Association between hepatic angiosarcoma and end-stage renal disease: nationwide population-based evidence and enriched mutational signature of aristolochic acid exposure.

Hepatic angiosarcoma (HAS) is an aggressive mesenchymal malignancy that remains underexplored with respect to its etiology and mutational landscapes. To clarify the association between HAS and end-stage renal disease (ESRD), we used nationwide data of the National Health Insurance Research Database (NHIRD) in Taiwan, covering ~99% of the population, from 2001 to 2016. To investigate molecular signatures, we performed whole-exome sequencing (WES) in 27 surgical specimens, including nine ESRD-associated cases. The NHIRD analysis demonstrated that HAS ranked second among all angiosarcomas in Taiwan, with the incidence rates of HAS being 0.08, 2.49, and 5.71 per 100,000 person-years in the general population, chronic kidney disease (CKD), and ESRD patients, respectively. The standardized incidence ratios of HAS in CKD and ESRD patients were 29.99 and 68.77, respectively. In comparison with nonhepatic angiosarcoma, the multivariate regression analysis of our institutional cohort confirmed CKD/ESRD as an independent risk factor for HAS (odds ratio: 9.521, 95% confidence interval: 2.995-30.261, p < 0.001). WES identified a high tumor mutation burden (TMB; median: 8.66 variants per megabase) and dominant A:T-to-T:A transversion in HAS with frequent TP53 (81%) and ATRX (41%) mutations, KDR amplifications/gains (56%), and CDKN2A/B deletions (48%). Notably, ESRD-associated HAS had a significantly higher TMB (17.62 variants per megabase, p = 0.01) and enriched mutational signatures of aristolochic acid exposure (COSMIC SBS22, p < 0.001). In summary, a significant proportion of HAS in Taiwan is associated with ESRD and harbors a distinctive mutational signature, which concomitantly links nephrotoxicity and mutagenesis resulting from exposure to aristolochic acid or related compounds. A high TMB may support the eligibility for immunotherapy in treating ESRD-associated HAS. © 2023 The Pathological Society of Great Britain and Ireland.

Global Genome Diversity and Recombination in Mycoplasma pneumoniae.

Genomic changes in Mycoplasma pneumoniae caused by adaptation to environmental or ecologic pressures are poorly understood. We collected M. pneumoniae from children who had confirmed pneumonia in Taiwan during 2017-2020. We used whole-genome sequencing to compare these isolates with a worldwide collection of current and historical clinical strains for characterizing population structures. A phylogenetic tree for 284 strains showed that all sequenced strains consisted of 5 clades: T1-1 (sequence type [ST]1), T1-2 (mainly ST3), T1-3 (ST17), T2-1 (mainly ST2), and T2-2 (mainly ST14). We identified a putative recombination block containing 6 genes (MPN366‒371). Macrolide resistance involving 23S rRNA mutations was detected for each clade. Clonal expansion of macrolide resistance occurred mostly within subtype 1 strains, of which clade T1-2 showed the highest recombination rate and genome diversity. Functional characterization of recombined regions provided clarification of the biologic role of these recombination events in the evolution of M. pneumoniae.

SNAP29 mediates the assembly of histidine-induced CTP synthase filaments in proximity to the cytokeratin network.

Under metabolic stress, cellular components can assemble into distinct membraneless organelles for adaptation. One such example is cytidine 5'-triphosphate synthase (CTPS, for which there are CTPS1 and CTPS2 forms in mammals), which forms filamentous structures under glutamine deprivation. We have previously demonstrated that histidine (His)-mediated methylation regulates the formation of CTPS filaments to suppress enzymatic activity and preserve the CTPS protein under glutamine deprivation, which promotes cancer cell growth after stress alleviation. However, it remains unclear where and how these enigmatic structures are assembled. Using CTPS-APEX2-mediated in vivo proximity labeling, we found that synaptosome-associated protein 29 (SNAP29) regulates the spatiotemporal filament assembly of CTPS along the cytokeratin network in a keratin 8 (KRT8)-dependent manner. Knockdown of SNAP29 interfered with assembly and relaxed the filament-induced suppression of CTPS enzymatic activity. Furthermore, APEX2 proximity labeling of keratin 18 (KRT18) revealed a spatiotemporal association of SNAP29 with cytokeratin in response to stress. Super-resolution imaging suggests that during CTPS filament formation, SNAP29 interacts with CTPS along the cytokeratin network. This study links the cytokeratin network to the regulation of metabolism by compartmentalization of metabolic enzymes during nutrient deprivation.

