Abstract PR05: The effect of chemotherapy on HER2+ breast cancer heterogeneity measured by STAR-FISH: Detection of PIK3CA mutation and HER2 amplification at single-cell level in situ

Current therapies in HER2-positive breast cancer are effective in only a subset of cases and part of the resistance is attributed to single nucleotide mutation H1047R in PIK3CA. Conventional PIK3CA mutation detection methods require isolation of DNA from the tumor bulk, which requires relatively large amount of tissue and may not detect mutations in rare cancer cells.We developed a novel method, Specific-To-Allele PCR-FISH (STAR-FISH), which allows for in situ detection of point mutation and gene amplification at single cell level. The assay consists of in situ PCR steps with mutation specific primers, followed by hybridization of a fluorescently labeled DNA probe homologous to PCR primer overhangs and probes for genomic regions of interest. The STAR-FISH signals present in intact formalin-fixed paraffin embedded (FFPE) samples are imaged and quantified in each individual nucleus within a tissue, with false discovery rate at 0.001, which facilitates identification of sub-populations of cells with different genetic makeup. The method was validated against FACS, immunofluorescence, droplet digital PCR, and MassArray; high correlation of the results was observed (R2=0.901 -0.9037, p<0.001).We applied STAR-FISH for PIK3CA hot-spot mutation and HER2 amplification to FFPE samples of HER2 positive breast tumors from 22 patients. For each case a chemotherapy naïve core needle biopsy and a post-neoadjuvant chemotherapy sample upon tumor resection were collected. STAR-FISH a...
Source: Molecular Cancer Research - Category: Cancer & Oncology Authors: Tags: Tumor Heterogeneity (Intratumor and Intertumoral): Oral Presentations - Proffered Abstracts Source Type: research