MRNA expression profiles of colorectal liver metastases as a novel biomarker for early recurrence after partial hepatectomy

Background: Identification of specific risk groups for recurrence after surgery for isolated colorectal liver metastases (CRLM) remains challenging due to the heterogeneity of the disease. Classical clinicopathologic parameters have limited prognostic value. The aim of this study was to identify a gene expression signature measured in CRLM discriminating early from late recurrence after partial hepatectomy. Methods: CRLM from two patient groups were collected: I) with recurrent disease ≤12 months after surgery (N = 33), and II) without recurrences and disease free for ≥36 months (N = 30). The patients were clinically homogeneous; all had a low clinical risk score (0-2) and did not receive (neo-) adjuvant chemotherapy. Total RNA was hybridised to Illumina arrays, and processed for analysis. A leave-one-out cross validation (LOOCV) analysis was performed to identify a prognostic gene expression signature. Results: LOOCV yielded an 11-gene profile with prognostic value in relation to recurrent disease ≤12 months after partial hepatectomy. This signature had a sensitivity of 81.8%, with a specificity of 66.7% for predicting recurrences (≤12 months) versus no recurrences for at least 36 months after surgery (X2 P < 0.0001). Conclusion: The current study yielded an 11-gene signature at mRNA level in CRLM discriminating early from late or no relapse after partial hepatectomy

Histopathological growth patterns of colorectal liver metastasis exhibit little heterogeneity and can be determined with a high diagnostic accuracy

Colorectal liver metastases (CRLM) exhibit distinct histopathological growth patterns (HGPs) that are indicative of prognosis following surgical treatment. This study aims to assess the reliability and replicability of this histological biomarker. Within and between metastasis HGP concordance was analysed in patients who underwent surgery for CRLM. An independent cohort was used for external validation. Within metastasis concordance was assessed in CRLM with ≥ 2 tissue blocks. Similarly, concordance amongst multiple metastases was determined in patients with ≥ 2 resected CRLM. Diagnostic accuracy [expressed in area under the curve (AUC)] was compared by number of blocks and number of metastases scored. Interobserver agreement (Cohen’s k) compared to the gold standard was determined for a pathologist and a PhD candidate witho

Salvage treatment for recurrences after first resection of colorectal liver metastases: the impact of histopathological growth patterns

The majority of patients recur after resection of colorectal liver metastases (CRLM). Patients with CRLM displaying a desmoplastic histopathological growth pattern (dHGP) have a better prognosis and lower probability of recurrence than patients with non-dHGP CRLM. The current study evaluates the impact of HGP type on the pattern and treatment of recurrences after first resection of CRLM. A retrospective cohort study was performed, including patients with known HGP type after complete resection of CRLM. All patients were treated between 2000 and 2015. The HGP was determined on the CRLM resected at first partial hepatectomy. The prognostic value of HGPs, in terms of survival outcome, in the current patient cohort were previously published. In total 690 patients were included, of which 492 (71%) developed recurrent disease. CRLM displaying dHGP were observed in 103 patients (21%). Amongst patients with dHGP CRLM diagnosed with recurrent disease, more liver-limited recurrences were seen (43% vs. 31%, p=0.030), whereas patients with non-dHGP more often recurred at multiple locations (34% vs. 19%, p=0.005). Patients with dHGP CRLM were more likely to undergo curatively intended local treatment for recurrent disease (adjusted odds ratio: 2.37; 95% confidence interval (CI) [1.46–3.84]; p<0.001) compared to patients with non-dHGP. The present study demonstrates that liver-limited disease recurrence after complete resection o

Semiautomated isolation and molecular characterisation of single or highly purified tumour cells from CellSearch enriched blood samples using dielectrophoretic cell sorting

