkynyl" intends an alkynyl group of 2 to 6 carbon atoms. The term "substituted alkynyl" refers to alkynyl substituted with one or more substituent groups, and the terms "heteroatom-containing alkynyl" and "heteroalkynyl" refer to alkynyl in which at least one carbon atom is replaced with a heteroatom. If not otherwise indicated, the terms "alkynyl" and "lower alkynyl" include linear, branched, unsubstituted, substituted, and / or heteroatom- containing alkynyl and lower alkynyl, respectively.

[0138] The terms "alkyl", "alkenyl", and "alkynyl" are intended to include moieties which are diradicals, i.e., having two points of attachment. A nonlimiting example of such an alkyl moiety that is a diradical is --CH2CH2--, i.e., a C2 alkyl group that is covalently bonded via each terminal carbon atom to the remainder of the molecule.

[0139] The term "alkoxy" refers to an alkyl group bound through a single, terminal ether linkage; that is, an "alkoxy" group may be represented as --O-alkyl where alkyl is as defined above. A "lower alkoxy" group intends an alkoxy group containing 1 to 6 carbon atoms, and includes, for example, methoxy, ethoxy, n-propoxy, isopropoxy, t-butyloxy, etc. Preferred substituents identified as "C1-C6alkoxy" or "lower alkoxy" herein contain 1 to 3 carbon atoms, and particularly preferred such substituents contain 1 or 2 carbon atoms (i.e., methoxy and ethoxy).

[0140] The term "aryl" refers to an aromatic substituent containing a single aromatic ring or multiple aromatic rings that are fused together, directly linked, or indirectly linked (such that the different aromatic rings are bound to a common group such as a methylene or ethylene moiety). Aryl groups can contain 5 to 20 carbon atoms, and particularly preferred aryl groups can contain 5 to 14 carbon atoms. Examples of aryl groups include benzene, phenyl, pyrrole, furan, thiophene, thiazole, isothiazole, imidazole, triazole, tetrazole, pyrazole, oxazole, isooxazole, pyridine, pyrazine, pyridazine, and pyrimidine, and the like. Furthermore, the term "aryl" includes multicyclic aryl groups, e.g., tricyclic, bicyclic, e.g., naphthalene, benzoxazole, benzodioxazole, benzothiazole, benzoimidazole, benzothiophene, methylenedioxyphenyl, quinoline, isoquinoline, napthridine, indole, benzofuran, purine, benzofuran, deazapurine, or indolizine. Those aryl groups having heteroatoms in the ring structure may also be referred to as "aryl heterocycles", "heterocycles," "heteroaryls" or "heteroaromatics". The aromatic ring can be substituted at one or more ring positions with such substituents as described above, as for example, halogen, hydroxyl, alkoxy, alkylcarbonyloxy, arylcarbonyloxy, alkoxycarbonyloxy, aryloxycarbonyloxy, carboxylate, alkylcarbonyl, alkylaminocarbonyl, aralkylaminocarbonyl, alkenylaminocarbonyl, alkylcarbonyl, arylcarbonyl, aralkylcarbonyl, alkenylcarbonyl, alkoxycarbonyl, aminocarbonyl, alkylthiocarbonyl, phosphate, phosphonato, phosphinato, cyano, amino (including alkylamino, dialkylamino, arylamino, diaryl amino, and alkylaryl amino), acylamino (including alkylcarbonylamino, arylcarbonylamino, carbamoyl and ureido), amidino, imino, sulfhydryl, alkylthio, arylthio, thiocarboxylate, sulfates, alkylsulfinyl, sulfonato, sulfamoyl, sulfonamido, nitro, trifluoromethyl, cyano, azido, heterocyclyl, alkylaryl, or an aromatic or heteroaromatic moiety. Aryl groups can also be fused or bridged with alicyclic or heterocyclic rings, which are not aromatic so as to form a multicyclic system (e.g., tetralin, methylenedioxyphenyl). If not otherwise indicated, the term "aryl" includes unsubstituted, substituted, and / or heteroatom-containing aromatic substituents.

[0141] The terms "heterocyclyl" or "heterocyclic group" include closed ring structures, e.g., 3- to 10-, or 4- to 7-membered rings, which include one or more heteroatoms. "Heteroatom" includes atoms of any element other than carbon or hydrogen. Examples of heteroatoms include nitrogen, oxygen, sulfur and phosphorus.

[0142] Heterocyclyl groups can be saturated or unsaturated and include pyrrolidine, oxolane, thiolane, piperidine, piperazine, morpholine, lactones, lactams, such as azetidinones and pyrrolidinones, sultams, and sultones. Heterocyclic groups such as pyrrole and furan can have aromatic character. They include fused ring structures, such as quinoline and isoquinoline. Other examples of heterocyclic groups include pyridine and purine. The heterocyclic ring can be substituted at one or more positions with such substituents as described above, as for example, halogen, hydroxyl, alkylcarbonyloxy, arylcarbonyloxy, alkoxycarbonyloxy, aryloxycarbonyloxy, carboxylate, alkylcarbonyl, alkoxycarbonyl, aminocarbonyl, alkylthiocarbonyl, alkoxyl, phosphate, phosphonato, phosphinato, cyano, amino (including alkyl amino, dialkylamino, arylamino, diarylamino, and alkylarylamino), acylamino (including alkylcarbonylamino, arylcarbonylamino, carbamoyl and ureido), amidino, imino, sulfhydryl, alkylthio, arylthio, thiocarboxylate, sulfates, sulfonato, sulfamoyl, sulfonamido, nitro, trifluoromethyl, cyano, azido, heterocyclyl, or an aromatic or heteroaromatic moiety. Heterocyclic groups can also be substituted at one or more constituent atoms with, for example, a lower alkyl, a lower alkenyl, a lower alkoxy, a lower alkylthio, a lower alkylamino, a lower alkylcarboxyl, a nitro, a hydroxyl, --CF3, or --CN, or the like.

[0143] The term "halo" or "halogen" refers to fluoro, chloro, bromo, and iodo. "Counterion" is used to represent a small, negatively charged species such as fluoride, chloride, bromide, iodide, hydroxide, acetate, and sulfate. The term sulfoxide refers to a sulfur attached to 2 different carbon atoms and one oxygen and the S-O bond can be graphically represented with a double bond (S=O), a single bond without charges (S-O) or a single bond with charges [S(+)-O(-)].

[0144] The terms "substituted" as in "substituted alkyl," "substituted aryl," and the like, as alluded to in some of the aforementioned definitions, is meant that in the alkyl, aryl, or other moiety, at least one hydrogen atom bound to a carbon (or other) atom is replaced with one or more non-hydrogen substituents. Examples of such substituents include, without limitation: functional groups such as halo, hydroxyl, silyl, sulfhydryl, C1-C24alkoxy, C2-C24alkenyloxy, C2-C24 alkynyloxy, C5-C20 aryloxy, acyl (including C2-C24 alkylcarbonyl (-CO-alkyl) and C6-C20arylcarbonyl (-CO-aryl)), acyloxy (-O-acyl), C2-C24alkoxycarbonyl (-(CO)-O-alkyl), C6-C20 aryloxycarbonyl (-(CO)-O-aryl), C2-C24 alkylcarbonato (-O-(CO)-O-alkyl), C6-C20arylcarbonato (-O-(CO)-O-aryl), carboxy (-COOH), carboxylato(-COO-), carbamoyl (-(CO)-NH2), mono-(C1-C24 alkyl)-substituted carbamoyl (-(CO)- NH(C1-C24alkyl)), di-(C1-C4alkyl)-substituted carbamoyl (-(CO)--N(C1-C24alkyl)2), mono-substituted arylcarbamoyl (-(CO)-NH-aryl), thiocarbamoyl (-(CS)-NH2), carbamido (-NH-(CO)-NH2), cyano(-CN), isocyano (-N+C-), cyanato (-O--CN), isocyanato (-ON+C-), isothiocyanato (-S-CN), azido (-N=N+=N-), formyl (-(CO)--H), thioformyl (-(CS)-H), amino (-NH2), mono- and di-(C1-C24alkyl)-substituted amino, mono- and di-(C5-C20aryl)- substituted amino, C2-C24 alkylamido (-NH-(CO)-alkyl), C6-C20 arylamido (-NH-(CO)-aryl), imino (-CR=NH where R=hydrogen, C1-C24alkyl, C5-C20aryl, C6-C24alkaryl, C6-C24aralkyl, etc.), alkylimino (--CR=N(alkyl), where R=hydrogen, alkyl, aryl, alkaryl, etc.), arylimino (-CR=N(aryl), where R=hydrogen, alkyl, aryl, alkaryl, etc.), nitro (-NO2), nitroso (-NO), sulfo (-SO2 -OH), sulfonato (-SO2-O-), C1-C24 alkylsulfanyl (-S-alkyl; also termed "alkylthio"), arylsulfanyl (-S-aryl; also termed "arylthio"), C1-C24 alkylsulfinyl (--(SO)-alkyl), C5-C20 arylsulfinyl (-(SO)-aryl), C1-C24 alkylsulfonyl (-SO2-alkyl), C5-C20 arylsulfonyl (-SO2 -aryl), phosphono (-P(O)(OH)2), phosphonato (-P(O)(O-)2), phosphinato (-P(O)(O-)), phospho (-PO2), and phosphino (-PH2); and the hydrocarbyl moieties C1-C24alkyl, C2-C24alkenyl, C2-C24 alkynyl, C5-C20 aryl, C6-C24 alkaryl, and C6-C24 aralkyl.

[0145] In addition, the aforementioned functional groups may, if a particular group permits, be further substituted with one or more additional functional groups or with one or more hydrocarbyl moieties such as those specifically enumerated above. Analogously, the above-mentioned hydrocarbyl moieties may be further substituted with one or more functional groups or additional hydrocarbyl moieties such as those specifically enumerated.

[0146] When the term "substituted" appears prior to a list of possible substituted groups, it is intended that the term apply to every member of that group. For example, the phrase "substituted alkyl, alkenyl, and aryl" is to be interpreted as "substituted alkyl, substituted alkenyl, and substituted aryl." Analogously, when the term "heteroatom- containing" appears prior to a list of possible heteroatom-containing groups, it is intended that the term apply to every member of that group. For example, the phrase "heteroatom- containing alkyl, alkenyl, and aryl" is to be interpreted as "heteroatom-containing alkyl, substituted alkenyl, and substituted aryl.

[0147] "Optional" or "optionally" means that the subsequently described circumstance may or may not occur, so that the description includes instances where the circumstance occurs and instances where it does not. For example, the phrase "optionally substituted"means that a non-hydrogen substituent may or may not be present on a given atom, and, thus, the description includes structures wherein a non-hydrogen substituent is present and structures wherein a non-hydrogen substituent is not present.

[0148] The terms "stable compound" and "stable structure" are meant to indicate a compound that is sufficiently robust to survive isolation, and as appropriate, purification from a reaction mixture, and formulation into an efficacious therapeutic agent.

[0149] The terms "free compound" is used herein to describe a compound in the unbound state.

[0150] Throughout the description, where compositions are described as having, including, or comprising, specific components, it is contemplated that compositions also consist essentially of, or consist of, the recited components. Similarly, where methods or processes are described as having, including, or comprising specific process steps, the processes also consist essentially of, or consist of, the recited processing steps. Further, it should be understood that the order of steps or order for performing certain actions is immaterial so long as the compositions and methods described herein remains operable. Moreover, two or more steps or actions can be conducted simultaneously.

[0151] The term "small molecule" is an art-recognized term. In certain embodiments, this term refers to a molecule, which has a molecular weight of less than about 2000 amu, or less than about 1000 amu, and even less than about 500 amu.

[0152] All percentages and ratios used herein, unless otherwise indicated, are by weight.

[0153] Embodiments described herein relate to compositions and methods of attenuating and / or treating opioid addiction, withdrawal and / or dependance in a subject in need thereof and / or preventing loss of opioid induced analgesia and / or occurrence of opioid induced hyperalgesia, and particularly relates to the use of thiol-based compounds in compositions and methods of attenuating and / or treating opioid withdrawal and / or dependance and / or preventing loss of opioid induced analgesia and / or occurrence of opioid induced hyperalgesia.

[0154] Opioids are substances that act by binding to opioid receptors, which receptors are found principally in the central and peripheral nervous system and the gastrointestinal tract. These receptors mediate both the psychoactive and the somatic effects of opioids.Medically, opioids are primarily used for pain relief, including anesthesia. Other medical uses include suppression of diarrhea and suppressing cough.

[0155] Opioids include opiates, which are alkaloid compounds naturally found in the opium poppy plant (i.e., Papaver somniferum). The psychoactive compounds found in the opium plant include opium, heroin, morphine, codeine and thebaine. Examples of synthetic, or semi-synthetic, opioids include hydrocodone; oxycodone; fentanyl; methadone; pethidine and hydromorphone. Opioid therapy is the treatment of a human subject with opioids, typically a prolonged treatment with opioids, typically to achieve analgesic effects.

[0156] In some embodiments, the methods described herein include the administration of a pharmaceutical composition to a human subject, in need thereof, in an amount which is effective to inhibit the physical dependence on opioids by the subject. A human subject in need thereof is a human who is to receive, or is receiving, opioid therapy. Administration includes administration by a physician or by self-administration.

[0157] Physical dependence on an opioid is a state of adaptation by a patient who has received the opioid for a period of time and who experiences, or would experience, withdrawal syndrome if the opioid is abruptly withdrawn or if a narcotic antagonist (e.g., naloxone) is administered. Physical dependence is a normal physiological response.

[0158] Opioid withdrawal or opioid withdrawal syndrome includes symptoms which may range from mild to severe, depending on how dependent the subject is on the opioid. Dependency can be directly tied to the length of time taking an opioid, dosage amount, which particular opioid was taken, route of administration, underlying medical conditions, mental health, and certain biological and environmental factors, such as family history of addiction, previous trauma, and stressful surroundings.

[0159] Withdrawal from an opioid may roughly adhere to the following timeline, although it can vary from subject to subject. Early withdrawal symptoms typically start within 6-12 hours after last dose is taken for short-acting opioids, and start within 30 hours after last dose is taken for longer-acting opioids. Early withdrawal symptoms include: lacrimation, muscle aches, agitation, insomnia, excessive yawning, anxiety, panic, rhinorrhea, sweating, tachycardia, hypertension and fever. Late withdrawal symptoms typically peak within 72 hours after last dose is taken, usually lasting about a week, and include nausea and vomiting, diarrhea, piloerection, stomach cramps, depression, and drug cravings. Other withdrawal symptoms may include, for example, mydriasis, restlessness; tremor; involuntarymovements; muscle twitches; abdominal cramps; cold flashes; substantial physical and mental fatigue; dysphoric mood; drowsiness; salivation; loss of appetite; headache; dizziness; fainting; malaise; shivering; muscle / joint pain; irritability; poor concentration; confusion; flu- like symptoms; and the like.

[0160] In some embodiments, administration of the thiol based compounds described herein to a subject undergoing and / or about to undergo opioid withdrawal and / or exhibiting or experiencing opioid withdrawal symptoms, such as subject dependent or addicted to an opioid, can prevent, attenuate, and / or treat opioid withdrawal symptoms.

[0161] In other embodiments, administration of the thiol based compounds described herein to a subject undergoing and / or about to undergo opioid therapy can to prevent, attenuate, and / or treat opioid addiction, dependence, and / or withdrawal symptoms and / or prevent loss of opioid induced analgesia and / or occurrence of opioid induced hyperalgesia.

[0162] In some embodiments, the thiol based compound that is used to prevent, attenuate, and / or treat opioid addiction, dependence, and / or withdrawal symptoms and / or prevent loss of opioid induced analgesia and / or occurrence of opioid induced hyperalgesia can include a compound having a structure of formula (I):a pharmaceutically acceptable salt, a tautomer, or a solvate thereof; wherein: X is -OR2or -N(R3)2; X1is -SR4or -S(O)nOR5; R1is H, alkyl optionally substituted with one or more halogen, or -C(O)-alkyl optionally substituted with one or more halogen; R2, R5, and R6are each independently H or alkyl optionally substituted with one or more halogen; each R3is independently H or alkyl optionally substituted with one or more halogen; R4is -N=O or -OR6; and n is 1 or 2.

[0163] In some embodiments, R1is H, C1-C6 alkyl optionally substituted with one or more halogen, or -C(O)-(C1-C6alkyl) optionally substituted with one or more halogen. For example, R1is H, methyl, ethyl, propyl, butyl, -C(O)-methyl, -C(O)-ethyl, -C(O)-propyl, or - C(O)-butyl, each optionally substituted with one or more halogen.

[0164] In some R2and R6are each independently H or C1-C6 alkyl optionally substituted with one or more halogen. For example, R2and R6are each independently H, methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

[0165] In some embodiments, each R3is independently H or C1-C6alkyl optionally substituted with one or more halogen. For example, one of each R3is H and the other of R3is methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

[0166] In some embodiments, X is OR2, R1is H, C1-C6 alkyl optionally substituted with one or more halogen, or -C(O)-(C1-C6 alkyl) optionally substituted with one or more halogen, and R2is H or C1-C6 alkyl optionally substituted with one or more halogen. For example, X is OR2, R1is H, methyl, ethyl, propyl, butyl, -C(O)-methyl, -C(O)-ethyl, -C(O)-propyl, or - C(O)-butyl, each optionally substituted with one or more halogen, and R2is H, methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

[0167] In other embodiments, X is N(R3)2; R1is H, C1-C6alkyl optionally substituted with one or more halogen, or -C(O)-(C1-C6 alkyl) optionally substituted with one or more halogen; and each R3is independently H or C1-C6alkyl optionally substituted with one or more halogen. For example, X is N(R3)2; R1is H, methyl, ethyl, propyl, butyl, -C(O)- methyl, -C(O)-ethyl, -C(O)-propyl, or -C(O)-butyl, each optionally substituted with one or more halogen, and one R3is H and the other R3is methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

[0168] In some embodiments, X1is SR4and R4is -N=O.

[0169] In other embodiments, X1is SR4, R4is -OR6, and R6is H or C1-C6 alkyl optionally substituted with one or more halogen. For example, X1is SR4, R4is -OR6, and R6is H, methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

[0170] In some embodiments, X1is -S(O)nOR5and R5is H or C1-C6alkyl optionally substituted with one or more halogen. For example, X1is -S(O)nOR5and R5is H, methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

[0171] In some embodiments, the compound can have a structure of formula (II):(II), a pharmaceutically acceptable salt, a tautomer, or a solvate thereof; wherein: X is -OR2or -N(R3)2; R1is H, alkyl optionally substituted with one or more halogen, or -C(O)-alkyl optionally substituted with one or more halogen; R2is independently H or alkyl optionally substituted with one or more halogen; each R3is independently H or alkyl optionally substituted with one or more halogen.

[0172] In some embodiments, R1is H, C1-C6alkyl optionally substituted with one or more halogen, or -C(O)-(C1-C6 alkyl) optionally substituted with one or more halogen. For example, R1is H, methyl, ethyl, propyl, butyl, -C(O)-methyl, -C(O)-ethyl, -C(O)-propyl, or - C(O)-butyl, each optionally substituted with one or more halogen.

[0173] In some embodiments, R2is H or C1-C6alkyl optionally substituted with one or more halogen. For example, R2is H, methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

[0174] In some embodiments, each R3is independently H or C1-C6 alkyl optionally substituted with one or more halogen. For example, one of each R3is H and the other of R3is methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

[0175] In some embodiments, the compound can have a structure of formula (III):a pharmaceutically acceptable salt, a tautomer, or a solvate thereof; wherein: X is -OR2or -N(R3)2; R1is H, alkyl optionally substituted with one or more halogen, or -C(O)-alkyl optionally substituted with one or more halogen; R2and R6are each independently H or alkyl optionally substituted with one or more halogen; andeach R3is independently H or alkyl optionally substituted with one or more halogen.

[0176] In some embodiments, R1is H, C1-C6 alkyl optionally substituted with one or more halogen, or -C(O)-(C1-C6alkyl) optionally substituted with one or more halogen. For example, R1is H, methyl, ethyl, propyl, butyl, -C(O)-methyl, -C(O)-ethyl, -C(O)-propyl, or - C(O)-butyl, each optionally substituted with one or more halogen.

[0177] In some embodiments, R2and R6are each independently H or C1-C6 alkyl optionally substituted with one or more halogen. For example, R2and R6are each independently H, methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

[0178] In some embodiments, each R3is independently H or C1-C6 alkyl optionally substituted with one or more halogen.

[0179] In some embodiments, where one of each R3is H, the other of R3is methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

[0180] In other embodiments, the compound can have a structure of formula (IV):a pharmaceutically acceptable salt, a tautomer, or a solvate thereof; wherein: X is -OR2or -N(R3)2; R1is H, alkyl optionally substituted with one or more halogen, or -C(O)-alkyl optionally substituted with one or more halogen; R2and R5are each independently H or alkyl optionally substituted with one or more halogen; and each R3is independently H or alkyl optionally substituted with one or more halogen.

[0181] In some embodiments, R1is H, C1-C6alkyl optionally substituted with one or more halogen, or -C(O)-(C1-C6 alkyl) optionally substituted with one or more halogen. For example, R1is H, methyl, ethyl, propyl, butyl, -C(O)-methyl, -C(O)-ethyl, -C(O)-propyl, or - C(O)-butyl, each optionally substituted with one or more halogen.

[0182] In some embodiments, R2and R5are each independently H or C1-C6 alkyl optionally substituted with one or more halogen. For example, R2and R5are each independently H, methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

[0183] In some embodiments, each R3is independently H or C1-C6 alkyl optionally substituted with one or more halogen. For example, one of each R3is H and the other of R3is methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

[0184] In some embodiments, the compound can have a structure of formula (V):a pharmaceutically acceptable salt, a tautomer, or a solvate thereof; wherein: X is -OR2or -N(R3)2; R1is H, alkyl optionally substituted with one or more halogen, or -C(O)-alkyl optionally substituted with one or more halogen; R2and R5are each independently H or alkyl optionally substituted with one or more halogen; and each R3is independently H or alkyl optionally substituted with one or more halogen.

[0185] In some embodiments, R1is H, C1-C6 alkyl optionally substituted with one or more halogen, or -C(O)-(C1-C6alkyl) optionally substituted with one or more halogen. For example, R1is H, methyl, ethyl, propyl, butyl, -C(O)-methyl, -C(O)-ethyl, -C(O)-propyl, or - C(O)-butyl, each optionally substituted with one or more halogen.

[0186] In some embodiments, R2and R5are each independently H or C1-C6alkyl optionally substituted with one or more halogen. For example, R2and R5are each independently H, methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

[0187] In some embodiments, each R3is independently H or C1-C6alkyl optionally substituted with one or more halogen. For example, one of each R3is H and the other of R3is methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

[0188] In some embodiments, the compound can have a structure selected from:pharmaceutically acceptable salt, a tautomer, or a solvate thereof; or a combination thereof.

[0189] Still other embodiments described herein relate to a pharmaceutical composition for use in treating addiction, withdrawal and / or dependence and / or preventing loss of opioidinduced analgesia and occurrence opioid induced hyperalgesia in a subject in need thereof includes a compound having a structure of formulas:adduct, a pharmaceutically acceptable salt, a tautomer, or a solvate thereof; wherein: X is -OR2or -N(R3)2; X1is -SR4or -S(O)nOR5; X2is -OR9or -N(R10)2; X3is -OR11or -N(R12)2; R1, R7, and R8are each independently H, alkyl optionally substituted with one or more halogen, or -C(O)-alkyl optionally substituted with one or more halogen; R2, R5, R6, R9and R11are each independently H or alkyl optionally substituted with one or more halogen; each R3, R10, and R12are independently H or alkyl optionally substituted with one or more halogen; R4is H, -N=O or -OR6; and n is 1 or 2.

[0190] In some embodiments, X is not -OR2if R1is H.

[0191] In other embodiments, X3is not -OR11if X2is -OR9and R7and R8are H.

[0192] In some embodiments, the adduct of the compound of formulas I or VI is biologically active and includes at least one of an albumin adduct, a glucose adduct, an L- cysteine adduct, an L-glutathione adduct, or a D-cysteine adduct.

[0193] In some embodiments, R1, R7, and R8are each independently H, C1-C6alkyl optionally substituted with one or more halogen, or -C(O)-(C1-C6 alkyl) optionally substituted with one or more halogen. For example, R1, R7, and R8are each independently H, methyl, ethyl, propyl, butyl, -C(O)-methyl, -C(O)-ethyl, -C(O)-propyl, or -C(O)-butyl, each optionally substituted with one or more halogen.

[0194] In some embodiments, R2, R5, R6, R9and R11are each independently H or C1-C6 alkyl optionally substituted with one or more halogen. For example, R2, R5, R6, R9and R11are each independently H, methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

[0195] In some embodiments, each R3, R10, and R12is independently H or C1-C6 alkyl optionally substituted with one or more halogen.

[0196] In some embodiments, one of each R3, R10, and R12is H and the other of R3, R6, and R9is methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

[0197] In some embodiments, X is -OR2, R1is H, C1-C6 alkyl optionally substituted with one or more halogen, or -C(O)-(C1-C6 alkyl) optionally substituted with one or more halogen, and R2is H or C1-C6 alkyl optionally substituted with one or more halogen. For example, X is -OR2, R1is H, methyl, ethyl, propyl, butyl, -C(O)-methyl, -C(O)-ethyl, -C(O)- propyl, or -C(O)-butyl, each optionally substituted with one or more halogen, and R2is H, methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

[0198] In some embodiments, X is -N(R3)2; R1is H, C1-C6 alkyl optionally substituted with one or more halogen, or -C(O)-(C1-C6alkyl) optionally substituted with one or more halogen; and each R3is independently H or C1-C6 alkyl optionally substituted with one or more halogen. For example, X is -N(R3)2; R1is H, methyl, ethyl, propyl, butyl, -C(O)- methyl, -C(O)-ethyl, -C(O)-propyl, or -C(O)-butyl, each optionally substituted with one or more halogen, and one R3is H and the other R3is methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

[0199] In some embodiments, X1is -SR4and R4is -N=O.

[0200] In other embodiments, X1is -SR4and R4is H.

