Perioperative toripalimab and chemotherapy in locally advanced gastric or gastro-esophageal junction cancer: a randomized phase 2 trial.

Perioperative chemotherapy is the standard treatment for locally advanced gastric or gastro-esophageal junction cancer, and the addition of programmed cell death 1 (PD-1) inhibitor is under investigation. In this randomized, open-label, phase 2 study (NEOSUMMIT-01), patients with resectable gastric or gastro-esophageal junction cancer clinically staged as cT3-4aN + M0 were randomized (1:1) to receive either three preoperative and five postoperative 3-week cycles of SOX/XELOX (chemotherapy group, n = 54) or PD-1 inhibitor toripalimab plus SOX/XELOX, followed by toripalimab monotherapy for up to 6 months (toripalimab plus chemotherapy group, n = 54). The primary endpoint was pathological complete response or near-complete response rate (tumor regression grade (TRG) 0/1). The results showed that patients in the toripalimab plus chemotherapy group achieved a higher proportion of TRG 0/1 than those in the chemotherapy group (44.4% (24 of 54, 95% confidence interval (CI): 30.9%-58.6%) versus 20.4% (11 of 54, 95% CI: 10.6%-33.5%)), and the risk difference of TRG 0/1 between toripalimab plus chemotherapy group and chemotherapy group was 22.7% (95% CI: 5.8%-39.6%; P = 0.009), meeting a prespecified endpoint. In addition, a higher pathological complete response rate (ypT0N0) was observed in the toripalimab plus chemotherapy group (22.2% (12 of 54, 95% CI: 12.0%-35.6%) versus 7.4% (4 of 54, 95% CI: 2.1%-17.9%); P = 0.030), and surgical morbidity (11.8% in the toripalimab plus chemotherapy group versus 13.5% in the chemotherapy group) and mortality (1.9% versus 0%), and treatment-related grade 3-4 adverse events (35.2% versus 29.6%) were comparable between the treatment groups. In conclusion, the addition of toripalimab to chemotherapy significantly increased the proportion of patients achieving TRG 0/1 compared to chemotherapy alone and showed a manageable safety profile. ClinicalTrials.gov registration: NCT04250948 .

Deep learning-derived spatial organization features on histology images predicts prognosis in colorectal liver metastasis patients after hepatectomy.

Histopathological images of colorectal liver metastases (CRLM) contain rich morphometric information that may predict patients' outcomes. However, to our knowledge, no study has reported any practical deep learning framework based on the histology images of CRLM, and their direct association with prognosis remains largely unknown. In this study, we developed a deep learning-based framework for fully automated tissue classification and quantification of clinically relevant spatial organization features (SOFs) in H&E-stained images of CRLM. The SOFs based risk-scoring system demonstrated a strong and robust prognostic value that is independent of the current clinical risk score (CRS) system in independent clinical cohorts. Our framework enables fully automated tissue classification of H&E images of CRLM, which could significantly reduce assessment subjectivity and the workload of pathologists. The risk-scoring system provides a time- and cost-efficient tool to assist clinical decision-making for patients with CRLM, which could potentially be implemented in clinical practice.

Radiomics-based survival risk stratification of glioblastoma is associated with different genome alteration.

BACKGROUND: Glioblastoma (GBM) is a remarkable heterogeneous tumor with few non-invasive, repeatable, and cost-effective prognostic biomarkers reported. In this study, we aim to explore the association between radiomic features and prognosis and genomic alterations in GBM. METHODS: A total of 180 GBM patients (training cohort: n = 119; validation cohort 1: n = 37; validation cohort 2: n = 24) were enrolled and underwent preoperative MRI scans. From the multiparametric (T1, T1-Gd, T2, and T2-FLAIR) MR images, the radscore was developed to predict overall survival (OS) in a multistep postprocessing workflow and validated in two external validation cohorts. The prognostic accuracy of the radscore was assessed with concordance index (C-index) and Brier scores. Furthermore, we used hierarchical clustering and enrichment analysis to explore the association between image features and genomic alterations. RESULTS: The MRI-based radscore was significantly correlated with OS in the training cohort (C-index: 0.70), validation cohort 1 (C-index: 0.66), and validation cohort 2 (C-index: 0.74). Multivariate analysis revealed that the radscore was an independent prognostic factor. Cluster analysis and enrichment analysis revealed that two distinct phenotypic clusters involved in distinct biological processes and pathways, including the VEGFA-VEGFR2 signaling pathway (q-value = 0.033), JAK-STAT signaling pathway (q-value = 0.049), and regulation of MAPK cascade (q-value = 0.0015/0.025). CONCLUSIONS: Radiomic features and radiomics-derived radscores provided important phenotypic and prognostic information with great potential for risk stratification in GBM.

