EBV-Positive Inflammatory Follicular Dendritic Cell Sarcoma of the Spleen: Report of an Aggressive Form With Molecular Characterization.

EBV-positive inflammatory follicular dendritic cell sarcoma (EBV+ inflammatory FDCS) is a rare neoplasm almost exclusively located in the spleen or liver. It is characterized by a proliferation of EBV-positive spindle-shaped cells bearing follicular dendritic cell markers, associated with an abundant lymphoplasmacytic infiltrate. EBV+ inflammatory FDCS is often asymptomatic or responsible for mild symptoms. It usually displays an indolent course and its prognosis is excellent after tumor removal, although relapsing and metastatic forms exist. Herein, we describe an aggressive form of splenic EBV+ inflammatory FDCS in a 79-year-old woman presenting with abdominal pain, deterioration of general health status, major inflammatory syndrome, and symptomatic hypercalcemia. A splenectomy was performed leading to a rapid improvement in her clinical condition and normalization of laboratory abnormalities. Unfortunately, her symptoms and laboratory abnormalities reappeared 4 months later. Computed tomography showed a mass in the splenectomy site and multiple liver and peritoneal nodules. Further analyses were performed on tumor tissue and showed positive phospho-ERK staining of tumoral cells indicating activation of MAPK pathway. Inactivating mutations were found on CDKN2A and NF1 genes. Subsequently, the patient's condition deteriorated rapidly. Since interleukin-6 levels were dramatically increased, tocilizumab was used but only had a transient effect on the patient's symptoms and inflammatory syndrome. Antitumor agent gemcitabine was initiated but her clinical condition continued to deteriorate and the patient died 2 weeks later. The management of aggressive forms of EBV+ inflammatory FDCS remains challenging. However, since these tumors seem to display genetic alterations, better characterization could lead to molecular targeted therapies.

A Dysfunctional T-cell Gene Signature for Predicting Nonresponse to PD-1 Blockade in Non-small Cell Lung Cancer That Is Suitable for Routine Clinical Diagnostics.

PURPOSE: Because PD-1 blockade is only effective in a minority of patients with advanced-stage non-small cell lung cancer (NSCLC), biomarkers are needed to guide treatment decisions. Tumor infiltration by PD-1T tumor-infiltrating lymphocytes (TIL), a dysfunctional TIL pool with tumor-reactive capacity, can be detected by digital quantitative IHC and has been established as a novel predictive biomarker in NSCLC. To facilitate translation of this biomarker to the clinic, we aimed to develop a robust RNA signature reflecting a tumor's PD-1T TIL status. EXPERIMENTAL DESIGN: mRNA expression analysis using the NanoString nCounter platform was performed in baseline tumor samples from 41 patients with advanced-stage NSCLC treated with nivolumab that were selected on the basis of PD-1T TIL infiltration by IHC. Samples were included as a training cohort (n = 41) to develop a predictive gene signature. This signature was independently validated in a second cohort (n = 42). Primary outcome was disease control at 12 months (DC 12 m), and secondary outcome was progression-free and overall survival. RESULTS: Regularized regression analysis yielded a signature using 12 out of 56 differentially expressed genes between PD-1T IHC-high tumors from patients with DC 12 m and PD-1T IHC-low tumors from patients with progressive disease (PD). In the validation cohort, 6/6 (100%) patients with DC 12 m and 23/36 (64%) with PD were correctly classified with a negative predictive value (NPV) of 100% and a positive predictive value of 32%. CONCLUSIONS: The PD-1T mRNA signature showed a similar high sensitivity and high NPV as the digital IHC quantification of PD-1T TIL. This finding provides a straightforward approach allowing for easy implementation in a routine diagnostic clinical setting.

Retrospective analysis of real-world data to evaluate actionability of a comprehensive molecular profiling panel in solid tumor tissue samples (REALM study).