[0201] In some embodiments, X1is -SR4, R4is -OR6, and R6is H or C1-C6 alkyl optionally substituted with one or more halogen. For example, X1is -SR4, R4is -OR6, and R6is H, methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

[0202] In some embodiments, X1is -S(O)nOR5and R5is H or C1-C6alkyl optionally substituted with one or more halogen. For example, X1is -S(O)nOR5and R5is H, methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

[0203] In some embodiments, X2is -OR9, R7is H, C1-C6 alkyl optionally substituted with one or more halogen, or -C(O)-(C1-C6alkyl) optionally substituted with one or morehalogen, R9is H or C1-C6 alkyl optionally substituted with one or more halogen; X3is OR11, R8is H, C1-C6alkyl optionally substituted with one or more halogen, or -C(O)-(C1-C6alkyl) optionally substituted with one or more halogen, and R11is H or C1-C6 alkyl optionally substituted with one or more halogen. For example, X2is OR9, R7is H, methyl, ethyl, propyl, butyl, -C(O)-methyl, -C(O)-ethyl, -C(O)-propyl, or -C(O)-butyl, each optionally substituted with one or more halogen, R9is H, methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen, X3is OR11; R8is H, methyl, ethyl, propyl, butyl, -C(O)-methyl, -C(O)-ethyl, -C(O)-propyl, or -C(O)-butyl, each optionally substituted with one or more halogen, and R11is H, methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

[0204] In other embodiments, X2is N(R10)2, R7is H, C1-C6 alkyl optionally substituted with one or more halogen, or -C(O)-(C1-C6 alkyl) optionally substituted with one or more halogen, each R10is independently H or C1-C6 alkyl optionally substituted with one or more halogen, X3is N(R12)2, R8is H, C1-C6 alkyl optionally substituted with one or more halogen, or -C(O)-(C1-C6alkyl) optionally substituted with one or more halogen, and each R12is independently H or C1-C6 alkyl optionally substituted with one or more halogen. For example, X2is N(R10)2, R7is H, methyl, ethyl, propyl, butyl, -C(O)-methyl, -C(O)-ethyl, - C(O)-propyl, or -C(O)-butyl, each optionally substituted with one or more halogen, one R10is H and the other R10is methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen, X3is N(R12)2; R8is H, methyl, ethyl, propyl, butyl, -C(O)-methyl, -C(O)- ethyl, -C(O)-propyl, or -C(O)-butyl, each optionally substituted with one or more halogen, and one R12is H and the other R12is methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

[0205] In some embodiments, the compound is not cysteine, cystine, a cysteine alkylester, or cystine dialkylester.

[0206] In some embodiments, the compound is not N-acetylcysteine.

[0207] In some embodiments, the compound has a structure selected from:pharmaceutically acceptable salt, a tautomer, or a solvate thereof; or a combination thereof.

[0208] In some embodiment, the pharmaceutical composition further includes a pharmaceutically acceptable carrier or excipient.

[0209] In some embodiments, the subject has an addiction, withdrawal and / or dependence from at least one of alcohol, amphetamine, cocaine, methamphetamine, nicotine, or opioid.

[0210] In some embodiments, the subject has an opioid withdrawal, dependence, and / or addiction.

[0211] In some embodiments, the subject has neonatal opioid withdrawal syndrome.

[0212] In some embodiments, the subject has an opioid use disorder (OUD).

[0213] In some embodiments, the opioid comprises at least one of alfentanil, buprenorphine, butorphanol, carfentanil, codeine, diamorphine, dextromoramide, dezocine, dihydrocodeine, fentanyl, hydrocodone, hydromorphone, levorphanol, meperidine, meptazinol, methadone, morphine, nalbuphine, nalorphine, opium, oxycodone, oxymorphone, pentazocine, propoxyphene, remifentanil, sufentanil, tapentadol, and tramadol, and pharmaceutically acceptable salts thereof. For example, the opioid is carfentanil, fentanyl, remifentanil, or sufentanil.

[0214] In some embodiments, the subject has been administered an opioid antagonist and the composition inhibits opioid antagonist withdrawal. The opioid antagonist caninclude, for example, naloxone, an oxymorphol analog of naloxone, a naloxone salt, or a naloxone dihydrate.

[0215] In some embodiments, the composition can be administered concurrently with opioid administration and / or up to about 10 minutes, up to about 20 minutes, up to about 30 minutes, up to about 40 minutes, up to about 50 minutes, up to about 60 minutes, up to about 70 minutes, up to about 80 minutes, up to about 90 minutes, up to about 100 minutes, up to about 110 minutes, or up to about 120 minutes before or after initiation of opioid administration.

[0216] Still other embodiments described herein relate to a compound of formula (I), (II), (III), (IV), (V), (VI) or a pharmaceutically acceptable salt or solvate thereof, or a pharmaceutical composition comprising any of the aforementioned compounds and a pharmaceutically acceptable excipient, for use in the treatment or prevention of a disease / disorder selected from: pain, including, e.g., acute pain, chronic pain, postsurgical pain (or postoperative pain or incisional pain), cancer pain, inflammatory pain (or pain associated with an inflammatory disease / disorder), rheumatoid arthritis-associated pain, neuropathic pain, or diabetes-associated pain; opioid-induced hyperalgesia (e.g., morphine- induced hyperalgesia or fentanyl-induced hyperalgesia) or analgesic tolerance associated with chronic opioid administration; or addiction, including, e.g., substance addiction (or drug addiction), particularly alcohol addiction, amphetamine addiction, cocaine addiction, methamphetamine addiction, methylphenidate addiction, nicotine addiction, or opioid addiction, behavioral addiction (or a compulsive control disease / disorder), particularly pathological forms of any one of gambling addiction, food or overeating addiction (or compulsive overeating), sex or sexual intercourse addiction, pornography addiction, electronic communication devices addiction, mobile phone addiction, computer addiction, internet addiction, videogames addiction, internet gaming addiction, digital media addiction, physical exercise addiction (or compulsive overexercising), shopping addiction (or compulsive spending), or work addiction (or compulsive overworking), or an obsessive- compulsive spectrum disorder, particularly obsessive-compulsive disorder, anorexia (or anorexia nervosa), bulimia (or bulimia nervosa), binge eating disorder, impulse control disorder, intermittent explosive disorder, kleptomania, pyromania, compulsive hoarding, a body-focused repetitive behavior disorder, or trichotillomania.

[0217] Compositions comprising a thiol compound of formula (I), (II), (III), (IV), (V), (VI), or an adduct, a pharmaceutically acceptable salt, a tautomer, or a solvate thereof described herein can be administered to a subject prior to or during opioid induced addiction, dependence and / or withdrawal. In some embodiments, the composition is administered to a human subject, in need thereof, during opioid therapy, optionally, slightly before the commencement of opioid therapy. For example, administration is begun at most about 48 hours before the first dose of an opioid or at the time of the first dose of an opioid, and is substantially continued for the duration of the opioid therapy. Alternatively, administration can be begun at any point during opioid therapy.

[0218] In some embodiments, administration of a composition comprising the thiol compound of formula (I), (II), (III), (IV), (V), (VI), or an adduct, a pharmaceutically acceptable salt, a tautomer, or a solvate thereof can inhibit, reduce, prevent, and / or shorten the duration of opioid dependance in the subject. That is, the methods described herein are considered to be effective if they cause one or more of: a reduction / prevention of dependence on an opioid and / or shortening of the duration of any dependence to an opioid. For example, dependence would be inhibited if upon cessation of opioid therapy, withdrawal symptoms are inhibited.

[0219] Inhibition of dependence can be assessed by comparing the magnitude and / or duration of dependence in a subject at two different occasions, that is, i) when administered the pharmaceutical composition during an opioid therapy; and ii) when not administered the pharmaceutical composition during an opioid therapy. An assessment is made as to the severity of withdrawal symptoms once the opioid is discontinued at the different occasions.

[0220] Inhibition of dependence can also be assessed by comparing the magnitude and / or duration of dependence in different subjects being treated with the same opioid, some of whom were administered the pharmaceutical composition during a therapy and some whom were not administered the pharmaceutical composition during a therapy. An assessment is made as to the severity of withdrawal symptoms once the opioid is discontinued between the different subjects.

[0221] Typically, dependence and / or withdrawal symptoms can be inhibited by at least about 10%, at least about 20%, at least about 30%, at least about 40%, at least about 50%, at least about 60%, at least about 70%, at least about 80%, at least about 90%, or about 100%.

[0222] In some embodiments, the subject treated with a composition comprising the thiol compound of formula (I), (II), (III), (IV), (V), (VI), or an adduct, a pharmaceutically acceptable salt, a tautomer, or a solvate thereof is a neonatal subject that has or at risk of neonatal opioid withdrawal syndrome (NOWS). (NOWS) is a subset of neonatal abstinence syndrome (NAS) and refers to neonatal withdrawal from opioid drugs and can occur in the presence of other drug withdrawal syndromes. NOWS is a generalized multisystem disorder that predominantly involves the central and autonomic nervous systems and the gastrointestinal tract. In the United States, neonatal withdrawal following in-utero opioid exposure is typically the result of prolonged maternal use and / or abuse of illicit or prescribed opioids during pregnancy. Birth leads to the abrupt cessation of fetal substance exposure and can precipitate acute withdrawal symptoms that can result in severe health complications with prolonged recovery and longer hospitalization. Withdrawal can be both severe and intense, with an estimated 60% to 80% of exposed neonates requiring pharmacological intervention to control symptoms (see Kraft, Walter K.; Stover, Megan W.; Davis, Jonathan M. Neonatal abstinence syndrome: Pharmacologic strategies for the mother and infant Seminars in Perinatology 40.3 (Apr.1, 2016): 203-212, and Tolia V N, Patrick S W, Bennett M M, et al. Increasing incidence of the neonatal abstinence syndrome in U.S. neonatal ICUs. N Engl J Med.2015; 372(22): 2118-2126, which are incorporated herein by reference in their entireties).

[0223] While the signs and symptoms of NOWS are similar to those experienced by adults undergoing acute opiate withdrawal, they present a higher risk to the neonate due to the infant's dependence on others for all aspects of well-being.

[0224] NOWS presenting signs include central nervous system hyperirritability (tremors, jitteriness, irritability, hyperactive muscle reflexes, and excessive high-pitched cry), autonomic nervous system deregulation and instability (tachypnea, nasal flaring, hyperphagia, temperature instability, insomnia, sweating, mottle skin, yawning, and sneezing), and gastrointestinal symptoms (diarrhea, vomiting, and poor feeding) (see Hudak M L, Tan R C. The Committee on Drugs and the Committee on Fetus and Newborn. Neonatal Drug Withdrawal. Pediatrics.2012; 129; e540, and Kocheriakota P. Neonatal Abstinence Syndrome. Pediatrics.2014; 134(2): e547-561, which are incorporated herein by reference in their entireties). Seizures are generally rare, although they have been reported occurring in 2% to 11% of neonate cases in the early stage of severe opioid withdrawal (seeDoberczak T M, Kandall S R, Wilets I. Neonatal opiate abstinence syndrome in term and preterm infants. J Pediatr 1991; 118: 933-7, which is incorporated herein by reference in its entirety, especially if medical treatment has been delayed. A recent large observational cohort example of Medicaid babies in 46 US States reported an incidence of seizures was 2.7% among the 1705 observed cases of NAS (see Desai, R. J., Hernandez-Diaz, S., Bateman, B. T. & Huybrechts, K. F. Increase in prescription opioid use during pregnancy among Medicaid-enrolled women. Obstet. Gynecol.123, 997-1002 (2014), which is incorporated herein by reference in its entirety. Neonates with NOWS can also experience weight loss or failure to thrive, which often results from a combination of poor feeding, vomiting, nausea, and diarrhea (see Kocheriakota P. Neonatal Abstinence Syndrome. Pediatrics.2014; 134(2): e547-561, which is incorporated herein by reference in its entirety). If left untreated, some cases of NOWS may even lead to death. Some of the less severe signs and symptoms of opiate withdrawal may persist for several months (see Hudak ML, Tan RC. The Committee on Drugs and the Committee on Fetus and Newborn. Neonatal Drug Withdrawal. Pediatrics.2012;129; e540, which is incorporated herein by reference in its entirety)

[0225] In some embodiments, administration of a composition including the thiol compound of formula (I), (II), (III), (IV), (V), (VI), or an adduct, a pharmaceutically acceptable salt, a tautomer, or a solvate thereof can inhibit, reduce, prevent, and / or shorten the duration of NOWS in a neonatal subject.

[0226] In some embodiments, compositions including the thiol compound of formula (I), (II), (III), (IV), (V), (VI), or adduct, a pharmaceutically acceptable salt, tautomer, or solvate thereof can be administered to the subject in combination with at least one additional compound, agent, and / or therapeutic agent useful for treating the subject or opioid use, opioid dependence, and / or opioid withdrawal. These additional compounds, agents, and / or therapeutic agents can include commercially available agents or compounds, known to treat, prevent, or reduce opioid use, opioid dependence, and / or opioid withdrawal in the subject.

[0227] In some embodiments, the at least one additional therapeutic agent can include an opioid antagonist, such as naloxone (naloxone, chemically known as 1-N-allyl-14- hydroxynordihydromorphinone). Naloxone can block the euphorigenic activity of an opioid and eliminate the development of psychological dependence. The inhibition of opiate effects by naloxone also prevents the development of physical dependence. U.S. Pat. No.3,773,955incorporated herein by reference, describes the oral combination of naloxone with a number of opiates particularly methadone. U.S. Pat. No.4,457,933 describes the protection with naloxone of oral dosage forms of various opioids against both oral and parenteral abuse. U.S. Pat. No.4,661,492 mentions the incorporation of 1-3 mg of naloxone in an oral unit dose of buprenorphine (2 mg).

[0228] In some embodiments, the compositions include an oxymorphol analog of naloxone, a naloxone salt, or a naloxone dihydrate.

[0229] Other compositions can include naltrexone (1-N-cyclopropylmethyl-14- hydroxynordihydromorphinone). Naltrexone is a pure opiate antagonist which, when administered orally as a maintenance drug for opiate addicts, blocks the effects of self- administered opiates thereby contributing to the extinction of drug craving.

[0230] In some embodiments, the composition can include nalmefene (also known as nalmetrene), another opioid antagonist. Nalmefene is similar in both structure and activity to naltrexone. Reported advantages of nalmefene relative to naltrexone include longer half-life, greater oral bioavailability and no observed dose-dependent liver toxicity. As with other opioid antagonists of the sort, nalmefene may precipitate acute withdrawal symptoms in patients who are dependent on opioid drugs, or post-operatively, to counteract the effects of strong opioids used in surgery. Any other opioid antagonist that can potentiate or enhance the effectiveness of a composition including the thiol compound of formula (I), (II), (III), (IV), (V), (VI), or adduct, pharmaceutically acceptable salt, tautomer, or solvate thereof is contemplated.

[0231] In still other embodiments a composition including the thiol compound of formula (I), (II), (III), (IV), (V), (VI), or adduct, pharmaceutically acceptable salt, tautomer, or solvate thereof can include an additional agent selected from an opioid, doxapram and enantiomers thereof, acetazolamide, almitrine, theophylline, caffeine, methylprogesterone and related compounds, sedatives that decrease arousal threshold in sleep disordered breathing patients, sodium oxybate, benzodiazepine receptor agonists, orexin antagonists, tricyclic antidepressants, serotonergic modulators, adenosine and adenosine receptor and nucleoside transporter modulators, cannabinoids, orexins, melatonin agonists, ampakines, or combinations thereof.

[0232] In one embodiment, the composition including the thiol compound of formula (I), (II), (III), (IV), (V), (VI), or adduct, pharmaceutically acceptable salt, tautomer, or solvatethereof described herein and an additional agent are physically mixed in the composition. In another embodiment, the composition a composition including the thiol compound of formula (I), (II), (III), (IV), (V), (VI), or an adduct, a pharmaceutically acceptable salt, a tautomer, or a solvate thereof described herein and the additional agent are physically separated in the composition.

[0233] In some embodiments, a composition including the thiol compound of formula (I), (II), (III), (IV), (V), (VI), or adduct, pharmaceutically acceptable salt, tautomer, or solvate thereof described herein may be packaged with at least one opioid capable of inducing dependence or addiction in a subject. The amount of the thiol compound of formula (I), (II), (III), (IV), (V), (VI), or adduct, pharmaceutically acceptable salt, tautomer, or solvate thereof described herein can be effective to prevent the opioid induced dependance or withdrawal.

[0234] In some embodiments, an effective amount (i.e., dose) of the compound of formula (I), (II), (III), (IV), (V), (VI) or a pharmaceutically acceptable salt or solvate thereof described herein to be administered to a subject can be determined depending upon, for example, age, body weight, symptom, the desired therapeutic effect, the route of administration, and the duration of the treatment. Exemplary doses can be from about 0.01 to about 1000 mg, by oral administration. Examples of dose ranges can include from a minimum dose of about 0.01, 0.10, 0.50, 1, 5, 10, 25, 50, 100, 125, 150, 200, or 250 mg to a maximum dose of about 300, 400, 500, 600, 700, 800, 900, or 1000 mg, wherein the dose range can include from any one of the foregoing minimum doses to any one of the foregoing maximum doses. Specific examples of particular effective amounts contemplated via oral administration can include about 0.02, 0.03, 0.04, 0.05, 0.10, 0.15, 0.20, 0.25, 0.30, 0.35, 0.40, 0.45, 0.50, 0.55, 0.60, 0.65, 0.70, 0.75, 0.80, 0.85, 0.90, 0.95, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 105, 110, 115, 120, 125, 130, 135, 140, 145, 150, 155, 160, 165, 170, 175, 180, 185, 190, 195, 200, 205, 210, 215, 220, 225, 230, 235, 240, 245, 250, 255, 260, 265, 270, 275, 280, 285, 290, 295, 300, 305, 310, 315, 320, 325, 330, 335, 340, 345, 350, 355, 360, 365, 370, 375, 380, 385, 390, 395, 400, 405, 410, 415, 420, 425, 430, 435, 440, 445, 450, 455, 460, 465, 470, 475, 480, 485, 490, 495, 500, 505, 510, 515, 520, 525, 530, 535, 540, 545, 550, 555, 560, 565, 570, 575, 580, 585,590, 595, 600, 605, 610, 615, 620, 625, 630, 635, 640, 645, 650, 655, 660, 665, 670, 675, 680, 685, 690, 695, 700, 705, 710, 715, 720, 725, 730, 735, 740, 745, 750, 755, 760, 765, 770, 775, 780, 785, 790, 795, 800, 805, 810, 820, 825, 830, 835, 840, 845, 850, 855, 860, 865, 870, 875, 880, 885, 890, 895, 900, 905, 910, 915, 920, 925, 930, 935, 940, 945, 950, 955, 960, 965, 970, 975, 980, 985, 990, 995, 1000 mg or more. The oral dose can be administered once daily, twice daily, three times daily, or more frequently.

[0235] The dose of the compound of formula (I), (II), (III), (IV), (V), (VI) or a pharmaceutically acceptable salt or solvate thereof described herein for use in parenteral administration (e.g., intravenous administration) is generally from about 0.01 to about 300 mg / kg body weight. Examples of dose ranges can include from a minimum dose of about 0.01, 0.10, 0.50, 1, 5, 10, 25, 50, or 100 mg / kg body weight to a maximum dose of about 125, 150, 175, 200, 250, 275, or 300 mg / kg body weight, wherein the dose range can include from any one of the foregoing minimum doses to any one of the foregoing maximum doses. Specific examples of effective amounts contemplated include about 0.02, 0.03, 0.04, 0.05, 0.10, 0.15, 0.20, 0.25, 0.30, 0.35, 0.40, 0.45, 0.50, 0.55, 0.60, 0.65, 0.70, 0.75, 0.80, 0.85, 0.90, 0.95, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 100, 105, 110, 115, 120, 125, 130, 135, 140, 145, 150, 155, 160, 165, 170, 175, 180, 185, 190, 195, 200, 205, 210, 215, 220, 225, 230, 235, 240, 245, 250, 255, 260, 265, 270, 275, 280, 285, 290, 295, 300 mg / kg body weight or more. Continuous intravenous administration is also contemplated for from 1 to 24 hours per day to achieve a target concentration from about 0.01 mg / L blood to about 100 mg / L blood. Exemplary dose ranges can include from a minimum dose of about 0.01, 0.10, 0.25, 0.50, 1, 5, 10, or 25 mg / L blood to a maximum dose of about 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, or 100 mg / L, wherein an exemplary dose ranges can include from any one of the foregoing minimum doses to any one of the foregoing maximum doses. Specific examples of particular effective amounts contemplated via this route include about 0.02, 0.03, 0.04, 0.05, 0.10, 0.15, 0.20, 0.25, 0.30, 0.35, 0.40, 0.45, 0.50, 0.55, 0.60, 0.65, 0.70, 0.75, 0.80, 0.85, 0.90, 0.95, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100 mg / L blood or more. Thedose to be used can depend upon various conditions, and there may be cases wherein doses lower than or greater than the ranges specified above are used.

[0236] In some embodiments, the composition is administered concurrently with opioid administration and / or up to about 10 minutes, up to about 20 minutes, up to about 30 minutes, up to about 40 minutes, up to about 50 minutes, up to about 60 minutes, up to about 70 minutes, up to about 80 minutes, up to about 90 minutes, up to about 100 minutes, up to about 110 minutes, or up to about 120 minutes before or after initiation of opioid administration.

[0237] The thiol compound of formula (I), (II), (III), (IV), (V), (VI), or adduct, pharmaceutically acceptable salt, tautomer, or solvate thereof described herein may be administered in the form of, for example, solid compositions, liquid compositions, or other compositions for oral administration, injections, liniments, or suppositories for parenteral administration. Solid compositions for oral administration include compressed tablets, pills, capsules, dispersible powders, and granules. Capsules include hard capsules and soft capsules. In such solid compositions, the thiol compound of formula (I), (II), (III), (IV), (V), (VI), or adduct, pharmaceutically acceptable salt, tautomer, or solvate thereof can be admixed with an excipient (e.g., lactose, mannitol, glucose, microcrystalline cellulose, or starch), combining agents (e.g., hydroxypropyl cellulose, polyvinyl pyrrolidone, or magnesium metasilicate aluminate), disintegrating agents (e.g., cellulose calcium glycolate), lubricating agents (e.g., magnesium stearate), stabilizing agents, agents to assist dissolution (e.g., glutamic acid or aspartic acid), or the like. The agents may, if desired, be coated with coating agents (e.g., sugar, gelatin, hydroxypropyl cellulose, or hydroxypropylmethyl cellulose phthalate), or be coated with two or more films. Further, coating may include containment within capsules of absorbable materials such as gelatin.

[0238] Liquid compositions for oral administration include pharmaceutically acceptable solutions, suspensions, emulsions, syrups, and elixirs. In such compositions, the thiol compound of formula (I), (II), (III), (IV), (V), (VI), or adduct, pharmaceutically acceptable salt, tautomer, or solvate thereof is dissolved, suspended, or emulsified in a commonly used diluent (e.g., purified water, ethanol, or mixture thereof). Furthermore, such liquid compositions may also comprise wetting agents, suspending agents, emulsifying agents, flavoring agents (e.g., flavor-masking agents) sweetening agents, perfuming agents, preserving agents, buffer agents, or the like.

[0239] Injections for parenteral administration include solutions, suspensions, emulsions, and solids, which are dissolved or suspended. For injections, the thiol compound of formula (I), (II), (III), (IV), (V), (VI), or adduct, pharmaceutically acceptable salt, tautomer, or solvate thereof can be dissolved, suspended, and / or emulsified in a solvent. The solvents are, for example, distilled water for injection, physiological salt solution, vegetable oil, propylene glycol, polyethylene glycol, alcohol such as ethanol, or a mixture thereof. Moreover the injections also can include stabilizing agents, agents to assist dissolution (e.g., glutamic acid, aspartic acid, or POLYSORBATE 80), suspending agents, emulsifying agents, soothing agents, buffer agents, preserving agents, etc. The compositions are sterilized in the final process or manufactured and prepared by sterile procedure. The compositions also can be manufactured in the form of sterile solid compositions, such as a freeze-dried composition, and can be sterilized or dissolved immediately before use in sterile distilled water for injection or some other solvent.

[0240] Other compositions for parenteral administration include liquids and ointments for external use, endermic liniments, compositions for inhalation, sprays, suppositories for rectal administration, and pessaries for vaginal administration, which compositions include a cystine ester and are administered by methods known in the art.

[0241] Compositions comprising the thiol compound of formula (I), (II), (III), (IV), (V), (VI), or adduct, pharmaceutically acceptable salt, tautomer, or solvate thereof for inhalation or sprays may comprise additional substances other than diluents, such as, e.g., stabilizing agents (e.g., sodium sulfite hydride), isotonic buffers (e.g., sodium chloride, sodium citrate or citric acid). See, for example, the methods described in U.S. Pat. Nos. 2,868,691 and 3,095,355. The thiol compound of formula (I), (II), (III), (IV), (V), (VI), or adduct, pharmaceutically acceptable salt, tautomer, or solvate thereof can be effectively distributed by inhalation or spray using a self-propelling composition that includes a solution or dispersion of the thiol compound of formula (I), (II), (III), (IV), (V), (VI), or adduct, pharmaceutically acceptable salt, tautomer, or solvate thereof in micronized form. For example, an effective dispersion of finely divided drug particles can be accomplished with the use of very small quantities of a suspending agent, present as a coating on micronized drug particles. Evaporation of the propellant from the aerosol particles after spraying from the aerosol container leaves finely divided drug particles coated with a fine film of the suspending agent. In the micronized form, the average particle size can be less than about 5microns. The propellant composition can employ, as the suspending agent, a fatty alcohol such as oleyl alcohol. Propellants that may be employed include hydrofluoroalkane propellants and chlorofluorocarbon propellants. Dry powder inhalation also can be employed.

[0242] The route(s) of administration will be readily apparent to the skilled artisan and will depend upon any number of factors including the type and severity of the disease being treated, the type and age of the veterinary or human patient being treated, and the like.

[0243] The formulations of the pharmaceutical compositions described herein may be prepared by any method known or hereafter developed in the art of pharmacology and pharmaceutics. In general, such preparatory methods include the step of bringing the active ingredient into association with a carrier or one or more other accessory ingredients, and then, if necessary or desirable, shaping or packaging the product into a desired single-dose or multi-dose unit.

[0244] Although the descriptions of pharmaceutical compositions provided herein are principally directed to pharmaceutical compositions, which are suitable for ethical administration to humans, it will be understood by the skilled artisan that such compositions are generally suitable for administration to animals of all sorts. Modification of pharmaceutical compositions suitable for administration to humans in order to render the compositions suitable for administration to various animals is well understood, and the ordinarily skilled veterinary pharmacologist can design and perform such modification with merely ordinary, if any, experimentation. Subjects to which administration of the pharmaceutical compositions is contemplated include, but are not limited to, humans and other primates, mammals including commercially relevant mammals such as cattle, pigs, horses, sheep, cats, and dogs.

