Pan-Cancer Proteomics Analysis Reveals Wiskott-Aldrich Syndrome Protein as a Potential Regulator of Programmed Death-Ligand 1.

The programmed death-ligand 1 (PD-L1) is a key mediator of immunosuppression in the tumor microenvironment. The expression of PD-L1 in cancer cells is useful for the clinical determination of an immune checkpoint blockade (ICB). However, the regulatory mechanism of the PD-L1 abundance remains incompletely understood. Here, we integrated the proteomics of 52 patients with solid tumors and examined immune cell infiltration to reveal PD-L1-related regulatory modules. Wiskott-Aldrich syndrome protein (WASP) was identified as a potential regulator of PD-L1 transcription. In two independent cohorts containing 164 cancer patients, WASP expression was significantly associated with PD-L1. High WASP expression contributed to immunosuppressive cell composition, including cells positive for immune checkpoints (PD1, CTLA4, TIGIT, and TIM3), FoxP3+ Treg cells, and CD163+ tumor-associated macrophages. Overexpression of WASP increased, whereas knockdown of WASP decreased the protein level of PD-L1 in cancer cells without alteration of PD-L1 protein stability. The WASP-mediated cell migration and invasion were markedly attenuated by the silence of PD-L1. Collectively, our data suggest that WASP is a potential regulator of PD-L1 and the WASP/PD-L1 axis is responsible for cell migration and an immunosuppressive microenvironment.

Immune cell patterns before and after neoadjuvant immune checkpoint blockade combined with chemoradiotherapy in locally advanced esophageal squamous cell carcinoma.

BACKGROUND: Neoadjuvant immune checkpoint blockade (ICB) combined with chemoradiotherapy offers high pathologic complete response (pCR) rate for patients with locally advanced esophageal squamous cell carcinomas (ESCC). But the dynamic tumor immune microenvironment modulated by such neoadjuvant therapy remains unclear. PATIENTS AND METHODS: A total of 41 patients with locally advanced ESCC were recruited. All patients received neoadjuvant toripalimab combined with concurrent chemoradiotherapy. Matched pre- and post-treatment tissues were obtained for fluorescent multiplex immunohistochemistry (mIHC) and IHC analyses. The densities and spatial distributions of immune cells were determined by HALO modules. The differences of immune cell patterns before and after neoadjuvant treatment were investigated. RESULTS: In the pre-treatment tissues, more stromal CD3 + FoxP3 + Tregs and CD86+/CD163 + macrophages were observed in patients with residual tumor existed in the resected lymph nodes (pN1), compared with patients with pCR. The majority of macrophages were distributed in close proximity to tumor nest in pN1 patients. In the post-treatment tissues, pCR patients had less CD86 + cell infiltration, whereas higher CD86 + cell density was significantly associated with higher tumor regression grades (TRG) in non-pCR patients. When comparing the paired pre- and post-treatment samples, heterogeneous therapy-associated immune cell patterns were found. Upon to the treatment, CD3 + T lymphocytes were slightly increased in pCR patients, but markedly decreased in non-pCR patients. In contrast, a noticeable increase and a less obvious decrease of CD86 + cell infiltration were respectively depicted in non-pCR and pCR patients. Furthermore, opposite trends of the treatment-induced alterations of CD8 + and CD15 + cell infiltrations were observed between pN0 and pN1 patients. CONCLUSIONS: Collectively, our data demonstrate a comprehensive picture of tumor immune landscape before and after neoadjuvant ICB combined with chemoradiotherapy in ESCC. The infiltration of CD86 + macrophage may serve as an unfavorable indicator for neoadjuvant toripalimab combined with chemoradiotherapy.

Quantification of eosinophilic area and its potential molecular feature in clear cell renal cell carcinoma.

