Dosing of polysubunit opioid prodrugs resistant to overdose and abuse

Abstract

Provided is a pharmaceutical composition comprising two different molecules, wherein each molecule comprises at least one GI enzyme-labile opioid agonist-releasing subunit comprising an opioid agonist, wherein the at least one GI enzyme-labile opioid agonist-releasing subunit is covalently linked to at least one GI enzyme-inhibitor subunit, wherein each GI enzyme-labile opioid agonist-releasing subunit releases the opioid agonist, and the resulting opioid agonist plasma Cmax concentrations are less than dose proportional compared to an analogous opioid agonist and is below the lethal exposure of an analogous free opioid agonist when administered at supra-therapeutic dosages.

Description

DOSING OF POLYSUBUNIT OPIOID PRODRUGS RESISTANTTO OVERDOSE AND ABUSECROSS REFERENCE

[0001] This application claims the benefit of U.S. Provisional Application Serial No. 63 / 632,294 filed April 10, 2024, and U.S. Provisional Application No. 63 / 634,039 filed April 15, 2024 which is hereby incorporated by reference in their entirety.STATEMENT AS TO FEDERALLY SPONSORED RESEARCH

[0002] This invention was made with the support of the United States government under SBIR Grant numbers 1R44DA037900, 1R43DA046302, and 1R01DA046102 by the National Institute on Drug Abuse (NIDA), one of the National Institutes of Health (NIH) in the U.S. Department of Health and Human Services.SUMMARY

[0003] In an aspect, provided herein is a pharmaceutical composition comprising two different molecules, wherein each molecule comprises at least one GI enzyme-labile opioid agonist-releasing subunit comprising an opioid agonist, wherein the at least one GI enzyme- labile opioid agonist-releasing subunit is covalently linked to at least one GI enzyme-inhibitor subunit, wherein each GI enzyme-labile opioid agonist-releasing subunit releases the opioid agonist, and the resulting opioid agonist plasma Cmaxconcentrations are less than dose proportional compared to an analogous opioid agonist.

[0004] In another aspect, provided herein is a pharmaceutical composition comprising two different molecules, wherein each molecule comprises at least one GI enzyme-labile opioid agonist-releasing subunit comprising an opioid agonist, wherein the at least one GI enzyme-labile opioid agonist-releasing subunit is covalently linked to at least one GI enzymeinhibitor subunit, wherein the GI enzyme-labile opioid agonist-releasing subunit releases the opioid agonist, and the opioid agonist provides an analgesic effect for about 24 hours to about 48 hours. In some embodiments, the opioid agonist provides an analgesic effect for about 12 hours to about 24 hours. In some embodiments, the onset of analgesia occurs within 2 hours after administration. In some embodiments, the onset of analgesia occurs within 1 hour after administration.

[0005] In another aspect, provided herein is a pharmaceutical composition comprising two different molecules, wherein each molecule comprises at least one GI enzyme-labileopioid agonist-releasing subunit comprising an opioid agonist, wherein the at least one GI enzyme-labile opioid agonist-releasing subunit is covalently linked to at least one GI enzymeinhibitor subunit, wherein the GI enzyme-labile opioid agonist-releasing subunit releases the opioid agonist, and the opioid agonist achieves a Cmaxafter about 6 hours. In some embodiments, the opioid agonist achieves a mean max plasma concentration after about 6 hours to about 10 hours. In some embodiments, the opioid agonist achieves a mean max plasma concentration after about 6 hours to about 8 hours. In some embodiments, the opioid agonist achieves a mean max plasma concentration after about 8 hours.

[0006] In another aspect, provided herein is a pharmaceutical composition comprising two different molecules, wherein each molecule comprises at least one GI enzyme-labile opioid agonist-releasing subunit comprising an opioid agonist, wherein the at least one GI enzyme-labile opioid agonist-releasing subunit is covalently linked to at least one GI enzymeinhibitor subunit, wherein the GI enzyme-labile opioid agonist-releasing subunit releases the opioid agonist, wherein the Cmaxof the opioid agonist occurs after about 6 hours upon administration the pharmaceutical composition to a subject, and is reduced by at least 50% about 24 hours subsequent to the administration.

[0007] In another aspect, provided herein is a pharmaceutical composition comprising two different molecules, wherein each molecule comprises at least one GI enzyme-labile opioid agonist-releasing subunit comprising an opioid agonist, wherein the at least one GI enzyme-labile opioid agonist-releasing subunit is covalently linked to at least one GI enzymeinhibitor subunit, wherein the GI enzyme-labile opioid agonist-releasing subunit releases the opioid agonist, wherein the Cmaxof the opioid agonist is significantly reduced by about at least 40% at a supra-therapeutic dosage compared to an analogous free opioid agonist. In some embodiments, the Cmaxof the opioid agonist is significantly reduced by about at least 50% at a supra-therapeutic dosage compared to an analogous free opioid agonist.

[0008] In another aspect, provided herein is a pharmaceutical composition comprising two different molecules, wherein each molecule comprises at least one GI enzyme-labile opioid agonist-releasing subunit comprising an opioid agonist, wherein the at least one GI enzyme-labile opioid agonist-releasing subunit is covalently linked to at least one GI enzymeinhibitor subunit, wherein the GI enzyme-labile opioid agonist-releasing subunit releases the opioid agonist, wherein the opioid agonist has a plasma concentration of about 1 ng / mL or greater about 24 hours after administration of one dose. In some embodiments, the opioid agonist has a plasma concentration of about 3 ng / mL or greater about 24 hours after administration of one dose.

[0009] In some embodiments, the two different molecules are independently selected from the group consisting of a structure of Formula (IF), (IG), (IH), (II), and salts thereof:and salts thereof; wherein:D is an opioid agonist;R1of R1-Z1- is independently selected from the group consisting ofwherein Y is amidine, guanidine, aminomethyl,substituted amidine, substituted guanidine, substituted aminomethyl, amidinomethyl, guanidinomethyl, substituted amidinomethyl, or substituted guanidinomethyl, and Q is hydrogen;R’ is selected from the group consisting of methyl, lower alkyl, substituted alkyl, aryl, substituted aryl, heteroalkyl, a natural or a non-natural amino acid, a polypeptide chain comprising natural or non-natural amino acids up to 10 amino acidsin length, a linear or a branched polyethylene glycol chain up to 5 kDa, benzyloxy,R” is selected from the group consisting of acetyl, substituted acyl, a natural or a non-natural amino acid, and a polypeptide chain comprising natural or non-natural amino acids up to 10 amino acids in length;AA is a natural or a non-natural amino acid side chain that is recognized by trypsin; andZ1of R1-Z1- is independently selected from the group consisting of R1-O-CH2-CH2- and R1-CH2-C(O)-NH-CH2-CH2-.

[0010] In some embodiments, the first opioid agonist comprises morphine, hydromorphone, hydrocodone, oxycodone, codeine, levorphanol, meperidine, methadone, oxymorphone, dihydrocodeine, tramadol, tapentadol, buprenorphine, cebranopadol, or pharmaceutically acceptable salts, prodrugs, and mixtures thereof. In some embodiments, the first opioid agonist comprises morphine, or pharmaceutically acceptable salts thereof. In some embodiments, the first opioid agonist comprises hydrocodone, or pharmaceutically acceptable salts thereof. In some embodiments, the first opioid agonist comprises oxycodone, or pharmaceutically acceptable salts thereof.

[0011] In some embodiments, the two different molecules is a structure of Compound A and Compound B :wherein the ratio of Compound A to Compound B is between 1 :4 w / w and 1 : 1 w / w, and the pharmaceutical composition is administered once or twice daily.

[0012] In another aspect, provided herein is a pharmaceutical composition comprising a first opioid agonist and one or two different molecules, wherein each molecule comprises at least one Gl-enzyme-labile opioid agonist-releasing subunit comprising a second opioid agonist, wherein the at least one GI enzyme-labile opioid agonist-releasing subunit is covalently linked to at least one GI enzyme inhibitor subunit wherein the first opioid agonist reduces the onset of action time, wherein the first opioid agonist comprises morphine, hydromorphone, hydrocodone, oxycodone, codeine, levorphanol, meperidine, methadone, oxymorphone, dihydrocodeine, tramadol, tapentadol, buprenorphine, cebranopadol, or pharmaceutically acceptable salts, prodrugs, and mixtures thereof, wherein the one or two different molecules are independently selected from the group consisting of a structure of Formula (IF), (IG), (IH), (II), and salts thereof:wherein:D is an opioid agonist;R1of R1-Z1- is independently selected from the group consisting of, wherein Y is amidine, guanidine, aminomethyl, substituted amidine, substituted guanidine, substituted aminomethyl, amidinomethyl, guanidinomethyl, substituted amidinomethyl, or substituted guanidinomethyl, and Q is hydrogen;R’ is selected from the group consisting of methyl, lower alkyl, substituted alkyl, aryl, substituted aryl, heteroalkyl, a natural or a non-natural amino acid, a polypeptide chain comprising natural or non-natural amino acids up to 10 amino acids in length, a linear or a branched polyethylene glycol chain up to 5 kDa, benzyloxy,R” is selected from the group consisting of acetyl, substituted acyl, a natural or a non-natural amino acid, and a polypeptide chain comprising natural or non-natural amino acids up to 10 amino acids in length;AA is a natural or a non-natural amino acid side chain that is recognized by trypsin; andZ1of R1-Z1- is independently selected from the group consisting of R1-O-CH2-CH2- and R1-CH2-C(O)-NH-CH2-CH2-.

[0013] In some embodiments, the two different molecules comprise a structure of Compound A and Compound B:the ratio of Compound A to Compound B is between 1 :6 w / w and 6: 1 w / w, and the pharmaceutical composition is administered once or twice daily.

[0014] In some embodiments, the two different molecules are in a ratio of 1 :4 w / w. In some embodiments, the two different molecules are in a ratio of 1 :3 w / w. In some embodiments, the two different molecules are in a ratio of 1 :2 w / w. In some embodiments, the two different molecules are in a ratio of 1 :1 w / w. In some embodiments, the pharmaceutical composition comprises about 10 mg to 1000 mg of the two different molecules. In some embodiments, the pharmaceutical composition comprises 60 mg of the two different molecules. In some embodiments, the pharmaceutical composition comprises 120 mg of the two different molecules. In some embodiments, the pharmaceutical composition comprises 240 mg of the two different molecules. In some embodiments, the pharmaceutical composition comprises 480 mg of the two different molecules. In some embodiments, the pharmaceutical composition comprises 960 mg of the two different molecules. In some embodiments, the first opioid agonist comprises between about 0.1 mg to about 20 mg in the pharmaceutical composition. In some embodiments, the first opioid agonist comprises no more than 10 mg in the pharmaceutical composition. In some embodiments, the first opioid agonist comprises about 6 mg in the pharmaceutical composition. In some embodiments, the first opioid agonist comprises about 3 mg in the pharmaceutical composition. In some embodiments, the first opioid agonist comprises about 1.5 mg in the pharmaceutical composition. In some embodiments, the first opioid agonist comprises about 0.5 mg in the pharmaceutical composition. In some embodiments, the pharmaceutical composition is administered once daily. In some embodiments, the pharmaceutical composition is administered twice daily.

[0015] In another aspect, provided herein is a method of treating pain in a subject, the method comprising administering to the subject a pharmaceutical composition described herein. In some embodiments, the pain is acute pain. In some embodiments, the pain is chronic pain. In some embodiments, the Cmaxof the free opioid agonist is reached between 6 and 10 hours. In some embodiments, the opioid agonist plasma concentration within the subject is below the lethal amount. In some embodiments, the opioid agonist plasma concentration in the subject demonstrates reduced dose proportionality when compared to the free opioid agonist delivered by the one or two different molecules. In some embodiments, the administration of a therapeutically effective dose of a pharmaceutical composition described herein reduces events of abuse, misuse, and overdose, when compared to the administration of the free opioid agonist.BRIEF DESCRIPTION OF THE DRAWINGS

[0016] Various aspects of the disclosure are set forth with particularity in the appended claims. A better understanding of the features and advantages of the present disclosure will be obtained by reference to the following detailed description that sets forth illustrative embodiments, in which the principles of the disclosure are utilized, and the accompanying drawings below.

[0017] FIG. 1 illustrates regioisomeric chemical structures of 1,4-substituted Compound A (ETR028) and 1,3 -substituted Compound B (ETR029).

[0018] FIG. 2 illustrates key structural components of Compound A and B including hydrocodone (A), opioid delivery subunit (B), trypsin inhibition subunit (C), and lysine derived linker (D).

[0019] FIG. 3 illustrates hydrocodone plasma concentration vs. time data for escalated doses of Compound A and B alone.

[0020] FIG. 4 illustrates hydrocodone plasma concentration vs. time data for representative Compound [A + B] blend ratios and doses in humans.

[0021] FIG. 5 illustrates hydrocodone plasma concentration vs. time data for [30 mg + 30 mg] and [60 mg + 60 mg] Compound [A + B] blend doses vs. 5 and 10 mg hydrocodone bitartrate comparator in humans.