[0245] The regimen of administration may affect what constitutes an effective amount. The therapeutic formulations may be administered to the patient either prior to or after the onset of acute or post-acute withdrawal. Further, several divided dosages, as well as staggered dosages may be administered daily or sequentially, or the dose may be continuously infused, or may be a bolus injection. Further, the dosages of the therapeutic formulations may be proportionally increased or decreased as indicated by the exigencies of the therapeutic or prophylactic situation.

[0246] Actual dosage levels of the active ingredients in the pharmaceutical compositions may be varied so as to obtain an amount of the active ingredient that is effective to achieve the desired therapeutic response for a particular patient, composition, and mode of administration, without being toxic to the patient.

[0247] A medical doctor, e.g., physician or veterinarian, having ordinary skill in the art may readily determine and prescribe the effective amount of the pharmaceutical composition required. For example, the physician or veterinarian could start doses of the compounds employed in the pharmaceutical composition at levels lower than that required in order to achieve the desired therapeutic effect and gradually increase the dosage until the desired effect is achieved.

[0248] The invention is further illustrated by the following example, which is not intended to limit the scope of the claims. Example 1

[0249] There are numerous major problems facing clinicians treating subjects with opioid use disorder (OUD) and other substance use disorders (SUDs). With regard to managing pain in the general population, key issues are how to provide µ-opioid receptor (µ- OR) agonist (e.g., fentanyl)-induced analgesia without eliciting (1) euphoria, (2) physical dependence and / or psychological addiction, (3) hyperalgesia, and (4) any combination of the above conditions. With respect to treating moderate to severe OUD (DSM-5 terminology for opioid addiction in 10-20% of people who have liability for SUD), the key issues are (1) how to manage the often severe opioid withdrawal (all current strategies to manage opioid withdrawal are inadequate); (2) how to manage patients having gone through withdrawal and need medication to block euphoria and / or physical dependence, with the three medications available, naltrexone, buprenorphine and methadone, all having strengths but major weaknesses; and (3) how to avoid opioid euphoria and / or physical dependence in patients with moderate to severe OUD who currently are sober, but require µ-OR agonist analgesia. With respect to effectively treating SUD involving opioids and other substances (e.g., alcohol, cannabinoids, benzodiazepines and psycho-stimulants, in 10-20% of the population prone to SUD), the important issue is how to provide (a yet to be developed) effective therapeutic that will attenuate / block dopamine surge-mediated euphoria of each of these families of brain-reward drugs, as an adjunct to treating addictive diseases. With respect tomaking opioid analgesics safer, we lack drugs that effectively modulate the actions of opioids to improve their analgesic profile. The therapeutics should (1) not interfere with or indeed promote opioid analgesia, (2) prevent the acquisition of physical dependence and psychological addiction to opioids, (3) block opioid-induced respiratory depression (OIRD), or (4) stop the development of hyperalgesia.

[0250] We hypothesized that co-administration of cell-permeant versions of L-cysteine such as L-cysteine ethyl ester (L-CYSee) may prevent acquisition of physical dependence to morphine and reverse established dependence to the opioid. We found that L-CYSee, L- cysteine methyl ester, and other thiolesters and related compounds prevent and / or reverse the adverse effects of morphine and fentanyl on ventilatory parameters, arterial blood-gas chemistry (pH, pCO2, pO2 and sO2) and Alveolar-arterial gradient (index of alveolar gas- exchange in the lungs) in freely moving rats without compromising opioid-induced analgesia or sedation. We now provide evidence that administration of L-CYSee prevents the acquisition of physical dependence, as measured by markedly fewer withdrawal phenomana in response to administration of the opioid receptor antagonist, naloxone HCl (NLX) in freely-moving male rats and reverses established dependence. The lack of effect of L- cysteine and L-serine ethyl ester (L-SERee, oxygen atom instead of a sulfur atom as in L- CYSee) in these paradigms suggests that the efficacy of L-CYSee involves its cell penetrability in brain regions vital to expression of morphine dependence and points to the vital role of thiol biochemistry in the biological efficacy of L-CYSee. Materials and Methods Permissions, Rats, And Surgical Procedures

[0251] All studies were carried out in strict accordance with the NIH Guide for Care and Use of Laboratory Animals (NIH Publication No.80-23) revised in 1996, and in strict compliance with the ARRIVE (Animal Research: Reporting of In Vivo Experiments) guidelines (http: / / www.nc3rs.org.uk / page. asp? id=1357). All protocols involving the use of rats were approved by the Animal Care and Use Committees of Galleon Pharmaceuticals, Case Western Reserve University, and the University of Virginia. Adult male Sprague Dawley rats were purchased from Harlan Industries (Madison, WI, USA). The rats were given five days to recover from transportation before being subject to surgeries as will be described below. (+)-Morphine sulfate was obtained from Baxter Healthcare (Deerfield, IL,USA). L-CYSee HCl powder was obtained from Sigma-Aldrich (St. Louis, MO, USA) and divided into 100 mg amounts under N2gas and stored at 4oC. Solutions of L-CYSee (dissolved in saline and brought to pH 7.2 with 0.1M NaOH at room temperature) were prepared immediately before use (see below). Naloxone HCl (Sigma-Aldrich, St. Louis, MO, USA) was dissolved in normal saline. On the day of example, all arterial and venous catheters were flushed with 0.3 ml of phosphate-buffered saline (0.1 M, pH 7.4) 3-4h before commencement of the example. All studies were done in a quiet room with relative humidity of 50 ± 2% and room temperature of 21.3 ± 0.2 °C. Protocols to determine the effects of L-CYSee on physical dependence to morphine Prevention of morphine dependence Behavioral studies

[0252] At 2 am of the day of surgery, two groups of rats received a jugular vein catheter (PE-10 connected to PE-50) under 2-3% isoflurane anesthesia. The jugular vein catheter was connected to a primed ALZET osmotic minipump (Model 2002, ALZA Corporation, California, U.S.A.) positioned at the back of the neck to allow continuous infusion of vehicle (20 µL / h, IV), L-cysteine (20.8 µmol / kg / h, IV), L-CYSee (20.8 µmol / kg / h, IV) or L-SERee (20.8, µmol / kg / h, IV), as described previously. Physical dependence was induced by a slow- release subcutaneous depot of morphine emulsion (150 mg / kg, SC) injected at the left side of the neck as described in detail by Fennessy and colleagues. In brief, morphine base was precipitated from a solution of (+)-morphine sulfate by titrating to pH 9 with 1 mol / L NaOH. After several distilled water washes, pure base was collected in a filter funnel and dried. Morphine slow-release emulsion was prepared by suspending a weighed amount of base in liquid paraffin and Arlacel A. This mixture was then emulsified with an equal volume of normal saline as initially described by Collier et al (1972). All wounds were sutured closed and the rats were returned to their warmed home cages. After 35.5h of morphine administration, the rats were placed in individual opaque plastic boxes and after 30 min of acclimatization, they received an intraperitoneal (IP) injection of NLX (1.5 mg / kg) and behavioral phenomena were scored for 45 min by at least 3 scorers. The scored phenomena were: Jumping behavior - all 4 paws of the ground - jumps; Wet dog shakes -whole body shakes as if to shed water from fur; Rearing behavior - rearing on hind legs - rears; Episodesof fore-paw licking – FPL; Circling behavior - Complete 360orotation; Writhes – full body contortion; Sneezes - episodes of sneezing – abrupt expulsion of air that often disturbed the fine bedding material. Plethysmography ventilatory studies

[0253] Groups of rats were prepared as above except that rats received a second catheter into the jugular vein as described by Getsy et al (2020f) to give a bolus injection of NLX. After 35h, rats were put in individual whole body plethysmography chambers and the free end of the exteriorized venous catheter was connected to a swivel assembly housed in the lid of the plethysmography chamber and after a 60 min acclimatization period, the rats were given an intravenous injection of NLX (1.5 mg / kg). Ventilatory parameters including frequency of breathing, tidal volume, minute ventilation and non-eupneic breathing indices were recorded (to be reported elsewhere) with the number of apneas (> 1.5 sec between breaths) reported here. Cardiovascular studies

[0254] Groups of rats were prepared as above except that the rats received a second catheter into the jugular vein to administer NLX and a catheter into a femoral artery to continuously record mean arterial blood pressure (MAP) and heart rate as described previously. After 35h, the rats were placed in placed in individual opaque plastic boxes and the free end of the exteriorized jugular vein catheter was connected to an injection line to deliver NLX. The free end of the arterial line was connected to tubing attached to a computer-coupled pressure transducer (Cabe Lab, Inc.) to record pulsatile arterial blood pressure. After a 60 min acclimatization period, the rats received a bolus injection of NLX (1.5 mg / kg, IV) and MAP and heart rate were recorded continuously for 45 min. Body temperature and body weight studies

[0255] Groups of rats without a second jugular catheter were prepared as described above. After 35h, the rats were placed in individual opaque plastic boxes and a thermistor probe connected to a telethermometer (Yellow Springs Instruments) to record body temperature was inserted 5-6 cm into the rectum and taped to the tail. The body weights of the rats and their body temperatures were recorded every 15 min during the acclimatization period to establish accurate baseline values. After the 60 min acclimatization, the ratsreceived an intraperitoneal injection of NLX (1.5 mg / kg) and body temperatures and body weights were recorded every 15 min for 90 min. Reversal of morphine dependence Behavioral studies

[0256] At 2 pm of the day of surgery, two groups of rats received a slow-release subcutaneous depot of morphine emulsion (150 mg / kg, SC) injected at the left side of the neck as described above. After 36h of morphine administration, the rats were anesthetized (2% isoflurane) and received a jugular vein catheter connected to a primed ALZET osmotic minipump positioned at the back of the neck for continuous infusion of vehicle (20 µL / h, IV), L-cysteine (20.8, µmol / kg / h, IV), L-CYSee (20.8 µmol / kg / h, IV) or L-SERee (20.8, µmol / kg / h, IV) as above. All wounds were then sutured closed and the rats were returned to their warmed home cages. After 11.5h, the rats were placed in individual opaque plastic boxes and after a 30 min period of acclimatization, the rats received an intraperitoneal injection of NLX (1.5 mg / kg) and behavioral phenomena (as detailed above) were scored for 45 min by at least 3 scorers. Plethysmography ventilatory studies

[0257] Groups of rats were prepared as above except that the rats received a second catheter into the jugular vein to allow for bolus injection of NLX. After 47h, rats were placed in individual whole body plethysmography chambers and the free end of the exteriorized jugular vein catheter was connected to a swivel on the lid of the plethysmography chamber. After 60 min of acclimatization, the rats received a bolus injection of NLX (1.5 mg / kg, IV). Ventilatory parameters and non-eupneic breathing indices were recorded with the number of apneas (> 1.5 sec between breaths) to be reported here. Cardiovascular studies

[0258] Groups of rats were prepared as above except that the rats received a second catheter into the jugular vein to administer NLX and a catheter into a femoral artery to record MAP) and heart. After 47h, the rats were placed in placed in individual opaque plastic boxes and the free end of the exteriorized jugular vein catheter was connected to an injection line to give NLX. The free end of the arterial line was connected to tubing attached to a computer-coupled pressure transducer to record pulsatile arterial blood pressure. After a 60 min acclimatization period, the rats received a bolus injection of NLX (1.5 mg / kg, IV) and MAP and heart rate were recorded continuously for 45 min. Body temperature and body weight studies

[0259] Groups of rats without a second jugular catheter were prepared as described above. After 47h, the rats were placed in individual opaque plastic boxes and a thermistor probe connected to a telethermometer (Yellow Springs Instruments) to record body temperature was inserted 5-6 cm into the rectum and taped to the tail. The body weights of the rats and body temperatures were recorded every 15 min during acclimatization to establish baseline values. After the 60 min acclimatization, the rats received an intraperitoneal injection of NLX (1.5 mg / kg). Body temperature and weights were recorded every 15 min for 90 min. Data Analyses

[0260] The directly recorded and arithmetically-derived parameters were statistically analyzed. All data are presented as mean ± SEM and were evaluated using one-way and two- way ANOVA followed by Bonferroni corrections for multiple comparisons between means using the error mean square terms from each ANOVA analysis. A P < 0.05 value denoted the initial level of statistical significance that was modified according to the number of comparisons between means. The modified t-statistic is t = (mean group 1 - mean group 2) / [s x (1 / n1 + 1 / n2)1 / 2] where s2= the mean square within groups term from the ANOVA (the square root of this value is used in the modified t-statistic formula) and n1and n2are the number of rats in each group under comparison. Based on Bonferroni's inequality, a conservative critical value for modified t-statistics can be obtained from tables of t- distribution using a significance level of P / m, where m is the number of comparisons between groups to be made. The degrees of freedom are those of the mean square for within group variation from the ANOVA table. The critical Bonferroni value can be approximated from tables of the normal curve by t* = z + (z + z3) / 4n, with n being the degrees of freedom and z being the critical normal curve value for P / m. Wallenstein et al (1980) first demonstrated that the Bonferroni procedure is preferable for general use because it provides critical values that are lower than those of other procedures when the number of comparisons can be limited andwill be slightly larger than those of other procedures if many comparisons are made. Statistical analyses were performed with the aid of GraphPad Prism software (GraphPad Software, Inc., La Jolla, CA). F- and P-statistics associated with ANOVA analyses of the data in Figs.1-5 are given in the respective figure legends. RESULTS L-CYSee prevention of physical dependence to morphine – 36h studies

[0261] The behavioral withdrawal phenomena elicited by injection of NLX (1.5 mg / kg, IP) in rats that were receiving morphine (150 mg / kg, SC) plus a continuous infusion of vehicle (saline, 20 μL / h, IV) or L-cysteine (20.8 μmol / kg / h, IV) or L-CYSee (20.8 μmol / kg / h, IV) for 36h are shown in Fig.1. As can be seen, the injection of NLX to rats receiving the infusion of vehicle elicited substantial increases in jumping behavior (jumps), wet-dog shakes (WDS) rearing behavior (rears) fore-paw licking (FPL), circling behavior (circles), full-body writhing (writhes) and sneezing (sneezes). These responses were similar in magnitude in rats that were receiving an infusion of L-cysteine or L-SERee. In contrast, NLX-precipitated phenomena (except for sneezing) were dramatically diminished in rats that were receiving the continuous infusion of L-CYSee. In addition, the NLX-precipitated withdrawal signs were fully expressed in rats that were receiving an infusion of L-SERee (see Table 1). Table 1 - Naloxone-precipitated withdrawal signs in morphine-treated rats receiving continuous infusion of vehicle or L-serine ethyl esterL-SERee, L-serine ethyl ester. The data are presented as mean ± SEM. There were no between-group differences in body weights or withdrawal signs (P > 0.05, for all comparisons)

[0262] The increases in apneic events and elevations in MAP and heart rate elicited by injection of NLX (1.5 mg / kg) in rats that were receiving morphine (150 mg / kg, SC) and continuous infusion of vehicle or L-cysteine or L-CYSee are summarized in Fig.2. The injection of NLX in rats that were receiving vehicle elicited substantial increases in the number of apneas and increases in MAP (sustained hypertension) and in heart rate (HR, tachycardia). These NLX-precipitated events were similar in magnitude in the rats that were receiving infusion of L-cysteine. In contrast, the NLX-precipitated withdrawal phenomena were markedly reduced in rats receiving the continuous infusion of L-CYSee. Full details of the actual values for MAP and heart rate before and after injection of NLX in morphine- treated rats receiving infusions of vehicle, L-cysteine or L-CYSee are shown in Table 2. Resting MAP and heart rate values before the injection of NLX were similar in the 3 groups of rats. The injection of NLX elicited rises in MAP and heart rate as described above. Table 2 - Changes in mean arterial blood pressure and heart rate elicited by injection of naloxone HCl in freely-moving male rats receiving morphine with co-infusion of vehicle, L- CYSee or L-cysteine for 36 hours

[0263] The changes in body temperature and body weights elicited by injection of NLX (1.5 mg / kg) in rats that were receiving morphine (150 mg / kg, SC) and continuous infusion of vehicle or L-cysteine or L-CYSee are summarized in the left-hand panels of Fig.3. The injection of NLX elicited marked decreases in body temperature and body weights that were similar in magnitude in rats receiving infusion of vehicle or L-cysteine. The NLX-induced responses were markedly smaller in the rats that were receiving an infusion of L-CYSee. Full details of the actual body temperature and body weight before and after injection of NLX in morphine-treated rats that were receiving infusions of vehicle, L-cysteine or L-CYSee are shown in Table 3. Resting body temperature and body weight values before the injection of NLX were similar in the 3 groups of rats. Body weights were similar in all 3 groups. After 36h of morphine treatment, body temperatures were elevated by just over 0.5oC in rats receiving infusions of vehicle or L-cysteine. Body temperature was not raised in rats receiving L-CYSee. The injection of NLX elicited substantial decreases in body temperature and body weights in the vehicle- or L-cysteine-infusion groups and much smaller responses in the rats receiving the infusion of L-CYSee. L-CYSee reversal of physical dependence to morphine – 48h studies

[0264] The behavioral withdrawal phenomena elicited by the injection of NLX (1.5 mg / kg, IP) in rats that were receiving morphine (150 mg / kg, SC) for 48h plus a continuous infusion of vehicle (saline, 20 μL / h, IV) or L-cysteine (20.8 μmol / kg / h, IV) or L-CYSee (20.8 μmol / kg / h, IV) beginning after 36h are shown in Fig.4. The injection of NLX to rats receiving vehicle infusion elicited substantial increases in withdrawal behaviors. These responses were similar in magnitude in rats receiving infusion of L-cysteine whereas withdrawal phenomena (except for sneezing) were dramatically diminished in rats receiving infusion of L-CYSee for 12h. NLX-precipitated withdrawal signs in rats receiving an infusion of L-SERee were similar to those receiving infusion of vehicle (see Table 3).Table 3 - Changes in body temperature and body weights elicited by the injection of naloxone HCl in rats treated receiving morphine with co-infusion of vehicle, L-cysteine or L-CYSee for 36 hours

[0265] The increases in apneic events and elevations in MAP and heart rate elicited by injection of NLX (1.5 mg / kg, IP) in rats receiving morphine (150 mg / kg, SC) for 48h and continuous infusion of vehicle or L-cysteine or L-CYSee beginning at 36h are shown in Fig. 5. NLX elicited substantial increases in apneas and elevations in MAP (hypertension) and in heart rate (HR, tachycardia) in rats receiving infusion of vehicle and similar responses in rats receiving infusion of L-cysteine. In contrast, NLX-precipitated withdrawal phenomena were markedly reduced in rats receiving L-CYSee infusion of. Full details of the actual values for MAP and heart rate before and after injection of NLX in morphine-treated rats receiving infusions of vehicle, L-cysteine or L-CYSee with are shown in Table 4. Resting MAP andheart rate values before injection of NLX were similar in the 3 groups. Injection of NLX elicited rises in MAP and heart rate as described above. Table 4 - Changes in mean arterial blood pressure and heart rate elicited by the injection of naloxone HCl in rats receiving morphine for 48 hours with co-infusion of vehicle, L-cysteine or L-CYSee for 12 hours starting at 36 hours of morphine administration

[0266] The changes in body temperature and body weights elicited by injection of NLX (1.5 mg / kg) in rats receiving morphine (150 mg / kg, SC) and infusion of vehicle or L-cysteine or L-CYSee are summarized in the right-hand panels of Fig.3. The injection of NLX elicited pronounced decreases in body temperature and body weights that were similar in magnitude in rats receiving vehicle or L-cysteine. These withdrawal responses were markedly smaller in rats receiving L-CYSee. Full details of the body temperature and body weight before and after injection of NLX in morphine-treated rats that were receiving infusions of vehicle, L- cysteine or L-CYSee are shown in Table 5. Resting body temperature and body weight values before the injection of NLX were similar in the 3 groups of rats. After 48h of morphine treatment, body temperatures were elevated by just over 0.5oC in rats receiving the infusions of vehicle or L-cysteine. Body weights were similar in all 3 groups. Body temperature was not elevated in rats receiving infusion of L-CYSee. The injection of NLX elicited substantial decreases in body temperature and body weights in the vehicle- or L-cysteine-infusion groups and much smaller responses in the rats receiving the infusion of L- CYSee. Table 5 - Changes in body temperature and body weights elicited by the injection of naloxone HCl in rats treated with morphine for 48 hours with co-infusion of L-CYSee or L-cysteine for 12 hours starting at 36 hours of morphine administration

[0267] The first set of major observations of this example was that co-infusion of the L- thiol ester, L-CYSee, markedly reduced the expression of multiple withdrawal signs (behavioral, cardiorespiratory, body weight loss and hypothermia) elicited by injection of the opioid receptor antagonist, NLX, in male Sprague-Dawley rats treated for 36h with slow- release morphine emulsion. Behavioral withdrawal signs indicative of the rats having become physically-dependent on morphine such as jumping, wet-dog shakes, rearing, fore-paw licking, circling, writhing and sneezing (rapid expulsions of air) as well as the decreases in body weight and body temperature were consistent with previous reports with this slow-release morphine model and with a wide variety of other administration protocols used to induce morphine dependence. The increases in MAP and heart rate elicited by NLX are new findings in our morphine-dependence model but are in full agreement with evidence that NLX-precipitated withdrawal is associated with hypertension and tachycardia in experimental animals and humans that is due to globalized activation of the sympathetic nervous system. Finally, our finding that NLX elicited a substantial increase in apneic events is new to our morphine-dependence model but consistent with such findings in rats and humans. The inability of L-cysteine to modify the NLX-precipitated withdrawal phenomena certainly suggests that the efficacy of L-CYSee involves entry of this cell-penetrant L-thiol ester. Moreover, the inability of L-SERee to prevent the acquisition of physical dependence to morphine indicates that the sulfur atom is vital to the ability of L-CYSee to prevent the intracellular processes within the brain by which morphine induces physical dependence.

[0268] At present, we do not know how L-CYSee prevents the development of physical dependence to morphine. The mechanisms by which L-thiolesters exert their biological effects are likely to be multi-factorial and possibly include (1) direct binding of L-CYSee to plasma membrane / intracellular proteins such as ion-channels, receptors and enzymes that alters the activities of the proteins by mechanisms not associated with changes in redox status of the proteins (yet to be substantiated) (2) formation of thiol adducts such as D-glucose:L- cysteine and mixed disulfides in the blood , (3) modulation of redox status (e.g., reduction of L-cystine to L-cysteine) and activity of plasma membrane proteins such as Kv1.2 K+-channels and after entry into cells, redox modulation of functional intracellular proteins, (4) formation of S-thiolated proteins such as S-cysteinylated, S-cysteinylglycinylated and S- glutathionylated proteins in plasma membranes and cells, (5) conversion of L-CYSee to L- cysteine by membrane associated esterases, which then enters into multiple metabolic pathways including those that generate hydrogen sulfide via the sequential actions of L- cysteine aminotransferase and cystathionine γ-lyase in peripheral and central tissues including the carotid bodies, (6) conversion of L-thiolesters to cysteine sulfenics, sulfonics and sulfonics via cysteine dioxygenase, and (7) formation of S-nitroso-L-cysteine, an endogenous S-nitrosothiol with many substantial roles in intracellular signaling cascades including those controlling cardiorespiratory function and those involved in the attenuation of OIRD. Any or all of these mechanisms (and possibly those not mentioned above) mayinteract with proposed signaling pathways involved in the acquisition of physical dependence to opioids such as morphine and the expression of the NLX-precipitated withdrawal syndrome including those involving N-methyl D-aspartate (NMDA) glutamatergic receptors corticotropin releasing factor (CRF) receptor CRF1, tachykinin receptors, voltage-gated Ca2+- channels, adenylyl cyclase superactivation and opioid receptor phosphorylation and the nitric oxide-cGMP signaling cascade. Since L-CYSee blunted the expression of all NLX- precipitated behavioral (except for sneezing), physical (body weight loss, hypothermia), and cardiorespiratory (hypertension, tachycardia, incidence of apneas) phenomena, it is tempting to assume that the L-CYSee interrupts some fundamental intracellular processes(es) that are essential to the development of physical dependence to morphine.

[0269] The second set of novel set of findings was that the introduction of L-CYSee infusion 36h into the morphine administration period appeared to reverse the established physical dependence to the opioid as assessed at 48h (i.e., within 12h). Specifically, NLX- precipitated behavioral phenomena (except for sneezing), hypertension, tachycardia, apneas, hypothermia and body weight loss were markedly fewer in the rats that had received L- CYSee for 12h. Again, the lack of effect of L-cysteine and L-SERee suggests that the intracellular delivery of L-CYSee and its sulfur atom (and associated thiol chemistry) are essential to the ability of the L-thiol ester to reverse established physical dependence to morphine. Again, we do not know how L-CYSee reverses physical dependence to morphine but any / none of the mechanisms discussed above including its potent antioxidant properties may be involved. The therapeutics and bioactive compounds that reverse established physical dependence include, L-histidine and certain histamine receptor sub-type agonists, melatonin, the antioxidant quercetin, the serotonin-reuptake inhibitor, fluoxetine, the nitric oxide synthase inhibitor, L-NG-nitroarginine methyl ester, inhibitors of Ca2+ / calmodulin- dependent protein kinase II, the β2-AR antagonist, butoxamine, adrenomedullin receptor antagonists, the antipsychotic (dopamine D2 receptor antagonist) haloperidol and positive allosteric modulators of AMPA (α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid) glutamatergic receptors. The ability of L-CYSee to reverse established physical dependence to morphine is of great clinical relevance and opens the way for future studies on this and other bioactive L,D-thiol esters and related compounds with respect to their the ability toreverse physical dependence to morphine and other opioids including heroin and fentanyl and establishing mechanisms of action.