INTRODUCTION: Considering the growing interest in matched cancer treatment, our aim was to evaluate the ability of a comprehensive genomic profiling (CGP) assay to propose at least one targeted therapy given an identified genomic alteration or signature (actionability), and to collect the treatment modifications based on the CGP test results in clinical practise for solid tumors. METHODS: This retrospective, multicentre French study was conducted among 25 centres that participated in a free of charge program between 2017 and 2019 for a tissue CGP test. Data were collected on the patient, disease, tumor genomic profile, treatment suggested in the report (related to the genomic profile results) and subsequent therapeutic decisions according to the physician's declaration. RESULTS: Among the 416 patients, most had lung cancer (35.6%), followed by biliary tract cancer (11.5%) or rare cancers (11.1%); 75% had a metastatic disease. The actionability was 75.0% (95% CI [70.6%-78.9%]) for all patients, 85.1% and 78.4%, respectively in lung cancer and metastatic patients. After exclusion of clinical trial suggestions, the actionability decreased to 62.3% (95% CI [57.5%-66.8%]). Treatment modification based on the test results was observed in 17.3% of the patients and was more frequent in metastatic disease (OR = 2.73, 95% CI [1.31-5.71], p = 0.007). The main reasons for no treatment modification were poor general condition (33.2%) and stable disease or remission (30.2%). The genomic-directed treatment changes were performed mostly during the first six months after the CGP test, and interestingly a substantial part was observed from six to 24 months after the genomic profiling. CONCLUSION: This French study provides information on the real-life actionability of a CGP test based on tissue samples, and trends to confirm its utility in clinical practice across the course of the disease, in particularly for patients with lung cancer and/or advanced disease.

Detection of Nine Oncogenes Amplification in Lung and Colorectal Cancer Formalin-Fixed Paraffin-Embedded Tissue Samples using Combined Next-Generation Sequencing-Based Script and Digital Droplet Polymerase Chain Reaction.

INTRODUCTION: Gene copy number variations have theranostic impact and require reliable methods for their identification. We aimed to evaluate the reliability of combined next-generation sequencing (NGS) and digital droplet PCR (ddPCR) method for gene amplification evaluation. METHODS: We conducted a retrospective multicentric observational study. MET/ERBB2 amplifications were assessed in patients with lung or colorectal carcinoma (cohort A), from 2016 to 2020, by fluorescence in situ hybridization (FISH)/immunohistochemistry (IHC), NGS and ddPCR. NGS-based script and ddPCR were then used to detect amplifications of 7 additional oncogenes (EGFR, KRAS, BRAF, FGFR1, FGFR2, FGFR3, PIK3CA) in a cohort of patients (cohort B). RESULTS: 55 patients (9 control, 25 ERBB2-amplified and 21 MET-amplified) out of 3779 patients tested were included in cohort A. Correlation coefficient between NGS-based script and FISH/IHC results were .88 for MET (P < .001) and .89 (P < .001) for ERBB2. Using a threshold ratio of 1.56 with the NGS-based script, the sensitivity was 100% for both genes and the specificity 69% for MET and 90% for ERBB2, respectively. With an alternative 1.76 threshold, sensitivity was 94% for MET and 96% for ERBB2, while specificity was 85% for MET and 90% for ERBB2. Correlation coefficient between FISH and ddPCR ratio was .90 for MET and .88 for ERBB2. In both cohorts, NGS-based script and ddPCR results were significantly correlated regarding all genes (P < .001). CONCLUSION: Combined NGS-based script and ddPCR method is reliable and easily feasible for the detection of gene amplifications, providing useful data for guided therapy in cancer.

Acute Influenza Infection Promotes Lung Tumor Growth by Reprogramming the Tumor Microenvironment.

One billion people worldwide get flu every year, including patients with non-small cell lung cancer (NSCLC). However, the impact of acute influenza A virus (IAV) infection on the composition of the tumor microenvironment (TME) and the clinical outcome of patients with NSCLC is largely unknown. We set out to understand how IAV load impacts cancer growth and modifies cellular and molecular players in the TME. Herein, we report that IAV can infect both tumor and immune cells, resulting in a long-term protumoral effect in tumor-bearing mice. Mechanistically, IAV impaired tumor-specific T-cell responses, led to the exhaustion of memory CD8+ T cells and induced PD-L1 expression on tumor cells. IAV infection modulated the transcriptomic profile of the TME, fine-tuning it toward immunosuppression, carcinogenesis, and lipid and drug metabolism. Consistent with these data, the transcriptional module induced by IAV infection in tumor cells in tumor-bearing mice was also found in human patients with lung adenocarcinoma and correlated with poor overall survival. In conclusion, we found that IAV infection worsened lung tumor progression by reprogramming the TME toward a more aggressive state.

