Controlling the confounding effect of metabolic gene expression to identify actual metabolite targets in microsatellite instability cancers.

BACKGROUND: The metabolome is the best representation of cancer phenotypes. Gene expression can be considered a confounding covariate affecting metabolite levels. Data integration across metabolomics and genomics to establish the biological relevance of cancer metabolism is challenging. This study aimed to eliminate the confounding effect of metabolic gene expression to reflect actual metabolite levels in microsatellite instability (MSI) cancers. METHODS: In this study, we propose a new strategy using covariate-adjusted tensor classification in high dimensions (CATCH) models to integrate metabolite and metabolic gene expression data to classify MSI and microsatellite stability (MSS) cancers. We used datasets from the Cancer Cell Line Encyclopedia (CCLE) phase II project and treated metabolomic data as tensor predictors and data on gene expression of metabolic enzymes as confounding covariates. RESULTS: The CATCH model performed well, with high accuracy (0.82), sensitivity (0.66), specificity (0.88), precision (0.65), and F1 score (0.65). Seven metabolite features adjusted for metabolic gene expression, namely, 3-phosphoglycerate, 6-phosphogluconate, cholesterol ester, lysophosphatidylethanolamine (LPE), phosphatidylcholine, reduced glutathione, and sarcosine, were found in MSI cancers. Only one metabolite, Hippurate, was present in MSS cancers. The gene expression of phosphofructokinase 1 (PFKP), which is involved in the glycolytic pathway, was related to 3-phosphoglycerate. ALDH4A1 and GPT2 were associated with sarcosine. LPE was associated with the expression of CHPT1, which is involved in lipid metabolism. The glycolysis, nucleotide, glutamate, and lipid metabolic pathways were enriched in MSI cancers. CONCLUSIONS: We propose an effective CATCH model for predicting MSI cancer status. By controlling the confounding effect of metabolic gene expression, we identified cancer metabolic biomarkers and therapeutic targets. In addition, we provided the possible biology and genetics of MSI cancer metabolism.

Dual role of sprouty2 as an inhibitor of RAS/ERK-driven proliferation and a promoter of cancer invasion in KRAS wild-type colorectal cancer.

Sprouty2 (SPRY2) is known to inhibit the RAS/MAPK/ERK pathway, and is a potential study target for cancer. The effect of SPRY2 in colorectal cancer (CRC) and whether it is influenced by KRAS mutation are not known. We manipulated SPRY2 gene expression and used an activating KRAS-mutant plasmid to determine its effect on CRC cell function in vitro and/or in vivo. We performed SPRY2 immunohistochemical staining in 143 CRC specimens and analyzed the staining results with various clinicopathological characteristics in relation to KRAS mutation status. SPRY2 knockdown in Caco-2 cells carrying the wild-type (WT) KRAS gene upregulated phosphorylated ERK (p-ERK) levels and increased cell proliferation in vitro, but inhibited cell invasion. However, SPRY2 knockdown in SW480 cells (activating KRAS mutant) or Caco-2 cells transfected with KRAS-mutant plasmid did not significantly alter p-ERK levels, cell proliferation, or invasion. The xenografts of SPRY2-knockdown Caco-2 cells were larger with less deep muscle invasion than those of control cells. The clinical cohort study revealed a positive association of SPRY2 protein expression with pT status, lymphovascular invasion, and perineural invasion in KRAS-WT CRCs. However, the associations were not observed in KRAS-mutant CRCs. Interestingly, high SPRY2 expression was related to shorter cancer-specific survival in both KRAS-WT and KRAS-mutant CRC patients. Our study demonstrated the dual role of SPRY2 as an inhibitor of RAS/ERK-driven proliferation and as a promoter of cancer invasion in KRAS-WT CRC. SPRY2 may promote the invasion and progression of KRAS-WT CRC, and might also enhance KRAS-mutant CRC progression through pathways other than invasion.

Distinct Features of Plasma Ultrashort Single-Stranded Cell-Free DNA as Biomarkers for Lung Cancer Detection.

