Comprehensive genomic and transcriptomic characterization of high-grade gastro-entero-pancreatic neoplasms.

BACKGROUND: High-grade gastro-entero-pancreatic neoplasms (HG GEP-NENs) can be stratified according to their morphology and Ki-67 values into three prognostic classes: neuroendocrine tumors grade 3 (NETs G3), neuroendocrine carcinomas with Ki-67 < 55% (NECs <55) and NECs with Ki-67 ≥ 55% (NECs ≥55). METHODS: We analyzed a cohort of 49 HG GEP-NENs by targeted Next-Generation Sequencing (TrueSight Oncology 500), RNA-seq, and immunohistochemistry for p53, Rb1, SSTR-2A, and PD-L1. RESULTS: Frequent genomic alterations affected TP53 (26%), APC (20%), KRAS and MEN1 (both 11%) genes. NET G3 were enriched in MEN1 (p = 0.02) mutations, while both NECs groups were enriched in TP53 (p = 0.001), APC (p = 0.002) and KRAS (p = 0.02) mutations and tumors with TMB ≥ 10 muts/Mb (p = 0.01). No differentially expressed (DE) gene was found between NECs <55% and NECs ≥55%, while 1129 DE genes were identified between NET G3 and NECs. A slight enrichment of CD4+ and CD8+ T cells in NECs and of cancer-associated fibroblasts and macrophages (M2-like) in NET G3. Multivariate analysis identified histologic type and Rb1 loss as independent prognostic factors for overall survival. CONCLUSIONS: This study showed that GEP-NET G3 and GEP-NECs exhibit clear genomic and transcriptomic differences, differently from GEP-NECs <55% and GEP-NECs ≥55%, and provided molecular findings with prognostic and potentially predictive value.

HER2 expression and genOmic characterization of rESected brain metastases from colorectal cancer: the HEROES study.

BACKGROUND: Little is known about prognostic factors of brain metastases (BM) from colorectal cancer (CRC). HER2 amplification/overexpression (HER2+) was previously described; its impact on prognosis remains uncertain. METHODS: In the translational study HEROES, extensive molecular analysis was performed on primary CRC (prCRC) and their matched resected BM by means of NGS comprehensive genomic profiling and HER2 status as assessed by immunohistochemical/ in situ hybridization. Count of tumour-infiltrating lymphocytes (TILs) was also performed. PRIMARY OBJECTIVE: to describe the molecular landscape of paired BM/prCRC. SECONDARY OBJECTIVES: to search for new prognostic biomarkers of outcome after BM resection: intracranial-only Progression-Free Survival (BM-iPFS), Progression-Free Survival (BM-PFS), and Overall Survival (BM-OS). RESULTS: Out of 22 patients having paired samples of prCRC and BM, HER2+ was found on 4 (18%) BM, 3 (75%) of which also HER2+ in matched prCRC. Lower tumour mutation burden (HR 3.08; 95%CI 1.06-8.93; p = 0.0386) and HER2-negative BM (HER2neg) (HR 7.75;95%CI 1.97-30.40; p = 0.0033) were associated with longer BM-iPFS; HER2neg BM (HR 3.44; 95%CI 1.03-11.53; p = 0.0449) and KRASmut BM (HR 0.31; 95%CI 0.12-0.80; p = 0.0153) conferred longer BM-PFS. Longer BM-OS was found in pts with TILs-enriched (≥1.6/HPF) BM (HR 0.11; 95%CI0.01-0.91; p = 0.0403). CONCLUSIONS: This study shows HER2+ enrichment in both BM and their prCRC. TILs-enriched BM conferred better BM-OS.

Immune-Related Diarrhea and Colitis in Non-small Cell Lung Cancers: Impact of Multidisciplinary Management in a Real-World Setting.