An interim analysis of a gestational weight gain intervention in military personnel and other TRICARE beneficiaries.

OBJECTIVE: Despite military fitness regulations, women in the military frequently experience overweight/obesity, excessive gestational weight gain (GWG), and the postpartum implications. This interim analysis of the Moms Fit 2 Fight study examines GWG outcomes among active-duty personnel and other TRICARE beneficiaries who received a stepped-care GWG intervention compared with those who did not receive a GWG intervention. METHOD: Participants (N = 430; 32% identified with an underrepresented racial group, 47% were active duty) were randomized to receive a GWG intervention or the comparison condition, which did not receive a GWG intervention. RESULTS: Retention was 88% at 32 to 36 weeks' gestation. Participants who received the GWG intervention gained less weight compared with those who did not (mean [SD] = 10.38 [4.58] vs. 11.80 [4.87] kg, p = 0.0056). Participants who received the intervention were less likely to have excessive GWG compared with those who did not (54.6% vs. 66.7%, p = 0.0241). The intervention effects were significant for participants who identified as White, but not for those of other racial identities. There were no significant differences between the conditions in maternal/neonatal outcomes. CONCLUSIONS: The intervention successfully reduced excessive GWG, particularly among participants who identified as White. Should this intervention be found cost-effective, it may be sustainably integrated throughout the military prenatal care system.

The strategic roles of four enzymes in the interconnection between metabolism and oncogene activation in non-small cell lung cancer: Therapeutic implications.

NSCLC is the leading cause of cancer mortality and represents a major challenge in cancer therapy. Intrinsic and acquired anticancer drug resistance are promoted by hypoxia and HIF-1α. Moreover, chemoresistance is sustained by the activation of key signaling pathways (such as RAS and its well-known downstream targets PI3K/AKT and MAPK) and several mutated oncogenes (including KRAS and EGFR among others). In this review, we highlight how these oncogenic factors are interconnected with cell metabolism (aerobic glycolysis, glutaminolysis and lipid synthesis). Also, we stress the key role of four metabolic enzymes (PFK1, dimeric-PKM2, GLS1 and ACLY), which promote the activation of these oncogenic pathways in a positive feedback loop. These four tenors orchestrating the coordination of metabolism and oncogenic pathways could be key druggable targets for specific inhibition. Since PFK1 appears as the first tenor of this orchestra, its inhibition (and/or that of its main activator PFK2/PFKFB3) could be an efficacious strategy against NSCLC. Citrate is a potent physiologic inhibitor of both PFK1 and PFKFB3, and NSCLC cells seem to maintain a low citrate level to sustain aerobic glycolysis and the PFK1/PI3K/EGFR axis. Awaiting the development of specific non-toxic inhibitors of PFK1 and PFK2/PFKFB3, we propose to test strategies increasing citrate levels in NSCLC tumors to disrupt this interconnection. This could be attempted by evaluating inhibitors of the citrate-consuming enzyme ACLY and/or by direct administration of citrate at high doses. In preclinical models, this "citrate strategy" efficiently inhibits PFK1/PFK2, HIF-1α, and IGFR/PI3K/AKT axes. It also blocks tumor growth in RAS-driven lung cancer models, reversing dedifferentiation, promoting T lymphocytes tumor infiltration, and increasing sensitivity to cytotoxic drugs.

Discovery and validation of a transcriptional signature identifying homologous recombination-deficient breast, endometrial and ovarian cancers.