BACKGROUND: Using broad range cell-free DNA sequencing (BRcfDNA-Seq), a nontargeted next-generation sequencing (NGS) methodology, we previously identified a novel class of approximately 50 nt ultrashort single-stranded cell-free DNA (uscfDNA) in plasma that is distinctly different from 167 bp mononucleosomal cell-free DNA (mncfDNA). We hypothesize that uscfDNA possesses characteristics that are useful for disease detection. METHODS: Using BRcfDNA-Seq, we examined both cfDNA populations in the plasma of 18 noncancer controls and 14 patients with late-stage nonsmall cell lung carcinoma (NSCLC). In comparison to mncfDNA, we assessed whether functional element (FE) peaks, fragmentomics, end-motifs, and G-Quadruplex (G-Quad) signatures could be useful features of uscfDNA for NSCLC determination. RESULTS: In noncancer participants, compared to mncfDNA, uscfDNA fragments showed a 45.2-fold increased tendency to form FE peaks (enriched in promoter, intronic, and exonic regions), demonstrated a distinct end-motif-frequency profile, and presented with a 4.9-fold increase in G-Quad signatures. Within NSCLC participants, only the uscfDNA population had discoverable FE peak candidates. Additionally, uscfDNA showcased different end-motif-frequency candidates distinct from mncfDNA. Although both cfDNA populations showed increased fragmentation in NSCLC, the G-Quad signatures were more discriminatory in uscfDNA. Compilation of cfDNA features using principal component analysis revealed that the first 5 principal components of both cfDNA subtypes had a cumulative explained variance of >80%. CONCLUSIONS: These observations indicate that the distinct biological processes of uscfDNA and that FE peaks, fragmentomics, end-motifs, and G-Quad signatures are uscfDNA features with promising biomarker potential. These findings further justify its exploration as a distinct class of biomarker to augment pre-existing liquid biopsy approaches.

CHK2 activation contributes to the development of oxaliplatin resistance in colorectal cancer.

BACKGROUND: Colorectal cancer (CRC), the most common cancer type, causes high morbidity and mortality. Patients who develop drug resistance to oxaliplatin-based regimens have short overall survival. Thus, identifying molecules involved in the development of oxaliplatin resistance is critical for designing therapeutic strategies. METHODS: A proteomic screen was performed to reveal altered protein kinase phosphorylation in oxaliplatin-resistant (OR) CRC tumour spheroids. The function of CHK2 was characterised using several biochemical techniques and evident using in vitro cell and in vivo tumour models. RESULTS: We revealed that the level of phospho-CHK2(Thr68) was elevated in OR CRC cells and in ~30% of tumour samples from patients with OR CRC. We demonstrated that oxaliplatin activated several phosphatidylinositol 3-kinase-related kinases (PIKKs) and CHK2 downstream effectors and enhanced CHK2/PARP1 interaction to facilitate DNA repair. A phosphorylation mimicking CHK2 mutant, CHK2T68D, but not a kinase-dead CHK2 mutant, CHK2D347A, promoted DNA repair, the CHK2/PARP1 interaction, and cell growth in the presence of oxaliplatin. Finally, we showed that a CHK2 inhibitor, BML-277, reduced protein poly(ADP-ribosyl)ation (PARylation), FANCD2 monoubiquitination, homologous recombination and OR CRC cell growth in vitro and in vivo. CONCLUSION: Our findings suggest that CHK2 activity is critical for modulating oxaliplatin response and that CHK2 is a potential therapeutic target for OR CRC.

Comprehensive assessments of germline deletion structural variants reveal the association between prognostic MUC4 and CEP72 deletions and immune response gene expression in colorectal cancer patients.

BACKGROUND: Functional disruptions by large germline genomic structural variants in susceptible genes are known risks for cancer. We used deletion structural variants (DSVs) generated from germline whole-genome sequencing (WGS) and DSV immune-related association tumor microenvironment (TME) to predict cancer risk and prognosis. METHODS: We investigated the contribution of germline DSVs to cancer susceptibility and prognosis by silicon and causal inference models. DSVs in germline WGS data were generated from the blood samples of 192 cancer and 499 non-cancer subjects. Clinical information, including family cancer history (FCH), was obtained from the National Cheng Kung University Hospital and Taiwan Biobank. Ninety-nine colorectal cancer (CRC) patients had immune response gene expression data. We used joint calling tools and an attention-weighted model to build the cancer risk predictive model and identify DSVs in familial cancer. The survival support vector machine (survival-SVM) was used to select prognostic DSVs. RESULTS: We identified 671 DSVs that could predict cancer risk. The area under the curve (AUC) of the receiver operating characteristic curve (ROC) of the attention-weighted model was 0.71. The 3 most frequent DSV genes observed in cancer patients were identified as ADCY9, AURKAPS1, and RAB3GAP2 (p < 0.05). The DSVs in SGSM2 and LHFPL3 were relevant to colorectal cancer. We found a higher incidence of FCH in cancer patients than in non-cancer subjects (p < 0.05). SMYD3 and NKD2DSV genes were associated with cancer patients with FCH (p < 0.05). We identified 65 immune-associated DSV markers for assessing cancer prognosis (p < 0.05). The functional protein of MUC4 DSV gene interacted with MAGE1 expression, according to the STRING database. The causal inference model showed that deleting the CEP72 DSV gene affect the recurrence-free survival (RFS) of IFIT1 expression. CONCLUSIONS: We established an explainable attention-weighted model for cancer risk prediction and used the survival-SVM for prognostic stratification by using germline DSVs and immune gene expression datasets. Comprehensive assessments of germline DSVs can predict the cancer risk and clinical outcome of colon cancer patients.

