CT-based conventional radiomics and quantification of intratumoral heterogeneity for predicting benign and malignant renal lesions.

BACKGROUND: With the increasing incidence of renal lesions, pretreatment differentiation between benign and malignant lesions is crucial for optimized management. This study aimed to develop a machine learning model utilizing radiomic features extracted from various regions of interest (ROIs), intratumoral ecological diversity features, and clinical factors to classify renal lesions. METHODS: CT images (arterial phase) of 1,795 renal lesions with confirmed pathology from three hospital sites were split into development (1184, 66%) and test (611, 34%) cohorts by surgery date. Conventional radiomic features were extracted from eight ROIs of arterial phase images. Intratumoral ecological diversity features were derived from intratumoral subregions. The combined model incorporating these features with clinical factors was developed, and its performance was compared with radiologists' interpretation. RESULTS: Combining intratumoral and peritumoral radiomic features, along with ecological diversity features yielded the highest AUC of 0.929 among all combinations of features extracted from CT scans. After incorporating clinical factors into the features extracted from CT images, our combined model outperformed the interpretation of radiologists in the whole (AUC = 0.946 vs 0.823, P < 0.001) and small renal lesion (AUC = 0.935 vs 0.745, P < 0.001) test cohorts. Furthermore, the combined model exhibited favorable concordance and provided the highest net benefit across threshold probabilities exceeding 60%. In the whole and small renal lesion test cohorts, the AUCs for subgroups with predicted risk below or above 95% sensitivity and specificity cutoffs were 0.974 and 0.978, respectively. CONCLUSIONS: The combined model, incorporating intratumoral and peritumoral radiomic features, ecological diversity features, and clinical factors showed good performance for distinguishing benign from malignant renal lesions, surpassing radiologists' diagnoses in both whole and small renal lesions. It has the potential to save patients from unnecessary invasive biopsies/surgeries and to enhance clinical decision-making.

Proteome-wide mendelian randomization identifies novel therapeutic targets for chronic kidney disease.

There is an urgent need to pinpoint novel targets for drug discovery in the context of chronic kidney disease (CKD), and the proteome represents a significant pool of potential therapeutic targets. To address this, we performed proteome-wide analyses using Mendelian randomization (MR) and colocalization techniques to uncover potential targets for CKD. We extracted summary-level data from the ARIC study, focusing on 7213 European American (EA) individuals and 4657 plasma proteins. To broaden our analysis, we incorporated genetic association data from Icelandic cohorts, thereby enhancing our investigation into the correlations with chronic kidney disease (CKD), creatinine-based estimated glomerular filtration rate (eGFRcrea), and estimated glomerular filtration rate (eGFR). We utilized genetic association data from the GWAS Catalog, including CKD (765,348, 625,219 European ancestry and 140,129 non-European ancestry), eGFRcrea (1,004,040, European ancestry), and eGFR (567,460, European ancestry). Employing MR analysis, we estimated the associations between proteins and CKD risk. Additionally, we conducted colocalization analysis to evaluate the existence of shared causal variants between the identified proteins and CKD. We detected notable correlations between levels predicted based on genetics of three circulating proteins and CKD, eGFRcrea, and eGFR. Notably, our colocalization analysis provided robust evidence supporting these associations. Specifically, genetically predicted levels of Transcription elongation factor A protein 2 (TCEA2) and Neuregulin-4 (NRG4) exhibited an inverse relationship with CKD risk, while Glucokinase regulatory protein (GCKR) showed an increased risk of CKD. Furthermore, our colocalization analysis also supported the associations of TCEA2, NRG4, and GCKR with the risk of eGFRcrea and eGFR.

CRISPR-Cas9 applications in T cells and adoptive T cell therapies.

