Deciphering the prognostic features of bladder cancer through gemcitabine resistance and immune-related gene analysis and identifying potential small molecular drug PIK-75.

BACKGROUND: Bladder cancer (BCa) stands out as a prevalent and highly lethal malignancy worldwide. Chemoresistance significantly contributes to cancer recurrence and progression. Traditional Tumor Node Metastasis (TNM) stage and molecular subtypes often fail to promptly identify treatment preferences based on sensitivity. METHODS: In this study, we developed a prognostic signature for BCa with uni-Cox + LASSO + multi-Cox survival analysis in multiple independent cohorts. Six machine learning algorithms were adopted to screen out the hub gene, RAC3. IHC staining was used to validate the expression of RAC3 in BCa tumor tissue. RT-qPCR and Western blot were performed to detect and quantify the mRNA and protein levels of RAC3. CCK8, colony formation, wound healing, and flow cytometry analysis of apoptosis were employed to determine cell proliferation, migration, and apoptosis. Molecular docking was used to find small target drugs, PIK-75. 3D cell viability assay was applied to evaluate the ATP viability of bladder cancer organoids before and after PIK-75 treated. RESULTS: The established clinical prognostic model, GIRS, comprises 13 genes associated with gemcitabine resistance and immunology. This model has demonstrated robust predictive capabilities for survival outcomes across various independent public cohorts. Additionally, the GIRS signature shows significant correlations with responses to both immunotherapy and chemotherapy. Leveraging machine learning algorithms, the hub gene, RAC3, was identified, and potential upstream transcription factors were screened through database analysis. IHC results showed that RAC3 was higher expressed in GEM-resistant BCa patients. Employing molecular docking, the small molecule drug PIK-75, as binding to RAC3, was identified. Experiments on cell lines, organoids and animals validated the biological effects of PIK-75 in bladder cancer. CONCLUSIONS: The GIRS signature offers a valuable complement to the conventional anatomic TNM staging system and molecular subtype stratification in bladder cancer. The hub gene, RAC3, plays a crucial role in BCa and is significantly associated with resistance to gemcitabine. The small molecular drug, PIK-75 having the potential as a therapeutic agent in the context of gemcitabine-resistant and immune-related pathways.

Genetically engineered membrane-based nanoengagers for immunotherapy of pancreatic cancer.

Modulating macrophages presents a promising avenue in tumor immunotherapy. However, tumor cells have evolved mechanisms to evade macrophage activation and phagocytosis. Herein, we introduced a bispecific antibody-based nanoengager to facilitate the recognition and phagocytosis of tumor cells by macrophages. Specifically, we genetically engineered two single chain variable fragments (scFv) onto cell membrane: anti-CD40 scFv for engaging with macrophages and anti-Claudin18.2 (CLDN18.2) scFv for interacting with tumor cells. These nanoengagers were further constructed by coating scFv-anchored membrane into PLGA nanoparticle core. Our developed nanoengagers significantly boosted immune responses, including increased recognition and phagocytosis of tumor cells by macrophages, enhanced activation and antigen presentation, and elevated cytotoxic T lymphocyte activity. These combined benefits resulted in enhancing antitumor efficacy against highly aggressive "cold" pancreatic cancer. Overall, this study offers a versatile nanoengager design for immunotherapy, achieved through genetically engineering to incorporate antibody-anchored membrane.

PTEN Mediates the Silencing of Unintegrated HIV-1 DNA.