PD-L1 Expression in High-Risk Early-Stage Colorectal Cancer-Its Clinical and Biological Significance in Immune Microenvironment.

Programmed death-ligand 1 (PD-L1) is an immune checkpoint molecule that can regulate immune responses in the tumor microenvironment (TME); however, the clinical applications of PD-L1 in early-stage colorectal cancer (CRC) remain unclear. In this study, we aimed to investigate the relationship between PD-L1 expression and survival outcome and explore its relevant immune responses in CRC. PD-L1 expression was evaluated by immunohistochemical staining to determine the tumor proportion score and combined positive score (CPS) in a Taiwanese CRC cohort. The oncomine immune response research assay was conducted for immune gene expression analyses. CRC datasets from the TCGA database were reappraised for PD-L1-associated gene enrichment analyses using GSEA. The high expression of PD-L1 (CPS ≥ 5) was associated with longer recurrence-free survival (p = 0.031) and was an independent prognostic factor as revealed by multivariate analysis. High PD-L1 expression was related to six immune-related gene signatures, and CXCL9 is the most significant overexpressed gene in differential analyses. High CXCL9 expression correlated with increased infiltration levels of immune cells in the TME, including CD8+ T lymphocytes and M1 macrophages. These findings suggest that high PD-L1 expression is a prognostic factor of early-stage CRC, and CXCL9 may play a key role in regulating PD-L1 expression.

Sequential and co-occurring DNA damage response genetic mutations impact survival in stage III colorectal cancer patients receiving adjuvant oxaliplatin-based chemotherapy.

BACKGROUND: Certain sequences of genomic mutations can lead to cancer formation and affect treatment outcomes and drug resistance. We constructed a cancer evolutionary tree using bulk-targeted deep sequencing to explore the impact of sequential and co-occurring somatic mutations on patients with stage III colorectal cancer (CRC). METHODS: A total of 108 stage III CRC patients from National Cheng Kung University Hospital (NCKUH) were recruited for this study between Jan. 2014 and Jan. 2019. Clinical information and tumor-targeted deep sequencing data were collected. Phylogenetic trees were reconstructed for evolutionary trajectories. We used a machine learning model for survival analysis. RESULTS: Six sequential somatic mutations stratified patients into seven subgroups based on survival. Patients carrying sequential germline followed by DNA damage response-related ATM or BRCA2 somatic mutations or non-TP53, APC somatic mutations had a better outcome than those without such mutations. The 4-year recurrence-free survival (RFS) probability was 88% in the low-risk group (G1) and 46% in the high-risk group (G2) (log-rank p-value 2e-05). The predictive efficacy by the area under the curve (AUC) was 0.73, 0.7, 0.797, and 0.88 at 2, 4, 6, and 8 years, respectively. The mutation status of mismatch repair (MMR) genes was not associated with RFS. Different genomic features were found between the groups. The orders of APC, KRAS and APC, BRCA2 sequential somatic mutations were associated with clinical outcomes. The occurrence of somatic mutations in BRCA2, such as TP53 somatic mutations, affected recurrence-free survival. CONCLUSIONS: According to the evolution model, DNA damage response (DDR)-related ATM or BRCA2 somatic mutations are promising biomarkers for assessing the response of stage III CRC patients to oxaliplatin-based chemotherapy. The sequential order and co-occurring DDR somatic mutations are associated with recurrence-free survival.