Adult NTRK-rearranged spindle cell neoplasms of the viscera: with an emphasis on rare locations and heterologous elements.

NTRK-rearranged mesenchymal neoplasms mostly affect the soft tissues of pediatric patients. Given the responsiveness to selective NTRK inhibitors, it remains critical to identify those ultra-rare cases occurring in the viscera of adults. In five females and two males aged 18-53 years, we characterized visceral mesenchymal tumors harboring TPM3-NTRK1 [uterine cervix (N = 2), pleura, prostate], LMNA-NTRK1 (lung), SQSTM1-NTRK3 (heart), and NTRK3 rearrangement with unknown fusion partner (colon/mesocolon) with RNA sequencing, FISH, RT-PCR, and immunohistochemistry. The tumors exhibited spindled to ovoid/epithelioid or pleomorphic cells, often arranged in fascicles, and were low-to-intermediate-grade and high-grade in three and four cases, respectively. Keloid-like stromal collagen and perivascular hyalinization was noted in five. Adenosarcoma-like appearances were observed in two, manifesting frond-like protrusions in one cervical tumor and phyllodes-like architecture in the prostatic tumor. Abrupt high-grade transformation into pleomorphic liposarcoma was found in another cervical tumor, while the pleural tumor contained intermixed rhabdomyoblasts. Pan-TRK immunostaining was positive in all cases. All cases expressed CD34, while five were S100-positive. CDKN2A homozygous deletion with concomitant p16 loss occurred in 4/7. Whole-exome sequencing identified TP53 mutation (c.672+2T>C, involving a splice site, with concomitant protein loss) in a cervical sarcoma, limited to its heterologous liposarcomatous component. At least moderate pan-TRK immunoreactivity was present in varying proportions of potential pathologic mimics, with BCOR-positive sarcoma (56%, 5/9), undifferentiated uterine sarcoma (50%, 3/6), and spindle cell/sclerosing rhabdomyosarcoma (33%, 2/6) being among the most frequent. This underscored the unsatisfactory specificity of pan-TRK immunohistochemistry and warranted molecular confirmation in the diagnosis of adult NTRK-rearranged visceral mesenchymal neoplasms. The current report highlights the ever-expanding clinicopathologic and genetic spectrum of this entity by describing the unprecedented cardiac and pleural locations and heterologous differentiation, as well as the second NTRK-rearranged "prostatic stromal sarcoma," while substantiating CDKN2A deletion as a frequent occurrence.

Identification of Salivary Biomarkers for Oral Cancer Detection with Untargeted and Targeted Quantitative Proteomics Approaches.

Oral cavity squamous cell carcinoma (OSCC) is one of the most common cancers worldwide. In Taiwan, OSCC is the fifth leading cause of cancer-related mortality and leads to 2800 deaths per year. The poor outcome of OSCC patients is principally ascribed to the fact that this disease is often advanced at the time of diagnosis, suggesting that early detection of OSCC is urgently needed. Analysis of cancer-related body fluids is one promising approach to identify biomarker candidates of cancers. To identify OSCC biomarkers, salivary proteomes of OSCC patients, individuals with oral potentially malignant disorders (OPMDs), and healthy volunteers were comparatively profiled with isobaric tags for relative and absolute quantitation (iTRAQ)-based mass spectrometry (MS). The salivary levels of 67 and 18 proteins in the OSCC group are elevated and decreased compared with that in the noncancerous group (OPMD and healthy groups), respectively. The candidate biomarkers were further selected using the multiple reaction monitoring (MRM)-MS and validated with the immunoassays. More importantly, the higher salivary level of three proteins, complement factor H (CFH), fibrinogen alpha chain (FGA), and alpha-1-antitrypsin (SERPINA1) was correlated with advanced stages of OSCC. Our results indicate that analysis of salivary proteome is a feasible strategy for biomarker discovery, and the three proteins are potential salivary markers for OSCC diagnosis.

