SMAD4 Mutations Research Articles

Beyond Khorana score 2: Molecular correlates of venous thromboembolism in pancreatic cancer.

751 Background: Pancreatic ductal adenocarcinoma (PDAC) is associated with a hypercoagulable state leading to thrombosis. Risk models such as the Khorana score automatically classify PDAC as intermediate-high risk, and recent guidelines recommend consideration of thromboprophylaxis. However, little is known about the molecular correlates of PDAC for venous thromboembolism (VTE). Methods: We examined clinical and genomic data from the prospective multi-institution COMPASS trial (NCT02750657), which enrolled patients with treatment-naïve metastatic PDAC who underwent a fresh tumor biopsy for real-time whole genome and transcriptome sequencing. Laser capture microdissection was performed. Patients underwent restaging scans at 8-week intervals. Detailed chart review was conducted to focus on VTE risk factors, timing of VTE diagnosis, and anticoagulation. We also compared clinical and molecular factors based on timing of VTE diagnosis. Overall survival was defined as time from VTE to death. Results: Of 268 patients enrolled in the COMPASS trial, 166 patients had detailed clinical data available regarding VTE status. 82 patients (49%) developed a VTE, where 16 (20%) had breakthrough clots. Baseline epidemiological variables were similar for those with and without VTE, including no differences between age, sex, BMI, and baseline CA 19-9. No patients were on routine prophylactic anticoagulation. SMAD4 mutations were more frequently seen in the VTE subgroup (57% vs. 40%, p= 0.04), but no differences were seen in other driver genes (KRAS, TP53, CDKN2A), Moffitt subtype, or homologous recombination deficiency status. Patients who developed an early VTE (within 3 months) had a higher baseline CA 19-9 (median 4015 vs. 891, p= 0.02), and none were port-associated. A higher incidence of KRAS wildtype cases (10%) were observed in the early vs. later VTE groups (p= 0.03), but no differences in KRAS allelic status. There were no differences in burden of SNVs, indels, or SVs. Early VTE occurred in 46% of all basal cases, which are typically more aggressive, and only in 37% of classical subtypes. Overall survival was shorter with early VTE (HR 1.74, p= 0.02). Conclusions: A higher incidence of SMAD4 alterations were observed among patients with metastatic PDAC who developed VTE. VTE diagnosed earlier were associated with shorter survival, suggesting that early thromboprophylaxis should be considered at the time of diagnosis. Clinical trial information: NCT02750657 .

Clinicopathologic and genomic characteristics of biliary tract carcinomas with TERT promoter mutations among East Asian population.

Telomerase reverse transcriptase gene promoter (TERT) mutations are biomarkers that predict survival and responses to immune checkpoint inhibitors in various malignancies. However, their prevalence and clinicopathologic characteristics in biliary tract carcinomas are largely unknown. We performed a comprehensive genomic profiling of formalin-fixed paraffin-embedded tumor tissue from 485 carcinomas, including intrahepatic (n = 220), perihilar (n = 54), distal biliary tract (n = 110), and gallbladder (n = 101) cancers, using next-generation sequencing. TERT mutations were observed in 50 out of 485 biliary tract cancers (10.3 %) consisting of 39 C228T (78.0 %) and 11 C250T (22.0 %) variants. Among the different anatomic locations, TERT mutations were most frequent in the gallbladder (20.8 %), followed by perihilar (9.3 %), intrahepatic (7.7 %), and distal bile ducts (6.4 %) (p < 0.01). Genetically, TERT mutations were significantly associated with TP53 mutations (p = 0.04), ERBB2 amplification (p < 0.01), and high tumor mutational burdens (TMB) (p < 0.01); moreover, they were negatively correlated with KRAS (p < 0.01), SMAD4 (p = 0.01), and PBRM1 mutations (p = 0.01). In addition, TERT mutations were associated with a poor progression-free survival (PFS, p = 0.01). Specifically, in cases of intrahepatic cholangiocarcinoma, TERT mutations were more frequent in patients with cirrhosis (p = 0.01), hepatitis B virus infection (p = 0.04), and advanced disease stages (p < 0.01). In gallbladder carcinoma, TERT mutations were also associated with poor PFS. In conclusion, TERT mutations in biliary tract carcinomas had unique clinicopathologic and genetic characteristics. Despite its poor PFS, the concomitant presence of ERBB2 amplification and a high TMB indicated a potential for targeted therapy and immunotherapy in this specific subtype.

