# Overcoming Pyrazinamide Resistance with Pyrazinoate-Cephalosporin Conjugates

> **NIH NIH R21** · UNIVERSITY OF MINNESOTA · 2020 · $229,632

## Abstract

SUMMARY
Mycobacterium tuberculosis (Mtb), the principal etiological agent of tuberculosis (TB), infects over one-third of
humanity and is now the leading cause of infectious disease mortality by a single pathogen. Pyrazinamide
(PZA) is a first-line sterilizing anti-tubercular drug that is anticipated to be an irreplaceable component of future
TB treatment regimens. Previous studies have demonstrated that PZA is a pro-drug which is converted to the
active form of pyrazinoic acid (POA) by the M. tuberculosis amidase PncA, and that loss of function mutations
in pncA account for the vast majority of PZA resistance. Resistance to PZA in turn is associated with
significantly higher treatment failures and relapse rates. Attempts to directly administer POA in humans and
mice have been unsuccessful because POA cannot attain sufficient concentrations within Mtb at the infection
site. Prodrug approaches employing simple POA esters have been unable to overcome the intrinsic drug
disposition liabilities of POA as conventional prodrugs are rapidly hydrolyzed by serum esterases. In
preliminary data, we have developed a novel prodrug strategy for POA through conjugation to a β-lactam
promoiety leading to selective release of POA by the unique mycobacterial beta-lactamase BlaC and not by
serum esterases. This strategy overcomes PZA-resistance by circumventing PncA-mediated activation and
renders blaC conditionally essential since loss of function mutations to blaC, restores susceptibility of Mtb to
the β-lactam promoiety. Consequently, we hypothesize the genetic barrier for development of spontaneous
resistance to our POA-β-lactam conjugates will be high. The objectives of this application are to synthesize and
evaluate β-lactam prodrugs of POA, which are orally bioavailable and selectively release POA within Mtb-
infected macrophages. We will accomplish the overall objectives of this application by pursuing two specific
aims. In aim, we will synthesize β-lactam prodrugs of POA, which are selectively activated by Mtb to release
POA and not by commensal microbiota. The prodrugs will be biochemically and microbiologically characterized
with a panel of β-lactamases and microorganisms. In aim 2, we will measure prodrug stability in serum and
stimulated gastric fluid and permeability in Caco-2 cells. We will next determine complete pharmacokinetic
parameters and the extent of POA release by the gut microbiome. Finally, we plan to evaluate one compound
in a murine TB infection model.

## Key facts

- **NIH application ID:** 9895968
- **Project number:** 1R21AI144501-01A1
- **Recipient organization:** UNIVERSITY OF MINNESOTA
- **Principal Investigator:** Courtney C Aldrich
- **Activity code:** R21 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2020
- **Award amount:** $229,632
- **Award type:** 1
- **Project period:** 2020-01-25 → 2021-12-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/9895968

## Citation

> US National Institutes of Health, RePORTER application 9895968, Overcoming Pyrazinamide Resistance with Pyrazinoate-Cephalosporin Conjugates (1R21AI144501-01A1). Retrieved via AI Analytics 2026-05-23 from https://api.ai-analytics.org/grant/nih/9895968. Licensed CC0.

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