T cell immunity is central to contemporary cancer and autoimmune therapies, encompassing immune checkpoint blockade and adoptive T cell therapies. Their diverse characteristics can be reprogrammed by different immune challenges dependent on antigen stimulation levels, metabolic conditions, and the degree of inflammation. T cell-based therapeutic strategies are gaining widespread adoption in oncology and treating inflammatory conditions. Emerging researches reveal that clustered regularly interspaced palindromic repeats-associated protein 9 (CRISPR-Cas9) genome editing has enabled T cells to be more adaptable to specific microenvironments, opening the door to advanced T cell therapies in preclinical and clinical trials. CRISPR-Cas9 can edit both primary T cells and engineered T cells, including CAR-T and TCR-T, in vivo and in vitro to regulate T cell differentiation and activation states. This review first provides a comprehensive summary of the role of CRISPR-Cas9 in T cells and its applications in preclinical and clinical studies for T cell-based therapies. We also explore the application of CRISPR screen high-throughput technology in editing T cells and anticipate the current limitations of CRISPR-Cas9, including off-target effects and delivery challenges, and envisioned improvements in related technologies for disease screening, diagnosis, and treatment.

Enhanced RNA knockdown efficiency with engineered fusion guide RNAs that function with both CRISPR-CasRx and hammerhead ribozyme.

BACKGROUND: CRISPR-Cas13 is a newly emerging RNA knockdown technology that is comparable to RNAi. Among all members of Cas13, CasRx degrades RNA in human cells with high precision and effectiveness. However, it remains unclear whether the efficiency of this technology can be further improved and applied to gene therapy. RESULTS: In this study, we fuse CasRx crRNA with an antisense ribozyme to construct a synthetic fusion guide RNA that can interact with both CasRx protein and ribozyme and tested the ability of this approach in RNA knockdown and cancer gene therapy. We show that the CasRx-crRNA-ribozyme system (CCRS) is more efficient for RNA knockdown of mRNAs and non-coding RNAs than conventional methods, including CasRx, shRNA, and ribozyme. In particular, CCRS is more effective than wild-type CasRx when targeting multiple transcripts simultaneously. We next use bladder cancer as a model to evaluate the anticancer effects of CCRS targeting multiple genes in vitro and in vivo. CCRS shows a higher anticancer effect than conventional methods, consistent with the gene knockdown results. CONCLUSIONS: Thus, our study demonstrates that CCRS expands the design ideas and RNA knockdown capabilities of Cas13 technology and has the potential to be used in disease treatment.

Laparoscopic placement of left renal vein extravascular stenting in treatment of nutcracker syndrome: Techniques and long-term outcomes.

OBJECTIVES: We aimed to assess the feasibility and efficacy of laparoscopic extravascular stent in treatment of nutcracker syndrome by transperitoneal or retroperitoneal approach. METHODS: Seventy-six patients with nutcracker syndrome were retrospectively enrolled from a tertiary referral center, and underwent transperitoneal (63 patients) or retroperitoneal (13 patients) laparoscopic extravascular stent from March 2011 to December 2020. Surgical parameters, complications, imaging and clinical outcomes were collected and analyzed. RESULTS: All procedures were successfully carried out without open conversion. The median operation time, estimated blood loss, and postoperative hospital day were 120 (interquartile range [IQR]: 90-144) min, 20 (IQR: 10-30) ml, and 7 (IQR: 6-9) days. At a median follow-up of 52 (range: 9-127) months, 60 (79%) patients had complete symptom resolution, 14 (18%) patients had significant symptom improvement, and 2 (3%) patients reported no symptom improvement. Ninety-four percent (50/53) of hematuria, 91% (30/33) of proteinuria, and 89% (25/28) of flank/abdominal pain resolved after extravascular LRV stenting. No significant differences were detected in surgery parameters and recovery rates of clinical symptoms between two approaches (each p > 0.05). However, patients with transperitoneal approach need longer to achieve complete recovery compared with retroperitoneal approach (8.7 vs. 1.5 months, p = 0.016). CONCLUSIONS: Laparoscopic extravascular stent performed either transperitoneally or retroperitoneally is a feasible and effective option in treatment of nutcracker syndrome. Retroperitoneal laparoscopic extravascular stent required shorter time to achieve complete recovery, which should be considered whenever possible in surgical decision-making.

The efficacy of modified binding technique for renorrhaphy during robotic partial nephrectomy: surgical and functional outcomes from single-center experience.

