A novel anti-gonorrhea immunotherapeutic

Wycoff, Keith; Ram, Sanjay

Abstract

Antimicrobial resistance is a major public health problem worldwide. Neisseria gonorrhoeae (Ng), the causative agent of the sexually transmitted infection gonorrhea, has become multidrug-resistant and has achieved ?superbug? status. Novel therapeutics against Ng are urgently needed. Complement (C?) is a key arm of innate immune defenses. A mechanism used by several pathogens, including Ng, to escape C? is to bind to a host C? inhibitor called factor H (FH). FH comprises 20 domains, arranged in an extended head-to-tail fashion. Only the four N-terminal domains (domains 1-4) possess C? inhibiting activity; the remainder of the molecule is important for recognition of host surfaces. Many pathogens, including Ng, bind FH through domains 6-7 and/or 19-20. A recombinant fusion of FH domains 18-20 to IgG Fc (FH18-20/Fc) promotes C?-dependent killing of Ng. Domains 19 and 20, in the context of full-length FH, bind to host cells and protect them from damage by C?. Introducing a D?G amino acid mutation in domain 19 of FH18-20/Fc to yield FH(18-20)*/Fc retained anti-Ng efficacy but abrogated C?-mediated lysis of host cells. Topically administered FH(18-20)*/Fc attenuated Ng infection in the mouse vaginal colonization model. Planet Biotechnology Inc. (PBI) has pioneered the production of functional antibodies and Fc-fusion proteins in tobacco plants (Nicotiana benthamiana). Production of large quantities of functional FH(18-20)*/Fc in plants at relatively low costs compared to traditional mammalian cell culture systems constitutes an ideal platform for developing anti- infective immunotherapeutics.
In Aim 1, PBI will produce four FH(18-20)*/Fc molecules differing in their Fc: three with human Fc (hFc) for studies in vitro and one with mouse Fc (mFc) for studies in mice. The three different hFc's fused to FH(18-20)* will be: i) Fc from IgG1 (equivalent to what has been produced in CHO cells); ii) a linear ?? chimera, comprising IgG1 Fc plus CH2 and CH3 from IgA2; and iii) Fc from IgA2. We expect human IgA2 Fc to increase ADCC by engaging Fc?RI on human neutrophils.
In Aim 2, UMass will examine the ability of the three FH(18-20)*/hFc's described above to i) bind to, ii) deposit C3 on, iii) effect C? dependent killing of and iv) support opsonophagocytosis of drug-resistant Ng. The efficacy of FH(18-20)*/mFc against Ng will be tested in the mouse vaginal colonization model using human FH transgenic BALB/c mice. The use of these novel mice will evaluate the therapeutic in the context of human FH, as would occur in humans. We also speculate that FH(18-20)*/Fc may guide bacteria to Fc receptors on antigen presenting cells and elicit an immune response that may protect mice against reinfection. This possibility will be addressed by re-infecting the mice following FH(18-20)*/Fc (or control) treatment. We envision the use of FH(18-20)*/Fc either as a topical prophylactic in women at high risk for gonorrhea, or as a parenteral adjunctive anti-infective. While this proposal focuses on Ng, we believe FH(18-20)*/Fc will be useful against several medically important drug-resistant pathogens that bind to human FH.

Public Health Relevance

Gonorrhea, a sexually transmitted infection that adversely affects the reproductive health of women worldwide, has become resistant to almost every conventional antibiotic. In an effort to develop new therapies against gonorrhea, we have created a chimeric Fc fusion protein that activates complement on and kills gonococci. In this proposal, we will develop additional chimeric molecules with novel Fc modifications and produce these fusion proteins in tobacco plants, which will provide an economical alternative to develop adjunctive treatments against multidrug-resistant gonorrhea.