The pressure towards innovation and creation of new model systems in regenerative medicine and cancer research has fostered the development of novel potential therapeutic applications. Kidney ...injuries provoke a high request of organ transplants making it the most demanding system in the field of regenerative medicine. Furthermore, renal cancer frequently threaten patients' life and aggressive forms still remain difficult to treat. Ethical issues related to the use of embryonic stem cells, has fueled research on adult, patient-specific pluripotent stem cells as a model for discovery and therapeutic development, but to date, normal and cancerous renal experimental models are lacking. Several research groups are focusing on the development of organoid cultures. Since organoids mimic the original tissue architecture in vitro, they represent an excellent model for tissue engineering studies and cancer therapy testing. We established normal and tumor renal cell carcinoma organoids previously maintained in a heterogeneous multi-clone stem cell-like enriching medium. Starting from adult normal kidney specimens, we were able to isolate and propagate organoid 3D-structures composed of both differentiated and undifferentiated cells while expressing nephron specific markers. Furthermore, we were capable to establish organoids derived from cancer tissues although with a success rate inferior to that of their normal counterpart. Cancer cultures displayed epithelial and mesenchymal phenotype while retaining tumor specific markers. Of note, tumor organoids recapitulated neoplastic masses when orthotopically injected into immunocompromised mice. Our data suggest an innovative approach of long-term establishment of normal- and cancer-derived renal organoids obtained from cultures of fleshly dissociated adult tissues. Our results pave the way to organ replacement pioneering strategies as well as to new models for studying drug-induced nephrotoxicity and renal diseases. Along similar lines, deriving organoids from renal cancer patients opens unprecedented opportunities for generation of preclinical models aimed at improving therapeutic treatments.
Abstract
Extracellular vesicles (EVs) and their cargo represent an intriguing source of cancer biomarkers for developing robust and sensitive molecular tests by liquid biopsy. Prostate cancer (PCa) ...is still one of the most frequent and deadly tumor in men and analysis of EVs from biological fluids of PCa patients has proven the feasibility and the unprecedented potential of such an approach. Here, we exploited an antibody-based proteomic technology, i.e. the Reverse-Phase Protein microArrays (RPPA), to measure key antigens and activated signaling in EVs isolated from sera of PCa patients. Notably, we found tumor-specific protein profiles associated with clinical settings as well as candidate markers for EV-based tumor diagnosis. Among others, PD-L1, ERG, Integrin-β5, Survivin, TGF-β, phosphorylated-TSC2 as well as partners of the MAP-kinase and mTOR pathways emerged as differentially expressed endpoints in tumor-derived EVs. In addition, the retrospective analysis of EVs from a 15-year follow-up cohort generated a protein signature with prognostic significance. Our results confirm that serum-derived EV cargo may be exploited to improve the current diagnostic procedures while providing potential prognostic and predictive information. The approach proposed here has been already applied to tumor entities other than PCa, thus proving its value in translational medicine and paving the way to innovative, clinically meaningful tools.
Intratumor heterogeneity (ITH) and tumor evolution have been well described for clear cell renal cell carcinomas (ccRCC), but they are less studied for other kidney cancer subtypes. Here we ...investigate ITH and clonal evolution of papillary renal cell carcinoma (pRCC) and rarer kidney cancer subtypes, integrating whole-genome sequencing and DNA methylation data. In 29 tumors, up to 10 samples from the center to the periphery of each tumor, and metastatic samples in 2 cases, enable phylogenetic analysis of spatial features of clonal expansion, which shows congruent patterns of genomic and epigenomic evolution. In contrast to previous studies of ccRCC, in pRCC, driver gene mutations and most arm-level somatic copy number alterations (SCNAs) are clonal. These findings suggest that a single biopsy would be sufficient to identify the important genetic drivers and that targeting large-scale SCNAs may improve pRCC treatment, which is currently poor. While type 1 pRCC displays near absence of structural variants (SVs), the more aggressive type 2 pRCC and the rarer subtypes have numerous SVs, which should be pursued for prognostic significance.