Neoadjuvant PD-1 blockade with sintilimab in mismatch-repair deficient, locally advanced rectal cancer: an open-label, single-centre phase 2 study.

BACKGROUND: The current standard treatment for locally advanced rectal cancer is neoadjuvant chemoradiotherapy followed by radical surgery, but this approach can lead to multiple complications. We aimed to investigate the clinical activity and safety of neoadjuvant therapy with sintilimab, a single-agent PD-1 antibody, in patients with mismatch-repair deficient locally advanced rectal cancer. METHODS: This open-label, single-arm, phase 2 study was done at the Sun Yat-sen University Cancer Center, Guangzhou, China. Patients aged 18-75 years with mismatch-repair deficient or microsatellite instability-high locally advanced rectal cancer were enrolled and received neoadjuvant sintilimab monotherapy (200 mg by intravenous infusion) every 21 days. After an initial four cycles of treatment, patients and clinicians could choose one of the following options: total mesorectal excision surgery, followed by four cycles of adjuvant sintilimab with or without CapeOX chemotherapy (capecitabine 1000 mg/m2, orally administered twice daily on days 1-14; oxaliplatin 130 mg/m2, intravenously administered on day 1 every 3 weeks), determined by clinicians; or another four cycles of sintilimab followed by radical surgery or observation (only for patients with a clinical complete response; also known as the watch and wait strategy). The primary endpoint was the complete response rate, which included both a pathological complete response after surgery and a clinical complete response after completion of sintilimab treatment. Clinical response was evaluated by digital rectal examination, MRI, and endoscopy. Response was assessed in all patients who received treatment at least until the first tumour response assessment, after the first two cycles of sintilimab. Safety was analysed in all patients who received at least one dose of treatment. This trial is closed to enrolment and is registered with ClinicalTrials.gov (NCT04304209). FINDINGS: Between Oct 19, 2019, and June 18, 2022, 17 patients were enrolled and received at least one dose of sintilimab. The median age was 50 years (IQR 35-59) and 11 (65%) of 17 patients were male. One patient was excluded from efficacy analyses because they were lost to follow-up after the first sintilimab cycle. Of the remaining 16 patients, six underwent surgery, of whom three had a pathological complete response. Nine other patients had a clinical complete response and chose the watch and wait strategy. One patient had a serious adverse event and discontinued treatment; this patient did not have a complete clinical response and refused to undergo surgery. A complete response was thus noted for 12 (75%; 95% CI 47-92) of 16 patients. One of the three patients who underwent surgery but did not have a pathological complete response showed an increase in tumour volume after the initial four cycles of sintilimab (at which point they underwent surgery); this patient was deemed to have primary resistance to immune checkpoint inhibitors. After a median follow-up of 17·2 (IQR 8·2-28·5) months, all patients were alive and none had disease recurrence. Only one (6%) patient had a grade 3-4 adverse event, which was deemed a serious adverse event (grade 3 encephalitis). INTERPRETATION: The preliminary results of this study suggest that anti-PD-1 monotherapy is effective and tolerable for patients with mismatch-repair deficient locally advanced rectal cancer and could potentially spare some patients from radical surgery. Longer treatment courses might be needed to achieve maximum effects in some patients. Longer follow-up is also needed to observe the duration of response. FUNDING: The National Natural Science Foundation of China, CAMS Innovation Fund for Medical Sciences, Science and Technology Program of Guangzhou, and Innovent Biologics.

