Microsecond Simulation of the Proteoglycan-like Region of Carbonic Anhydrase IX and Design of Chemical Inhibitors Targeting pH Homeostasis in Cancer Cells

Arun John; Umashankar Vetrivel; Muthukumaran Sivashanmugam; Sulochana Konerirajapuram Natarajan

doi:10.1021/acsomega.9b04203

Microsecond Simulation of the Proteoglycan-like Region of Carbonic Anhydrase IX and Design of Chemical Inhibitors Targeting pH Homeostasis in Cancer Cells

ACS Omega. 2020 Feb 20;5(8):4270-4281. doi: 10.1021/acsomega.9b04203. eCollection 2020 Mar 3.

Authors

Arun John^{1

2}, Umashankar Vetrivel¹, Muthukumaran Sivashanmugam¹, Sulochana Konerirajapuram Natarajan³

Affiliations

¹ Centre for Bioinformatics, Kamalnayan Bajaj Institute for Research in Vision and Ophthalmology, Vision Research Foundation, Sankara Nethralaya, Chennai 600 006, Tamil Nadu, India.
² School of Chemical and Biotechnology, SASTRA Deemed University, Thanjavur, Tamil Nadu, India.
³ R.S. Mehta Jain Department of Biochemistry and Cell Biology, Kamalnayan Bajaj Institute for Research in Vision and Ophthalmology, Vision Research Foundation, Sankara Nethralaya, Chennai 600 006, Tamil Nadu, India.

Abstract

Carbonic anhydrase IX (CAIX) is a membrane-bound enzyme associated with tumor hypoxia and found to be over expressed in various tumor conditions. Targeting CAIX catalytic activity is proven to be efficient modality in modulating pH homeostasis in cancer cells. Proteoglycan-like (PG) region is unique to CAIX and is proposed to serve as an antenna enhancing the export of protons in conjunction with facilitated efflux of lactate ions via monocarboxylate transporters. Moreover, the PG region is also reported to contribute to the assembly and maturation of focal adhesion links during cellular attachment and dispersion on solid supports. Thus, drug targeting of this region shall efficiently modulate pH homeostasis and cell adhesion in cancer cells. As the PG region is intrinsically disordered, the complete crystal structure is not elucidated. Hence, in this study, we intend to sample the conformational landscape of the PG region at microsecond scale simulation in order to sample the most probable conformations that shall be utilized for structure-based drug design. In addition, the sampled conformations were subjected to high-throughput virtual screening against NCI and Maybridge datasets to identify potential hits based on consensus scoring and validation by molecular dynamics simulation. Further, the identified hits were experimentally validated for efficacy by in vitro and direct enzymatic assays. The results reveal 5-(2-aminoethyl)-1,2,3-benzenetriol to be the most promising hit as it showed significant CAIX inhibition at all levels of in silico and experimental validation.