[0022] FIG. 6 illustrates mean maximum hydrocodone plasma exposures (Cmax) vs. dose data for escalating Compound [A 30 mg + B 30 mg] blend doses in humans vs. extrapolated dose proportional exposures.

[0023] FIG. 7 illustrates mean maximum hydrocodone plasma exposures (Cmax) vs. dose data for escalating Compound [A 30 mg + B 30 mg] blend doses vs. HCBT dose proportional exposures relative to a potentially lethal hydrocodone exposure in humans.DETAILED DESCRIPTION

[0024] In today’s opioid crisis, there are two groups suffering: (i) patients dealing with moderate-to-severe pain, and (ii) those with Opioid Use Disorder due to the inherent risks of prescription opioids.

[0025] Opioid agonists provide unmatched analgesic efficacy especially when compared to non-opioid alternatives. Opioid agonists, when taken as prescribed, offer safe and effective pain relief to patients suffering from a wide spectrum of pain pathologies.

[0026] A new class of opioids is needed that reduce suffering from fatal opioid overdose, opioid-use disorder, and moderate-to-severe acute pain that cannot be effectively treated with non-opioid analgesics.

[0027] Despite a multiple-decades long quest to develop non-opioid analgesics with comparable potency to opioid agonists, current and emerging non-opioid analgesics continue to fail to meet clinical efficacy endpoints or demonstrate lackluster efficacy against moderate- to-severe pain for which opioids are highly effective. Further, non-opioid analgesics (e.g., Vioxx™, Tylenol™, Motrin™) often demonstrate serious side-effects which are frequently dose-limiting.

[0028] The vast majority (>90%) of the abuse of short-acting prescription opioids is via the ingestion of multiple intact tablets in excess of the prescribed dose (“supratherapeutic dose”). This is reinforced by two key factors (i) the dosing regimen of short-acting opioids necessitates a high number of prescribed tablets to provide the required duration of analgesia resulting in multiple opportunities to reinforce abuse; and, (ii) the incentive for abusers to ingest supratherapeutic doses for increased euphoric effects, often resulting in opioid use disorders and potentially lethal overdoses.

[0029] Herein is present oral overdose protected (O2P) opioid prodrugs designed to effectively mitigate non-oral and oral abuse while safely delivering an effective and well- known FDA approved opioid agonist. O2P opioids are indicated for the effective treatment of moderate-to-severe pain that cannot be effectively relieved with non-opioid analgesics.

[0030] O2P hydrocodone is indicated for the treatment of moderate-to-severe acute pain and offer a highly sought-after and unprecedented profile, including:Efficient delivery of therapeutic hydrocodone exposures at prescribed doses;Attenuation of hydrocodone exposures (e.g., Cmax) when supratherapeutic doses are ingested; Hydrocodone pharmacokinetics consistent with once-daily dosing, which reduce the number of pills prescribed; thus, reducing the risk of abuse and diversion;Protection against non-oral abuse and overdose (i.e., hydrocodone not activated unless orally administered); andRobust chemical stability that renders O2P molecules highly tamper-resistant.

[0031] O2P opioid prodrug technology is broadly applicable beyond hydrocodone to all currently prescribed oral opioid agonists.O2P Mechanism of Action

[0032] O2P prodrugs leverage bifunctional hydrocodone prodrug molecules that comprise (i) a trypsin-activated opioid delivery subunit that efficiently releases therapeutic levels of hydrocodone only when exposed to the digestive enzyme trypsin in the lumen of the small intestine, and (ii) a trypsin inhibitor subunit that progressively inhibits trypsin, and thereby the trypsin-mediated release of hydrocodone, when supratherapeutic doses are ingested. This unique mechanism has been demonstrated to (i) thwart abuse via non-oral routes of administration; (ii) provide unprecedented oral overdose protection in humans; and, (iii) significantly extend the duration of action, allowing for a substantial reduction in prescribed tablets.Pharmaceutical Composition

[0033] In an aspect, described herein is a pharmaceutical composition comprising two different molecules, wherein each molecule comprises at least one GI enzyme-labile opioid agonist-releasing subunit comprising an opioid agonist, wherein the at least one GI enzyme- labile opioid agonist-releasing subunit is covalently linked to at least one GI enzyme-inhibitor subunit, wherein each GI enzyme-labile opioid agonist-releasing subunit releases the opioid agonist, and the resulting opioid agonist plasma Cmaxconcentrations are less than dose proportional compared to an analogous opioid agonist.

[0034] In another aspect, described herein is a pharmaceutical composition comprising two different molecules, wherein each molecule comprises at least one GI enzyme-labile opioid agonist-releasing subunit comprising an opioid agonist, wherein the at least one GI enzyme-labile opioid agonist-releasing subunit is covalently linked to at least one GI enzymeinhibitor subunit, wherein the GI enzyme-labile opioid agonist-releasing subunit releases the opioid agonist, and the opioid agonist achieves a Cmaxafter about 6 hours.

[0035] In an aspect, provided herein is a pharmaceutical composition comprising two different molecules, wherein each molecule comprises at least one GI enzyme-labile opioid agonist-releasing subunit comprising an opioid agonist, wherein the at least one GI enzyme- labile opioid agonist-releasing subunit is covalently linked to at least one GI enzyme-inhibitor subunit, wherein the GI enzyme-labile opioid agonist-releasing subunit releases the opioid agonist, and the opioid agonist achieves a mean plasma concentration after about 6 hours.

[0036] In some embodiments, the opioid agonist achieves a mean max plasma concentration after about 6 hours to about 10 hours. In some embodiments, the opioid agonist achieves a mean max plasma concentration after about 6 hours to about 8 hours. In some embodiments, the opioid agonist achieves a mean max plasma concentration after about 10 hours. In some embodiments, the opioid agonist achieves a mean max plasma concentration after about 8 hours. In some embodiments, the opioid agonist achieves a mean max plasma concentration after about 6 hours.

[0037] In another aspect, provided herein is a pharmaceutical composition comprising two different molecules, wherein each molecule comprises at least one GI enzyme-labile opioid agonist-releasing subunit comprising an opioid agonist, wherein the at least one GI enzyme-labile opioid agonist-releasing subunit is covalently linked to at least one GI enzymeinhibitor subunit, wherein the GI enzyme-labile opioid agonist-releasing subunit releases the opioid agonist, and the opioid agonist provides an analgesic effect for about 24 hours to about 48 hours.

[0038] In some embodiments, the opioid agonist provides an analgesic effect for about 6 hours to about 48 hours. In some embodiments, the opioid agonist provides an analgesic effect for about 24 hours to about 48 hours. In some embodiments, the opioid agonist provides an analgesic effect for about 12 hours to about 24 hours. In some embodiments, the opioid agonist provides an analgesic effect for about 8 hours to about 10 hours. In some embodiments, the opioid agonist provides an analgesic effect for about 6 hours to about 12 hours. In some embodiments, the opioid agonist provides an analgesic effect for about 48 hours. In some embodiments, the opioid agonist provides an analgesic effect for about 36 hours. In some embodiments, the opioid agonist provides an analgesic effect for about 24 hours. In some embodiments, the opioid agonist provides an analgesic effect for about 12 hours. In some embodiments, the opioid agonist provides an analgesic effect for about 10 hours. In some embodiments, the opioid agonist provides an analgesic effect for about 6 hours.

[0039] In some embodiments, the onset of analgesia occurs within 2 hours after administration of the pharmaceutical composition. In some embodiments, the onset of analgesia occurs within 1 hour after administration of the pharmaceutical composition. In some embodiments, the onset of analgesia occurs 30 minutes after administration of the pharmaceutical composition. In some embodiments, the onset of analgesia occurs 10 minutes after administration of the pharmaceutical composition. In some embodiments, the onset of analgesia occurs 5 minutes after administration of the pharmaceutical composition.

[0040] In another aspect, described herein is a pharmaceutical composition comprising two different molecules, wherein each molecule comprises at least one GI enzyme-labile opioid agonist-releasing subunit comprising an opioid agonist, wherein the at least one GI enzyme-labile opioid agonist-releasing subunit is covalently linked to at least one GI enzymeinhibitor subunit, wherein the GI enzyme-labile opioid agonist-releasing subunit releases the opioid agonist, wherein the Cmaxof the opioid agonist occurs after about 6 hours upon administration the pharmaceutical composition to a subject, and is reduced by at least 50% from about 12 to 24 hours subsequent to the administration. In some embodiments, the Cmaxof the opioid agonist occurs after about 6 hours upon administration the pharmaceutical composition to a subject, and is reduced by at least 50% about 24 hours subsequent to the administration. In some embodiments, the Cmaxof the opioid agonist occurs after about 6 hours upon administration the pharmaceutical composition to a subject, and is reduced by at least 40% about 24 hours subsequent to the administration. In some embodiments, the Cmaxof the opioid agonist occurs after about 6 hours upon administration the pharmaceutical composition to a subject, and is reduced by at least 50% about 12 hours subsequent to the administration. In some embodiments, the Cmaxof the opioid agonist occurs after about 6 hours upon administration the pharmaceutical composition to a subject, and is reduced by at least 40% about 12 hours subsequent to the administration.

[0041] In another aspect, described herein is a pharmaceutical composition comprising two different molecules, wherein each molecule comprises at least one GI enzyme-labile opioid agonist-releasing subunit comprising an opioid agonist, wherein the at least one GI enzyme-labile opioid agonist-releasing subunit is covalently linked to at least one GI enzymeinhibitor subunit, wherein the GI enzyme-labile opioid agonist-releasing subunit releases the opioid agonist, wherein the Cmaxof the opioid agonist is significantly reduced when compared to an analogous free opioid agonist.

[0042] In some embodiments, the Cmaxof the opioid agonist is reduced by about 50% when compared to an analogous free opioid agonist. In some embodiments, the Cmaxof theopioid agonist is reduced by about 40% when compared to an analogous free opioid agonist. In some embodiments, the Cmaxof the opioid agonist is reduced by about 30% when compared to an analogous free opioid agonist. In some embodiments, the Cmaxof the opioid agonist is reduced by about 25% when compared to an analogous free opioid agonist. In some embodiments, the Cmaxof the opioid agonist is reduced by about 20% when compared to an analogous free opioid agonist. In some embodiments, the Cmaxof the opioid agonist is reduced by about 10% when compared to an analogous free opioid agonist.

[0043] In some embodiments, the Cmaxof the opioid agonist is significantly reduced by about at least 50% at a supra-therapeutic dosage compared to an analogous free opioid agonist. In some embodiments, the Cmaxof the opioid agonist is significantly reduced by about at least 40% at a supra-therapeutic dosage compared to an analogous free opioid agonist. . In some embodiments, the Cmaxof the opioid agonist is significantly reduced by about at least 30% at a supra-therapeutic dosage compared to an analogous free opioid agonist. . In some embodiments, the Cmaxof the opioid agonist is significantly reduced by about at least 25% at a supra-therapeutic dosage compared to an analogous free opioid agonist. In some embodiments, the opioid agonist demonstrates reduced dose proportionality when compared to an analogous free opioid agonist. In some embodiments, the dose proportionality is not linear.

[0044] In another aspect, described herein is a pharmaceutical composition comprising two different molecules, wherein each molecule comprises at least one GI enzyme-labile opioid agonist-releasing subunit comprising an opioid agonist, wherein the at least one GI enzyme-labile opioid agonist-releasing subunit is covalently linked to at least one GI enzymeinhibitor subunit, wherein the GI enzyme-labile opioid agonist-releasing subunit releases the opioid agonist, wherein the opioid agonist maintains an effective plasma concentration for about 24 hours after administration of one dose. In some embodiments, the opioid agonist maintains an effective plasma concentration for about 12 hours after administration of one dose. In some embodiments, the opioid agonist maintains an effective plasma concentration for about 10 hours after administration of one dose. In some embodiments, the opioid agonist maintains an effective plasma concentration for about 8 hours after administration of one dose. In some embodiments, the opioid agonist maintains an effective plasma concentration for about 6 hours after administration of one dose.

[0045] In some embodiments, the opioid agonist has a plasma concentration of about 0.5 ng / mL, about 1 ng / mL, about 1.5 ng / mL, about 2 ng / mL, about 3 ng / mL or greater, about 5 ng / mL, about 10 ng / mL, about 20 ng / mL, about 30 ng / mL, about 40 ng / mL, or about 50ng / mL. In some embodiments, the opioid agonist has a plasma concentration of about 1 ng / mL or greater about 24 hours after administration of one dose. In some embodiments, the opioid agonist has a plasma concentration of about 1.5 ng / mL or greater about 24 hours after administration of one dose. In some embodiments, the opioid agonist has a plasma concentration of about 2 ng / mL or greater about 24 hours after administration of one dose. In some embodiments, the opioid agonist has a plasma concentration of about 3 ng / mL or greater about 24 hours after administration of one dose. In some embodiments, the opioid agonist has a plasma concentration of about 5 ng / mL or greater about 24 hours after administration of one dose. In some embodiments, the opioid agonist has a plasma concentration of about 10 ng / mL or greater about 24 hours after administration of one dose. In some embodiments, the opioid agonist has a plasma concentration of about 20 ng / mL or greater about 24 hours after administration of one dose. In some embodiments, the opioid agonist has a plasma concentration of about 30 ng / mL or greater about 24 hours after administration of one dose.