Background: Molecular characterisation of single circulating tumour cells (CTCs) holds considerable promise for predictive biomarker assessment and to explore CTC heterogeneity. We evaluate a new method, the DEPArray system, that allows the dielectrophoretic manipulation and isolation of single and 100% purified groups of CTCs from pre-enriched blood samples and explore the feasibility of their molecular characterisation.Methods:Samples containing known numbers of two cell populations were used to assess cell loss during sample loading. Cultured breast cancer cells were isolated from spiked blood samples using CellSearch CTC and Profile kits. Single tumour cells and groups of up to 10 tumour cells were recovered with the DEPArray system and subjected to transcriptional and mutation analysis.Results:On average, 40% cell loss was observed when loading samples to the DEPArray system. Expected mutations in clinically relevant markers could be obtained for 60% of single recovered tumour cells and all groups of tumour cells. Reliable gene expression profiles were obtained from single cells and groups of up to 10 cells for 2 out of 3 spiked breast cancer cell lines.Conclusion:We describe a semiautomated workflow for the isolation of small groups of 1 to 10 tumour cells from whole blood samples and provide proof of principle for the feasibility of their comprehensive molecular characterisation

Enrichment of the tumour immune microenvironment in patients with desmoplastic colorectal liver metastasis

Background: Patients with resected colorectal liver metastasis (CRLM) who display only the desmoplastic histopathological growth pattern (dHGP) exhibit superior survival compared to patients with any non-desmoplastic growth (non-dHGP). The aim of this study was to compare the tumour microenvironment between dHGP and non-dHGP. Methods: The tumour microenvironment was investigated in three cohorts of chemo-naive patients surgically treated for CRLM. In cohort A semi-quantitative immunohistochemistry was performed, in cohort B intra

Angiogenic desmoplastic histopathological growth pattern as a prognostic marker of good outcome in patients with colorectal liver metastases

Abstract Background In patients with resectable colorectal liver metastases (CRLM), distinct histopathological growth patterns (HGPs) develop at the interface between the tumour and surrounding tissue. The desmoplastic (d) HGP is characterised by angiogenesis and a peripheral fibrotic rim, whereas non-angiogenic HGPs co-opt endogenous sinusoidal hepatic vasculature. Evidence from previous studies has suggested that patients with dHGP in their CRLM have improved prognosis as compared to patients with non-desmoplastic HGPs. However, these studies were relatively small and applied arbitrary cut-off values for the determination of the predominant HGP. We have now investigated the

Decreased expression of ABAT and STC2 hallmarks ER-positive inflammatory breast cancer and endocrine therapy resistance in advanced disease

Background: Patients with Estrogen Receptor α-positive (ER+) Inflammatory Breast Cancer (IBC) are less responsive to endocrine therapy compared with ER+ non-IBC (nIBC) patients. The study of ER+ IBC samples might reveal biomarkers for endocrine resistant breast cancer. Materials & methods: Gene expression profiles of ER+ samples from 201 patients were explored for genes that discriminated between IBC and nIBC. Classifier genes were applied onto clinically annotated expression data from 947 patients with ER+ breast cancer and validated with RT-qPCR for 231 patients treated with first-line tamoxifen. Relationships with metastasis-free survival (MFS) and progression-free survival (PFS) following adjuvant and first-line endocrine treatment, respectively, were investigated using Cox regression analysis. Results: A metagene of six genes including the genes encoding for 4-aminobutyrate aminotransferase (ABAT) and Stanniocalcin-2 (STC2) were identified to distinguish 22 ER+ IBC from 43 ER+ nIBC patients and remained discriminatory in an independent series of 136 patients. The metagene and two genes were not prognostic in 517 (neo)adjuvant untreated lymph node-negative ER+ nIBC breast cancer patients. Only ABAT was related to outcome in 250 patients treated with adjuvant tamoxifen. Three independent series of in total 411 patients with advanced disease showed increased metagene scores and decreased expression of ABAT and STC2 to be correlated with poor first-line endocrine therapy outcome. The biomarkers remained predictive for first-line tamoxifen treatment outcome in multivariate analysis including traditional factors or published signatures. In an exploratory analysis, ABAT and STC2 protein expression levels had no relation with PFS after first-line tamoxifen. Conclusions: This study utilized ER+ IBC to identify a metagene including ABAT and STC2 as predictive biomarkers for endocrine therapy resistance

Histopathological growth patterns as biomarker for adjuvant systemic chemotherapy in patients with resected colorectal liver metastases