Genomic Insight into the Spread of Meropenem-Resistant Streptococcus pneumoniae Spain23F-ST81, Taiwan.

Incidence of invasive pneumococcal disease caused by antimicrobial-resistant Streptococcus pneumoniae types not included in pneumococcal conjugate vaccines has increased, including a penicillin- and meropenem-resistant serotype 15A-ST63 clone in Japan. During 2013-2017, we collected 206 invasive pneumococcal isolates in Taiwan for penicillin and meropenem susceptibility testing. We found serotypes 15B/C-ST83 and 15A-ST63 were the most prevalent penicillin- and meropenem-resistant clones. A transformation study confirmed that penicillin-binding protein (PBP) 2b was the primary meropenem resistance determinant, and PBP1a was essential for high-level resistance. The rate of serotype 15B/C-ST83 increased during the study. All 15B/C-ST83 isolates showed an ermB macrolide resistance genotype. Prediction analysis of recombination sites revealed 12 recombination regions in 15B/C-ST83 compared with the S. pneumoniae Spain23F-ST81 genome. Pneumococcal clones rapidly recombine to acquire survival advantages and undergo local expansion under the selective pressure exerted by vaccines and antimicrobial drugs. The spread of 15B/C-ST83 is alarming for countries with high antimicrobial pressure.

Tumor-associated intronic editing of HNRPLL generates a novel splicing variant linked to cell proliferation.

Processing of the eukaryotic transcriptome is a dynamic regulatory mechanism that confers genetic diversity, and splicing and adenosine to inosine (A-to-I) RNA editing are well-characterized examples of such processing. Growing evidence reveals the cross-talk between the splicing and RNA editing, but there is a paucity of substantial evidence for its mechanistic details and contribution in a physiological context. Here, our findings demonstrate that tumor-associated differential RNA editing, in conjunction with splicing machinery, regulates the expression of variants of HNRPLL, a gene encoding splicing factor. We discovered an HNRPLL transcript variant containing an additional exon 12A (E12A), which is a substrate of ADAR1 and ADAR2. Adenosine deaminases acting on RNA (ADAR) direct deaminase-dependent expression of the E12A transcript, and ADAR-mediated regulation of E12A is largely splicing-based, and does not affect the stability or nucleocytoplasmic distribution of the transcript. Furthermore, ADAR-mediated modification of exon 12A generates an enhancer for the oncogenic splicing factor SRSF1 and consequently promotes the frequency of alternative splicing. Gene expression profiling by RNA-seq revealed that E12A acts distinctly from HNRPLL and regulates a set of growth-related genes, such as cyclin CCND1 and growth factor receptor TGFBR1 Accordingly, silencing E12A expression leads to impaired clonogenic ability and enhanced sensitivity to doxorubicin, thus highlighting the significance of this alternative isoform in tumor cell survival. In summary, we present the interplay of RNA editing and splicing as a regulatory mechanism of gene expression and also its physiological relevance. These findings extend our understanding of transcriptional dynamics and provide a mechanistic explanation to the link of RNA editors to tumorigenesis.

Redefining aberrant P53 expression of gastric cancer and its distinct clinical significance among molecular-histologic subtypes.

BACKGROUND: Numerous studies have demonstrated a correlation between p53 overexpression and diminished survival in gastric cancer patients. However, conflicting findings exist, and we hypothesize that these discrepancies arise from the cancer's complexity and heterogeneity, coupled with a lack of consensus on aberrant p53 expression. METHODS: We enrolled a cohort of 187 patients with surgically resected gastric cancer. Patient categorization was based on Epstein-Barr virus (EBV), microsatellite instability (MSI), and Lauren classification (intestinal, diffuse and mixed). Utilizing an incremental algorithm, we evaluated p53 immunohistochemical (IHC) patterns in all 187 cases, while next-generation sequencing was successfully performed on 152 cases to identify TP53 mutations (mutTP53). RESULTS: MutTP53 was identified in 32 % of the 152 cases, comprising 36 missense, 5 nonsense, and 7 frameshift alterations. Missense mutations predominantly correlated with p53 overexpression, while nonsense and frameshifting alterations related to null expression. Trial calculations indicated that null expression and a p53 IHC cutoff at >40 % offered the best prediction of mutTP53 (kappa coefficient, 0.427), with the highest agreement (0.524) observed in diffuse type and the lowest (0.269) in intestinal type. Null expression and a p53 IHC cutoff at >10 %, but not mutTP53 per se, provided the optimal prediction of survival outcome (p = 0.043), particularly in diffuse type (p = 0.044). Multivariate analysis showed that aberrant p53 IHC expression was not an independent prognostic factor. CONCLUSIONS: P53 IHC patterns are predictive biomarkers for mutTP53 and gastric cancer outcomes, where a prerequisite involves a nuanced approach considering cutoff values and molecular-histologic subtyping.

