Peripheral T-cell lymphoma comprises a heterogeneous group of mature non-Hodgkin lymphomas. Their diagnosis is challenging, with up to 30% of cases remaining unclassifiable and referred to as "not ...otherwise specified". We developed a reverse transcriptase-multiplex ligation-dependent probe amplification gene expression profiling assay to differentiate the main T-cell lymphoma entities and to study the heterogeneity of the "not specified" category. The test evaluates the expression of 20 genes, including 17 markers relevant to T-cell immunology and lymphoma biopathology, one Epstein-Barr virus-related transcript, and variants of
(G17V) and
(R172K/T). By unsupervised hierarchical clustering, our assay accurately identified 21 of 21 ALK-positive anaplastic large cell lymphomas, 16 of 16 extranodal natural killer (NK)/T-cell lymphomas, 6 of 6 hepatosplenic T-cell lymphomas, and 13 of 13 adult T-cell leukemia/lymphomas. ALK-negative anaplastic lymphomas (n=34) segregated into one cytotoxic cluster (n=10) and one non-cytotoxic cluster expressing Th2 markers (n=24) and enriched in
-rearranged cases. The 63 T
-derived lymphomas divided into two subgroups according to a predominant T
(n=50) or an enrichment in Th2 (n=13) signatures. We next developed a support vector machine predictor which attributed a molecular class to 27 of 77 not specified T-cell lymphomas: 17 T
, five cytotoxic ALK-negative anaplastic and five NK/T-cell lymphomas. Among the remaining cases, we identified two cell-of-origin subgroups corresponding to cytotoxic/Th1 (n=19) and Th2 (n=24) signatures. A reproducibility test on 40 cases yielded a 90% concordance between three independent laboratories. This study demonstrates the applicability of a simple gene expression assay for the classification of peripheral T-cell lymphomas. Its applicability to routinely-fixed samples makes it an attractive adjunct in diagnostic practice.
IDH1 and IDH2 somatic mutations have been identified in solid tumors and blood malignancies. The development of inhibitors of mutant IDH1 and IDH2 in the past few years has prompted the development ...of a fast and sensitive assay to detect IDH1R132, IDH2R140 and IDH2R172 mutations to identify patients eligible for these targeted therapies. This study aimed to compare two new multiplexed PCR assays – an automated quantitative PCR (qPCR) on the PGX platform and a droplet digital PCR (ddPCR) with next‐generation sequencing (NGS) for IDH1/2 mutation detection. These assays were evaluated on 102 DNA extracted from patient peripheral blood, bone marrow and formalin‐fixed paraffin‐embedded tissue samples with mutation allelic frequency ranging from 0.6% to 45.6%. The ddPCR assay had better analytical performances than the PGX assay with 100% specificity, 100% sensitivity and a detection limit down to 0.5% on IDH1R132, IDH2R140 and IDH2R172 codons, and a high correlation with NGS results. Therefore, the new highly multiplexed ddPCR is a fast and cost‐effective assay that meets most clinical needs to identify and follow cancer patients in the era of anti‐IDH1/2‐targeted therapies.
This study presents the first evaluation of two new highly multiplexed PCR assays as compared with next‐generation sequencing (NGS): an automated qPCR on the PGX platform and an allele‐specific droplet digital PCR (ddPCR) assay, both designed to detect most IDH1 and IDH2 hotspot mutations. The ddPCR assay was the best assay in terms of sensitivity, specificity, robustness, cost and timeliness, regardless of patient sample processing. It is, therefore, an appealing alternative to NGS to screen patients eligible for IDH1/2 targeted therapies.
Exonucleasic domain POLE (edPOLE) mutations, which are responsible for a hypermutated tumor phenotype, occur in 1–2% of colorectal cancer (CRC) cases. These alterations represent an emerging ...biomarker for response to immune checkpoint blockade. This study aimed to assess the molecular characteristics of edPOLE‐mutated tumors to facilitate patient screening. Based on opensource data analysis, we compared the prevalence of edPOLE mutations in a control group of unselected CRC patients (n = 222) vs a group enriched for unusual BRAF/RAS mutations (n = 198). Tumor mutational burden (TMB) and immune infiltrate of tumors harboring edPOLE mutations were then analyzed. In total, 420 CRC patients were analyzed: 11 edPOLE‐mutated tumors were identified, most frequently in microsatellite (MMR)‐proficient young (< 70 years) male patients, with left‐sided tumors harboring noncodon 12 KRAS mutation. The prevalence of edPOLE‐mutated tumors in the control vs the experimental screening group was, respectively, 0.45% (n = 1) vs 5.0% (n = 10). Among the 11 edPOLE‐mutated cases, two had a low TMB, three were hypermutated, and six were ultramutated. EdPOLE‐mutated cases had a high CD8+ tumor‐infiltrating lymphocyte (TIL) infiltration. These clinicopathological and molecular criteria may help to identify edPOLE mutations associated with a high TMB in CRC, and improve the selection of patients who could benefit from immunotherapy.
