Parenchymal obliteration by renal masses: Functional and oncologic implications.

OBJECTIVES: Most renal tumors merely displace nephrons while others can obliterate parenchyma in an invasive manner. Substantial parenchymal volume replacement (PVR) by renal cell carcinoma (RCC) may have oncologic implications; however, studies regarding PVR remain limited. Our objective was to evaluate the oncologic implications associated with PVR using improved methodology including more accurate and objective tools. PATIENTS/METHODS: A total of 1,222 patients with non-metastatic renal tumors managed with partial nephrectomy (PN) or radical nephrectomy (RN) at Cleveland Clinic (2011-2014) with necessary studies were retrospectively evaluated. Parenchymal volume analysis via semiautomated software was used to estimate split renal function and preoperative parenchymal volumes. Using the contralateral kidney as a control, %PVR was defined: (parenchymal volumecontralateral-parenchymal volumeipsilateral) normalized by parenchymal volumecontralateral x100%. PVR was determined preoperatively and not altered by management. Patients were grouped by degree of PVR: minimal (<5%, N = 566), modest (5%-25%, N = 414), and prominent (≥25%, N = 142). Kaplan-Meier was used to evaluate survival outcomes relative to degree of PVR. Multivariable Cox-regression models evaluated predictors of recurrence-free survival (RFS). RESULTS: Of 1,122 patients, 801 (71%) were selected for PN and 321 (29%) for RN. Overall, median tumor size was 3.1 cm and 6.8 cm for PN and RN, respectively, and median follow-up was 8.6 years. Median %PVR was 15% (IQR = 6%-29%) for patients selected for RN and negligible for those selected for PN. %PVR correlated inversely with preoperative ipsilateral GFR (r = -0.49, P < 0.01) and directly with advanced pathologic stage, high tumor grade, clear cell histology, and sarcomatoid features (all P < 0.01). PVR≥25% associated with shortened recurrence-free, cancer-specific, and overall survival (all P < 0.01). Male sex, ≥pT3a, tumor grade 4, positive surgical margins, and PVR≥25% independently associated with reduced RFS (all P < 0.02). CONCLUSIONS: Obliteration of normal parenchyma by RCC substantially impacts preoperative renal function and patient selection. Our data suggests that increased PVR is primarily driven by aggressive tumor characteristics and independently associates with reduced RFS, although further studies will be needed to substantiate our findings.

Limitations of Parenchymal Volume Analysis for Estimating Split Renal Function and New Baseline Glomerular Filtration Rate After Radical Nephrectomy.

PURPOSE: Accurately predicting new baseline glomerular filtration rate (NBGFR) after radical nephrectomy (RN) can improve counseling about RN vs partial nephrectomy. Split renal function (SRF)-based models are optimal, and differential parenchymal volume analysis (PVA) is more accurate than nuclear renal scans (NRS) for this purpose. However, there are minimal data regarding the limitations of PVA. Our objective was to identify patient-/tumor-related factors associated with PVA inaccuracy. MATERIALS AND METHODS: Five hundred and ninety-eight RN patients (2006-2021) with preoperative CT/MRI were retrospectively analyzed, with 235 also having NRS. Our SRF-based model to predict NBGFR was: 1.25 × (GlobalGFRPre-RN × SRFContralateral), where GFR indicates glomerular filtration rate, with SRF determined by PVA or NRS, and with 1.25 representing the median renal functional compensation in adults. Accuracy of predicted NBGFR within 15% of observed was evaluated in various patient/tumor cohorts using multivariable logistic regression analysis. RESULTS: PVA and NRS accuracy were 73%/52% overall, and 71%/52% in patients with both studies (n = 235, P < .001), respectively. PVA inaccuracy independently associated with pyelonephritis, hydronephrosis, renal vein thrombosis, and infiltrative features (all P < .03). Ipsilateral hydronephrosis and renal vein thrombosis associated with PVA underprediction, while contralateral hydronephrosis and increased age associated with PVA overprediction (all P < .01). NRS inaccuracy was more common and did not associate with any of these conditions. Even among cohorts where PVA inaccuracy was observed (22% of our patients), there was no significant difference in the accuracies of NRS- and PVA-based predictions. CONCLUSIONS: PVA was more accurate for predicting NBGFR after RN than NRS. Inaccuracy of PVA correlated with factors that distort the parenchymal volume/function relationship or alter renal functional compensation. NRS inaccuracy was more common and unpredictable, likely reflecting the inherent inaccuracy of NRS. Awareness of cohorts where PVA is less accurate can help guide clinical decision-making.

