Cholangiocarcinoma (CC) is an aggressive cancer with limited prognostic biomarkers. Aquaporin 1 (AQP1), a water channel protein, is implicated in cancer progression. Moreover, DNA methylation plays a key role in epigenetic gene regulation. Therefore, the present study aimed to investigate the prognostic significance of AQP1 promoter methylation haplotypes in CC using high-resolution bisulfite amplicon sequencing. AQP1 mRNA expression in 102 patients was evaluated first to examine its prognostic impact. Subsequently, methylation haplotypes in non-neoplastic and neoplastic tissues from 96 patients were analyzed. Methylation heterogeneity was assessed using t-distributed stochastic neighbor embedding (t-SNE) and k-means clustering. Although AQP1 mRNA expression alone was not a robust independent prognostic factor, a t-SNE-based classification of the methylation signature identified two subgroups with distinct overall survival in both non-neoplastic and neoplastic tissues. Multivariate analysis using Firth’s penalized Cox regression indicated that this methylation signature was independently associated with prognosis (hazard ratio 0.322, p = 0.001). Similar haplotype alterations were observed in non-neoplastic and neoplastic tissues, consistent with the possibility of an epigenetic field effect. These findings suggest that evaluating the methylation signature of the AQP1 promoter may support future less-invasive prognostic assessment and risk stratification, suggesting its potential utility as a candidate biomarker in CC.

Evaluating cancer gene mutations is critical for effective therapeutic selection. Although massive parallel sequencing can efficiently detect gene mutations, most are variants of uncertain significance (VUS). Saturation genome editing (SGE) can facilitate VUS analysis by leveraging CRISPR-Cas9 and homology-directed repair to simultaneously introduce abundant gene mutations. Chronic myelogenous leukemia-derived HAP1 cells are widely used in SGE because of their clear genotype-phenotype relationship; however, the sole use of haploid cells limits SGE applicability in cancer research. Therefore, we developed an SGE-based system for evaluating KRAS mutations in diploid HCT 116 colon carcinoma cells. Single-nucleotide variants (SNVs) in KRAS codons A11-V14 were generated using Cas9-based SGE. Massive parallel sequencing revealed increased abundance of KRASG12 and KRASG13 SNVs and decreased abundance of the KRASG12C SNV after KRASG12C inhibitor treatment in SGE pooled cells. These results indicate that SGE is applicable to diploid HCT 116 cells and useful for evaluating SNV population changes and drug sensitivity. Thus, although haploid HAP1 cells are the primary models for SGE, the successful application of SGE to diploid HCT 116 colon carcinoma cells provides a practical framework for implementing SGE in KRAS-dependent carcinoma cells.

In the 2021 WHO Central Nervous System tumor classification, the “Glioblastoma, IDH-wildtype” diagnosis changed markedly. In a Japanese cohort, we compared the clinical backgrounds and prognoses of molecular glioblastoma (mGBM) and conventional glioblastoma (histological glioblastoma, hGBM). We included 270 patients with glioblastoma treated at five institutions during 2011–2023. Driver gene analysis was performed using a brain tumor-specific custom gene panel to verify the association between molecular and clinical information. Patients with mGBM had better preoperative KPS, lower Ki-67, and lower removal rates than did those with hGBM. Overall survival was longer in patients with mGBM than in those with hGBM (1207 vs 599 days, p = 0.037). TP53 mutation (hazard ratio: 5.33, 95