CD40 Pathway and IL-2 Expression Mediate the Differential Outcome of Colorectal Cancer Patients with Different CSF1R c.1085 Genotypes.

Colony-stimulating factor 1 receptor (CSF-1R) acts as the receptor for colony stimulating factor 1, a cytokine that controls the production, differentiation, and function of macrophages. Prior studies showed cancer patients harboring germline CSF1R c.1085A>G genetic variant had better survival. Here, primary tumor samples from a stage III colorectal cancer (CRC) cohort were analyzed by a targeted gene expression assay containing 395 immune-related genes to study the immune mechanism underlying the different outcomes. CRC patients with CSF1R c.1085 genotype A_G had a better disease-free and overall survival than those with CSF1R genotype A_A. Compared to the group of patients without CSF1R variant, higher CD40LG expression, a surface marker of T cells, was found in the tumor tissues of patients with CSF1R c.1085 variant. In parallel with the higher CD40LG gene expression, immunofluorescent staining also showed more CD3+CD40L+ T cell infiltrates in tumors with CSF1R c.1085 genotype A_G. Moreover, higher IL-2 expression, known to be regulated by CD40 pathway, was also observed in tumors with CSF1R c.1085 genotype A_G than genotype A_A. Higher IL-2 expression generated by the interaction of CD40 ligand and CD40 between T cells and macrophages with CSF1R c.1085A>G variant is the potential mechanism explaining the different outcomes.

Treatment monitoring of colorectal cancer by integrated analysis of plasma concentration and sequencing of circulating tumor DNA.

Circulating cell-free DNA (cfDNA) analysis is an important tool for cancer monitoring. The patient-specific mutations identified in colorectal cancer (CRC) tissues are usually used to design the cfDNA analysis. Despite high specificity in predicting relapse, the sensitivity in most studies is around 40-50%. To improve this weakness, we designed a cfDNA panel according to the CRC genomic landscape and recurrent-specific mutations. The pathological variants in cfDNA samples from 60 CRC patients were studied by a next-generation sequencing (NGS) method incorporating the dual molecular barcode. Interestingly, patients in the disease positive group had a significantly higher cfDNA concentration than those in the disease negative group. Based on receiver operating characteristic analysis, the cfDNA concentration of 7 ng/mL was selected into the analytical workflow. The sensitivity in determining the disease status was 72.4%, which represented a considerable improvement on prior studies, and the specificity remained high at 80.6%. Compared to standard imaging and laboratory studies, earlier detection of residual disease and clinical benefits were shown on two cases by this cfDNA assay. We conclude this integrative framework of cfDNA analytical pipeline with a satisfactory sensitivity and specificity could be used in postoperative CRC surveillance.

Emergence of RAS mutations in patients with metastatic colorectal cancer receiving cetuximab-based treatment: a study protocol.

BACKGROUND: In the management of patients with RAS wild-type metastatic colorectal cancer (mCRC), anti-epidermal growth factor receptor (EGFR) therapies have demonstrated a clinical benefit, with longer survival. However, the correlation between the emergence of circulating RAS mutations and secondary resistance to anti-EGFR therapies requires further elucidation. In this study, we aim to examine evolutionary changes in RAS mutations through liquid biopsy in patients with mCRC during and after anti-EGFR therapy. METHODS: A total of 120 patients diagnosed with RAS wild-type mCRC will be enrolled in this study. Patients will receive a cetuximab-based infusional 5-fluorouracil regimen as first-line treatment. Cetuximab-based treatment is expected to continue until disease progression, intolerable toxic effects, or withdrawal of consent. Blood samples from enrolled patients will be collected before and then every 3 months during cetuximab-based treatment and also at disease progression. These blood samples will be evaluated for RAS resistance mutations by using the MassARRAY platform. The primary endpoint is the percentage of RAS mutations detected in circulating DNA from patients during cetuximab treatment. The correlation between the tumor response and survival outcomes of these patients and the emergence of circulating RAS mutations will be further analyzed. DISCUSSION: Liquid biopsy is a powerful technology that can represent tumor heterogeneity in a relatively noninvasive manner. Because RAS mutations play a major role in resistance to anti-EGFR therapy for mCRC, examining evolutionary changes in these mutations during such treatment through liquid biopsy would be useful. After comprehensively analyzing the emergence of circulating RAS mutations and its clinical relevance in this study, our results should provide practical guidance on anti-EGFR therapy for mCRC. TRIAL REGISTRATION: The date of trial registration ( NCT03401957 ) in this study was January 17, 2018.