OBJECTIVE: Previous studies have acknowledged the presence of eosinophilic cytoplasm in clear cell renal cell carcinoma, yet the precise quantification method and potential molecular attributes in clear cell renal cell carcinoma remain elusive. This study endeavours to precisely quantify the eosinophilic attribute and probe into the molecular mechanisms governing its presence in clear cell renal cell carcinoma. METHODS: Data from cohorts of clear cell renal cell carcinoma patients who underwent nephrectomy, comprising The Cancer Genome Atlas cohort (n = 475) and Sun Yat-sen University Cancer Center cohort (n = 480), were aggregated to assess the eosinophilic attribute. Additionally, Omics data from Clinical Proteomic Tumor Analysis Consortium (CPTAC) (n = 58) were leveraged to explore the potential molecular features associated with eosinophilic clear cell renal cell carcinoma. Employing receiver operating characteristic curve analysis, the proportion of tumour cells with eosinophilic cytoplasm was determined, leading to the classification of each cohort into distinct groups: a clear group (<5%) and an eosinophilic group (≥5%). RESULTS: In both cohorts, the eosinophilic feature consistently correlated with higher International Society of Urological Pathology (ISUP) grade, elevated tumor stage, and the presence of necrosis. Furthermore, the Kaplan-Meier method demonstrated that patients in the eosinophilic group exhibited shorter overall survival or disease-free survival compared with those in the clear group, a pattern reaffirmed in various stratified survival analyses. Intriguingly, within The Cancer Genome Atlas cohort, the pathological characterization of cell cytoplasm (eosinophilic vs. clear) emerged as an independent risk factor for overall survival (hazard ratio = 2.507 [95% confidence interval: 1.328-4.733], P = 0.005) or disease-free survival (hazard ratio = 1.730 [95% confidence interval: 1.062-2.818], P = 0.028) via Cox regression analysis. Moreover, multi-Omics data unveiled frequent BAP1 mutations and down-regulation of Erythroblast Transformation-Specific-Related Gene associated with the eosinophilic feature in clear cell renal cell carcinoma. Additionally, patients with low expression of Erythroblast Transformation-Specific-Related Gene showed worse overall survival (P < 0.001). CONCLUSIONS: The quantification of the eosinophilic feature serves as a robust predictor of clinical prognosis in clear cell renal cell carcinoma. Furthermore, the manifestation of this feature may be linked to BAP1 mutations and the down-regulation of Erythroblast Transformation-Specific-Related Gene in clear cell renal cell carcinoma. Significantly, the expression levels of Erythroblast Transformation-Specific-Related Gene manifest as an exemplary prognostic marker, providing exceptional predictive accuracy for the clinical prognosis in clear cell renal cell carcinoma.

EFHD1, a novel mitochondrial regulator of tumor metastasis in clear cell renal cell carcinoma.

The biological function of many mitochondrial proteins in mechanistic detail has not been well investigated in clear cell renal cell carcinoma (ccRCC). A seven-mitochondrial-gene signature was generated by Lasso regression analysis to improve the prediction of prognosis of patients with ccRCC, using The Cancer Genome Atlas and Clinical Proteomic Tumor Analysis Consortium cohort. Among those seven genes, EFHD1 is less studied and its role in the progression of ccRCC remains unknown. The decreased expression of EFHD1 was validated in clinical samples and was correlated with unfavorable outcome. Overexpression of EFHD1 in ccRCC cells resulted in the reduction of mitochondrial Ca2+ , and the inhibition of cell migration and invasion in vitro and tumor metastasis in vivo. Mechanistically, EFHD1 physically bound to the core mitochondrial calcium transporter (mitochondrial calcium uniporter, MCU) through its N-terminal domain. The interaction between EFHD1 and MCU suppressed the uptake of Ca2+ into mitochondria, and deactivated the Hippo/YAP signaling pathway. Further data revealed that the ectopic expression of EFHD1 upregulated STARD13 to enhance the phosphorylation of YAP protein at Ser-127. The knockdown of STARD13 or the overexpression of MCU partly abrogated the EFHD1-mediated induction of phosphorylation of YAP at Ser-127 and suppression of cell migration. Taken together, the newly identified EFHD1-MCU-STARD13 axis participates in the modulation of the Hippo/YAP pathway and serves as a novel regulator in the progression of ccRCC.

Regular gastroscopy and colonoscopy during the evaluation of urachal cancer: do we really need them?