Introduction To identify prognostic biomarkers that could predict how well patients will respond to lenvatinib/pembrolizumab (LEN/PEM). The utility of certain inflammatory biomarkers in endometrial liquid-based cytology (LBC) or peripheral blood samples, such as neutrophil counts, lymphocyte counts, and neutrophil-to-lymphocyte ratio (NLR) were explored. Methods The study included 25 patients with advanced or recurrent endometrial cancer who had received LEN/PEM between August 2018 and March 2024. Predictors for overall response (OR), disease control, and progression-free survival, based on neutrophil/lymphocyte counts, NLR scores of the endometrial LBC prior to initial treatment, and peripheral blood prior to initial treatment and prior to LEM/PEM treatment were compared using a receiver operating characteristic curve. Significant predictors were evaluated using the log-rank test, and multivariate analysis. Results Although neutrophil counts and NLR score in endometrial LBC prior to initial treatment were better effective predictors for OR, the most accurate predictor of a progression-free status was NLR score in peripheral blood prior to LEM/PEM (0.722, 95% CI: 0.45-0.99, sensitivity: 57.1%, specificity: 94.4%). In peripheral blood prior to LEN/PEM, the lower NLR (NLR <5.39) group had a significantly longer PFS than the higher NLR (5.39 <= NLR) group (p = 0.023, median survival: 13.5 vs. 3.0 months), and tended to be independently correlated with PFS (hazard ratio = 2.571; 95% CI = 0.857-7.719; p = 0.092). Conclusion Inflammatory biomarkers in endometrial LBC failed to predict the efficacy of LEN/PEM, while peripheral blood NLR score sampled prior to LEN/PEM potentially could be a significant predictor.

Comprehensive genomic profiling (CGP) is increasingly used as a clinical laboratory test and being applied to cancer treatment; however, standardization and external quality assessments (EQA) have not been fully developed. This study performed cost-effective EQA and proficiency tests (PT) for CGP testing among multiple institutions those belong to the EQA working group of Japan Association for Clinical Laboratory Science (JACLS). This study revealed that preanalytical processes, such as derived nucleic acids (NA) extraction from formalin fixed paraffine embedded (FFPE) samples, are critical. First, EQA with extracted DNA from cell lines showed a detection rate of 100% (9 out of 9) in KRAS (c.38G > A; p.G13D), PIK3CA (p.H1047R), and B-Raf proto-oncogene, serine/threonine kinase (BRAF) (c.1799 T > A; p.V600E) in cases of > 10% variant allele frequency (VAF). However, BRAF (c.1799 T > A; p.V600E) detection decreased to 67% (6 out of 9) for a VAF of 4.9%. Second, when DNA was extracted from FFPE samples, pathogenic variants and variants with companion diagnostic indications were detected in all 10 participating laboratories. Each variant had < 20% VAFs on average (8.1-19.1%) and wide variability among laboratories was observed (relative standard deviation, 13-60%). Nonetheless, BRAF (c.1798_1799delinsAA; p.V600K) of 8.1% VAF, EGFR (c.2235_2249del; p.E746_A750del) of 9.7% VAF, and EGFR (c.2254_2277del; p.S752_I759del) of 9.8% VAF were detected with 70% (7/10), 70% (7/10), and 60% (6/10) frequency, respectively. Therefore, 10% VAF in pre-analytic processing for DNA extraction from FFPE was critical for variant detection in CGP analysis. Further, incorrect results were reported in case independent variant calling of BRAF; c.1798_1799delinsAA (p.V600K) was mistakenly interpreted as c.1798G > A, and c.1799 T > A was on the other strand. In conclusion, the EQA/PT among 10 institutes with common samples revealed the importance of VAF in pre-analysis and helped us understand the significance of the pipeline and common pitfalls usually ignored by the internal quality control in a single institute.