Genomic and transcriptomic signatures of sequential carcinogenesis from papillary neoplasm to biliary tract cancer.

Papillary neoplasms of the biliary tree, including intraductal papillary neoplasms (IPN) and intracholecystic papillary neoplasms (ICPN), are recognized as precancerous lesions. However, the genetic characteristics underlying sequential carcinogenesis remain unclear. Whole-exome sequencing was performed on 166 neoplasms (33 intrahepatic IPNs, 44 extrahepatic IPNs, and 89 ICPNs), and 41 associated carcinomas. Nine available cases were also subjected to spatial transcriptomic analysis. Mutations in the MAPK (48%), genomic integrity maintenance (42%), and Wnt/β-catenin (33%) pathways were prevalent in intrahepatic IPNs, extrahepatic IPNs, and ICPNs, respectively. KRAS mutations were enriched in intrahepatic IPN (42%, P<0.001), whereas SMAD4 mutations were enriched in extrahepatic IPN (21%, P=0.005). ICPNs frequently exhibit CTNNB1 mutations, particularly in low-grade lesions. Mutational signature analysis revealed that SBS1 and SBS5 signatures were homogeneously enriched in intrahepatic IPN, in contrast to the heterogeneous distribution of SBS1, SBS2, SBS5, SBS13, SBS7b, and SBS23 in extrahepatic IPN and ICPN. Copy number aberrations gradually increased from low- to high-grade intraepithelial neoplasia and eventually to carcinoma. Phylogenetic analysis revealed that 89% of carcinomas were derived from IPN/ICPN through sequential carcinogenesis, with the majority sharing driver mutations between IPN/ICPN and carcinoma. Furthermore, multifocal, independent carcinogenesis events were observed in IPNs/ICPNs, resulting in mutationally distinct carcinoma lesions. Carcinogenesis of IPN/ICPN occurs in multiple subclones through mutational accumulation and transcriptomic alterations that affect vascular development, cell morphogenesis, extracellular matrix organization, and growth factor response. With the largest IPN/ICPN cohort reported to date, our study provides a genome- and spatial transcriptome-level portrait of sequential carcinogenesis and differences in the anatomical location of biliary papillary neoplasms. Biliary tract cancer is a fatal malignancy. However, its genome-level sequential carcinogenesis from intraepithelial neoplasia to carcinoma has not yet been evaluated in a sufficiently large cohort. Papillary lesions of the bile duct and gallbladder are collectively termed intraductal papillary neoplasms (IPN) of the bile duct and intracholecystic papillary neoplasms (ICPN), respectively. They are primarily diagnosed based on histopathological studies. This study provides a comprehensive mutational and spatial transcriptomic landscape of papillary neoplasms of the bile duct and gallbladder. The results of this study offer insights into the mechanism of sequential carcinogenesis in papillary biliary tract tumors, pathology-genomics correlation, and potential therapeutic targets.

Genomic Profiling of Biliary Tract Cancers: Comprehensive Assessment of Anatomic and Geographic Heterogeneity, Co-Alterations and Outcomes.

Biliary tract cancers (BTCs) represent distinct biological and genomic entities. Anatomic and geographic heterogeneity in genomic profiling of BTC subtypes, genomic co-alterations, and their impact on long-term outcomes are not well defined. Genomic data to characterize alterations among patients with BTCs were derived from the AACR GENIE registry (v15.1) and other genomic data sets. Patterns of mutational co-occurrence, frequency of co-alterations, and their impact on long-term outcomes among BTC patients were examined. Alterations in IDH1 and FGFR2 genes were mostly noted among intrahepatic cholangiocarcinoma (iCCA) samples, TP53, ERBB2/HER2, and SMAD4 mutations were more frequent among gallbladder cancer (GBC) samples while extrahepatic cholangiocarcinoma (eCCA) more commonly harbored KRAS mutations (all Q < 0.001). Alterations in IDH1 and FGFR2 genes were more frequent among iCCA samples from Western vs. Eastern populations, while KRAS, SMAD4, and ERBB2 mutations were more commonly observed among Eastern populations(all Q < 0.05). FGFR2 gene was frequently co-mutated with BAP1 (log2OR: 1.55, Q < 0.001), while IDH1 gene was commonly co-mutated with PBRM1 (log2OR: 1.09, Q < 0.001). Co-alteration rate among patients with IDH1-mutant iCCAs, FGFR2-rearranged iCCAs, KRAS-mutant eCCA, and HER2-mutant GBCs were 80.8%, 85.2%, 76.7%, and 100%, respectively. Among patients with iCCA and FGFR2 fusions/rearrangements, harboring co-alterations in the TP53 pathway or PI3K pathway correlated with worse overall survival (OS), while patients with IDH1-mutant iCCA had worse OS when harboring co-alterations in the cell cycle pathway. Marked genomic heterogeneity exists among patients with BTCs based on anatomic and geographic location. The overwhelming majority of BTC patients with clinically significant mutations had concurrent genomic co-alterations. The current study highlights the molecular complexity of BTCs with multiple alterations that commonly co-exist and could potentially be targeted to treat BTCs.

