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Molecular characterization of tumor cells from the cerebrospinal fluid and matched primary tumors from metastatic breast cancer patients with leptomeningeal disease

ABSTRACT: We purified tumor cells in the CSF (“CSFTC”) from 15 metastatic breast cancer patients diagnosed with leptomeningeal disease using a two-step method involving immunomagnetic enrichment and fluorescence-activated cell sorting (IE/FACS). Magnetic beads coated with mAb to the epithelial cell adhesion marker (EPCAM) were used to enrich for tumor cells and were further purified by FACS analysis. For DNA profiling, isolated CSFTC were subjected to molecular characterization through genome-wide copy number analyses. Genomic analyses were then compared with those performed on the corresponding archival primary tumors. For RNA profiling, isolated CSFTC were then subjected to molecular characterization through gene expression profiling via QPCR analysis of 64 cancer-related genes CGH: 17 CSFTC samples from 13 patients were successfully profiled ,1 patient had 5 timepoints, 6 of 13 patients had matched to copy number data archival tumors RNA: 18 samples from 5 patients had successful gene expression data of the 64 genes measured in triplicates. For non-tumor controls, 9 of the samples had matching gene expression data from sorted leuckocytes (+CD45 cells) obtained from the same draw.

ORGANISM(S): Homo sapiens

SUBMITTER: Magbanua MJ

PROVIDER: S-ECPF-GEOD-46068 | biostudies-other | 2013 Dec

REPOSITORIES: biostudies-other

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Project description:BackgroundBreast cancer brain metastases (BrMs) are defined by complex adaptations to both adjuvant treatment regimens and the brain microenvironment. Consequences of these alterations remain poorly understood, as does their potential for clinical targeting. We utilized genome-wide molecular profiling to identify therapeutic targets acquired in metastatic disease.MethodsGene expression profiling of 21 patient-matched primary breast tumors and their associated brain metastases was performed by TrueSeq RNA-sequencing to determine clinically actionable BrM target genes. Identified targets were functionally validated using small molecule inhibitors in a cohort of resected BrM ex vivo explants (n = 4) and in a patient-derived xenograft (PDX) model of BrM. All statistical tests were two-sided.ResultsConsiderable shifts in breast cancer cell-specific gene expression profiles were observed (1314 genes upregulated in BrM; 1702 genes downregulated in BrM; DESeq; fold change > 1.5, Padj < .05). Subsequent bioinformatic analysis for readily druggable targets revealed recurrent gains in RET expression and human epidermal growth factor receptor 2 (HER2) signaling. Small molecule inhibition of RET and HER2 in ex vivo patient BrM models (n = 4) resulted in statistically significantly reduced proliferation (P < .001 in four of four models). Furthermore, RET and HER2 inhibition in a PDX model of BrM led to a statistically significant antitumor response vs control (n = 4, % tumor growth inhibition [mean difference; SD], anti-RET = 86.3% [1176; 258.3], P < .001; anti-HER2 = 91.2% [1114; 257.9], P < .01).ConclusionsRNA-seq profiling of longitudinally collected specimens uncovered recurrent gene expression acquisitions in metastatic tumors, distinct from matched primary tumors. Critically, we identify aberrations in key oncogenic pathways and provide functional evidence for their suitability as therapeutic targets. Altogether, this study establishes recurrent, acquired vulnerabilities in BrM that warrant immediate clinical investigation and suggests paired specimen expression profiling as a compelling and underutilized strategy to identify targetable dependencies in advanced cancers.

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Molecular characterization of tumor cells from the cerebrospinal fluid and matched primary tumors from metastatic breast cancer patients with leptomeningeal disease

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