Here, we present a rare case of multicentric glioblastoma in which two spatially distinct tumors exhibited different molecular profiles. A 56-year-old woman presented with progressive headache and speech difficulties. Magnetic resonance imaging (MRI) of the brain revealed two separate contrast-enhancing lesions, one in the left superior temporal gyrus and the other in the right middle temporal gyrus, with no connecting tract or edema on fluid-attenuated inversion recovery (FLAIR) imaging. Multicentric glioblastoma was suspected, and the patient underwent two-stage surgical resection of both tumors to achieve gross total removal. Histopathology confirmed both lesions as isocitrate dehydrogenase (IDH)-wildtype glioblastoma (World Health Organization (WHO) Grade 4), with typical features including extensive necrosis and microvascular proliferation. However, targeted next-generation sequencing revealed distinct genetic alterations in the two tumors. The left temporal lobe lesion harbored TERT promoter (TERTp) and BRAF p.Val600Glu (V600E) mutations. In contrast, the right temporal lobe lesion harbored a TERTp mutation, epidermal growth factor receptor (EGFR) amplification, MDM4 amplification, and a homozygous deletion of CDKN2A/B. The patient underwent standard postoperative chemoradiotherapy. One year after surgery, the right tumor recurred, whereas no recurrence was observed in the left temporal lobe. The BRAF p.Val600Glu (V600E) mutation in the left-sided tumor is a potential target for molecular therapy and may have contributed to the lack of recurrence. Multicentric glioblastomas are extremely rare, and only a few cases have been genetically profiled to date. Notably, most reported multicentric glioblastomas exhibit monoclonal driver mutations across lesions, suggesting a single origin. In the present case, the two tumors had different molecular signatures. This case underscores the importance of comprehensive genetic analysis of multicentric glioblastomas and highlights how divergent molecular pathologies can inform the prognosis and open opportunities for targeted treatment.

BACKGROUND An ileal neobladder is a standardized form of urinary diversion that provides acceptable outcomes in terms of long-term quality of life. Urothelial carcinomas (UCs) arising in the ileal neobladder are extremely rare, and few reports on this have been published in the English language. CASE SUMMARY We report a case of UC that developed in the ileal neobladder of a 63-year-old man. The patient was diagnosed with UC in situ and underwent radical cystoprostatectomy and ileal neobladder creation. Ten years after the surgery and neoadjuvant chemotherapy, an UC developed in the ileal neobladder. CONCLUSION Ileal neobladder urothelial carcinoma can originate from the implanted urothelium and the intestinal mucosa can migrate intraluminally.

Objective L-methyl-11C-methionine (MET)- and 18F-fluorodeoxyglucose (FDG)-positron emission tomography (PET) are used to detect gliomas. However, the efficacy of MET-PET and FDG-PET in detecting gene alterations in gliomas remains unclear. Therefore, in this study, we evaluated the relationship between genetic alterations and PET tracer uptake in diffuse astrocytic glioma. Methods Thirty-two patients who had been newly diagnosed with astrocytic gliomas at Kagoshima University and Kyushu University and had undergone MET-PET and FDG-PET were enrolled. They underwent analysis of glioma-related gene expression using a customized 48-gene panel. Results The tumors identified in this study were classified as follows: glioblastomas, isocitrate dehydrogenase (IDH) wildtype (n = 15); astrocytic glioma, IDH-mutant, World Health Organization (WHO) grade 4 (n = 2); astrocytic glioma, IDH-mutant, WHO grade 3 (n = 4); astrocytic glioma, IDH-mutant, WHO grade 2 (n = 7); and diffuse astrocytic glioma, not elsewhere classified (n = 4). Astrocytic tumors with IDH mutations, ATRX mutations, and/or loss of function (mut/loss) had a significantly lower tumor-to-normal tissue (T/N) ratio on the MET-PET and FDG-PET images compared with those without these alterations. Astrocytic tumors with CDKN2A/B homozygous deletions (HD), EGFR mutation and/or amplification (mut/amp), or PTEN mut/loss had a significantly higher T/N ratio on the MET-PET images compared with those without these alterations. Astrocytic tumors with NF1 mut/loss had a significantly higher T/N ratio on their FDG-PET images compared with those without these alterations. The cut-off T/N ratio for the MET-PET images for the identification of EGFR mut/amp was 4.50 (sensitivity: 95%; specificity: 56%, AUC: 0.77), and that for detecting CDKN2A/B HDwas 2.32 (sensitivity: 72%; specificity: 86%; AUC: 0.85). Conclusion These findings from our small, retrospective cohort study suggest that MET-PET and FDG-PET are potentially valuable approaches for preoperatively predicting the molecular status of gliomas, particularly for assessing tumors with EGFR mut/amp and CDKN2A/B HD. Preoperative genetic alteration prediction in astrocytic gliomas based on PET tracer uptake may provide accurate information for patients and inform clinical decision-making. Multicenter prospective trials are essential.