Leveraging Tumor Mutation Profiles to Forecast Immune Checkpoint Blockade Resistance in Melanoma, Lung, Head and Neck, Bladder and Renal Cancers

Immune checkpoint blockade (ICB), radiotherapy, chemotherapy and surgery are currently used as therapeutic strategies against melanoma, lung, bladder and renal cancers, but their efficacy is limited. Thus, I need to predict treatment response and resistance to address this challenge. In this study, I analyzed 350 lung cancer, 320 melanoma, 215 bladder cancer, 139 head and neck cancer and 151 renal carcinoma patients treated with ICB to identify tumor mutations associated with response and resistance to treatment. I identified several tumor mutations linked with a difference in survival outcomes following ICB. In lung cancer, missense mutations in ABL1, ASXL1, EPHA3, EPHA5, ERBB4, MET, MRE11A, MSH2, NOTCH1, PAK7, PAX5, PGR, ZFHX3, PIK3C3 and REL genes were indicative of favorable responses to ICB. Conversely, mutations in TGFBR2, ARID5B, CDKN2C, HIST1H3I, RICTOR, SMAD2, SMAD4 and TP53 genes were associated with shorter overall survival post-ICB treatment. In melanoma, mutations in FBXW7, CDK12, CREBBP, CTNNB1, NOTCH1 and RB1 genes predict resistance to ICB, whereas missense mutations in FAM46C and RHOA genes are associated with extended overall survival. In bladder cancer, mutations in HRAS genes predict resistance to ICB, whereas missense mutations in ERBB2, GNAS, ATM, CDKN2A and LATS1 genes, as well as nonsense mutations in NCOR1 and TP53 genes, are associated with extended overall survival. In head and neck cancer, mutations in genes like PIK3CA and KRAS correlated with longer survival, while mutations in genes like TERT and TP53 were linked to shorter survival. In renal carcinoma, mutations such as EPHA5, MGA, PIK3R1, PMS1, TSC1 and VHL were linked to prolonged overall survival, while others, including total splice mutations and mutations in B2M, BCOR, JUN, FH, IGF1R and MYCN genes were associated with shorter overall survival following ICB. Then, I developed predictive survival models by machine learning that correctly forecasted cancer patient survival following ICB within an error between 5 and 8 months based on their distinct tumor mutational attributes. In conclusion, this study advocates for personalized immunotherapy approaches in cancer patients.

Genetic and microenvironmental evolution of colorectal liver metastases under chemotherapy.

Drug resistance limits the efficacy of chemotherapy for colorectal cancer liver metastasis (CRLM). However, the evolution of CRLM during drug treatment remains poorly elucidated. Multi-omics and treatment response data from 115 samples of 49 patients with CRLM undergoing bevacizumab (BVZ)-based chemotherapy show little difference in genomic alterations in 92% of cases, while remarkable differences are observed at the transcriptomic level. By decoupling intrinsic and acquired resistance, we find that hepatocyte and myeloid cell infiltration contribute to 38.5% and 23.1% of acquired resistance, respectively. Importantly, SMAD4 mutations and chr20q copy-number gain are associated with intrinsic chemoresistance. Gene interference experiments suggest that SMAD4R361H/C mutations confer BVZ and 5-fluorouracil (5-FU) resistance through STAT3 signaling. Notably, supplementing BVZ and 5-FU with the STAT3 inhibitor GB201 restores therapeutic efficacy in SMAD4R361H/C cancer cells. Our study uncovers the evolutionary dynamics of CRLM and its microenvironment during treatment and offers strategies to overcome drug resistance.

Genomic profiles and their associations with microsatellite instability status, tumor mutational burden, and programmed death ligand 1 expression in Chinese patients with colorectal cancer.