BACKGROUND: To compare the traditional single-layer and double-layer suture renorrhaphy with modified "Binding" suture renorrhaphy (whole rim of the wound was closed by the all-layer flow suture starting from the parenchyma cut edges to hilum, followed by the final defect closure) in robotic partial nephrectomy (RPN) for treating localized renal cell carcinoma in our large institutional experience. METHODS: We retrospectively reviewed clinical data of 406 consecutive patients who underwent RPN from May 2018 and December 2020 in our center. The demographic and oncologic outcome variables were compared between different renal reconstruction groups and the effect of these suture techniques on renal function outcomes was also evaluated. RESULTS: For the single-layer group, median operative time and warm ischemic time were significantly less than that of the double-layer and "Binding" groups (p < 0.001), while the significantly lower eGFR drop (p = 0.014) was also detected within postoperative 3 months from baseline, but this difference lost its statistical significance from 3th month to the last follow-up. The changes in postoperative creatinine values were clinically insignificant among the three groups. In a sub-analysis over 258 patients with moderate/high nephrometry score, those patients who underwent "Binding" suture had an undifferentiated warm ischemic time, estimated blood loss, and length of hospitalization stay with a decreased risk of Grade III complications (postoperative hemorrhage requiring intervention) and improved renal function recovery during the whole follow-up. CONCLUSION: Single-layer suture renorrhaphy may be associated with better renal functional preservation and could prove to be reliable in patients with low-complexity tumor (RENAL score ≤ 6). Patients with moderate/high-complexity tumor (RENAL score ≥ 7) might represent a subgroup of patients having a functional benefit after "Binding" suture renorrhaphy even in the long-term period.

[Expression and significance analysis of GDF3 in testicular cancer based on TCGA and GTEx databases].

OBJECTIVE: To investigate the expression and significance of GDF3 in testicular cancer through bioinformatics analysis. METHODS: Using the TCGA and GTEx databases, differential expression analysis and pan-cancer analysis were performed to identify the target gene GDF3, and the clinical relevance of GDF3 in testicular cancer was analyzed using the UALCAN database. Based on the R packages "org.Hs.eg.db" and "clusterProfiler," gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses were conducted to explore the potential functions of GDF3 in testicular cancer. The correlation of GDF3 with immune chemokines and immune inhibitors in testicular cancer was investigated using the TISIDB database. RESULTS: The GDF3 was significantly upregulated in testicular cancer (P<0.001) and closely associated with clinical staging (P<0.05) and tumor subtypes (P<0.001). The immune-related analysis revealed that GDF3 was strongly correlated with immune chemokines CCL26 (rho=0.599, P<0.001), CCL7 (rho=0.525, P<0.001), immune inhibitor ADORA2A (rho=0.723, P<0.001), and PVRL2 (rho=0.585, P<0.001). CONCLUSION: The GDF3 is closely related to the occurrence, development, and immune microenvironment of testicular cancer.

CRISPR signal conductor 2.0 for redirecting cellular information flow.

A key challenge in designing intelligent artificial gene circuits is generating flexible connections between arbitrary components and directly coupling them with endogenous signaling pathways. The CRISPR signal conductor based on conditionally inducible artificial transcriptional regulators can link classic cellular protein signals with targeted gene expression, but there are still problems with multiple signal processing and gene delivery. With the discovery and characterization of new Cas systems and long noncoding RNA (lncRNA) functional motifs, and because of the compatibility of guide RNA with noncoding RNA elements at multiple sites, it is increasingly possible to solve these problems. In this study, we developed CRISPR signal conductor version 2.0 by integrating various lncRNA functional motifs into different parts of the crRNA in the CRISPR-dCasΦ system. This system can directly regulate the expression of target genes by recruiting cellular endogenous transcription factors and efficiently sense a variety of protein signals that are not detected by a classical synthetic system. The new system solved the problems of background leakage and insensitive signaling responses and enabled the construction of logic gates with as many as six input signals, which can be used to specifically target cancer cells. By rewiring endogenous signaling networks, we further demonstrated the effectiveness and biosafety of this system for in vivo cancer gene therapy.

Patient-Derived Upper Tract Urothelial Carcinoma Organoids as a Platform for Drug Screening.