BACKGROUND: Molecular alterations leading to homologous recombination deficiency (HRD) are heterogeneous. We aimed to identify a transcriptional profile shared by endometrial (UCEC), breast (BRCA) and ovarian (OV) cancers with HRD. METHODS: Genes differentially expressed with HRD genomic score (continuous gHRD score) in UCEC/BRCA/OV were identified using edgeR, and used to train a RNAseq score (ridge-regression model) predictive of the gHRD score (PanCanAtlas, N = 1684 samples). The RNAseq score was applied in independent gynaecological datasets (CARPEM/CPTAC/SCAN/TCGA, N = 4038 samples). Validations used ROC curves, linear regressions and Pearson correlations. Overall survival (OS) analyses used Kaplan-Meier curves and Cox models. RESULTS: In total, 656 genes were commonly up/downregulated with gHRD score in UCEC/BRCA/OV. Upregulated genes were enriched for nuclear/chromatin/DNA-repair processes, while downregulated genes for cytoskeleton (gene ontologies). The RNAseq score correlated with gHRD score in independent gynaecological cancers (R² = 0.4-0.7, Pearson correlation = 0.64-0.86, all P < 10-11), and was predictive of gHRD score >42 (RNAseq HRD profile; AUC = 0.95/0.92/0.78 in UCEC/BRCA/OV). RNAseq HRD profile was associated (i) with better OS in platinum-treated advanced TP53-mutated-UCEC (P < 0.001) and OV (P = 0.013), and (ii) with poorer OS (P < 0.001) and higher benefit of adjuvant chemotherapy in Stage I-III BRCA (interaction test, P < 0.001). CONCLUSIONS: UCEC/BRCA/OV with HRD-associated genomic scars share a common transcriptional profile. RNAseq signatures might be relevant for identifying HRD-gynaecological cancers, for prognostication and for therapeutic decision.

SMARCA4-deficient lung carcinoma is an aggressive tumor highly infiltrated by FOXP3+ cells and neutrophils.

INTRODUCTION: SMARCA4/BRG1 loss of expression occurs in 5-10% of non-small cell lung carcinomas (NSCLC). We investigated the pathological, molecular and immune environment characteristics of this deficiency among NSCLC, its impact on overall survival (OS) of resected patients and the sensitivity to anti-PD1 inhibitors in metastatic patients. MATERIALS AND METHODS: BRG1 expression was assessed by immunohistochemistry to identify SMARCA4-deficient NSCLC (SD-NSCLC) from the cancer tissue collection of Cochin Hospital (Paris, France). Molecular profiles were analyzed by targeted NGS covering 28 genes in 63 resected SD-NSCLC. The balance of immune cells between CD8+, FOXP3+ cells and neutrophils (CD66b+) was characterized by multiplex immunohistochemistry and compared to non-SD NSCLC. Clinical outcome after anti-PD-1 therapy was evaluated in 7 SD-NSCLC out of 77 NSCLC patients. RESULTS: SD-NSCLCs were more commonly found in TTF1-negative high-grade adenocarcinomas and pleomorphic carcinomas. They were associated with few targetable alterations (KRAS G12C and MET amplification). Their immune environment was characterized by an increased of FOXP3+ cell and neutrophil densities, but not of CD8+ T cells, compared to non-SD NSCLC. SD-NSCLC patients had a significantly shorter OS in early stages of resected patients and in metastatic patients treated by anti-PD1 treatment. CONCLUSION: BRG1-loss in NSCLC confers a poor prognosis and is associated with an immunosuppressive environment that could be responsible of limited efficacy to anti-PD1 inhibitors. The identification of SD-NSCLC by BRG1 immunohistochemistry is desirable for an optimal management of NSCLC patients.

Transcriptome in paraffin samples for the diagnosis and prognosis of adrenocortical carcinoma.