INTRODUCTION: Immune-related adverse events (irAEs) constitute a challenge in the clinical management of solid tumors. This study aims to collect real-world data on the occurrence of immune-mediated diarrhea and colitis (IMDC) in advanced non-small cell lung cancer (aNSCLC) treated with immune checkpoint inhibitors (ICIs) and to assess the clinical impact of a multidisciplinary approach (MDA) on IMDC management. METHODS: We retrospectively collected data on patients with aNSCLC consecutively treated with ICIs, either as single agent or in combination with chemotherapy, between September 2013 and July 2022. Among patients developing IMDC, we conducted blinded revision of colonic biopsies and evaluated the clinical impact of the introduction of MDA through predefined indicators. RESULTS: Among the 607 patients included, 84 (13.8%) experienced IMDC. Pathological review highlighted a high prevalence of microscopic colitis (28%), with a collagenous pattern linked to longer symptoms duration (P = .01). IMDC occurred more frequently in females (P = .05) and PD-L1 expressors (P = .014) and was correlated with longer progression-free survival (17.0 vs 5.8, P < .001) and overall survival (28.3 vs 9.5, P < .001). The introduction of MDA was associated with increased employment of diagnostical tools such as fecal calprotectin test (P < .001), colonoscopy (P < .001), and gastroenterological evaluation (P = .017) and a significant decrease in both grade 3 conversion rate (P = .046) and recurrence after rechallenge (P = .016). Hospitalization rate dropped from 17.2% to 3.8% (P: ns). CONCLUSION: These findings highlight the clinical relevance of IMDC and support the incorporation of a MDA to optimize the clinical management of this irAE to improve patient care. Prospective validation has been planned.

Gastric Carcinoma in Autoimmune Gastritis: A Histopathologic and Molecular Study.

Patients with autoimmune gastritis (AIG) have a 13-fold risk of developing type-1 neuroendocrine tumors, whereas the risk for gastric adenocarcinoma is still uncertain. Here we describe the clinicopathologic and molecular features of a series of gastric carcinomas (GC) arising in the context of AIG. A total of 26 AIG-associated GC specimens were collected from 4 Italian Institutions. Immunohistochemistry for MUC1, MUC2, MUC5AC, MUC6, CDX2, HER2, PD-L1, CLDN18, mismatch repair (MMR) proteins, and p53 and EBV-encoded RNA (EBER) in situ hybridization were performed. Histologic and immunohistochemical features were jointly reviewed by 5 expert gastrointestinal pathologists. Next-generation sequencing analysis (TrueSight Oncology 500, Illumina) of 523 cancer-related genes was performed on 19 cases. Most tumors were diagnosed as pT1 (52%) and they were located in the corpus/fundus (58%) and associated with operative link for gastritis assessment stage II gastritis (80.8%), absence of parietal cells, complete intestinal metaplasia, and enterochromaffin-like-cell micronodular hyperplasia. Only 4 (15.4%) GCs were diagnosed during follow-up for AIG. The following histotypes were identified: 20 (77%) adenocarcinomas; 3 (11%) mixed neuroendocrine-non-neuroendocrine neoplasms, and 2 (8%) high-grade solid adenocarcinomas with focal neuroendocrine component, 1 (4%) adenocarcinoma with an amphicrine component. Overall, 7 cases (27%) showed MMR deficiency, 3 (12%) were positive (score 3+) for HER2, 6 (23%) were CLDN18 positive, and 11 (42%) had PD-L1 combined positive score ≥ 10. EBER was negative in all cases. Molecular analysis revealed 5/19 (26%) microsatellite instability (MSI) cases and 7 (37%) tumor mutational burden (TMB) high. The most frequently altered genes were TP53 (8/19, 42%), RNF43 (7/19, 37%), ERBB2 (7/19, 37% [2 amplified and 5 mutated cases]), ARID1A (6/19, 32%), and PIK3CA (4/19, 21%). In summary, AIG-associated GCs are often diagnosed at low stage in patients with longstanding misrecognized severe AIG; they often display a neuroendocrine component or differentiation, have relatively higher rates of MMR deficiency, and TMB high.

Application of a pattern-based approach to histological diagnosis in very early onset IBD (VEO-IBD) in a multicentric cohort of children with emphasis on monogenic disease with IBD-like morphology.