Exonucleasic domain POLE (edPOLE) mutations, which occur in 1–2% of colorectal cancer, represent an emerging biomarker for response to immunotherapy. This study aimed to assess the molecular characteristics of edPOLE‐mutated tumors. Results show that the prevalence of edPOLE‐mutated tumors is higher in microsatellite‐proficient young Male patients harboring non‐codon 12 KRAS mutations. EdPOLE‐mutated cases had a high CD8+ lymphocyte infiltration.
•Colorectal cancer liver metastases (CRCLM) should be resected when feasible•Preoperative choice of intentional microscopic incomplete CRCLM resection is valid•Such surgical strategy must be provided ...to highly selected patients
Clinical outcomes of colorectal cancer (CRC) patients after an incomplete microscopic (R1) resection of liver metastases may not differ from those following a microscopically margin negative (R0) resection, when the latest is not feasible because of anatomic issues. We aimed at comparing the clinical outcomes of CRC patients with an intentional R1 or with a R0 resection of liver metastases.
All patients with advanced in CRC and liver metastases consecutively treated by liver resection between February 2005 and January 2019 at in the department of Digestive and Hepatobiliary Surgery of Henri Mondor University Hospital (Créteil, France) were included in this retrospective case-control study. Overall survival (OS) and event-free survival (EFS) were compared between patients who underwent an intentional (pre-operative decision) R1 resection (iR1) to those who had a R0 resection of liver metastases. To account for confounding, comparison between the 2 groups was performed after adjustment using propensity score analysis.
Twenty-six CRC patients treated by iR1 resection of liver metastases were compared to 98 patients treated by R0 resection. Median OS reached 39 months 95% confidence interval (CI): 25-67 and 63 months 95% CI: 52-76 in the iR1 and R0 groups, respectively. After adjustment by inverse probability of treatment weighting, patients’ OS and EFS did not differ significantly between the iR1 and R0 groups (hazard ratio (HR): 1.19 0.54-2.62 and 1.67 0.93-3.03), respectively.
iR1 resection of liver metastases in advanced CRC patients is an acceptable therapeutic strategy, when R0 resection is not feasible.
The Epstein-Barr virus (EBV) is associated with various lymphoproliferative disorders and lymphomas. We have previously demonstrated that treating wild-type TP53-expressing B cell lines with the TP53 ...pathway activator nutlin-3 induced apoptosis in EBV-negative and EBV-positive latency I cells whereas EBV-positive latency III cells remained much more apoptosis-resistant. Here, we report a constitutively high level of autophagy in these resistant cells which express high levels of the proautophagic protein BECN1/Beclin 1 based, at least in part, on the activation of the NFKB signaling pathway by the viral protein LMP1. Following treatment with nutlin-3, several autophagy-stimulating genes were upregulated both in EBV-negative and EBV-positive latency III cells. However the process of autophagy was only triggered in the latter and was associated with an upregulation of SESN1/sestrin 1 and inhibition of MTOR more rapid than in EBV-negative cells. A treatment with chloroquine, an inhibitor of autophagy, potentiated the apoptotic effect of nutlin-3, particularly in those EBV-positive cells which were resistant to apoptosis induced by nutlin-3 alone, thereby showing that autophagy participates in this resistant phenotype. Finally, using immunohistochemical staining, clinical samples from various B cell lymphoproliferations with the EBV-positive latency II or III phenotype were found to harbor a constitutively active autophagy.
