NIH grant: Engineered Probiotics as a Strategy to Induce Oral Tolerance to AAV-Delivered Broadly Neutralizing Antibodies (1R01AI203544-01)

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NIH grant 1R01AI203544-01: Engineered Probiotics as a Strategy to Induce Oral Tolerance to AAV-Delivered Broadly Neutralizing Antibodies
Institute: {'code': 'AI', 'abbreviation': 'NIAID', 'name': 'National Institute of Allergy and Infectious Diseases', 'admin_org_id': '12500003', 'admin_funding_url': ''}. PI: James Mario Termini. Award: $457128.
Start: 2026-07-16T00:00:00.
Project Summary/Abstract
Given the vast assortment of human monoclonal antibodies with potent neutralizing activity, one can envision
long-term delivery of a combination of antibodies to achieve sterilizing immunity against most circulating HIV
strains. Adeno-associated virus (AAV) vectors are well suited for such sustained antibody delivery. Our group
has demonstrated high circulating levels of broadly neutralizing antibodies (bNAbs) following a single
intramuscular AAV injection in rhesus macaques, as well as long-term virologic suppression for more than nine
years after AAV-mediated delivery of 10-1074 and 3BNC117. These studies have not only demonstrated that
long-term expression of AAV-delivered bNAbs can lead to a functional cure but can also eliminate the viral
reservoir. In three macaques, intact proviral DNA levels were at or below the limit of detection five years
following AAV delivery. Despite these successes, host immune responses against the delivered antibodies
have frequently limited long-term transgene expression.
 To address the crippling effects of anti-drug antibody (ADA) responses, this proposal seeks to induce
antigen-specific oral tolerance using engineered probiotics. We aim to determine whether pretreatment with
these probiotics can prevent, and potentially reverse, immune responses to AAV-delivered bNAbs, thereby
enabling sustained antibody expression in both uninfected and HIV-infected hosts. To test this concept, we will
develop and validate designer probiotic strains that can be delivered orally, colonize the gut, and secrete
functional bNAb immunoadhesins. Bifidobacterium longum will be engineered to secrete immunoadhesins
derived from the bNAbs 3BNC117 and 10-1074, while Lactococcus lactis will be engineered to secrete the
cytokine IL-10 to enhance tolerogenic signaling and support oral tolerance induction.
 Rhesus macaques will be pretreated with weekly oral probiotics prior to intramuscular AAV9 delivery.
This approach will be evaluated longitudinally to assess ADA formation, regulatory T cell induction, microbial
persistence, and the durability of AAV-mediated bNAb expression. This approach will be directly compared to
oral delivery of purified bNAbs, as classically used in oral tolerance studies. In addition, we will assess whether
sustained probiotic exposure can reprogram established humoral immunity and restore bNAb expression
following AAV delivery. Finally, probiotic pretreatment will be evaluated in SHIV chronically infected rhesus
macaques receiving antiretroviral therapy. This model is more clinically relevant and will allow us to determine
whether oral tolerance can be established in the setting of chronic immune inflammation and dysregulation.
 If successful, this project will establish a microbiome-based, non-immunosuppressive strategy for
overcoming the last critical barrier to AAV-mediated bNAb delivery. These studies could enable the
development of long-term, ART-free HIV treatment and prevention and would provide a broadly applicable
approach for improving the durability of AAV-delivered biologics across many diseases.

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