Parenchymal volume preservation during partial nephrectomy: improved methodology to assess impact and predictive factors.

OBJECTIVE: To rigorously evaluate the impact of the percentage of parenchymal volume preserved (PPVP) and how well the preserved parenchyma recovers from ischaemia (Recischaemia ) on functional outcomes after partial nephrectomy (PN) using an accurate and objective software-based methodology for estimating parenchymal volumes and split renal function (SRF). A secondary objective was to assess potential predictors of the PPVP. PATIENTS AND METHODS: A total of 894 PN patients with available studies (2011-2014) were evaluated. The PPVP was measured from cross-sectional imaging at ≤3 months before and 3-12 months after PN using semi-automated software. Pearson correlation evaluated relationships between continuous variables. Multivariable linear regression evaluated predictors of ipsilateral glomerular filtration rate (GFR) preserved and the PPVP. Relative-importance analysis was used to evaluate the impact of the PPVP on ipsilateral GFR preserved. Recischaemia was defined as the percentage of ipsilateral GFR preserved normalised by the PPVP. RESULTS: The median tumour size and R.E.N.A.L. nephrometry score were 3.4 cm and 7, respectively. In all, 49 patients (5.5%) had a solitary kidney. In all, 538 (60%)/251 (28%)/104 (12%) patients were managed with warm/cold/zero ischaemia, respectively. The median pre/post ipsilateral GFRs were 40/31 mL/min/1.73 m2 , and the median (interquartile range [IQR]) percentage of ipsilateral GFR preserved was 80% (71-88%). The median pre/post ipsilateral parenchymal volumes were 181/149 mL, and the median (IQR) PPVP was 84% (76-92%). In all, 330 patients (37%) had a PPVP of <80%, while only 34 (4%) had a Recischaemia of <80%. The percentage of ipsilateral GFR preserved correlated strongly with the PPVP (r = 0.83, P < 0.01) and loss of parenchymal volume accounted for 80% of the loss of ipsilateral GFR. Multivariable analysis confirmed that the PPVP was the strongest predictor of ipsilateral GFR preserved. Greater tumour size and endophytic and nearness properties of the R.E.N.A.L. nephrometry score were associated with a reduced PPVP (all P ≤ 0.01). Solitary kidney and cold ischaemia were associated with an increased PPVP (all P < 0.05). CONCLUSIONS: A reduced PPVP predominates regarding functional decline after PN, although a low Recischaemia can also contribute. Tumour-related factors strongly influence the PPVP, while surgical efforts can improve the PPVP as observed for patients with solitary kidneys.

Thermal Ablation Versus Partial Nephrectomy for cT1 Renal Mass in a Solitary Kidney: A Matched Cohort Comparative Analysis.

