Analytic validation of an assay to detect androgen receptor splice variant ARv7 protein expression on circulating tumor cells from prostate cancer patients
An immunofluorescence assay that scores ARv7 on circulating tumor cells is only interpretable once its own sensitivity, specificity and repeatability are measured; this AACR 2020 poster reports that analytic validation, cell line by cell line, against ARv7-high, ARv7-low and ARv7-negative cells spiked into blood from healthy normal donors.
Presented by RareCyte and the University of Washington.
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- Analytical performance was measured cell line by cell line against a single MFI threshold of 100, chosen to hold specificity at a minimum of 0.9. Specificity on the ARv7-negative BT-474 line was 0.976; on the ARv7-high 22Rv1 line sensitivity reached 0.832, while on the ARv7-low LNCaP line sensitivity fell to 0.245. These are analytical figures measured on cell lines spiked into blood from healthy normal donors, not clinical performance in patients.
- Repeatability was measured across three separate stainer runs of 7 slide replicates each. On the ARv7-high 22Rv1 line the mean ARv7-positive rate across runs was 83.4% with an inter-stainer CV of 3.0%, and mean ARv7 MFI was 460 with an inter-stainer CV of 10.8%; the ARv7-negative BT-474 line averaged 3.5% positive across the same three runs.
- In a small clinical comparison, the ARv7 assay found at least as many CTCs as the base CTC detection assay used as the gold standard. Across three prostate cancer patients the gold standard counted 1, 0 and 3 CTCs against the ARv7 assay’s 1, 0 and 4, totalling 4 and 5; the authors state that a larger clinical study will be required to determine the clinical cutoff for ARv7.