The integration of viral DNA into a host genome is an important step in HIV-1 replication. However, due to the high failure rate of integration, the majority of viral DNA exists in an unintegrated state during HIV-1 infection. In contrast to the robust expression from integrated viral DNA, unintegrated HIV-1 DNA is very poorly transcribed in infected cells, but the molecular machinery responsible for the silencing of unintegrated HIV-1 DNA remains poorly characterized. In this study, we sought to characterize new host factors for the inhibition of expression from unintegrated HIV-1 DNA. A genome-wide CRISPR-Cas9 knockout screening revealed the essential role of phosphatase and tensin homolog (PTEN) in the silencing of unintegrated HIV-1 DNA. PTEN's phosphatase activity negatively regulates the PI3K-Akt pathway to inhibit the transcription from unintegrated HIV-1 DNA. The knockout (KO) of PTEN or inhibition of PTEN's phosphatase activity by point mutagenesis activates Akt by phosphorylation and enhances the transcription from unintegrated HIV-1 DNA. Inhibition of the PI3K-Akt pathway by Akt inhibitor in PTEN-KO cells restores the silencing of unintegrated HIV-1 DNA. Transcriptional factors (NF-κB, Sp1, and AP-1) are important for the activation of unintegrated HIV-1 DNA in PTEN-KO cells. Finally, the knockout of PTEN increases the levels of active epigenetic marks (H3ac and H3K4me3) and the recruitment of PolII on unintegrated HIV-1 DNA chromatin. Our experiments reveal that PTEN targets transcription factors (NF-κB, Sp1, and AP-1) by negatively regulating the PI3K-Akt pathway to promote the silencing of unintegrated HIV-1 DNA.

A novel missense mutation in the MECOM gene in a Chinese boy with radioulnar synostosis with amegakaryocytic thrombocytopenia.

Radioulnar synostosis with amegakaryocytic thrombocytopenia (RUSAT) type 2, caused by MDS1 and EVI1 complex locus (MECOM) gene mutations, is a rare inherited bone marrow failure syndrome (IBMFS) with skeletal anomalies, characterized by varying presentation of congenital thrombocytopenia (progressing to pancytopenia), bilateral proximal radioulnar synostosis, and other skeletal abnormalities. Due to limited knowledge and heterogenous manifestations, clinical diagnosis of the disease is challenging. Here we reported a novel MECOM mutation in a Chinese boy with typical clinical features for RUSAT-2. Trio-based whole exome sequencing of buccal swab revealed a novel heterozygous missense mutation in exon 11 of the MECOM gene (chr3:168818673; NM_001105078.3:c.2285G > A). The results strongly suggest that the variant was a germline mutation and disease-causing mutation. The patient received matched unrelated donor hematopoetic stem cell transplantation (HSCT). This finding was not only expanded the pathogenic mutation spectrum of MECOM gene, but also provided key information for clinical diagnosis and treatment of RUSAT-2.

Resveratrol alleviates fumonisin-induced intestinal cytotoxicity by modulating apoptosis, tight junction, and inflammation in IPEC-J2 porcine intestinal epithelial cells.

Fumonisins are common contaminants in the global food and environment, pose a variety of health risks to humans and animals. However, the method of mitigating fumonisin toxicity is still unclear. Resveratrol is a natural compound with antioxidant and anti-inflammatory properties. In this study, the protective effect of resveratrol against fumonisin-induced intestinal toxicity was investigated by the porcine intestinal epithelial cell line (IPEC-J2). The cells were treated with 0-40 µM fumonisin for 24 or 48 h with or without the 24 h resveratrol (15 µM) pretreatment. The data showed that resveratrol could alleviate the fumonisin B1 (FB1)-induced decrease in cell viability and amplify in membrane permeability. At the same time, it could reduce the accumulation of intracellular reactive oxygen species and increase the expression ranges of Nrf2 and downstream genes (SOD1 and NQO-1), thereby counteracting FB1-induced apoptosis. Furthermore, resveratrol was able to reduce the expression levels of inflammatory factors (TNF-α, IL-1ß, and IL-6), increase the expression levels of tight junction proteins (Claudin-1, Occludin, and ZO-1), and the integrity of the IPEC-J2 monolayer. Our data also showed that resveratrol could attenuate the toxicity of the co-occurrence of three fumonisins. It is implied that resveratrol represents a promising protective approach for fumonisin, even other mycotoxins in the future. This provided a new strategy for further blocking and controlling the toxicity of fumonisin, subsequently avoiding adverse effects on the human and animal health.

Current status and prospect of immunotherapy for colorectal cancer.