BACKGROUND: Nephron-sparing approaches are preferred for renal mass in a solitary kidney (RMSK), with partial nephrectomy (PN) generally prioritized. Thermal ablation (TA) also is an option for small renal masses in this setting; however, comparative functional/survival outcomes are not well-defined. METHODS: A retrospective study of 504 patients (1975-2022) with cT1 RMSK managed with PN (n = 409)/TA (n = 95) with necessary data for analysis was performed. Propensity score was used for matching patients, including age, preoperative glomerular filtration rate (GFR), tumor diameter, R.E.N.A.L. ((R)adius (tumor size as maximal diameter), (E)xophytic/endophytic properties of tumor, (N)earness of tumor deepest portion to collecting system or sinus, (A)nterior (a)/posterior (p) descriptor, and (L)ocation relative to polar lines), and comorbidities. Functional outcomes were compared, and Kaplan-Meier was used to analyze survival. RESULTS: The matched cohort included 132 patients (TA = 66/PN = 66), with median tumor diameter of 2.4 cm, R.E.N.A.L. of 6, and preoperative GFR of 52 ml/min/1.73 m2. Acute kidney injury occurred in 11%/61% in the TA/PN cohorts, respectively (p < 0.01). After recovery, median GFR preserved was 89%/83% for TA/PN, respectively (p = 0.02), and 5-year dialysis-free survival was 96% in both cohorts. Median follow-up was 53 months. Five-year recurrence-free survival (RFS) was 62%/86% in the TA/PN cohorts, respectively (p < 0.01). Five-year local recurrence (LR)-free survival was 74%/95% in the TA/PN cohorts, respectively (p < 0.01). Five-year cancer-specific survival (CSS) was 96%/98% in the TA/PN cohorts, respectively (p = 0.7). Local recurrence was observed in nine of 36 (25%) and five of 30 (17%) patients managed with laparoscopic versus percutaneous TA, respectively. For TA with LR (n = 14), nine patients presented with multifocality and/or cT1b tumors. Twelve LR were managed with salvage TA, and seven remained cancer-free, while five developed systemic recurrence, three with concomitant LR. CONCLUSIONS: Functional outcomes for TA for RMSK were improved compared with PN. Local recurrence was more common after TA and often was associated with the laparoscopic approach, multifocality, and large tumor size. Improved patient selection and greater experience with TA should improve outcomes. Salvage of LR was not always possible. Partial nephrectomy remains the reference standard for RMSK.

Practical Prediction of New Baseline Renal Function After Partial Nephrectomy.

BACKGROUND: Partial nephrectomy (PN) is generally preferred for localized renal masses due to strong functional outcomes. Accurate prediction of new baseline glomerular filtration rate (NBGFR) after PN may facilitate preoperative counseling because NBGFR may affect long-term survival, particularly for patients with preoperative chronic kidney disease. Methods for predicting parenchymal volume preservation, and by extension NBGFR, have been proposed, including those based on contact surface area (CSA) or direct measurement of tissue likely to be excised/devascularized during PN. We previously reported that presuming 89% of global GFR preservation (the median value saved from previous, independent analyses) is as accurate as the more subjective/labor-intensive CSA and direct measurement approaches. More recently, several promising complex/multivariable predictive algorithms have been published, which typically include tumor, patient, and surgical factors. In this study, we compare our conceptually simple approach (NBGFRPost-PN = 0.90 × GFRPre-PN) with these sophisticated algorithms, presuming that an even 90% of the global GFR is saved with each PN. PATIENTS AND METHODS: A total of 631 patients with bilateral kidneys who underwent PN at Cleveland Clinic (2012-2014) for localized renal masses with available preoperative/postoperative GFR were analyzed. NBGFR was defined as the final GFR 3-12 months post-PN. Predictive accuracies were assessed from correlation coefficients (r) and mean squared errors (MSE). RESULTS: Our conceptually simple approach based on uniform 90% functional preservation had equivalent r values when compared with complex, multivariable models, and had the lowest degree of error when predicting NBGFR post-PN. CONCLUSIONS: Our simple formula performs equally well as complex algorithms when predicting NBGFR after PN. Strong anchoring by preoperative GFR and minimal functional loss (≈ 10%) with the typical PN likely account for these observations. This formula is practical and can facilitate counseling about expected postoperative functional outcomes after PN.

Functional recovery after partial nephrectomy in a solitary kidney.

