Author ORCID Identifier

https://orcid.org/0000-0002-8141-4760

Date of Graduation

12-2021

Document Type

Thesis (MS)

Program Affiliation

Experimental Therapeutics

Degree Name

Masters of Science (MS)

Advisor/Committee Chair

Kendra Carmon

Committee Member

Qingyun (Jim) Liu

Committee Member

Ali Azhdarinia

Committee Member

Eduardo Vilar-Sanchez

Committee Member

Venkata Lokesh Battula

Abstract

LGR5 Regulation of STAT3 Signaling and Drug Resistance in Colorectal Cancer

Tressie Alexandra Capri Posey B.S.

Advisory Professor: Kendra Carmon, Ph.D.

The greatest difficulty in treating colorectal cancer (CRC) is the development of drug resistance which leads to relapse after treatment and progression to metastasis. Cancer stem cells (CSCs) are believed to drive relapse because of their capacity to self-renew, acquire resistance mechanisms, and differentiate promoting tumor growth and heterogeneity. Leucine-rich repeat-containing G protein-coupled receptor 5 (LGR5), is a bona-fide marker of CSCs and has been considered a viable target for CSC specific therapeutic development. While we showed targeting LGR5 with antibody-drug conjugates (ADCs) led to tumor regression in CRC xenografts, once treatment was ceased the tumor ultimately relapsed, possibly due to the reemergence of LGR5+ CSCs. Recent studies have shown that LGR5+ CSCs undergo plasticity converting from an LGR5+ state to a more chemo-resistant LGR5- state which can lead to relapse. Recently, we reported that loss of LGR5 expression, through shRNA knockdown (KD) or CRISPR knockout (KO) in multiple CRC cell lines, increased both chemotherapy resistance and proliferation, however the complete mechanisms by which this occurs is unknown. Through comprehensive western analysis, we discovered that loss of LGR5 expression increased phosphorylation of STAT3(Y705) and MET(Y1234/1235). Interestingly, we also found that LGR5+ CRC cells treated with irinotecan or LGR5-targeted ADCs, converted these cells to an LGR5- state with increased MET/STAT3 activation. LGR5 overexpression in a CRC cell line which doesn’t endogenously express LGR5, resulted in decreased phosphorylation of MET/STAT3 and enhanced sensitivity to chemotherapy. We next showed that LGR5- CRC cells are dependent on MET/STAT3 signaling for their acquired drug resistance and blockade of this pathway through chemical inhibition can be used to enhance chemotherapy and ADC efficacy in vitro. Also, combination treatment with STAT3 inhibitor, stattic, and irinotecan in vivo resulted in increased tumor inhibition and survival compared to either monotherapy alone. Our findings suggest that loss of LGR5 expression in CRC cells promotes drug resistance, at least in part, through the activation of MET/STAT3 signaling and thus inhibition of STAT3 activity may restore drug sensitivity of LGR5- CRC cells. Co-targeting LGR5+ CSCs along with molecular mechanisms that drive drug resistance and cancer cell plasticity may be essential for a more effective CRC treatment.

Keywords

Colorectal Cancer, LGR5, STAT3, Stem Cell Plasticity, Drug Resistance, In Vitro, In Vivo

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