Colorectal cancer (CRC) is among the most prevalent malignancies globally, with a rising incidence observed in younger demographics. Despite surgical resection remaining the cornerstone of treatment, metastatic CRC poses significant therapeutic challenges. Immunotherapy, a mode of treatment that leverages the patient's immune system, presents a promising frontier in CRC management, particularly for late-stage cases with limited treatment options. The study was aimed to elucidate the relationships between genetic profiles and predictive biomarkers in CRC patients to inform immunotherapy decisions and improve outcomes. We conducted a large-scale study involving 660 patients with CRC, analyzing genetic profiles and predictive biomarkers for immune checkpoint inhibitors (ICIs) using next-generation sequencing (NGS) and immunohistochemistry (IHC). The study focused on assessing the association between gene mutations and markers such as microsatellite instability (MSI) status, tumor mutational burden (TMB), and programmed death ligand 1 (PD-L1) expression. Analysis revealed a diverse mutational landscape in CRC, with TP53 (73.64%), APC (67.58%), and KRAS (46.82%) being the most frequently mutated genes. We observed significant associations between KRAS mutations and co-occurrences with FBXW7, PIK3CA, and SMAD4 mutations, while KRAS mutations were mutually exclusive with TP53 mutations. KRAS mutations were enriched in the PD-L1 tumor proportion score (TPS) ≥1% population (P=0.03), whereas APC mutations were enriched in the PD-L1 TPS <1% population (P=0.10) as compared to their wild types. Additionally, specific mutations such as KRAS p.A146T, PIK3CA p.H1047R, and BRAF p.V600E were significantly associated with higher TMB and MSI-high status, indicating potential benefits from ICI therapy. Our findings underscore the importance of genetic profiling in guiding treatment decisions for patients with CRC, particularly in the era of immunotherapy. Understanding the complex interplay between genetic alterations and immune markers is critical for optimizing therapeutic strategies and improving clinical outcomes. Further research is warranted to validate these findings and explore personalized treatment approaches in CRC.

A Targeted Octahedral DNA Nanostructure Co-delivers siME3 and Doxorubicin to Enhance Collateral Lethality in ME2-Deficient Pancreatic Cancer.

The genetic characteristics of pancreatic cancer (PC) are being revealed, but treatment strategies based on these profiles are developing slowly. About one-third of PC patients harbor SMAD4 mutations, with its homozygous deletions often accompanied by deletions of the malic enzyme 2 (ME2) gene, leading to upregulation of malic enzyme 3 (ME3) to eliminate reactive oxygen species (ROS). We designed an aptamer-modified octahedral DNA nanostructure for targeted co-delivery of siRNA targeting ME3 (siME3) and doxorubicin (DOX). This nanostructure targets the epidermal growth factor receptor (EGFR) on the membrane of PC cells. Upon internalization, siME3 and DOX are released intracellularly. The siME3 effectively inhibited ME3 expression, diminishing the tumor cells' capacity to clear ROS. Moreover, DOX further increases the level of cellular ROS, and the sustained accumulation of ROS ultimately leads to apoptosis of ME2-deficient PC cells. This targeting nanostructure shows potential for enhancing collateral lethality in this PC subgroup.

The application of statistical techniques in assessing the impact of medical interventions: A case study of diabetes management

Abstract. Esophageal cancer (EC) ranks seventh among all cancers in terms of frequency and is the sixth leading cause of cancer-related deaths worldwide. The 5-year relative survival rate for esophageal cancer stands at a dismal 21%, which is nearly the lowest among all cancers. In China alone, over 477,900 individuals are diagnosed with EC annually. Advances in whole-genome sequencing technologies have enabled researchers to identify numerous gene mutations across the entire gene sequence in EC patients. This paper aims to consolidate current knowledge on several key genes implicated in EC, as revealed by whole-genome sequencing of patient biopsies. By examining research papers published within the last decade, this study captures the main findings and elucidates the influence of mutations in SOX2, TP53, NOTCH1, SMAD4, and CDKN2A on the formation, progression, and prognosis of both esophageal adenocarcinomas and squamous cell carcinomas. The insights gained are expected to contribute positively to the overall comprehension of esophageal cancer.

Overlap syndrome of hereditary hemorrhagic telangiectasia and juvenile polyposis syndrome: ten years follow-up-case series and review of literature.