[0270] A key question arising from these novel studies relates to the potential use of D- thiolesters as therapeutics for key clinical problems associated with opioid analgesics: With respect to use in humans, (1) if L-CYSee attenuates / blocks self-administration of opioids in OUD patients, adding it to prescription opioids may result in lower abuse or addiction potential; (2) If L-CYSee attenuates or blocks development of physical dependence to opioids, then adding it to prescription opioids will minimize and may potentially eliminate physical dependence in individuals who receive opioids long-term (day in and day out for weeks / months); (3) If L-CYSee attenuates / blocks tachyphylaxis to opioid analgesia or hyperalgesia caused by opioids in many people, then the addition of L-CYSee to prescription opioids will maintain their analgesic efficacy over long periods of time, eliminating the development of tolerance, need for escalating doses, and potential complications of hyperalgesia; (4) If L-CYSee has several of the advantageous effects observed in rodents, then adding it to opioid analgesics would multiply the beneficial aspects of the opioids; (5) If L-CYSee prevents development of physical dependence, and especially if it is introduced to an individual with physical dependence and attenuates / blocks opioid withdrawal, it could be used as an outpatient / inpatient medication to manage opioid withdrawal in those who are iatrogenically physically-dependent (long-term opioid prescriptions) or those who are addicted and physically dependent; (6) If L-CYSee attenuates / blocks euphoria and / or the development of physiological dependence to opioids, then it would be a good medication for medication-assisted treatment (MAT) and a potentially good drug for harm reduction interventions in people with OUD who are not interested in the psychosocial aspects of counseling and treatment; (7) As some patients with a history of OUD who are currently sober need opioids for treatment of acute or chronic pain syndromes, this L-thiolester, if it attenuates or blocks euphoria and physical dependence, could be added to opioid analgesics when given to people with a history of OUD, thereby eliminating the risk of opioid analgesics precipitating euphoria, drug cravings and their markedly increased risk of relapse; (8) If L- CYSee attenuates / blocks euphoria from chemically mediated dopamine surges within the ventral tegmentum, nucleus accumbens, or medial prefrontal cortex, where brain rewarding euphoria-producing dopamine surge happens from all drugs of abuse / addiction then it will be useful in treatment of OUD and other SUDs; (9) If L-CYSee attenuates / blocks euphoria fromchemically-mediated dopamine surges, it could be combined with or added to all controlled prescription drugs resulting in an abuse-resistant or non-abusable form of prescribed opioids, benzodiazepines, and psychostimulants, for example. In relation to point (1), we recently showed that co-administration of the D isomer, D-cysteine ethyl ester, with fentanyl prevents the development of fentanyl-induced conditioned place preference in both male and female rats. Thus, L,D-thiol esters likely reduce the rewarding properties of opioids and reduce their addictive potential.

[0271] This example demonstrates that systemic infusion of the membrane-permeable L-thiol ester, L-CYSee, prevents the development of physical dependence to morphine in male Sprague-Dawley rats by mechanisms dependent on thiol biochemistry. As important, this example demonstrates that L-CYSee reverses established dependence to morphine in such rats also by thiol-dependent processes. Delineating the exact thiol-dependent signaling pathways will add greatly to our understanding of the processes by which opioid induce dependence and how the bioactive L-thiol esters exert their effects. The exact thiol- dependent processes. Our example was spurred by the ground-breaking work of Trivedi, Deth and colleagues which added greatly to our understanding of the mechanisms by which opioids cause physical dependence and psychological addiction. Their evidence that morphine may cause dependence / addiction by blocking the entry of L-cysteine into neurons by inhibition of EAA3 / EAAC1 transporter prompted our studies with the membrane- permeable, L-thiol ester, L-CYSee. The findings that L-CYSee markedly reduced the majority of NLX-precipitated withdrawal phenomena speaks to the key role that decreased levels of L-cysteine entry into cells plays in establishing physical dependence to morphine. The lone withdrawal phenomenon that was not ameliorated by L-CYSee was sneezing, a key feature of the opioid withdrawal response in humans and experimental animals. We are currently trying to understand the current state of knowledge about the neural mechanisms responsible for sneezing to see if that can give insights into the signaling pathways that are / are not involved in the actions of L-CYSee. The present findings add to our knowledge about the efficacy of L,D-thiolesters, such as L-CYSee L-GSHee, D-CYSee, D-cystine di(m)ethylester and the free radical-superoxide anion scavenger, Tempol on the pharmacological actions of opioids.Example 2

[0272] Avariety of highly cell-permeable analogues of L-NAC including L-NACme, L- NAC ethyl ester, and L-NAC propyl ester, have been developed (see Fig.9 for chemical structures). The rapid conversion of L-NACme to L-cysteine and L-glutathione in cells, would in theory overcome the loss of these L-thiols elicited by opioids and restore biochemical / redox signaling including by potential formation of S-nitrosylated analogues of these thiols such as S-nitroso-L-NACme, S-nitroso-L-cysteine, which markedly reduce respiratory depression induced by fentanyl and morphine, and S-nitroso-L-glutathione. To date, the possibility that L-NAC or L-NACme prevent / attenuate acquisition of psychological addiction / physical dependence to fentanyl has not been determined. Numerous administration / dosage paradigms have been employed to develop addiction to fentanyl in male and female rats (self-administration studies), and physical dependence via continuous infusion and multiple injections. We decided to first determine (1) whether co-injections of L-NAC or L-NACme diminish / prevent development of physical dependence elicited by injections of fentanyl in male Sprague-Dawley rats, as assessed by the strength of the withdrawal response elicited by injection of the µ-OR antagonist, naloxone HCl (NLX), and (2) whether introduction of co-injections of L-NAC or L-NACme to rats that are physically- dependent on fentanyl could reverse this dependence, again assessed by the expression of the NLX-precipitated withdrawal responses. The fentanyl dose-regime consisting of twice daily intravenous injections of 125 µmol / kg, IV) was based on daily dose ranges employed by others. The behavioral, cardiorespiratory, thermoregulatory and body weight changes were chosen on the basis of previous studies determining the scope and types of expected withdrawal phenomena. Materials and Methods Permissions, rats, and surgical procedures

[0273] All studies were performed as per the NIH Guide for Care and Use of Laboratory Animals (NIH Publication No.80-23) revised in 1996, and ARRIVE (Animal Research: Reporting of In Vivo Experiments) guidelines (http: / / www.nc3rs.org.uk / page. asp? id=1357). All protocols involving rats were approved by the Animal Care and Use Committees of the University of Virginia and Galleon Pharmaceuticals. Adult male SpragueDawley rats purchased from Harlan Industries (Madison, WI, USA) were given five days to recover from transportation before being subject to surgeries. Fentanyl citrate, L-NAC and L- NACme powders were obtained from Sigma-Aldrich (St. Louis, MO, USA) and divided into 100 mg amounts under N2gas and stored at 4oC. Solutions of L-NAC and L-NACme (dissolved in saline and brought to pH 7.2 with 0.1M NaOH at room temperature) were prepared immediately before use. Naloxone HCl (Sigma-Aldrich, St. Louis, MO, USA) was dissolved in normal saline. All arterial and venous catheters were flushed with 0.3 ml of phosphate-buffered saline (0.1 M, pH 7.4) 3-4h before commencement of the example. All studies were done in a room with relative humidity of 49 ± 2% and temperature of 21.4 ± 0.2°C. Protocols to determine the abilities of L-NAC or L-NACme to prevent the development of physical dependence to fentanyl Behavioral studies

[0274] Three groups of rats received a jugular vein catheter of PE-10 connected to PE- 50 (Intramedic; Becton & Dickinson, Franklin Drive, NJ, USA) under 2-3% isoflurane anesthesia, to allow injections of test agents. Rats were given 4 days to recover from surgery. Example 1 - 5 co-injections: Groups of rats (n=9 per group) received co-injections of vehicle (100 µL / 100g body weight) + fentanyl (125 µg / kg, IV), L-NAC (500 µmol / kg, IV) + fentanyl (125 µg / kg, IV) or L-NACme (500 µmol / kg) + fentanyl (125 µg / kg, IV) given apart at 8 am and 8 PM on days 1 and 2 and at 8 AM on day 3. Example 1 - 10 co-injections: Other groups of rats (n=9 per group) received co-injections vehicle (100 µL / 100g body weight) + fentanyl (125 µg / kg, IV), L-NAC (500 µmol / kg, IV) + fentanyl (125 µg / kg, IV) or L-NACme (500 µmol / kg) + fentanyl (125 µg / kg, IV) given 90 sec apart at 8 AM and 8 PM on days 1 to 4. The rats received injection 9 at 8 AM on day 5 and injection 10 at 2 PM to allow for subsequent NLX challenges to be given. Ninety min after the 5th or 10th set of co-injections, rats were placed in individual opaque plastic boxes and after 30 min, they were injected with NLX (1.5 mg / kg) and behavioral phenomena were scored for 45 min by 3 scorers. Scored phenomena were: Jumping behavior - all 4 paws of the ground - jumps; Wet dog shakes - whole body shakes as if to shed water from fur; Rearing behavior - rearing on hind legs - rears; Episodes of fore-paw licking – FPL; Circling behavior - Complete 360orotation;Writhes – full body contortion; Sneezes - episodes of sneezing – abrupt expulsion of air that often disturbed the fine bedding material. Plethysmography ventilatory studies

[0275] One hour before co-injections 5 or 10 were to be given (as described above), rats were placed into individual whole body plethysmography chambers to record ventilatory parameters. The free end of the exteriorized venous catheter was connected to a swivel assembly housed in the lid of the plethysmography chamber and after 60 min acclimatization, the rats received co-injections 5 or 10 and after 90 min they received an injection of NLX (1.5 mg / kg, IV). Parameters including frequency of breathing, tidal volume, minute ventilation, peak inspiratory and expiratory flows and a variety of non-eupneic breathing indices were recorded (to be reported in detail elsewhere) with the number of apneas of > 1.5 sec in duration reported here. Cardiovascular studies

[0276] Groups of rats (n=9 rats per group) were implanted with a jugular vein catheter to inject drugs and a catheter into a femoral artery to continuously record mean arterial blood pressure (MAP) and heart rate. The rats were given 3 days to recover from surgery before the protocols began. The arterial lines were kept patent throughout the acclimatization period and 3 or 5 day example protocols by connecting the arterial line to an infusion pump (Standard Infuse-Withdraw Pump 11 Pico Plus Elite Programmable Syringe Pump; Harvard Apparatus, MA, USA) delivering normal saline at 20 µL / h). One hour before co-injections 5 or 10 were to be given (as described above), the rats were placed in individual opaque plastic boxes and prepared to give drugs and to continuously record pulsatile arterial blood pressure, MAP and heart rate. After 60 min acclimatization, the rats received co-injections 5 or 10 and then after 90 min they received NLX (1.5 mg / kg, IV) and cardiovascular parameters recorded continuously for a further 90 min. Body temperature and body weight studies

[0277] Groups of rats (n=9 per group) were placed in individual opaque plastic boxes 1h before co-injections 5 or 10 of drug combinations were to be given (as above). A thermistor probe connected to a telethermometer (Yellow Springs Instruments) to record body temperature was inserted 5-6 cm into the rectum and taped to the tail. Body weights of therats and body temperatures were recorded every 15 min during acclimatization to establish baseline values and at 15 min intervals throughout the injection protocols. After 60 min acclimatization, the rats received co-injections 5 or 10 and after 90 min an injection of NLX (1.5 mg / kg, IV) and body weights and body temperatures were recorded for another 90 min. Protocols to determine the abilities of L-NAC or L-NACme to reverse fentanyl dependence Behavioral studies

[0278] Groups of rats (n=9 per group) received 5 injections of fentanyl (125 µg / kg, IV) at 8 AM and 8 PM as described above. These rats then received co-injections 6-10 of fentanyl (125 µg / kg, IV) + vehicle, L-NAC (500 µmol / kg, IV) + fentanyl (125 µg / kg, IV) or L-NACme (500 µmol / kg) + fentanyl (125 µg / kg, IV) given 90 sec apart. Co-injections 6 were given at 8 PM, co-injections 7 at 8 AM, co-injections 8 at 8 PM, co-injections 9 at 8 AM were given at 8 AM and co-injection 10 was given at 2 PM to allow for the experiments to be performed. Immediately after co-injections 10 we given, the rats were placed in individual opaque plastic boxes and after a 90 min period of acclimatization, the rats received an injection of NLX (1.5 mg / kg, IV) and behavioral phenomena (as detailed above) were scored for 45 min by at least 3 scorers. Plethysmography ventilatory studies

[0279] One hour before co-injections 10 were to be given, rats were put into individual whole body plethysmography chambers to record ventilatory parameters. The free end of the exteriorized venous catheter was connected to the swivel assembly and after 60 min acclimatization, the rats received the 10thset of co-injections and after 90 min they received NLX (1.5 mg / kg, IV). Ventilatory parameters and non-eupneic breathing indices were recorded (to be reported elsewhere) with the number of apneas of > 1.5 sec in length reported here. Cardiovascular studies

[0280] One hour before the 10thset of co-injections were given, groups of rats (n=9 rats per group) were put in individual opaque plastic boxes and the free end of the exteriorized jugular vein catheter was connected to an injection line to give drugs. The free end of the arterial catheter was connected to tubing attached to a computer-coupled pressure transducer(Cabe Lab, Inc.) to continuously record pulsatile arterial blood pressure to derive MAP and heart rate. After 60 min acclimatization, rats received co-injections 5 or 10 and then after 90 min an injection of NLX (1.5 mg / kg, IV) and cardiovascular parameters were recorded continuously for a further 90 min. Body temperature and body weight studies

[0281] Groups of rats (n=9 per group) were placed in individual opaque plastic boxes 1h before the 10thset of co-injections were to be given (as described above). A thermistor probe connected to a telethermometer (Yellow Springs Instruments) to record body temperature was inserted 5-6 cm into the rectum and taped to the tail. Body weights of the rats and body temperatures were recorded every 15 min during acclimatization to establish baseline values and at 15 min intervals throughout the injection protocols. After 60 min acclimatization, the rats received co-injections 10 and after 90 min they were injected with NLX (1.5 mg / kg, IV) and body weights and body temperatures were recorded for another 90 min. Data Analyses

[0282] All data are presented as mean ± SEM and were analyzed by one-way and two- way ANOVA followed by Bonferroni corrections for multiple comparisons between means using the error mean square terms from each ANOVA analysis as described previously. A P < 0.05 value denoted the initial level of significance that was modified as per the number of comparisons between means. The modified t-statistic is t = (mean group 1 - mean group 2) / [s x (1 / n1+ 1 / n2)1 / 2] where s2= mean square within groups term from the ANOVA (the square root of this value is used in the modified t-statistic formula) and n1 and n2 are the number of rats in each group under comparison. Based on Bonferroni's inequality, a conservative critical value for modified t-statistics is obtained from tables of t-distribution using a significance level of P / m, where m is the number of comparisons between groups to be performed. The degrees of freedom are those for the mean square for within group variation from the ANOVA table. In most situations, the critical Bonferroni value cannot be found in conventional tables of the t-distribution but can be approximated from tables of the normal curve by t* = z + (z + z3) / 4n, with n being the degrees of freedom and z being the critical normal curve value for P / m. Wallenstein et al. demonstrated that the Bonferroni procedurethe widest range of applications because it provides critical values that are lower than those of other procedures when the number of comparisons can be limited (and will be slightly larger than those of other procedures if many comparisons are made. Statistical analyses were performed with the aid of GraphPad Prism software (GraphPad Software, Inc., La Jolla, CA). F- and P-statistics associated with analyses of the data in Fig.6-8 are given in Table 6. Results L-NAC and L-NACme prevention of physical dependence to fentanyl

[0283] The behavioral phenomena elicited by the injection of NLX (1.5 mg / kg, IV) in rats that had received 5 co-injections of fentanyl (125 μg / kg, IV) + vehicle or L-NAC (500 μmol / kg, IV) or L-NACme (500 μmol / kg, IV) are shown in panel A of Fig.6. Injection of NLX to rats that received fentanyl + vehicle elicited jumping behavior (jumps), wet-dog shakes (WDS) rearing behavior (rears) fore-paw licking (FPL), circling behavior (circles), full-body writhing (writhes) and episodes of sneezing (sneezes). NLX-precipitated phenomena (except for sneezing) were diminished in rats that received fentanyl + L-NAC and markedly diminished in rats that received fentanyl + L-NACme. As seen in Panel B, NLX (1.5 mg / kg)-induced increases in MAP, heart rate and incidence of apneic events (> 1.5 sec) greater were reduced by L-NAC and markedly reduced by L-NACme. As seen in Panels C and D, the NLX-induced falls in body weight and body temperature, respectively, were less in rats that received fentanyl + L-NAC and markedly less in rats that received fentanyl + L- NACme. Behavioral phenomena elicited by the injection of NLX (1.5 mg / kg, IV) in rats that had received 10 co-injections of fentanyl (125 μg / kg, IV) + vehicle or L-NAC (500 μmol / kg, IV) or L-NACme (500 μmol / kg, IV) are shown in panel A of Fig.7. The injection of NLX to rats that received fentanyl + vehicle elicited qualitatively similar responses as described above except that they were more numerous. The NLX-precipitated withdrawal phenomena (except for the sneezing) were diminished in rats that had received fentanyl + L-NAC and markedly diminished in rats that had received fentanyl + L-NACme. As seen in Panel B, the NLX (1.5 mg / kg)-induced increases in MAP, heart rate and incidence of apneic events (> 1.5 sec) were reduced by L-NAC and markedly diminished by L-NACme. As seen in Panels C and D, the NLX-induced decreases in body weight and body temperature were diminished in rats that had received fentanyl + L-NAC and markedly diminished in rats that had received fentanyl + L-NACme.L-NAC and L-NACme reversal of established physical dependence to fentanyl

[0284] The behavioral responses elicited by the injection of NLX (1.5 mg / kg, IV) in rats that had received 10 injections of fentanyl (125 µg / kg, IV) plus 5 co-injections of vehicle, L-NAC (500 µmol / kg, IV) or L-NACme (500 µmol / kg, IV) beginning at fentanyl injection 6 are summarized in Panel A of Fig.8.

[0285] Sneezes aside, the NLX-precipitated phenomena were reduced in rats that received co-injections of L-NAC and markedly reduced in rats that received L-NACme. As seen in Panel B, the NLX-precipitated increases in MAP, heart rate and incidence of apneas (> 1.5 sec) in duration, were reduced in rats that received L-NAC and markedly reduced in those that received L-NACme. As seen in Panels C and D, the NLX-induced decreases in body weight and body temperature, were not as marked in rats that received fentanyl + L- NAC and were markedly less in rats that received fentanyl + L-NACme. Changes in variables during the progression of the protocols

[0286] The actual body weights and arithmetic changes in weights at key points of Studies 1-4 for co-injections 1-5 and co-injections 1-10 studies are shown in Table 6. The rats that received fentanyl + vehicle lost body weight (arithmetic change from pre-drug values). Body weights increased in rats that received L-NAC and especially L-NACme. Regarding the effects of NLX (Groups 4), the decreases in body weight were less in L-NAC- treated rats and markedly less in L-NACme-treated rats. Actual body weights and arithmetic changes in weights at key points of Studies 1-4 for co-injections 6-10 studies are shown in Table 6. In rats that received fentanyl +vehicle, the loss in body weight after 10 injections were greater than after 5 injections whereas the increases in body weights in the rats that received fentanyl + L-NAC or fentanyl + L-NACme after co-injections 10 were greater than after 5 co-injections. Regarding the effects of NLX (Groups 4), the decreases in body weight were less in L-NAC-treated rats and markedly less in L-NACme-treated rats. Actual body temperatures and arithmetic changes in temperatures at key points of for co-injections 1-5 studies, co-injections 1-10 studies and co-injection 6-10 studies are shown in Table 7. The rats that received 5 or co-injections of fentanyl + vehicle displayed a hyperthermia that was less pronounced in rats that received fentanyl + L-NAC and especially fentanyl +L-NACme. Similarly, the rats that received 10 injections of fentanyl plus co-injections of vehicle, L- NAC or L-NACme with injections 6-10 of fentanyl displayed hyperthermia that was lessintense in rats that received L-NAC and especially in those that received L-NACme. Again, the NLX-precipitated falls in body temperature seen in rats that received vehicle + L-NAC and especially L-NACme were smaller than in fentanyl + vehicle-injected rats. Table 6 - Actual body Weights and their arithmetic changes at key points of exampleTable 7 - Actual body temperatures and their arithmetic changes at key points of example

[0287] The actual MAP, heart rate and ∆Heart Rate / ∆MAP (cardiovascular index) values before and after administration of NLX plus arithmetic changes in these parameters for co-injections 1-5, co-injections 1-10 and co-injections 6-10 studies are shown in Table 8. There were no between-group differences in resting parameters prior to the administration of NLX (note that the injections of fentanyl elicited decreases in MAP and heart rate that had fully resolved by the time the pre-NLX measurements were taken. The development of tolerance to the cardiovascular effects of fentanyl will be reported elsewhere). The NLX- precipitated increases in MAP and heart rate in rats that received 10 co-injections of fentanyl + vehicle were greater than those that received 5 co-injections of fentanyl + vehicle. The NLX-precipitated increases in MAP and heart rate were smaller in rats that received fentanyl + L-NAC and especially fentanyl + L-NACme in the co-injections 1-5, co-injections 1-10 and co-injections 6-10 studies. The arithmetic changes in ∆Heart Rate / ∆MAP values (see column denoted Delta) were enhanced in the rats that received co-injections 1-10 and co- injections 6-10 of fentanyl + vehicle. These ratios were markedly diminished in rats that received co-injections of fentanyl + L-NACme for the co-injections 1-5, co-injections 1-10 and co-injections 6-10 studies. Table 8 -Cardiorespiratory responses elicited by the injection of naloxone HCl

[0288] This example demonstrates that twice-daily injections of fentanyl (5 or 10 injections of 125 µg / kg, IV) elicit physical dependence in male Sprague-Dawley rats on the basis of the pronounced withdrawal syndrome elicited by injection of NLX. The behavioral withdrawal signs indicative of the rats having become physically-dependent on fentanyl such as jumping, wet-dog shakes, rearing, circling, fore-paw licking, writhing and sneezing as well as the decreases in body weight and body temperature are consistent with reports on the patterns of NLX-precipitated phenomena observed in fentanyl-administration protocols and in a wide variety of other opioid administration protocols used to induce dependence. The increases in MAP and heart rate elicited by NLX are new findings with respect to our fentanyl-dependence model but are consistent with reports that NLX-precipitated withdrawal is associated with hypertension and tachycardia in experimental animals and humans due to globalized activation of sympathetic nerve activity. Our finding that NLX elicited a substantial increase in apneic events (> 1.5 sec) is a novel finding with respect to fentanyl-dependence model but is consistent with such findings in opioid withdrawal paradigms in rats and humans. The first novel observations in this example were that co-injections of L-NAC and L-NACme appeared to diminish the development of physical dependence to fentanyl on the basis that the NLX-precipitated withdrawal phenomena (behavioral responses, hypertension and tachycardia, hypothermia and body weight loss) were far less than in rats that received co-injections of fentanyl and vehicle. This may have been expected / predicted on the basis that co-injections of L-NAC and L-NACme reduced the body weight loss and development of hyperthermia observed in the rats that received co-injections of fentanyl and vehicle. The findings that L-NACme was much more effective than L-NAC in preventing the development of dependence to fentanyl is most likely due to it being more cell-penetrant than L-NAC and especially into neurons of brain regions involved in the acquisition of physical dependence and addiction.

[0289] At present, we do not know the mechanisms by which L-NAC (and presumably L-NACme) ameliorate the development of physical dependence to fentanyl. The mechanisms may include (1) their antioxidant / reductant properties, resulting in modulation of redox status (e.g., reduction of L-cystine to L-cysteine) and activity of plasma membrane proteins such as Kv1.2K+-channels and after entry into cells, redox modulation of functional intracellular proteins, (2) direct binding of L-NAC and L-NACme to plasma membrane / intracellular proteins such as ion-channels, receptors and enzymes that alters the activities of the proteins by mechanisms not due to redox status of the proteins (yet to be substantiated), (3) formation of thiol adducts such as D-glucose:L-NAC and D-glucose:L-NACme akin to D-glucose:L- cysteine and formation of the disulfides (covalent bond between the sulfur atoms) of L-NAC and L-NACme and mixed disulfides in the blood, and upon deacetylayion to L-cysteine via a family of deacetylases, (1) formation of S-thiolated proteins such as S-cysteinylated, S- cysteinylglycinylated and S-glutathionylated proteins in cell membranes and cells, (2) generation of hydrogen sulfide via the sequential actions of L-cysteine amino-transferase and cystathionine γ-lyase in peripheral and central tissues, including the carotid bodies, (3) conversion of L-thiolesters to cysteine sulfenics, sulfonics and sulfonics via cysteine dioxygenase, and (4) formation of S-nitroso-L-cysteine, an endogenous S-nitrosothiol with many roles in intracellular signaling cascades including those controlling cardiorespiratory function and those involved in the attenuation of OIRD. These mechanisms may interactwith signaling pathways involved in acquisition of physical dependence to opioids such as morphine and expression of the NLX-precipitated withdrawal syndrome including those involving N-methyl D-aspartate (NMDA) glutamatergic receptors, muscarinic receptors, corticotropin releasing factor (CRF) receptor CRF1, tachykinin receptors, voltage-gated Ca2+- channels, adenylyl cyclase super-activation and opioid receptor phosphorylation, oxidative stress, and the nitric oxide-cGMP signaling cascade. Since L-NAC and L-NACme blunted the expression of all NLX-precipitated behavioral (except for sneezing), physical (body weight loss, hypothermia), and cardiorespiratory (hypertension, tachycardia, incidence of apneas) phenomena, it is tempting to assume that they interrupt fundamental intracellular processes essential to the development of morphine dependence. It is important to note that the ability of the redox regulator, α-lipoic acid, to diminish the development of morphine dependence and to modulate NLX-induced biochemical alterations in morphine-dependent mice was enhanced by concurrent administration of L-NAC.

[0290] The second novel finding of this example was that the introduction of co- injections of L-NAC and L-NACme beginning with the 6thand continuing with the 7th-10thinjections of fentanyl appeared to reverse established physical dependence to the opioid. More specifically, the NLX-precipitated behavioral phenomena (except sneezing), hypertension, tachycardia, apneic events, hypothermia and body weight loss were fewer in the rats that had received the co-injections of L-NAC and markedly fewer in those that received L-NACme. Again, we do not know how L-NAC and L-NACme reverse physical dependence to fentanyl but any / none of the mechanisms discussed above including their potent antioxidant properties and abilities to boost intracellular levels of L-cysteine and L- glutathione may be involved (see above). Agents that show efficacy at reversing established physical dependence include, L-histidine and histamine receptor sub-type agonists, melatonin, the antioxidant quercetin, the serotonin-reuptake inhibitor, fluoxetine, the nitric oxide synthase inhibitor, L-NG-nitroarginine methyl ester, inhibitors of Ca2+-calmodulin- dependent protein kinase II, the β2-AR antagonist, butoxamine, adrenomedullin receptor antagonists, the antipsychotic (dopamine D2 receptor antagonist) haloperidol, ATP- dependent K+-channel modulators, and allosteric modulators of AMPA (α-amino-3-hydroxy- 5-methyl-4-isoxazolepropionic acid) glutamate receptors. The ability of L-NAC and especially L-NACme to reverse established physical dependence to fentanyl is of greatclinical relevance and opens the way for future studies on this and other bioactive L,D-thiol esters and related compounds with respect to their the ability to reverse physical dependence to fentanyl and other opioids such as heroin and oxycodone and establishing pharmacological mechanisms of action.