Whole exome sequencing reveals the genetic heterogeneity and evolutionary history of primary gliomas and matched recurrences.

Diffuse glioma is a highly heterogeneous central nervous system tumor that is refractory to conventional therapy. Residual glioma cells escape from surgery and chemoradiotherapy, leading to lethal recurrence. Understanding the molecular mechanism of this recurrence process is critical to the development of successful therapies. Here, we analyzed whole-exome sequencing (WES) data of 97 paired primary and recurrent samples from 46 patients with glioma via a uniform pipeline. Clonality and phylogenetic analyses revealed that branching evolution was widespread in the recurrent process of gliomas. Recurrent tumors continued to evolve independently with chemoradiotherapy and harbored multiple recurrence-selected genetic alterations, such as amplification of PPFIBP1, PDE4DIP, and KRAS, deletion of TNFRSF14, DCC, CDKN2A, and MSH6, and mutations in ATRX, ARID1A, KEL, TP53, MSH6, and KMT2B. Meanwhile, truncal variants within partial driver genes were identified among primary and recurrent gliomas, suggesting that they might be ideal therapeutic targets. Intriguingly, the immunogenicity of recurrent gliomas did not increase significantly compared to the primary tumors. Genomic analysis of recurrent gliomas provided an opportunity to identify potentially clinically informative alterations not detected in clinically sampled primary tumors.

Driver mutations in ADGRL3 are involved in the evolution of ependymoma.

Although there have been recent advances in the molecular pathology of ependymomas, little is known about the underlying molecular evolution during its development. Here, we assessed the clinical, pathological and molecular evolutionary process of ependymoma recurrence in a 9-year-old patient who had seven recurrences of supratentorial ependymoma and died from intracranial multiregional recurrences at the age of 19 years old. Whole-genome sequencing (WGS) of 7 tumor samples (1 primary and 6 subsequent recurrent tumors) was performed to elucidate the mutation landscape and identify potential driver mutations for tumor evolution. The genetic profiles of the seven tumor specimens showed significant heterogeneity and suggested a highly branched evolutionary pattern. The mutational signatures and chromothripsis changed with treatments. Strikingly, adhesion G protein-coupled receptor L3 (ADGRL3, also known as Latrophilins 3, LPNH3) was found to be consistently mutated during the entire disease process. However, Sanger sequencing of other 78 ependymoma patients who underwent surgery at our institution showed no genetic alteration of ADGRL3, as found in the present case. The mRNA levels of ADGRL3 were significantly lower in ependymomas (n = 36), as compared with normal brain tissue (n = 3). Grade III ependymomas had the lowest ADGRL3 expression. Moreover, ependymomas with lower mRNA level of ADGRL3 had shorter overall survival. Our findings, therefore, demonstrate a rare evolutionary process of ependymoma involving ADGRL3.

Growth differentiation factor 1-induced tumour plasticity provides a therapeutic window for immunotherapy in hepatocellular carcinoma.

Tumour lineage plasticity is an emerging hallmark of aggressive tumours. Tumour cells usually hijack developmental signalling pathways to gain cellular plasticity and evade therapeutic targeting. In the present study, the secreted protein growth and differentiation factor 1 (GDF1) is found to be closely associated with poor tumour differentiation. Overexpression of GDF1 suppresses cell proliferation but strongly enhances tumour dissemination and metastasis. Ectopic expression of GDF1 can induce the dedifferentiation of hepatocellular carcinoma (HCC) cells into their ancestral lineages and reactivate a broad panel of cancer testis antigens (CTAs), which further stimulate the immunogenicity of HCC cells to immune-based therapies. Mechanistic studies reveal that GDF1 functions through the Activin receptor-like kinase 7 (ALK7)-Mothers against decapentaplegic homolog 2/3 (SMAD2/3) signalling cascade and suppresses the epigenetic regulator Lysine specific demethylase 1 (LSD1) to boost CTA expression. GDF1-induced tumour lineage plasticity might be an Achilles heel for HCC immunotherapy. Inhibition of LSD1 based on GDF1 biomarker prescreening might widen the therapeutic window for immune checkpoint inhibitors in the clinic.