PURPOSE: Colorectal cancer is the most common gastrointestinal tumor in China. While significant progress has been achieved in traditional chemotherapy, radiotherapy, and targeted therapy, the prognosis of advanced colorectal cancer is poor, and the five-year survival rate is unsatisfactory. There is an urgent need to explore new treatment modalities. In this review, we examined the latest progress of colorectal cancer immunotherapy and discussed its future prospects. METHODS: We conducted a literature review to sort out the current status of immunotherapy for different types of colorectal cancer and discussed potential combination therapy options. Results Subsequent line therapy, first-line therapy and neoadjuvant therapy for MSI-H/dMMR colorectal cancer are discussed. In addition, combination therapy options for patients with MSS/pMMR colorectal cancer are presented. Finally, current valuable biomarkers for immunotherapy are highlighted. RESULTS: Subsequent line therapy, first-line therapy and neoadjuvant therapy for MSI-H/dMMR colorectal cancer are discussed. In addition, combination therapy options for patients with MSS/pMMR colorectal cancer are presented. Finally, current valuable biomarkers for immunotherapy are highlighted. CONCLUSION: This review discussed the current status of immunotherapy for different types of colorectal cancer and biomarkers for immunotherapy.

lncRNA ENST00000422059 promotes cell proliferation and inhibits cell apoptosis in breast cancer by regulating the miR-145-5p/KLF5 axis.

Krüppel-like zinc-finger transcription factor 5 (KLF5) is a vital regulator of breast cancer (BC) onset and progression. The mechanism by which KLF5 regulates BC is still not clearly known. In this study, bioinformatics analysis shows that BC-affected individuals with elevated KLF5 expression levels have poor clinical outcomes. We further verify that miR-145-5p regulated KLF5 expression to promote cell apoptosis and inhibit cell proliferation in BC via dual-luciferase reporter assay, western blot analysis, qRT-PCR, CCK-8 assay and cell apoptosis assay. In addition, based on bioinformatics analysis, the binding of ENST00000422059 with miR-145-5p is confirmed by dual-luciferase reporter assay. Subsequently, FISH, western blot analysis, qRT-PCR, CCK-8 and cell apoptosis assays verified that ENST00000422059 increases KLF5 protein expression by sponging miRNA to promote cell proliferation and inhibit cell apoptosis. Finally, ENST00000422059 is found to accelerate tumor progression by regulating the miR-145-5p/KLF5 axis in vivo. In conclusion, this study suggests that ENST00000422059 upregulates KLF5 by sponging miR-145-5p to promote BC progression.

Autologous Airway Basal Cell Transplantation Alleviates Airway Epithelium Defect in Recurrent Benign Tracheal Stenosis.

BACKGROUND: Airway epithelium defects are a hallmark of recurrent benign tracheal stenosis (RBTS). Reconstructing an intact airway epithelium is of great importance in airway homeostasis and epithelial wound healing and has great potential for treating tracheal stenosis. METHODS: An experimental study was conducted in canines to explore the therapeutic effect of autologous basal cell transplantation in restoring airway homeostasis. First, airway mucosae from human patients with recurrent tracheal stenosis were analyzed by single-cell RNA sequencing. Canines were then randomly divided into tracheal stenosis, Stent, Stent + Cells, and Stent + Cells + Biogel groups. Autologous airway basal cells of canines in the Stent + Cells and Stent + Cells + Biogel groups were transplanted onto the stenotic airway after modeling. A biogel was coated on the airway prior to basal cell transplantation in the Stent + Cells + Biogel group. After bronchoscopic treatments, canines were followed up for 16 weeks. RESULTS: Single-cell RNA sequencing demonstrated packed airway basal cells and an absence of normal airway epithelial cells in patients with RBTS. Autologous airway basal cell transplantation, together with biogel coating, was successfully performed in the canine model. Follow-up observation indicated that survival time in the Stent + Cells + Biogel group was significantly prolonged, with a higher (100%) survival rate compared with the other groups. In terms of pathological and bronchoscopic findings, canines that received autologous basal cell transplantation showed a reduction in granulation hyperplasia as well as airway re-epithelialization with functionally mature epithelial cells. CONCLUSIONS: Autologous airway basal cell transplantation might serve as a novel regenerative therapy for airway re-epithelialization and inhibit recurrent granulation hyperplasia in benign tracheal stenosis.

Effect of the tyrosine kinase inhibitors on the growth in children with Philadelphia chromosome-positive acute lymphoblastic leukemia: a case-control study.