Monoclonality and cytogenetic abnormalities in hyaline vascular Castleman disease.

Hyaline vascular Castleman disease is traditionally regarded as a reactive hyperplastic process. Occasional cases, however, have been reported with cytogenetic anomalies bringing this concept into question. In this study, we used conventional and methylation-specific polymerase chain reaction methods to assess the human androgen receptor α (HUMARA) gene in 29 female patients with hyaline vascular Castleman disease and compared the results with three cases of plasma cell Castleman disease and 20 cases of age-matched lymphoid hyperplasia. We also assessed for immunoglobulin gene and T-cell receptor gene rearrangements, and conventional cytogenetic analysis was performed in three cases of hyaline vascular Castleman disease. In cases with informative results, conventional and methylation-specific human androgen receptor α gene analyses yielded a monoclonal pattern in 10 of 19 (53%) and 17 of 23 (74%) cases of hyaline vascular Castleman disease, respectively. A monoclonal pattern was also detected in three cases of plasma cell Castleman disease but not in cases of lymphoid hyperplasia. The frequency of monoclonality was higher for lesions >5 cm in size (100%) and for the stromal-rich variant (91%). Cytogenetic abnormalities in stromal cells were revealed in two cases of hyaline vascular Castleman disease and no cases showed monoclonal immunoglobulin or T-cell receptor gene rearrangements. Follow-up data showed persistent disease in 4 of 23 (17%) patients. We conclude that hyaline vascular Castleman disease is often a monoclonal proliferation, most likely of lymph node stromal cells.

Tight junction protein cingulin variant is associated with cancer susceptibility by overexpressed IQGAP1 and Rac1-dependent epithelial-mesenchymal transition.

BACKGROUND: Cingulin (CGN) is a pivotal cytoskeletal adaptor protein located at tight junctions. This study investigates the link between CGN mutation and increased cancer susceptibility through genetic and mechanistic analyses and proposes a potential targeted therapeutic approach. METHODS: In a high-cancer-density family without known pathogenic variants, we performed tumor-targeted and germline whole-genome sequencing to identify novel cancer-associated variants. Subsequently, these variants were validated in a 222 cancer patient cohort, and CGN c.3560C > T was identified as a potential cancer-risk allele. Both wild-type (WT) (c.3560C > C) and variant (c.3560C > T) were transfected into cancer cell lines and incorporated into orthotopic xenograft mice model for evaluating their effects on cancer progression. Western blot, immunofluorescence analysis, migration and invasion assays, two-dimensional gel electrophoresis with mass spectrometry, immunoprecipitation assays, and siRNA applications were used to explore the biological consequence of CGN c.3560C > T. RESULTS: In cancer cell lines and orthotopic animal models, CGN c.3560C > T enhanced tumor progression with reduced sensitivity to oxaliplatin compared to the CGN WT. The variant induced downregulation of epithelial marker, upregulation of mesenchymal marker and transcription factor, which converged to initiate epithelial-mesenchymal transition (EMT). Proteomic analysis was conducted to investigate the elements driving EMT in CGN c.3560C > T. This exploration unveiled overexpression of IQGAP1 induced by the variant, contrasting the levels observed in CGN WT. Immunoprecipitation assay confirmed a direct interaction between CGN and IQGAP1. IQGAP1 functions as a regulator of multiple GTPases, particularly the Rho family. This overexpressed IQGAP1 was consistently associated with the activation of Rac1, as evidenced by the analysis of the cancer cell line and clinical sample harboring CGN c.3560C > T. Notably, activated Rac1 was suppressed following the downregulation of IQGAP1 by siRNA. Treatment with NSC23766, a selective inhibitor for Rac1-GEF interaction, resulted in the inactivation of Rac1. This intervention mitigated the EMT program in cancer cells carrying CGN c.3560C > T. Consistently, xenograft tumors with WT CGN showed no sensitivity to NSC23766 treatment, but NSC23766 demonstrated the capacity to attenuate tumor growth harboring c.3560C > T. CONCLUSIONS: CGN c.3560C > T leads to IQGAP1 overexpression, subsequently triggering Rac1-dependent EMT. Targeting activated Rac1 is a strategy to impede the advancement of cancers carrying this specific variant.