The Prognostic Value of Locoregional Interventions for BRAF V600E Metastatic Colorectal Cancer: A Retrospective Cohort Analysis.

The prognostic heterogeneity in patients with BRAF V600E metastatic colorectal cancer (mCRC) remains poorly defined. Real-world data of 93 BRAF V600E mCRC patients from Sun Yat-sen University Cancer Center were evaluated using the prognostic factors affecting overall survival (OS). Treatment of metastases served as an independent prognosticator, where curative locoregional interventions (LRIs) were associated with superior clinical outcomes (adjusted hazard ratio (HR): 0.46, 95% confidence interval (CI): 0.22-0.98; p = 0.044). The LRIs group showed an improved median OS of 49.4 months versus 18.3 months for the palliative treatments (PTs) group. The median OS of patients with colorectal liver metastasis (CRLM) was significantly prolonged after undergoing LRIs (42.4 vs. 23.7 months; HR: 0.11, 95% CI: 0.01-1.22; p = 0.030), and patients in the LRIs plus liver-limited or lung-limited metastasis (LLM) group benefited more than those in the LRIs plus non-LLM group when compared to the PTs group (LLM from LRIs vs. PTs, HR: 0.16, 95% CI: 0.04-0.68; p = 0.006. Non-LLM from LRIs vs. PTs, HR: 0.47, 95% CI: 0.21-1.05; p = 0.074). In conclusion, we confirmed the positive prognostic value of LRIs in BRAF V600E mCRC, particularly in patients with CRLM or LLM.

Mutational landscape of primary pulmonary salivary gland-type tumors through targeted next-generation sequencing.

OBJECTIVES: Primary pulmonary salivary gland-type tumors (PSGTs) mainly comprise of mucoepidermoid carcinoma (MEC) and adenoid cystic carcinoma (ACC), which are rare and molecularly poorly understood. This study aimed to profile the molecular alterations of PSGTs via targeted next-generation sequencing (NGS). MATERIAL AND METHODS: Immunohistochemistry was used to screen PSGTs in 32 patients and MAML2 and MYB rearrangements were detected using fluorescence in situ hybridization. 1021-Genepanel of targeted NGS was conducted to profile genomic mutations in all the PSGT patients. RESULTS: Among the 32 patients, 25 had MEC and 7 had ACC. MAML2 and MYB rearrangements were detected in 80.0% (20/25) of the MEC and 71.4% (5/7) of the ACC patients. Among the MEC patients, 10 (40.0%) had ≥1 mutation, and 6 of them had 11 isolated mutations with abundance >5%, namely NFE2L2, MYOD1, INPP4B, CCND2, SNTG1, HSPD1, TGFBR1, RBM10, NOTCH4, ASXL1, and PTPRD mutations. The remaining 4 patients had 9 mutations with abundance <5%, namely KMT2A, PDCD11, FLT1, BRCA2, APC, SLX4, FOXP1, FGFR1, and HRAS mutations. All the ACC patients had mutations, which were enriched in 5 pathways including the PI3K and NOTCH pathways, chromatin and cytoskeleton remodeling, and DNA damage. These results explain PSGTs harbor distinct driver features of MAML2 or MYB rearrangement, accompanied with wide mutational diversity with very low rate of somatic mutation. Several important pathways, including the NOTCH and PI3K pathways, and chromatin remodeling could be targeted to improve the survival in patients with ACC.

Morphologic, Immunohistochemical, and Genetic Differences Between High-grade and Low-grade Fetal Adenocarcinomas of the Lung.

