A liquid biopsy for bronchopulmonary/lung carcinoid diagnosis

Abstract

No effective blood biomarker exists to detect and clinically manage bronchopulmonary (BP) neuroendocrine tumors (NET). We developed a blood-based 51 NET-specific transcript set for diagnosis and monitoring and evaluated clinical performance metrics. It accurately diagnosed the tumor and differentiated stable from progressive disease as determined by RECIST criteria. Gene expression was evaluated in: a) publicly available BPNET transcriptomes (GSE35679); b) two BPNET cell-lines; and c) BPNET tissue with paired blood (n = 7). Blood gene expression was assessed in 194 samples including controls, benign lung diseases, malignant lung diseases and small bowel NETs. A separate validation study in 25 age- and gender-matched BPNETs/controls was performed. Gene expression measured by real-time PCR was scored (0–100%; normal: < 14%). Regression analyses, Principal Component Analysis (PCA), hierarchical clustering, Fisher's and non-parametric evaluations were undertaken. All 51 genes were identified in BPNET transcriptomes, tumor samples and cell-lines. Significant correlations were evident between paired tumor and blood (R2:0.63–0.91, p < 0.001). PCA and hierarchical clustering identified blood gene expression was significantly different between lung cancers and benign diseases, including BPNETs. Gene expression was highly correlated (R2: 0.91, p = 1.7 × 10-15) between small bowel and BPNET. For validation, all 25 BPNETs were positive compared to 20% controls (p < 0.0001). Scores were significantly elevated (p < 0.0001) in BPNETs (57 ± 28%) compared to controls (4 ± 5%). BPNETs with progressive disease (85 ± 11%) exhibited higher scores than stable disease (32 ± 7%, p < 0.0001). Blood measurements accurately diagnosed bronchopulmonary carcinoids, distinguishing stable from progressive disease. This marker panel will have clinical utility as a diagnostic liquid biopsy able to define disease activity and progression in real-time.

NETest Liquid Biopsy Is Diagnostic of Lung Neuroendocrine Tumors and Identifies Progressive Disease

“…A comparison of these transcriptomes with BPC confirmed the similarity of matched tumors and lung neuroendocrine neoplasia cell cultures [16]. A correlation of these tumor transcripts in matched circulating blood accurately ( R 2 > 0.8) represented those identified in BPCs [16]. …”

“…This is a separate patient cohort than the one previously reported [16, 17]. We calculated diagnostic accuracy and metrics (AUROC, sensitivity, and specificity) for the ­NETest in each group.…”

“…Tumor tissue ( n = 20; Table 1) was collected at the time of surgery as previously described [16]. Samples were snap frozen in liquid nitrogen.…”

“…Details of the PCR methodology, mathematical analysis, and validation have been published in detail, comprising a 2-step protocol (RNA isolation/cDNA production and q-PCR) from EDTA-collected whole blood [16-19]. The assay is undertaken in a US clinically certified laboratory (Wren Laboratories CL-0704, CLIA 07D2081388).…”

“…The evaluation of gastrointestinal NET transcriptomes enabled the development of a liquid biopsy for small-bowel and pancreatic tumors [16]. A comparison of these transcriptomes with BPC confirmed the similarity of matched tumors and lung neuroendocrine neoplasia cell cultures [16].…”