Background: First-generation ABL-targeted tyrosine kinase inhibitor (TKI) imatinib is known to retard growth in children but it is not known if the second-generation ABL-targeted TKI dasatinib has the same effect. We aimed to determine the impact of the first- or second-generation TKI on the growth of children treated for Philadelphia chromosome-positive (Ph+) childhood acute lymphoblastic leukemia (ALL). Methods: We evaluated the longitudinal growth changes in 140 children with Ph+ ALL treated with imatinib or dasatinib in additional to intensive cytotoxic chemotherapy and 280 matched controls treated with the same intensity of cytotoxic chemotherapy without TKI on Chinese Children's Cancer Group ALL-2015 protocol between 2015 and 2019. We retrospectively reviewed the height data obtained during routine clinic visits at 4 time points: at diagnosis, the end of therapy, 1 year and 2 years off therapy. Height z Scores were derived with the aid of WHO Anthro version 3.2.2 and WHO AnthroPlus version 1.0.4, global growth monitoring tool. Findings: This study consisted only patients who have completed all treatment in continuous complete remission without major events, including 33 patients randomized to receive imatinib, 43 randomized to receive dasatinib, and 64 assigned to receive dasatinib. Similar degree of loss of height z scores from diagnosis to the end of therapy was observed for the 33 imatinib- and the 107 dasatinib-treated patients (median △ = -0.84 vs. -0.88, P = 0.41). Adjusting for height z score at diagnosis, puberty status, and sex, there was no significant difference in the longitudinal mean height z scores between patients treated with imatinib and those with dasatinib (0.08, 95% CI, -0.22 to 0.38, P = 0.60). The degree of loss of height z scores from diagnosis to end of therapy was significantly greater in the 140 TKI-treated patients than the 280 controls (median △ = -0.88 vs. -0.18, P < 0.001). The longitudinal mean height z scores in the TKI-treated patients were significantly lower than those of the controls (-0.84, 95% CI, -0.98 to -0.69; P < 0.001). Interpretation: These data suggest that dasatinib and imatinib have the similar adverse impact on the growth of children with Ph+ ALL. Funding: This study was supported by the National Natural Science Foundation of China (grant 81670136 [JCai and JT]), the fourth round of Three-Year Public Health Action Plan (2015-2017; GWIV-25 [SS]), Shanghai Health Commission Clinical Research Project (202140161 [JCai]), the US National Cancer institute (CA21765 [C-H Pui]), and the American Lebanese Syrian Associated Charities (CC, JJY, and C-HP). The content of this paper is solely the responsibility of the authors and does not necessarily represent the official views of the US National Institutes of Health.

Simultaneous thulium laser resection of the prostate and transperineal prostate biopsy in clinically diagnosed metastatic prostate cancer with bladder outlet obstruction.

Metastatic prostate cancer (mPCa) patients complicated with bladder outlet obstruction (BOO) are often referred to a urologist. Androgen deprivation therapy (ADT) combined with indwelling catheter usually be the initial management. To retrospectively analysis the safety and efficacy of simultaneous thulium laser resection of the prostate (TmLRP) and transperineal prostate biopsy in metastatic prostate cancer with bladder outlet obstruction. From January 2016 to December 2021, 67 clinically diagnosed mPCa with BOO patients were included in this study. All patients were preoperatively assessed with international prostate symptom score (IPSS), QoL, serum prostate-specific antigen (PSA), prostate volume evaluation by transrectal ultrasound, postvoid residual urine volume (PVR), and maximum flow rate (Qmax). Preoperative and perioperative parameters at 1-, 3-, and 6-month follow-up were also evaluated. All complications were recorded. Simultaneous TmLRP and transperineal prostate biopsy had obvious advantages for clinically diagnosed mPCa patients with BOO, including short overall operation time (52 ± 23.3 min), little hemoglobin decrease (0.6 ± 0.7 g/l), and short hospital stay (average 3.8 days). In addition, simultaneous TmLRP and transperineal prostate biopsy also brought them significant improvement on IPSS, QoL score, Qmax, and PVR volume (P < 0.001) at 1-, 3-, and 6-month follow-up after operation compared to preoperative parameters. Complications were in a low incidence. Simultaneous TmLRP and transperineal prostate biopsy is a bloodless operation with immediate effect and little perioperative complication. Importantly, it is a promising technology in the diagnosis and treatment of clinically diagnosed mPCa patients with BOO.