Background Molecular classification of endometrial cancer is useful for predicting prognosis. Genomic examinations are performed using formalin-fixed paraffin-embedded (FFPE) tissues; however, we previously reported that liquid-based cytology (LBC) specimens can be used for next-generation sequencing (NGS). In this study, we evaluated long-term storage effects of LBC specimens on NGS-based genomic profiling, including gene mutations, tumor mutation burden (TMB), and microsatellite instability (MSI).Methods Four LBC fixatives (CellPrep, ThinPrep, CytoRich Red, and SurePath) were used to prepare NGS samples from cultured endometrioid carcinoma HEK-251 cells. Twelve endometrial LBC specimens from patients with endometrioid carcinoma were fixed with CytoRich Red. The TMB, MSI, and gene mutations were analyzed after 1 week, 6 months, and 12 months of storage in cultured HEK-251 cells. Paired LBC and FFPE specimens of endometrioid carcinoma stored for 15-45 months were subjected to NGS-based analysis, and their molecular profiles were compared to those at the initial diagnosis.Results The TMB and MSI did not differ during the storage periods for any of the LBC fixatives in the cultured cells; in addition to gene mutations, they were comparable between the initial and second analyses of the clinical FFPE and LBC specimens. There were no changes in the integrative diagnosis of the endometrioid carcinoma subtype classification.Conclusion LBC specimens, which preserved high-quality genomes for molecular classification after long-term storage, may be an alternative source of genomic examination for the integrative diagnosis of endometrial cancer.

This report presents a rare case involving an extreme epithelial-to-mesenchymal transition, in which a specific type of sarcoma developed heterochronically as a recurrence of endometrioid carcinoma. A female in her 50’s presented with abnormal genital bleeding, and an endometrial biopsy revealed endometrioid carcinoma. Following the diagnosis of stage IA endometrioid carcinoma according to the 2008 classification system of the International Federation of Gynecology and Obstetrics, a robot-assisted simple hysterectomy, bilateral salpingo-oophorectomy, and sentinel lymph node navigation surgery were performed. Six months postoperatively, a tumor mass developed in the pelvis. A transrectal needle biopsy revealed spindle cell proliferation, and pelvic tumor resection was conducted for diagnostic therapy. The patient received no adjuvant chemotherapy or radiotherapy after the second surgery and remained free of tumor recurrence for 8 months. The resected yellowish solid tumor mass, measuring 16 × 12 × 9 cm, exhibited hemorrhage, necrosis, and cystic degeneration and was composed of fascicular proliferation of spindle tumor cells showing nuclear pleomorphism and frequent mitotic figures within a myxoid and inflammatory stroma. No epithelial component or organoid patterns were observed. Immunohistochemically, the tumor cells were positive for factor XIIIa, CD10, and cyclin D1, but negative for keratins (AE1/AE3 and CAM5.2) and other specific markers, supporting a diagnosis of high-grade myxoinflammatory fibroblastic sarcoma (MIFS). Genomic analysis revealed identical mutations in PTEN, PIK3R1, CDKN2 A, and TP53 in both the primary uterine endometrioid carcinoma and heterochronic pelvic MIFS. An integrative approach involving histology, immunohistochemistry, and genomic analysis is critical for elucidating the pathogenesis of rare pelvic and uterine tumors.