Hereditary Hemorrhagic Telangiectasia (HHT) is an autosomal vascular dysplasia characterized by the presence of mucocutaneous telangiectasia and arteriovenous malformations in solid organs. The Curaçao criteria and/or detection of ALK1, ENG, and SMAD4 gene mutations are used for diagnosis. Juvenile Polyposis Syndrome (JPS) is diagnosed according to the number and localization of juvenile polyps, and family history of JPS. Both entities have a low prevalence. Mutation of SMAD4 leads to a combined syndrome of these two conditions called HHT-JPS Overlap Syndrome. We aim to describe the clinical characteristics associated with this condition focusing on long term follow up and review of the literature. A cross-sectional descriptive study of HHT-JPS cases from an HHT Institutional Registry was designed. Patients were eligible for this case series if they fulfilled both HHT and JPS diagnostic criteria and/or mutation on SMAD4. A comprehensive review was conducted on the clinical phenotype associated with HHT and its gastrointestinal involvement. Fourteen patients from eleven families in 788 previously HHT-diagnosed patients met the inclusion criteria. The ages ranged between 25 and 70 years old and 12 were females. In addition to the typical signs/symptoms of HHT, two distinct phenotypes were observed. Nine patients predominantly exhibited initially upper, while five showed predominantly initially lower gastrointestinal involvement. Numerous musculoskeletal and cardiovascular anomalies were also identified. The observed phenotypic diversity, particularly in gastrointestinal involvement, underscores the need for tailored clinical approaches. Comprehensive assessments identified associated musculoskeletal and cardiovascular anomalies, emphasizing the systemic nature of HHT-JPS.

Mucinous Borderline Tumors of the Pelvicalyceal System: Report of Two Tumors With Molecular Findings and Review of Literature.

Mucinous borderline tumors of the pelvicalyceal system are exceedingly rare tumors with very little information regarding their clinicopathological characteristics and molecular underpinnings in the published literature. Herein, we report two mucinous borderline tumors and review the literature pertaining to these tumors. The patients were women who presented with flank pain without any history of nephrolithiasis. Abdominal computed tomography (CT) showed multiloculated cystic lesions without mural nodules involving the kidney (Bosniak category III). The nephrectomy specimens showed multiloculated cysts filled with mucoid material, without any solid areas. On microscopic examination, the cystic spaces were lined predominantly by a single layer of mucinous epithelium with variable foci of epithelial stratification. Extensive sampling of the tumors did not show any evidence of invasion. The tumor cells were positive for keratin 7, keratin 20, and CDX2 and were negative for urothelial markers. Molecular analysis performed on one of the tumors showed loss of function mutation of SMAD4 p.S32fs. The presence of SMAD4 alterations alludes to the potential role of the SMAD4-TGFß pathway in the pathogenesis of this tumor. On clinical follow-up, both patients are well with no evidence of disease. Our case report expands on the clinical characteristics of these rare tumors and provides insight into the previously unreported molecular landscape of this tumor.

SMAD4 Limits PARP1 dependent DNA Repair to Render Pancreatic Cancer Cells Sensitive to Radiotherapy

Dysregulation of SMAD4 (i.e. somatic mutation) is strongly associated with poor pancreatic ductal adenocarcinoma (PDAC) prognosis, yet the molecular mechanisms remain underlying this relationship obscure. Previously, we discovered that SMAD4 mutation renders pancreatic cancer resistant to radiotherapy via promotion of autophagy. In the current work, we observed a downregulation of the protein level of SMAD4 in PDAC as compared with adjacent normal tissue, and that such SMAD4low PDAC failed to benefit from chemotherapy. Furthermore, we observed that SMAD4 depletion dramatically enhanced DNA repair capacity in response to irradiation (IR) or a radiomimetic chemical. Interestingly, we found the radiomimetic chemical having induced a robust translocation of SMAD4 into the nucleus, where a direct interaction was shown to occur between the MH1 domain of SMAD4 and the DBD domain of PARP1. Functionally, the SMAD4-PARP1 interaction was found to perturb the recruitment of PARP1 to DNA damage sites. Accordingly, the combination of olaparib and radiotherapy was indicated in vivo and in vitro to specifically reduce the growth of SMAD4-deficient PDAC by attenuating PARP1 activity. Collectively, our results revealed a novel molecular mechanism for the involvement of the SMAD4-PARP1 interaction in DNA repair with a vital role in radiotherapy response in PDAC. Based on our set of findings, our findings offer a new combined therapeutic strategy for SMAD4 deficient PDAC that can significantly reduce pancreatic cancer radiotherapy resistance.

Adequacy of EUS-guided fine-needle aspiration and fine-needle biopsy for next-generation sequencing in pancreatic malignancies: A systematic review and meta-analysis.