In the era of minimally invasive surgery, partial adrenalectomy has certainly been underused. We aimed to report surgical technique and perioperative, pathologic, and early functional outcomes of a ...two-center robot-assisted partial adrenalectomy (RAPA) series.
To detail surgical technique of RAPA for unilateral aldosterone-producing adenoma (UAPA), and to report perioperative and 1-yr functional outcomes.
Data of 10 consecutive patients who underwent RAPA for UAPA at two centers from June 2014 to April 2017 were prospectively collected and reported.
RAPA was performed using a standardized technique with the da Vinci Si in a three-arm configuration.
Baseline and perioperative data were reported. One-year functional outcomes were assessed according to primary aldosteronism surgery outcome guidelines. A descriptive statistical analysis was performed.
All cases were completed robotically. Median nodule size was 18mm (interquartile range IQR 16–20). Intraoperative blood loss was negligible. A single (10%) postoperative Clavien grade 2 complication occurred. Median hospital stay was 3 d (IQR 2–3). Patients became normotensive immediately after surgery (median pre- and postoperative blood pressure: 150/90 and 120/70mmHg, respectively). At both 3-mo and 1-yr functional evaluation, all patients achieved biochemical success (aldosterone level, plasmatic renin activity, and aldosterone-renin ratio within normal range). Complete clinical success was achieved in nine patients, but one required low-dose amlodipine at 6-mo evaluation. At a median follow-up of 30.5 mo (IQR 19–42), neither symptoms nor imaging recurrence was observed.
We demonstrated feasibility and safety of RAPA for UAPA; this technique had very low risk of complications and excellent functional results. Increased availability of robotic platform and increasing robotic skills among urologists make RAPA a treatment option with potential for widespread use in urologic community.
Robot-assisted partial adrenalectomy is a safe, feasible, and minimally invasive surgical approach. Promising perioperative and functional outcomes suggest an increasing adoption of this technique in the near future.
Robot-assisted partial adrenalectomy is a feasible and safe surgical option for small, isolated unilateral aldosterone-producing adenoma. The role of partial adrenalectomy (PA) versus total adrenalectomy is still a matter of debate, and indications for PA should be discussed in a multidisciplinary setting.
The use of robotic surgery (RS) in urology has grown exponentially in the last decade, but RS training has lagged behind. The launch of new robotic platforms has paved the way for the creation of ...innovative robotics training systems. The aim of our study is to test the new training system from Hugo™ RAS System–Medtronic. Between July 2020 and September 2022, a total of 44 residents from urology, gynaecology and general surgery at our institution participated in advanced robotic simulation training using the Hugo™ RAS simulator. Information about sex, age, year of residency, hours spent playing video games, laparoscopic or robotic exposure and interest in robotics (90.9% declared an interest in robotics) was collected. The training program involved three robotic exercises, and the residents performed these exercises under the guidance of a robotics tutor. The residents’ performance was assessed based on five parameters: timing, range of motion, panoramic view, conflict of instruments and exercise completion. Their performance was evaluated according to an objective Hugo system form and a subjective assessment by the tutor. After completing the training, the residents completed a Likert scale questionnaire to gauge their overall satisfaction. The rate of the residents’ improvement in almost all parameters of the three exercises between the first and the last attempts was statistically significant (p < 0.02), indicating significant progress in the residents’ robotic surgical skills during the training. The mean overall satisfaction score ± standard deviation (SD) was 9.4 ± 1.2, signifying a high level of satisfaction among the residents with the training program. In conclusion, these findings suggest that the training program utilizing the Hugo™ RAS System is effective in enhancing robotic surgical skills among residents and holds promise for the development of standardized robotics training programs in various surgical specialties.
Objectives
To compare long‐term functional outcomes of off‐clamp or on‐clamp partial nephrectomy patients of two high‐volume centers with cT1–2/N0 M0 renal tumors and baseline estimated glomerular ...filtration rate >60 mL/min.