Design: Molecular classification is important for the diagnosis and prognosis of adrenocortical tumors (ACT). Transcriptome profiles separate adrenocortical adenomas 'C2' from carcinomas, and identify two groups of carcinomas 'C1A' and 'C1B', of poor and better prognosis respectively. However, many ACT cannot be profiled because of improper or absent freezing procedures, a mandatory requirement so far. The main aim was to determine transcriptome profiles on formalin-fixed paraffin-embedded (FFPE) samples, using the new 3'-end RNA-sequencing technology. A secondary aim was to demonstrate the ability of this technique to explore large FFPE archives, by focusing on the rare oncocytic ACT variants. Methods: We included 131 ACT: a training cohort from Cochin hospital and an independent validation cohort from Wuerzburg hospital. The 3' transcriptome was generated from FFPE samples using QuantSeq (Lexogen, Vienna, Austria) and NextSeq500 (Illumina, San Diego, CA, USA). Results: In the training cohort, unsupervised clustering identified three groups: 'C1A' aggressive carcinomas (n = 28, 29%), 'C1B' more indolent carcinomas (n = 28, 29%), and 'C2' adenomas (n = 39, 41%). The prognostic value of FFPE transcriptome was confirmed in the validation cohort (5-year OS: 26% in 'C1A' (n = 26) and 100% in 'C1B' (n = 10), P = 0.003). FFPE transcriptome was an independent prognostic factor in a multivariable model including tumor stage and Ki-67 (OS HR: 7.5, P = 0.01). Oncocytic ACT (n = 19) did not form any specific cluster. Oncocytic carcinomas (n = 6) and oncocytic ACT of uncertain malignant potential (n = 4) were all in 'C1B'. Conclusions: The 3' RNA-sequencing represents a convenient solution for determining ACT molecular class from FFPE samples. This technique should facilitate routine use and large retrospective studies.

Highly Specific Droplet-Digital PCR Detection of Universally Methylated Circulating Tumor DNA in Endometrial Carcinoma.

BACKGROUND: No circulating biomarker is available for endometrial carcinoma (EC). We aimed to identify DNA positions universally hypermethylated in EC, and to develop a digital droplet PCR (ddPCR) assay for detection of hypermethylated circulating tumor DNA (meth-ctDNA) in plasma from patients with EC. METHODS: DNA positions hypermethylated in EC, and without unspecific hypermethylation in tissue/cell types releasing circulating cell-free DNA in plasma, were identified in silico from TCGA/Gene Expression Omnibus (GEO) data. A methylation-specific ddPCR (meth-ddPCR) assay following bisulfite conversion of DNA extracted from plasma was optimized for detection of meth-ctDNA according to dMIQE guidelines. Performances were validated on a retrospective cohort (n = 78 tumors, n = 30 tumor-adjacent tissues), a prospective pilot cohort (n = 33 stage I-IV patients), and 55 patients/donors without cancer. RESULTS: Hypermethylation of zinc finger and SCAN domain containing 12 (ZSCAN12) and/or oxytocin (OXT) classified EC samples from multiple noncancer samples with high diagnostic specificity/sensitivity [>97%; area under the curve (AUC) = 0.99; TCGA/GEO tissues/blood samples]. These results were confirmed in the independent retrospective cohort (AUC = 0.99). Meth-ddPCR showed a high analytical specificity (limit of blank = 2) and sensitivity (absolute lower threshold of detection = 50 pgmethDNA/mLplasma). In the pilot cohort, meth-ctDNA was detected in pretreatment plasma samples from 9/11 and 5/20 patients with advanced and non-advanced EC, respectively. 2 of 9 patients had ctDNA detected after macroscopic complete surgery and experienced progression within 6 months. No healthy donors had any copy of hypermethylated DNA detected in plasma. CONCLUSIONS: Meth-ddPCR of ZSCAN12/OXT allows a highly specific and sensitive detection of ctDNA in plasma from patients with EC and appears promising for personalized approaches for these patients.

Cracking the homologous recombination deficiency code: how to identify responders to PARP inhibitors.

DNA double-strand breaks are the most critical DNA damage to cells, and their repair is tightly regulated to maintain cellular integrity. Some cancers exhibit homologous recombination deficiency (HRD), a faithful double-strand break repair system, making them more sensitive to poly (ADP ribose) polymerase inhibitors (PARPi). PARPi have shown substantial efficacy in BRCA-mutated ovarian cancer for several years, and their indication has gradually been extended to other tumour locations such as breast, prostate and pancreas. More recently, PARPi were demonstrated to be effective in cancers with an HRD phenotype beyond BRCA mutations. Today, a major challenge is developing tests capable of detecting the HRD phenotype of cancers (HRD tests) and predicting sensitivity to PARPi to select patients likely to benefit from this therapy. This review provides a synthesis of the existing HRD tests, divided into three main approaches to detect HRD: the investigation of the HRD causes, the study of its consequences and the evaluation of the HR activity itself.