Germline MLH1 and MSH6 mutations from two Lynch syndrome families identified in a patient with early-onset of endometrial cancer: A case report.

Introduction: Lynch syndrome is caused by a germline mutation in mismatch repair (MMR) genes, leading to the loss of expression of MMR heterodimers, either MLH1/PMS2 or MSH2/MSH6, or isolated loss of PMS2 or MSH6. Concurrent loss of both heterodimers is uncommon, and patients carrying pathogenic variants affecting different MMR genes are rare, leading to the lack of cancer screening recommendation for these patients.Case presentation:Here, we reported a female with a family history of Lynch syndrome with MLH1 c.676C > T mutation. She developed endometrial cancer at 37 years old, with loss of MLH1/PMS2 expression. Immunohistochemical staining on tumor samples incidentally detected the additional loss of MSH6 expression. Whole exome sequencing on genomic DNA from peripheral blood revealed MSH6 c.2731C > T mutation, which was confirmed to be inherited from her mother, who had an early-onset ascending colon cancer without cancer family history. Conclusion: This is a rare case of the Lynch syndrome harboring germline mutations simultaneously in two different MMR genes inherited from two families with Lynch syndrome. The diagnosis of endometrial cancer at the age less than 40 years is uncommon for Lynch syndrome-related endometrial cancer. This suggests an earlier cancer screening for patients carrying two MMR mutations.

Circulating interleukin-6 level is a prognostic marker for survival in advanced nonsmall cell lung cancer patients treated with chemotherapy.

Lung cancer is the leading cause of cancer death worldwide as well as in Taiwan. Interleukin-6 (IL-6) is a multifunctional cytokine and has been implicated in tumor progression. This study recruited 245 patients with advanced (Stage 3B/4) nonsmall cell lung cancer (NSCLC) that had received chemotherapy, to evaluate associations between IL-6 and lung cancer-specific survival. Among these subjects, 112 gave blood samples before and 133 after the start of chemotherapy. Plasma IL-6 was measured using an enzyme linked-immunosorbent assay. The 33rd and 66th percentiles of IL-6 concentrations were 2.01 and 25.16 for the 245 patients and were defined as the cutoff points for dividing the patients into low, intermediate and high groups. Kaplan-Meier and Cox proportional-hazard models were used to evaluate the relationship between the IL-6 level and survival time. Results after adjusting for age, sex, smoking history, histologic type and stage of lung cancer revealed a significant relationship. For all patients, the hazard ratio with high IL-6 levels for lung cancer-specific survival was 2.10 [95% confidence interval (CI) = 1.49 - 2.96] compared with low IL-6 levels. The hazard ratio for patients who were recruited before and after the start of chemotherapy was1.25 (95% CI = 0.73 - 2.13) and 3.66 (95% CI = 2.18 - 6.15), respectively. Patients with high circulating IL-6 also responded poorly to chemotherapy. Therefore, a high level of circulating IL-6 was associated with an inferior response and survival outcome in NSCLC patients treated with chemotherapy.

Clinical Characteristics and Responses to Immune Checkpoint Inhibitors in RET-Aberrant Digestive Tract Tumours.

BACKGROUND: RET plays an oncogenic role, and its aberrations are potentially actionable. However, they have seldom been reported in tumours other than lung or thyroid cancers. The correlation of RET aberrations with clinical characteristics, co-occurring aberrations, and responses to immune checkpoint inhibitors (ICPi) have not been explored in digestive tract tumours. OBJECTIVES: The aim of the study was to elucidate the clinical characteristics, frequently co-altered genes, and treatment responses in RET-aberrant digestive tract tumours. PATIENTS AND METHODS: We retrospectively evaluated patients with digestive tract cancers for RET-aberrant tumours via FoundationOne CDx tumour-based selected genome sequencing from Jan 2016 to Jan 2021. RESULTS: In a median follow-up time of 51 months, a total of 453 patients were analysed. RET-aberrant tumours accounted for 4.4% in the studied population (n = 20), and 1.1% had an oncogenic fusion (n = 5). APC, KRAS, TP53, MSH6 and STK11 were the differentially co-altered genes (all false discovery rates <0.05). The presence of RET aberrations alone was not a significant prognostic factor. Eleven patients with RET-aberrant tumours received ICPi-based treatment and none achieved an objective response. In contrast, 47 patients with non-aberrant tumours received ICPi treatment and had an objective response rate of 27.7% and a significantly longer treatment duration (6.2 vs 2.8 months, p = 0.0008). CONCLUSIONS: Albeit rarely, RET aberrations can be found in digestive tract tumours. Patients with RET-aberrant tumours have a blunted response to ICPi and a comparable prognosis as compared with RET-wild type tumours. Together, these results provide insights into this rare but potentially actionable target in digestive tract tumours.