Noncanonical formation of SNX5 gene-derived circular RNA regulates cancer growth.

Oral squamous cell carcinoma (OSCC) is a prevalent cancer worldwide, exhibiting unique regional prevalence. Despite advancements in diagnostics and therapy, the 5-year survival rate for patients has seen limited improvement. A deeper understanding of OSCC pathogenesis, especially its molecular underpinnings, is essential for improving detection, prevention, and treatment. In this context, noncoding RNAs, such as circular RNAs (circRNAs), have gained recognition as crucial regulators and potential biomarkers in OSCC progression. Our study highlights the discovery of previously uncharacterized circRNAs, including a SNX5 gene-derived circRNA, circSNX5, through deep sequencing of OSCC patient tissue transcriptomes. We established circSNX5's tumor-specific expression and its strong correlation with patient survival using structure-specific and quantitative PCR analyses. In vitro and in vivo experiments underscored circSNX5 RNA's regulatory role in cancer growth and metastasis. Further, our omics profiling and functional assays revealed that ADAM10 is a critical effector in circSNX5-mediated cancer progression, with circSNX5 maintaining ADAM10 expression by sponging miR-323. This novel circRNA-miRNA-mRNA regulatory axis significantly contributes to oral cancer progression and malignancy. Moreover, we discovered that circSNX5 RNA is produced via noncanonical sequential back-splicing of pre-mRNA, a process negatively regulated by the RNA-binding protein STAU1. This finding adds a new dimension to our understanding of exonic circRNA biogenesis in the eukaryotic transcriptome. Collectively, our findings offer a detailed mechanistic dissection and functional interpretation of a novel circRNA, shedding light on the role of the noncoding transcriptome in cancer biology and potentially paving the way for innovative therapeutic strategies.

CAN008 prolongs overall survival in patients with newly diagnosed GBM characterized by high tumor mutational burden.

BACKGROUND: A previous phase 1 dose-escalation study in Taiwan indicated CAN008 (asunercept) with standard concurrent chemoradiotherapy (CCRT) improved progression-free survival (PFS) in newly diagnosed glioblastoma (GBM) patients. This study evaluates the efficacy of CAN008 in promoting overall survival (OS) and identifies genetic alterations associated with treatment responses. METHODS: We compared OS of 5-year follow-ups from 9 evaluable CAN008 cohort patients (6 received high-dose and 3 received low-dose) to a historical Taiwanese GBM cohort with 164 newly diagnosed patients. CAN008 treatment response-associated genetic alterations were identified by whole-exome sequencing and comparing variant differences between response groups. Associations among patient survival, tumor mutational burden (TMB), and genetic alterations were analyzed using CAN008 cohort and TCGA-GBM dataset. RESULTS: OS for high-dose CAN008 patients at 2 and 5 years was 83% and 67%, respectively, and 40.1% and 8.8% for the historical GBM cohort, respectively. Better OS was observed in the high-dose CAN008 cohort (without reaching the median survival) than the historical GBM cohort (median OS: 20 months; p=0.0103). Five high-dose CAN008 patients were divided into good and poor response groups based on their PFS. A higher variant count and TMB were observed in good response patients, whereas no significant association was observed between TMB and patient survival in the newly diagnosed TCGA-GBM dataset, suggesting TMB may modulate patient CAN008 response. CONCLUSION: CAN008 combined with standard CCRT treatment prolonged the PFS and OS of newly diagnosed GBM patients compared to standard therapy alone. Higher treatment efficacy was associated with higher TMB.