Analysis of the anatomical and biomechanical characteristics of the pelvic floor in cystocele.

INTRODUCTION: Stress urinary incontinence (SUI) occurs due to disruption of the pelvic floor anatomy; however, the complexity of the pelvic floor support structures and individual patient differences make it difficult to identify the weak points in the pelvic floor support that cause SUI to occur, develop, and recur. This study aimed to analyze the pelvic floor anatomy, structural features, and biomechanics of cystoceles to develop more effective treatment plans with individualized and precise healthcare. MATERIAL AND METHODS: In this observational case-controlled study (clinical trial identifier BOJI201855L), 102 women with normal pelvic floor function and 273 patients diagnosed with cystocele degrees I-III were identified at Shanghai General Hospital from October 2016 to December 2019. We combined ultrasound and vaginal tactile imaging (VTI) to assess the anatomy and biomechanical functions of the anterior and posterior vaginal walls. Both examinations included relaxation and muscle tension tests. RESULTS: Of the 42 VTI parameters, 13 were associated with the degree of cystocele, six with an increase in the urethral rotation angle (pointing to the mobility of the urethra), and six with a decrease in the retrovesical angle (pointing to hypsokinesis and decrease in bladder position). According to these data, the strength of tissues, especially the muscles in both the anterior and posterior compartments, contributes to the stability of the pelvic floor structure. The strength of the levator ani muscle (LAM) is important for the degree of cystocele, mobility of the urethra, hypsokinesis, and decrease in bladder position. CONCLUSIONS: In general, the biomechanical status of the pelvic floor in patients with cystocele is complex and involves various muscles, ligaments, tendons, and fascia. Of these, repair and exercise of the LAM have not received much attention in the treatment of patients with cystoceles, which may be an important risk factor for the high recurrence rate.

OX40L-Armed Oncolytic Virus Boosts T-cell Response and Remodels Tumor Microenvironment for Pancreatic Cancer Treatment.

Rationale: The resistance of pancreatic ductal adenocarcinoma (PDAC) to immunotherapies is caused by the immunosuppressive tumor microenvironment (TME) and dense extracellular matrix. Currently, the efficacy of an isolated strategy targeting stromal desmoplasia or immune cells has been met with limited success in the treatment of pancreatic cancer. Oncolytic virus (OV) therapy can remodel the TME and damage tumor cells either by directly killing them or by enhancing the anti-tumor immune response, which holds promise for the treatment of PDAC. This study aimed to investigate the therapeutic effect of OX40L-armed OV on PDAC and to elucidate the underlying mechanisms. Methods: Murine OX40L was inserted into herpes simplex virus-1 (HSV-1) to construct OV-mOX40L. Its expression and function were assessed using reporter cells, cytopathic effect, and immunogenic cell death assays. The efficacy of OV-mOX40L was then evaluated in a KPC syngeneic mouse model. Tumor-infiltrating immune and stromal cells were analyzed using flow cytometry and single-cell RNA sequencing to gain insight into the mechanisms of oncolytic virotherapy. Results: OV-mOX40L treatment delayed tumor growth in KPC tumor-bearing C57BL/6 mice. It also boosted the tumor-infiltrating CD4+ T cell response, mitigated cytotoxic T lymphocyte (CTL) exhaustion, and reduced the number of regulatory T cells. The treatment of OV-mOX40L reprogrammed macrophages and neutrophils to a more pro-inflammatory anti-tumor state. In addition, the number of myofibroblastic cancer-associated fibroblasts (CAF) was reduced after treatment. Based on single-cell sequencing analysis, OV-mOX40L, in combination with anti-IL6 and anti-PD-1, significantly extended the lifespan of PDAC mice. Conclusion: OV-mOX40L converted the immunosuppressive tumor immune microenvironment to a more activated state, remodeled the stromal matrix, and enhanced T cell response. OV-mOX40L significantly prolonged the survival of PDAC mice, either as a monotherapy or in combination with synergistic antibodies. Thus, this study provides a multimodal therapeutic strategy for pancreatic cancer treatment.

Regulation mechanism and pathogenic role of lncRNA plasmacytoma variant translocation 1 (PVT1) in human diseases.