AIMS: Very early-onset inflammatory bowel disease (VEO-IBD) is a clinical umbrella term referring to IBD-like symptoms arising in children before the age of 6 years, encompassing both 'pure' IBD, such as ulcerative colitis (UC) and Crohn's disease (CD) and monogenic diseases (MDs), the latter often involving genes associated with primary immunodeficiencies. Moreover, histological features in gastrointestinal (GI) biopsies in MD can also have IBD-like morphology, making differential diagnosis difficult. Correct diagnosis is fundamental, as MDs show a more severe clinical course and their inadequate/untimely recognition leads to inappropriate therapy. METHODS AND RESULTS: Biopsy samples from the lower and upper GI tract of 93 clinically diagnosed VEO-IBD children were retrospectively selected in a multicentre cohort and histologically re-evaluated by 10 pathologists blinded to clinical information. Each case was classified according to morphological patterns, including UC-like; CD-like; enterocolitis-like; apoptotic; eosinophil-rich; and IBD-unclassified (IBD-U). Nine (69%) MD children showed IBD-like morphology; only the IBD-U pattern correlated with MD diagnosis (P = 0.02) (available in 64 cases: 51 non-MD, true early-onset IBD/other; 13 MD cases). MD patients showed earlier GI symptom onset (18.7 versus 26.9 months) and were sent to endoscopy earlier (22 versus 37 months), these differences were statistically significant (P < 0.05). Upper GI histology was informative in 37 biopsies. CONCLUSIONS: The diagnosis of the underlying cause of VEO-IBD requires a multidisciplinary setting, and pathology, while being one of the fundamental puzzle pieces, is often difficult to interpret. A pattern-based histological approach is therefore suggested, thus aiding the pathologist in VEO-IBD reporting and multidisciplinary discussion.

Synchronous Endometrial and Ovarian Carcinomas: Pathologic and Molecular Analysis Highlights the Monoclonal Origin of the Lesions.

The diagnosis of synchronous carcinomas, involving both the endometrium and ovaries, is not a rare finding in gynecologic pathology and represents a challenge with implications on tumor staging and therapeutic decision-making. A mono-institutional series of 11 metastatic and 6 paired synchronous endometrial and ovarian carcinomas were reviewed by 2 expert pathologists based on previously published histopathologic criteria. The series was investigated for DNA mismatch repair proteins, p53, and POLE status and was subject to DNA-based next-generation sequencing targeting 67 cancer-related genes. Out of 17 pairs, 16 featured the same histotype (10 endometrioid, 4 serous high-grade, and 2 clear cells). By using WHO 2020 criteria, 11 couples of tumors were confirmed as metastatic and 6 couples were confirmed as independent. Based on next-generation sequencing analysis, 16 of 17 cases (11 metastatic and 5 independent) of our series showed evidence of a clonal relationship between endometrial and ovarian carcinomas. In metastatic cases, the adverse outcome was associated with nonendometrioid/high-grade endometrioid histotype and with the p53-abnormal molecular subtype. Four cases originally fulfilling clinicopathological criteria of independent endometrial and ovarian carcinomas were clonally related, low-grade endometrioid histotype and POLE-mut, mismatch repair deficient, and no specific molecular profile molecular subtypes; no adverse event was recorded in this group. In summary, the molecular characterization of synchronous gynecologic carcinomas confirms their clonal origin in most cases. However, the results of our study point out that the clinical behavior of these tumors seems to be determined by the presence of high-risk WHO 2020 histologic criteria and molecular features (i.e. p53-abnormal), rather than the monoclonal origin.

Molecular Determinants of Peritoneal Dissemination in Gastric Adenocarcinoma.

BACKGROUND: Peritoneal dissemination represents a poor prognostic indicator in gastric cancer. Despite a comprehensive molecular characterization of this disease, no peritoneal dissemination-specific signature has been identified, limiting the tailoring of the surgical and oncological treatments. In this review, we outline the available literature focusing on the role of the different molecular pathways involved in the acquisition of peritoneal metastatic dissemination. SUMMARY: According to our results, several molecular determinants are associated with peritoneal carcinomatosis and are involved in several cellular and molecular carcinogenetic processes. However, a comprehensive understanding of the complex molecular landscape of gastric carcinosis is still lacking. KEY MESSAGES: More efforts should be made toward the integration of molecular and histologic data to perform a risk prediction assessment of peritoneal dissemination based on molecular profiling and histological evaluation.

Histopathological Landscape of Precursor Lesions of Gastro-Entero-Pancreatic Neuroendocrine Neoplasms.