Fetal adenocarcinoma of the lung (FLAC) is a rare lung tumor classified into low-grade fetal adenocarcinoma of the lung (LG-FLAC) and high-grade fetal adenocarcinoma of the lung (HG-FLAC). It remains debatable whether HG-FLAC is a subset of FLAC or a distinct subtype of the conventional lung adenocarcinoma (CLA). In this study, samples of 4 LG-FLAC and 2 HG-FLAC cases were examined, and the clinicopathologic, immunohistochemical (IHC), and mutational differences between the 2 subtypes were analyzed using literature review. Morphologically, LG-FLACs had a pure pattern with complex glandular architecture composed of cells with subnuclear and supranuclear vacuoles, mimicking a developing fetal lung. In contrast, HG-FLACs contained both fetal lung-like (FLL) and CLA components. With regard to IHC markers, ß-catenin exhibited a nuclear/cytoplasmic staining pattern in LG-FLACs but a membranous staining pattern in HG-FLACs. Furthermore, p53 was expressed diffusely and strongly in HG-FLACs, whereas in LG-FLACs, p53 staining was completely absent. Using next-generation sequencing targeting a 1021-gene panel, mutations of CTNNB1 and DICER1 were detected in all 4 LG-FLAC samples, and a novel mutation, MYCN P44L, was discovered in 2 LG-FLAC samples. DNA samples of the FLL and CLA components of HG-FLACs were separately extracted and sequenced. The FLL component harbored no CTNNB1, DICER1, or MYCN mutations; moreover, the FLL genetic profile largely overlapped with that of the CLA component. The morphologic, IHC, and genetic features of HG-FLAC indicate that it is a variant of CLA rather than a subset of FLAC. Thus, HG-FLAC should be treated differently from LG-FLAC.

Targeting tumor lineage plasticity in hepatocellular carcinoma using an anti-CLDN6 antibody-drug conjugate.

Tumor lineage plasticity is emerging as a critical mechanism of therapeutic resistance and tumor relapse. Highly plastic tumor cells can undergo phenotypic switching to a drug-tolerant state to avoid drug toxicity. Here, we investigate the transmembrane tight junction protein Claudin6 (CLDN6) as a therapeutic target related to lineage plasticity for hepatocellular carcinoma (HCC). CLDN6 was highly expressed in embryonic stem cells but markedly decreased in normal tissues. Reactivation of CLDN6 was frequently observed in HCC tumor tissues as well as in premalignant lesions. Functional assays indicated that CLDN6 is not only a tumor-associated antigen but also conferred strong oncogenic effects in HCC. Overexpression of CLDN6 induced phenotypic shift of HCC cells from hepatic lineage to biliary lineage, which was more refractory to sorafenib treatment. The enhanced tumor lineage plasticity and cellular identity change were potentially induced by the CLDN6/TJP2 (tight junction protein 2)/YAP1 (Yes-associated protein 1) interacting axis and further activation of the Hippo signaling pathway. A de novo anti-CLDN6 monoclonal antibody conjugated with cytotoxic agent (Mertansine) DM1 (CLDN6-DM1) was developed. Preclinical data on both HCC cell lines and primary tumors showed the potent antitumor efficiency of CLDN6-DM1 as a single agent or in combination with sorafenib in HCC treatment.

A Genotype Signature for Predicting Pathologic Complete Response in Locally Advanced Rectal Cancer.

