ORCID

https://orcid.org/0000-0002-8381-1620

Date of Award

2026

Document Type

Dissertation

Degree Name

Philosophy (Ph.D)

Department

Pharmaceutical Sciences

First Advisor

Zhe-Sheng Chen

Second Advisor

Sandra E. Reznik

Third Advisor

Xingguo Cheng

Abstract

Multidrug resistance (MDR) is a major barrier to effective chemotherapy, largely driven by the overexpression of the ATP binding cassette transporter ABCG2, which actively extrudes anticancer drugs and reduces intracellular drug accumulation. In this study, we examined whether vactosertib, a potent ALK5 (TGF β type I receptor) inhibitor, could overcome ABCG2 mediated MDR and explored its underlying mechanisms. Using drug selected resistant cell lines (NCI H460/TPT10, S1 M1 80) and ABCG2 transfected HEK293 variants, we found that vactosertib at non-toxic concentrations markedly restored sensitivity to mitoxantrone and topotecan, achieving a reversal effect comparable to Ko 143. In a long-term 3D multicellular tumor spheroid model, vactosertib induced mitoxantrone mediated spheroid shrinkage and reduced the viable cell fraction from approximately 70% to below 20% in resistance cell line, demonstrating strong efficacy in a physiologically relevant environment. Mechanistic studies revealed that vactosertib significantly increased intracellular drug retention by inhibiting ABCG2 mediated efflux, consistent with molecular docking predictions showing high affinity binding within the transporter cavity and ATPase assays indicating a characteristic biphasic modulation pattern. Beyond functional inhibition, vactosertib also downregulated ABCG2 mRNA and protein levels in a concentration- and time-dependent manner without altering membrane localization, suggesting transcriptional suppression rather than trafficking disruption. Importantly, vactosertib showed no effect on non ABCG2 substrates such as cisplatin and exhibited no reversal activity in ABCG2 knockout cells, confirming its selectivity. Together, these findings demonstrate that vactosertib effectively reverses ABCG2 mediated multidrug resistance through a dual mechanism involving direct transporter inhibition and sustained suppression of ABCG2 expression, supporting its potential as a promising ABCG2 targeted agent for combination cancer therapy.

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