BACKGROUND: Despite the important advances in research on neuroendocrine neoplasms of the gastro-entero-pancreatic tract, their precursor lesions are much less well known. SUMMARY: This review analyzes the preneoplastic neuroendocrine lesions of the gastro-entero-pancreatic tract, by adopting a coherent anatomical benchmark. In particular, the settings in which neuroendocrine precursor lesions represent well-recognized pathophysiological and morphological entities (with eventual molecular correlates) have been distinguished from the ones in which the nature of preneoplastic changes is still obscure. KEY MESSAGES: The aim of the paper was to summarize what is known about precursor lesions of gastro-entero-pancreatic neuroendocrine tumors, with the goal of providing a useful tool for future research aimed at obtaining a fuller understanding of the underlying biology and early development of these diseases.

Concordance of microsatellite instability and mismatch repair status in paired biopsies and surgical specimens of resectable gastroesophageal adenocarcinoma: time for a call to action.

BACKGROUND: Reliability of mismatch repair proteins and microsatellite instability assessment is essential in order to define treatment strategy and identify candidates to immune checkpoint inhibitors in locally advanced gastroesophageal carcinoma. We evaluated the concordance of deficient mismatch repair (dMMR) and microsatellite instability-high (MSI-H) status between endoscopic biopsies and surgical specimens. METHODS: Consecutive patients with resectable gastric or gastroesophageal junction adenocarcinoma classified as MSI-H/dMMR by polymerase chain reaction (PCR) or immunohistochemistry (IHC) and operated at three referral Institutions were included. The primary endpoint was the rate of concordance between biopsy and surgical samples. If needed, central revision by IHC/PCR was performed by specialized pathologists from coordinating Institutions. RESULTS: Thirteen (19.7%) out of 66 patients showed discordant MSI-H/dMMR results in the original pathology reports. In most cases (11, 16.7%) this was due to the diagnosis of proficient mismatch repair status on biopsies. Among the ten cases available for central review, four were due to sample issues, four were reclassified as dMMR, one case showed dMMR status but was classified as microsatellite stable by PCR, one was linked to misdiagnosis of endoscopic biopsy by the local pathologist. Heterogeneity of mismatch repair proteins staining was observed in two cases. CONCLUSIONS: Available methods can lead to conflicting results in MSI-H/dMMR evaluation between endoscopic biopsies and surgical samples of gastroesophageal adenocarcinoma. Strategies aiming to improve the reliability of assessment should be primarily focused on the optimization of tissue collection and management during endoscopy and adequate training of dedicated gastrointestinal pathologists within the multidisciplinary team.

The Day-To-Day Practice of MMR and MSI Assessment in Colorectal Adenocarcinoma: What We Know and What We Still Need to Explore.

BACKGROUND: The DNA mismatch repair (MMR) system is a highly preserved protein complex recognizing short insertions, short deletions, and single base mismatches during DNA replication and recombination. MMR protein status is identified using immunohistochemistry. Deficit in one or more MMR proteins, configuring deficient MMR status (dMMR), leads to frameshift mutations particularly clustered in microsatellite repeats. Thus, microsatellite instability (MSI) is the epiphenomenon of dMMR. In colorectal cancer (CRC), MMR/MSI status is a biomarker with prognostic and predictive value of resistance to 5-fluorouracil and response to immune checkpoint inhibitor therapy. SUMMARY: In this Review, we describe the challenges the practicing pathologist may face in relation to the assessment of MMR/MSI status and any open issues which still need to be addressed, focusing on pre-analytic issues, pitfalls in the interpretation, and technical aspects of the different assays. KEY MESSAGES: The current methods of detecting dMMR/MSI status have been optimized for CRCs, and whether these techniques can be applied to all tumor and specimen types is still not fully understood. Following the Food and Drug Administration (FDA), tissue/site agnostic drug approval of pembrolizumab for advanced/metastatic MSI tumors, MMR/MSI status in gastrointestinal tract is a common request from the oncologist. In this setting, several issues still need to be addressed, including criteria for sample adequacy.

Preoperative endoscopy and pathology report of the specimen to be recommended in sleeve gastrectomy?

