Bispecific antibody for topical administration to prevent gonorrhea

1 Gonorrhea affects over 80 million individuals globally, annually. Over 700,000 cases were reported to the CDC 2 in 2021, a relentless rise since 2009. Women suffer the most serious consequences of this disease, including 3 pelvic inflammatory disease, ectopic pregnancy and infertility. The etiologic agent, Neisseria gonorrhoeae (Ng)

4 has become resistant to almost every antibiotic in clinical use. The emergence of ceftriaxone-resistant isolates 5 in all regions of the world portends an era of untreatable gonorrhea. There is no licensed vaccine against 6 gonorrhea. Thus, safe and effective preventive measures are urgently needed. Ng evade killing by complement

7 (C’) by sialylating its lipooligosaccharide (LOS), which results in recruitment of the C’ inhibitor, factor H (FH). To 8 exploit this virulence mechanism, we fused the Ng-binding fragment of FH (that lacks C’ inhibitory activity) to the 9 C-terminus of human IgG3 Fc to produce Fc3/FH*. Fc3/FH* killed all 46 Ng isolates that expressed the PorB1B

10 allele of the major outer membrane porin B (PorB) protein in a C’-dependent bactericidal assay, and diminished 11 the duration and burden of Ng in the mouse vaginal colonization model. However, Fc3/FH was bactericidal 12 against only 2 of 15 Ng strains that expressed the PorB1A allele. Although responsible for only a minority of

13 infections, PorB1A isolates have a relatively high propensity to disseminate through the bloodstream. To reliably 14 cover PorB1A isolates, we will create a novel bispecific mAb where the C-terminus of FH will be fused to an anti- 15 LOS mAb called 2C7. mAb 2C7 recognizes an epitope distinct from LOS sialic acid, which is critical for Fc3/FH

16 binding. Further, mAb 2C7 binds independently of the PorB allele expressed. The 2C7 LOS epitope is critical for 17 Ng colonization, and therefore expressed by >95% of Ng in vivo. A chimeric version of mAb 2C7 shows C’- 18 dependent killing of all minimally passaged Ng tested and attenuates mouse vaginal colonization by both PorB1A

19 and PorB1B Ng. The advantages of the bispecific 2C7/FH* mAb include: 1) broad activity against PorB1A and 20 PorB1B Ng; 2) a substantially raised threshold for the development of drug-resistance by targeting two distinct 21 epitopes and virulence factors; and 3) reduced cost of production compared to producing two separate molecules

22 – a critical consideration for Ng therapeutics because gonorrhea rates are highest among socio-economically 23 underprivileged and marginalized populations and in low- and middle-income countries. In Aim 1, we will produce 24 four bispecific mAbs that contains FH domains 19-20 or domain 20 alone fused to the C-terminus of either the

25 heavy or light chain of chimeric mAb 2C7. A 6-month accelerated stability study will be carried out on the

26 bispecifics. In Aim 2, we will compare the efficacy of the four bispecifics in vitro using C’-dependent bactericidal 27 assays against a small panel of Ng isolates. The most effective lead molecule will be tested against a broader 28 panel of Ng isolates to confirm breadth of coverage. In Aim 3, in vivo efficacy of the lead candidate against

29 PorB1A and PorB1B Ng will be established in the mouse vaginal colonization model using transgenic mice that 30 express the C’ inhibitors FH and C4b-binding protein (C4BP) to better simulate a human-like C’ environment.