Pharmacological and Molecular Mechanisms Behind the Sterilizing Activity of Pyrazinamide

Abstract

Inclusion of pyrazinamide (PZA) in the tuberculosis (TB) drug regimen during the 1970s enabled a reduction in treatment duration from 12 to 6 months. PZA has this remarkable effect in patients despite displaying poor potency against Mycobacterium tuberculosis (Mtb) in vitro. The pharmacological basis for the in vivo sterilizing activity of the drug has remained obscure and its bacterial target controversial. Recently it was shown that PZA penetrates necrotic caseous TB lung lesions and kills nongrowing, drug-tolerant bacilli. Furthermore, it was uncovered that PZA inhibits bacterial Coenzyme A biosynthesis. It may block this pathway by triggering degradation of its target, aspartate decarboxylase. The elucidation of the pharmacological and molecular mechanisms of PZA provides the basis for the rational discovery of the next-generation PZA with improved in vitro potency while maintaining attractive pharmacological properties.

Keywords: drug tolerance; lesion penetration; pyrazinamide; target degradation; tuberculosis.

Figure 1.

Pharmacological basis of PZA’s sterilizing activity. (A) MALDI mass spectrometry ion maps of PZA and clofazimine, showing that PZA effectively penetrates the cellular and necrotic compartments of TB lesions, at concentrations identical to those achieved in plasma, in contrast to clofazimine which diffuses poorly into non-vascularized caseum. The left panel shows hematoxylin & eosin staining of the adjacent tissue section revealing the underlying lesion architecture and histology. The necrotic or caseous core of the lesion is highlighted in white or black contour lines. (B) Mtb bacilli do not replicate in ex vivo caseum for the 7-day duration of the cidal assay. Bacterial burden is shown as black circles, and cumulative burden measured as chromosome equivalent is shown as empty circles, indicating that the observed no-net growth is true no-growth rather than the result of balanced death and growth. (C) PZA kills non-growing drug tolerant persister Mtb present in necrotic caseous lesion material ex vivo. Clofazimine is shown as an example of a drug that is not active against caseum Mtb. Experimental details are described in.

Figure 2, Key Figure.

Proposed antibacterial mechanism of action of PZA. (A) PZA, once converted into POA by the bacterial amidase PncA, blocks synthesis of the essential cofactor Coenzyme A at the aspartate decarboxylase PanD-catalyzed step. (B) PanD level is post-translationally regulated by the caseinolytic protease complex ClpC1-ClpP and binding of POA to PanD triggers increased degradation of the protein. Upper part: PanD (blue circle) contains a C-terminal protease degradation tag. The tag is recognized by ClpC1 (light orange) which unfolds PanD for degradation by the ClpP protease (dark orange). Lower part: Binding of POA to PanD causes conformational changes resulting in increased exposure of the degradation tag and hence increased degradation of PanD by ClpC1-ClpP.