The incidence of fatal overdoses has increased worldwide due to the widespread access to illicit fentanyl and its potent analogues. Vaccines offer a promising strategy to reduce the prevalence of opioid use disorders (OUDs) and to prevent toxicity from accidental and deliberate exposure to fentanyl and its derivatives. This study describes the development and characterization of vaccine formulations consisting of novel fentanyl-based haptens conjugated to carrier proteins. Vaccine efficacy was tested against opioid-induced behavior and toxicity in mice and rats challenged with fentanyl and its analogues. Prophylactic vaccination reduced fentanyl- and sufentanil-induced antinociception, respiratory depression, and bradycardia in mice and rats. Therapeutic vaccination also reduced fentanyl intravenous self-administration in rats. Because of their selectivity, vaccines did not interfere with the pharmacological effects of commonly used anesthetics nor with methadone, naloxone, oxycodone, or heroin. These preclinical data support the translation of vaccines as a viable strategy to counteract fentanyl use disorders and toxicity.

Vaccines may offer a new treatment strategy for opioid use disorders and opioid-related overdoses. To speed translation, this study evaluates opioid conjugate vaccines containing components suitable for pharmaceutical manufacturing and compares analytical assays for conjugate characterization. Three oxycodone-based haptens (OXY) containing either PEGylated or tetraglycine [(Gly)4] linkers were conjugated to a keyhole limpet hemocyanin (KLH) carrier protein via carbodiimide (EDAC) or maleimide chemistry. The EDAC-conjugated OXY(Gly)4-KLH was most effective in reducing oxycodone distribution to the brain in mice. Vaccine efficacy was T cell-dependent. The lead OXY hapten was conjugated to the KLH, tetanus toxoid, diphtheria cross-reactive material (CRM), as well as the E. coli-expressed CRM (EcoCRM) and nontoxic tetanus toxin heavy chain fragment C (rTTHc) carrier proteins. All vaccines induced early hapten-specific B cell expansion and showed equivalent efficacy against oxycodone in mice. However, some hapten-protein conjugates were easier to characterize for molecular weight and size. Finally, heroin vaccines formulated with either EcoCRM or KLH were equally effective in reducing heroin-induced antinociception and distribution to the brain of heroin and its metabolites in mice. This study identifies vaccine candidates and vaccine components for further development

Drug-related fatal overdoses have significantly increased in the past decade due to the widespread availability of illicit fentanyl and other potent synthetic opioids such as carfentanil. Deliberate or accidental consumption or exposure to carfentanil, fentanyl, and their mixture induces respiratory depression and bradycardia that can be difficult to reverse with the opioid receptor antagonist naloxone. Vaccines offer a promising strategy to reduce the incidence of fatalities associated with fentanyl-related substances, as well as treatment for opioid use disorder (OUD). This study reports monovalent and bivalent vaccination strategies that elicit polyclonal antibody responses effective in protecting against the pharmacological actions of carfentanil, fentanyl, or carfentanil/fentanyl mixtures. Rats were prophylactically immunized with individual conjugate vaccines containing either carfentanil- or fentanyl-based haptens, or their combination in bivalent vaccine formulations, and then challenged with carfentanil, fentanyl, or their mixture. First, these studies identified a lead vaccine protective against carfentanil-induced antinociception, respiratory depression, and bradycardia. Then, efficacy against both carfentanil and fentanyl was achieved through bivalent vaccination strategies that combined lead anti-carfentanil and anti-fentanyl vaccines via either heterologous prime/boost or co-administration immunization regimens. These preclinical data support the development of vaccines as a viable strategy to prevent toxicity from exposure to excessive doses of carfentanil, fentanyl, or their mixtures.

The ongoing public health emergency of opioid use disorders (OUD) and overdose in the United States is largely driven by fentanyl and its related analogues and has resulted in over 75 673 deaths in 2021. Immunotherapeutics such as vaccines have been investigated as a potential interventional strategy complementary to current pharmacotherapies to reduce the incidence of OUD and opioid-related overdose. Given the importance of targeting structurally distinct fentanyl analogues, this study compared a previously established lead conjugate vaccine (F 1 –CRM) to a series of novel vaccines incorporating haptens derived from alfentanil and acetylfentanyl (F 8, 9a, 9b, 10 ), and evaluated their efficacy against drug-induced pharmacological effects in rats. While no vaccine tested provided significant protection against alfentanil, lead formulations were effective in reducing antinociception, respiratory depression, and bradycardia elicited by fentanyl, sufentanil, and acetylfentanyl. Compared with control, vaccination with F 1 –CRM also reduced drug levels in the brain of rats challenged with lethal doses of fentanyl. These data further support investigation of F 1 –CRM as a candidate vaccine against fentanyl and selected analogues.