Adjuvant systemic chemotherapy (CTx) is widely administered in patients with colorectal liver metastases (CRLM). Histopathological growth patterns (HGPs) are an independent prognostic factor for survival after complete resection. This study evaluates whether HGPs can predict the efectiveness of adjuvant CTx in patients with resected CRLM. Two main types of HGPs can be distinguished; the desmoplastic type and the non-desmoplastic type. Uni- and multivariable analyses for overall survival (OS) and disease-free survival (DFS) were performed, in both patients treated with and without preoperative chemotherapy. A total of 1236 patients from two tertiary centers (Memorial Sloan Kettering Cancer Center, New York, USA; Erasmus MC Cancer Institute, Rotterdam, The Netherlands) were included (period 2000–2016). A total of 656 patients (53.1%) patients received preoperative chemotherapy. Adjuvant CTx was only associated with a superior OS in non-desmoplastic patients that had not been pretreated (adjusted hazard ratio (HR) 0.52, 95% confdence interval (CI) 0.37–0.73, p<0.001), and not in desmoplastic patients (adjusted HR 1.78, 95% CI 0.75–4.21, p=0.19). In pretreated patients no signifcant efect of adjuvant CTx was observed, neither in the desmoplastic group (adjusted HR 0.83, 95% CI 0.49–1.42, p=0.50) nor in the non-desmoplastic group (adjusted HR 0.96, 95% CI 0.71–1.29, p=0.79). Similar results were found for DFS, with a superior DFS in non-desmoplastic patients treated with adjuvant CTx (HR 0.71, 95% CI 0.55–0.93, p<0.001) that were not pretreated. Adjuvant CTx seems to improve OS and DFS after resection of non-desmoplastic CRLM. However, this efect was only observed in patients that were not treated with chemotherap

Gene expression profiles associated with the presence of a fibrotic focus and the growth pattern in lymph node-negative breast cancer

Purpose: A fibrotic focus, the scar-like area found in the center of an invasive breast tumor, is a prognostic parameter associated with an expansive growth pattern, hypoxia, and (lymph) angiogenesis. Little is known about the molecular pathways involved. Experimental Design: Sixty-five patients were selected of whom microarray data of the tumor and H&E slides for histologic analysis were available. The growth pattern and the presence and size of a fibrotic focus were assessed. Differences in biological pathways were identified with global testing. The correlations of growth pattern and fibrotic focus with common breast cancer signatures and with clinicopathologic variables and survival were investigated. Results: Tumors with a large fibrotic focus showed activation of Ras signaling and of the hypoxia-inducible factor-1 α pathway. Furthermore, unsupervised hierarchical cluster analysis with hypoxia- and (lymph) angiogenesis-related genes showed that hypoxia-inducible factor-1 α vascular endothelial growth factor A, and carbonic anhydrase 9 were overexpressed. The presence of a fibrotic focus, especially a large fibrotic focus, was associated with the basal-like subtype (P = 0.009), an activated wound-healing signature (P = 0.06), and a poor-prognosis 76-gene signature (P = 0.004). The presence of a fibrotic focus (P = 0.02) and especially of a large fibrotic focus (P = 0.004) was also associated with early development of distant metastasis. Conclusions: Our results sustain the hypothesis that hypoxia-driven angiogenesis is essential in the biology of a fibrotic focus. Ras and Akt might play a role as downstream modulators. Our data furthermore suggest that vascular endothelial growth factor A does not only drive angiogenesis but also lymphangiogenesis in tumors with a fibrotic focus. Our data also show an association between the presence of a fibrotic focus and infaust molecular signatures

Landscape of somatic mutations in 560 breast cancer whole-genome sequences

We analysed whole-genome sequences of 560 breast cancers to advance understanding of the driver mutations conferring clonal advantage and the mutational processes generating somatic mutations. We found that 93 protein-coding cancer genes carried probable driver mutations. Some non-coding regions exhibited high mutation frequencies, but most have distinctive structural features probably causing elevated mutation rates and do not contain driver mutations. Mutational signature analysis was extended to genome rearrangements and revealed twelve base substitution and six rearrangement signatures. Three rearrangement signatures, characterized by tandem duplications or deletions, appear associated with defective homologous-recombination-based DNA repair: one with deficient BRCA1 function, another with deficient BRCA1 or BRCA2 function, the cause of the third is unknown. This analysis of all classes of somatic mutation across exons, introns and intergenic regions highlights the repertoire of cancer genes and mutational processes operating, and progresses towards a comprehensive account of the somatic genetic basis of breast cancer