[0046] In some embodiments, the opioid agonist has a plasma concentration of about 1 ng / mL or greater after administration of one dose. In some embodiments, the opioid agonist has a plasma concentration of about 3 ng / mL or greater after administration of one dose. In some embodiments, the opioid agonist has a plasma concentration of about 5 ng / mL or greater after administration of one dose. In some embodiments, the opioid agonist has a plasma concentration of about 10 ng / mL or greater after administration of one dose. In some embodiments, the opioid agonist has a plasma concentration of about 20 ng / mL or greater after administration of one dose. In some embodiments, the opioid agonist has a plasma concentration of about 30 ng / mL or greater after administration of one dose.

[0047] In some embodiments, the GI enzyme-labile opioid agonist-releasing subunits comprises morphine, hydromorphone, hydrocodone, oxycodone, codeine, levorphanol, meperidine, methadone, oxymorphone, dihydrocodeine, tramadol, tapentadol, buprenorphine, cebranopadol, or pharmaceutically acceptable salts, prodrugs, and mixtures thereof. In some embodiments, at least one of the GI enzyme-labile opioid agonist-releasing subunits comprises morphine, or pharmaceutically acceptable salts thereof. In some embodiments, at least one of the GI enzyme-labile opioid agonist-releasing subunits comprises hydrocodone, or pharmaceutically acceptable salts thereof. In some embodiments, at least one of the GI enzyme-labile opioid agonist-releasing subunits comprises oxycodone, or pharmaceutically acceptable salts thereof. In some embodiments, at least one of the GI enzyme-labile opioid agonist-releasing subunits comprises codeine, or pharmaceutically acceptable salts thereof. Insome embodiments, at least one of the GI enzyme-labile opioid agonist-releasing subunits comprises levorphanol, or pharmaceutically acceptable salts thereof. In some embodiments, at least one of the GI enzyme-labile opioid agonist-releasing subunits comprises meperidine, or pharmaceutically acceptable salts thereof. In some embodiments, at least one of the GI enzyme-labile opioid agonist-releasing subunits comprises methadone, or pharmaceutically acceptable salts thereof. In some embodiments, at least one of the GI enzyme-labile opioid agonist-releasing subunits comprises methadone, or pharmaceutically acceptable salts thereof. In some embodiments, at least one of the GI enzyme-labile opioid agonist-releasing subunits comprises oxymorphone, or pharmaceutically acceptable salts thereof. In some embodiments, at least one of the GI enzyme-labile opioid agonist-releasing subunits comprises tramadol, or pharmaceutically acceptable salts thereof. In some embodiments, at least one of the GI enzyme-labile opioid agonist-releasing subunits comprises tapentadol, or pharmaceutically acceptable salts thereof.

[0048] In another aspect, provided herein is a pharmaceutical composition comprising a first opioid agonist and one or two different molecules, wherein each molecule comprises at least one Gl-enzyme-labile opioid agonist-releasing subunit comprising a second opioid agonist, wherein the at least one GI enzyme-labile opioid agonist-releasing subunit is covalently linked to at least one GI enzyme inhibitor subunit wherein the first opioid agonist reduces the onset of action time.

[0049] In some embodiments, the first opioid agonist comprises morphine, hydromorphone, hydrocodone, oxycodone, codeine, levorphanol, meperidine, methadone, oxymorphone, dihydrocodeine, tramadol, tapentadol, buprenorphine, or pharmaceutically acceptable salts, prodrugs, and mixtures thereof. In some embodiments, the first opioid agonist comprises morphine, or pharmaceutically acceptable salt thereof. In some embodiments, the first opioid agonist comprises hydromorphone, or a pharmaceutically acceptable salt thereof. In some embodiments, the first opioid agonist comprises hydrocodone, or a pharmaceutically acceptable salt thereof. In some embodiments, the first opioid agonist comprises oxycodone, or a pharmaceutically acceptable salt thereof. In some embodiments, the first opioid agonist comprises codeine, or a pharmaceutically acceptable salt thereof. In some embodiments, the first opioid agonist comprises levorphanol, or a pharmaceutically acceptable salt thereof. In some embodiments, the first opioid agonist comprises meperidine, or a pharmaceutically acceptable salt thereof. In some embodiments, the first opioid agonist comprises methadone, or a pharmaceutically acceptable salt thereof. In some embodiments, the first opioid agonist comprises oxymorphone, or apharmaceutically acceptable salt thereof. In some embodiments, the first opioid agonist comprises dihydrocodeine, or a pharmaceutically acceptable salt thereof. In some embodiments, the first opioid agonist comprises oxymorphone, or a pharmaceutically acceptable salt thereof. In some embodiments, the first opioid agonist comprises tramadol, or a pharmaceutically acceptable salt thereof. In some embodiments, the first opioid agonist comprises tapentadol, or a pharmaceutically acceptable salt thereof. In some embodiments, the first opioid agonist comprises buprenorphine, or a pharmaceutically acceptable salt thereof. In some embodiments, the first opioid agonist comprises cebranopadol, or a pharmaceutically acceptable salt thereof.

[0050] In some embodiments, the second opioid agonist is morphine, hydromorphone, hydrocodone, oxycodone, codeine, levorphanol, meperidine, methadone, oxymorphone, dihydrocodeine, tramadol, tapentadol, buprenorphine, or pharmaceutically acceptable salts, prodrugs, and mixtures thereof. In some embodiments, the second opioid agonist is morphine, hydrocodone, or oxycodone, or pharmaceutically acceptable salts, prodrugs, and mixtures thereof. In some embodiments, the second opioid agonist is hydrocodone, or pharmaceutically acceptable salts, prodrugs, and mixtures thereof.

[0051] In some embodiments, the first and / or second opioid agonist comprises between about 0.1 mg to about 20 mg in the pharmaceutical composition. In some embodiments, the first and / or second opioid agonist comprises between about 0.5 mg to about 10 mg in the pharmaceutical composition.

[0052] In some embodiments, the first opioid agonist comprises no more than 20 mg in the pharmaceutical composition. In some embodiments, the first opioid agonist comprises no more than 10 mg in the pharmaceutical composition. In some embodiments, the first opioid agonist comprises no more than 8 mg in the pharmaceutical composition. In some embodiments, the first opioid agonist comprises no more than 6 mg in the pharmaceutical composition. In some embodiments, the first opioid agonist comprises no more than 5 mg in the pharmaceutical composition. In some embodiments, the first opioid agonist comprises no more than 10 mg in the pharmaceutical composition. In some embodiments, the first opioid agonist comprises no more than 3 mg in the pharmaceutical composition, the first opioid agonist comprises no more than 2 mg in the pharmaceutical composition. In some embodiments, the first opioid agonist comprises no more than 1.5 mg in the pharmaceutical composition. In some embodiments, the first opioid agonist comprises no more than 1 mg in the pharmaceutical composition. In some embodiments, the first opioid agonist comprises no more than 0.5 mg in the pharmaceutical composition. In some embodiments, the first opioidagonist comprises no more than 0.1 mg in the pharmaceutical composition. In some embodiments, the first opioid agonist comprises no more than 0.01 mg in the pharmaceutical composition.

[0053] In some embodiments, the two different molecules are in a ratio of 1 :20 w / w to 1 : 1 w / w. In some embodiments, the two different molecules are in a ratio of 1 : 10 w / w to 1 : 1 w / w. In some embodiments, the two different molecules are in a ratio of 1 :8 w / w to 1 : 1 w / w. In some embodiments, the two different molecules are in a ratio of 1 :6 w / w to 1 : 1 w / w. In Some embodiments, the two different molecules are in a ratio of 1 :5 w / w to 1 : 1 w / w. In some embodiments, the two different molecules are in a ratio of about 1 :20 w / w. In some embodiments, the two different molecules are in a ratio of about 1 : 15 w / w. In some embodiments, the two different molecules are in a ratio of about 1 : 10 w / w. In some embodiments, the two different molecules are in a ratio of about 1 :8 w / w. In some embodiments, the two different molecules are in a ratio of about 1 :6 w / w. In some embodiments, the two different molecules are in a ratio of about 1 :5 w / w. In some embodiments, the two different molecules are in a ratio of about 1 :4 w / w. In some embodiments, the two different molecules are in a ratio of about 1 :3 w / w. In some embodiments, the two different molecules are in a ratio of about 1 :2 w / w. In some embodiments, the two different molecules are in a ratio of about 1 : 1 w / w.

[0054] In some embodiments, the pharmaceutical composition comprises about 0.1 mg to about 2000 mg of the one, two, or more different molecules. In some embodiments, the pharmaceutical composition comprises about 1 mg to about 2000 mg of the one, two, or more different molecules. In some embodiments, the pharmaceutical composition comprises about 5 mg to about 2000 mg of the one, two, or more different molecules. In some embodiments, the pharmaceutical composition comprises about 10 mg to about 1500 mg of the one, two, or more different molecules. In some embodiments, the pharmaceutical composition comprises about 20 mg to about 1250 mg of the one, two, or more different molecules. In some embodiments, the pharmaceutical composition comprises about 25 mg to about 1000 mg of the one, two, ore more different molecules.

[0055] In some embodiments, the pharmaceutical composition comprises 0.5 mg of the one, two, or more different molecules. In some embodiments, the pharmaceutical composition comprises 1 mg of the one or two different molecules. In some embodiments, the pharmaceutical composition comprises 3 mg of the one, two, or more different molecules. In some embodiments, the pharmaceutical composition comprises 5 mg of the one, two, or more different molecules. In some embodiments, the pharmaceutical composition comprises10 mg of the one, two, or more different molecules. In some embodiments, the pharmaceutical composition comprises 15 mg of the one, two, or more different molecules. In some embodiments, the pharmaceutical composition comprises 20 mg of the one, two, or more different molecules. In some embodiments, the pharmaceutical composition comprises 30 mg of the one, two, or more different molecules. In some embodiments, the pharmaceutical composition comprises 40 mg of the one, two, or more different molecules. In some embodiments, the pharmaceutical composition comprises 50 mg of the one, two, or more different molecules. In some embodiments, the pharmaceutical composition comprises 60 mg of the one, two, or more different molecules. In some embodiments, the pharmaceutical composition comprises 80 mg of the one, two, or more different molecules. In some embodiments, the pharmaceutical composition comprises 100 mg of the one, two, or more different molecules. In some embodiments, the pharmaceutical composition comprises 120 mg of the one, two, or more different molecules. In some embodiments, the pharmaceutical composition comprises 160 mg of the one, two, or more different molecules. In some embodiments, the pharmaceutical composition comprises 180 mg of the one, two, or more different molecules. In some embodiments, the pharmaceutical composition comprises 200 mg of the one, two, or more different molecules. In some embodiments, the pharmaceutical composition comprises 220 mg of the one, two, or more different molecules. In some embodiments, the pharmaceutical composition comprises 240 mg of the one, two, or more different molecules. In some embodiments, the pharmaceutical composition comprises 280 mg of the one, two, or more different molecules. In some embodiments, the pharmaceutical composition comprises 300 mg of the one, two, or more different molecules. In some embodiments, the pharmaceutical composition comprises 340 mg of the one, two, or more different molecules. In some embodiments, the pharmaceutical composition comprises 380 mg of the one, two, or more different molecules. In some embodiments, the pharmaceutical composition comprises 400 mg of the one, two, or more different molecules. In some embodiments, the pharmaceutical composition comprises 440 mg of the one, two, or more different molecules. In some embodiments, the pharmaceutical composition comprises 480 mg of the one, two, or more different molecules. In some embodiments, the pharmaceutical composition comprises 500 mg of the one, two, or more different molecules. In some embodiments, the pharmaceutical composition comprises 600 mg of the one, two, or more different molecules. In some embodiments, the pharmaceutical composition comprises 700 mg of the one, two, or more different molecules. In some embodiments, the pharmaceutical composition comprises 800 mg of the one, two, or more different molecules.In some embodiments, the pharmaceutical composition comprises 900 mg of the one, two, or more different molecules. In some embodiments, the pharmaceutical composition comprises 1000 mg of the one, two, or more different molecules. In some embodiments, the pharmaceutical composition comprises 960 mg of the one, two, or more different molecules. In some embodiments, the pharmaceutical composition comprises 1500 mg of the one, two, or more different molecule. In some embodiments, the pharmaceutical composition comprises 2000 mg of the one, two, or more different molecule.

[0056] In some embodiments, the pharmaceutical composition comprises one molecule. In some embodiments, the pharmaceutical composition comprises two different molecules. In some embodiments, the pharmaceutical composition comprises two or more different molecules.