Plasmacytoma variant translocation 1 (PVT1) is a long non-coding RNA (lncRNA) gene identified as a recurrent breakpoint of Burkitt's lymphomas. Human PVT1 gene is located on region 8q24.21, a well-known cancer risk region, and encodes at least 26 linear ncRNA isoforms and 26 circular RNA isoforms, as well as 6 microRNAs. Several PVT1 functioning models have been reported recently such as competing endogenous RNA (ceRNA) activity and regulating protein stability of oncogenes, especially MYC oncogene. The promoter of PVT1 gene is a boundary element of tumor-suppressor DNA. CircPVT1 derived from PVT1 gene is also a critical non-coding oncogenic RNA. Although substantial advancements have been made in understanding the roles of PVT1 in cancer recently, the detailed mechanisms underlying its functions remain unclear. Herein, we summarize the recent progressions on the mechanisms underlying PVT1 regulated gene expression at different levels. We also discuss the interaction between lncRNA and protein, RNA and DNA, as well as the potential cancer therapy strategy by targeting these networks.

Outcomes after HSCT for mucolipidosis II (I-cell disease) caused by novel compound heterozygous GNPTAB mutations.

Background: Mucolipidosis type II (MLII), or I-cell disease, is a rare lysosomal storage disease (LSD) caused by variants in the GNPTAB gene. MLII patients exhibit clinical phenotypes in the prenatal or neonatal stage, such as marked dysmorphic features, cardiac involvement, respiratory symptoms, dysostosis multiplex, severe growth abnormalities, and mental and motor developmental abnormalities. The median age at diagnosis for MLII is 0.7 years, the median survival is 5.0 years, and the median age at death is 1.8 years. No cure for MLII exists. Methods: Sanger sequencing of the GNPTAB gene identified the compound heterozygous mutations c.673C > T in exon 7 and c.1090C > T in exon 9, which were novel double heterozygous mutations first reported in China. For the first time, we describe our experience in the use of HSCT for MLII. Our patient underwent HSCT with cells from a 9/10 human leukocyte antigen (HLA)-matched unrelated donor at 12 months of age. Myeloid neutrophil and platelet engraftment occurred on Days 10 and 11, respectively. Results: The patient's limb muscle tension was significantly reduced, and his gross and fine motor skills were improved four months after transplantation. DST(Developmental Screen Test) results showed that the patient's fine motor skills and mental development were improved compared with before HSCT. Conclusion: MLII is a very severe lysosomal storage disease, to date, only 3 cases have been reported on the use of HSCT to treat MLII. Our data show that HSCT is a potential way to prolong the life of patients and improve their quality of life. Due to the lack of comparable data and time, the exact benefit remains unclear in MLII patients. Longer-term follow-up and in-depth prospective studies are indispensable.

Rapamycin and Low-dose IL-2 Mediate an Immunosuppressive Microenvironment to Inhibit Benign Prostatic Hyperplasia.

Benign prostatic hyperplasia (BPH) is a condition that becomes more common with age and manifests itself primarily as the expansion of the prostate and surrounding tissues. However, to date, the etiology of BPH remains unclear. In this respect, we performed single-cell RNA sequencing of prostate transition zone tissues from elderly individuals with different prostate volumes to reveal their distinct tissue microenvironment. Ultimately, we demonstrated that a reduced Treg/CD4+ T-cell ratio in the large-volume prostate and a relatively activated immune microenvironment were present, characterized partially by increased expression levels of granzymes, which may promote vascular growth and profibrotic processes and further exacerbate BPH progression. Consistently, we observed that the prostate gland of patients taking immunosuppressive drugs usually remained at a smaller volume. Furthermore, in mouse models, we confirmed that both suppression of the immune system with rapamycin and induction of Treg proliferation with low doses of IL-2 therapy indeed prevented the progression of BPH. Taken together, our findings suggest that an activated immune microenvironment is necessary for prostate volume growth and that Tregs can reverse this immune activation state, thereby inhibiting the progression of BPH.

"Sandwich" protocol based on modified SMILE regimen for children with newly extranodal NK/T cell lymphoma, nasal type: a single-arm, single-center clinical study.

