Pre-existence of invasive cells in breast ductal carcinoma in situ.

Abstract

10508 Background: The treatment of DCIS is controversial because its true malignant nature is unclear. Very little is known about factors that promote survival of breast ductal carcinoma in situ (DCIS) neoplastic cells within the duct or induce the emergence of invasive carcinoma. To address this question we have studied the malignant phenotype of fresh living human DCIS lesions. We reproduciblely cultured human epithelial cells and spheroids from pure fresh human DCIS lesions suggesting that invasive cells pre-exist within the ductal niche. Methods: We procured fresh living human DCIS lesions, obtained under consent after surgical therapy as leftover tissue not required for diagnosis. We characterized the DCIS cells by organ culture, xenograft transplantation, full genome molecular cytogenetics (300k SNPs, Illumina HumanCytoSNP profile), and signal pathway profiling (Reverse Phase Protein Microarray). Results: Ex vivo organoid culture of pure DCIS lesions, without enzymatic treatment or sorting, induced the emergence of neoplastic epithelial cells exhibiting the following characteristics: a) spontaneous generation of hundreds of spheroids and duct- like 3-D structures in culture within 2 weeks, b) tumorigenicity in NOD SCID mice, c) cytogenetically abnormal (copy number loss/gain in chromosomes including 1, 5, 6, 8, 13, 17) compared to the normal karyotype of the non-neoplastic cells in the source patient's breast tissue, d) in vitro migration and invasion of autologous breast stroma, and e) up-regulation of signal pathways linked to, and components of, cellular autophagy. Multiple autophagy markers were present in the patient's original DCIS lesion and mouse xenografts. Autophagy is a survival mechanism. Treatment with a lysosomotropic inhibitor (chloroquine phosphate) of autophagy completely suppressed the generation of DCIS spheroids/3-D structures, suppressed invasion of autologous stroma, induced apoptosis, suppressed autophagy and eliminated cytogenetically abnormal cells from the organ culture. Conclusions: These findings support the hypothesis that cytogenetically abnormal invasive cells pre-exist in human DCIS, but are held in check in the duct microenvironment. Autophagy constitutes a novel treatment target for DCIS. No significant financial relationships to disclose.