In the spectrum of colorectal tumors, microsatellite-stable (MSS) tumors with DNA polymerase ε (POLE) mutations exhibit a hypermutated profile, holding the potential to respond to immunotherapy ...similarly to their microsatellite-instable (MSI) counterparts. Yet, due to their rarity and the associated testing costs, systematic screening for these mutations is not commonly pursued. Notably, the histopathological phenotype resulting from POLE mutations is theorized to resemble that of MSI. This resemblance not only could facilitate their detection by a transformer-based Deep Learning (DL) system trained on MSI pathology slides, but also indicates the possibility for MSS patients with POLE mutations to access enhanced treatment options, which might otherwise be overlooked. To harness this potential, we trained a Deep Learning classifier on a large dataset with the ground truth for microsatellite status and subsequently validated its capabilities for MSI and POLE detection across three external cohorts. Our model accurately identified MSI status in both the internal and external resection cohorts using pathology images alone. Notably, with a classification threshold of 0.5, over 75% of POLE driver mutant patients in the external resection cohorts were flagged as "positive" by a DL system trained on MSI status. In a clinical setting, deploying this DL model as a preliminary screening tool could facilitate the efficient identification of clinically relevant MSI and POLE mutations in colorectal tumors, in one go.
Skin biopsies of 41 angioimmunoblastic T-cell lymphoma patients were retrospectively analyzed for the expression of follicular helper T-cell (TFH) markers, Epstein-Barr virus (EBV), and the presence ...of RHOA (p.G17V) and IDH2 (p.R172K/S) mutations using allele-specific polymerase chain reaction. We categorized cases into 4 distinctive patterns: (1) low-density lymphocytic perivascular infiltrates (n=11), (2) dense perivascular infiltrates with atypical cells and occasional inflammatory cells (n=13), (3) diffuse infiltrates reminiscent of angioimmunoblastic T-cell lymphoma (n=4), or (4) other aspects (n=13). Two EBV and 2 plasmacytoid lymphoproliferative disorders were seen. We observed variable expression of TFH markers (CD10 50%, BCLB6 84%, PD1 94%, CXCL13 84%, and ICOS 97.5%), and EBV B-blasts (26%). A TFH phenotype was identified in 82% and 73%, respectively, of cases with the most challenging patterns 1 and 2. TFH markers and EBV can thus help for diagnosis and are detected in samples with low-density infiltrates. We found RHOA G17V and IDH2 R172K/S mutations in the skin in 14/18 (78%) and 3/16 (19%) cases, respectively. The RHOA G17V mutation was identified in a proportion of biopsies with patterns 1 and 2, which represent a diagnostic challenge. The RHOA G17V mutation was detected both in the skin and lymph node (LN) biopsies in 7/9 (64%) cases, and in only the skin or the LN of 1 sample each. The frequency of RHOA G17V mutation was similar to that reported in LNs. It may represent a sensitive diagnostic marker in the skin, helpful in cases with low-density infiltrates.
Anaplastic lymphoma kinase (ALK) rearrangement is reported in 3% to 8% of patients with lung adenocarcinoma and can be detected by fluorescent in situ hybridization (FISH) or indirectly by ...immunohistochemistry. In FISH assay, isolated 5′ signal (loss of 3′ signal) is usually considered negative. We report three young nonsmoking patients with stage IV lung adenocarcinoma. Strong ALK expression in tumor cells detected by immunohistochemistry was observed in all cases, but FISH revealed an isolated 5′ signal pattern. Massive parallel “next-generation” sequencing was performed in two patients and confirmed ALK rearrangement. The three patients were treated and responded to crizotinib after 14, 10, and 31 months.
Aims
Bile duct adenomas (BDA) and bile duct hamartomas (BDH) are benign bile duct lesions considered neoplastic or secondary to ductal plate malformation, respectively. We have reported previously a ...high prevalence of BRAF V600E mutations detected by allele‐specific polymerase chain reaction assay in BDA, and suggested that BDA may be precursors to a subset of intrahepatic cholangiocarcinomas harbouring V600E mutations. The aim of the present study was to assess the existence of BRAF V600E mutations, using immunohistochemical methods, in additional BDA as well as in BDH.
Methods and results
Fifteen BDA and 35 BDH were retrieved from the archives of the pathology departments of two French university hospitals. All cases were reviewed by two pathologists specialized in liver diseases. BRAF V600E mutational status was investigated by immunohistochemistry. Mutated BRAF mutant protein was detected in 53% of the BDA and in none of the cases of BDH.
Conclusion
Our findings suggest that BDA and BDH are different processes, and that BDA represent true benign neoplasms. They also support the hypothesis that mutated BDA might precede the development of the subset of intrahepatic cholangiocarcinomas harbouring BRAF V600E mutations.
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