HER2 amplification in colorectal cancer with brain metastasis: A propensity score matching study.

BACKGROUND: The association between human epidermal growth factor receptor-2 (HER2) amplification and brain metastasis (BM) in patients having colorectal cancer (CRC) has been suggested but not yet established. This study investigated the expression patterns of HER2, its association with BM, and its prognostic value in patients having CRC. METHODS: We retrospectively identified 99 patients having metastatic CRC (mCRC) and BM (the BM cohort) and compared them with a cohort of 249 patients having mCRC and without BM (the stage IV cohort) by propensity score matching. Immunohistochemical studies of HER2 on all available paraffin-embedded tumour samples, either from the primary tumour, the metastasis (brain and/or extracranial sites) or both, were performed and analysed. HER2 fluorescent in situ hybridisation was applied when necessary. The expression of HER2 was compared and correlated with survival. RESULTS: HER2 amplifications were detected in 16 (18.4%) of 87 and 9 (3.6%) of 249 patients who had specimens available in the BM and stage IV cohorts, respectively (P < .001). After propensity score matching, HER2 amplification was significantly associated with BM (odds ratio: 5.38, P = .003). HER2 heterogeneity was frequently observed not only at the single tumour level but also in paired tumour samples. A marginally significant longer survival since BM was found in patients having HER2-amplified mCRC than in those without (P = .07). CONCLUSIONS: HER2 amplification was significantly associated with BM in patients having mCRC and might have prognostic value for survival since BM. Given the heterogeneity of HER2 expression, the testing of HER2 status on available tissues from both primary and metastatic tumours should be encouraged.

Betulinic acid decreases specificity protein 1 (Sp1) level via increasing the sumoylation of sp1 to inhibit lung cancer growth.

Previous studies have shown that the inhibitory effect of betulinic acid (BA) on specificity protein 1 (Sp1) expression is involved in the prevention of cancer progression, but the mechanism of this effect remains to be delineated. In this study, we determined that BA treatment in HeLa cells increased the sumoylation of Sp1 by inhibiting sentrin-specific protease 1 expression. The subsequent recruitment of E3 ubiquitin-protein ligase RING finger protein 4 resulted in ubiquitin-mediated degradation in a 26S-proteosome-dependent pathway. In addition, both BA treatment and mithramycin A (MMA) treatment inhibited lung tumor growth and down-regulated Sp1 protein expression in Kras(G12D)-induced lung cancers of bitransgenic mice. In gene expression profiles of Kras(G12D)-induced lung cancers in bitransgenic mice with and without Sp1 inhibition, 542 genes were affected by MMA treatment. One of the gene products, cyclin A2, which was involved in the S and G(2)/M phase transition during cell cycle progression, was investigated in detail because its expression was regulated by Sp1. The down-regulation of cyclin A2 by BA treatment resulted in decreased retinoblastoma protein phosphorylation and cell cycle G(2)/M arrest. The BA-mediated cellular Sp1 degradation and antitumor effect were also confirmed in a xenograft mouse model by using H1299 cells. The knockdown of Sp1 in lung cancer cells attenuated the tumor-suppressive effect of BA. Taken together, the results of this study clarify the mechanism of BA-mediated Sp1 degradation and identify a pivotal role for Sp1 in the BA-induced repression of lung cancer growth.

Low incidence of thromboembolism in relapsed/refractory myeloma patients treated with thalidomide without thromboprophylaxis in Taiwan.