[0057] In some embodiments, the one, two, or more different molecules are independently selected from the group consisting of a structure of Formula (IF), (IG), (IH), (II), and salts thereof:wherein:D is an opioid agonist;R1of R1-Z1- is independently selected from the group consisting of, wherein Y is amidine, guanidine, aminomethyl, substituted amidine, substituted guanidine, substituted aminomethyl, amidinomethyl, guanidinomethyl, substituted amidinomethyl, or substituted guanidinomethyl, and Q is hydrogen;R’ is selected from the group consisting of methyl, lower alkyl, substituted alkyl, aryl, substituted aryl, heteroalkyl, a natural or a non-natural amino acid, a polypeptide chain comprising natural or non-natural amino acids up to 10 amino acids in length, a linear or a branched polyethylene glycol chain up to 5 kDa, benzyloxy,R” is selected from the group consisting of acetyl, substituted acyl, a natural or a non-natural amino acid, and a polypeptide chain comprising natural or non-natural amino acids up to 10 amino acids in length;AA is a natural or a non-natural amino acid side chain that is recognized by trypsin; andZ1of R1-Z1- is independently selected from the group consisting of R1-O-CH2-CH2- and R1-CH2-C(O)-NH-CH2-CH2-.

[0058] In some embodiments, Y is amidine, aminomethyl, or guanidine.

[0059] In some embodiments, R1of R1-Z1- is independently selected from the group consisting of:

[0060] In some embodiments, the one, two, or more different molecules are independently selected from the group consisting of a structure of Formula (II- A), (II-B), (II- C), (II-D), (II-E), (II-F), (II-G), (II-H), (II-I), (II- J), (II-K), (II-L), (II-M), (II-N), (II-O), (II- P), (II-Q), (II-R), (II-S), (II-T), (II-U), (II-V), (II-W), (II-X), and salts thereof:wherein:R is selected from the group consisting ofX is hydrogen or -OH.

[0061] In some embodiments, AA is the side chain of lysine or arginine.

[0062] In some embodiments, R’ is methyl, benzyloxy,

[0063] In some embodiments, R” is acetyl, -Gly-NAc, or -Ala-NAc.

[0064] In some embodiments, the pharmaceutical composition comprises:or salts thereof; wherein: R' is independently, methyl, benzyloxy,AA is independently, the side chain of lysine or arginine;R" is independently, acetyl, -Gly-NAc, or -Ala-NAc; and X is independently, hydrogen or -OH.

[0065] In some embodiments, the pharmaceutical composition comprises:

[0066] In some embodiments, the pharmaceutical composition comprises:

[0067] In some embodiments, the pharmaceutical composition comprises:

[0068] In some embodiments, the pharmaceutical composition comprises:

[0069] In some embodiments, the two different molecules is a structure of Compound A and Compound B :the ratio of Compound A to Compound B is between 1 :6 w / w and 6: 1 w / w, and the pharmaceutical composition is administered once or twice daily.

[0070] In some embodiments, the ratio of Compound A to Compound B is between 1 : 10 w / w and 10: 1 w / w, and the pharmaceutical composition is administered once or twice daily.

[0071] In some embodiments, the ratio of Compound A to Compound B is between 1 :4 w / w and 1 : 1 w / w, and the pharmaceutical composition is administered once or twice daily.

[0072] In some embodiments, the ratio of Compound A to Compound B is 1 : 10 w / w. In some embodiments, the ratio of Compound A to Compound B is 1 :8 w / w. In some embodiments, the ratio of Compound A to Compound B is 1 :6 w / w. In some embodiments, the ratio of Compound A to Compound B is 1 :5 w / w. In some embodiments, the ratio of Compound A to Compound B is 1 :4 w / w. In some embodiments, the ratio of Compound A toCompound B is 1 :3 w / w. In some embodiments, the ratio of Compound A to Compound B is 1 :2 w / w. In some embodiments, the ratio of Compound A to Compound B is 1 : 1 w / w. In some embodiments, the ratio of Compound A to Compound B is 2: 1 w / w. In some embodiments, the ratio of Compound A to Compound B is 3 : 1 w / w. In some embodiments, the ratio of Compound A to Compound B is 4: 1 w / w. In some embodiments, the ratio of Compound A to Compound B is 5: 1 w / w. In some embodiments, the ratio of Compound A to Compound B is 6: 1 w / w. In some embodiments, the ratio of Compound A to Compound B is 8: 1 w / w. In some embodiments, the ratio of Compound A to Compound B is 10: 1 w / w.

[0073] In some embodiments, the pharmaceutical composition comprises two different molecules, wherein each molecule comprises at least one GI enzyme-labile opioid agonistreleasing subunit comprising an opioid agonist, wherein the at least one GI enzyme-labile opioid agonist-releasing subunit is covalently linked to at least one GI enzyme-inhibitor subunit, wherein the two different molecules are in a ratio of 1 : 10 w / w to 1 : 1 w / w.

[0074] In some embodiments, the two different molecules are a first GI enzyme-labile opioid agonist-releasing subunit comprising a first Gl-enzyme inhibitor subunit and a first opioid agonist; and a second GI enzyme-labile opioid agonist-releasing subunit comprising a second Gl-enzyme inhibitor subunit and a second opioid agonist.

[0075] In some embodiments, the first GI enzyme-labile opioid agonist-releasing subunit and the second GI enzyme-labile opioid agonist-releasing subunit are in a ratio of 1 : 10 w / w. In some embodiments, the first GI enzyme-labile opioid agonist-releasing subunit and the second GI enzyme-labile opioid agonist-releasing subunit are in a ratio of 1 :4 w / w. In some embodiments, the first GI enzyme-labile opioid agonist-releasing subunit and the second GI enzyme-labile opioid agonist-releasing subunit are in a ratio of 1 : 1 w / w.

[0076] In some embodiments, the pharmaceutical composition is administered no more than once daily. In some embodiments, the pharmaceutical composition is administered once daily. In some embodiments, the pharmaceutical composition is administered twice daily.

[0077] In some embodiments, the disclosure provides for compositions containing two or more polysubunit molecules that comprise subunits that interact with gastrointestinal (GI) or digestive enzymes. In some cases, such a composition can be specifically hydrolyzed by at least one of any of the GI enzymes disclosed herein. The GI enzyme can be, for example, pepsin, trypsin, chymotrypsin, colipase, elastase, aminopeptidases, dipeptidylaminopeptidase IV, tripeptidase, enteropeptidases, carboxypeptidases, dipeptidal aminopeptidases, pteroyl polyglutamate hydrolyase, gamma-glutamyl transferase, aminoaspartate aminopeptidases, amino-oligopeptidase, membrane Gly-Leu peptidase, and zinc stable Asp-Lys peptidase.

[0078] An example of a GI enzyme subunit is a protease substrate, such as a trypsin substrate, or a chymotrypsin substrate. In some embodiments,

[0079] As used herein, the term “trypsin substrate” refers to any agent capable of being hydrolyzed by trypsin, and includes salts of trypsin substrates. The ability of an agent to be a substrate for trypsin can be measured using assays well known in the art. For example, in a typical assay, one can directly measure the rate and extent of expected hydrolysis products formed in incubations containing specified concentrations of digestive enzymes and enzyme substrates using common HPLC or spectrophotometric detection methods.

[0080] There are many trypsin substrates known in the art, and include substrates that are specific to trypsin and those that are specific to other proteases such as chymotrypsin.Trypsin substrates include natural, synthetic, and semi-synthetic compounds. The disclosure provides for trypsin substrates that are proteins, peptides, and small molecules. The disclosure also provides for trypsin substrates that are hydrolyzed via “normal” or “inverse” substrate mechanisms. A trypsin substrate can be an arginine mimic or lysine mimic. In certain embodiments, the trypsin substrate is an arginine mimic or a lysine mimic, wherein the arginine mimic or lysine mimic is a synthetic compound. As used herein, an arginine mimic or lysine mimic can include moi eties capable of being recognized by, and binding to, the specificity pocket of trypsin and / or interacting with the trypsin active site functionalities. The arginine or lysine mimic can comprise a cleavable moiety. In some embodiments, cleavage of the cleavable moiety will directly, or indirectly result in release of an opioid agonist from the substrate moiety. In some cases, when supra-therapeutic doses (overdoses) are ingested, the presence of GI enzyme inhibitor subunits can saturate or inhibit the capacity of the enzyme to cleave the cleavable moieties that directly, or indirectly, release opioid agonists resulting in overdose protection.

[0081] Examples of trypsin substrates which are arginine mimics and / or lysine mimics include a cationic specificity pocket binding moiety designed to be recognized by, and bind to, the negatively charged specificity pocket of the enzyme, and a hydrolyzable functionality that is cleaved by the active site of the enzyme. Cationic specificity pocket binding moieties include, but are not limited to, alkyl-amines, alkylguanidines, alkylamidines, arylguanidines, benzamidines, benzylamines, naphthylamidines, naphthylguanidines, naphthylamines, and the like. Hydrolyzable functionalities include, but are not limited to, amide, ester, carbamate, thioester, carbonate, and the like.

[0082] In one aspect of the invention, the opioid agonist releasing GI enzyme substrate subunit(s) and the GI enzyme inhibitor subunit(s) are covalently linked via a covalent bond,an atom, or via a scaffold, for example a polymeric, oligomeric, or molecular scaffold. The opioid releasing and non-opioid releasing subunits can be linked directly, or indirectly, via a wide range of atoms or linkers as described herein. The particular linkages and linkage chemistries employed will depend upon the specific functional groups available on the opioid releasing and the enzyme inhibiting subunits, and the available complimentary functional groups present on the linker or scaffold moiety components. The presence of suitable functional groups within the opioid releasing and enzyme inhibiting subunits, and scaffold moiety components, and useful chemistry for linking strategies involving these suitable functional groups can be readily determined by one skilled in the art based upon the guidance presented herein. Particular examples of unimolecular compositions comprised of linkers (Z), opioid releasing subunits, enzyme inhibiting subunits, and opioid antagonist subunits, and atomic, molecular, oligomeric or polymeric scaffolds are disclosed herein.

[0083] Compositions of the invention are not required to have, and preferably do not have opioid agonist activity. Thus, in one aspect of the invention, a composition in accordance with the invention will retain from about 0% to about 30% of the specific agonist activity of the delivered opioid agonist compound. Such activity may be determined using suitable in- vivo, or in-vitro functional assays, depending upon the known activity of the particular opioid parent compound. For example, a functional opioid receptor-based assay, or an in vivo hotplate or tail-flick analgesia assay can be used to assess the level of agonist activity of the polymer conjugates of the invention. Thus, compositions of the invention will possess a specific activity of about 0% or less than about 0.25%, 0.5%, 0.75%, 1% 2%, 5%, 10%, 15%, 25%, 30% or 50% relative to that of the delivered opioid agonist, when measured in a suitable model, such as those well known in the art.

[0084] In another aspect of the invention, compositions of the invention are not required to be, and preferably are not, able to efficiently traverse the blood-brain barrier and gain access to the central nervous system (CNS). For example, compositions of the invention may not efficiently penetrate the central nervous system. Thus, in one aspect of the invention, a composition in accordance with the invention will retain from about 0% to about 30% of the CNS penetration of the delivered opioid agonist. CNS penetration can be determined using suitable in-vivo assays. Thus, a composition described herein will possess CNS penetration of about 0% or less than about 0.25%, 0.5%, 0.75%, 1% 2%, 5%, 10%, 15%, 25%, or 30% relative to that of the unmodified parent opioid, when measured in a suitable model, such as those well known in the art.

[0085] In another aspect, the present disclosure provides a compound represented by the structure of Formula (I):or a salt thereof, wherein: each R1is independently a GI enzyme inhibitor subunit; each R2is independently a GI enzyme-labile opioid agonist releasing subunit; each R3is independently an opioid antagonist releasing subunit;M is, an atom, or a scaffold moiety; each Z1, Z2, and Z3is independently absent or a linking moiety; each h, k, g, d, and e is independently an integer ranging from 1 to 10, 1 to 100, 1 to 1,000, 1 to 100,000, 1 to 1,000,000, or 1 to 1,000,000,000; and q is an integer ranging from 0 to 10, 0 to 100, 0 to 1,000, 0 to 100,000, 0 to 1,000,000, or 0 to 1,000,000,000.

[0086] In some embodiments, for the compound or salt of Formula (I), M is an optionally substituted heteroalkyl group.

[0087] In some embodiments, for the compound or salt of Formula (I), M is an optionally substituted peptide.

[0088] In some embodiments, for the compound or salt of Formula (I), M is an optionally substituted peptide with from 1 to 500 amino acids. In some embodiments, for the compound or salt of Formula (I), M is an optionally substituted peptide with from 1 to 50 amino acids.In some embodiments, for the compound or salt of Formula (I), M is an optionally substituted peptide with from 1 to 10 amino acids. In some embodiments, for the compound or salt of Formula (I), M is an optionally substituted peptide with from 1 to 3 amino acids.

[0089] In some embodiments, a compound or salt of Formula (I) is represented by a structure of Formula (IA), (IB), (IC), or (ID):wherein:M is an atom, a functional group, a substituted functional group, or a molecular scaffold; andW is selected from hydrogen, optionally substituted alkyl, optionally substituted acyl, and optionally substituted alkoxycarbonyl, or

[0090] In some embodiments, the compound or salt, wherein R1is independently selected at each occurrence from a GI enzyme inhibitor. In some embodiments, for the compound or salt of Formula (I), R1at each occurrence is a serine protease inhibitor. In some embodiments, for the compound or salt of Formula (I), R1at each occurrence is a trypsin inhibitor.