PURPOSE: Urachal cancer is similar to gastrointestinal adenocarcinoma in histology, and gastroscopy/colonoscopy is often administered during perioperative evaluation. However, gastroscopy and colonoscopy have corresponding disadvantages. This study discusses whether gastroscopy/colonoscopy is truly necessary for patients with urachal cancer. PATIENTS AND METHODS: A total of 166 bladder adenocarcinoma cases diagnosed at Sun Yat-sen University Cancer Center were retrospectively reviewed and divided into two groups (urachal cancer and nonurachal cancer), and perioperative evaluations were retrieved. RESULTS: There were 78 patients with urachal cancer, the median age was 48 years, and 59 were male. Perioperative gastroscopy/colonoscopy revealed 5 intestinal polyps and 1 adenoma during these evaluations, and no primary gastrointestinal cancer was found. Meanwhile, preoperative imaging evaluation did not detect significant gastrointestinal lesions. For 88 patients with nonurachal cancer, including primary bladder adenocarcinoma and metastatic tumors from gastrointestinal cancer, the median age was 56 years, and 64 were male. Preoperative imaging evaluation demonstrated 36 cases of gastrointestinal lesions, and 32 were confirmed by gastroscopy/colonoscopy; the other 4 were negative. Another 4 cases of colon cancer were detected by regular colonoscopy for suspected primary bladder adenocarcinoma. In all, 35 cases of colon cancer and 1 case of gastric cancer were identified by endoscopic examination. The diagnostic consistency of imaging and gastrointestinal endoscopy was favorable (P < 0.001), and the negative predictive value and diagnostic efficiency of imaging were 96.9% and 94.6%, respectively. CONCLUSIONS: The vast majority of gastrointestinal cancer cases can be identified by assessment of the patient's clinical symptoms, meticulous physical examination, and imaging evaluation. We recommend that gastroscopy/colonoscopy only be applied to patients with urachal cancer when the above examinations are positive.

Bladder paraganglioma: basic characteristics and new perspectives on perioperative management.

PURPOSE: Paraganglioma and pheochromocytoma are rare neuroendocrine tumors with severe metabolic and cardiovascular complications. Bladder PGLs are rare, and their clinical management is not precise. Here, we discuss the basic characteristics and perioperative management of bladder PGLs. METHODS: We retrospectively reviewed 20 bladder PGL cases diagnosed at Sun Yat-sen University Cancer Center. Case notes were reviewed, clinical presentations, therapies, and outcomes were collected, and data analysis was performed. RESULTS: Ten male and ten female patients with a median age of 47.5 years (range 14-69 years) were included. Most patients (65%) had no symptoms, and PGL was detected incidentally during medical checkups. All patients were treated surgically; 4 (20%) underwent transurethral resection of bladder tumor (TURBT), and 16 (80%) underwent partial cystectomy. Strong intraoperative blood pressure fluctuations were observed in 13 patients (65%). Two patients who were treated preoperatively with α-receptor blockers also experienced severe intraoperative blood pressure fluctuations. Postoperative measurements of troponin I were available for 3 patients, and all were significantly elevated. All patients were diagnosed with bladder PGL on postoperative pathological examination. The median follow-up time was 51 months (range 2-147 months), and 2 patients were lost to follow-up at 1 and 3 months; 16 (88.9%) survived without recurrence, 2 patients (11.1%) experienced recurrence, and 1 patient died. CONCLUSION: Most bladder paragangliomas are easily mistaken for bladder urothelial carcinoma, and robust hemodynamic instability during surgery might be a challenge for urologists. Postoperative monitoring of troponin I, regardless of the presence of clinical symptoms, is recommended for patients with bladder PGL.

Impact of immunohistochemistry-based molecular subtype on predicting chemotherapy response and survival in patients with T1 stage bladder cancer after bladder-preserving treatment.