[0291] A key question arising from these novel studies relates to the potential use of L- or D-thiolesters as therapeutics for key clinical problems associated with opioid analgesics: With respect to use in humans, (1) if L-NACme attenuates / blocks self-administration of opioids in OUD patients, adding it to prescription opioids may result in lower abuse or addiction potential; (2) if L-NACme attenuates or blocks development of physical dependence to opioids, then adding it to prescription opioids will minimize and may potentially eliminate physical dependence in individuals who receive opioids long-term (day in and day out for weeks and for months); (3) if L-NACme attenuates / blocks tachyphylaxis to opioid analgesia or hyperalgesia caused by opioids in many people, then adding L-NACme to prescription opioids will maintain their analgesic efficacy over long periods of time, eliminating development of tolerance, the need for escalating doses, and potential complications of hyperalgesia; (4) if L-NACme has several of the advantageous effects found in rodents, then adding it to opioid analgesics would multiply the beneficial aspects of opioids; (5) if L-NACme prevents THE development of physical dependence, and especially if it is introduced to an individual with physical dependence and attenuates / blocks opioid withdrawal, it could be used as an outpatient / inpatient medication to manage opioid withdrawal in those iatrogenically physically-dependent (long-term opioid prescriptions) or those psychologically addicted and / or physically dependent; (6) if L-NACme attenuates / blocks euphoria and / or the development of physiological dependence to opioids, then it would be a good medication for medication-assisted treatment (MAT) and a good drug for harm reduction interventions in people with OUD who are not interested in the psychosocial aspects of counseling and treatment; (7) as some patients with a history of OUD who are currently sober need opioids for treatment of acute or chronic pain syndromes, L- NACme, if it attenuates / blocks euphoria and physical dependence, could be added to opioid analgesics when given to people with a history of OUD, thereby eliminating the risk of opioid analgesics precipitating euphoria, drug cravings and their markedly increased risk of relapse; (8) if L-NACme attenuates / blocks euphoria from chemically mediated dopamine surges within ventral tegmentum, nucleus accumbens, or medial prefrontal cortex, where brainrewarding euphoria-producing dopamine surge happens from all drugs of abuse / addiction then it will be useful in treatment of OUD and other SUDs; (9) if L-NACme attenuates / blocks euphoria from chemically-mediated dopamine surges, it could be combined with or added to all controlled prescription drugs resulting in an abuse-resistant or non- abusable form of prescribed opioids, benzodiazepines, and psychostimulants, for example.

[0292] In conclusion, this example demonstrates that systemic injection of L-NAC and especially the membrane-permeable L-thiol ester, L-NACme, prevents the development of physical dependence to fentanyl and equally importantly that L-NACme reverses established dependence to fentanyl in male Sprague-Dawley rats. Delineating the exact thiol-dependent signaling pathways by which L-NAC and L-NACme exert their effects will add to our understanding of the processes by which opioid induce dependence and how bioactive L-thiol esters exert their effects. Our example was spurred by the ground-breaking work of Trivedi, Deth and colleagues which added greatly to our understanding of the mechanisms by which opioids cause physical dependence and psychological addiction. Their evidence that morphine may cause dependence / addiction by blocking the entry of L-cysteine into neurons by inhibition of EAA3 / EAAC1 transporter prompted our studies with the membrane- permeable, L-cysteine ethyl ester (findings to be submitted), L-NAC (because of immediate clinical applicability) and L-NACme. The findings that L-NACme markedly reduced the large majority of NLX-precipitated withdrawal phenomena speaks to the key role that the loss of L-cysteine entry into cells has in establishing physical dependence to fentanyl. The one withdrawal event not ameliorated by L-NACme was sneezing, a key feature of the opioid withdrawal response in humans and experimental animals. We are trying to develop an understanding of the neural mechanisms responsible for sneezing to gain insights into the signaling pathways that are and are not involved in the actions of L-NACme. The present findings add to our knowledge about the efficacy of L,D-thiolesters on the pharmacological actions of opioids.

[0293] This example provides a strong rationale for consideration of L-NACme and other bioactive membrane-permeable L-thiolesters such as such as L-cysteine (m)ethyl ester, L-glutathione ethyl ester, γ-L-glutamylcysteine ethyl ester and L-cystine di(m)ethylester (as therapeutics to prevent or reverse physical dependence to opioids.Example 3

[0294] We examined whether co-administration of the cell-penetrant L-thiol ester, L- cysteine ethyl ester (L-CYSee), would reduce physical dependence to morphine in male Sprague-Dawley rats and reverse established dependence to the opioid. With respect to preventing the acquisition of dependence, we found that the injection of the opioid-receptor antagonist, naloxone HCl (NLX; 1.5 mg / kg, IP), elicited pronounced withdrawal phenomena in rats which received a subcutaneous depot of morphine (150 mg / kg) for 36h and a continuous infusion of saline (20 µL / h, IV) via osmotic minipumps for this 36h period. Withdrawal phenomena included wet-dog shakes (WDS), jumping, rearing, forepaw licking (FPL), 360ocircling, writhing, apneas, cardiovascular (pressor, tachycardic) responses, hypothermia and body weight loss. Remarkably, NLX elicited substantially fewer withdrawal phenomena in rats that received an infusion of L-CYSee (20.8 μmol / kg / h, IV) for 36h. With respect to reversing acquired dependence, we first established that NLX precipitated a marked withdrawal syndrome in rats that had received subcutaneous depots of morphine (150 mg / kg) for 48h) and a co-infusion of vehicle. Again, the NLX-precipitated withdrawal phenomena were markedly reduced in morphine-treated (150 mg / kg for 48h) rats that began receiving an infusion of L-CYSee (20.8 µmol / kg / h, IV) at 36h. Of equal importance was that the infusion of L-cysteine or L-serine ethyl ester (both at 20.8 µmol / kg / h, IV) were not able to mimic the effects of L-CYSee. As such, it appears that L-CYSee attenuates the development of physical dependence to morphine in male rats and reverses dependence acquired prior to administration of L-CYSee, most likely by intracellular actions within the brain. The lack of effect of L-serine ethyl ester (oxygen atom instead of sulfur atom) implicates thiol biochemistry in the efficacy of L-CYSee. This attribute L-CYSee is matched by the ability of this and other L-thiolesters such as L-cysteine methyl ester and L- glutathione ethyl ester to prevent and / or reverse the actions of morphine and fentanyl on ventilatory parameters, arterial blood-gas chemistry and Alveolar-arterial gradient (index of alveolar gas-exchange) in freely moving rats without compromising opioid-induced analgesia or sedation. In pursuing potential mechanisms of action of the L-thiol esters we found somewhat unexpectedly that D-thiol esters such as D-cysteine ethyl ester (D-CYSee), D- cystine diethyl ester (D-CYSdiee) and D-cysteine dimethyl ester (D-CYSdime) were also able to reverse the adverse effects of morphine on breathing without compromising analgesia.

[0295] We recently reported that the co-administration of D-CYSee with fentanyl prevented the development of fentanyl-induced conditioned place preference in male and female rats. Accordingly, D-CYSee likely reduces the rewarding properties of fentanyl and therefore reduces its addictive potential. The question that arose from these studies was whether D-CYSee is able to prevent the acquisition of physical dependence to fentanyl and / or reverse established dependence to the powerful synthetic opioid. Another important question was whether D-cysteine ethyl amide (D-CYSea) has greater efficacy than D-CYSee based on its expected greater resistance to plasma carboxylesterases that potentially convert D-CYSee to D-cysteine. The present example demonstrates that co-injections of D-CYSee and D- CYSea prevent and reverse the acquisition of fentanyl in male rats and that D-CYSea appears to me more efficacious than D-CYSee. Accordingly, these D-cysteine analogues may represent a novel class of therapeutics that ameliorate the development of physical dependence to opioids in humans. Materials and Methods Permissions, rats, and surgical procedures

[0296] All studies were done according to the NIH Guide for Care and Use of Laboratory Animals (NIH Publication No.80-23) revised in 1996, and incompliance with ARRIVE (Animal Research: Reporting of In Vivo Experiments) guidelines (http: / / www.nc3rs.org.uk / page. asp? id=1357). All protocols involving the use of rats were approved by the Animal Care and Use Committees of the University of Virginia and Galleon Pharmaceuticals. Adult male Sprague Dawley rats from Harlan Industries (Madison, WI, USA) were given 5 days to recover from transportation before undergoing surgery. Fentanyl citrate powder was obtained from Sigma-Aldrich (St. Louis, MO, USA). D-CYSee and D- CYSea were obtained from ChemImpex (Wood Dale, Illinois, USA) and Olon RIcerca (Concord, Ohio, USA) and divided into 100 mg amounts under N2gas and stored at 4oC. Solutions of D-CYSee and D-CYSea (dissolved in saline and brought to pH 6.8 with 0.1M NaOH at room temperature) were prepared immediately before use. Naloxone HCl (Sigma- Aldrich, St. Louis, MO, USA) was dissolved in normal saline. All arterial and venous catheters were flushed with 0.3 ml of phosphate-buffered saline (0.1 M, pH 7.4) 3-4h before starting the example. All studies were done in a room with relative humidity of 49 ± 2% and temperature of 21.4 ± 0.2 °C. Every example group contained 9 rats.D-CYSee or D-CYSea prevention of the development of physical dependence to fentanyl Behavioral studies

[0297] Three groups of rats were implanted with a catheter of PE-10 connected to PE- 50 (Intramedic; Becton & Dickinson, Franklin Drive, NJ, USA) into the jugular vein under 2- 3% isoflurane anesthesia, to allow injections of test agents. Rats were given 5 days to recover from surgery before use in experiments. Example 1 - 5 co-injections: Rats received co-injections of vehicle (100 µL / 100g body weight) + fentanyl (125 µg / kg, IV), D-cysteine (250 µmol / kg, IV) + fentanyl (125 µg / kg, IV), D-CYSee (250 µmol / kg) + fentanyl (125 µg / kg, IV) or D-CYSea (100 µmol / kg) + fentanyl (125 µg / kg, IV) given 90 sec apart (fentanyl given second in all instances) at 8 am and 8 PM on days 1 and 2 and at 8 AM on day 3. Example 1 - 10 co-injections: Other rats received co-injections of vehicle (100 µL / 100g body weight) + fentanyl (125 µg / kg, IV), D-cysteine (250 µmol / kg, IV) + fentanyl (125 µg / kg, IV), D-CYSee (250 µmol / kg) + fentanyl (125 µg / kg, IV) or D-CYSea (100 µmol / kg) + fentanyl (125 µg / kg, IV) given 90 sec apart at 8 AM and 8 PM on days 1 to 4. The rats were given injection 9 at 8 AM on day 5 and injection 10 at 2 PM to allow for NLX challenges to be given. Ninety min after the 5th or 10th set of co-injections, rats were placed in individual opaque plastic boxes and after 30 min, they were injected with NLX (1.5 mg / kg) and behavioral phenomena were scored for 45 min by 3 scorers. Scored behavioral phenomena were: Jumping behavior - all 4 paws of the ground - jumps; Wet dog shakes - whole body shakes as if to shed water from fur; Rearing behavior - rearing on hind legs - rears; Episodes of fore-paw licking – FPL; Circling behavior - Complete 360orotation; Writhes – full body contortion; Sneezes - episodes of sneezing – abrupt expulsion of air that often disturbed the fine bedding material. Plethysmography ventilatory studies

[0298] One hour before co-injections 5 or 10 were to be given (as described above), rats were placed into individual whole body plethysmography chambers to record ventilatory parameters. The free end of the venous catheter was attached to a swivel assembly in the lid of the chamber and after 60 min of acclimatization, the rats received co-injections 5 or 10 and after 90 min they received an injection of NLX (1.5 mg / kg, IV). Ventilatory parameters including frequency of breathing, tidal volume, minute ventilation and non-eupneic breathingindices were recorded (to be detailed elsewhere) with apnea frequency (> 1.5 sec in duration between breaths) to be reported here. Cardiovascular studies

[0299] Rats were implanted with a jugular vein catheter to inject drugs and a femoral artery catheter to record mean arterial blood pressure (MAP) and heart rate. The rats were given 4 days to recover. The patency of arterial lines was maintained by connecting the line to an infusion pump (Standard Infuse-Withdraw Pump 11 Pico Plus Elite Programmable Syringe Pump; Harvard Apparatus, MA, USA) delivering saline at 20 µL / h). Sixty min before co-injections 5 or 10 were to be administered, the rats were put into individual opaque plastic boxes in order to administer drugs and to actively record pulsatile arterial blood pressure, MAP and heart rate. After 60 min acclimatization, the rats received co-injections 5 or 10 and then after 90 min they received NLX (1.5 mg / kg, IV) and cardiovascular parameters recorded for a further 90 min. Body temperature and body weight studies

[0300] Rats were put into individual opaque plastic boxes 1h before co-injections 5 or 10 of the fentanyl + drug combinations were to be given. A thermistor probe attached to a telethermometer (Yellow Springs Instruments) to record body temperature was inserted 5-6 cm into the rectum and taped to the tail. Body weights and body temperatures were recorded every 15 min during the acclimatization period to establish baseline values and at 15 min intervals during the injection protocols. After 60 min of acclimatization, rats received co- injections 5 or 10 and after 90 min an injection of NLX (1.5 mg / kg, IV) and body weights and body temperatures were recorded for another 90 min. Protocols to determine the abilities of D-CYSee or D-CYSea to reverse fentanyl dependence Behavioral studies

[0301] Rats received 5 injections of fentanyl (125 µg / kg, IV) at 8 AM and 8 PM as described above. The rats then received co-injections 6-10 of fentanyl (125 µg / kg, IV) + vehicle, D-cysteine (250 µmol / kg, IV) + fentanyl (125 µg / kg, IV) or D-CYSee (250 µmol / kg) + fentanyl (125 µg / kg, IV) or D-CYSea (100 µmol / kg) + fentanyl (125 µg / kg, IV) given 90 sec apart. Co-injections 6 were given at 8 PM, co-injections 7 at 8 AM, co-injections 8 at 8 PM, co-injections 9 at 8 AM were given at 8 AM and co-injection 10 was given at 2 PM toallow for the experiments to be performed. Immediately after co-injections 10 we given, the rats were placed in individual opaque plastic boxes and after a 90 min acclimatization period, the rats received an injection of NLX (1.5 mg / kg, IV) and behavioral phenomena (as detailed above) were scored for 45 min by at least 3 independent scorers. Plethysmography ventilatory studies

[0302] One hour before co-injections 10 were to be given, rats were put into individual whole body plethysmography chambers to record ventilatory parameters. The free end of the externalized venous catheter was connected to the swivel assembly and after 60 min acclimatization, the rats received the 10thset of co-injections and after 90 min they received NLX (1.5 mg / kg, IV). Ventilatory parameters and non-eupneic breathing indices were recorded (to be reported elsewhere) with the number of apneas of > 1.5 sec in duration reported here. Cardiovascular studies

[0303] One hour before the 10thset of co-injections were given, groups of rats (n=9 rats per group) were put in individual opaque plastic boxes and the free end of the exteriorized jugular vein catheter was connected to an injection line to give drugs. The free end of the arterial catheter was connected to tubing attached to a computer-coupled pressure transducer (Cabe Lab, Inc.) to continuously record pulsatile arterial blood pressure to derive MAP and heart rate. After 60 min acclimatization, rats received co-injections 5 or 10 and then after 90 min an injection of NLX (1.5 mg / kg, IV) and cardiovascular parameters were recorded continuously for a further 90 min. Body temperature and body weight studies

[0304] Rats were put into individual opaque plastic boxes 1h before the 10thset of co- injections were to be given. A thermistor probe for body temperature recordings was placed as above. Rat body weights and temperatures were recorded every 15 min during acclimatization to establish baseline values and at 15 min intervals throughout the injection protocols. After 60 min, the rats received co-injections 10 and after 90 min they were injected with NLX (1.5 mg / kg, IV) and body weights and body temperatures were recorded for another 90 min.Data Analyses

[0305] All data are presented as mean ± SEM and were analyzed by one-way and two- way ANOVA and Bonferroni corrections for multiple comparisons between means using the error mean square terms from the ANOVA analyses. A P < 0.05 value was the initial level of significance that was modified by the number of between-mean comparisons. The modified t-statistic for 2 groups for instance is t = (mean group 1 - mean group 2) / [s x (1 / n1+ 1 / n2)1 / 2] where s2= mean square within groups term from the ANOVA and n1 and n2 are the number of rats in each group being compared. Statistics were done with GraphPad Prism software (GraphPad Software, Inc., La Jolla, CA). F- and P-statistics related to Figs.10-12 are provided in the relevant figure legends. Results D-CYSee and D-CYSea prevent physical dependence to fentanyl

[0306] The behaviors elicited by the injection of NLX in rats that had received 5 co- injections of fentanyl + vehicle or D-cysteine or D-CYSee or D-CYSea are summarized in panel A of Fig.10. The injection of NLX in fentanyl + vehicle-injected rats produced jumps, wet-dog shakes (WDS) rearing, fore-paw licking (FPL), circling, full-body writhing and episodes of sneezing. These NLX-precipitated responses were similar in rats that had received co-injections of fentanyl + D-cysteine. Except for sneezing, these NLX-precipitated behaviors were reduced in rats that received fentanyl + D-CYSee and much more so in rats that had received fentanyl + D-CYSea. As summarized in Panel B, the injection of NLX produced sustained increases in MAP and heart rate and elicited a large increase in apneas (> 1.5 sec between breaths). The NLX-induced responses were reduced in rats that received co- injections of fentanyl + D-CYSee and much more so in rats that received co-injections of fentanyl + D-CYSea. As summarized in Panels C and D, the NLX-induced decreases in body weight and body temperature were smaller in rats that received fentanyl + D-CYSee and markedly less in rats that received fentanyl + D-CYSea but not in those that received D- cysteine. Behaviors elicited by NLX in rats that received 10 co-injections of fentanyl + vehicle or D-cysteine, D-CYSee or D-CYSea are summarized in panel A of Fig.11. Administration of NLX to rats that received fentanyl + vehicle produced qualitatively similar responses as described above except that they were more intense. NLX-precipitatedwithdrawal phenomena were not diminished in rats that received co-injections of D-cysteine and except for sneezing, the responses were reduced in rats that had received fentanyl + D- CYSee. All NLX-precipitated responses (including sneezing) were markedly reduced in rats that received co-injections of D-CYSea and these responses were less than in those that received D-CYSee. As summarized in Panel B, the NLX-induced increases in MAP, heart rate and incidence of apneic events (> 1.5 sec between breaths) were reduced in rats that received co-injections of D-CYSee (but not D-cysteine) and markedly so in the rats that received D-CYSea. As summarized in Panels C and D, the NLX-precipitated falls in body weight and body temperature were reduced in rats that had received fentanyl + D-CYSee and markedly reduced in rats that had received fentanyl + D-CYSea. D-CYSee and D-CYSea reverse established physical dependence to fentanyl

[0307] The behaviors elicited by injection of NLX in rats that had received 10 injections of fentanyl plus 5 co-injections of vehicle, D-cysteine, D-CYSee or D-CYSea starting with fentanyl injection 6 are summarized in Panel A of Fig.12. These responses were similar in rats that received co-injections of vehicle or D-cysteine. Except for sneezing, the NLX-precipitated behaviors were reduced in rats that received co-injections of D-CYSee. All behaviors were markedly diminished in rats that had received co-injections of D-CYSea. As summarized in Panel B, the NLX-precipitated elevations in MAP, heart rate and incidence of apneas (> 1.5 sec between breaths), were diminished in rats that received D-CYSee and markedly diminished in rats that received D-CYSea. As summarized in Panels C and D, the NLX-induced falls in body weight and body temperature were less in rats that received fentanyl + D-CYSee and markedly diminished in rats that received fentanyl D-CYSea. Changes in variables during the progression of the protocols

[0308] Body weights and arithmetic changes in these weights at key points of the three studies are shown in Table 9. The rats that received the 5 or 10 co-injections of fentanyl + vehicle or fentanyl + D-cysteine lost body weight (arithmetic change from pre-drug values). In contrast, body weights rose in rats that received co-injections of D-CYSee and especially D-CYSea. In rats that received fentanyl +vehicle, the loss in body weight after 10 injections were greater than after 5 injections whereas the increases in body weights in rats that received fentanyl + D-CYSee or fentanyl + D-CYSea after co-injections 10 were greater than after the5 co-injections. The decreases in body weight elicited by NLX were similar in rats that received vehicle or D-cysteine but markedly less in rats that received co-injections of D- CYSee or D-CYSea. With respect to the co-injection 6-10 studies, the rats that received co- injections of fentanyl + vehicle and fentanyl + D-cysteine lost body weight whereas those that received co-injections of ventanyl + D-CYSee or D-CYSea gained weight. The NLX- induced decreases in body weight seen in the rats that received co-injections of D-cysteine were similar to those that received co-injections of vehicle. The falls in body weight were smaller in the rats that received co-injections of D-CYSee and even less in the rats that received co-injections of D-CYSea. Actual and arithmetic changes in body temperatures at key points of for co-injections 1-5 and 1-10 studies and co-injection 6-10 studies are summarized in Table 10. The rats that received 5 or 10 co-injections of fentanyl + vehicle developed a hyperthermia that also occurred in rats that received fentanyl + D-cysteine but not in rats that received fentanyl + D-CYSee or D-CYSea. The injection of NLX elicited a profound hypothermia in rats that received co-injections of vehicle or D-cysteine but not in those that received D-CYSee or D-CYSea. With respect to the co-injection 6-10 studies, the rats that received co-injections of fental + vehicle or D-cysteine showed a pronounced hyperthermia whereas those that received D-CYSee or D-CYSea did not. A NLX- precipitated hypothermia was seen in the rats that received co-injections of vehicle or D- cysteine. This hypothermia was less in rats that received co-injections of D-CYSee and substantially less in rats that received D-CYSea.

[0309] MAP, heart rate, ∆Heart Rate / ∆MAP values before and after injection of NLX and arithmetic changes in these parameters for co-injections 1-5, co-injections 1-10 and co- injections 6-10 studies are presented in Table 11. There were no between-group differences in resting parameters prior to the administration of NLX (the injections of fentanyl elicited transient decreases in MAP and heart rate that had fully resolved by the time the pre-NLX values were recorded). The NLX-precipitated increases in MAP and heart rate in rats that received 10 co-injections of fentanyl + vehicle were substantially greater than in rats given 5 co-injections of fentanyl + vehicle. The NLX-precipitated increases in MAP and heart rate were smaller in rats that received fentanyl + D-CYSee and especially fentanyl + D-CYSea in the studies listed as co-injections 1-5, co-injections 1-10 and co-injections 6-10. Arithmetic changes in ∆Heart Rate / ∆MAP values (see column Delta) were enhanced in rats that receivedco-injections 1-10 and co-injections 6-10 of fentanyl + vehicle. These ratios were markedly diminished in rats that received co-injections of fentanyl + D-CYSee or D-CYSea for all three studies. Table 9 - Body Weights at key points of exampleTable 10 - Body Temperatures at key points of exampleTable 11 - Cardiorespiratory responses elicited by the injection of naloxone HCl

[0310] We show here that the injection of NLX elicited a pronounced withdrawal syndrome consisting of behavioral and cardiorespiratory responses and falls in body temperature and body weight, in male rats that received twice-daily co-injections of fentanyl(125 µg / kg, IV) + vehicle. These withdrawal phenomena signs that are strongly suggestive of the rats having become physically-dependent on fentanyl are in agreement with the patterns are consistent with reports on the patterns of NLX-precipitated withdrawal signs seen in a variety of fentanyl-administration protocols and in other protocols used to induce physical dependence to opioids. Moreover, the NLX-induced hypertension and tachycardia are consistent with previous studies showing that NLX-precipitated withdrawal is associated with hypertension and tachycardia due to activation of the sympathetic nervous system in humans. Our observation that the NLX produced a marked increase in apneas (> 1.5 sec between breaths) is also in agreement with similar results from opioid withdrawal paradigms in humans.

[0311] The first new conclusion of this example was that co-injections of D-CYSee and D-CYSea reduced the development of physical dependence to fentanyl. This conclusion was based on the findings that the withdrawal phenomena elicited by NLX (behaviors, elevations in MAP and heart rate, falls in body weight and temperature) were substantially less than in rats were co-injected with fentanyl and vehicle. The finding that D-cysteine was ineffective certainly suggests that the cell-penetrability of D-CYSee and D-CYSea is an important factor in their efficacy. The enhanced potency of D-CYSea over D-CYSee may results from greater resistance to plasma carboxylesterases that convert thiol esters such as D-CYSee to parent thiols. Accordingly, more D-CYSea than D-CYSee may enter brain neurons involved in acquisition of physical dependence and addiction. However, it should be noted that whereas it was evident that a lower dose of D-CYSea (100 µmol / kg, IV) was more effective than a higher dose of D-CYSee (250 µmol / kg, IV) in preventing the development of dependence (markedly reduced NLX-precipitated withdrawal phenomena) we observed that D-CYSea markedly diminished NLX-precipitated sneezing whereas D-CYSee did not. Sneezing is an important phenomenon in opioid withdrawal syndromes in humans and animals. One possibility therefore is that D-CYSea interacts with signaling pathways driving sneezing whereas D-CYSee does not. The activity of D-CYSea raises the possibility that this and other ethyl amides such as the antioxidant N-acetyl-L-cysteine (L-NAC) ethyl amide may show efficacy in human trials. We have no direct evidence as to the cellular mechanisms by which D-CYSee and D-CYSea blunt the development of physical dependence to fentanyl. These mechanisms may involve (1) their reducing potential (e.g., reduction of Fe3+to Fe2+,free L-cystine to L-cysteine) and reduction of protein bound L-cystine to L-cysteine in plasma membrane ion-channels including K+-, Ca2+- and non-selective cation channels and major ligand-gated ion channel receptors such as N-methyl-D-aspartate (NMDA) glutamatergic receptors and γ-aminobutyric acid (GABA) receptors, (2) upon entering cells, the redox regulation of an array of functional proteins, (3) conversion of D-CYSee and D- CYSea to D-cysteine in cells which enters into enzymatic processes generating H2S sequentially by D-aminoacid oxidase and 3-mercaptopyruvate sulfur-transferase such as the carotid bodies, and (4) conversion to S-nitroso-D-CYSee and S-nitroso-D-CYSea, which exert effects similar to endogenous S-nitrosothiols, S-nitroso-L-cysteine, which controls various intracellular processes including those that control cardiorespiratory functions and those diminishing opioid-induced respiratory depression (OIRD). These actions of D-CYSee and D-CYSea may affect the cell-signaling processes thought important in acquisition of physical dependence to opioids including those involving NMDA receptors, voltage-gated Ca2+-channels, oxidative stress and the nitric oxide-initiated cGMP-mediated signaling cascades. It is likely that the redox effects of D-CYSee and D-CYSea are important since a primary intracellular redox regulator, α-lipoic acid, diminishes the development of morphine dependence in mice and the NLX-induced biochemical alterations in morphine-dependent mice, and moreover, the actions of α-lipoic acid were enhanced by concurrent treatment with N-acetyl-L-cysteine.