In glioblastoma (GBM), methylation of the O6-methylguanine-DNA methyltransferase (MGMT) gene promoter is a critical predictive biomarker for temozolomide (TMZ) therapy. Conventional qualitative assessment by methylation-specific PCR (MSP) cannot determine the degree of methylation and risks false positives. While quantitative methods like pyrosequencing are now available, the clinical significance of low-level methylation remains debated. This study aimed to investigate the impact of MGMT promoter methylation on gene expression and prognosis in greater detail using Bisulfite Amplicon Sequencing (BSAS), a high-resolution method employing next-generation sequencing (NGS). DNA was extracted from GBM patient tumor tissues and treated with bisulfite. The methylation status of the MGMT promoter region was then quantified at single-base resolution using BSAS. MGMT mRNA expression levels in the same samples were quantified using RNA-sequencing (RNA-seq). Correlations between methylation levels, mRNA expression, and clinical data were statistically analyzed. Detailed analysis by BSAS revealed that the direct quantitative correlation between MGMT promoter methylation level and mRNA expression was much weaker than previously thought. Nevertheless, the presence of even a low level of promoter methylation was confirmed to be a powerful, independent prognostic factor significantly correlated with improved overall survival. These findings suggest that MGMT promoter methylation may function less as a direct “switch” that significantly suppresses gene expression, and more as a sensitive “biomarker” reflecting a favorable biological state of the tumor. This study highlights the importance of quantitative methylation assessment and suggests a more complex biological role for this key epigenetic mark.

Cholangiocarcinoma (CC) is a highly lethal malignancy that urgently requires reliable prognostic biomarkers. Although MUC1 expression and promoter methylation have been implicated in CC, the clinical significance of promoter methylation pattern composition, beyond average methylation levels, remains unclear. Here, we investigated the relationship between MUC1 promoter methylation heterogeneity, MUC1 mRNA expression, and prognosis in CC. We analyzed bisulfite amplicon sequencing data and mRNA expression of MUC1, DNA methylation-related enzymes (TET1, TET2, TET3, Dnmt1, and Dnmt3a), and tumor microenvironment stress markers in 131 CC tissues. In the neoplastic region, high MUC1 mRNA expression was associated with poor overall survival (HR = 0.131, 95% CI: 0.02 to 0.95, p = 0.042) and correlated with the abundance of completely unmethylated promoter patterns (r = 0.386, p < 0.001). Among the enzymes analyzed, only TET3 expression significantly correlated with the abundance of completely unmethylated patterns in the neoplastic region (Cohen’s f2 = 0.108, p = 0.009), suggesting a potential region-specific regulatory association. We visualized beta-diversity in methylation pattern composition using t-SNE and classified samples into two groups based on a linear decision boundary in the t-SNE space. This classification stratified prognosis independently of clinical factors (HR = 0.291, 95% CI: 0.06 to 0.94, p = 0.037; multivariate p = 0.021). These findings propose a novel, composition-based epigenetic stratification framework in CC, revealing that MUC1 promoter methylation pattern structure—rather than average methylation level—has prognostic relevance. Our results highlight the potential of pattern-resolved methylation profiling in the development of clinically applicable epigenetic biomarkers.

A male in his seventies presented with lung cancer in the right lower lobe. The surgically resected specimen revealed a pleomorphic carcinoma featuring an adenocarcinoma component with lepidic, acinar, and papillary patterns, alongside a spindle cell component spreading along the pulmonary artery wall, resembling intimal sarcoma. The spindle tumor cells were positive for keratins, TTF-1, napsin A, and vimentin, but negative for p40, CK14, desmin, alpha-smooth muscle actin, CDK4, and MDM2. This profile indicates that the spindle cells originated from the adenocarcinoma cells via epithelial-mesenchymal transition. The vascular spread of pleomorphic carcinoma, mimicking pulmonary artery intimal sarcoma, represents an extremely rare mode of tumor invasion, which could be termed "arterial carcinomatosis". Immunohistochemical analysis is crucial for distinguishing pleomorphic carcinoma from coexisting intimal sarcoma, as the latter's prognosis is markedly poor.