OBJECTIVE: To explore the immunohistochemistry-based molecular subtypes of bladder cancer, and their impact on the prognosis and the chemotherapy response between gemcitabine plus cisplatin intra-arterial chemotherapy and epirubicin-inducted intravesical chemotherapy, in patients with T1 stage bladder cancer after bladder-preserving treatment. METHODS: One hundred and seventy-six patients with T1 stage bladder cancer were selected for this study. Thirty-three patients underwent radical cystectomy, 43 received gemcitabine plus cisplatin intra-arterial chemotherapy and 100 received intravesical chemotherapy. The markers labeled with luminal (GATA3, Uroplakin II, CK20) and basal (CK5/6, CK14, CD44) phenotypes were chosen as candidate markers. RESULTS: One hundred and seventy-six patients were divided into 76 patients as basal/squamous (BASQ), 45 as the luminal A and 55 as the luminal B. Compared with the luminal B and BASQ tumors, the luminal A tumors showed a trend for better recurrence-free survival (P = 0.105) and progression-free survival (P = 0.093). The combination of CK20 and GATA3 was practical to identify the molecular phenotypes with total 84.9% accuracy and significantly associated with recurrence-free survival (P = 0.025) and progression-free survival (P = 0.004). The patient with BASQ tumors who received intravesical chemotherapy showed a trend for worse progression-free survival than the patient who received gemcitabine plus cisplatin intra-arterial chemotherapy or radical cystectomy. Furthermore, the patients with BASQ tumors experienced a significant improvement in progression-free survival after gemcitabine plus cisplatin intra-arterial chemotherapy compared with the patients who received intravesical chemotherapy (P = 0.011). CONCLUSIONS: The immunohistochemistry-based molecular subtypes could predict the patient's prognosis and clinically different chemotherapeutic survival outcomes in patients with T1 stage bladder cancer after bladder-preserving treatment.

Perivesical Fat Invasive Pattern as Prognostic Factor and Predictor of Response to Adjuvant Chemotherapy in T3 Stage Bladder Cancer.

BACKGROUND: The prognostic value of the distinction between microscopic (pT3a) and macroscopic (pT3b) perivesical fat invasions remains a subject of debate. To explore whether the pattern of perivesical fat invasion can serve as a prognostic factor to better subgroup T3 stage bladder cancer. MATERIALS AND METHODS: One hundred and forty-nine patients diagnosed with T3 stage bladder cancer at Sun Yat-sen University Cancer Center (SYSUCC) were selected for the experimental cohort in this study. Ninety-seven T3 stage bladder cancer patients with pathological slices at the Cancer Genome Atlas (TCGA) were selected as validation cohort in this study. The perivesical fat invasive pattern was examined with hematoxylin and eosin-stained pathological slides by two pathologists independently. Two different perivesical fat invasive patterns, fibrous-surrounded (FS) pattern, and nonfibrous-surrounded (NFS) pattern were assessed. RESULTS: Perivesical fat invasion pattern had a significant influence on overall survival in T3 stage bladder cancer. Compared to the NFS pattern, the FS pattern was related to a better prognosis in both the SYSUCC cohort and TCGA cohort. The patients with NFS pattern tumor who underwent cisplatin-based adjuvant chemotherapy experienced an obvious improvement compared to observation after radical cystectomy in overall survival in the SYSUCC cohort. CONCLUSION: The perivesical fat invasion pattern could predict prognosis and clinically different chemotherapeutic survival outcomes in patients with T3 stage bladder cancer after radical cystectomy.

Optimal interval timing between transurethral resection of bladder tumors and Bacillus Calmette-Guerin perfusion.

OBJECTIVE: There is no consensus regarding the best interval time between transurethral resection of a bladder tumor and Bacillus Calmette-Guerin (BCG) perfusion. This study was to explore whether the interval time has an impact on the prognosis and adverse effects. METHODS: We retrospectively reviewed the clinical data of patients who received BCG intravesical perfusion at Sun Yat-sen University Cancer Center (SYSUCC) from September 2015 to October 2021. Recurrence-free survival (RFS) and progression-free survival were the primary endpoints. Cox regression was used to explore independent predictors. The association between interval time and adverse effect grade was detected by logistic regression. Propensity score matching (PSM) was performed. RESULTS: A total of 403 patients were enrolled, the median interval time was 24 days (6-163 days), and the follow-up was 28 months (7-82 months). Eighty-eight (20.9%) patients relapsed, and 40 patients (10.0%) suffered progression. The multivariate Cox regression analysis confirmed that interval time was an independent predictor of RFS (p = 0.017). Notably, when the interval time was less than or equal to 26 days, there was a trend toward better RFS, PSM resulted in 65 matched pairs in each group, and Kaplan-Meier analysis showed that there was a significant difference in RFS between groups (p = 0.009). The logistic regression analysis showed that there was no correlation between interval time and adverse effects and their grades (p > 0.05). CONCLUSIONS: We considered that the first BCG perfusion could be performed within 2-4 weeks after surgery.