Redifferentiating Effect of Larotrectinib in NTRK-Rearranged Advanced Radioactive-Iodine Refractory Thyroid Cancer.

Metastatic thyroid cancers may dedifferentiate and become radioactive-iodine (RAI) resistant. A redifferentiating effect can be observed with inhibitors of the mitogen-activated protein kinase pathway in thyroid cancers with point mutation in oncogenes. This effect allows RAI reuptake that may lead to a therapeutic effect different from the antitumoral effect of the inhibitor. The potential redifferentiating effect of inhibitors targeting oncogenic fusion-genes was suggested by one adult and one pediatric patient using larotrectinib in NTRK-rearranged tumors. We report on three consecutive adult patients with metastatic RAI-resistant NTRK-rearranged thyroid cancer who received larotrectinib for disease progression and for whom the redifferentiating effect was examined. Larotrectinib-induced RAI reuptake in all or part of the metastatic disease for two patients and no reuptake was noted for the other patient. We demonstrate that redifferentiation of NTRK-rearranged RAI-resistant thyroid cancer with larotrectinib may exist but does not occur in all patients.

Development and validation of a RNAseq signature for prognostic stratification in endometrial cancer.

BACKGROUND: Despite recent advances in endometrial carcinoma (EC) molecular characterization, its prognostication remains challenging. We aimed to assess whether RNAseq could stratify EC patient prognosis beyond current classification systems. METHODS: A prognostic signature was identified using a LASSO-penalized Cox model trained on TCGA (N = 543 patients). A clinically applicable polyA-RNAseq-based work-flow was developed for validation of the signature in a cohort of stage I-IV patients treated in two Hospitals [2010-2017]. Model performances were evaluated using time-dependent ROC curves (prediction of disease-specific-survival (DSS)). The additional value of the RNAseq signature was evaluated by multivariable Cox model, adjusted on high-risk prognostic group (2021 ESGO-ESTRO-ESP guidelines: non-endometrioid histology or stage III-IVA orTP53-mutated molecular subgroup). RESULTS: Among 209 patients included in the external validation cohort, 61 (30%), 10 (5%), 52 (25%), and 82 (40%), had mismatch repair-deficient, POLE-mutated, TP53-mutated tumors, and tumors with no specific molecular profile, respectively. The 38-genes signature accurately predicted DSS (AUC = 0.80). Most disease-related deaths occurred in high-risk patients (5-years DSS = 78% (95% CI = [68%-89%]) versus 99% [97%-100%] in patients without high-risk). A composite classifier accounting for the TP53-mutated subgroup and the RNAseq signature identified three classes independently associated with DSS: RNAseq-good prognosis (reference, 5-years DSS = 99%), non-TP53 tumors but with RNAseq-poor prognosis (adjusted-hazard ratio (aHR) = 5.75, 95% CI[1.14-29.0]), and TP53-mutated subgroup (aHR = 5.64 [1.12-28.3]). The model accounting for the high-risk group and the composite classifier predicted DSS with AUC = 0.84, versus AUC = 0.76 without (p = 0.01). CONCLUSION: RNA-seq profiling can provide an additional prognostic information to established classification systems, and warrants validation for potential RNAseq-based therapeutic strategies in EC.

The alternative RelB NF-κB subunit is a novel critical player in diffuse large B-cell lymphoma.