[0091] In some embodiments, for the compound or salt of Formula (I), each R1is independently selected from the group consisting of:wherein:Y is amidine, guanidine, aminomethyl, substituted amidine, substituted guanidine, substituted aminomethyl, amidinomethyl, guanidinomethyl, substituted amidinomethyl, or substituted guanidinomethyl; andQ is independently selected from hydrogen, cyano, nitro, halogen, alkyl and alkoxy.

[0092] In some embodiments, for the compound or salt of Formula (I), R1— Z1— is represented by the formula:

[0093] In some embodiments, for the compound or salt of Formula (I), Z1at each occurrence is selected from a cleavable or non-cleavable linker including from 2 to 15 atoms.

[0094] In some embodiments, for the compound or salt of Formula (I), Z1is -O-CH2- CH2-NH- or -O-CH2-CH2-O-.

[0095] In some embodiments, for the compound or salt of Formula (I), g, d, or e is independently selected from 1 to 20. In some embodiments, for the compound or salt of Formula (I), g, d, or e is independently selected from 1 to 10. In some embodiments, for the compound or salt of Formula (I), g, d, or e is independently selected from 1 to 3.

[0096] In some embodiments, for the compound or salt of Formula (I), R2- is independently selected at each occurrence from:wherein:D is an opioid agonist;R101is independently selected from an amino acid side chain, or an amino acid sidechain mimic that is recognized by a GI enzyme; andR102is independently selected from hydrogen, alkyl, aryl, substituted alkyl, substituted aryl, heteroalkyl, substituted heteroalkyl, acyl, substituted acyl group,polyethylene glycol containing acyl, polyethylene glycol containing moiety, or a natural or unnatural amino acid, or an amino acid mimic.

[0097] In some embodiments, for the compound or salt of Formula (I), R101is selected from an amino acid side chain and R102is optionally substituted alkyl. In some embodiments, for the compound or salt of Formula (I), R101is selected from an arginine or lysine side chain and R102is optionally substituted methyl. In some embodiments, for the compound or salt of Formula (I), R102is methyl substituted with -NH-acetyl, or dimethyl substituted with -NH- acetyl.

[0098] In some embodiments, linking moieties Z1, Z2, and Z3are independently represented by the general formulae:and can also be defined by

[0099] Exemplary terminal linker functionalities “F” can each or independently be as shown below:where “L” is shown in the structures of “F”, illustrated above, to indicate the connectivity of F and L, and wherein: each R is independently hydrogen, methyl, lower alkyl, aryl, or arylalkyl;X is carbon, oxygen, or nitrogen; andL is a linear, branched, or multivalent scaffold which is alkyl, aryl, substituted alkyl, substituted aryl, heteroalkyl, substituted heteroalkyl, polyalkylene glycol, polypeptide, polyamide, polycarbamate, polyurea, or polycarbonate.

[0100] In some embodiments, L is formed of 0-100 atoms. In some embodiments, L is formed of 1-50 non-hydrogen atoms as well as additional hydrogen atoms. Such atoms may be, for example, C, N, O, P or S. In other embodiments, L may connect two or more groups comprising 1 to 50 consecutive bonds between the groups. L may have 1 to 40, 1 to 30, 1 to 20, 1 to 10, 1 to 5, 5 to 25, or 5 to 20 such consecutive bonds.

[0101] In some embodiments, for the compound or salt of Formula (I), D is selected from morphine, hydromorphone, hydrocodone, oxycodone, codeine, levorphanol, meperidine, methadone, oxymorphone, dihydrocodeine, tramadol, tapentadol, and buprenorphine. In some embodiments, for the compound or salt of Formula (I), D is represented by the formula:

[0102] In some embodiments, for the compound or salt of Formula (I), k is selected from 1 to 20. In some embodiments, for the compound or salt of Formula (I), k is selected from 1 to 10. In some embodiments, for the compound or salt of Formula (I), k is 1 to 3.

[0103] In some embodiments, the present disclosure provides a method of treating pain in a subject in need thereof, the method comprising administrating to the subject a therapeutically effective amount of two or more compound(s) or salt(s) of Formula (I).

[0104] In some embodiments, the present disclosure provides two or more compound(s) or salt(s) of Formula (I) and one or more pharmaceutically acceptable excipient(s).

[0105] In some embodiments, the present disclosure provides a pharmaceutical formulation comprising two or more polysubunit molecules with each molecule comprising: an opioid prodrug; a gastrointestinal enzyme inhibitor; and a scaffold moiety, wherein the opioid prodrug and the inhibitor are covalently attached to the scaffold moiety.

[0106] In some embodiments, the present disclosure provides a pharmaceutical composition, the composition comprising: two or more polysubunit molecules each comprising GI enzyme labile opioid releasing subunit(s), and GI enzyme inhibitor subunit(s) wherein the GI enzyme labile opioid releasing subunit(s) and the GI enzyme inhibitor subunit(s) are covalently linked via a covalent bond, an atom, or a scaffold.

[0107] In some embodiments the disclosure provides for polysubunit compounds comprising one GI enzyme labile opioid releasing subunit covalently linked to one GI enzyme inhibitor subunit represented by formulae II(A-X) below:wherein:R is selected from the group consisting of :R’ can be methyl, lower alkyl, substituted alkyl, aryl, substituted aryl, heteroalkyl, a natural or non-natural amino acid, a polypeptide chain comprising natural and / or nonnatural amino acids up to 10 amino acids in length, a linear or branched polyethylene glycol chain up to 5 kDa, benzyloxy, and the like; R” can be an acetyl, substituted acyl, a natural or non-natural amino acid, or a polypeptide chain comprising natural and / or non-natural amino acids up to 10 amino acids in length; AA is a natural or non- natural amino acid side chain capable of being recognized by trypsin; and X is hydrogen or OH.

[0108] In some embodiments, the disclosure provides for compositions comprising two or more of the aforementioned compounds of Formula II wherein:R can be R’ can bemethyl or benzyloxy; R” can be an acetyl or a substituted acyl, a natural or nonnatural amino acid or a di- or tri-peptide comprising natural or non-natural aminoacids; AA is a natural or non-natural amino acid side chain capable of being recognized by trypsin; and X is hydrogen or OH.

[0109] In some embodiments, the disclosure provides for compositions comprising two or more of the aforementioned compounds of Formula II wherein: R can beR' can be methyl or benzyloxy; R” can be an acetyl, a natural or non-natural amino acid or a dipeptide comprising natural or non-natural amino acids; AA is the side chain of lysine or arginine; and X is hydrogen or OH.

[0110] In some embodiments, the disclosure provides for compositions comprising two or more of the aforementioned compounds of Formula II wherein: R can be R'can be methyl or benzyloxy; R” can be acetyl, -Ala-NAc or -Gly-NAc; AA is the side chain of lysine or arginine; and X is hydrogen or OH.

[0111] Table la illustrates various hydrocodone, hydromorphone, oxycodone, and oxymorphone containing compounds of Formula II- A, II-B, II-G, and II-H contemplated by the present disclosure.

[0112] Table 2a illustrates various morphine containing compounds of Formula II-C and II-I contemplated by the present disclosure.

[0113] Table 3a illustrates various hydrocodone, hydromorphone, oxycodone, and oxymorphone containing compounds of Formula II-D, II-E, II- J, and II-K contemplated by the present disclosure.

[0114] Table 4a illustrates various morphine containing compounds of Formula II-F and II-L contemplated by the present disclosure.

[0115] In one aspect, the present disclosure provides a pharmaceutical composition, the composition comprising: one, two, or more polysubunit molecules each comprising GI enzyme labile opioid releasing subunit(s), and gastrointestinal enzyme inhibitor subunit(s) wherein the GI enzyme labile opioid releasing subunit(s) and the GI enzyme inhibitor subunit(s) are covalently linked via a covalent bond, an atom, or a scaffold moiety.

[0116] In some embodiments the disclosure provides for polysubunit compounds comprising one GI enzyme labile opioid releasing subunit covalently linked to one GI enzyme inhibitor subunit represented by formulae III(A-L) below:wherein: R can be R'can be methyl, lower alkyl, substituted alkyl, aryl, substituted aryl, heteroalkyl, a natural or non-natural amino acid, a polypeptide chain comprising natural and / or nonnatural amino acids up to 10 amino acids in length, a linear or branched polyethylene glycol chain up to 5 kDa, benzyloxy, and the like; R” can be an acetyl, substitutedacyl, a natural or non-natural amino acid, or a polypeptide chain comprising natural and / or non-natural amino acids up to 10 amino acids in length; AA is a natural or nonnatural amino acid side chain recognized by trypsin; and X is hydrogen or OH.

[0117] In some embodiments the disclosure provides for compositions comprising one, two, or more of the aforementioned compounds of Formula II wherein: R can be R'can be methyl or benzyloxy; R” can be an acetyl or a substituted acyl, a natural or non-natural amino acid or a di- or tri-peptide comprising natural or non-natural amino acids; AA is a natural or non-natural amino acid side chain recognized by trypsin; and X is hydrogen or OH.

[0118] In yet other embodiments the disclosure provides one, two, or more of the aforementioned compounds of Formula II wherein: R can beR' can be methyl or benzyloxy; R” can be an acetyl, a natural or non-natural amino acid or a dipeptide comprising natural or non-natural amino acids; AA is the side chain of lysine or arginine; and X is hydrogen or OH.

[0119] In yet another embodiment the disclosure provides one or more of the aforementioned compounds of Formula II wherein: R can be R' can bemethyl or benzyloxy; R” can be acetyl, -Ala-NAc or -Gly-NAc; AA is the side chain of lysine or arginine; and X is hydrogen or OH.Methods

[0120] In some embodiments, provided herein is a method of treating pain in a subject, comprising administering to the subject (e.g., a therapeutically effective amount of) the pharmaceutical composition described herein. In some embodiments, the pharmaceutical composition comprises a compound represented by the structure of Formula (I). In some embodiments, the pharmaceutical composition comprises Compound A and / or Compound B. In some embodiments, the pharmaceutical composition comprises Compound A. In some embodiments, the pharmaceutical composition comprises Compound B.

[0121] In some embodiments, provided herein is a method of treating pain in a subject, the method comprising administering to the subject a pharmaceutical composition comprising two different molecules, wherein each molecule comprises at least one GI enzyme-labile opioid agonist-releasing subunit comprising an opioid agonist, wherein the at least one GI enzyme-labile opioid agonist-releasing subunit is covalently linked to at least one GI enzymeinhibitor subunit, wherein the GI enzyme-labile opioid agonist-releasing subunit releases the opioid agonist, and the opioid agonist achieves a mean max plasma concentration after about 6 hours. In some embodiments, wherein the opioid agonist achieves a mean max plasma concentration after about 6 hours to about 10 hours. In some embodiments, the opioid agonist achieves a mean max plasma concentration after about 6 hours to about 8 hours. In some embodiments, the opioid agonist achieves a mean max plasma concentration after about 8 hours. In some embodiments, the opioid agonist achieves a mean max plasma concentration after about 10 hours.

[0122] In some embodiments, provided herein is a method of treating pain in a subject, the method comprising administering to the subject a therapeutically effective amount of a pharmaceutical composition comprising two different molecules, wherein each molecule comprises at least one GI enzyme-labile opioid agonist-releasing subunit comprising an opioid agonist, wherein the at least one GI enzyme-labile opioid agonist-releasing subunit is covalently linked to at least one GI enzyme-inhibitor subunit, wherein the two different molecules are in a ratio of 1 :4 w / w to 1 : 1 w / w, and wherein the pharmaceutical composition is administered once daily.

[0123] In some embodiments, the pain is acute pain. In some embodiments, the pain is chronic pain.

[0124] In some embodiments, the pharmaceutical composition described herein is administered to in an amount sufficient to provide a Cmaxof the free opioid agonist between 5 and 11 hours. In some embodiments, the Cmaxof the free opioid agonist is reached between 5 and 11.

[0125] In some embodiments, the pharmaceutical composition described herein provides opioid agonist plasma concentration within the subject is below the lethal amount. In some embodiments, the pharmaceutical composition provides opioid agonist plasma concentration within the subject is below the lethal amount in a supra-therapeutic dosage. In some embodiments, the hydrocodone plasma concentration in the subject is lower than about 110 ng / mL.

[0126] In some embodiments, an opioid agonist plasma concentration within the subject is not dose-proportional.

[0127] In some embodiments, an opioid agonist plasma concentration in the subject demonstrates reduced dose proportionality when compared to the free opioid agonist delivered by the one or two different molecules.

[0128] In some embodiments, the administration of a therapeutically effective amount of a pharmaceutical composition described herein reduces events of abuse, misuse, and misdirection, when compared to the administration of the free opioid agonist.

[0129] In another aspect, provided herein is a method of preventing opioid abuse or treating opioid overdose in a subject, comprising administering to the subject a therapeutically effective amount of a pharmaceutical composition described herein.