Comparison of RNAscope and immunohistochemistry for evaluation of the UPK2 status in urothelial carcinoma tissues.

BACKGROUND: UPK2 exhibits excellent specificity for urothelial carcinoma (UC). UPK2 evaluation can be useful in making the correct diagnosis of UC. However, UPK2 detection by immunohistochemistry (IHC) has relatively low sensitivity. This paper aimed to compare the diagnostic sensitivity of RNAscope and IHC for evaluation of the UPK2 status in UC. METHODS: Tissue blocks from 127 conventional bladder UCs, 45 variant bladder UCs, 24 upper tract UCs and 23 metastatic UCs were selected for this study. IHC and RNAscope were used to detect the UPK2 status in UCs. Then, comparisons of the two methods were undertaken. RESULTS: There was no significant difference between RNAscope and IHC for the evaluation of the UPK2 positivity rate in UC (68.0% vs. 62.6%, P = 0.141). Correlation analysis revealed a moderate positive correlation for detection of UPK2: RNAscope vs. IHC (P < 0.001, R = 0.441). Our results showed a trend toward a higher positive UPK2 rate detected by RNAscope (53.3%) than by IHC (35.6%) in variant bladder UCs. Disappointingly, the P value did not indicate a significant difference (P = 0.057). CONCLUSIONS: RNAscope for UPK2 appeared to perform similarly to IHC, with a marginally higher positive rate, suggesting it could be used as an alternative or adjunct to UPK2 IHC.

Ginsenoside Rg1 improves cognitive capability and affects the microbiota of large intestine of tree shrew model for Alzheimer's disease.

Ginsenoside Rg1 (Rg1) is traditional Chinese medicine with neuroprotective activity. Previous studies have demonstrated that Rg1 improves Alzheimer's disease (AD) and alters gut microbiology, but its mechanism remains to be elucidated, and thus far, its use in the treatment of AD has not been satisfactory. The present study investigated the improvement effects of Rg1 and its association with the microbiota of the large intestine. Following treatment with Rg1 in AD tree shrews, the treatment group demonstrated significantly shorter escape latency and crossed a platform more frequently in a water maze test. Western blotting demonstrated that Rg1 inhibited the expression of ß-secretase 1, while increasing microtubule-associated protein 2 and Fox-3 in the hippocampus. Immunohistochemical analysis revealed that Rg1 decreased the expression of amyloid ß, tau phosphorylated at serine 404 and pro-apoptotic factor Bax, while increasing the expression of Bcl-2 in the hippocampus and cortex. High throughput sequencing of 16S rRNA demonstrated that Rg1 altered the microbiota abundance of the large intestine. In conclusion, Rg1 affected the expression of apoptosis proteins, possessed a neuroprotective effect and may have a close association with the microbiota of large intestine by significantly reducing the abundance of Bacteroidetes and increasing the energy requirement of tree shrews.

Improving Alzheimer's disease by altering gut microbiota in tree shrews with ginsenoside Rg1.

Ginsenoside Rg1 (GRg1) has neuroprotective effects on Alzheimer's disease (AD). The occurrence and progression of AD are closely related to gut microbiota. Few studies have learned the direct relationship between GRg1 and gut microbiota. In this study, we found an original way to research this relationship by using GRg1 in the AD model of tree shrews. Morris water maze and immunohistochemistry were performed to test the cognition repairing function of GRg1 by tree shrews and 16S ribosomal RNA sequencing was used to explore the composition and abundance of gut microbiota. After GRg1 treatment, the result of Morris water maze showed an improvement in cognitive function, and immunohistochemistry revealed a decrease in tau protein. Moreover, 16SrRNA sequencing results showed the abundances of Proteobacteria and Verrucomicrobia were significantly different, and Lactobacillaceae was significantly increased in the GRg1 treatment group. It also showed that the gut microbiome with middle and high doses of GRg1 was close to the normal group. In conclusion, this study suggests that GRg1 at middle and high doses may change the abundance of gut microbiota to improve AD, and thatProteobacteria and Verrucomicrobia are key microbiota. This is the first report that has ever studied the relationship between GRg1 and gut microbiota in tree shrews.

