Abstract PD2-08: Detecting subtype switching in metastatic breast cancer with circulating tumor DNA methylation profiling

Abstract Background: Subtype switching in metastatic breast cancer (BC), for example switching between ER positive and triple negative subtypes, is a well-recognized change in tumor phenotype which can be a major source of treatment failure if not identified. Guidelines1,2 recommend repeat tissue biopsy of metastasis to retest ER+/PR+/HER2+ status at recurrence. Yet, such biopsies may not always be clinically feasible, single-site tumour biopsies sample only a single metastasis, and subtype may change later during metastatic therapy. We developed a non-invasive blood test to monitor the evolution of metastatic BC subtype based on circulating tumor DNA (ctDNA) methylation profiling. Methods: A capture based enzymatic ctDNA based methylation profile previously described3 was used to develop three independent binary Machine learning classifiers to discriminate between ER positive v ER negative, and HER-2 positive v HER-2 negative and TNBC vs non-TNBC from ctDNA methylation profiles. A 10-fold stratified nested cross-validation was used to assess classifier characteristics. Liquid biopsy subtype was compared with metastatic tissue biopsy subtype, in independent test folds, split by whether the subtype was the same or different in the original primary cancer. Subtype was defined by immunohistochemistry (IHC). Results: There was a total of 191 metastatic liquid biopsy samples taken from 86 patients where multiple biopsies had been taken, with a median 63 months between first and repeat tissue biopsy (range 20-203 months). Of these 37/191 (19%) samples were from 17 patients that had switched subtype based on tissue. The liquid biopsy correctly identified the subtype switch in 89.2% (33/37) of samples, with plasma samples taken a median of 7 months from the tissue biopsy. Focussing on the samples that were initially ER+HER2-. Of the cancers that were ER+HER2- on both tissue biopsies, 96% (129/134) were liquid ER+HER2- and 0% (0/134) were liquid TNBC. Conversely of the cancers that were ER+HER2- on initial biopsy and TNBC on the later biopsy, only 9% (1/11) were liquid ER+HER2-, and 91% (10/11) were liquid TNBC including one sample that was mixed ER+/TNBC liquid subtype (p