A majority of pancreatic malignancies are unresectable at the time of presentation and require EUS-guided fine-needle aspiration or fine-needle biopsy (EUS-FNA/FNB) for diagnosis. With the advent of precision therapy, there is an increasing need to use EUS-FNA/FNB sample for genetic analysis. Next-generation sequencing (NGS) is a preferred technology to detect genetic mutations with high sensitivity in small specimens. We performed a meta-analysis to evaluate the adequacy of EUS-FNA/FNB for NGS in pancreatic malignancies. PubMed, Embase, Cochrane Library, and Web of Science were searched from database inception to November 11, 2023. The primary outcome was the proportion of sufficient sample acquired by EUS-FNA/FNB in pancreatic malignancies for NGS. Secondary outcomes were the proportion of sufficient sample for NGS in pancreatic ductal adenocarcinoma (PDAC) and the detection rates of mutations in KRAS, TP53, CDKN2A, and SMAD4 and actionable mutations in PDAC. The pooled proportions were calculated using a random-effects model. Potential sources of heterogeneity were investigated with subgroup analyses and meta-regression. Twenty studies with 881 samples were included. The pooled adequacy of EUS-FNA/FNB sample for NGS was 89.9% (95% CI, 80.8%-96.7%) in pancreatic malignancies and 92.0% (95% CI, 81.3%-98.8%) in PDAC. Screening sample suitability before NGS testing was associated with lower adequacy in subgroup analysis (79.7% vs. 98.4%, P = 0.001). The pooled prevalences of mutations in KRAS, TP53, CDKN2A, and SMAD4 in PDAC were 87.4% (95% CI, 83.2%-91.2%), 62.6% (95% CI, 53.2%-71.7%), 20.6% (95% CI, 11.9%-30.8%), and 19.4% (95% CI, 11.2%-29.1%), respectively. The pooled prevalence of potentially actionable mutations in PDAC was 14.5% (95% CI, 8.2%-22.0%). In the majority of cases, EUS-FNA/FNB can acquire adequate sample for NGS and identify tumor-specific mutations in patients with pancreatic malignancies. Strict pre-analysis screening criteria may negatively impact the sample adequacy and the success rate for NGS.

Large cell neuroendocrine carcinoma in pancreatoblastoma with TP53 and SMAD4 mutations: a clinicopathologic study of a rare entity

Abstract Pancreatoblastoma, a rare pancreatic tumor, exhibits diverse differentiation pathways, including acinar, ductal, and neuroendocrine lineages, often with distinct squamoid nests [3]. We present a notable case of pancreatoblastoma coexisting with large cell neuroendocrine carcinoma in a 10-year-old boy, presenting with abdominal discomfort, weight loss, and lesions in the pancreas, spleen, and liver visible on imaging. A liver biopsy revealed a poorly differentiated carcinoma with neuroendocrine features, while a splenic biopsy showed acinar cell differentiation, raising possible diagnoses of pancreatoblastoma or acinar cell carcinoma. Subsequent surgical resection after chemotherapy revealed diverse components within the pancreatoblastoma, including well-differentiated acinar and neuroendocrine cells, squamoid nests, and a high-grade neuroendocrine carcinoma. Genetic analysis detected pathogenic variants in TP53 and SMAD4, rarely found in pancreatoblastomas. This juxtaposition of large cell neuroendocrine carcinoma and pancreatoblastoma suggests a potential evolution from well-differentiated neuroendocrine tumors to poorly-differentiated carcinomas within pancreatoblastomas.

Roth spots in a Rendu-Osler-Weber syndrome.

To describe the molecular diagnosis and atypical ocular presentation of a patient who suffered for a Rendu-Osler-Weber syndrome associated with juvenile polyposis (JP) syndrome. This is a case report of a patient that underwent fundus examen, brain Magnetic Resonance Imaging (MRI) and arteriography. Genetic testing was performed by next-generation-sequencing (NGS). A 35-year-old woman presented with right hemiplegia with right homonymous lateral hemianopia and homolateral complete sensory deficit. She also had Roth spots in her left fundus. Genetic testing revealed a pathogenic variation in the heterozygous state in the SMAD-4 gene (c.1245_1248del). Hereditary Hemorrhagic Telangiectasia (HTT) also known as Rendu-Osler-Weber syndrome is a rare autosomal dominant disease which reveals mostly with epistaxis and cutaneous telangiectasias. Our clinical case reports Roth spots in the context of HTT associated with juvenile polyposis syndrome. SMAD-4 mutation may explain the presence of a carotid-ophthalmic aneurysm which is not a lesion usually found in HTT.