Immunophenotypes Based on the Tumor Immune Microenvironment Allow for Unsupervised Penile Cancer Patient Stratification.

The tumor immune microenvironment (TIME) plays an important role in penile squamous cell carcinoma (peSCC) pathogenesis. Here, the immunophenotype of the TIME in peSCC was determined by integrating the expression patterns of immune checkpoints (programmed cell death-1 (PD-1)/programmed cell death ligand-1 (PD-L1), cytotoxic T lymphocyte antigen 4 (CTLA-4), and Siglec-15) and the components of tumor-infiltrating lymphocytes, including CD8+ or Granzyme B+ T cells, FOXP3+ regulatory T cells, and CD68+ or CD206+ macrophages, in 178 patients. A high density of Granzyme B, FOXP3, CD68, CD206, PD-1, and CTLA-4 was associated with better disease-specific survival (DSS). The patients with diffuse PD-L1 tumor cell expression had worse prognoses than those with marginal or negative PD-L1 expression. Four immunophenotypes were identified by unsupervised clustering analysis, based on certain immune markers, which were associated with DSS and lymph node metastasis (LNM) in peSCC. There was no significant relationship between the immunophenotypes and high-risk human papillomavirus (hrHPV) infection. However, the hrHPV-positive peSCC exhibited a higher density of stromal Granzyme B and intratumoral PD-1 than the hrHPV-negative tumors (p = 0.049 and 0.002, respectively). In conclusion, the immunophenotypes of peSCC were of great value in predicting LNM and prognosis, and may provide support for clinical stratification management and immunotherapy intervention.

Prevalence of human papillomavirus and implication on survival in Chinese penile cancer.

We assessed the prevalence of HPV DNA in a large series of Chinese penile cancer and examine its association with the histological subtype, p16INK4a expression, and prognosis. We pathologically categorized 226 invasive penile squamous cell carcinomas and assessed HPV genotyping by real-time PCR and p16INK4a immunohistochemistry. The results were correlated with histopathological and clinical parameters and disease-specific survival (DSS). HPV DNA was detected in 32.7% (74/226) of penile cancer cases. The most frequent genotype was HPV 16 (64/74, 86.5%), followed by HPV 18 (6/74, 8.1%). Fifty-nine (26.1%) cases were positive for the p16INK4a expression, and p16INK4a expression had a sensitivity of 56.8% (95% CI, 45.2-68.3%) and a specificity of 88.8% (95% CI, 83.8-93.9%) for defining HPV status. HPV DNA (P = 0.019), p16INK4a (P = 0.038), age (P = 0.018), grade of differentiation (P = 0.001), lymph nodes (P < 0.001), T stage (P < 0.001), M stage (P < 0.001), and lymphovascular invasion (LVI, P = 0.001) were prognostic factors for DSS. HPV-positivity (HR 0.334; 95% CI, 0.158-0.705, P = 0.004) was still a significant prognostic factor for DSS in the multivariate Cox regression model. HPV DNA was observed in one third of Chinese penile carcinoma cases. The p16INK4a expression can indicate high-risk human papillomavirus (HR-HPV). HPV-positive penile tumors confer a survival benefit over HPV-negative tumors.

Hydrothermal synthesis of graphitic carbon nitride-Bi2WO6 heterojunctions with enhanced visible light photocatalytic activities.

Graphitic carbon nitride (C3N4) was hybridized by Bi2WO6 via a hydrothermal method. The high-resolution transmission electron microscopy (HR-TEM) results reveal that an intimate interface between C3N4 and Bi2WO6 forms in the heterojunctions. The UV-vis diffuse reflection spectra show that the resulting C3N4-Bi2WO6 heterojunctions possess more intensive absorption within the visible light range in comparison with pure Bi2WO6. These excellent structural and spectral properties endowed the C3N4-Bi2WO6 heterojunctions with enhanced photocatalytic activities. Significantly, the optimum photocatalytic activity of the 0.5C3N4-0.5Bi2WO6 heterojunction for the degradation of methyl orange (MO) was almost 3 and 155 times higher than those of either individual C3N4 or Bi2WO6. The possible photocatalytic mechanism with superoxide radical species as the main active species in photocatalysis is proposed on the basis of experimental results. Moreover, the heterojunction depicted high stability and durability during six successive cycles.