TNP Analogues Inhibit the Virulence Promoting IP3-4 Kinase Arg1 in the Fungal Pathogen Cryptococcus neoformans

Desmarini Desmarini; Daniel Truong; Lorna Wilkinson-White; Chandrika Desphande; Mario Torrado; Joel P Mackay; Jacqueline M Matthews; Tania C Sorrell; Sophie Lev; Philip E Thompson; Julianne Teresa Djordjevic

doi:10.3390/biom12101526

TNP Analogues Inhibit the Virulence Promoting IP_3-4 Kinase Arg1 in the Fungal Pathogen Cryptococcus neoformans

Biomolecules. 2022 Oct 20;12(10):1526. doi: 10.3390/biom12101526.

Authors

Desmarini Desmarini^{1

2}, Daniel Truong³, Lorna Wilkinson-White⁴, Chandrika Desphande^{4

5}, Mario Torrado⁴, Joel P Mackay⁵, Jacqueline M Matthews⁵, Tania C Sorrell², Sophie Lev^{1

2}, Philip E Thompson³, Julianne Teresa Djordjevic^{1

2

6}

Affiliations

¹ Centre for Infectious Diseases and Microbiology, The Westmead Institute for Medical Research, Westmead, NSW 2145, Australia.
² Sydney Institute for Infectious Diseases, Faculty of Medicine and Health, University of Sydney, Sydney, NSW 2006, Australia.
³ Medicinal Chemistry, Monash Institute of Pharmaceutical Sciences, Monash University, 381 Royal Parade, Parkville, VIC 3052, Australia.
⁴ Sydney Analytical, Core Research Facilities, The University of Sydney, Sydney, NSW 2006, Australia.
⁵ School of Life & Environmental Sciences, The University of Sydney, Sydney, NSW 2006, Australia.
⁶ Western Sydney Local Health District, Westmead, NSW 2145, Australia.

Abstract

New antifungals with unique modes of action are urgently needed to treat the increasing global burden of invasive fungal infections. The fungal inositol polyphosphate kinase (IPK) pathway, comprised of IPKs that convert IP₃ to IP₈, provides a promising new target due to its impact on multiple, critical cellular functions and, unlike in mammalian cells, its lack of redundancy. Nearly all IPKs in the fungal pathway are essential for virulence, with IP_3-4 kinase (IP_3-4K) the most critical. The dibenzylaminopurine compound, N2-(m-trifluorobenzylamino)-N6-(p-nitrobenzylamino)purine (TNP), is a commercially available inhibitor of mammalian IPKs. The ability of TNP to be adapted as an inhibitor of fungal IP_3-4K has not been investigated. We purified IP_3-4K from the human pathogens, Cryptococcus neoformans and Candida albicans, and optimised enzyme and surface plasmon resonance (SPR) assays to determine the half inhibitory concentration (IC₅₀) and binding affinity (K_D), respectively, of TNP and 38 analogues. A novel chemical route was developed to efficiently prepare TNP analogues. TNP and its analogues demonstrated inhibition of recombinant IP_3-4K from C. neoformans (CnArg1) at low µM IC₅₀s, but not IP_3-4K from C. albicans (CaIpk2) and many analogues exhibited selectivity for CnArg1 over the human equivalent, HsIPMK. Our results provide a foundation for improving potency and selectivity of the TNP series for fungal IP_3-4K.

Keywords: Cryptococcus neoformans; IP3-4K; TNP; antifungal drug discovery; dibenzylaminopurine; enzyme assay; fungal pathogens; inositol polyphosphate kinase; structure activity relationship; surface plasmon resonance.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Animals
Antifungal Agents / chemistry
Candida albicans
Cryptococcosis* / drug therapy
Cryptococcosis* / microbiology
Cryptococcus neoformans*
Humans
Inositol / metabolism
Mammals
Purines / metabolism
Virulence

Substances

Antifungal Agents
Inositol
Purines

Grants and funding

This project was supported by seed grants from the University of Sydney Drug Discovery Institute and the Sydney Institute for Infectious Diseases, an Industry PhD scholarship from NSW Health and the University of Sydney and project grants to J.T. Djordjevic from the National Health and Medical Research Council (NHMRC) of Australia (APP1183939, APP1058779). Daniel Truong is a recipient of an RTP scholarship from the Australian Government.