Objective: Preoperative upper gastrointestinal endoscopy (UGIE) and postoperative histopathological examination (HPE) of resected specimens are still controversial issues in bariatric surgery. Methods: A retrospective review of prospectively collected laparoscopic sleeve gastrectomies (SG) performed at our institution for morbid obesity was carried out. All patients underwent pre-operative UGIE with biopsy, post-operative HPE and conventional post-operative follow-up. Results: From January 2019 through January 2021 we performed a total of 501 laparoscopic SG. A total of 12 (2.4%) neoplasms were found, 2 evident at preoperative UGIE, 4 detected during operation, and 6 at HPE. Eight of these 12 cases had some malignant potential and 5 would not have been detected without HPE of the specimen. The most significant unexpected case was a fundic gland type adenocarcinoma in a 64-year-old female with severe obesity. Conclusion: On the basis of our clinical experience, we recommend both preoperative endoscopic assessment and postoperative HPE of the specimen to provide the best available treatment to these patients.

FGFR2 testing in cholangiocarcinoma: translating molecular studies into clinical practice.

Cholangiocarcinoma (CCA) is a heterogeneous group of neoplasms burdened by a dismal prognosis. Several studies have investigated the genomic profile of CCA and identified numerous druggable genetic alterations, including FGFR2 fusions/rearrangements. Approximately 5-7% of CCAs and 10-20% of intrahepatic iCCAs harbor FGFR2 fusions. With the recent advent of FGFR-targeting therapies into clinical practice, a standardization of molecular testing for FGFR2 alterations in CCA will be necessary. In this review, we describe the technical aspects and challenges related to FGFR2 testing in routine practice, focusing on the comparison between Next-Generation Sequencing (NGS) and FISH assays, the best timing to perform the test, and on the role of liquid biopsy.

A practical approach for PD-L1 evaluation in gastroesophageal cancer.

PD-L1 is an established predictive immunohistochemical biomarker of response to immune checkpoint inhibitors. At present, PD-L1 is routinely assessed on biopsy samples of advanced gastroesophageal cancer patients before initiating first-line treatment. However, PD-L1 is still a suboptimal biomarker, due to changing cut-off values and scoring systems, interobserver and interlaboratory variability.This practical illustrated review discusses the range of staining patterns of PD-L1 and the potential pitfalls and challenges that can be encountered when evaluating PD-L1, focusing on gastric and gastroesophageal adenocarcinoma (G/GEA) and esophageal squamous cell carcinoma (ESCC).

The ideal reporting of RAS testing in colorectal adenocarcinoma: a pathologists' perspective.

RAS gene mutational status represents an imperative predictive biomarker to be tested in the clinical management of metastatic colorectal adenocarcinoma. Even if it is one of the most studied biomarkers in the era of precision medicine, several pre-analytical and analytical factors may still impasse an adequate reporting of RAS status in clinical practice, with significant therapeutic consequences. Thus, pathologists should be aware on the main topics related to this molecular evaluation: (i) adopt diagnostic limit of detections adequate to avoid the interference of sub-clonal cancer cell populations; (ii) choose the most adequate diagnostic strategy according to the available sample and its qualification for molecular testing; (iii) provide all the information regarding the mutation detected, since many RAS mutation-specific targeted therapeutic approaches are in development and will enter into routine clinical practice. In this review, we give a comprehensive description of the current scenario about RAS gene mutational testing in the clinic focusing on the pathologist's role in patient selection for targeted therapies.

Comprehensive Genomic Profiling (CGP)-Informed Personalized Molecular Residual Disease (MRD) Detection: An Exploratory Analysis from the PREDATOR Study of Metastatic Colorectal Cancer (mCRC) Patients Undergoing Surgical Resection.