[0312] The second new conclusion from this example was that co-injections of D- CYSee and D-CYSea beginning with the 6thand continuing to the 10thinjection of fentanyl reverse established physical dependence to the opioid (again on the basis of diminished NLX- precipitated responses). The NLX-precipitated behaviors (except for sneezes), increases in MAP and heart rate and the numbers of apneas, and falls in body temperature and body weight were fewer / smaller in magnitude in rats that received the co-injections of D-CYSee. Notably, the NLX-precipitated withdrawal phenomena including sneezes were markedly attenuated in rats that received co-injections of D-CYSea. Again, the potential mechanisms by which D-CYSee and D-CYSea reverse acquired physical dependence to fentanyl but some of the mechanisms discussed above including their antioxidant properties may be involved. Although none of the following have proven to be tenable therapeutics, the drugs and bioactive agents that have shown some ability to reverse established physical dependenceinclude, the antioxidants quercetin and melatonin; allosteric modulators of AMPA receptors; the dopamine D2 receptor antagonist, haloperidol; adrenomedullin receptor antagonists; the β2-AR antagonist, butoxamine, histamine receptor agonists, a 5-hydroxytryptamine-reuptake inhibitor, fluoxetine, inhibitors of Ca2+ / calmodulin-dependent protein kinase II, and the nitric oxide synthase inhibitor, L-NG-nitroarginine methyl ester (see Introduction). The clear lack of rationale (driving hypotheses) for the use of these speaks to our minimal understanding about the mechanisms underlying physical dependence to opioids. Our findings with D- CYSee and D-CYSea with respect to physical dependence to fentanyl and our finding that D- CYSee prevents the development of fentanyl-induced conditioned place preference (addictive potential) in male and female rats certainly supports the concept that alterations in thiol chemistry within cells may be an important common feature of fentanyl-induced addiction and physical dependence. The ability of D-CYSee and DCYSea to reverse established physical dependence to fentanyl has potential clinical relevance in that these and other bioactive L,D-thiol esters / amides and related compounds such as L-NAC, Tempol and S- nitrosothiols may be able to reverse acquired physical dependence to fentanyl and other opioids in humans. In particular, if D-CYSee and D-CYSea for example can block opioid- induced dopamine surges in the brain structures (e.g., medial prefrontal cortex, ventral tegmentum and nucleus accumbens) in which the rewarding euphoria-producing dopamine surge happens for drugs of abuse / addiction then then they could be useful in treatment of opioid use disorder (OUD) as stand-alone therapies. Moreover, maternal opioid use is an ever-growing public health issue and babies born to mothers dependent on opioids display often severe withdrawal symptoms that require hospitalization. Current treatment strategies of this neonatal opioid withdrawal syndrome (NOWS) are inadequate and the infants develop numerous behavioral, cognitive social problems as they grow older. Accordingly, novel therapies and better understanding of the mechanisms by which they benefit the immediate and long-term consequences of NOWS are desperately needed. The compelling findings with L-NAC and L-NAC suggest that they may be of therapeutic benefit in preventing the development of NOWS.

[0313] Using the degree of NLX-induced withdrawal phenomena as a measure, we have provided evidence here that D-CYSee and D-CYSea prevent the development of physical dependence to fentanyl and and reverses acquired dependence to the synthetic opioidin adult male Sprague-Dawley rats. The enhanced efficacy of D-CYSea is potentially due to its greater resistance to carboxylesterases that may convert D-CYSee to D-cysteine. Our example with D-CYSee and D-CYSea was in large part due to the pioneering work of Trivedi, Deth and colleagues which greatly contributed to our knowledge about the cell processes by which opioids cause addiction and dependence. In particular, the possibility that opioids may cause psychological addiction and physical dependence by opioid-receptor- mediated blockade of EAA3 (also known as EAAC1) transporter-mediated entry of L- cysteine into neurons prompted our pharmacological studies with the membrane-permeable, L-cysteine ethyl ester (manuscript submitted), as well as D-CYSee and D-CYSea and other D,L-thiol esters. The findings that D-CYSee and D-CYSea both markedly reduced the large majority of NLX-precipitated withdrawal phenomena speaks to the key role that the loss of L-cysteine entry into cells has in establishing physical dependence to fentanyl. The enhanced potency of D-CYSea points to an important strategy in the development of therapeutically effective thiol drugs that not only brings enhanced (non-carrier-mediated) cell-penetrability but also potential protection from plasma carboxylesterases. Determining the thiol-dependent signaling pathways by which D-CYSee and D-CYSea exert their therapeutic actions will add to our understanding of the processes by which these two classes of thiol analogues exert their effects. Example 4

[0314] Clinicians face several serious challenges in treating subjects with opioid use disorder (OUD) and substance use disorders (SUDs) in general. They also must consider related issues in the treatment of pain. To minimize the potential misuse of pain medications, key issues are how to provide opioid (e.g., fentanyl)-induced analgesia without producing euphoric effects, psychological addiction and / or physical dependence, and hyperalgesic states. Regarding treatment of moderate to severe OUD, terminology from the American Psychiatric Association for opioid addiction in 10-20% of people with SUD liability, key problems are (1) how to treat severe opioid withdrawal, as current strategies have many substantial weaknesses, (2) how to manage subjects who have gone through withdrawal and require therapeutics to block dependence and euphoria, with current therapeutics available to the public, namely, methadone, buprenorphine and naltrexone, having strengths but also substantial weaknesses regarding this objective; and (3) how to minimize opioid-inducedeuphoria and dependence in those with moderate to severe OUD who are currently off opioids but need opioid analgesia. In treating SUD related to opioid use, cannabinoids alcohol, psycho-stimulants and benzodiazepines in 10-20% of people prone to SUD, a vital problem is how to provide an effective therapeutic that will attenuate dopamine-mediated euphoria of each brain-reward drug, as a vital adjunct to treating addictive disease states. In regard to making opioid analgesics safer, there are current therapeutics that improve the analgesic efficacy of opioids. Ideally, drugs should (1) promote opioid analgesia while preventing the development of hyperalgesia, (2) prevent the acquisition of psychological addiction and / or physical dependence to the opioid, and (3) prevent and reverse opioid- induced respiratory depression (OIRD).

[0315] The development of morphine-induced dependence and addiction may involve redox-based changes in global DNA methylation and retrotransposon transcription via µ- opioid receptor (µ-OR)-mediated inhibition of excitatory amino acid transporter type 3 (EAA3; also referred to as EAAC1)-facilitated cysteine uptake into central neurons. As presented in Fig.5 of Trivedi et al., the sequence of events proposed from the studies of Trivedi et al. and others are (1) morphine-induced reduction in the active uptake of L- cysteine into neurons by Gprotein-mediated blockade of EAA3 activity, (2) decreases in the levels of L-cysteine and L- glutathione within the brain neurons, (3) fall in S-adenosyl- methionine / S-adenosyl-homocysteine (SAM / SAH ratio, methylation index), (4) reduced methylation of global CpG (regions of DNA in which a cytosine nucleotide is followed by a guanine nucleotide) and decreases in CpG methylation of long interspersed nuclear element - 1 (LINE-1) retrotransposon regulatory regions, (5) stimulation of transcription of previously silenced LINE-1 gene. It occurred to us that co-administration of cell-permeant analogues of L-cysteine such as L-cysteine ethyl ester (L-CYSee) may prevent / diinish the acquisition of physical dependence to morphine and reverse established dependence to morphine. Indeed, we recently reported that the intravenous infusion of L-CYSee prevented the acquisition of morphine dependence in male rats and reversed acquired dependence to morphine. The inability of L-cysteine or L-serine ethyl ester (oxygen atom rather than sulfur atom as in L- CYSee) suggests that the efficacy of L-CYSee is due to its high cell penetrability into brain regions vital to expression of morphine dependence and points to the vital role of thiol biochemical pathways in the efficacy of the L-thiolester. These results are complimented byour findings that L-CYSee, L-cysteine methyl ester, other thiolesters and related compounds such as S-nitrosothiols and the free radical scavenger, Tempol prevent and / or reverse the adverse actions of morphine and fentanyl on breathing, arterial blood-gas chemistry (pH, pCO2, pO2and sO2) and Alveolar-arterial gradient (index of alveolar gas-exchange) in freely- moving rats without affecting opioid-induced analgesia or sedation.

[0316] Since L-CYSee and D-CYSee effectively prevent and reverse OIRD, the main issue we addressed here was whether stereoselectivity is a key feature in the efficacy of L- CYSee with respect to modulating physical dependence to morphine, or more simply, can D- CYSee behave like L-CYSee? Key findings were that D-CYSee prevented / reversed the acquisition of morphine dependence in male rats, based on diminished withdrawal symptoms elicited by the opioid receptor antagonist, naloxone HCl (NLX) whereas D-cysteine was ineffective. In studies designed to define the role of thiol chemistry in the action of D- CYSee, we found that D-serine ethylester (D-SERee) prevented the development of dependence to morphine but was unable to reverse established dependence in these rats. The inability of D-SERee to reverse morphine dependence implicates thiol biochemistry in the mechanisms by which D-CYSee reverses morphine dependence. D-CYSee and analogues may be novel therapeutics ameliorate the development / reversal of physical dependence to opioids. Materials and Methods Permissions, rats, surgical procedures

[0317] All studies were done in accordance with the NIH Guide for Care and Use of Laboratory Animals (NIH Publication No.80-23) revised in 1996, and in strict compliance with ARRIVE (Animal Research: Reporting of In Vivo Experiments) guidelines (http: / / www.nc3rs.org.uk / page. asp? id=1357). All protocols involving rats were approved by the Animal Care and Use Committees of Galleon Pharmaceuticals, Case Western Reserve University, and the University of Virginia. Adult male Sprague Dawley rats were purchased from Harlan Industries (Madison, WI, USA). The rats were given 5 days to recover from transportation before being subject to surgeries. (+)-Morphine sulfate was from Baxter Healthcare (Deerfield, IL, USA). Powder forms of D-CYSee HCl, D-cysteine HCl, and D- serine HCl were purchased from Sigma-Aldrich (St. Louis, MO, USA) and D-SERee HCl was purchased from Neta Scientific (Hainesport, NJ). The powders were divided into 100 mgamounts under N2 gas and stored at 4oC. Solutions of these compounds (dissolved in saline and brought to pH 7.2 with 0.1M NaOH at room temperature) were prepared immediately before use. Naloxone HCl (NLX; Sigma-Aldrich, St. Louis, MO, USA) was dissolved in normal saline. On the day of example, all arterial and venous catheters were flushed with 0.3 ml of phosphate-buffered saline (0.1 M, pH 7.4) 3-4h before commencement of the example. All studies were done in a quiet room with relative humidity of 50 ± 2% and room temperature of 21.3 ± 0.2 °C. Protocols to determine the effects of D-CYSee on physical dependence to morphine Prevention of morphine dependence Behavioral studies

[0318] At 2 am of the day of surgery, groups of rats received a jugular vein catheter (PE-10 connected to PE-50) under 2-3% isoflurane anesthesia. The jugular vein catheter was connected to a primed ALZET osmotic minipump (Model 2002, ALZA Corporation, California, U.S.A.) positioned at the back of the neck to allow for the continuous infusion of vehicle (20 µL / h, IV), D-cysteine, D-CYSee, D-serine or D-SERee (all at 20.8 µmol / kg / h, IV), as described previously. Physical dependence was induced by a slow-release subcutaneous depot of morphine emulsion (150 mg / kg, SC) injected at the left side of the neck as described in detail by Fennessy and colleagues. In brief, morphine base was precipitated from a solution of (+)-morphine sulfate by titrating to pH 9 with 1 mol / L NaOH. After several distilled water washes, pure base was collected in a filter funnel and dried. Morphine slow-release emulsion was prepared by suspending a weighed amount of base in liquid paraffin and Arlacel A. This mixture was emulsified with an equal volume of normal saline as initially described by Collier et al. All wounds were sutured closed and the rats were returned to their home cages. After 35.5h of morphine exposure, the rats were placed in individual opaque plastic boxes and after 30 min of acclimatization, they received an intraperitoneal (IP) injection of NLX (1.5 mg / kg) and behaviors were scored for 45 min by 3 scorers. The scored phenomena were: Jumping behavior - all 4 paws of the ground - jumps; Wet dog shakes -whole body shakes as if to shed water from fur; Rearing behavior - rearing on hind legs - rears; Episodes of fore-paw licking – FPL; Circling behavior - Complete 360orotation; Writhes – full body contortion; Sneezes - episodes of sneezing – abrupt expulsion of air that often disturbed the fine bedding material. Plethysmography ventilatory studies

[0319] Rats were prepared as above except that they received a second catheter into the jugular vein as described by Getsy et al. to give a bolus injection of NLX. After 35h, rats were put in individual whole body plethysmography chambers and the free end of the exteriorized venous catheter was connected to a swivel assembly within the lid of each of the plethysmography chambers and after a 60 min acclimatization period, the rats were given an intravenous injection of NLX (1.5 mg / kg). Ventilatory parameters including frequency of breathing, tidal volume, minute ventilation and non-eupneic breathing indices were recorded (to be reported elsewhere) with the number of apneas (> 1.5 sec between breaths) reported here. Cardiovascular studies

[0320] Rats were prepared as above except that they received a second catheter in the jugular vein to administer NLX and a catheter into a femoral artery to continuously record mean arterial blood pressure (MAP) and heart rate as described previously. After 35h, the rats were placed in placed in individual opaque plastic boxes and the free end of the exteriorized jugular vein catheter was connected to an injection line to deliver NLX. The free end of the arterial line was connected to tubing attached to a computer-coupled pressure transducer (Cabe Lab, Inc.) to record pulsatile arterial blood pressure. After a 60 min acclimatization period, the rats received a bolus injection of NLX (1.5 mg / kg, IV) and MAP and heart rate were recorded continuously for 90 min. Body temperature and body weight studies

[0321] Groups of rats without a second jugular catheter were prepared as described above. After 35h, the rats were placed in individual opaque plastic boxes and a thermistor probe connected to a telethermometer (Yellow Springs Instruments) to record body temperature was inserted 5-6 cm into the rectum and taped to the tail. The body weights of the rats and body temperatures were recorded every 15 min during acclimatization to establish baseline values. After 60 min acclimatization, the rats received an intraperitonealinjection of NLX (1.5 mg / kg). Body temperature and weights were recorded every 15 min for 90 min. Reversal of morphine dependence Behavioral studies

[0322] At 2 pm on the day of surgery, rats received a slow-release subcutaneous depot of morphine emulsion (150 mg / kg, SC) injected at the left side of the neck as described above. After 36h of morphine administration, the rats were anesthetized (2% isoflurane) and received a jugular vein catheter connected to a primed ALZET osmotic minipump positioned at the back of the neck for continuous infusion of vehicle (20 µL / h, IV), D-cysteine, D- CYSee, D-serine or D-SERee (all at 20.8 µmol / kg / h, IV). All wounds were sutured closed and the rats were returned to their warmed home cages. After 11.5h, the rats were placed in individual opaque plastic boxes and after a 30 min period of acclimatization, the rats received an intraperitoneal injection of NLX (1.5 mg / kg) and behavioral phenomena (as detailed above) were scored for 45 min by at least 3 scorers. Plethysmography ventilatory studies

[0323] Rats were prepared as above except that the rats received a second catheter into the jugular vein as described by Getsy et al. to allow for bolus injection of NLX. After 47h, rats were placed in individual whole body plethysmography chambers and the free end of the exteriorized jugular vein catheter that was connected tightly to a swivel on the lid of the plethysmography chamber. After 60 min of acclimatization, the rats received a bolus injection of NLX (1.5 mg / kg, IV). Ventilatory parameters and non-eupneic breathing indices were recorded with the number of apneas (> 1.5 sec between breaths) to be reported here. Cardiovascular studies

[0324] Rats were prepared as above except that rats received a second catheter into the jugular vein to give NLX and a catheter into a femoral artery to record MAP and heart rate. After 47h, the rats were placed in placed in individual opaque plastic boxes and the free end of the exteriorized jugular vein catheter was connected to an injection line to give NLX. The free end of the arterial line was connected to tubing attached to a computer-coupled pressure transducer to record pulsatile arterial blood pressure. After a 60 min acclimatization period,the rats received a bolus injection of NLX (1.5 mg / kg, IV) and MAP and heart rate were recorded continuously for 45 min. Body temperature and body weight studies

[0325] Groups of rats without a second jugular catheter were prepared as described above. After 47h, the rats were placed in individual opaque plastic boxes and a thermistor probe connected to a telethermometer (Yellow Springs Instruments) to record body temperature was inserted 5-6 cm into the rectum and taped to the tail. The body weights of the rats and body temperatures were recorded every 15 min during acclimatization to establish baseline values. After the 60 min acclimatization, the rats received an intraperitoneal injection of NLX (1.5 mg / kg). Body temperature and weights were recorded every 15 min for 90 min. Data Analyses

[0326] All data are shown as mean ± SEM and were evaluated by one-way and two- way ANOVA followed by Bonferroni corrections for multiple comparisons between means using the error mean square terms from each ANOVA analysis as detailed previously. A P < 0.05 value denoted the initial level of statistical significance that was modified according to the number of comparisons between means. The modified t-statistic is t = (mean group 1 - mean group 2) / [s x (1 / n1 + 1 / n2)1 / 2] where s2= mean square within groups term from the ANOVA (the square root of this value is used in the modified t-statistic formula) and n1and n2 are the number of rats in each group under comparison. Based on elementary (Bonferroni's) inequality), a conservative critical value for modified t-statistics obtained from tables of t-distribution using a significance level of P / m, where m is the number of comparisons between groups to be performed. The degrees of freedom are those for the mean square for within group variation from the ANOVA table. In most situations, the critical Bonferroni value cannot be found in conventional tables of the t- distribution but can be approximated from tables of the normal curve by t* = z + (z + z3) / 4n, with n being the degrees of freedom and z being the critical normal curve value for P / m.91-93Wallenstein et al. demonstrated that the Bonferroni procedure is preferable for general use since it is easy to apply, has the widest range of applications, and because it provides critical values that are lower than those of other procedures when the investigator can limit the number ofcomparisons (and will be slightly larger than those of other procedures if many comparisons are made. Statistical analyses were performed with the aid of GraphPad Prism software (GraphPad Software, Inc., La Jolla, CA). Results D-CYSee prevention of physical dependence to morphine – 36h studies

[0327] Behavioral phenomena produced by injection of NLX (1.5 mg / kg, IP) in rats receiving morphine (150 mg / kg, SC) plus a continuous infusion of vehicle (saline, 20 μL / h, IV) or L-cysteine (20.8 μmol / kg / h, IV) or L-CYSee (20.8 μmol / kg / h, IV) for 36h are shown in Fig.13. The injection of NLX to rats receiving vehicle-infusion produced jumping behavior (jumps), wet-dog shakes (WDS) rearing behavior (rears) fore-paw licking (FPL), circling behavior (360ocircles), full-body writhes and sneezes. These NLX-precipitated withdrawal phenomena were similar in rats that were receiving L-cysteine infusion . In contrast, the withdrawal phenomena (except sneezing) were substantially reduced in rats that were receiving the continuous infusion of L-CYSee. The NLX-precipitated withdrawal signs were reduced in rats receiving an infusion of L-SERee (to a much lesser degree than by D- CYSee) but not in rats receiving an infusion of L-serine (see Table 12). Table 12 - Naloxone-precipitated withdrawal signs in rats treated for 36 hours with morphine and continuous infusion of vehicle or D-serine or D-serine ethyl ester

[0328] Expression of apneas (> 1.5 sec between breaths) and elevations in MAP and heart rate produced by injection of NLX (1.5 mg / kg) in rats receiving morphine (150 mg / kg,SC) and continuous infusion of vehicle or D-cysteine or D-CYSee are presented in Fig.13. Injection of NLX in rats that receiving vehicle produced number of apneic events (breath- holds) and elevations in MAP and heart rate (HR). These NLX-precipitated events were numerically similar in rats receiving L-cysteine infusion whereas withdrawal phenomena were substantially diminished in rats receiving L-CYSee. MAP and heart rate values before and after injection of NLX in morphine-treated rats receiving infusions of vehicle, L-cysteine or L-CYSee are shown in Table 14. Resting MAP and heart rate values before injection of NLX were similar in the 3 groups of rats. The injection of NLX produced elevations in MAP and heart rate as described above. In addition, NLX-initiated elevations in MAP and heart rate were reduced in rats receiving an infusion of L-SERee (to a much lesser degree than by D-CYSee) but not in rats receiving L-serine (see Table 14). Table 13 - Changes in mean arterial blood pressure and heart rate elicited by the injection of naloxone HCl in rats treated with morphine with infusions of vehicle, D-cysteine or D-CYSee or for 36 hours

[0329] The decreases in body temperature and body weights elicited by NLX (1.5 mg / kg) in rats receiving morphine (150 mg / kg, SC) and infusion of vehicle or L-cysteine or L-CYSee are presented in the left-hand panels of Fig.15. NLX produced substantialdecreases in body temperature and body weights that were similar in rats receiving infusion of vehicle or L-cysteine. These responses were markedly smaller in rats receiving an infusion of L-CYSee. The body temperatures and body weights before and after injection of NLX in morphine-treated rats that were receiving infusions of vehicle, L-cysteine or L- CYSee are shown in Table 14. Resting body temperatures and body weights before injection of NLX were similar in the 3 groups. After 36h of morphine treatment, body temperatures were elevated by about a 0.5oC in rats receiving infusions of vehicle or L-cysteine whereas body temperature was not raised in rats receiving L-CYSee. Injection of NLX elicited substantial falls in body temperature and body weight in rats receiving vehicle- or L-cysteine and much smaller responses in rats receiving L-CYSee infusion. NLX decreased body temperatures and body weights as described above. In addition, the NLX-initiated decreases in body temperature and body weights were less in rats receiving infusion of L-SERee (to a lesser degree than by L-CYSee) but not in rats receiving L-serine (see Table 15). Body temperatures and body weights before and after injection of NLX in morphine-treated rats receiving continuous infusions of vehicle, L-serine or L-SERee are shown in Table 15. Resting body temperatures and body weights before injection of NLX were similar in each group. After 36h of morphine treatment, body temperatures were elevated by about a 0.5oC in rats receiving infusions of vehicle, L-serine or L-SERee. The NLX-induced decreases in body temperatures and body weights were smaller in the rats receiving L-SERee infusion as described above.Table 14 - Changes in body temperature and body weights elicited by the injection of naloxone HCl in rats treated with morphine with infusions of vehicle, D-cysteine or D-CYSee for 36 hoursTable 15 - Changes in body temperature and body weights elicited by the injection of naloxone HCl in rats treated with morphine with infusions of vehicle, D-serine or D-SERee for 36 hoursL-CYSee reversal of physical dependence to morphine – 48h studies

[0330] The behavioral l phenomena elicited by injection of NLX (1.5 mg / kg, IP) in rats receiving morphine (150 mg / kg, SC) for 48h plus infusion of vehicle (saline, 20 μL / h, IV) or L-cysteine (20.8 μmol / kg / h, IV) or L-CYSee (20.8 μmol / kg / h, IV) beginning at 36h of morphine administration are shown in Fig.15. The injection of NLX to rats receiving infusion of vehicle elicited substantial numbers of withdrawal behaviors. These withdrawal phenomena were similar in rats receiving infusion of L-cysteine whereas they were (except for sneezing) markedly diminished in rats receiving L-CYSee for 12h. The NLX-precipitated withdrawal signs in rats receiving infusions of L-serine or L-SERee were similar to those receiving vehicle (see Table 13).

[0331] The incidence in apneas and changes in MAP and heart rate produced by the injection of NLX (1.5 mg / kg, IP) in rats receiving morphine (150 mg / kg, SC) for 48h and continuous infusion of vehicle or L-cysteine or L-CYSee beginning at 36h of morphine exposure are shown in Fig.17. NLX elicited substantial increases in apneas and elevations in MAP and in heart rate (HR) in rats receiving vehicle infusion. These responses were similar in rats receiving infusion of L-cysteine whereas they were markedly diminished in rats receiving infusion of L-CYSee. Actual values for MAP and heart rate before and after injection of NLX in morphine-treated rats receiving infusions of vehicle, L-cysteine or L- CYSee are shown in Table 16. Resting MAP and heart rate values before injection of NLX were similar in the 3 groups. Injection of NLX elicited the increases in MAP and heart rate as described above. The NLX-precipitated increases in apneas, MAP and heart rate in rats receiving infusion of L-serine or L-SERee were similar to those receiving vehicle (see Table 17). Table 16 - changes in mean arterial blood pressure and heart rate elicited by injection of naloxone HCl in rats treated with morphine for 48h with co-administration of vehicle, D- cysteine or D-CYSee for 12 hours starting at 36h of morphine administrationIV) or D-cysteine (20.8 µmol / kg / h, IV) or D-cysteine ethyl ester (D-CYSee, 20.8 µmol / kg / h, IV) There were 9 rats in each group. The data are presented as mean ± SEM. *P < 0.05, significant response.†P < 0.05, D-cysteine or D-CYSee versus Vehicle. Table 17 - Naloxone-precipitated withdrawal signs in rats treated with morphine for 48 hours and which received infusions of vehicle, D-serine or D-serine ethyl ester for 12 hours starting at 36 hours of morphine administrationD-SERee, D-serine ethyl ester. The data are presented as mean ± SEM. There were no between-group differences in starting body weights (P > 0.05, for all comparisons) *P < 0.05, D-serine or D-SERee versus Vehicle.