[0130] In some embodiments, the subject is experiencing symptoms from overdose of an opioid agonist. In some embodiments, the subject is suffering from opioid use disorder. In some embodiments, the subject is suffering from alcohol use disorder. In some embodiments, the subject is suffering from an impulse control disorder.Definitions

[0131] Unless defined otherwise, all terms of art, notations and other technical and scientific terms or terminology used herein are intended to have the same meaning as is commonly understood by one of ordinary skill in the art to which the claimed subject matter pertains. In some cases, terms with commonly understood meanings are defined herein for clarity and / or for ready reference, and the inclusion of such definitions herein should not necessarily be construed to represent a substantial difference over what is generally understood in the art.

[0132] Throughout this application, various embodiments may be presented in a range format. It should be understood that the description in range format is merely for convenience and brevity and should not be construed as an inflexible limitation on the scope of the disclosure. Accordingly, the description of a range should be considered to have specifically disclosed all the possible subranges as well as individual numerical values within that range. For example, description of a range such as from 1 to 6 should be considered to have specifically disclosed subranges such as from 1 to 3, from 1 to 4, from 1 to 5, from 2 to 4, from 2 to 6, from 3 to 6 etc., as well as individual numbers within that range, for example, 1, 2, 3, 4, 5, and 6. This applies regardless of the breadth of the range.

[0133] As used in the specification and claims, the singular forms “a”, “an” and “the” include plural references unless the context clearly dictates otherwise. For example, the term “a sample” includes a plurality of samples, including mixtures thereof.

[0134] As used herein, the term “trypsin substrate” refers to any agent capable of being hydrolyzed by trypsin, and includes salts of trypsin substrates. The ability of an agent to be a substrate for trypsin can be measured using assays well known in the art. For example, in a typical assay, one can directly measure the rate and extent of expected hydrolysis products formed in incubations containing specified concentrations of digestive enzymes and enzyme substrates using common HPLC or spectrophotometric detection methods.

[0135] As used herein, the term “supra-therapeutic” refers to any dose that is greater than required to achieve the intended therapeutic effect, such as, e.g., a dose of an opioid agonist that is greater than required to achieve analgesia.

[0136] As used herein, the term “Cmax” refers to the maximum concentration that a compound achieves in the plasma after the compound has been administered. Cmaxmay be referenced either as an amount (e.g., 110 ng / mL) or a time (e.g., 6-8 hours), wherein the amount refers to the peak concentration achieved and the time refers to the time at which the maximum plasma concentration has been reached upon administration.

[0137] As used herein, the term “enzyme-labile” refers to a structure, compound, bond, or functional group, that is susceptible to reactivity with an enzyme. Reactivity can include, for example, alteration, cleavage, addition, or substitution. Additionally, the reactivity can include reversible and irreversible reactivity.

[0138] As used herein, the term “agonist-releasing” refers to a composition, mixture, compound, or structure that delivers an agonist upon interaction with an appropriate substrate, such as, e.g., the delivery of an opioid agonist upon the interaction of a trypsin- labile opioid agonist-releasing subunit with trypsin.

[0139] As used herein, the terms “abuse,” “misuse,” and “diversion” refer to consumption of a therapeutic agent in a manner that differs from the guidance provided by a healthcare professional. Examples of “abuse” and “misuse” include: consuming therapeutic agents in a larger dose than prescribed, consuming therapeutic agents in a smaller dose than prescribed, consuming therapeutic agents in a higher frequency than prescribed, consuming the therapeutic agent in a lower frequency than prescribed, and administering the therapeutic agent through a route that differs from what was prescribed, among others. An example of “diversion” includes: consuming a therapeutic agent that was prescribed to another.

[0140] The term “dose proportionality” refers to the relationship between the administered dose and the resulting exposures (e.g., Cmaxor AUC) of a drug. If a given drugs exposure can be linearly extrapolated based on the administered dose (i.e., dose-dependent exposures), then the drug is described as dose-proportional. There are several mathematical treatments used to determine dose proportionality including directly comparing the dose exposure relationship of several doses to confirm that the values for exposure (e.g., Cmax, AUC) divided by dose over a range of doses are equivalent:or via implementation of a power model:whereby beta (the slope) measures the proportionality between the dose and resulting PK exposure Y. If beta equals 1.0 then the drug is demonstrated to be dose proportional.

[0141] The term “dose proportionality” refers to a situation when a drug exhibits a proportional increase in the amount of drug reaching the systemic circulation.

[0142] The section headings used herein are for organizational purposes only and are not to be construed as limiting the subject matter described.EXAMPLES

[0143] The following illustrative examples are representative of embodiments of the stimulation, systems, and methods described herein and are not meant to be limiting in any wayExample 1. Chemical Structures

[0144] The structures of ETR028 (para-isomer) and ETR029 (meta isomer) are provided in FIG. 1. An expanded structure of ETR028 and ETR029 depicting their key structural components is provided in FIG. 2. These key structural components include A) the delivered opioid (hydrocodone). Note that hydrocodone can be exchanged for other opioid agonists (e.g., oxycodone, oxymorphone, hydromorphone, morphine, methadone, buprenorphine, and any suitably functionalized drug of choice); B) the opioid delivery subunit that is activated by the digestive enzyme trypsin in the lumen of the small intestine. The opioid delivery subunit has been designed and demonstrated to completely release the appended opioid (e.g. hydrocodone) within 10 minutes upon exposure to active trypsin in vitro., C) the regio- isomeric trypsin inhibitor subunit. Importantly, the para-isomer present in ETR028 isapproximately lOOx more potent than the meta-isomer present in ETR029; D) the lysinederived linker subunit that conjoins the opioid delivery subunit and the trypsin inhibitor subunit. This enables “blending” of ETR028 with ETR029 to modulate the overall trypsin inhibition potency.

[0145] The molecular mechanisms for the trypsin mediated release of hydrocodone from ETR028 and ETR029 are identical and are depicted in Scheme 1. This mechanism involves the initial recognition and cleavage of the C-terminus of the Arg-Gly-NAc residue by the digestive enzyme trypsin (arrow a) in the lumen of the small intestine to release Arg-Gly- NAc and form an aninomethylpiperidine intermediate. The aninomethylpiperidine intermediate then undergoes a spontaneous cyclization-release reaction (arrow b) that releases hydrocodone and forms a cyclic urea product in the lumen of the small intestine. The released hydrocodone is subsequently absorbed into the systemic circulation where is exerts its desired analgesic effects.

[0146] When exposed to active human trypsin in vitro, the release of hydrocodone from the opioid delivery subunit present in ETR028 and ETR029 is complete within 10 minutes.

[0147] Scheme 1: The Molecular Mechanism for the Release of Hydrocodone from ETR028 and ETR029Example 2. Human Studies

[0148] A phase 1, randomized, open-label, 2-part study to evaluate the safety and pharmacokinetics of oral O2P Hydrocodone vs. hydrocodone bitartrate (HCBT) in healthy adult subjects under fasted conditions with naltrexone blockade (O2P-001) was studied. A total of 93 subjects were enrolled in the study. Key objectives of the study were to:

[0149] Evaluate the safety, tolerability, and pharmacokinetics of oral O2P Hydrocodone relative to a HCBT comparator following single oral doses in healthy adult subjects.

[0150] Demonstrate reduced maximum plasma exposures (Cmax) of hydrocodone delivered from escalated oral doses of O2P Hydrocodone relative to an escalated comparator dose of HCBT (i.e., oral overdose protection).

[0151] Identify appropriate doses of oral O2P Hydrocodone for subsequent clinical studies.POPULATION:

[0152] The population for this study included male and female subjects, 18 to 55 years of age, inclusive, who were in general good health based on medical history, physical examination, vital signs, laboratory profile (serum chemistry, hematology, and urinalysis), and a 12-lead electrocardiogram (ECG). Subjects had a body mass index of 18 kg / m2to 32 kg / m2 (> 45 kg), inclusive, at the Screening Visit.STUDY DESIGN:

[0153] 93 healthy adult subjects were enrolled for the study with each subject participating in 1 treatment period. Six subjects were included in each treatment period. Safety and tolerability were evaluated throughout the study. Safety assessments included incidence of adverse events (AEs), physical examination findings, vital signs, electrocardiogram (ECG) parameters, and clinical laboratory tests.

[0154] A follow-up visit occurred approximately 7 days after the last treatment period dosing. Subjects were administered one 50 mg naltrexone tablet 12 hours pre-dose, 1 hour pre-dose, and 12 hours after dosing for each treatment period. A provisional PK and safety review (PPSR) was performed following the completion of all treatment periods prior to initiating subsequent treatment periods.

[0155] Blood sampling and subsequent PK analyses were also performed during the study. For each treatment period PK samples were collected at pre-dose and at 0.5, 1, 1.5, 2, 2.5, 3, 4, 6, 8, 10, 12, 24, 36, and 48 hours post-dose. The LLOQ for hydrocodone was established at 0.050 ng / mL.

[0156] Unexpected correlation between trypsin inhibition and duration of action and unexpected synergistic aspects of dosing regimen. The crux of why the pharmacokinetic profiles demonstrated by ETR028 alone, ETR029 alone, and the various blends studied in humans were “unexpected” is based on the fact that there is no ability to predict how the trypsin inhibition potencies of various doses of ETR028 alone, ETR029 alone, and the blend ratios and doses studied would affect the pharmacokinetics of delivered hydrocodone in the (i) presence of endogenous trypsin in humans, and (ii) in the context of the unique anatomical structure of the human gastrointestinal tract.Onset time

[0157] Onset time relates to the time required for a patient to perceive pain relief (i.e., analgesic effects) following an oral dose of an opioid agonist (e.g., hydrocodone). This canoccur as early as when plasma concentrations reach 25% of their maximum level (T25), or more commonly when plasma concentrations reach 50% of their maximum level (T50). When opioid agonists are dosed properly it is not necessary to achieve the maximum plasma concentration (Cmax) for the patient to experience pain relief. Onset time as used herein refers to time following an oral dose for a patient to experience pain relief. The plasma concentrations required for a patient to experience pain relief will vary depending on the type and severity of their pain and their prescribed opioid dose. Patients who are prescribed a 5 mg dose of hydrocodone will achieve a maximum plasma concentration (Cmax) of ~12 ng / mL at approximately 1.5 hours post-dose (Tmax). The T25and T50values will occur at 15 and 30 minutes, respectively. The plasma concentrations at T25and T50will be approximately 3 and 6 ng / mL, respectively.ETR028 and ETR029 alone

[0158] The PK profiles of released hydrocodone for escalating doses of ETR028 and ETR029 alone are presented in FIG. 3.

[0159] ETR028 demonstrated a slow onset time (i.e., the rate of appearance of hydrocodone in the plasma). This is likely due to the unexpectedly strong inhibition of ETR028 in the presence of endogenous human trypsin levels. Of particular importance was the unexpected inability to increase the early hydrocodone plasma exposures by escalating the dose of ETR028 alone.

[0160] ETR029 demonstrated a more rapid onset time than ETR028 due to its reduced trypsin inhibition potency. Further, hydrocodone exposures resulting from escalating doses of ETR029 were demonstrated to be dose proportional enabling the titration of therapeutic doses.

[0161] Human PK parameters for HCBT, ETR028, ETR029 alone are presented in Table 5.Table 5. Human PK parameters for HCBT Comparator, ETR028, ETR029, andETR028 / ETR028 BlendsETR028 / ETR029 Blends

[0162] The potential for decreasing the onset time (i.e., increasing the rate of appearance of hydrocodone in the plasma) and decreasing the Tmaxof ETR028 by blending ETR028 with ETR029 was also demonstrated in humans. The ability of various [ETR028 + ETR029] blend ratios and doses to alter the onset time, Tmax, and total exposures (i.e., area under the curve - AUC0-t) of hydrocodone in humans is depicted in Figure 4. Human PK parameters for the {ETR028 + ETR029} blends depicted in FIG. 4 are presented in Table 5.Duration of Action

[0163] When dosed to achieve comparable Cmaxexposure values - ETR028, ETR029, and the [ETR028 + ETR029] blends studied demonstrated longer plasma exposure times (i.e., greater AUC0-tvalues) than the commonly prescribed 5 and 10 mg dose strengths of a shortacting hydrocodone bitartrate (HCBT) comparator (see Table 5). This result was unexpected and was not observed in preclinical studies in rats and dogs. The pharmacokinetic profile of HCBT presented in FIG. 5 is representative of the hydrocodone pharmacokinetic profile demonstrated by currently marketed hydrocodone containing drug products indicated for the treatment of moderate to severe pain.

[0164] This increased exposure time extends the duration of analgesic action following an oral dose. The comparative pharmacokinetic profiles for the 30 mg + 30 mg and 60 mg ± 60 mg [ETR028 + ETR029] blends and the 5 and 10 mg HCBT are presented in FIG. 5.

[0165] The pharmacokinetic profile of [ETR028 + ETR029] blends support a once or twice daily dosing regimen. This is in stark contrast to the prescribed dosing regimen forcommonly prescribed hydrocodone containing drug products that entail more frequent dosing (i.e., one to two 5 mg tablets every 4-6 hours, or one 10 mg tablet every 4-6 hours to maintain effective analgesia).

