# Screening for rAAV transduction enhancers

> **NIH NIH R44** · MICROBIOTIX, INC · 2021 · $999,976

## Abstract

ABSTRACT
 Recombinant adeno-associated virus (rAAV) is an established vector for gene therapy and
vaccines with the potential to radically change the face of modern healthcare. Although extremely
promising, with two examples already approved for clinical use by the FDA, rAAV therapeutics
are hampered by the necessity of large vector doses and the serious logistical and safety
challenges resulting from these doses, including detrimental innate and adaptive immune
responses to capsid and transgene products. The overall goal of this proposal is to optimizie
small molecule drugs that enhance rAAV transduction and enable the therapeutic application of
this technology at vector doses below the threshold of deleterious responses.
 As the result of a successful Phase I, which was a collaborative effort by Microbiotix, Inc.
(MBX) and Dr. Michael Farzan’s laboratory at TSRI, 11 compounds in five chemical series were
prioritized as validated rAAV transduction enhancers. After validation testing, the furopyrimidine
scaffold was selected as the highest priority series, and members of isoindoline and
pyridopyrimidine scaffolds were selected as backups. The furopyrimidine MBXC-4409 exhibited
multiple favorable features, inlcuding: (a) highly significant dose-dependent activity (EC50 <10 µM)
and transduction enhancements (≥3-fold) that are independent of the transgene (Gaussia or
Firefly luciferase), the assay signal (luminescence or qRT-PCR), the cell line (HT1080 and HeLa),
or the capsid serotype (serotypes 1, 2, 8, and 9); (b) CC50 ≥100 µM; SI (CC50/EC50) >20; (c) drug-
like structures of ≥98% purity (NMR) and of correct mass (LC/MS); (d) not promiscuous in other
unrelated screens and no significant inhibition or enhancement of infection by other viruses (ZIKV
and VSV); (e) time-of-addition studies indicate early action; and (f) favorable in vitro ADME
properties (solubility ≥100 µM; stable in murine serum; serum protein binding ≤93%).
 This Phase II effort will continue to include Dr. Michael Farzan’s laboratory at TSRI. The
potency and drug-like properties of the furopyrimidine inhibitor series will be optimized, with the
isoindoline and pyridopyrimidine series as backups, to produce in vivo-validated preclinical
candidates for development as adjunctive agents to enhance rAAV therapeutics. Analogs will be
evaluated through an assay funnel designed to prioritize them by potency, selectivity, and specific
non-toxic mechanisms of action. Prioritized analogs will be formulated for in vivo studies. The
maximum tolerated dose of the most promising lead compounds will be determined in mouse
tolerability studies, and pharmacokinetic analyses will be used to further prioritize compounds and
optimize dosing strategies. Inhibitors will be tested for efficacy in combination with rAAV-gLuc in
a murine model of transduction with whole body luminescence and qRT-PCR analysis of genome
copies in tissues. The major milestone of this proposal is to select an in vivo-validated...

## Key facts

- **NIH application ID:** 10157565
- **Project number:** 2R44AI125043-03
- **Recipient organization:** MICROBIOTIX, INC
- **Principal Investigator:** Terry L. Bowlin
- **Activity code:** R44 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2021
- **Award amount:** $999,976
- **Award type:** 2
- **Project period:** 2017-01-27 → 2024-01-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10157565

## Citation

> US National Institutes of Health, RePORTER application 10157565, Screening for rAAV transduction enhancers (2R44AI125043-03). Retrieved via AI Analytics 2026-05-22 from https://api.ai-analytics.org/grant/nih/10157565. Licensed CC0.

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