Production and characterization of a human antisperm monoclonal antibody against CD52g for topical contraception in women.

BACKGROUND: Approximately 40% of human pregnancies are unintended, indicating a need for more acceptable effective contraception methods. New antibody production systems make it possible to manufacture reagent-grade human monoclonal antibodies (mAbs) for clinical use. We used the Nicotiana platform to produce a human antisperm mAb and tested its efficacy for on-demand topical contraception. METHODS: Heavy and light chain variable region DNA sequences of a human IgM antisperm antibody derived from an infertile woman were inserted with human IgG1 constant region sequences into an agrobacterium and transfected into Nicotiana benthamiana. The product, an IgG1 mAb ["Human Contraception Antibody" (HCA)], was purified on Protein A columns, and QC was performed using the LabChip GXII Touch protein characterization system and SEC-HPLC. HCA was tested for antigen specificity by immunofluorescence and western blot assays, antisperm activity by sperm agglutination and complement dependent sperm immobilization assays, and safety in a human vaginal tissue (EpiVaginal™) model. FINDINGS: HCA was obtained at concentrations ranging from 0.4 to 4 mg/ml and consisted of > 90% IgG monomers. The mAb specifically reacted with a glycan epitope on CD52g, a glycoprotein produced in the male reproductive tract and found in abundance on sperm. HCA potently agglutinated sperm under a variety of relevant physiological conditions at concentrations ≥ 6.25 µg/ml, and mediated complement-dependent sperm immobilization at concentrations ≥ 1 µg/ml. HCA and its immune complexes did not induce inflammation in EpiVaginal™ tissue. INTERPRETATION: HCA, an IgG1 mAb with potent sperm agglutination and immobilization activity and a good safety profile, is a promising candidate for female contraception. FUNDING: This research was supported by grants R01 HD095630 and P50HD096957 from the National Institutes of Health.

Salinity-dependent expression of ncc2 in opercular epithelium and gill of mummichog (Fundulus heteroclitus).

Mummichogs (Fundulus heteroclitus) can tolerate abrupt changes in environmental salinity because of their ability to rapidly adjust the activities of ionocytes in branchial and opercular epithelia. In turn, the concerted expression of sub-cellular effectors of ion transport underlies adaptive responses to fluctuating salinities. Exposure to seawater (SW) stimulates the expression of Na+/K+/2Cl- cotransporter 1 (nkcc1) and cystic fibrosis transmembrane regulator (cftr) mRNAs in support of ion extrusion by SW-type ionocytes. Given the incomplete understanding of how freshwater (FW)-type ionocytes actually operate in mummichogs, the transcriptional responses essential for ion absorption in FW environments remain unresolved. In a subset of species, a 'fish-specific' Na+/Cl- cotransporter denoted Ncc2 (Slc12a10) is responsible for the uptake of Na+ and Cl- across the apical surface of FW-type ionocytes. In the current study, we identified an ncc2 transcript that is highly expressed in gill filaments and opercular epithelium of FW-acclimated mummichogs. Within 1 day of transfer from SW to FW, ncc2 levels in both tissues increased in parallel with reductions in nkcc1 and cftr. Conversely, mummichogs transferred from FW to SW exhibited marked reductions in ncc2 concurrent with increases in nkcc1 and cftr. Immunohistochemical analyses employing a homologous antibody revealed apical Ncc2-immunoreactivity in Na+/K+-ATPase-immunoreactive ionocytes of FW-acclimated animals. Our combined observations suggest that Ncc2/ncc2-expressing ionocytes support the capacity of mummichogs to inhabit FW environments.