[0166] The most prevalent mode of abuse of short-acting prescription opioids is via the oral route. Ingestion of supratherapeutic doses can result in a euphoric experience that can reinforce further abuse and addiction, or cause death.

[0167] ETR028 and ETR029 are bifunctional hydrocodone prodrug molecules that comprise (i) a trypsin-activated opioid delivery subunit that efficiently releases therapeutic levels of hydrocodone only when exposed to the digestive enzyme trypsin in the lumen of the small intestine, and (ii) a trypsin inhibitor subunit that progressively inhibits trypsin, and thereby the trypsin-mediated release of hydrocodone, when supratherapeutic doses are ingested. This unique mechanism has been demonstrated to provide unprecedented oral overdose protection in humans.

[0168] Escalated doses of a [30 mg ETR028 + ETR029] blend were evaluated to assess the ability to attenuate hydrocodone exposures when supratherapeutic doses are ingested. The escalated doses of the 1 : 1 ratio ETR028 / 029 included 30 / 30 mg (single dose), 60 / 60 mg (2- fold dose), 120 / 120 mg (4-fold dose), 240 / 240 mg (8-fold dose), and 480 / 480 mg (16-fold dose). The pharmacokinetic profile (e.g., Cmax) of the escalated doses of the 1 : 1 ratio ETR028ZETR029 blend compared to Cmaxexposures extrapolated based on linear dose proportionality are presented in FIG. 6.

[0169] The 30 / 30 mg ETR029ZETR028 blend demonstrated reduced Cmaxexposures at the 4-fold, 8-fold, and 16-fold doses of 19%, 58%, and 60%, vs. the respective extrapolated dose proportional exposures.

[0170] Importantly, the single and 2-fold doses were dose proportional enabling titration of therapeutic exposures with the prescribed doses.

[0171] In addition, escalated doses of 5 mg (single dose), 10 mg (2 -fold dose), and 80 mg (16-fold dose) of HCBT were also evaluated for comparative purposes. The mean maximum hydrocodone plasma exposures (Cmax) vs. dose data for escalating ETR028 / 029 blend doses vs. extrapolated dose proportional exposures relative to a potentially lethal hydrocodone exposure in humans is presented in FIG. 7.

[0172] The mean maximum hydrocodone plasma exposures (Cmax) vs. dose data for escalating ETR028 / 029 blend doses vs. extrapolated dose proportional exposures relative to a potential lethal hydrocodone exposure in humans is presented in FIG. 7. The Cmaxvalues forthe 4- and 8-fold doses of HCBT were extrapolated from the Cmaxvs. dose curve established using the single, 2-fold and 16-fold HCBT doses evaluated in the study.

[0173] In stark contrast to the HCBT comparator, dose escalation of the 1 : 1 ratio of ETR028 / 029 did not exceed potentially lethal hydrocodone exposure.Key Benefits vs. currently prescribed hydrocodone

[0174] Significant Reduction in The Number of Prescribed Doses to Mitigate Misuse, Abuse and Diversion. The dosing regimens for currently marketed hydrocodone products are summarized below:

[0175] Tablets containing 5 mg Hydrocodone: The usual adult dosage is one or two tablets every four to six hours as needed for pain - a maximum of 12 tablets / day.

[0176] Tablets containing 10 mg Hydrocodone: The usual adult dosage is one tablet every four to six hours as needed for pain - a maximum of 6 tablets / day.

[0177] The sustained plasma exposure of hydrocodone following oral ingestion of O2P hydrocodone (FIG. 5) is consistent with a once or twice daily dosing regimen. Consequently, the number of prescribed 5 or 10 mg hydrocodone equivalent dose (HED) tablets for O2P hydrocodone is significantly reduced vs. the prescribed number of tablets for currently marketed hydrocodone products.

[0178] Limits The Number of Euphoric Experiences that Serve to Initiate and Reinforce Abuse Behavior. The PK / PD relationship for hydrocodone drug liking has been well characterized in multiple human abuse liability (HAL) studies. A dose of 20 mgs of hydrocodone has been demonstrated to produce a euphoric effect that is well-liked by individuals who engage in recreational abuse of hydrocodone. A 20 mg dose of hydrocodone produces a Cmaxof approximately 40 ng / ml. Based on the attenuated hydrocodone exposures with O2P hydrocodone at supratherapeutic doses, a dose of 480 mg of O2P hydrocodone would be required to achieve a comparable Cmax. Consequently, the number of 20 mg hydrocodone equivalent doses (i.e., number of euphoric experiences) for a O2P hydrocodone prescription is dramatically reduced vs. currently prescribed hydrocodone.

[0179] Significantly Reduces the Risk of Fatal Overdose. The unique pharmacokinetic profile of O2P hydrocodone significantly reduces the risk of fatal overdose. The demonstrated oral overdose protection which reduces hydrocodone exposures when supratherapeutic doses are ingested in concert with a once or twice daily dosing regimen (i.e., fewer tablets per prescription) serve to significantly reduce the risk of potentially fatal overdose.

[0180] While preferred embodiments of the present invention have been shown and described herein, it will be obvious to those skilled in the art that such embodiments are provided by way of example only. Numerous variations, changes, and substitutions will now occur to those skilled in the art without departing from the invention. It should be understood that various alternatives to the embodiments of the invention described herein may be employed in practicing the invention. It is intended that the following claims define the scope of the invention and that methods and structures within the scope of these claims and their equivalents be covered thereby.Example 3. Simulations of Steady-State Exposure

[0181] The data was generated using the well-established principle of superposition. Under the assumption of linear conditions, this principle can be used to simulate steady-state concentrations. The superposition principle states that under linear conditions (i.e., constant clearance) the total concentration of drug in the body is the sum of the remaining concentrations from each administered dose at that point in time when a measurement is made. As drug accumulates, drug elimination accelerates since it is a first-order process dependent on the amount of drug in the body.

[0182] Specific doses and specific dosing regimens were evaluated to best simulate the steady-state pharmacokinetics of the 5 mg and 10 mg (q 4 hours x 8 days) doses and dosing regimens for currently prescribed hydrocodone.

[0183] The PK modeling and steady-state simulations were performed using R (version 4.3.2 or later) software package / programming language.Table 6. Summary of Hydrocodone Concentrations by TreatmentTable 7. Summary of Simulated Hydrocodone AUCs by TreatmentTable 8. Power Model Analysis of Dose Proportionality of ETR028 and ET029 BlendedPK Parameter analysis (Cmax)Table 9. Power Model Analysis of Dose Proportionality of HCBT PK ParameterAnalysis (Cmax)

[0184] While preferred embodiments of the present invention have been shown and described herein, it will be obvious to those skilled in the art that such embodiments are provided by way of example only. Numerous variations, changes, and substitutions will now occur to those skilled in the art without departing from the invention. It should be understood that various alternatives to the embodiments of the invention described herein may be employed in practicing the invention. It is intended that the following claims define the scope of the invention and that methods and structures within the scope of these claims and their equivalents be covered thereby.

Claims

CLAIMSWhat is claimed is:

1. A pharmaceutical composition comprising two different molecules, wherein each molecule comprises at least one GI enzyme-labile opioid agonist-releasing subunit comprising an opioid agonist, wherein the at least one GI enzyme-labile opioid agonistreleasing subunit is covalently linked to at least one GI enzyme-inhibitor subunit, wherein each GI enzyme-labile opioid agonist-releasing subunit releases the opioid agonist, and the resulting opioid agonist plasma Cmaxconcentrations are less than dose proportional compared to an analogous opioid agonist.

2. A pharmaceutical composition comprising two different molecules, wherein each molecule comprises at least one GI enzyme-labile opioid agonist-releasing subunit comprising an opioid agonist, wherein the at least one GI enzyme-labile opioid agonistreleasing subunit is covalently linked to at least one GI enzyme-inhibitor subunit, wherein the GI enzyme-labile opioid agonist-releasing subunit releases the opioid agonist, and the opioid agonist provides an analgesic effect for about 24 hours to about 48 hours.

3. The pharmaceutical composition of claim 2, wherein the opioid agonist provides an analgesic effect for about 12 hours to about 24 hours.

4. The pharmaceutical composition of any one of claims 1 to 3, wherein an onset of analgesia occurs within 2 hours after administration.

5. The pharmaceutical composition of any one of claims 1 to 3, wherein an onset of analgesia occurs within 1 hour after administration.

6. A pharmaceutical composition comprising two different molecules, wherein each molecule comprises at least one GI enzyme-labile opioid agonist-releasing subunit comprising an opioid agonist, wherein the at least one GI enzyme-labile opioid agonistreleasing subunit is covalently linked to at least one GI enzyme-inhibitor subunit, wherein the GI enzyme-labile opioid agonist-releasing subunit releases the opioid agonist, and the opioid agonist achieves a mean plasma concentration after about 6 hours.

7. The pharmaceutical composition of claim 6, wherein the opioid agonist achieves a mean max plasma concentration after about 6 hours to about 10 hours.

8. The pharmaceutical composition of claim 6, wherein the opioid agonist achieves a mean max plasma concentration after about 6 hours to about 8 hours.

9. The pharmaceutical composition of claim 6, wherein the opioid agonist achieves a mean max plasma concentration after about 8 hours.

10. A pharmaceutical composition comprising two different molecules, wherein each molecule comprises at least one GI enzyme-labile opioid agonist-releasing subunit comprising an opioid agonist, wherein the at least one GI enzyme-labile opioid agonistreleasing subunit is covalently linked to at least one GI enzyme-inhibitor subunit, wherein the GI enzyme-labile opioid agonist-releasing subunit releases the opioid agonist, wherein the Cmaxof the opioid agonist occurs after about 6 hours upon administration of the pharmaceutical composition to a subject, and is reduced by at least 50% about 12 to about 24 hours subsequent to the administration.

11. A pharmaceutical composition comprising two different molecules, wherein each molecule comprises at least one GI enzyme-labile opioid agonist-releasing subunit comprising an opioid agonist, wherein the at least one GI enzyme-labile opioid agonistreleasing subunit is covalently linked to at least one GI enzyme-inhibitor subunit, wherein the GI enzyme-labile opioid agonist-releasing subunit releases the opioid agonist, wherein a supra-therapeutic dosage of the opioid agonist demonstrates reduced dose proportionality when compared to an analogous free opioid agonist.

12. A pharmaceutical composition comprising two different molecules, wherein each molecule comprises at least one GI enzyme-labile opioid agonist-releasing subunit comprising an opioid agonist, wherein the at least one GI enzyme-labile opioid agonistreleasing subunit is covalently linked to at least one GI enzyme-inhibitor subunit, wherein the GI enzyme-labile opioid agonist-releasing subunit releases the opioid agonist, wherein the opioid agonist maintains an effective plasma concentration for about 24 hours after administration of one dose.

13. The pharmaceutical composition of claim 12, wherein the opioid agonist maintains an effective plasma concentration for about 12 hours after administration of one dose.

14. The pharmaceutical composition of claim 12 or 13, wherein the opioid agonist has an effective plasma concentration of about 1 ng / mL or greater.

15. The pharmaceutical composition of claim 12 or 13, wherein the opioid agonist has an effective plasma concentration of about 10 ng / mL or greater.

16. The pharmaceutical composition of any one of claims 1 to 15, wherein the two different molecules are in a ratio of 1 :4 w / w.

17. The pharmaceutical composition of any one of claims 1 to 15, wherein the two different molecules are in a ratio of 1 :3 w / w.

18. The pharmaceutical composition of any one of claims 1 to 15, the two different molecules are in a ratio of 1 :2 w / w.

19. The pharmaceutical composition of any one of claims 1 to 15, wherein the two different molecules are in a ratio of 1 : 1 w / w.

20. The pharmaceutical composition of any one of claims 1 to 19, wherein the pharmaceutical composition comprises 10 mg to 1000 mg of the two different molecules.

21. The pharmaceutical composition of claim 20, wherein the pharmaceutical composition comprises 20 mg of the two different molecules.

22. The pharmaceutical composition of claim 20, wherein the pharmaceutical composition comprises 60 mg of the two different molecules.

23. The pharmaceutical composition of claim 20, wherein the pharmaceutical composition comprises 120 mg of the two different molecules.

24. The pharmaceutical composition of claim 20, wherein the pharmaceutical composition comprises 240 mg of the two different molecules.

25. The pharmaceutical composition of claim 20, wherein the pharmaceutical composition comprises 480 mg of the two different molecules.

26. The pharmaceutical composition of claim 20, wherein the pharmaceutical composition comprises 960 mg of the two different molecules.

27. The pharmaceutical composition of any one of claims 1 to 26, wherein the GI enzyme-labile opioid agonist-releasing subunits comprises morphine, hydromorphone, hydrocodone, oxycodone, codeine, levorphanol, meperidine, methadone, oxymorphone, dihydrocodeine, tramadol, tapentadol, buprenorphine, cebranopadol, or pharmaceutically acceptable salts, prodrugs, and mixtures thereof.

28. The pharmaceutical composition of claim 27, wherein at least one of the GI enzyme-labile opioid agonist-releasing subunits comprises morphine, or pharmaceutically acceptable salts thereof.