OBJECTIVES: Partial nephrectomy (PN) is the reference standard for renal mass in a solitary kidney (RMSK), although factors determining functional recovery in this setting remain poorly defined. PATIENTS/METHODS: Single center, retrospective analysis of 841 RMSK patients (1975-2022) managed with PN with functional data, including 361/435/45 with cold/warm/zero ischemia, respectively. A total of 155 of these patients also had necessary studies for detailed analysis of parenchymal volume preserved. Acute kidney injury (AKI) was classified by RIFLE (Risk/Injury/Failure/Loss/Endstage). Recovery-from-ischemia (Rec-Ischemia) was defined as glomerular filtration rate (GFR) saved normalized by parenchymal volume saved. Logistic regression identified predictive factors for AKI and predictors of Rec-Ischemia were analyzed by multivariable linear regression. RESULTS: Overall, median preoperative GFR was 56.7 ml/min/1.73m2 and new-baseline and 5-year GFRs were 43.1 and 44.5 ml/min/1.73m2, respectively. Median follow-up was 55 months; 5-year dialysis-free survival was 97%. In the detailed analysis cohort, a primary focus of this study, median warm (n = 70)/cold (n = 85) ischemia times were 25/34 minutes, respectively; and median preoperative, new-baseline and 5-year GFRs were 57.8, 45.0, and 41.7 ml/min/1.73m2, respectively. Functional recovery correlated strongly with parenchymal volume preserved (r = 0.84, p < 0.001). Parenchymal volume loss accounted for 69% of the total median GFR decline associated with PN, leaving only 3 to 4 ml/min/1.73m2 attributed to ischemia and other factors. AKI occurred in 52% of patients and the only independent predictor of AKI was ischemia time. Independent predictors of reduced Rec-Ischemia were increased age, warm ischemia, and AKI. CONCLUSION: The main determinant of functional recovery after PN in RMSK is parenchymal volume preservation. Type/duration of ischemia, AKI, and age also correlated, although altogether their contributions were less impactful. Our findings suggest multiple opportunities for optimizing functional outcomes although preservation of parenchymal volume remains predominant. Long-term function generally remains stable with dialysis only occasionally required.

Neocapsular reconstruction for challenging partial nephrectomy: A feasibility study in a porcine model and recent clinical experience.

OBJECTIVES: Multifocal renal masses and large central-endophytic tumors can be challenging for partial nephrectomy (PN) due to the paucity of capsule remaining after tumor removal. Our objective was to develop a neocapsule to provide tamponade and test its feasibility/safety in a porcine model. METHODS: Eight live pigs (50-70 kg) underwent unclamped open right flank PN. Renal defects were 1 cm deep and had moderate ongoing venous bleeding. A 6 × 9 inch sheet of Nu-knit® was used for neocapsular reconstruction with Fibrillar™ packing to provide modest tamponade and preclude ongoing bleeding. Blood chemistry and hemoglobin (Hb) levels were drawn preoperatively and postoperative Days 3/5/8. On postoperative Day 8, euthanasia was performed, and both kidneys were inspected and analyzed for histologic changes. RESULTS: PN defects ranged from 1 × 1 × 1 cm to 4 × 2 × 1 cm; four pigs had PN performed in both poles and four in one pole. Neocapsular reconstruction was successful (n = 8), with no perioperative complications. Median baseline Hb was 10.4 g/dL, and median Hb postoperative Days 3/5/8 were 10.0/10.8/10.6 g/dL, respectively. Median baseline serum creatinine (SCr) was 1.9 mg/dL, and median SCr postoperative Days 3/5/8 were 1.5/1.4/1.5 mg/dL, respectively. At sacrifice, no significant hematomas were observed. Other than adjacent to the PN site, there were no significant histologic changes in the parenchyma for operative kidneys versus controls. Based on our experience, we recently performed neocapsular reconstruction safely/effectively after extensive PN for multifocal tumors and for an allograft with difficult-to-manage subcapsular hematoma. CONCLUSIONS: Neocapsular reconstruction after PN or capsular trauma appears feasible and safe and may be considered to reduce the risk of perioperative bleeding. However, further study will be needed to confirm the utility/efficacy of this approach.

MicroRNAs as Potential Regulators of GSK-3ß in Renal Cell Carcinoma.

