Genomic comparison of early-passage conditionally reprogrammed breast cancer cells to their corresponding primary tumors

Abstract

Conditionally reprogrammed cells (CRCs) are epithelial cells that are directly isolated from patients’ specimens and propagated in vitro with feeder cells and a Rho kinase inhibitor. A number of these cells have been generated from biopsies of breast cancer patients, including ductal carcinoma in situ and invasive carcinomas. The characterization of their genomic signatures is essential to determine their ability to reflect the natural biology of their tumors of origin. In this study, we performed the genomic characterization of six newly established invasive breast cancer CRC cultures in comparison to the original patients’ primary breast tumors (PBT) from which they derived. The CRCs and corresponding PBTs were simultaneously profiled by genome-wide array-CGH, targeted next generation sequencing and global miRNA expression to determine their molecular similarities in the patterns of copy number alterations (CNAs), gene mutations and miRNA expression levels, respectively. The CRCs’ epithelial cells content and ploidy levels were also evaluated by flow cytometry. A similar level of CNAs was observed in the pairs of CRCs/PBTs analyzed by array-CGH, with >95% of overlap for the most frequently affected cytobands. Consistently, targeted next generation sequencing analysis showed the retention of specific somatic variants in the CRCs as present in their original PBTs. Global miRNA profiling closely clustered the CRCs with their PBTs (Pearson Correlation, ANOVA paired test, P<0.05), indicating also similarity at the miRNA expression level; the retention of tumor-specific alterations in a subset of miRNAs in the CRCs was further confirmed by qRT-PCR. These data demonstrated that the human breast cancer CRCs of this study maintained at early passages the overall copy number, gene mutations and miRNA expression patterns of their original tumors. The further characterization of these cells by other molecular and cellular phenotypes at late cell passages, are required to further expand their use as a unique and representative ex-vivo tumor model for basic science and translational breast cancer studies.

Murine neuroblastoma cell lines developed by conditional reprogramming preserve heterogeneous phenotypes observed in vivo

“…27 In CR cultures from tumor tissues, phenotypic and genotypic features of the primary tumor are maintained. 26,28 Importantly, CR tumor cells have been used to identify an appropriate therapy for a pediatric HPV-positive lung cancer and to identify both genetic and non-genetic alterations in lung tumors that correlate with patient's drug resistance pattern. 47,48 Overall, CR has been successful in generating several types of human tumor models and other human and mouse cell cultures used for various research purposes.…”

“…27 CR induces cancer cells to grow at a rapid rate while retaining their tumorigenic properties and characteristic genetic features. 28 Overall, CR has been very successful in generating human tumor cell cultures derived from epithelial cancers. To our knowledge, this is the first demonstration of the use of this method to develop NB cell lines.…”

Conditional reprogramming: next generation cell culture

“…Mahajan et al. 148 characterized the molecular and cellular phenotypes of CR breast cancer cells and found that the CR cells represented the heterogeneity of primary breast tumor cells, making it a unique representative breast cancer model for further study. At present, there are many studies on female breast cancer, while only a few focused on molecular mechanisms for the development of male breast cancer.…”

Culture and application of conditionally reprogrammed primary tumor cells

“…These reprogrammed immortalized cells of malignant tumors [ 10 ], such as bladder cancer [ 11 ], prostate cancer [ 12 ], pancreatic cancer [ 13 ], breast carcinoma [ 14 ], and hepatocellular carcinoma [ 15 ], without genetic manipulation or chromosomal abnormalities, represent an adult stem-cell-like state but express fairly low levels of Sox2 , Oct4 , and Klf4 [ 16 ], which are the pluripotent progenitor markers [ 17 ]. What is more, these non-tumorigenic cells can maintain intra-tumor heterogeneity [ 18 ] in addition to keeping their molecular features [ 19 , 20 ], and are only capable of differentiating into the native tissues in which they originated [ 16 , 21 ]. Therefore, CR is appropriate to effectively assess tumor biology, screen potential therapeutic targets, and preclinically evaluate the efficiency of antitumor drugs.…”