Diffuse large B-cell lymphoma (DLBCL) is the most frequent lymphoid malignancy affecting adults. The NF-κB transcription factor family is activated by 2 main pathways, the canonical and the alternative NF-κB activation pathway, with different functions. The alternative NF-κB pathway leads to activation of the transcriptionally active RelB NF-κB subunit. Alternative NF-κB activation status and its role in DLBCL pathogenesis remain undefined. Here, we reveal a frequent activation of RelB in a large cohort of DLBCL patients and cell lines, independently of their activated B-cell-like or germinal center B-cell-like subtype. RelB activity defines a new subset of patients with DLBCL and a peculiar gene expression profile and mutational pattern. Importantly, RelB activation does not correlate with the MCD genetic subtype, enriched for activated B-cell-like tumors carrying MYD88L265P and CD79B mutations that cooperatively activate canonical NF-κB, thus indicating that current genetic tools to evaluate NF-κB activity in DLBCL do not provide information on the alternative NF-κB activation. Furthermore, the newly defined RelB-positive subgroup of patients with DLBCL exhibits a dismal outcome after immunochemotherapy. Functional studies revealed that RelB confers DLBCL cell resistance to DNA damage-induced apoptosis in response to doxorubicin, a genotoxic agent used in the front-line treatment of DLBCL. We also show that RelB positivity is associated with high expression of cellular inhibitor of apoptosis protein 2 (cIAP2). Altogether, RelB activation can be used to refine the prognostic stratification of DLBCL and may contribute to subvert the therapeutic DNA damage response in a segment of patients with DLBCL.

Standardisation of pathogenicity classification for somatic alterations in solid tumours and haematologic malignancies.

BACKGROUND: The difficulty in interpreting somatic alterations is correlated with the increase in sequencing panel size. To correctly guide the clinical management of patients with cancer, there needs to be accurate classification of pathogenicity followed by actionability assessment. Here, we describe a specific detailed workflow for the classification of the pathogenicity of somatic variants in cancer into five categories: benign, likely benign, unknown significance, likely pathogenic and pathogenic. METHODS: Classification is obtained by combining a set of eight relevant criteria in favour of either a pathogenic or a benign effect (pathogenic stand-alone, pathogenic very strong, pathogenic strong, pathogenic moderate, pathogenic supporting, benign supporting, benign strong and benign stand-alone). RESULTS: Our guide is concordant with the ACMG/AMP 2015 guidelines for germline variants. Interpretation of somatic variants requires considering specific criteria, such as the disease and therapeutic context, co-occurring genomic events in the tumour when available and the use of cancer-specific variant databases. In addition, the gene role in tumorigenesis (oncogene or tumour suppressor gene) also needs to be taken into consideration. CONCLUSION: Our classification could contribute to homogenize best practices on somatic variant pathogenicity interpretation and improve interpretation consistency both within and between laboratories.

EGFR Exon 20 Insertion in Metastatic Non-Small-Cell Lung Cancer: Survival and Clinical Efficacy of EGFR Tyrosine-Kinase Inhibitor and Chemotherapy.

EGFR exon 20 insertions are rare genetic alterations in non-small-cell lung cancers (NSCLCs) that are usually unresponsive to approved EGFR tyrosine kinase inhibitors (TKIs). In this paper, we describe the clinical characteristics, efficacy of EFGR TKIs and chemotherapy, and resulting survival in this population. We retrospectively collected patients with EGFR exon 20 insertions (Exon20ins) from 11 French genetic platforms and paired them (1:2 ratio) with classic Exon 19/21 EGFR mutation patients (controls). Between 2012 and 2017, 35 Exon20ins patients were included. These patients were younger at diagnosis than the controls. All Exon20ins patients who were treated with first-line EGFR TKIs (n = 6) showed progressive disease as the best tumor response. There was no significant difference in the tumor response or the disease control rate with first-line platinum-based chemotherapy between the two groups. A trend towards shorter overall survival was observed in Exon20ins vs. controls (17 months (14-not reach(NR) 95% confidence interval(CI) vs. 29 months (17-NR 95%CI), p = 0.09), respectively. A significant heterogeneity in amino acid insertion in EGFR exon 20 was observed. EGFR exon 20 insertions are heterogeneous molecular alterations in NSCLC that are resistant to classic EGFR TKIs, which contraindicates their use as a first-line treatment.

c-Rel Is the Pivotal NF-κB Subunit in Germinal Center Diffuse Large B-Cell Lymphoma: A LYSA Study.