Suppressive cancer nonstop extension mutations increase C-terminal hydrophobicity and disrupt evolutionarily conserved amino acid patterns

Nonstop extension mutations, a.k.a. stop-lost or stop-loss mutations, convert a stop codon into a sense codon resulting in translation into the 3’ untranslated region until the next in-frame stop codon, thereby extending the C-terminus of a protein. In cancer, only nonstop mutations in SMAD4 have been functionally characterized, while the impact of other nonstop mutations remain unknown. Here, we exploit our pan-cancer NonStopDB dataset and test all 2335 C-terminal extensions arising from somatic nonstop mutations in cancer for their impact on protein expression. In a high-throughput screen, 56.1% of the extensions effectively reduce protein abundance. Extensions of multiple tumor suppressor genes like PTEN, APC, B2M, CASP8, CDKN1B and MLH1 are effective and validated for their suppressive impact. Importantly, the effective extensions possess a higher hydrophobicity than the neutral extensions linking C-terminal hydrophobicity with protein destabilization. Analyzing the proteomes of eleven different species reveals conserved patterns of amino acid distribution in the C-terminal regions of all proteins compared to the proteomes like an enrichment of lysine and arginine and a depletion of glycine, leucine, valine and isoleucine across species and kingdoms. These evolutionary selection patterns are disrupted in the cancer-derived effective nonstop extensions.

Molecular Profiling of Low-Grade Appendiceal Mucinous Neoplasms (LAMN).

Low-grade appendiceal mucinous neoplasia (LAMN) represents a relatively rare tumor of the appendix typically diagnosed incidentally through appendectomy for acute appendicitis. In cases where perforation occurs, mucinous content may disseminate into the abdominal cavity, leading to the development of pseudomyxoma peritonei (PMP). The primary objective of this study was to elucidate the molecular characteristics associated with various stages of LAMN and PMP. DNA was extracted from LAMN, primary PMPs, recurrent PMPs, and adenocarcinomas originating from LAMN. The subsequent analysis involved the examination of mutational hotspot regions within 50 cancer-related genes, covering over 2800 COSMIC mutations, utilizing amplicon-based next-generation sequencing (NGS). Our findings revealed activating somatic mutations within the MAPK-signaling pathway across all tumors examined. Specifically, 98.1% of cases showed mutations in KRAS, while one tumor harbored a BRAF mutation. Additionally, GNAS mutations were identified in 55.8% of tumors, with no significant difference observed between LAMN and PMP. While LAMN rarely displayed additional mutations, 42% of primary PMPs and 60% of recurrent PMPs showed additional mutations. Notably, both adenocarcinomas originating from LAMN showed mutations within TP53. Furthermore, 7.7% (4/52) of cases exhibited a potentially targetable KRAS G12C mutation. In four patients, NGS analysis was performed on both primary PMP and recurrent PMP/adenocarcinoma samples. While mutations in KRAS and GNAS were detected in almost all samples, 50% of recurrent cases displayed an additional SMAD4 mutation, suggesting a notable alteration during disease progression. Our findings indicate two key points: First, mutations within the MAPK pathway, particularly in KRAS, are evident across all tumors, along with a high frequency of GNAS mutations. Second, progression toward PMP or adenocarcinoma is associated with an accumulation of additional mutations within common oncogenic pathways.

Molecular genetic analysis of colorectal carcinoma with an aggressive extraintestinal immunohistochemical phenotype

Colorectal cancer (CRC) is a leading global cause of illness and death. There is a need for identification of better prognostic markers beyond traditional clinical variables like grade and stage. Previous research revealed that abnormal expression of cytokeratin 7 (CK7) and loss of the intestinal-specific Special AT-rich sequence-binding protein 2 (SATB2) are linked to poor CRC prognosis. This study aimed to explore these markers’ prognostic significance alongside two extraintestinal mucins (MUC5AC, MUC6), claudin 18, and MUC4 in 285 CRC cases using immunohistochemistry on tissue microarrays (TMAs). CK7 expression and SATB2-loss were associated with MUC5AC, MUC6, and claudin 18 positivity. These findings suggest a distinct "non-intestinal" immunohistochemical profile in CRC, often right-sided, SATB2-low, with atypical expression of CK7 and non-colorectal mucins (MUC5AC, MUC6). Strong MUC4 expression negatively impacted cancer-specific survival (hazard ratio = 2.7, p = 0.044). Genetic analysis via next-generation sequencing (NGS) in CK7 + CRCs and those with high MUC4 expression revealed prevalent mutations in TP53, APC, BRAF, KRAS, PIK3CA, FBXW7, and SMAD4, consistent with known CRC mutation patterns. NGS also identified druggable variants in BRAF, PIK3CA, and KRAS. CK7 + tumors showed intriguingly common (31.6%) BRAF V600E mutations corelating with poor prognosis, compared to the frequency described in the literature and databases. Further research on larger cohorts with a non-colorectal immunophenotype and high MUC4 expression is needed.