Recently, immune checkpoint inhibitors (ICIs) and cabozantinib, a tyrosine kinase inhibitor (TKI), have been used to treat renal cell carcinoma (RCC); the combination of these agents has become a standard treatment for RCC. TKIs generally target vascular endothelial growth factor. However, cabozantinib is characterized by its targeting of MET. Therefore, cabozantinib can be used as a late-line therapy for TKI-resistant RCC. According to data from The Cancer Genome Atlas (TCGA), heat shock transcription factor 4 (HSF4) expression is higher in RCC tissues than in normal renal tissues. HSF4 binds to the MET promoter in colorectal carcinoma to enhance MET expression and promote tumor progression. However, the functional role of HSF4 in RCC is unclear. We performed loss-of-function assays of HSF4, and our results showed that HSF4 knockdown in RCC cells significantly decreased cell functions. Moreover, MET expression was decreased in HSF4-knockdown cells but elevated in sunitinib-resistant RCC cells. The combination of cabozantinib and HSF4 knockdown reduced cell proliferation in sunitinib-resistant cells more than each monotherapy alone. Furthermore, HSF4 knockdown combined with an ICI showed synergistic suppression of tumor growth in vivo. Overall, our strategy involving HSF4 knockdown may enhance the efficacy of existing therapies, such as cabozantinib and ICIs.

BACKGROUND/OBJECTIVES:Krüppel-like factor 4 (KLF4) hotspot mutations are linked to non-invasive intraductal papillary mucinous neoplasms (IPMNs); however, the molecular mechanisms driving their progression remain unclear. Therefore, in this study, we aimed to identify the signaling pathways associated with KLF4 mutations and their roles in IPMNs. METHODS:Thirty-three resected specimens of IPMNs were collected. Associations between genetic alterations in KLF4 and the expression levels of KLF4 protein as well as its related signaling proteins, including p21, p27, and cyclin D, were assessed. RESULTS:KLF4 mutations were found in eight specimens of IPMNs (24 %), all of which were non-invasive lesions and non-intestinal type; of them, seven (87.5 %) showed GNAS mutations. p21 expression was significantly elevated in KLF4-mutated IPMNs (p < 0.01), whereas p27 and cyclin D1 levels were not significantly altered compared with KLF4 wild-type IPMNs. KLF4 and p21 expression levels were significantly elevated in non-invasive lesions compared with invasive lesions (p < 0.05). Higher KLF4 expression was observed in GNAS-mutated IPMNs than in GNAS wild-type IPMNs (p < 0.05). CONCLUSIONS:KLF4 mutations might enhance p21 expression, possibly influencing the indolent characteristics of non-invasive IPMNs and providing a new biomarker strategy for risk stratification in patients with IPMNs.

BACKGROUND AND AIMS:Gallbladder adenocarcinoma (GBAC) is associated with high mortality because of the difficulty in its early detection and treatment. Therefore, identifying prognostic factors is crucial for managing patients with GBAC. We aimed to elucidate the relationship between immunohistochemical profiles of mucin expression, clinicopathological behavior, and prognosis for curatively resected gallbladder adenocarcinoma (GBAC) and to prove that mucin expression is a prognostic factor for GBAC. METHODS:We examined the expression of mucins (MUC1, MUC2, and MUC4) using immunohistochemical analyses and compared the prevalence of each mucin with clinicopathological features in patients with early-stage (stage 0 to IIB) GBAC. RESULTS:MUC1 expression was significantly expressed in patients with GBAC with lymphatic invasion, vascular invasion, perineural invasion, and recurrence. MUC2 was significantly expressed in patients with GBAC with perineural invasion and recurrence but not with prognosis. Patients with MUC1-high expression exhibited worse prognoses than those with MUC1-low expression. In contrast, patients with positive MUC4 expression had significantly better prognoses than those without MUC4 expression. CONCLUSION:The expression of MUC1 and MUC4 in GBAC is an independent prognostic factor for survival and a useful marker for predicting the outcomes of patients with GBAC.

A 49-year-old woman presented with abdominal distention and right leg numbness. Bone biopsy from a huge tumor in her pelvis revealed chondrosarcoma, and proton beam therapy was performed. Although there were no abnormalities at 6 months, dry cough, dyspnea and fever with elevated inflammation appeared at 9 months. Computed tomography showed multiple consolidations in both lungs, different from typical images of nodular shadows with calcification in pulmonary metastases of chondrosarcoma. We first suspected infection or organizing pneumonia, but biopsy revealed chondrosarcoma metastases with severely infiltrated inflammatory cells. Imaging patterns we experienced may reflect inflammatory pathogenesis due to rapid cancer progression.