Relationships between c-Rel and GCB-DLBCLs remain unclear. We found that strong c-Rel DNA-binding activity was mostly found in GCBs on two independent series of 48 DLBCLs and 66 DLBCLs, the latter issued from the GHEDI series. c-Rel DNA-binding activity was associated with increased REL mRNA expression. Extending the study to the whole GHEDI and Lenz DLBCL published series of 202 and 233 cases, it was found that the c-Rel gene expression profile (GEP) overlapped partially (12%) but only with the GCB GEP and not with the GEP of ABC-DLBCLs. Cases with both overexpression of REL mRNA and c-Rel GEP were defined as those having a c-Rel signature. These cases were GCBs in 88 and 83% of the GHEDI or Lenz's DLBCL series respectively. The c-Rel signature was also associated with various recurrent GCB-DLBCL genetic events, including REL gains, BCL2 translocation, MEF2B, EZH2, CREBBP, and TNFRSF14 mutations and with the EZB GCB genetic subtype. By CGH array, the c-Rel signature was specifically correlated with 2p15-16.1 amplification that includes XPO1, BCL11A, and USP34 and with the 22q11.22 deletion that covers IGLL5 and PRAME. The total number of gene copy number aberrations, so-called genomic imbalance complexity, was decreased in cases with the c-Rel signature. These cases exhibited a better overall survival. Functionally, overexpression of c-Rel induced its constitutive nuclear localization and protected cells against apoptosis while its repression tended to increase cell death. These results show that, clinically and biologically, c-Rel is the pivotal NF-κB subunit in the GCB-DLBCL subgroup. Functionally, c-Rel overexpression could directly promote DLBCL tumorigenesis without need for further activation signals.

No evidence for a pathogen associated with pulmonary MALT lymphoma: a metagenomics investigation.

Mucosa-associated lymphoid tissue (MALT) lymphoma is generally associated with chronic antigen stimulation: auto-antigens or of microbial origin. Only one study suggested association between Achromobacter xylosoxidans and pulmonary MALT lymphoma. We aimed to investigate the presence of virus or any infectious agents in pulmonary MALT lymphoma by using metagenomic next-generation sequencing (mNGS).All lung samples were centrally reviewed. The t(11;18) (q21;q21) was evaluated by FISH analysis. The snap frozen large lung biopsies were analyzed by mNGS. After lung biopsies homogenization total nucleic acids (RNA and DNA) were extracted, amplified and classified according to their taxonomic assignment, after exclusion of host DNA.We included 13 samples from pulmonary MALT lymphoma (mean age: 60.3 years, 7 women, 3 with auto-immune background) and 10 controls. The diagnosis of MALT lymphoma was confirmed for the 13 samples, 3 showed API2-MALT1 translocation (23%). No evidence of the presence of a specific pathogen was clearly identified in the group of patients with pulmonary MALT lymphoma. We identifiedA. xylosoxidans sequence in 4/13 patients and in 4/10 controls.This study did not find evidence for a DNA or RNA virus, a fungi, a parasite or a bacteria associated with pulmonary MALT lymphoma either in the stroma or in tumor cells.

[Fusion transcripts: Therapeutic targets in thoracic oncology]. / Les réarrangements moléculaires : cibles thérapeutiques en cancérologie thoracique.

Five to ten percent of lung adenocarcinoma harbor chromosomal rearrangements affecting the ALK, ROS1, NTRK and RET genes. These rearrangements are associated with the production of fusion transcripts that lead to the synthesis of chimeric proteins with constitutive kinase activity. These abnormal proteins induce an oncogenic dependency that may be targeted by tyrosine kinase inhibitors. In this review, we will summarize the clinical and molecular epidemiology of chromosomal rearrangements affecting ALK, ROS1, NTRK and RET genes. We will describe the mechanisms of resistance to tyrosine kinase inhibitors that have been reported. We will present the molecular techniques that can be used to detect these rearrangements and the strategies set-up by the molecular oncology laboratories to diagnose these genetic alterations.