Extranodal NK/T-cell lymphoma, nasal type (ENKTL), which is a rare form of mature T/NK cell lymphoma in children, currently lacks a standardized first-line treatment approach. However, a treatment protocol known as the "sandwich" regimen has been used in children newly diagnosed with ENKTL. This protocol combines the administration of methotrexate, ifosfamide, etoposide, pegaspargase, and dexamethasone (referred to as SMILE) with the addition of radiotherapy (RT). From September 2017 to December 2020, a total of five patients were included in the study, consisting of three males and two females. The median age of onset was 10.6 years (range, 9.8 to 14.0 years). Among the patients, four had nasal/nasopharyngeal disease at stage II, while one patient had extra nasal disease involving the skin at stage IV. The median EBV-DNA level in plasma was 1.68 × 103 copies/ml (range, 0.44 to 21.1 × 103copies/ml). All the patients had good overall response after 2 cycles of chemotherapy and radiotherapy, including 4 of the patients who had a complete response and 1 of the patients with partial remission. The patient with stage IV received allogeneic hematopoietic stem cell transplantation after the EBV-DNA level was elevated again during treatment. One patient in the low-risk group experienced grade 4 oral mucositis, while no other severe complications or treatment-related deaths were observed. The median follow-up period was 22 months (range, 5 to 57 months). All five patients successfully completed their treatment, with four patients achieving event-free survival, and one patient was lost to follow-up. The median OS time and EFS time was 33 months (range: 18-57 months) and 20 months (range: 5-47 months), respectively. The sandwich protocol has demonstrated a high response rate, good tolerance to chemotherapy, and no treatment-related fatalities. However, further confirmation is necessary through additional clinical studies involving larger sample sizes. Clinical trial registration number: Due to modified SMILE regimens with sandwiched radiotherapy yielded promising outcomes in children ENKTL, we have carried out a phase II multicenter clinical trial (ChiCTR220005954) for children ENKTL in China to further verify the efficacy and safety.

Di-n-butyl phthalate induces oversecretion of vascular endothelium-derived NAP-2 and promotes epithelial-mesenchymal transition of urothelial cells in newborn hypospadias rats.

Di-n-butyl phthalate (DBP) is a plasticizer commonly used in industrial production and is present in our daily life. It has been confirmed that DBP causes genitourinary malformations, especially hypospadias. However, the research of hypospadias mainly focusses on the genital tubercle in previous studies. In this study, we found DBP could affect the exocrine function of the vascular endothelium which disturb the development of genital nodules and induced hypospadias. We used cytokine array to find that vascular endothelium-derived NAP-2 may be a major abnormal secreted cytokine with biological functions. The transcriptomic sequencing analysis showed that abnormal activation of the RhoA/ROCK signaling pathway was the main reason for increased NAP-2 secretion. The expression levels of epithelial-mesenchymal transition (EMT) biomarkers and NAP-2 in hypospadias animal models were detected with Immunohistochemistry, Western blot, Immunofluorescence, and ELISA methods. The expression levels of NAP-2, RhoA/ROCK signaling pathway related proteins, reactive oxygen species (ROS) levels in HUVEC cells, EMT biomarkers and migration capacity of urothelial cells cocultured with HUVEC were measured with ELISA, flow cytometry, Western blot or Transwell assay for further cell experiments. The results showed that DBP leaded to NAP-2 oversecretion from vascular endothelium mainly rely on the activation of RhoA/ROCK signaling pathway and ROS accumulation. The RhoA/ROCK inhibitor fasudil could partially decrease ROS production, and both fasudil and N-acetyl-L-cysteine (NAC) could decrease NAP-2 secretion. Meanwhile, the oversecretion of NAP-2 from HUVEC in coculture system promoted EMT and migration capacity of urothelial cells, and TGF-ß inhibitor LY219761 could block the aberrant activation of EMT process. Therefore, it could be concluded that DBP increase NAP-2 secretion from vascular endothelium by RhoA/ROCK/ROS pathway, and further promote EMT in urothelial cells through TGF-ß pathway. This study provided a novel direction for studying the occurrence of hypospadias and may provide a hypospadias predictive marker in the future.