A majority of patients with metastatic colorectal cancer (mCRC) experience recurrence post curative-intent surgery. The addition of adjuvant chemotherapy has shown to provide limited survival benefits when applied to all patients. Therefore, a biomarker to assess molecular residual disease (MRD) accurately and guide treatment selection is highly desirable for high-risk patients. This feasibility study evaluated the prognostic value of a tissue comprehensive genomic profiling (CGP)-informed, personalized circulating tumor DNA (ctDNA) assay (FoundationOne®Tracker) (Foundation Medicine, Inc., Cambridge, MA, USA) by correlating MRD status with clinical outcomes. ctDNA analysis was performed retrospectively on plasma samples from 69 patients with resected mCRC obtained at the MRD and the follow-up time point. Tissue CGP identified potentially actionable alterations in 54% (37/69) of patients. MRD-positivity was significantly associated with lower disease-free survival (DFS) (HR: 4.97, 95% CI: 2.67−9.24, p < 0.0001) and overall survival (OS) (HR: 27.05, 95% CI: 3.60−203.46, p < 0.0001). Similarly, ctDNA positive status at the follow-up time point correlated with a marked reduction in DFS (HR: 8.78, 95% CI: 3.59−21.49, p < 0.0001) and OS (HR: 20.06, 95% CI: 2.51−160.25, p < 0.0001). The overall sensitivity and specificity at the follow-up time point were 69% and 100%, respectively. Our results indicate that MRD detection using the tissue CGP-informed ctDNA assay is prognostic of survival outcomes in patients with resected mCRC. The concurrent MRD detection and identification of actionable alterations has the potential to guide perioperative clinical decision-making.

PD-L1 evaluation in the gastrointestinal tract: from biological rationale to its clinical application.

Immune-checkpoint inhibitors targeting the PD-1/PD-L1 axis have brought significant clinical benefit in many solid cancer types, including gastrointestinal malignancies. However, it has been estimated that only 20-40% of patients respond to treatment. The pattern of expression and potential predictive value of PD-L1 as an immunohistochemical biomarker has been extensively studied in gastrointestinal neoplasms. Until now, its predictive value has been demonstrated, and is currently in use only in upper gastrointestinal malignancies (gastroesophageal adenocarcinoma and esophageal squamous cell carcinoma).In this Review, we describe the technical aspects and challenges related to PD-L1 immunohistochemical assays, the current role of PD-L1 as a biomarker in clinical practice and we outline the main studies and clinical trials analyzing the prognostic and predictive value of PD-L1 in gastrointestinal cancers.

miRNAs Involved in Esophageal Carcinogenesis and miRNA-Related Therapeutic Perspectives in Esophageal Carcinoma.

MicroRNAs (miRNAs) are small non-coding RNAs that play a pivotal role in many aspects of cell biology, including cancer development. Within esophageal cancer, miRNAs have been proved to be involved in all phases of carcinogenesis, from initiation to metastatic spread. Several miRNAs have been found to be dysregulated in esophageal premalignant lesions, namely Barrett's esophagus, Barrett's dysplasia, and squamous dysplasia. Furthermore, numerous studies have investigated the alteration in the expression levels of many oncomiRNAs and tumor suppressor miRNAs in esophageal squamous cell carcinoma and esophageal adenocarcinoma, thus proving how miRNAs are able modulate crucial regulatory pathways of cancer development. Considering these findings, miRNAs may have a role not only as a diagnostic and prognostic tool, but also as predictive biomarker of response to anti-cancer therapies and as potential therapeutic targets. This review aims to summarize several studies on the matter, focusing on the possible diagnostic-therapeutic implications.

The Immunopathological and Histological Landscape of COVID-19-Mediated Lung Injury.

A complete understanding of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) physiopathology and related histopathologic lesions is necessary to improve treatment and outcome of coronavirus disease 2019 (COVID-19) patients. Many studies have focused on autopsy findings in COVID-19-related deaths to try and define any possible specific pattern. Histopathologic alterations are principally found within lungs and blood vessels, and these abnormalities also seem to have the highest clinical impact. Nevertheless, many of the morphological data collected so far are non-specific, fickle, and possibly associated with other co-existing factors. The aim of this minireview is to describe the main histopathological features related to COVID-19 and the mechanism known as "cytokine storm".

Molecular Landscapes of Gastric Pre-Neoplastic and Pre-Invasive Lesions.

Gastric carcinoma (GC) represents one of the most common and most lethal malignancies worldwide. The histopathological characterization of GC precursor lesions has provided great knowledge about gastric carcinogenesis, with the consequent introduction of effective strategies of primary and secondary prevention. In recent years, a large amount of data about the molecular events in GC development is emerging, flanking the histomorphological descriptions. In this review, we describe the landscape of molecular alterations in gastric pre-invasive lesions with a glance at their potential use in the diagnostic and therapeutic decision-making process.