PURPOSE: To construct and validate a predicting genotype signature for pathologic complete response (pCR) in locally advanced rectal cancer (PGS-LARC) after neoadjuvant chemoradiation. METHODS AND MATERIALS: Whole exome sequencing was performed in 15 LARC tissues. Mutation sites were selected according to the whole exome sequencing data and literature. Target sequencing was performed in a training cohort (n = 202) to build the PGS-LARC model using regression analysis, and internal (n = 76) and external validation cohorts (n = 69) were used for validating the results. Predictive performance of the PGS-LARC model was compared with clinical factors and between subgroups. The PGS-LARC model comprised 15 genes. RESULTS: The area under the curve (AUC) of the PGS model in the training, internal, and external validation cohorts was 0.776 (0.697-0.849), 0.760 (0.644-0.867), and 0.812 (0.690-0.915), respectively, and demonstrated higher AUC, accuracy, sensitivity, and specificity than cT stage, cN stage, carcinoembryonic antigen level, and CA19-9 level for pCR prediction. The predictive performance of the model was superior to clinical factors in all subgroups. For patients with clinical complete response (cCR), the positive prediction value was 94.7%. CONCLUSIONS: The PGS-LARC is a reliable predictive tool for pCR in patients with LARC and might be helpful to enable nonoperative management strategy in those patients who refuse surgery. It has the potential to guide treatment decisions for patients with different probability of tumor regression after neoadjuvant therapy, especially when combining cCR criteria and PGS-LARC.

PARP inhibitor Olaparib overcomes Sorafenib resistance through reshaping the pluripotent transcriptome in hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) is one of the most common human malignancies worldwide with very poor prognosis. Resistance to targeted therapeutic drugs such as sorafenib remains one of the major challenges in clinical treatment. In the present study, PARP1 was found to be highly expressed in human embryonic stem cells, but progressively decreased upon specified hepatic differentiation. Reactivation of PARP1 expression was also detected in HCC residual tumors after sorafenib treatment in xenograft mouse model, indicating the potential important roles of PARP1 in stem cell pluripotency and HCC sorafenib treatment resistance. Overexpression of PARP1 was frequently observed in HCC patients, and closely associated with poor clinical outcome. Treatment of Sorafenib induced activation of DNA damage repair signaling, which is highly active and essential for maintenance of stem cell pluripotency in HCC residual tumors. PARP inhibitor Olaparib extensively suppressed the DNA damage repair signaling, and significantly inhibited the global pluripotent transcriptional network. The repression of key pluripotent transcriptional factors and DNA damage repair signaling by Olaparib was mainly through CHD1L-mediated condensation of the chromatin structure at their promotor regions. The global reshaping of the pluripotent transcriptome by Olaparib might reinforce Sorafenib in eliminating HCC residual tumors and enhance therapeutic efficiency.

Histopathological growth patterns correlate with the immunoscore in colorectal cancer liver metastasis patients after hepatectomy.

Various scoring systems have been proposed to predict the postoperative prognosis of colorectal liver metastasis (CRLM), including the clinical risk score (CRS), the immunoscore and so on. Recently, histopathological growth patterns (HGPs) have been recognized. However, the correlation between HGPs and the immunoscore, and their prognostic values in patients with CRLM after liver resection remain undetermined. In this study, HGPs were retrospectively evaluated in H&E-stained slides from 166 CRLM patients. The immunoscore was calculated according to the densities of immunostained CD3 + and CD8 + cells. A risk score combining HGPs, the immunoscore and the CRS was defined and divided patients into the low-, medium- and high-risk group. Our results showed that the densities of CD3 + and CD8 + cells were higher in the desmoplastic HGP (dHGP) group than in the non-dHGP group, and the proportion of high immunoscores was also higher in the dHGP group (51.9% vs. 33.0%, respectively, P = 0.020). Patients with the dHGP had significantly longer relapse-free survival (RFS) and overall survival (OS) than those with the non-HGP. The low-risk group showed significantly higher 2-year RFS and 5-year OS rates than the other two groups (RFS: 76.2%, 43.7% and 33.1%, respectively; P < 0.001; OS: 89.7%, 54.4% and 33.3%, respectively; P < 0.001). In conclusion, the dHGP correlates with relatively high immunoscores, predicting a favorable prognosis independent of the immunoscore and CRS. A novel risk score combining HGPs, the immunoscore and the CRS may be used for the stratification of CRLM patients' survival.

L1CAM High Expression Associates with Poor Prognosis in Glioma but Does Not Correlate with C11orf95-RELA Fusion.

