Choice of antibody is critical for specific and sensitive detection of androgen receptor splice variant-7 in circulating tumor cells

Tanzila Khan; John G Lock; Yafeng Ma; David G Harman; Paul de Souza; Wei Chua; Bavanthi Balakrishnar; Kieran F Scott; Therese M Becker

doi:10.1038/s41598-022-20079-w

Choice of antibody is critical for specific and sensitive detection of androgen receptor splice variant-7 in circulating tumor cells

Sci Rep. 2022 Sep 28;12(1):16159. doi: 10.1038/s41598-022-20079-w.

Authors

Tanzila Khan^{1

2

3

4}, John G Lock^{5

6}, Yafeng Ma^{7

8

5

9}, David G Harman¹⁰, Paul de Souza^{7

8

5}, Wei Chua^{7

8

9

11}, Bavanthi Balakrishnar¹¹, Kieran F Scott^{7

8}, Therese M Becker^{12

13

14

15}

Affiliations

¹ School of Medicine, Western Sydney University, Campbelltown, NSW, 2560, Australia. 19153083@student.westernsydney.edu.au.
² Medical Oncology, Ingham Institute of Applied Medical Research, Liverpool, NSW, 2170, Australia. 19153083@student.westernsydney.edu.au.
³ Centre of Circulating Tumour Cells Diagnostics & Research, Ingham Institute for Applied Medical Research, Liverpool, NSW, 2170, Australia. 19153083@student.westernsydney.edu.au.
⁴ South Western Sydney Clinical School, University of New South Wales, Liverpool Hospital, Liverpool, NSW, 2170, Australia. 19153083@student.westernsydney.edu.au.
⁵ Centre of Circulating Tumour Cells Diagnostics & Research, Ingham Institute for Applied Medical Research, Liverpool, NSW, 2170, Australia.
⁶ School of Medical Sciences, University of New South Wales, Sydney, Australia.
⁷ School of Medicine, Western Sydney University, Campbelltown, NSW, 2560, Australia.
⁸ Medical Oncology, Ingham Institute of Applied Medical Research, Liverpool, NSW, 2170, Australia.
⁹ South Western Sydney Clinical School, University of New South Wales, Liverpool Hospital, Liverpool, NSW, 2170, Australia.
¹⁰ School of Science, Western Sydney University, Campbelltown, NSW, 2560, Australia.
¹¹ Department of Medical Oncology, Liverpool Hospital, Liverpool, NSW, 2170, Australia.
¹² School of Medicine, Western Sydney University, Campbelltown, NSW, 2560, Australia. therese.becker@inghaminstitute.org.au.
¹³ Medical Oncology, Ingham Institute of Applied Medical Research, Liverpool, NSW, 2170, Australia. therese.becker@inghaminstitute.org.au.
¹⁴ Centre of Circulating Tumour Cells Diagnostics & Research, Ingham Institute for Applied Medical Research, Liverpool, NSW, 2170, Australia. therese.becker@inghaminstitute.org.au.
¹⁵ South Western Sydney Clinical School, University of New South Wales, Liverpool Hospital, Liverpool, NSW, 2170, Australia. therese.becker@inghaminstitute.org.au.

Abstract

Androgen receptor variant 7 (AR-V7) is an important biomarker to guide treatment options for castration-resistant prostate cancer (CRPC) patients. Its detectability in circulating tumour cells (CTCs) opens non-invasive diagnostic avenues. While detectable at the transcript level, AR-V7 protein detection in CTCs may add additional information and clinical relevance. The aim of this study was to compare commercially available anti-AR-V7 antibodies and establish reliable AR-V7 immunocytostaining applicable to CTCs from prostate cancer (PCa) patients. We compared seven AR-V7 antibodies by western blotting and immmunocytostaining using a set of PCa cell lines with known AR/AR-V7 status. The emerging best antibody was validated for detection of CRPC patient CTCs enriched by negative depletion of leucocytes. The anti-AR-V7 antibody, clone E308L emerged as the best antibody in regard to signal to noise ratio with a specific nuclear signal. Moreover, this antibody detects CRPC CTCs more efficiently compared to an antibody previously shown to detect AR-V7 CTCs. We have determined the best antibody for AR-V7 detection of CTCs, which will open future studies to correlate AR-V7 subcellular localization and potential co-localization with other proteins and cellular structures to patient outcomes.

Publication types

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

MeSH terms

Cell Count
Humans
Male
Neoplastic Cells, Circulating* / pathology
Prostatic Neoplasms, Castration-Resistant* / diagnosis
Prostatic Neoplasms, Castration-Resistant* / drug therapy
Prostatic Neoplasms, Castration-Resistant* / genetics
Protein Isoforms / genetics
Protein Isoforms / metabolism
Receptors, Androgen / genetics
Receptors, Androgen / metabolism

Substances

Protein Isoforms
Receptors, Androgen