[0332] The changes in body temperatures and body weights produced by NLX (1.5 mg / kg) in rats receiving morphine (150 mg / kg, SC) and continuous infusion of vehicle or L- cysteine or L-CYSee from 36h of morphine exposure are shown in the right-hand panels of Fig.15. Injection of NLX elicited substantial decreases in body temperature and body weights of similar size in rats receiving vehicle or L-cysteine. The withdrawal responses were markedly smaller in rats receiving infusion of L-CYSee. Body temperature and body weight before and after injection of NLX in morphine-treated rats receiving infusions of vehicle, L-cysteine or L-CYSee are shown in Table 18. Resting body temperature and body weight values before injection of NLX were similar in the 3 groups. After 48h of morphine treatment, body temperatures were elevated by about 0.5oC in rats receiving the infusions of vehicle or L-cysteine. Body weights were similar in all 3 groups. Body temperature was not elevated in the rats receiving infusion of L-CYSee but was in those receiving L-cysteine. NLX elicited pronounced falls in body temperature and body weights in vehicle- or L- cysteine-infusion groups but markedly smaller responses in rats receiving L-CYSee. Actual body temperature and body weight before and after injection of NLX in morphine-treated rats receiving infusions of vehicle, L-serine or L-SERee are shown in Table 19. Body temperatures and body weights before injection of NLX were similar in each group. After 48h of morphine exposure, body temperatures were elevated by about a 0.5oC in rats receiving infusions of vehicle or L-serine or L-SERee. NLX-precipitated decreases in body temperatures and body weights were similar in all 3 groups. Table 18 - changes in body temperature and body weights elicited by the injection of naloxone HCl in rats treated with morphine for 48 hours with co-infusion of vehicle, D- cysteine or D-CYSee for 12 hours starting at 36 hours of morphine administrationTable 19 - changes in body temperature and body weights elicited by the injection of naloxone HCl in rats treated with morphine with infusions of vehicle, D-serine or D-SERee for 36 hoursThere were 9 rats in each group. The data are presented as mean ± SEM. *P < 0.05, significant response.†P < 0.05, D-serine or D-SERee versus Vehicle.

[0333] This example found that co-infusion of D-CYSee substantially diminished the expression of numerous withdrawal phenomena (behavioral responses, cardiorespiratory changes, body weight loss and hypothermia) initiated by the injection of the opioid receptor antagonist, NLX, in male Sprague-Dawley rats exposed to a slow-release morphine emulsion for 36h. The behavioral phenomena, which indicate that the rats were physically-dependent on morphine, included wet-dog shakes, jumping, fore-paw licking, rearing, writhing, circling and sneezing (rapid and forceful expulsions of air). These phenomena as well as the falls in body temperature and decreases in body weight and were consistent with previous findings in rats with this slow-release morphine method of inducing physical dependence and with a wide variety of other administration protocols used to induce morphine dependence. The substantial increases in MAP and heart rate elicited NLX are novel findings in our method of eliciting morphine dependence but are consistent with the results of studies showing that NLX-precipitated withdrawal causes hypertension and tachycardia in experimental animals and humans that is due to globalized activation of the sympathetic nervous system. Finally, our finding that NLX elicited a substantial increase in apneic events is new to our morphine- dependence model but consistent with such findings in rats and humans. The failure of L- cysteine to modify the NLX-precipitated withdrawal phenomena suggests that the more rapid entry of L-CYSee into cells underlies the efficacy of this highly cell-penetrant L-thiol ester and in particular entry into neurons involved in acquisition of physical dependence and addiction. At present, we do not know how L-CYSee modulates the intracellular processes within the brain by which morphine induces physical dependence. The mechanisms by which L-thiolesters exert their biological effects are likely to be multi-factorial and possibly include (1) direct binding of L-CYSee to plasma membrane / intracellular proteins such as ion- channels, receptors and enzymes that alters the activities of the proteins by mechanisms not associated with changes in redox status of the proteins (yet to be substantiated) (2) formation of thiol adducts such as D-glucose:D-cysteine and mixed disulfides in blood, (3) modulation of redox status (e.g., reduction of disulfides to the monothiol such as endogenous L-cystine to L-cysteine) and the activity of plasma membrane proteins such as Kv1.2 K+-channels and after entry into cells, redox modulation of functional intracellular proteins, (4) the formation of S- thiolated proteins such as S-cysteinylated, S-cysteinylglycinylated and S-glutathionylatedproteins in plasma membranes and cells, (5) potential conversion of D-CYSee to D-cysteine by membrane associated esterases, which then enters into metabolic pathways including those that generate hydrogen sulfide by sequential actions of D-aminoacid oxidase and 3- mercaptopyruvate sulfur-transferase in central and peripheral tissues including the carotid bodies, (6) conversion of L-thiolesters to cysteine sulfenics, sulfonics and sulfonics via cysteine dioxygenase, and (7) formation of the S-nitrosothiols, S-nitroso-D-cysteine ethyl ester and S-nitroso-D-cysteine, which may behave like the endogenous S-nitrosothiol, S- nitroso-L-cysteine, which has many roles in intracellular signaling pathways including those that regulate cardiovascular and ventilatory functions and those that blunt OIRD. These and other potential mechanisms may interact with intracellular signaling pathways involved in the acquisition of physical dependence to opioids and expression of NLX-precipitated withdrawal phenomena including pathways involving N-methyl D-aspartate (NMDA) glutamatergic receptors, muscarinic receptors, corticotropin releasing factor (CRF) receptor CRF1, tachykinin receptors, voltage-gated Ca2+-channels, adenylyl cyclase super-activation and phosphorylation of opioid receptor, oxidative stress and the nitric oxide-cGMP signaling pathway. Because D-CYSee markedly attenuated all the NLX-precipitated behavioral (except for sneezes), physical (hypothermia and body weight loss), and cardiorespiratory (hypertension, tachycardia and apneic episodes) phenomena, it is possible that D-CYSee modulates the fundamental intracellular processes(es) that are essential to the development of physical dependence to morphine in these male Sprague-Dawley rats.

[0334] The second group of novel and important findings was that the introduction of D-CYSee infusion 36h into the morphine administration period reversed pre-existing physical dependence to the opioid as assessed at 48h. More specifically, NLX-precipitated behavioral phenomena (except for sneezing), hypertension, tachycardia, apneic events, hypothermia and body weight loss were substantially diminished in rats that had received L-CYSee for 12h. The lack of effect of L-cysteine suggests that intracellular delivery of L-CYSee and most likely its sulfur atom (and thiol chemistry and associated signaling mechanisms) are essential for the L-thiol ester to reverse established morphine physical dependence. The therapeutics and bioactive compounds that reverse physical dependence include, L-histidine and histamine receptor sub-type agonists, melatonin, the antioxidant, quercetin, the serotonin-reuptake inhibitor, fluoxetine, a nitric oxide synthase inhibitor, L-NG-nitroarginine methyl ester, inhibitors of Ca2+ / calmodulin-dependent protein kinase II, the β2-AR antagonist,butoxamine, adrenomedullin receptor antagonists, the dopamine D2 receptor antagonist, haloperidol, insulin and selective KATPchannel modulators and positive allosteric modulators / regulators of α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) glutamatergic receptors. The finding that D-CYSee reverses established physical dependence to morphine is of immediate clinical relevance and provides the rationale for studies on D- CYSee and other active L,D-thiol esters and related agents to establish their ability to reverse physical dependence to morphine, heroin and fentanyl.

[0335] A fundamental question arising from this example relates to the potential use of D-thiolesters as therapeutics for clinical problems associated with long-term use of opioid analgesics in humans. With respect to the therapeutic use of D-CYSee, the issues are (1) if D-CYSee substantially attenuates self-administration of opioids in OUD patients, then adding it to prescribed opioids may result in lower addiction / abuse potential; (2) if D-CYSee attenuates the acquisition of physical dependence to opioids, then its addition to prescribed opioids would minimize / eliminate physical dependence in subjects taking opioids day in and day out for weeks and months); (3) if D-CYSee diminishes the development of tachyphylaxis to opioid analgesia or the switch to hyperalgesia caused by long-term opioid use, then adding D-CYSee to prescription opioids will maintain their analgesic efficacy over long periods of time, eliminating the development of tolerance, need for dose escalation, and the multiplicity of issues caused by hyperalgesia; (4) if D-CYSee has several of the beneficial actions seen in rodents, then adding it to prescribed opioid analgesics would multiply the beneficial effects of the opioids; (5) if D-CYSee prevents the development of physical dependence, and if it can be given to a subject with physical dependence and can block opioid withdrawal, it could be used as an inpatient and outpatient therapeutic to manage opioid withdrawal in those prescribed long-term opioid prescriptions (iatrogenically physically-dependent) or those addicted and physically dependent; (6) if D-CYSee attenuates euphoria and the development of psychological addiction to opioids, then it would be a much needed therapeutic for MAT (medication-assisted treatment) and a potentially game-changing therapeutic for harm reduction interventions in people with OUD who are not willing to engage in the psychosocial benefits of counseling and treatment; (7) since some patients with a history of OUD who are currently sober need opioids for the management of acute or chronic pain, then D-CYSee, if it attenuates euphoria and physical dependence, could be added to opioid analgesics when given to subjects with history of OUD, thereby eliminating risks of opioidanalgesics precipitating euphoria, drug cravings and associated highly-increased risk of relapse; (8) if D-CYSee attenuates / blocks euphoria from chemically mediated dopamine surges within ventral tegmentum, nucleus accumbens, or medial prefrontal cortex, where brain rewarding euphoria-producing dopamine surge happens from all drugs of abuse / addiction then it will be a much needed drug in the treatment of OUD and other SUDs; (9) If D-CYSee attenuates euphoria from chemically-mediated dopamine surges, it could be combined with or added to all controlled prescription drugs resulting in an abuse-resistant or non-abusable form of prescribed opioids, benzodiazepines, and psychostimulants, for example.

[0336] In an effort to provide evidence that the sulfur moiety of D-CYSee and accompanying thiol biochemistry is vital to the efficacy of the D-thiolester, we examined whether D-serine and D-SERee (oxygen for sulfur) would prevent or reverse morphine dependence. Although D-SERee has been used sparingly in vivo, systemic administration of D-SERee was shown to improve motor function in ataxic mice and was administered in vivo to determine whether it would reverse morphine-induced OIRD, which it did not. In the present example, we found that the infusion of L-SERee was able to prevent the development of dependence to morphine whereas the infusion of D-serine did not. This finding with D- serine is not consistent with evidence that co-injections of D-serine (600 mg / kg, IP) antagonized morphine (10 mg / kg, SC)-induced conditioned place preference (CPP, addiction behavioral test) in 7-9 week old rats. As such, it is possible that our infusion paradigm (20.8 µmol / kg / h = 3.224 mg / kg / h, IV) with the highly cell-permeable L-SERee delivers enough L- serine into key brain regions involved in the development of dependence to morphine whereas this infusion rate of D-serine does not. The possibility that morphine induces its affects by altering D-serine bioavailability / bioefficacy is supported by several studies. For example, morphine induces a slight but significant increase of D-serine levels in cortex, striatum and hippocampus but reduces extracellular D-serine levels in the nucleus accumbens, a vital brain structure in the development of opioid dependence addiction via blockade of Ca2+-dependent exocytosis of vesicular stores of D-serine. Indeed, D-serine is an endogenous neurotransmitter / neuromodulator that acts as a NMDA receptor co-agonist. Morphine alters NMDAR-mediated synaptic plasticity, reduces NMDA receptor-mediated excitatory post-synaptic currents and excitability of GABAergic neurons, and internalizesAMPA receptors. Several of these morphine-induced effects are reversed by D-serine. Although our present example did not find L-serine to be effective, the efficacy of L-SERee in preventing acquisition of dependence to morphine adds to thinking about potential therapeutic uses of D-serine and D-SERee. The inability of D-SERee to reverse previously acquired dependence to morphine suggests that the efficacy of D-CYSee to reverse this phenomenon involves thiol-dependent mechanisms, as discussed above. With the caveats mentioned above, our finding with D-SERee is consistent with evidence that whereas co- injections of D-serine prevent morphine-induced CPP it does not reverse established CPP. However, evidence that D-serine is essential for opioid-withdrawal long-term potentiation (opioid-induced hyperalgesia by amplification of synaptic strength at spinal C-fiber synapses after withdrawal from systemic opioids such as remifentanil) suggests a complicated role for D-serine in these dependence / withdrawal processes.

[0337] In conclusion, were report here that intravenous infusion of the membrane- permeable D-thiol ester, D-CYSee, prevents development of physical dependence to morphine and overcomes previously acquired dependence in in male Sprague-Dawley rats. This and our companion example with L-CYSee example was spurred by the studies of Trivedi and co-workers who provided evidence that morphine elicits psychological addiction / physical dependence by diminishing the uptake of L-cysteine into neurons / astrocytes by blocking the activity of the EAA3 / EAAC1 transporter. The findings that L-CYSee markedly diminished the large majority of the NLX-precipitated withdrawal phenomena speaks to the key role that the loss of L-cysteine entry into cells plays in establishing physical dependence to morphine whereas the efficacy of D-CYSee suggests that the processes by which L-CYSee exerts its effects on these pathways are not stereoselective. The lone withdrawal phenomenon that was not ameliorated by D-CYSee or L-CYSee was sneezing, a phenomenon of the opioid withdrawal in humans and animals. The finding that D-SERee markedly reduced the incidence of sneezing precipitated by the injection of NLX in rats that received morphine for 36h points to the possible involvement of NMDA receptors in the neural mechanisms responsible for sneezing during opioid withdrawal. The present findings add to our knowledge about the pharmacological actions of L,D-thiolesters, such as L-CYSee, L-cysteine methyl ester, L-GSHee, D-CYSee, D-cystine di(m)ethylester and a free radical-superoxide anion scavenger, Tempol, on the pharmacological actions of opioids. Our findings provide rationale for considering L-CYSee and other membrane-permeable L-thiolesters such as such as L-cysteine methyl ester, L-glutathione ethyl ester, γ-L- glutamylcysteine ethyl ester and L-cystine diethylester (see Supplemental Table 2 of Getsy et al62) as potential drugs to prevent and reverse physical dependence to opioids. N-acetyl-L- cysteine (L-NAC) reduces opioid withdrawal phenomena in neonatal rats perhaps via decreased brain oxidative stress in the brain. This raises the intriguing possibility that the readily cell-penetrant L-thiolester, L-NAC ethyl ester, which is a more effective anti-oxidant than L-NAC, will be more efficacious in neonates and perhaps be an effective drug in adults suffering from OUD. In addition, ethyl amide derivatives, such as L-cysteine ethyl amide and L-NAC ethylamide may have even greater biological efficacy than the ethyl esters based on their expected greater resistance to desterfication by plasma / tissue carboxylesterases. Example 5

[0338] The development of hyperalgesia (enhanced physical pain and / or psychological distress) during chronic fentanyl use plays a vital role in the continued and accelerated use of the opioid. Numerous biochemical changes in the brain including changes in redox status are thought to contribute to the development of hyperalgesia to opioids. The antioxidant N- acetyl-L-cysteine (L-NAC), has been used in numerous clinical scenarios and so this example addressed the possibility that L-NAC, which has relatively poor bioavailability, and its readily cell-penetrant analogue, L-NAC methyl ester (L-NACme) would affect the progressive loss of analgesia and appearance of hyperalgesia in adult male Sprague-Dawley rats. We determined the changes in tail-flick latencies (in response to a focused beam of light to the tail) following twice daily (8 AM and 8 PM) co-injections of fentanyl (125 µg / kg, IV) + vehicle or L-NAC (500 µmol / kg, IV) or L-NACme (500 µmol / kg, IV) in adult male Sprague-Dawley rats (10 co-injections over 5 days). The first co-injections of fentanyl (75 µg / kg, IV) + vehicle elicited a profound long-lasting analgesia (increases in tail-flick latencies expressed as increases in maximum possible effect, MPE,%) whereas the duration of analgesia dropped markedly with co-injections 5 and 10 with a robust hyperalgesia (decreases in tail-flick latencies expressed as decreases in MPE, %) appearing with co- injections 5 and more dramatically expressed following co-injections 6-10. The co-injections of L-NAC partially prevented the loss of analgesia and occurrence of the hyperalgesia whereas co-injections of L-NACme (which initially enhanced the duration of the analgesic effects of fentanyl) markedly reduced the decay in analgesia and prevented the appearance ofhyperalgesia. As such, this example provides compelling evidence that co-injections of L- NAC and L-NACme are able to beneficially affect the alterations in pain processing that occurs with sub-chronic administration of fentanyl. The higher efficacy of L-NACme is likely due to its greater cell penetrability in brain regions and interaction with intracellular signaling cascades including redox-dependent processes that mediate fentanyl-induced analgesia and hyperalgesia. Materials And Methods Permissions, rats, and surgical procedures

[0339] All studies were done in accordance with the NIH Guide for Care and Use of Laboratory Animals (NIH Publication No.80-23) revised in 1996, and incompliance with ARRIVE (Animal Research: Reporting of In Vivo Experiments) guidelines (http: / / www.nc3rs.org.uk / page.asp?id=1357). All protocols involving the use of rats were approved by the Animal Care and Use Committees of the University of Virginia and Galleon Pharmaceuticals. Adult male Sprague Dawley rats purchased from Harlan Industries (Madison, WI, USA) were given four days to recover from transportation before being subject to surgeries. Fentanyl citrate, L-NAC and L-NACme powders were obtained from Sigma-Aldrich (St. Louis, MO, USA). The L-NAC and L-NACme powders were divided into 100 mg amounts under N2 gas and stored at 4oC. Solutions of L-NAC and L-NACme (dissolved in saline and brought to pH 7.2 with 0.1M NaOH at room temperature) were prepared immediately before use. Naloxone HCl (Sigma-Aldrich, St. Louis, MO, USA) was dissolved in normal saline. All arterial and venous catheters were flushed with 0.3 ml of phosphate-buffered saline (0.1 M, pH 7.4) 3-4h before commencement of the example. All studies were done in a room with relative humidity of 48 ± 1% and temperature of 21.3 ± 0.2°C. Results

[0340] Figs.18(A-E) illustrate the changes in tail-flick latencies expressed as maximal possible effect (MPE, %) elicited by co-injections 1-10 of fentanyl (125 µg / kg, IV) + vehicle or fentanyl (125 µg / kg, IV) + naloxone HCl (NLX, 5 mg / kg, IV). The data are presented as mean ± SEM. There were 9 rats in each group. *P < 0.05, significant response.†P < 0.05, NLX versus vehicle.

[0341] Figs.19(A-E) illustrate the changes in tail-flick latencies expressed as maximal possible effect (MPE, %) elicited by co-injections 6-10 of fentanyl (125 µg / kg, IV) + vehicle or fentanyl (125 µg / kg, IV) + naloxone HCl (NLX, 5 mg / kg, IV). The data are presented as mean ± SEM. There were 9 rats in each group. *P < 0.05, significant response.†P < 0.05, NLX versus vehicle.

[0342] Figs.20(A-C) illustrate total (cumulative) changes in tail-flick latencies for analgesia or hyperalgesia expressed as the sum of the maximal possible effect (MPE, %) elicited by co-injections 1-10 of fentanyl (125 µg / kg, IV) + vehicle or fentanyl (125 µg / kg, IV) + naloxone HCl (NLX, 5 mg / kg, IV). C: Changes in Pre-values for tail-flick latencies expressed as arithmetic difference from Pre-injection 1. The data are presented as mean ± SEM. There were 9 rats in each group. *P < 0.05, significant response.†P < 0.05, NLX versus vehicle.

[0343] Figs.21(A-E) illustrate the changes in tail-flick latencies expressed as maximal possible effect (MPE, %) elicited by co-injections 1-5 of fentanyl (125 µg / kg, IV) + vehicle or fentanyl (125 µg / kg, IV) + N-acetyl-L-cysteine (L-NAC, 500 µmol / kg, IV) or fentanyl (125 µg / kg, IV) + L-NAC methyl ester (L-NACme, 500 µmol / kg, IV). The data are presented as mean ± SEM. There were 9 rats in each group. *P < 0.05, significant response.†P < 0.05, L-NAC or L-NACme versus vehicle. P < 0.05, L-NACme versus L-NAC.

[0344] Figs.22(A-E) illustrate the changes in tail-flick latencies expressed as maximal possible effect (MPE, %) elicited by co-injections 6-10 of fentanyl (125 µg / kg, IV) + vehicle or fentanyl (125 µg / kg, IV) + N-acetyl-L-cysteine (L-NAC, 500 µmol / kg, IV) or fentanyl (125 µg / kg, IV) + L-NAC methyl ester (L-NACme, 500 µmol / kg, IV). The data are presented as mean ± SEM. There were 9 rats in each group. *P < 0.05, significant response.†P < 0.05, L-NAC or L-NACme versus vehicle. P < 0.05, L-NACme versus L-NAC.

[0345] Figs.23(A-B) illustrate the total (cumulative) changes in tail-flick latencies for analgesia (Panel A) and for hyperalgesia (Panel B) expressed as the sum of the maximal possible effect (MPE, %) values elicited by co-injections 1-10 of fentanyl (125 µg / kg, IV) + vehicle or fentanyl (125 µg / kg, IV) + N-acetyl-L-cysteine (L-NAC, 500 µmol / kg, IV) or fentanyl (125 µg / kg, IV) + L-NAC methyl ester (L-NACme, 500 µmol / kg, IV). The dataare s mean ± SEM. There were 9 rats in each group. *P < 0.05, significant response.†P < 0.05, L-NAC or L-NACme versus vehicle. P < 0.05, L-NACme versus L-NAC.

[0346] Figs.24(A-B) illustrate (A) Baseline tail-flick latencies prior to each set of co- injections of fentanyl (125 µg / kg, IV) + vehicle or fentanyl (125 µg / kg, IV) + N-acetyl-L- cysteine (L-NAC, 500 µmol / kg, IV) or fentanyl (125 µg / kg, IV) + L-NAC methyl ester (L- NACme, 500 µmol / kg, IV). B: Changes in Pre-values for tail-flick latencies expressed as arithmetic difference from Pre-injection 1. The data are presented as mean ± SEM. There were 9 rats in each group. *P < 0.05, significant response.†P < 0.05, L-NAC or L-NACme versus vehicle. P < 0.05, L-NACme versus L-NAC.

[0347] Fig.25 are chemical structures of L-NAC and analogues.

[0348] Figs.26(A-H) illustrate the changes in tail-flick latencies expressed as maximal possible effect (MPE, %) elicited by co-injections 1-10 of vehicle + vehicle or vehicle + N- acetyl-L-cysteine (L-NAC, 500 µmol / kg, IV) or vehicle + L-NAC methyl ester (L-NACme, 500 µmol / kg, IV). The data are presented as mean ± SEM. There were 9 rats in each group. There were no differences from Pre values for ant administration protocol (P > 0.05, for all responses).

[0349] Figs.27(A-B) illustrate baseline tail-flick latencies prior to each set of co- injections of fentanyl (125 µg / kg, IV) + vehicle or fentanyl (125 µg / kg, IV) + naloxone HCl (NLX, 5 mg / kg, IV). Panel B: Changes in Pre-values for tail-flick latencies expressed as arithmetic difference from Pre-injection 1. The data are presented as mean ± SEM. There were 9 rats in each group. There were no changes in baseline values from the initial set of co-injections (P > 0.05, for all responses).

[0350] The present example demonstrates that the robust analgesic responses elicited by 10 injections of a relatively high dose of fentanyl (twice daily doses of 125 µg / kg, IV) were absent in rats that were pretreated with NLX (5 mg / kg, IV) given 15 min before each injection of fentanyl. The dose of NLX was higher than that usually employed in such studies since a more standard dose of 1.5 mg / kg elicited only a partial reduction in the analgesic responses (data not shown). We are assuming that the ability of the high dose fentanyl to elicit analgesia is due to the activation of opioid (preferentially µ-) receptors the high dose NLX to prevent fentanyl analgesia is due to blockade of opioid receptors.Example 6 Behavioral and neural effects of L-NAC and derivatives on prevention and reduction of synthetic opioid use disorder

[0351] Synthetic opioids, such as fentanyl, are clinically administered to alleviate severe chronic and acute pain, and induce anesthesia. However, their clinical efficiency is complicated by their inclination to induce analgesic tolerance, pleasure, and addictive states. In 2021, opioid use disorder (OUD) and overdose due to opioid-induced respiratory depression (OIRD) resulted in an estimated 109,000 deaths in the US, incurring a medical cost of more than 1-1.5 trillion dollars. OUD is a chronically relapsing disorder that is characterized by compulsive drug-seeking behavior that can be enhanced by drug abstinence and is maintained by encounters with drug-associated cues which elicit strong cravings and promote drug-seeking behaviors. Medication-assisted treatments for OUD (e.g., methadone, buprenorphine, and ER naltrexone), which operate through direct antagonistic or agonistic action on the mu-opioid receptor, prevent subsequent clinical use of opioids for analgesia or anesthesia. Moreover, these reactive intervention-based strategies for OUD are not preventative and burden patients with self-admittance and retention of treatment. In fact, only 11% of people suffering from OUD receive treatment and less than 20.7% of those show sustained abstinence for at least five years. Thus, treatments that prevent or alleviate OUD without reducing the analgesic properties of opioids would be optimal.

[0352] Cysteine derivatives have shown significant promise in countering substance abuse in animal models, but clinical efficacy has been limited. For example, N-acetyl-L- cysteine (L-NAC) reduces cocaine-seeking behavior and reinstatement of heroin self- administration in rats and mice. Treatment with L-NAC has been shown to normalize glutamate and restore glutamate uptake by the glutamate transporter 1 (GLT-1) resulting in a reduction of drug seeking in animal models. However, clinical translation has been limited with reduced cocaine seeking observed under specific scenarios such as cocaine-primed cocaine seeking but not placebo-primed cocaine seeking in cocaine users. Importantly, these data reveal a novel approach to treatment with L-NAC as a preventative, as L-NAC was more efficacious in the presence of cocaine. We have shown that L-NAC co-treatment with fentanyl reduced the development of dependence as assessed by a reduction in naloxone- induced withdrawal symptoms while maintaining analgesic properties. Thus, treatment withL-NAC during opioid administration could reduce the development of OUD. We have shown that pretreatment with a novel cysteine derivative, D-cysteine ethylester, prevents the establishment of fentanyl reward seeking in both male and female rats. We now have evidence that L-NAC effectively prevents acquisition of fentanyl reward seeking in rats (Fig. 28). Rats were conditioned across eight days (30 min each day) with four pairings of fentanyl and four pairings of saline. Rats received either saline or L-NAC 30 min or 120 min prior to fentanyl on fentanyl conditioning days. In saline-treated controls, a robust fentanyl CPP was observed across days, but treatment with L-NAC during acquisition impaired acquisition of a fentanyl CPP. Thus, L-NAC effectively inhibits fentanyl reward indicating that co-administration of L-NAC with fentanyl could reduce the development of OUD. Furthermore, both L-NAC and D-cysteine ethylester reverse OIRD in rats using naloxone- precipitated withdrawal. These data support the use of cysteine derivatives, such as L-NAC, in the prevention of OUD development and OIRD without affecting analgesic efficacy.

