A major problem in cancer chemotherapy is the existence of primary resistance and/or the acquisition of secondary resistance. Many cellular defects contribute to chemoresistance, but epigenetic ...changes can also be a cause.
A DNA methylation microarray was used to identify epigenetic differences in oxaliplatin-sensitive and -resistant colorectal cancer cells. The candidate gene SRBC was validated by single-locus DNA methylation and expression techniques. Transfection and short hairpin experiments were used to assess oxaliplatin sensitivity. Progression-free survival (PFS) and overall survival (OS) in metastasic colorectal cancer patients were explored with Kaplan-Meier and Cox regression analyses. All statistical tests were two-sided.
We found that oxaliplatin resistance in colorectal cancer cells depends on the DNA methylation-associated inactivation of the BRCA1 interactor SRBC gene. SRBC overexpression or depletion gives rise to sensitivity or resistance to oxaliplatin, respectively. SRBC epigenetic inactivation occurred in primary tumors from a discovery cohort of colorectal cancer patients (29.8%; n = 39 of 131), where it predicted shorter PFS (hazard ratio HR = 1.83; 95% confidence interval CI = 1.15 to 2.92; log-rank P = .01), particularly in oxaliplatin-treated case subjects for which metastasis surgery was not indicated (HR = 1.96; 95% CI = 1.13 to 3.40; log-rank P = .01). In a validation cohort of unresectable colorectal tumors treated with oxaliplatin (n = 58), SRBC hypermethylation was also associated with shorter PFS (HR = 1.90; 95% CI = 1.01 to 3.60; log-rank P = .045).
These results provide a basis for future clinical studies to validate SRBC hypermethylation as a predictive marker for oxaliplatin resistance in colorectal cancer.
Despite the remarkable activity of BTK inhibitors (BTKi) in relapsed B-cell non-Hodgkin lymphoma (B-NHL), no clinically-relevant biomarker has been associated to these agents so far. The relevance of ...phosphoproteomic profiling for the early identification of BTKi responders remains underexplored.
A set of six clinical samples from an ongoing phase I trial dosing patients with chronic lymphocytic leukemia (CLL) with TG-1701, a novel irreversible and highly specific BTKi, were characterized by phosphoproteomic and RNA sequencing (RNA-seq) analysis. The activity of TG-1701 was evaluated in a panel of 11 B-NHL cell lines and mouse xenografts, including two NF-κB- and BTK
-driven BTKi-resistant models. Biomarker validation and signal transduction analysis were conducted through real-time PCR, Western blot analysis, immunostaining, and gene knockout (KO) experiments.
A nonsupervised, phosphoproteomic-based clustering did match the early clinical outcomes of patients with CLL and separated a group of "early-responders" from a group of "late-responders." This clustering was based on a selected list of 96 phosphosites with Ikaros-pSer442/445 as a potential biomarker for TG-1701 efficacy. TG-1701 treatment was further shown to blunt Ikaros gene signature, including
and
, in early-responder patients as well as in BTKi-sensitive B-NHL cell lines and xenografts. In contrast, Ikaros nuclear activity and signaling remained unaffected by the drug
and
in late-responder patients and in BTK
, BTK
, and noncanonical NF-κB models.
These data validate phosphoproteomic as a valuable tool for the early detection of response to BTK inhibition in the clinic, and for the determination of drug mechanism of action.
The majority of mismatch repair (MMR) gene mutations causing Lynch syndrome (LS) occur either in MLH1 or MSH2. However, the relative contribution of PMS2 is less well defined. The aim of this study ...was to evaluate the role of PMS2 in LS by assessing the pathogenicity of variants of unknown significance (VUS) detected in the mutational analysis of PMS2 in a series of Spanish patients.
From a cohort of 202 LS suspected patients, 13 patients showing loss of PMS2 expression in tumours were screened for germline mutations in PMS2, using a long range PCR based strategy and multiplex ligation dependent probe amplification (MLPA). Pathogenicity assessment of PMS2 VUS was performed evaluating clinicopathological data, frequency in control population and in silico and in vitro analyses at the RNA and protein level.
Overall 25 different PMS2 DNA variants were detected. Fourteen were classified as polymorphisms. Nine variants were classified as pathogenic: seven alterations based on their molecular nature and two after demonstrating a functional defect (c.538-3C>G affected mRNA processing and c.137G>T impaired MMR activity). The c.1569C>G variant was classified as likely neutral while the c.384G>A remained as a VUS. We have also shown that the polymorphic variant c.59G>A is MMR proficient.
Pathogenic PMS2 mutations were detected in 69% of patients harbouring LS associated tumours with loss of PMS2 expression. In all, PMS2 mutations account for 6% of the LS cases identified. The comprehensive functional analysis shown here has been useful in the classification of PMS2 VUS and contributes to refining the role of PMS2 in LS.
Background & aims: there are few validated biomarkers that can be used to predict outcomes for patients with colorectal cancer. Part of the challenge is the genetic and molecular heterogeneity of ...colorectal tumors not only among patients, but also within tumors. We have explored intratumor heterogeneity at the epigenetic level, due to its dynamic nature. We analyzed DNA methylation profiles of the digestive tract surface and the central bulk and invasive front regions of colorectal tumors. Methods: we determined the DNA methylation profiles of >450,000 CpG sites in 3 macrodissected regions of 79 colorectal tumors and 23 associated liver metastases, obtained from 2 hospitals in Spain. We also analyzed samples for KRAS and BRAF mutations, 499,170 single nucleotide polymorphisms, and performed immunohistochemical analyses. Results: we observed differences in DNA methylation among the 3 tumor sections; regions of tumor−host interface differed the most from the other tumor sections. Interestingly, tumor samples collected from areas closer to the gastrointestinal transit most frequently shared methylation events with metastases. When we calculated individual coefficients to quantify heterogeneity, we found that epigenetic homogeneity was significantly associated with short time of relapse-free survival (log-rank P = .037) and short time of overall survival (log-rank P = .026) in patients with locoregional colorectal cancer. Conclusions: in an analysis of 79 colorectal tumors, we found significant heterogeneity in patterns of DNA methylation within each tumor; the level of heterogeneity correlates with times of relapse-free and overall survival.
CD44 is a polymorphic family of cell adhesion molecules that seems to be instrumental in the mechanism of tumor invasion and metastasis. Tumor cell expression of CD44, or lack thereof, may be one of ...the factors conditioning the highly disparate ability to penetrate the brain extracellular matrix (ECM) exhibited by glioblastoma multiforme (GM) and conventional meningioma. To assess the presence of CD44 in these two tumor types we have immunohistochemically investigated the expression of CD44 standard form (CD44s) and the variant isoforms containing the domain encoded by variant exon 3 (CD44v3) and variant exon 6 (CD44v6) in paraffin-embedded tissue from 10 conventional meningiomas and 10 GMs. A CD44s−/CD44v− phenotype was discerned in the meningioma cases, whereas GMs featured a CD44s+/CD44v− expression profile. Consequently, the growth patterns of meningioma and GM seem to be, at least in part, a reflection of their CD44 expression status. Paucity of CD44 in meningioma cells would render them unable to infiltrate the brain ECM, whereas CD44-rich glioma cells would successfully migrate through it. Conversely, lack of CD44v expression would contribute to explain the lack of metastatic potential characterizing both conventional meningioma and GM.
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