Immune Co-inhibitory Receptors CTLA-4, PD-1, TIGIT, LAG-3, and TIM-3 in Upper Tract Urothelial Carcinomas: A Large Cohort Study.

Programmed cell death 1 ligand 1), programmed cell death protein-1 (PD-1), cytotoxic T-lymphocyte antigen 4 (CTLA-4), T-cell immunoglobulin and mucin-domain containing-3, lymphocyte activation gene-3, and T-cell immunoglobulin and ITIM domain (TIGIT) are considered major immune co-inhibitory receptors (CIRs) and the most promising immunotherapeutic targets in cancer treatment, but they are largely unexplored in upper tract urothelial carcinoma (UTUC). The aim of this Cohort Study was to provide evidence concerning expression profiles and the clinical significance of CIRs among Chinese UTUC patients. A total of 175 UTUC patients who received radical surgery in our center were included. We used immunohistochemistry to evaluate CIR expressions in tissue microarrays (TMAs). Clinicopathological characteristics and prognostic correlations of CIR proteins were retrospectively analyzed. TIGIT, T-cell immunoglobulin and mucin-domain containing-3, PD-1, CTLA-4, Programmed cell death 1 ligand 1, and lymphocyte activation gene-3 high expression was examined in 136(77.7%), 86(49.1%), 57(32.6%), 18(10.3%), 28(16.0%), and 18(10.3%) patients, respectively. Log-rank tests and Multivariate Cox analysis both implied CTLA-4 and TIGIT expression was associated with worse relapse-free survival. In conclusion, this is the largest Chinese UTUC cohort study, and we analyzed the Co-inhibitory receptor expression profiles in UTUC. We identified CTLA-4 and TIGIT expression as promising biomarkers for tumor recurrence. Furthermore, a subset of advanced UTUCs are probably immunogenic, for which single or combined immunotherapy may be potential therapeutic approaches in the future.

Diagnosing and Treating Infertility via Transvaginal Natural Orifice Transluminal Endoscopic Surgery versus Laparoendoscopic Single-Site Surgery: A Retrospective Study.

OBJECTIVE: To evaluate the efficacy and safety of transvaginal natural orifice transluminal endoscopic surgery (vNOTES) in the treatment of female infertility. MATERIALS AND METHODS: This study includes 174 female patients with a history of long-standing female infertility. We retrospectively reviewed 41 patients who underwent hysterolaparoscopy (HL) via transvaginal natural orifice transluminal endoscopic surgery (vNOTES) and 133 patients who underwent laparoendoscopic single-site surgery (LESS). Demographic data, operation records, and pregnancy outcomes were collected and analyzed. The deadline for postoperative follow-up was June 2022. All the included patients were followed up for at least 18 months after surgery. RESULTS: Compared with the LESS group, the vNOTES group had a shorter postoperative bowel movement time and less pain at 4 and 12 h (p = 0.004 vs. 0.008); no differences were found in other perioperative indicators. The clinical pregnancy rates of the vNOTES and LESS groups were 87.80% and 74.43% (p = 0.073), respectively. CONCLUSIONS: vNOTES represents a new, less invasive approach for infertility diagnosis and treatment that is particularly suitable for women who have special esthetic requirements. vNOTES is safe and practical and may be an ideal choice for scarless infertility surgery.

Inherited mutations in Chinese patients with upper tract urothelial carcinoma.

Upper tract urothelial carcinoma (UTUC) accounts for 10% of urothelial carcinomas (UCs) and has a substantial hereditary component. However, the majority of our knowledge of germline spectrum comes from bladder cancer (BCa) data in White populations. Here, we sequence 309 Chinese UTUC cases and identify 71 germline pathogenic/likely pathogenic (P/LP) mutations in 62 patients (20.1%). Compared with White cases, we observe disparities and similarities in inherited mutational profiles. Association analysis reveals that germline P/LP mutations in MSH2, BRCA2, BRCA1, and BRIP1 significantly increase UTUC risk in Chinese populations. Furthermore, germline P/LP mutation in homologous recombination genes indicates poor prognosis for non-metastatic UTUC. Finally, we perform paired sequencing and observe significant correlations between germline mutation patterns and tumor subtypes. This study highlights the importance of genetic testing in patients with UTUC and calls for germline data from various ethnicities to better understand this disease.