The latest WHO guideline of CNS tumor defined a RELA fusion-positive ependymoma type with extremely poor prognosis, and the expression of L1CAM was correlated well with the presence of RELA fusion. However, the L1CAM protein expression in large sample gliomas other than ependymoma, its relationship with the RELA gene and its prognostic significance remained unknown. We examined the expression of L1CAM in 565 glioma cases (WHO grade I-IV). The L1CAM IHC-positive cases were selected to test RELA fusion with FISH break-apart probes. L1CAM was positive in 109 cases (19.29%) of all 565 glioma cases, with 18.27% in low-grade gliomas and 19.84% in high-grade gliomas, respectively. Unlike ependymoma, L1CAM protein expression was not correlated with the C11orf95-RELA fusion gene in other gliomas, but it had correction with the patient age (older than 45-year-old, p = 0.006), ATRX mutation (p = 0.003) and Ki67 (p = 0.007). High expression of L1CAM was an independent prognostic factor in our cohort. Further analysis demonstrated that L1CAM strong positive expression was significantly associated with poor prognosis in gliomas, both in our cohort (p < 0.001) and TCGA (p < 0.009) dataset. Although uncorrelated with C11orf95-RELA fusion, L1CAM was a significant poor prognostic marker in glioma patients. More aggressive treatment should be taken for these patients and L1CAM might be a promising therapeutic target in glioma.

Association between glioblastoma cell-derived vessels and poor prognosis of the patients.

BACKGROUND: Vessels with different microcirculation patterns are required for glioblastoma (GBM) growth. However, details of the microcirculation patterns in GBM remain unclear. Here, we examined the microcirculation patterns of GBM and analyzed their roles in patient prognosis together with two well-known GMB prognosis factors (O6 -methylguanine DNA methyltransferase [MGMT] promoter methylation status and isocitrate dehydrogenase [IDH] mutations). METHODS: Eighty GBM clinical specimens were collected from patients diagnosed between January 2000 and December 2012. The microcirculation patterns, including endothelium-dependent vessels (EDVs), extracellular matrix-dependent vessels (ECMDVs), GBM cell-derived vessels (GDVs), and mosaic vessels (MVs), were evaluated by immunohistochemistry (IHC) and immunofluorescence (IF) staining in both GBM clinical specimens and xenograft tissues. Vascular density assessments and three-dimensional reconstruction were performed. MGMT promoter methylation status was determined by methylation-specific PCR, and IDH1/2 mutations were detected by Sanger sequencing. The relationship between the microcirculation patterns and patient prognosis was analyzed by Kaplan-Meier method. RESULTS: All 4 microcirculation patterns were observed in both GBM clinical specimens and xenograft tissues. EDVs were detected in all tissue samples, while the other three patterns were observed in a small number of tissue samples (ECMDVs in 27.5%, GDVs in 43.8%, and MVs in 52.5% tissue samples). GDV-positive patients had a median survival of 9.56 months versus 13.60 months for GDV-negative patients (P = 0.015). In MGMT promoter-methylated cohort, GDV-positive patients had a median survival of 6.76 months versus 14.23 months for GDV-negative patients (P = 0.022). CONCLUSION: GDVs might be a negative predictor for the survival of GBM patients, even in those with MGMT promoter methylation.

Skp2 modulates proliferation, senescence and tumorigenesis of glioma.