Methods
A 3:1 propensity score‐matched analysis was used to select two homogeneous cohorts to compare off‐clamp versus on‐clamp partial nephrectomy. Joinpoint regression analysis was used to compare the 2–8‐year probabilities of estimated glomerular filtration rate modifications in both selected cohorts. The Kaplan–Meier method assessed the risk of developing a stage ≥3b chronic kidney disease during follow up. Multivariable analyses aimed to identify predictors of renal function deterioration. Perioperative complications and oncological outcomes were compared.
Results
Overall, 1073 patients were included (588 on‐clamp and 485 off‐clamp). After applying the propensity score‐matched analysis, the two cohorts of 157 on‐clamp and 472 off‐clamp patients did not differ for all covariates, except for warm ischemia time and last estimated glomerular filtration rate. At joinpoint analysis, the off‐clamp group showed higher probabilities of maintaining an unmodified estimated glomerular filtration rate (P = 0.02). The probability of developing a stage ≥3b chronic kidney disease was significantly higher (P < 0.001) in the on‐clamp cohort. At multivariable analysis, estimated glomerular filtration rate at discharge and off‐clamp approach were independent predictors of improved functional outcomes. Perioperative complications were comparable among the two cohorts (P = 0.67). There were not any statistically significant differences in terms of cancer‐specific survival (P = 0.26) and overall survival (P = 0.18).
Conclusions
Off‐clamp partial nephrectomy seems to offer a higher probability of maintaining 100% estimated glomerular filtration rate after surgery. In our cohort, patients undergoing on‐clamp partial nephrectomy presented a 7.3‐fold increased risk of developing a severe chronic kidney disease during follow up.
The Keap1/Nrf2 pathway is a master regulator of the cellular redox state through the induction of several antioxidant defence genes implicated in chemotherapeutic drugs resistance of tumor cells. An ...increasing body of evidence supports a key role for Keap1/Nrf2 pathway in kidney diseases and renal cell carcinoma (RCC), but data concerning the molecular basis and the clinical effect of its deregulation remain incomplete.Here we present a molecular profiling of the KEAP1 and NFE2L2 genes in five different Renal Cell Carcinoma histotypes by analysing 89 tumor/normal paired tissues (clear cell Renal Carcinoma, ccRCCs; Oncocytomas; Papillary Renal Cell Carcinoma Type 1, PRCC1; Papillary Renal Cell Carcinoma Type 2, PRCC2; and Chromophobe Cell Carcinoma).A tumor-specific DNA methylation of the KEAP1 gene promoter region was found as a specific feature of the ccRCC subtype (18/37, 48.6%) and a direct correlation with mRNA levels was confirmed by in vitro 5-azacytidine treatment. Analysis of an independent data set of 481 ccRCC and 265 PRCC tumors corroborates our results and multivariate analysis reveals a significant correlation among ccRCCs epigenetic KEAP1 silencing and staging, grading and overall survival.Our molecular results show for the the first time the epigenetic silencing of KEAP1 promoter as the leading mechanism for modulation of KEAP1 expression in ccRCCs and corroborate the driver role of Keap1/Nrf2 axis deregulation with potential new function as independent epigenetic prognostic marker in renal cell carcinoma.
Urine analysis via an electronic nose provides volatile organic compounds easily usable in the diagnosis of urological diseases. Although challenging and highly expensive for health systems ...worldwide, no useful markers are available in clinical practice that aim to anticipate prostate cancer (PCa) diagnosis in the early stages in the context of wide population screening. Some previous works suggested that dogs trained to smell urine could recognize several types of cancers with various success rates. We hypothesized that urinary volatilome profiling may distinguish PCa patients from healthy controls. In this study, 272 individuals, 133 patients, and 139 healthy controls participated. Urine samples were collected, stabilized at 37 °C, and analyzed using a commercially available electronic nose (Cyranose C320). Statistical analysis of the sensor responses was performed off-line using principal component (PCA) analyses, discriminant analysis (CDA), and ROC curves. Principal components best discriminating groups were identified with univariable ANOVA analysis. groups were identified with univariable ANOVA analysis. Here, 110/133 and 123/139 cases were correctly identified in the PCa and healthy control cohorts, respectively (sensitivity 82.7%, specificity 88.5%; positive predictive value 87.3%, negative predictive value 84.2%). The Cross Validated Accuracy (CVA 85.3%, p < 0.001) was calculated. Using ROC analysis, the area under the curve was 0.9. Urine volatilome profiling via an electronic nose seems a promising non-invasive diagnostic tool.