29. The pharmaceutical composition of claim 27, wherein at least one of the GI enzyme-labile opioid agonist-releasing subunits comprises hydrocodone, or pharmaceutically acceptable salts thereof.

30. The pharmaceutical composition of claim 27, wherein at least one of the GI enzyme-labile opioid agonist-releasing subunits comprises oxycodone, or pharmaceutically acceptable salts thereof.

31. The pharmaceutical composition of any one of claims 1 to 30, wherein the two or more different molecules are independently selected from the group consisting of a structure of Formula (IF), (IG), (IH), (II), and salts thereof:salts thereof; wherein:D is an opioid agonist;R1of R1-Z1- is independently selected from the group consisting of, wherein Y is amidine, guanidine, aminomethyl, substituted amidine, substituted guanidine, substituted aminomethyl, amidinomethyl, guanidinomethyl, substituted amidinomethyl, or substituted guanidinomethyl, and Q is hydrogen;R’ is selected from the group consisting of methyl, lower alkyl, substituted alkyl, aryl, substituted aryl, heteroalkyl, a natural or a non-natural amino acid, a polypeptide chain comprising natural or non-natural amino acids up to 10 amino acids in length, a linear or a branched polyethylene glycol chain up to 5 kDa, benzyloxy,R” is selected from the group consisting of acetyl, substituted acyl, a natural or a non-natural amino acid, and a polypeptide chain comprising natural or non-natural amino acids up to 10 amino acids in length;AA is a natural or a non-natural amino acid side chain that is recognized by trypsin; andZ1of R1-Z1- is independently selected from the group consisting of R1-O-CH2-CH2- and R1-CH2-C(O)-NH-CH2-CH2-.

32. The pharmaceutical composition of claim 31, or salts thereof, wherein Y is amidine, aminomethyl, or guanidine.

33. The pharmaceutical composition of claim 31 or 32, or salts thereof, wherein R1of R1-Z1- is independently selected from the group consisting of:

34. The pharmaceutical composition of any one of claims 31 to 33, wherein the two or more different molecules are independently selected from the group consisting of a structure of Formula (II-A), (II-B), (II-C), (II-D), (II-E), (II-F), (II-G), (II-H), (II-I), (II- J), (II- K), (II-L), (II-M), (II-N), (II-O), (II-P), (II-Q), (II-R), (II-S), (II-T), (II-U), (II-V), (II-W), (II- X), and salts thereof:wherein:R is selected from the group consisting ofX is hydrogen or -OH.

35. The pharmaceutical composition of claim 34, or salts thereof, wherein AA is the side chain of lysine or arginine.

36. The pharmaceutical composition of claim 34 or 35, or salts thereof, whereinR’ is methyl, benzyloxy,37. The pharmaceutical composition of any one of claims 34 to 36, or salts thereof, wherein R” is acetyl, -Gly-NAc, or -Ala-NAc.

38. The pharmaceutical composition of claim 34, comprising:or salts thereof; wherein:R' is independently, methyl, benzyloxy,AA is independently, the side chain of lysine or arginine;R" is independently, acetyl, -Gly-NAc, or -Ala-NAc; andX is independently, hydrogen or -OH.

39. The pharmaceutical composition of any one of claims 1 to 38, wherein the two different molecules is a structure of Compound A and Compound B, or pharmaceutically acceptable salts thereof:wherein the ratio of Compound A to Compound B is between 1 :4 w / w and 1 : 1 w / w, and the pharmaceutical composition is administered once or twice daily.

40. The pharmaceutical composition of claim 39, wherein the pharmaceutical composition is administered once daily.

41. The pharmaceutical composition of claim 39, wherein the pharmaceutical composition is administered twice daily.

42. The pharmaceutical composition of any one of claims 39 to 41, wherein the ratio of Compound A to Compound B is 1 : 1 w / w.

43. A pharmaceutical composition comprising a first opioid agonist and one or two different molecules, wherein each molecule comprises at least one Gl-enzyme-labileopioid agonist-releasing subunit comprising a second opioid agonist, wherein the at least one GI enzyme-labile opioid agonist-releasing subunit is covalently linked to at least one GI enzyme inhibitor subunit wherein the first opioid agonist reduces the onset of action time, wherein the first opioid agonist comprises morphine, hydromorphone, hydrocodone, oxycodone, codeine, levorphanol, meperidine, methadone, oxymorphone, dihydrocodeine, tramadol, tapentadol, buprenorphine, or pharmaceutically acceptable salts, prodrugs, and mixtures thereof, wherein the one or two different molecules are independently selected from the group consisting of a structure of Formula (IF), (IG), (IH), (II), and salts thereofwherein:D is an opioid agonist;R1of R1-Z1- is independently selected from the group consisting of, wherein Y is amidine, guanidine, aminomethyl, substituted amidine, substituted guanidine, substituted aminomethyl, amidinomethyl, guanidinomethyl, substituted amidinomethyl, or substituted guanidinomethyl, and Q is hydrogen;R’ is selected from the group consisting of methyl, lower alkyl, substituted alkyl, aryl, substituted aryl, heteroalkyl, a natural or a non-natural amino acid, a polypeptide chain comprising natural or non-natural amino acids up to 10 amino acids in length, a linear or a branched polyethylene glycol chain up to 5 kDa, benzyloxy,R” is selected from the group consisting of acetyl, substituted acyl, a natural or a non-natural amino acid, and a polypeptide chain comprising natural or non-natural amino acids up to 10 amino acids in length;AA is a natural or a non-natural amino acid side chain that is recognized by trypsin; andZ1of R1-Z1- is independently selected from the group consisting of R1-O-CH2-CH2- and R1-CH2-C(O)-NH-CH2-CH2-.

44. The pharmaceutical composition of claim 43, or salts thereof, wherein Y is amidine, aminomethyl, or guanidine.

45. The pharmaceutical composition of claim 43 or 44, or salts thereof, wherein R1of R1-Z1- is independently selected from the group consisting of:

46. The pharmaceutical composition of any one of claims 43 to 45, wherein the one or two different molecules are independently selected from the group consisting of a structure of Formula (II-A), (II-B), (II-C), (II-D), (II-E), (II-F), (II-G), (II-H), (II-I), (II- J), (II- K), (II-L), (II-M), (II-N), (II-O), (II-P), (II-Q), (II-R), (II-S), (II-T), (II-U), (II-V), (II-W), (II- X), and salts thereof:wherein:R is selected from the group consisting ofX is hydrogen or -OH.

47. The pharmaceutical composition of claim 46, or salts thereof, wherein AA is the side chain of lysine or arginine.

48. The pharmaceutical composition of claim 46 or 47, or salts thereof, whereinR’ is methyl, benzyloxy,49. The pharmaceutical composition of any one of claims 46 to 48, or salts thereof, wherein R” is acetyl, -Gly-NAc, or -Ala-NAc.

50. The pharmaceutical composition of any one of claims 43 to 49, wherein the pharmaceutical composition comprises about 10 mg to about 1000 mg of the one or two different molecules.

51. The pharmaceutical composition of claim 50, wherein the pharmaceutical composition comprises 20 mg of the one or two different molecules.

52. The pharmaceutical composition of claim 50, wherein the pharmaceutical composition comprises 40 mg of the one or two different molecules.

53. The pharmaceutical composition of claim 50, wherein the pharmaceutical composition comprises 60 mg of the one or two different molecules.

54. The pharmaceutical composition of claim 50, wherein the pharmaceutical composition comprises 120 mg of the one or two different molecules.

55. The pharmaceutical composition of claim 50, wherein the pharmaceutical composition comprises 240 mg of the one or two different molecules.

56. The pharmaceutical composition of claim 50, wherein the pharmaceutical composition comprises 480 mg of the one or two different molecules.

57. The pharmaceutical composition of claim 50, wherein the pharmaceutical composition comprises 960 mg of the one or two different molecules.

58. . The pharmaceutical composition of any one of claims 43 to 57, wherein the pharmaceutical composition comprises two different molecules.

59. The pharmaceutical composition of any one of claims 43 to 58, the two different molecules comprises:or salts thereof; wherein: R' is independently, methyl, benzyloxy,AA is independently, the side chain of lysine or arginine;R" is independently, acetyl, -Gly-NAc, or -Ala-NAc; andX is independently, hydrogen or -OH.

60. The pharmaceutical composition of any one of claims 43 to 59, wherein the pharmaceutical composition comprises two different molecules in a ratio of 1 :4 w / w.

61. The pharmaceutical composition of any one of claims 43 to 59, wherein the pharmaceutical composition comprises two different molecules in a ratio of 1 :3 w / w.

62. The pharmaceutical composition of any one of claims 43 to 59, the pharmaceutical composition comprises two different molecules in a ratio of 1 :2 w / w.

63. The pharmaceutical composition of any one of claims 43 to 59, wherein the pharmaceutical composition comprises two different molecules in a ratio of 1 : 1 w / w.

64. The pharmaceutical composition of any one of claims 49 to 57, wherein the one or two different molecules is a structure of Compound A and / or Compound B:wherein the ratio of Compound A to Compound B is between 1 :6 w / w and 6: 1 w / w, and the pharmaceutical composition is administered once or twice daily.

65. The pharmaceutical composition of claim 64, wherein the ratio of Compound A to Compound B is 1 :6 w / w.

66. The pharmaceutical composition of claim 64, wherein the ratio of Compound A to Compound B is 1 :4 w / w.

67. The pharmaceutical composition of claim 64, wherein the ratio of Compound A to Compound B is 1 : 1 w / w.

68. The pharmaceutical composition of claim 64, wherein the pharmaceutical composition comprises Compound A.

69. The pharmaceutical composition of claim 64, wherein the pharmaceutical composition comprises Compound B.

70. The pharmaceutical composition of any one of claims 43 to 69, wherein the first opioid agonist comprises between about 0.1 mg to about 20 mg in the pharmaceutical composition.

71. The pharmaceutical composition of claim 70, wherein the first opioid agonist comprises no more than 10 mg in the pharmaceutical composition.

72. The pharmaceutical composition of claim 70, wherein the first opioid agonist comprises about 6 mg in the pharmaceutical composition.

73. The pharmaceutical composition of claim 70, wherein the first opioid agonist comprises about 3 mg in the pharmaceutical composition.

74. The pharmaceutical composition of claim 70, wherein the first opioid agonist comprises about 1.5 mg in the pharmaceutical composition.

75. The pharmaceutical composition of claim 70, wherein the first opioid agonist comprises about 0.5 mg in the pharmaceutical composition.

76. The pharmaceutical composition of any one of claims 43 to 75, wherein the first opioid agonist comprises morphine, or pharmaceutically acceptable salts thereof.

77. The pharmaceutical composition of any one of claims 43 to 75, wherein the first opioid agonist comprises hydrocodone, or pharmaceutically acceptable salts thereof.

78. The pharmaceutical composition of any one of claims 43 to 75, wherein the first opioid agonist comprises oxycodone, or pharmaceutically acceptable salts thereof.

79. The pharmaceutical composition of any one of claims 43 to 78, wherein the pharmaceutical composition is administered once daily.

80. The pharmaceutical composition of any one of claims 43 to 79, wherein the pharmaceutical composition is administered twice daily.

81. A method of treating pain in a subj ect in need thereof, the method comprising administering to the subject a pharmaceutical composition of any one of claims 1 to 80.

82. A method of treating pain in a subject, the method comprising administering to the subject a pharmaceutical composition comprising a first opioid agonist and one or two different molecules, wherein each molecule comprises at least one Gl-enzyme-labile opioid agonist-releasing subunit comprising a second opioid agonist, wherein the at least one GI enzyme-labile opioid agonist-releasing subunit is covalently linked to at least one GI enzyme inhibitor subunit wherein the first opioid agonist reduces the onset of action time.

83. The method of claim 82, wherein the first opioid agonist is administered at the same time as the one or two different molecules.

84. The method of claim 82, wherein the first opioid agonist is administered before or after the one or two different molecules.

85. The method of any one of claims 82 to 84, wherein an opioid agonist plasma concentration within the subject is not dose-proportional.

86. The method of any one of claims 82 to 84, wherein the opioid agonist plasma concentration in the subject demonstrates reduced dose proportionality when compared to the free opioid agonist delivered by the one or two different molecules.

87. The method of any one of claims 82 to 84, wherein the opioid agonist plasma concentration within the subject is below a lethal amount.

88. The method of any one of claims 82 to 86, wherein the administration of a therapeutically effective dose of the pharmaceutical composition reduces events of abuse, misuse, and overdose, when compared to the administration of the free opioid agonist.

89. The method of any one of claims 82 to 87, wherein the pain is acute pain.

90. The method of any one of claims 82 to 87, wherein the pain is chronic pain.

91. A method of preventing opioid abuse or treating opioid overdose in a subject, comprising administering to the subject a therapeutically effective amount of a pharmaceutical composition of any one of claims 1 to 80.

92. The method of claim 91, wherein the subject is experiencing symptoms from overdose of an opioid agonist.

93. The method of claim 91, wherein the subject is suffering from opioid use disorder.

94. The method of claim 91, wherein the subject is suffering from alcohol use disorder.

95. The method of claim 91, wherein the subject is suffering from an impulse control disorder.

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