Clinical and Genomic Features of Classical and Basal Transcriptional Subtypes in Pancreatic Cancer.

Transcriptional profiling of pancreatic cancers has defined two main transcriptional subtypes: classical and basal. Initial data suggest shorter survival for patients with basal tumors and differing treatment sensitivity to FOLFIRINOX and gemcitabine plus nab-paclitaxel by transcriptional subtype. We examined 8,743 patients with RNA sequencing from pancreatic cancers performed at Caris Life Sciences. Classical and basal subtypes were identified using purity independent subtyping algorithm on RNA sequencing, and two cohorts were analyzed: (i) the biomarker cohort included patients with complete molecular profiling data (n = 7,250) and (ii) the outcome cohort included patients with metastatic disease with available survival outcomes (n = 5,335). A total of 3,842 patients were shared between the two cohorts. Kaplan-Meier curves and Cox proportional hazards regression were used to assess patient survival. In the biomarker cohort, 3,063 tumors (42.2%) were strongly classical (SC) and 2,015 tumors (27.8%) were strongly basal (SB). SC and SB tumors showed strong associations with histologic phenotypes and biopsy sites. SB tumors had higher rates of KRAS, TP53, and ARID1A mutations, lower rates of SMAD4 mutation, and transcriptional evidence of epithelial-mesenchymal transition. Sixty of 77 cases (78%) maintained their transcriptional subtype between temporally and/or spatially disparate lesions. In the outcome cohort, the SB subtype was associated with shorter overall survival time, regardless of whether they received FOLFIRINOX or gemcitabine plus nab-paclitaxel as first-line chemotherapy. The mutant KRAS allele type was prognostic of outcomes; however, this impact was restricted to SC tumors, whereas all mutant KRAS alleles had similarly poor outcomes in SB tumors. The SB subtype is a strong independent predictor of worse outcomes, regardless of the up-front chemotherapy regimen used. Clinical trials should further investigate pancreatic cancer transcriptional subtypes as a prognostic and predictive biomarker.

The Role of TGF-β1 and Mutant SMAD4 on Epithelial-Mesenchymal Transition Features in Head and Neck Squamous Cell Carcinoma Cell Lines.

The aim of the present study was to investigate possible differences in the sensitivity of HNSCC cells to known EMT regulators. Three HNSCC cell lines (UM-SCC-1, -3, -22B) and the HaCaT control keratinocyte cell line were exposed to transforming growth factor beta 1 (TGF-β1), a known EMT master regulator, and the cellular response was evaluated by real-time cell analysis (RTCA), Western blot, quantitative PCR, flow cytometry, immunocytochemistry, and the wound closure (scratch) assay. Targeted sequencing on 50 cancer-related genes was performed using the Cancer Hotspot Panel v2. Mutant, and wild type SMAD4 cDNA was used to generate recombinant SMAD4 constructs for expression in mammalian cell lines. The most extensive response to TGF-β1, such as cell growth and migration, β-actin expression, or E-cadherin (CDH1) downregulation, was seen in cells with a more epithelial phenotype. Lower response correlated with higher basal p-TGFβ RII (Tyr424) levels, pointing to a possible autocrine pre-activation of these cell lines. Targeted sequencing revealed a homozygous SMAD4 mutation in the UM-SCC-22B cell line. Furthermore, PCR cloning of SMAD4 cDNA from the same cell line revealed an additional SMAD4 transcript with a 14 bp insertion mutation, which gives rise to a truncated SMAD4 protein. Overexpression of this mutant SMAD4 protein in the highly epithelial control cell line HaCaT resulted in upregulation of TGF-β1 and vimentin. Consistent with previous reports, the invasive and metastatic potential of HNSCC tumor cells appears associated with the level of autocrine secretion of EMT regulators such as TGF-β1, and it could be influenced by exogenous EMT cytokines such as those derived from immune cells of the tumor microenvironment. Furthermore, mutant SMAD4 appears to be a significant contributor to the mesenchymal transformation of HNSCC cells.