To compare outcomes of sutureless (SL) vs. renorrhaphy (RR) off-clamp robotic partial nephrectomy (ocRPN), we retrospectively analyzed procedures performed at our center, from January 2017 to April ...2021, for cT1-2N0M0 renal masses. All the patients with a minimum follow-up < 1 month were excluded from the analysis. The trifecta rate defined surgical quality. Any worsening from chronic kidney disease (CKD) I-II to ≧ IIIa (from IIIa to ≧ IIIb, and from IIIb to ≧ IV) was considered as significant stage migration (sCKDsm). A 1:1 propensity score-matched (PSM) analysis minimized baseline imbalances between SL and RR cohorts in terms of age, gender, ASA score, baseline estimated glomerular filtration rate (eGFR), tumor size, and RENAL score. Logistic regression analyses identified predictors of trifecta achievement. Kaplan-Meier (KM) analysis assessed the impact of RR on significant chronic kidney disease sCKDsm-free survival (SMFS), while Cox regression analyses identified its predictors. Overall, 531 patients were included, with a median tumor size of 3.5 cm (IQR: 2.7-5); 70 (13%) presented with a cT2 mass. An SL approach was pursued in 180 cases, but 10 needed conversion to RR. After PSM analysis, patients receiving SL showed a higher trifecta rate (94% vs. 84%;
= 0.007). SMFS probabilities were comparable at KM analysis (log-rank = 0.69). Age (OR: 0.97; 95%CI: 0.95-0.99;
= 0.01), a RENAL score ≧ 10 (OR: 0.29; 95%CI: 0.15-0.57;
< 0.001), and RR (OR: 0.34; 95%CI: 0.17-0.67;
= 0.002) were independent predictors of trifecta achievement. Age (OR: 1.04; 95%CI: 1.003-1.07;
= 0.03) and baseline eGFR (OR: 0.99; 95%CI: 0.97-0.99;
= 0.05) independently predicted sCKDsm. Compared to RR, our experience seems to show that the SL approach significantly increased the probabilities of achieving the trifecta in the observed group of cases.
Few data are available on survival outcomes of partial nephrectomy performed for cystic renal tumors. We present the first long-term oncological outcomes of cystic (cystRCC) versus pure clear cell ...renal cell carcinoma (ccRCC) in a propensity score-matched (PSM) analysis. Our “renal cancer” prospectively maintained database was queried for “cystRCC” or “ccRCC” and “off-clamp robotic partial nephrectomy” (off-C RPN). The two groups were compared for age, gender, tumor size, pT stage, and Fuhrman grade. A 1:3 PSM analysis was applied to reduce covariate imbalance to <10% and two homogeneous populations were generated. Student t- and Chi-square tests were used for continuous and categorical variables, respectively. Ten-year oncological outcomes were compared between the two cohorts using log-rank test. Univariable Cox regression analysis was used to identify predictors of disease progression after RPN. Out of 859 off-C RPNs included, 85 cases were cystRCC and 774 were ccRCC at histologic evaluation. After applying the PSM analysis, two cohorts were selected, including 64 cystRCC and 170 ccRCC. Comparable 10-year cancer-specific survival probability (95.3% versus 100%, p = 0.146) was found between the two cohorts. Conversely, 10-year disease-free survival probability (DFS) was less favorable for pure ccRCC than cystRCC (66.69% versus 90.1%, p = 0.035). At univariable regression analysis, ccRCC histology was the only independent predictor of DFS probability (HR 2.96 95% CI 1.03–8.47, p = 0.044). At the 10-year evaluation, cystRCC showed favorable oncological outcomes after off-C RPN. Pure clear cell variant histology displayed a higher rate of disease recurrence than cystic lesions.
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