Identification of Arvicola terrestris scherman Sperm Antigens for Immune Contraceptive Purposes

The cyclical proliferation of the wild fossorial rodent Arvicola terrestris scherman (ATS) is critical in mid-mountain ecosystems of several European countries. Our goal is to develop an immunocontraceptive vaccine to control their fertility, as a sustainable alternative to chemical poisons currently used. Indeed, these chemicals cause the death of ATS predators and animals sharing their ecosystem, and current laws progressively limit their use, making the development of a targeted vaccination strategy an interesting and efficient alternative. In order to identify species-specific sperm antigens, male and female ATS received subcutaneous injections of whole ATS spermatozoa to elicit an immune response. The analysis of the immune sera led to the identification of 120 immunogenic proteins of sperm cells. Of these, 15 were strictly sperm-specific and located in different regions of the male gamete. Some of these antigens are proteins involved in molecular events essential to the reproductive process, such as sperm–egg interaction, acrosomal reaction, or sperm motility. This approach not only identified a panel of immunogenic proteins from ATS sperm cells, but also demonstrated that some of these proteins trigger an immune response in both male and female ATS. These spermatic antigens are good candidates for the development of a contraceptive vaccine.

Polyacrylate-GnRH Peptide Conjugate as an Oral Contraceptive Vaccine Candidate

Contraceptive vaccines are designed to elicit immune responses against major components of animal reproductive systems. These vaccines, which are most commonly administered via injection, typically target gonadotropin-releasing hormone (GnRH). However, the need to restrain animals for treatment limits the field applications of injectable vaccines. Oral administration would broaden vaccine applicability. We explored contraceptive vaccine candidates composed of GnRH peptide hormone, universal T helper PADRE (P), and a poly(methylacrylate) (PMA)-based delivery system. When self-assembled into nanoparticles, PMA-P-GnRH induced the production of high IgG titers after subcutaneous and oral administration in mice. PADRE was then replaced with pig T helper derived from the swine flu virus, and the vaccine was tested in pigs. High levels of systemic antibodies were produced in pigs after both injection and oral administration of the vaccine. In conclusion, we developed a simple peptide–polymer conjugate that shows promise as an effective, adjuvant-free, oral GnRH-based contraceptive vaccine.

Immunocontraceptive potential of a GnRH receptor-based fusion recombinant protein

Background The management of stray dog population has been of utmost importance due to their overpopulation, increase in dog bites incidence, and rabies. Contraceptive vaccines, a non-surgical alternative to spaying and neutering are viewed as a valuable option for the management of dog population. In this study, the contraceptive potential of a recombinant fusion protein containing the three genes GnRH, GnRH receptor, and ZP3 was explored. Results The gene fragment encoding GnRH, GnRHR, and ZP3 along with the antigenic epitopes of canine distemper virus and tetanus toxoid was assembled, synthesized, and cloned into pET28a expression vector. The resulting construct GVAC08 was successfully transformed into BL21DE3 strain of E. coli and confirmed by colony PCR. The recombinant GVAC08 protein was expressed and purified using Ni-NTA and was confirmed to be a 50-KDa protein by SDS PAGE and Western blot. Mice were immunized with the GVAC08 protein using Freund’s complete adjuvant followed by a booster using Freund’s incomplete adjuvant. This induced a high antibody titer against GnRH, GnRH receptor, and ZP3 which was determined by ELISA. Conclusion Mating studies showed that the GVAC08 recombinant protein was able to reduce the litter size in immunized mice showing improved efficacy. However, the vaccine candidate with further improvements will be a viable contraceptive vaccine.

Designing a Construct of Chimeric Multi-Epitopes Protein for Contraceptive Vaccine in Mice: An Immunoinformatics and In Silico Study

Background: Contraceptive vaccines (CVs) can be used as a valuable and alternative method for the prevention of gestation in humans and animals. These vaccines can have several targets, such as superficial sperm proteins. Vaccines based on sperm antigens are quite efficacious to create a contraceptive effect. However, multi-epitope vaccines are more effective in stimulating the immune system and producing more antibodies to reduce the infertility rate. Materials and Methods: This study aimed to design and evaluate a chimeric fusion protein containing IZUMO, SACA3, and PH-20 epitopes. IZUMO1, SACA3, and PH-20 were assessed, and appropriate regions were selected using various bioinformatics tools, including IEDB, I-TASSER, ProtParam, Asa-View, and Chimera software. Protein epitopes were selected based on various characters, including specificity, solvent accessibility, their weight and length, antigenic intensity, and topology. Epitopes with high antigenic potential were selected and joined together by linkers. The designed fusion protein was simulated using Molecular Dynamic, GROMACS 5, and Chimera 1.14 software. Results: The results demonstrated that all antigenic plots and availability of epitopes in the new construct remained constant. The spermatic antigens were combined using rigid linkers as a new construct and showed a stable formation with proper solvent accessibility validated by ProSA-web and PROCHECK. Also, comparing the new structure with its original one did not show any structural change. Conclusion: Based on bioinformatics results, the fusion protein that consists of three spermatic antigens has productive potential to stimulate the immune system and capable of producing more antibodies in circulation and reliable infertility.