BACKGROUND/AIM:As the pathophysiology of Microminipigs (μMPs) is similar to that of human, μMPs are useful in atherosclerosis research. To clarify the effect of methotrexate (MTX) on atherosclerosis, we investigated the pathology of MTX-induced atherosclerosis lesion exacerbation in μMPs fed a high-fat and high-cholesterol diet (HFHCD). MATERIALS AND METHODS:The μMPs were divided into four groups: HFHCD, HFHCD+MTX, HFHCD+MTX+leucovorin (LV), and HFHCD+MTX+folic acid (FA), and fed for two weeks. Laboratory tests including blood lipid, FA, and homocysteine (Hcy) levels, and pathological evaluation of the atherosclerosis lesion area and thickness were performed. Hepatic and jejunal gene expressions related to lipid and folate metabolism pathways including 5-methyltetrahydrofolate-homocysteine methyltransferase (MTR) were monitored using RT-PCR. RESULTS:The HFHCD+MTX group showed increased blood Hcy (p<0.01) and decreased FA levels (p<0.05) in accordance with increased hepatic MTR mRNA expression (p<0.1) and exacerbation of atherosclerosis (p=0.051 for lesion area and p=0.045 for lesion thickness) compared to the HFHCD group. Administration of LV or FA attenuated the MTX-induced increase in the Hcy level (p<0.01), atherosclerosis lesion thickness (p<0.1), and MTR mRNA expression (p<0.1 in HFHCD+MTX vs. HFHCD+MTX+LV groups). CONCLUSION:MTX exacerbated HFHCD-induced atherosclerosis mediated through reduced blood FA and the subsequent increase of Hcy in μMPs, indicating that the μMP model may advance cardio-oncology research by providing useful experimental approaches. As MTX is administered for rheumatoid arthritis and malignant tumors in humans, atherosclerosis exacerbation should be acknowledged as a possible adverse effect of MTX treatment.

Kinase-related gene fusion and point mutations play pivotal roles as drivers in cancer, necessitating optimized, targeted therapy against these alterations. The efficacy of molecularly targeted therapeutics varies depending on the specific alteration, with great success reported for such therapeutics in the treatment of cancer with kinase fusion proteins. However, the involvement of actionable alterations in solid tumors, especially regarding kinase fusions, remains unclear. Therefore, in this study, we aimed to compare the number of actionable alterations in patients with tyrosine or serine/threonine kinase domain fusions, mutations, and copy number alterations (CNAs). We analyzed 613 patients with 40 solid cancer types who visited our division between June 2020 and April 2024. Furthermore, to detect alterations involving multiple-fusion calling, we performed comprehensive genomic sequencing using FoundationOne® companion diagnostic (F1CDx) and FoundationOne® Liquid companion diagnostic (F1LCDx). Patient characteristics and genomic profiles were analyzed to assess the frequency and distribution of actionable alterations across different cancer types. Notably, 44 of the 613 patients had fusions involving kinases, transcriptional regulators, or tumor suppressors. F1CDx and F1LCDx detected 13 cases with kinase-domain fusions. We identified 117 patients with kinase-domain mutations and 58 with kinase-domain CNAs. The number of actionable alterations in patients with kinase-domain fusion, mutation, or CNA (median [interquartile range; IQR]) was 2 (1-3), 5 (3-7), and 6 (4-8), respectively. Patients with kinase fusion had significantly fewer actionable alterations than those with kinase-domain mutations and CNAs. However, those with fusion involving tumor suppressors tended to have more actionable alterations (median [IQR]; 4 [2-9]). Cancers with kinase fusions exhibited fewer actionable alterations than those with kinase mutations and CNAs. These findings underscore the importance of detecting kinase alterations and indicate the pivotal role of kinase fusions as strong drivers of cancer development, highlighting their potential as prime targets for molecular therapeutics.