Thromboembolism (TE) is a common complication in patients with multiple myeloma (MM). Immunomodulatory agents, e.g., thalidomide, have expanded the therapeutic options for treating myeloma; however, Western countries report a high incidence of thrombosis in thalidomide-treated MM patients who lack thromboprophylaxis. A Korean trial reported low TE incidence in thalidomide-treated myeloma patients (39% were given aspirin prophylactically). We aimed to elucidate the TE frequency in MM patients in Taiwan who were treated with thalidomide without TE prophylaxis. We retrospectively collected the records of MM patients who had used thalidomide from a single institute between 2004 and 2010, combined these records with two other Taiwanese studies, and compared all three with the Korean trial. In the current Taiwanese series, five of 144 patients (3.5%) developed TE as follows: three (2.1%) were venous and two (1.3%) were arterial. Only 6.1% of the patients had undergone TE prophylaxis, which is less than in the Korean trial (38.9%, p < 0.05). Of the patients in the relapsed/refractory cohort (n = 114) who were given thalidomide alone, none (0/52) developed venous TE (VTE); however, two patients (2/35, 5.7%) who were given thalidomide-dexamethasone as a salvage treatment developed VTE. In the thrombosis cohort, four patients (80%) were treated with thalidomide plus dexamethasone. In conclusion, the frequency of thalidomide-related TE in myeloma patients without effective TE prophylaxis was low in Taiwan. In relapsed/refractory myeloma patients, the VTE frequency was slightly lower compared with Western patients irrespective of treatment with thalidomide alone or combined with dexamethasone. Even in low TE incidence areas, thalidomide combined with dexamethasone was more thrombogenic compared with others.

Cutting-Edge AI Technologies Meet Precision Medicine to Improve Cancer Care.

To provide precision medicine for better cancer care, researchers must work on clinical patient data, such as electronic medical records, physiological measurements, biochemistry, computerized tomography scans, digital pathology, and the genetic landscape of cancer tissue. To interpret big biodata in cancer genomics, an operational flow based on artificial intelligence (AI) models and medical management platforms with high-performance computing must be set up for precision cancer genomics in clinical practice. To work in the fast-evolving fields of patient care, clinical diagnostics, and therapeutic services, clinicians must understand the fundamentals of the AI tool approach. Therefore, the present article covers the following four themes: (i) computational prediction of pathogenic variants of cancer susceptibility genes; (ii) AI model for mutational analysis; (iii) single-cell genomics and computational biology; (iv) text mining for identifying gene targets in cancer; and (v) the NVIDIA graphics processing units, DRAGEN field programmable gate arrays systems and AI medical cloud platforms in clinical next-generation sequencing laboratories. Based on AI medical platforms and visualization, large amounts of clinical biodata can be rapidly copied and understood using an AI pipeline. The use of innovative AI technologies can deliver more accurate and rapid cancer therapy targets.

The Expression Quantitative Trait Loci in Immune Response Genes Impact the Characteristics and Survival of Colorectal Cancer.

The impact of germline variants on the regulation of the expression of tumor microenvironment (TME)-based immune response genes remains unclear. Expression quantitative trait loci (eQTL) provide insight into the effect of downstream target genes (eGenes) regulated by germline-associated variants (eVariants). Through eQTL analyses, we illustrated the relationships between germline eVariants, TME-based immune response eGenes, and clinical outcomes. In this study, both RNA sequencing data from primary tumor and germline whole-genome sequencing data were collected from patients with stage III colorectal cancer (CRC). Ninety-nine high-risk subjects were subjected to immune response gene expression analyses. Seventy-seven subjects remained for further analysis after quality control, of which twenty-two patients (28.5%) experienced tumor recurrence. We found that 65 eQTL, including 60 germline eVariants and 22 TME-based eGenes, impacted the survival of cancer patients. For the recurrence prediction model, 41 differentially expressed genes (DEGs) achieved the best area under the receiver operating characteristic curve of 0.93. In total, 19 survival-associated eGenes were identified among the DEGs. Most of these genes were related to the regulation of lymphocytes and cytokines. A high expression of HGF, CCR5, IL18, FCER1G, TDO2, IFITM2, and LAPTM5 was significantly associated with a poor prognosis. In addition, the FCER1G eGene was associated with tumor invasion, tumor nodal stage, and tumor site. The eVariants that regulate the TME-based expression of FCER1G, including rs2118867 and rs12124509, were determined to influence survival and chromatin binding preferences. We also demonstrated that FCER1G and co-expressed genes in TME were related to the aggregation of leukocytes via pathway analysis. By analyzing the eQTL from the cancer genome using germline variants and TME-based RNA sequencing, we identified the eQTL in immune response genes that impact colorectal cancer characteristics and survival.