[0353] From the above description of the invention, those skilled in the art will perceive improvements, changes and modifications. Such improvements, changes and modifications within the skill of the art are intended to be covered by the appended claims. All references, publications, and patents cited in the present application are herein incorporated by reference in their entirety.

Claims

Having described the invention, we claim:

1. A compound having a structure of formula (I):a pharmaceutically acceptable salt, a tautomer, or a solvate thereof; wherein: X is -OR2or -N(R3)2; X1is -SR4or -S(O)nOR5; R1is H, alkyl optionally substituted with one or more halogen, or -C(O)-alkyl optionally substituted with one or more halogen; R2, R5, and R6are each independently H or alkyl optionally substituted with one or more halogen; each R3is independently H or alkyl optionally substituted with one or more halogen; R4is -N=O or -OR6; and n is 1 or 2.

2. The compound of claim 1, wherein R1is H, C1-C6 alkyl optionally substituted with one or more halogen, or -C(O)-(C1-C6alkyl) optionally substituted with one or more halogen.

3. The compound of claims 1 or 2, wherein R1is H, methyl, ethyl, propyl, butyl, - C(O)-methyl, -C(O)-ethyl, -C(O)-propyl, or -C(O)-butyl, each optionally substituted with one or more halogen.

4. The compound of any of claims 1 to 3, wherein R2and R6are each independently H or C1-C6 alkyl optionally substituted with one or more halogen.

5. The compound of any of claims 1 to 4, wherein R2and R6are each independently H, methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

6. The compound of any of claims 1 to 5, wherein each R3is independently H or C1-C6alkyl optionally substituted with one or more halogen.

7. The compound of any of claims 1 to 6, wherein one of each R3is H and the other of R3is methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

8. The compound of claim 1, wherein X is OR2, R1is H, C1-C6 alkyl optionally substituted with one or more halogen, or -C(O)-(C1-C6 alkyl) optionally substituted with one or more halogen, and R2is H or C1-C6 alkyl optionally substituted with one or more halogen.

9. The compound of claim 1, wherein X is OR2, R1is H, methyl, ethyl, propyl, butyl, -C(O)-methyl, -C(O)-ethyl, -C(O)-propyl, or -C(O)-butyl, each optionally substituted with one or more halogen, and R2is H, methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

10. The compound of claim 1, wherein X is N(R3)2; R1is H, C1-C6alkyl optionally substituted with one or more halogen, or -C(O)-(C1-C6 alkyl) optionally substituted with one or more halogen; and each R3is independently H or C1-C6alkyl optionally substituted with one or more halogen.

11. The compound of claim 1, wherein X is N(R3)2; R1is H, methyl, ethyl, propyl, butyl, -C(O)-methyl, -C(O)-ethyl, -C(O)-propyl, or -C(O)-butyl, each optionally substituted with one or more halogen, and one R3is H and the other R3is methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

12. The compound of claim 1, wherein X1is SR4and R4is -N=O.

13. The compound of claim 1, wherein X1is SR4, R4is -OR6, and R6is H or C1-C6 alkyl optionally substituted with one or more halogen.

14. The compound of claim 1, wherein X1is SR4, R4is -OR6, and R6is H, methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

15. The compound of claim 1, wherein X1is -S(O)nOR5and R5is H or C1-C6 alkyl optionally substituted with one or more halogen.

16. The compound of claim 1, wherein X1is -S(O)nOR5and R5is H, methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

17. A compound having a structure of formula (II): (II), a pharmaceutically acceptable salt, a tautomer, or a solvate thereof; wherein: X is -OR2or -N(R3)2; R1is H, alkyl optionally substituted with one or more halogen, or -C(O)-alkyl optionally substituted with one or more halogen; R2is independently H or alkyl optionally substituted with one or more halogen; each R3is independently H or alkyl optionally substituted with one or more halogen.

18. The compound of claim 17, wherein R1is H, C1-C6 alkyl optionally substituted with one or more halogen, or -C(O)-(C1-C6alkyl) optionally substituted with one or more halogen.

19. The compound of claims 17 or 18, wherein R1is H, methyl, ethyl, propyl, butyl, -C(O)-methyl, -C(O)-ethyl, -C(O)-propyl, or -C(O)-butyl, each optionally substituted with one or more halogen.

20. The compound of any of claims 17 to 19, wherein R2is H or C1-C6 alkyl optionally substituted with one or more halogen.

21. The compound of any of claims 17 to 20, wherein R2is H, methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

22. The compound of any of claims 17 to 21, wherein each R3is independently H or C1-C6 alkyl optionally substituted with one or more halogen.

23. The compound of any of claims 17 to 22, wherein one of each R3is H and the other of R3is methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

24. A compound having a structure of formula (III):a pharmaceutically acceptable salt, a tautomer, or a solvate thereof; wherein: X is -OR2or -N(R3)2; R1is H, alkyl optionally substituted with one or more halogen, or -C(O)-alkyl optionally substituted with one or more halogen; R2and R6are each independently H or alkyl optionally substituted with one or more halogen; and each R3is independently H or alkyl optionally substituted with one or more halogen.

25. The compound of claim 24, wherein R1is H, C1-C6 alkyl optionally substituted with one or more halogen, or -C(O)-(C1-C6alkyl) optionally substituted with one or more halogen.

26. The compound of claims 24 or 25, wherein R1is H, methyl, ethyl, propyl, butyl, -C(O)-methyl, -C(O)-ethyl, -C(O)-propyl, or -C(O)-butyl, each optionally substituted with one or more halogen.

27. The compound of any of claims 24 to 26, wherein R2and R6are each independently H or C1-C6 alkyl optionally substituted with one or more halogen. 28 The compound of any of claims 24 to 27, wherein R2and R6are each independently H, methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

29. The compound of any of claim 24 to 28, wherein each R3is independently H or C1-C6alkyl optionally substituted with one or more halogen.

30. The compound of any of claims 24 to 29, wherein one of each R3is H and the other of R3is methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

31. A compound having a structure of formula (IV):a pharmaceutically acceptable salt, a tautomer, or a solvate thereof; wherein: X is -OR2or -N(R3)2; R1is H, alkyl optionally substituted with one or more halogen, or -C(O)-alkyl optionally substituted with one or more halogen;R2and R5are each independently H or alkyl optionally substituted with one or more halogen; and each R3is independently H or alkyl optionally substituted with one or more halogen.

32. The compound of claim 31, wherein R1is H, C1-C6alkyl optionally substituted with one or more halogen, or -C(O)-(C1-C6 alkyl) optionally substituted with one or more halogen.

33. The compound of claims 31 or 22, wherein R1is H, methyl, ethyl, propyl, butyl, -C(O)-methyl, -C(O)-ethyl, -C(O)-propyl, or -C(O)-butyl, each optionally substituted with one or more halogen.

34. The compound of any of claims 31 to 33, wherein R2and R5are each independently H or C1-C6alkyl optionally substituted with one or more halogen.

35. The compound of any of claims 31 to 34, wherein R2and R5are each independently H, methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

36. The compound of any of claims 31 to 35, wherein each R3is independently H or C1-C6 alkyl optionally substituted with one or more halogen.

37. The compound of any of claims 31 to 36, wherein one of each R3is H and the other of R3is methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

38. A compound having a structure of formula (V):a pharmaceutically acceptable salt, a tautomer, or a solvate thereof; wherein: X is -OR2or -N(R3)2; R1is H, alkyl optionally substituted with one or more halogen, or -C(O)-alkyl optionally substituted with one or more halogen; R2and R5are each independently H or alkyl optionally substituted with one or more halogen; and each R3is independently H or alkyl optionally substituted with one or more halogen.

39. The compound of claim 38, wherein R1is H, C1-C6 alkyl optionally substituted with one or more halogen, or -C(O)-(C1-C6 alkyl) optionally substituted with one or more halogen.

40. The compound of claims 38 or 39, wherein R1is H, methyl, ethyl, propyl, butyl, -C(O)-methyl, -C(O)-ethyl, -C(O)-propyl, or -C(O)-butyl, each optionally substituted with one or more halogen.

41. The compound of any of claims 38 to 40, wherein R2and R5are each independently H or C1-C6alkyl optionally substituted with one or more halogen.

42. The compound of any of claims 38 to 41, wherein R2and R5are each independently H, methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

43. The compound of any of claims 38 to 42, wherein each R3is independently H or C1-C6alkyl optionally substituted with one or more halogen.

44. The compound of any of claims 38 to 43, wherein one of each R3is H and the other of R3is methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

45. The compound of claim 1, having a structure selected from:pharmaceutically acceptable salt; a tautomer; a solvate thereof; or a combination thereof.

46. A pharmaceutical composition comprising the compounds of any of claim 1 to 4 47. A pharmaceutical composition for use in treating addiction, withdrawal and / or dependence in a subject in need thereof comprising: a compound having a structure of formulas:adduct, a pharmaceutically acceptable salt, a tautomer, or a solvate thereof; wherein: X is -OR2or -N(R3)2; X1is -SR4or -S(O)nOR5; X2is -OR9or -N(R10)2; X3is -OR11or -N(R12)2; R1, R7, and R8are each independently H, alkyl optionally substituted with one or more halogen, or -C(O)-alkyl optionally substituted with one or more halogen; R2, R5, R6, R9and R11are each independently H or alkyl optionally substituted with one or more halogen; each R3, R10, and R12are independently H or alkyl optionally substituted with one or more halogen; R4is H, -N=O or -OR6; n is 1 or 2; and optionally X is not -OR2if R1is H; and X3is not -OR11if X2is -OR9and R7and R8are H.

48. The composition of claim 47, wherein the adduct of the compound of formulas I or II is biologically active and includes at least one of an albumin adduct, a glucose adduct, an L-cysteine adduct, an L-glutathione adduct, or a D-cysteine adduct.

49. The composition of claim 47 or 48, wherein R1, R7, and R8are each independently H, C1-C6alkyl optionally substituted with one or more halogen, or -C(O)-(C1- C6 alkyl) optionally substituted with one or more halogen.

50. The composition of any of claims 47 to 49, wherein R1, R7, and R8are each independently H, methyl, ethyl, propyl, butyl, -C(O)-methyl, -C(O)-ethyl, -C(O)-propyl, or - C(O)-butyl, each optionally substituted with one or more halogen.

51. The composition of any of claims 47 to 50, wherein R2, R5, R6, R9and R11are each independently H or C1-C6 alkyl optionally substituted with one or more halogen.

52. The composition of any of claim 47 to 51, wherein R2, R5, R6, R9and R11are each independently H, methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

53. The composition of any of claim 47 to 52, wherein each R3, R10, and R12is independently H or C1-C6alkyl optionally substituted with one or more halogen.

54. The composition of any of claims 47 to 53, wherein one of each R3, R10, and R12is H and the other of R3, R6, and R9is methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

55. The composition of claim 47, wherein X is -OR2, R1is H, C1-C6alkyl optionally substituted with one or more halogen, or -C(O)-(C1-C6 alkyl) optionally substituted with one or more halogen, and R2is H or C1-C6 alkyl optionally substituted with one or more halogen.

56. The composition of claim 47, wherein X is -OR2, R1is H, methyl, ethyl, propyl, butyl, -C(O)-methyl, -C(O)-ethyl, -C(O)-propyl, or -C(O)-butyl, each optionally substituted with one or more halogen, and R2is H, methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

57. The composition of claim 47, wherein X is -N(R3)2; R1is H, C1-C6 alkyl optionally substituted with one or more halogen, or -C(O)-(C1-C6alkyl) optionally substituted with one or more halogen; and each R3is independently H or C1-C6 alkyl optionally substituted with one or more halogen.

58. The composition of claim 47, wherein X is -N(R3)2; R1is H, methyl, ethyl, propyl, butyl, -C(O)-methyl, -C(O)-ethyl, -C(O)-propyl, or -C(O)-butyl, each optionally substituted with one or more halogen, and one R3is H and the other R3is methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

59. The composition of claim 47, wherein X1is -SR4and R4is -N=O.

60. The composition of claim 47, wherein X1is -SR4and R4is H.

61. The composition of claim 47, wherein X1is -SR4, R4is -OR6, and R6is H or C1-C6 alkyl optionally substituted with one or more halogen.

62. The composition of claim 47, wherein X1is -SR4, R4is -OR6, and R6is H, methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

63. The composition of claim 47, wherein X1is -S(O)nOR5and R5is H or C1-C6alkyl optionally substituted with one or more halogen.

64. The composition of claim 47, wherein X1is -S(O)nOR5and R5is H, methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

65. The composition of claim 47, wherein X2is -OR9, R7is H, C1-C6 alkyl optionally substituted with one or more halogen, or -C(O)-(C1-C6alkyl) optionally substituted with one or more halogen, R9is H or C1-C6 alkyl optionally substituted with one or more halogen; X3is OR11, R8is H, C1-C6alkyl optionally substituted with one or more halogen, or -C(O)-(C1-C6 alkyl) optionally substituted with one or more halogen, and R11is H or C1-C6 alkyl optionally substituted with one or more halogen.

66. The composition of claim 47, wherein X2is OR9, R7is H, methyl, ethyl, propyl, butyl, -C(O)-methyl, -C(O)-ethyl, -C(O)-propyl, or -C(O)-butyl, each optionally substituted with one or more halogen, R9is H, methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen, X3is OR11; R8is H, methyl, ethyl, propyl, butyl, -C(O)-methyl, -C(O)-ethyl, -C(O)-propyl, or -C(O)-butyl, each optionally substituted with one or more halogen, and R11is H, methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

67. The composition of claim 47, wherein X2is N(R10)2, R7is H, C1-C6 alkyl optionally substituted with one or more halogen, or -C(O)-(C1-C6 alkyl) optionally substituted with one or more halogen, each R10is independently H or C1-C6 alkyl optionally substituted with one or more halogen, X3is N(R12)2, R8is H, C1-C6 alkyl optionally substituted with one or more halogen, or -C(O)-(C1-C6 alkyl) optionally substituted with one or more halogen, and each R12is independently H or C1-C6 alkyl optionally substituted with one or more halogen.

68. The composition of claim 47, wherein X2is N(R10)2, R7is H, methyl, ethyl, propyl, butyl, -C(O)-methyl, -C(O)-ethyl, -C(O)-propyl, or -C(O)-butyl, each optionally substituted with one or more halogen, one R10is H and the other R10is methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen, X3is N(R12)2; R8is H, methyl, ethyl, propyl, butyl, -C(O)-methyl, -C(O)-ethyl, -C(O)-propyl, or -C(O)-butyl, each optionally substituted with one or more halogen, and one R12is H and the other R12is methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

69. The composition of any of claims 47 to 68, wherein the compound is not cysteine, cystine, a cysteine alkylester, or cystine dialkylester.

70. The composition of any of claims 47 to 68, wherein the compound is not N- acetylcysteine.. The composition of claim 47, wherein the compound has a structure selected from:pharmaceutically acceptable salt, a tautomer, or a solvate thereof; or a combination thereof.

72. The composition of any of claims 47 to 71, further comprising a pharmaceutically acceptable carrier or excipient.

73. The composition of any of claims 47 to 72, wherein the subject has an addiction, withdrawal and / or dependence from at least one of alcohol, amphetamine, cocaine, methamphetamine, nicotine, or opioid.

74. The composition of any of claims 47 to 73, wherein the subject has an opioid withdrawal, dependence, and / or addiction.

75. The composition of any of claims 47 to 74, wherein the subject has neonatal opioid withdrawal syndrome.

76. The composition of any of claims 47 to 74, wherein the subject has an opioid use disorder (OUD).

77. The composition of any of claims 73 to 76, wherein the opioid comprises at least one of alfentanil, buprenorphine, butorphanol, carfentanil, codeine, diamorphine, dextromoramide, dezocine, dihydrocodeine, fentanyl, hydrocodone, hydromorphone, levorphanol, meperidine, meptazinol, methadone, morphine, nalbuphine, nalorphine, opium, oxycodone, oxymorphone, pentazocine, propoxyphene, remifentanil, sufentanil, tapentadol, and tramadol, and pharmaceutically acceptable salts thereof.

78. The composition of any of claims 73 to 77, wherein the opioid is carfentanil, fentanyl, remifentanil, or sufentanil.

79. The composition of any of claims 73 to 78, wherein the subject has been administered an opioid antagonist and the composition inhibits opioid antagonist withdrawal.

80. The composition of claim 79, wherein the opioid antagonist is naloxone, an oxymorphol analog of naloxone, a naloxone salt, or a naloxone dihydrate.

81. The composition of claim 79, wherein the opioid antagonist is naloxone.

82. The composition of claim 73 to 79, being administered concurrently with opioid administration and / or up to about 10 minutes, up to about 20 minutes, up to about 30 minutes, up to about 40 minutes, up to about 50 minutes, up to about 60 minutes, up to about 70 minutes, up to about 80 minutes, up to about 90 minutes, up to about 100 minutes, up to about 110 minutes, or up to about 120 minutes before or after initiation of opioid administration.

83. A pharmaceutical composition for use in preventing loss of opioid induced analgesia and occurrence opioid induced hyperalgesia in a subject in need thereof comprising: a compound having a structure of formulas:adduct, a pharmaceutically acceptable salt, a tautomer, or a solvate thereof; wherein: X is -OR2or -N(R3)2; X1is -SR4or -S(O)nOR5; X2is -OR9or -N(R10)2; X3is -OR11or -N(R12)2; R1, R7, and R8are each independently H, alkyl optionally substituted with one or more halogen, or -C(O)-alkyl optionally substituted with one or more halogen; R2, R5, R6, R9and R11are each independently H or alkyl optionally substituted with one or more halogen; each R3, R10, and R12are independently H or alkyl optionally substituted with one or more halogen; R4is H, -N=O or -OR6; n is 1 or 2; and optionally X is not -OR2if R1is H; and X3is not -OR11if X2is -OR9and R7and R8are H.

84. The composition of claim 83, wherein the adduct of the compound of formulas I or II is biologically active and includes at least one of an albumin adduct, a glucose adduct, an L-cysteine adduct, an L-glutathione adduct, or a D-cysteine adduct.

85. The composition of claim 83 or 84, wherein R1, R7, and R8are each independently H, C1-C6alkyl optionally substituted with one or more halogen, or -C(O)-(C1- C6 alkyl) optionally substituted with one or more halogen.

86. The composition of any of claims 83 to 85, wherein R1, R7, and R8are each independently H, methyl, ethyl, propyl, butyl, -C(O)-methyl, -C(O)-ethyl, -C(O)-propyl, or - C(O)-butyl, each optionally substituted with one or more halogen.

87. The composition of any of claims 43 to 86, wherein R2, R5, R6, R9and R11are each independently H or C1-C6alkyl optionally substituted with one or more halogen.

88. The composition of any of claim 83 to 87, wherein R2, R5, R6, R9and R11are each independently H, methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

89. The composition of any of claim 83 to 88, wherein each R3, R10, and R12is independently H or C1-C6 alkyl optionally substituted with one or more halogen.

90. The composition of any of claims 83 to 89, wherein one of each R3, R10, and R12is H and the other of R3, R6, and R9is methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

91. The composition of claim 83, wherein X is -OR2, R1is H, C1-C6alkyl optionally substituted with one or more halogen, or -C(O)-(C1-C6 alkyl) optionally substituted with one or more halogen, and R2is H or C1-C6alkyl optionally substituted with one or more halogen.

92. The composition of claim 83, wherein X is -OR2, R1is H, methyl, ethyl, propyl, butyl, -C(O)-methyl, -C(O)-ethyl, -C(O)-propyl, or -C(O)-butyl, each optionally substituted with one or more halogen, and R2is H, methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

93. The composition of claim 83, wherein X is -N(R3)2; R1is H, C1-C6 alkyl optionally substituted with one or more halogen, or -C(O)-(C1-C6alkyl) optionally substituted with one or more halogen; and each R3is independently H or C1-C6 alkyl optionally substituted with one or more halogen.

94. The composition of claim 83, wherein X is -N(R3)2; R1is H, methyl, ethyl, propyl, butyl, -C(O)-methyl, -C(O)-ethyl, -C(O)-propyl, or -C(O)-butyl, each optionally substituted with one or more halogen, and one R3is H and the other R3is methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

95. The composition of claim 83, wherein X1is -SR4and R4is -N=O.

96. The composition of claim 83, wherein X1is -SR4and R4is H.

97. The composition of claim 83, wherein X1is -SR4, R4is -OR6, and R6is H or C1-C6 alkyl optionally substituted with one or more halogen.

98. The composition of claim 83, wherein X1is -SR4, R4is -OR6, and R6is H, methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

99. The composition of claim 83, wherein X1is -S(O)nOR5and R5is H or C1-C6 alkyl optionally substituted with one or more halogen.

100. The composition of claim 83, wherein X1is -S(O)nOR5and R5is H, methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

101. The composition of claim 83, wherein X2is -OR9, R7is H, C1-C6 alkyl optionally substituted with one or more halogen, or -C(O)-(C1-C6alkyl) optionally substituted with one or more halogen, R9is H or C1-C6 alkyl optionally substituted with one or more halogen; X3is OR11, R8is H, C1-C6 alkyl optionally substituted with one or more halogen, or -C(O)-(C1-C6alkyl) optionally substituted with one or more halogen, and R11is H or C1-C6alkyl optionally substituted with one or more halogen.

102. The composition of claim 83, wherein X2is OR9, R7is H, methyl, ethyl, propyl, butyl, -C(O)-methyl, -C(O)-ethyl, -C(O)-propyl, or -C(O)-butyl, each optionally substituted with one or more halogen, R9is H, methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen, X3is OR11; R8is H, methyl, ethyl, propyl,butyl, -C(O)-methyl, -C(O)-ethyl, -C(O)-propyl, or -C(O)-butyl, each optionally substituted with one or more halogen, and R11is H, methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

103. The composition of claim 83, wherein X2is N(R10)2, R7is H, C1-C6 alkyl optionally substituted with one or more halogen, or -C(O)-(C1-C6alkyl) optionally substituted with one or more halogen, each R10is independently H or C1-C6 alkyl optionally substituted with one or more halogen, X3is N(R12)2, R8is H, C1-C6alkyl optionally substituted with one or more halogen, or -C(O)-(C1-C6 alkyl) optionally substituted with one or more halogen, and each R12is independently H or C1-C6 alkyl optionally substituted with one or more halogen.

104. The composition of claim 83, wherein X2is N(R10)2, R7is H, methyl, ethyl, propyl, butyl, -C(O)-methyl, -C(O)-ethyl, -C(O)-propyl, or -C(O)-butyl, each optionally substituted with one or more halogen, one R10is H and the other R10is methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen, X3is N(R12)2; R8is H, methyl, ethyl, propyl, butyl, -C(O)-methyl, -C(O)-ethyl, -C(O)-propyl, or -C(O)-butyl, each optionally substituted with one or more halogen, and one R12is H and the other R12is methyl, ethyl, or propyl, or butyl, each optionally substituted with one or more halogen.

105. The composition of any of claims 83 to 105, wherein the compound is not cysteine, cystine, a cysteine alkylester, or cystine dialkylester.

106. The composition of any of claims 83 to 105, wherein the compound is not N- acetylcysteine.

107. The composition of claim 83, wherein the compound has a structure selected from:acceptable salt, a tautomer, a solvate thereof; or a combination thereof.

108. The composition of any of claims 83 to 107, further comprising a pharmaceutically acceptable carrier or excipient.

109. The composition of any of claims 83 to 107, wherein the opioid comprises at least one of alfentanil, buprenorphine, butorphanol, carfentanil, codeine, diamorphine, dextromoramide, dezocine, dihydrocodeine, fentanyl, hydrocodone, hydromorphone, levorphanol, meperidine, meptazinol, methadone, morphine, nalbuphine, nalorphine, opium, oxycodone, oxymorphone, pentazocine, propoxyphene, remifentanil, sufentanil, tapentadol, and tramadol, and pharmaceutically acceptable salts thereof.

110. The composition of any of claims 83 to 109, wherein the opioid is carfentanil, fentanyl, remifentanil, or sufentanil.

111. The composition of claim 83 to 110, being administered concurrently with opioid administration and / or up to about 10 minutes, up to about 20 minutes, up to about 30 minutes, up to about 40 minutes, up to about 50 minutes, up to about 60 minutes, up to about 70 minutes, up to about 80 minutes, up to about 90 minutes, up to about 100 minutes, up to about 110 minutes, or up to about 120 minutes before or after initiation of opioid administration.

112. A pharmaceutical composition comprising at least one opioid and an amount of a compound of any of claims 1 to 45 effective to reduce opioid addiction, withdrawal, and / or dependence and / or loss of opioid induced analgesia and / or occurrence opioid induced hyperalgesia.

113. The composition of claim 112, further comprising a pharmaceutically acceptable carrier or excipient.

114. The composition of claim 112 or 113, wherein the opioid comprises at least one of alfentanil, buprenorphine, butorphanol, carfentanil, codeine, diamorphine, dextromoramide, dezocine, dihydrocodeine, fentanyl, hydrocodone, hydromorphone, levorphanol, meperidine, meptazinol, methadone, morphine, nalbuphine, nalorphine, opium, oxycodone, oxymorphone, pentazocine, propoxyphene, remifentanil, sufentanil, tapentadol, and tramadol, and pharmaceutically acceptable salts thereof.

115. The composition of any of claims 112 to 114, wherein the opioid is carfentanil, fentanyl, remifentanil, or sufentanil.

116. The composition of any of claims 46 to 115, being formulated for systemic administration.

117. The composition of any of claims 46 to 115, being formulated for continuous intravenous infusion.

118. The composition of any of claims 46 to 115, being formulated for oral administration.

119. The composition of any of claims 46 to 115, being formulated for intranasal administration.

120. The composition of any of claims 46 to 115, being formulated for inhalation.

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