BACKGROUND: Gliomas represent the largest class of primary central nervous system neoplasms, many subtypes of which exhibit poor prognoses. Surgery followed by radiotherapy and chemotherapy has been used as a standard strategy but yielded unsatisfactory improvements in patient survival outcomes. The S-phase kinase protein 2 (Skp2), a critical component of the E3-ligase SCF complex, has been documented in tumorigenesis in various cancer types but its role in glioma has yet to be fully clarified. In this study, we investigated the function of Skp2 in the proliferation, stem cell maintenance, and drug sensitivity to temozolomide (TMZ) of glioma. METHODS: To investigate the role of Skp2 in the prognosis of patients with glioma, we first analyzed data in databases TCGA and GTEx. To further clarify the effect of Skp2 on glioma cell proliferation, we suppressed its level in glioblastoma (GBM) cell lines through knockdown and small molecule inhibitors (lovastatin and SZL-P1-41). We then detected cell growth, colony formation, sphere formation, drug sensitivity, and in vivo tumor formation in xenograft mice model. RESULTS: Skp2 mRNA level was higher in both low-grade glioma and GBM than normal brain tissues. The knockdown of Skp2 increased cell sensitivity to TMZ, decreased cell proliferation and tumorigenesis. In addition, Skp2 level was found increased upon stem cells enriching, while the knockdown of Skp2 led to reduced sphere numbers. Downregulation of Skp2 also induced senescence. Repurposing of lovastatin and novel compound SZL-P1-41 suppressed Skp2 effectively, and enhanced glioma cell sensitivity to TMZ in vitro and in vivo. CONCLUSION: Our data demonstrated that Skp2 modulated glioma cell proliferation in vitro and in vivo, stem cell maintenance, and cell sensitivity to TMZ, which indicated that Skp2 could be a potential target for long-term treatment.

Practice of the New Integrated Molecular Diagnostics in Gliomas: Experiences and New Findings in a Single Chinese Center.

Background: The latest WHO classification of CNS tumors using the integrated phenotypic and molecular parameters (IDH, ATRX, 1p19q, TERT etc.) have reestablished the CNS tumors classification in addition to traditional histology. The establishment of glioma molecular typing can more accurately predict prognosis, better guide individualized treatment to improve survival. Methods: The expression of IDH1, ATRX, PHH3, P53 and Ki67 was detected by IHC. Molecular status of IDH1/2 and TERT were analyzed using Sanger sequencing. MGMT was explored using methylation-specific PCR. 1p/19q codeletion status was firstly detected by FISH, then further confirmed by multiplex PCR-based next generation sequencing. Results: The mutation frequency of IDH1 was 68.7% (79/115) in WHO II astrocytoma, and 82 cases (82/344, 23.8%) were "triple-negative glioma" in our cohort. Multivariate COX analysis revealed that only IDH, 1p/19q, TERT and MGMT were independent prognostic factors. Noteworthily, we found 7 cases of the new molecular phenotype presented as "IDH wildtype and 1p/19q codeletion", not mentioned in the latest WHO guideline. Conclusion: We detected the newly recommended markers in a large cohort of Chinese glioma patients. Our data demonstrated a relatively lower frequency of IDH mutations and a higher prevalence of triple-negative glioma in Chinese compared with American and European, indicating ethnic and geographical difference in some markers. In addition, the new molecular phenotype "IDH wildtype and 1p/19q codeletion" glioma deserved special focus. These findings suggest that further stratification of infiltrating gliomas is needed for different treatment strategy and precision medicine.

Tenascin-cmediated vasculogenic mimicry formation via regulation of MMP2/MMP9 in glioma.

Vasculogenic mimicry (VM), the formation of vessel-like structures by highly invasive tumor cells, has been considered one of several mechanisms responsible for the failure of anti-angiogenesis therapy in glioma patients. Therefore, inhibiting VM formation might be an effective therapeutic method to antagonize the angiogenesis resistance. This study aimed to show that an extracellular protein called Tenascin-c (TNC) is involved in VM formation and that TNC knockdown inhibits VM in glioma. TNC was upregulated with an increase in glioma grade. TNC and VM formation are potential independent predictors of survival of glioma patients. TNC upregulation was correlated with VM formation, and exogenous TNC stimulated VM formation. Furthermore, TNC knockdown significantly suppressed VM formation and proliferation in glioma cells in vitro and in vivo, with a reduction in cellular invasiveness and migration. Mechanistically, TNC knockdown decreased Akt phosphorylation at Ser473 and Thr308 and subsequently downregulated matrix metalloproteinase 2 and 9, both of which are important proteins associated with VM formation and migration. Our results indicate that TNC plays an important role in VM formation in glioma, suggesting that TNC is a potential therapeutic target for anti-angiogenesis therapy for glioma.