Upper tract urothelial carcinomas (UTUCs) are rare entities that are usually diagnosed at advanced stages. Research on UTUC pathobiology and clinical management has been hampered by the lack of models accurately reflecting disease nature and diversity. In this study, a modified organoid culture system is used to generate a library of 25 patient-derived UTUC organoid lines retaining the histological architectures, marker gene expressions, genomic landscapes, and gene expression profiles of their parental tumors. The study demonstrates that the responses of UTUC organoids to anticancer drugs can be identified and the model supports the exploration of novel treatment strategies. This work proposes a modified protocol for generating patient-derived UTUC organoid lines that may help elucidate UTUC pathophysiology and assess the responses of these diseases to various drug therapies in personalized medicine.

METTL3/N6-methyladenosine/ miR-21-5p promotes obstructive renal fibrosis by regulating inflammation through SPRY1/ERK/NF-κB pathway activation.

Renal fibrosis induced by urinary tract obstruction is a common clinical occurrence; however, effective treatment is lacking, and a deeper understanding of the mechanism of renal fibrosis is needed. Previous studies have revealed that miR-21 impacts liver and lung fibrosis progression by activating the SPRY1/ERK/NF-kB signalling pathway. However, whether miR-21 mediates obstructive renal fibrosis through the same signalling pathway has not been determined. Additionally, studies have shown that N6-methyladenosine (m6 A) modification-dependent primary microRNA (pri-microRNA) processing is essential for maturation of microRNAs, but its role in the maturation of miR-21 in obstructive renal fibrosis has not yet been investigated in detail. To address these issues, we employed a mouse model of unilateral ureteral obstruction (UUO) in which the left ureters were ligated for 3, 7 and 14 days to simulate the fibrotic process. In vitro, human renal proximal tubular epithelial (HK-2) cells were transfected with plasmids containing the corresponding sequence of METTL3, miR-21-5p mimic or miR-21-5p inhibitor. We found that the levels of miR-21-5p and m6 A modification in the UUO model groups increased significantly, and as predicted, the SPRY1/ERK/NF-kB pathway was activated by miR-21-5p, confirming that miR-21-5p plays an important role in obstructive renal fibrosis by enhancing inflammation. METTL3 was found to play a major catalytic role in m6 A modification in UUO mice and drove obstructive renal fibrosis development by promoting miR-21-5p maturation. Our research is the first to demonstrate the role of the METTL3-m6 A-miR-21-5p-SPRY1/ERK/NF-kB axis in obstructive renal fibrosis and provides a deeper understanding of renal fibrosis.

Identification of an IRGP Signature to Predict Prognosis and Immunotherapeutic Efficiency in Bladder Cancer.

BACKGROUND: Identify immune-related gene pairs (IRGPs) signature related to the prognosis and immunotherapeutic efficiency for bladder cancer (BLCA) patients. MATERIALS AND METHODS: One RNA-seq dataset (The Cancer Genome Atlas Program) and two microarray datasets (GSE13507 and GSE31684) were included in this study. We defined these cohorts as training set to construct IRGPs and one immunotherapy microarray dataset as validation set. Identifying BLCA subclasses based on IRGPs by consensus clustering. The Lasso penalized Cox proportional hazards regression model was used to construct prognostic signature and potential molecular mechanisms were analyzed. RESULTS: This signature can accurately predict the overall survival of BLCA patients and was verified in the immunotherapy validation set. IRGP-signatures can be used as independent prognostic risk factor in various clinical subgroups. Use the CIBERSORT algorithm to assess the abundance of infiltrating immune cells in each sample, and combine the results of the gene set enrichment analysis of a single sample to explore the differences in the immune microenvironment between IRPG signature groups. According to the results of GSVA, GSEA, and CIBERSORT algorithm, we found that IRGP is strikingly positive correlated with tumor microenvironment (TME) stromal cells infiltration, indicating that the poor prognosis and immunotherapy might be caused partly by enrichment of stromal cells. Finally, the results from the TIDE analysis revealed that IRGP could efficiently predict the response of immunotherapy in BLCA. CONCLUSION: The novel IRGP signature has a significant prognostic value for BLCA patients might facilitate personalized for immunotherapy.