The prognosis of patients with advanced renal cell carcinoma (RCC) has improved with newer therapies, including molecular-targeted therapies and immuno-oncology agents. Despite these therapeutic advances, many patients with metastatic disease remain uncured. Inhibition of glycogen synthase kinase-3ß (GSK-3ß) is a promising new therapeutic strategy for RCC; however, the precise regulatory mechanism has not yet been fully elucidated. MicroRNAs (miRNAs) act as post-translational regulators of target genes, and we investigated the potential regulation of miRNAs on GSK-3ß in RCC. We selected nine candidate miRNAs from three databases that could potentially regulate GSK-3ß. Among these, hsa-miR-4465 (miR-4465) was downregulated in RCC cell lines and renal cancer tissues. Furthermore, luciferase assays revealed that miR-4465 directly interacted with the 3' untranslated region of GSK-3ß, and Western blot analysis showed that overexpression of miR-4465 significantly decreased GSK-3ß protein expression. Functional assays showed that miR-4465 overexpression significantly suppressed cell invasion of A498 and Caki-1 cells; however, cell proliferation and migration were suppressed only in Caki-1 and A498 cells, respectively, with no effect on cell cycle and apoptosis. In conclusion, miR-4465 regulates GSK-3ß expression but does not consistently affect RCC cell function as a single molecule. Further comprehensive investigation of regulatory networks is required in this field.

Selective Use of Neoadjuvant Targeted Therapy Is Associated with Greater Achievement of Partial Nephrectomy for High-complexity Renal Masses in a Solitary Kidney.

Background: Partial nephrectomy (PN) is preferred for a renal mass in a solitary kidney (RMSK), although tumors with high complexity can be challenging. Objective: To evaluate the evolution of RMSK management with a focus on achievement of PN. Design setting and participants: Patients with nonmetastatic RMSK (n = 499) were retrospectively reviewed; 133 had high tumor complexity, including 80 in the pre-tyrosine kinase inhibitor (TKI) era (1999-2008) and 53 in the TKI era (2009-2022). After 2009, 23/53 patients received neoadjuvant TKI and 30/53 had immediate-surgery. Outcome measurements and statistical analysis: Functional outcomes, adverse events and complications, dialysis-free survival, and recurrence-free survival (RFS) were the measures evaluated. Mann-Whitney and χ2 tests were used to compare cohorts, and the log-rank test was applied for survival analyses. Results and limitations: Overall, the median RENAL score was 10 and the median tumor diameter was 5.2 cm. Demographic characteristics, tumor diameter, and RENAL scores were similar between the pre-TKI-era and TKI-era groups. In the TKI era, 23/53 patients (43%) with clear-cell histology were selected for neoadjuvant TKI. These 23 patients had a greater median tumor diameter (7.1 vs 4.4 cm; p = 0.02) and RENAL score (11 vs 10; p = 0.07). After TKI treatment, the median tumor diameter decreased to 5.6 cm and the RENAL score to 9, and tumor volume was reduced by 59% (all p < 0.05). PN was accomplished in 21/23 (91%) the TKI-treated cases and in 27/30 (90%) of the immediate-surgery cases (2009-2022). PN was only accomplished in 52/80 (65%) of the patients from the pre-TKI era (p < 0.01). The 5-yr dialysis-free survival rate was 59% in the pre-TKI-era group and 91% in the TKI-era group. The 5-yr RFS rate was lower in the TKI-era group (59% vs 74%; p = 0.21), which was mostly related to more aggressive tumor biology, as reflected by a predominance of systemic rather than local recurrences. Conclusions: Management of RMSK with high tumor complexity is challenging. Selective use of TKI therapy was associated with greater use of PN, although a randomized study is needed. RFS mostly reflected aggressive tumor biology rather than failure of local management. Patient summary: For complex kidney tumors in patients with a single kidney, management is challenging. Use of drugs called tyrosine kinase inhibitors before surgery was associated with reductions in tumor size and greater ability to achieve partial kidney removal for cancer control. Most recurrences were metastatic, which reflects aggressive tumor biology rather than failure of surgery.

Targeting Pro-Survival Autophagy Enhanced GSK-3ß Inhibition-Induced Apoptosis and Retarded Proliferation in Bladder Cancer Cells.

Advanced bladder cancer (BC) (local invasive and/or metastatic) is not curable even with cytotoxic chemotherapy, immune checkpoint inhibitors, and targeted treatment. Targeting GSK-3ß is a promising novel approach in advanced BC. The induction of autophagy is a mechanism of secondary resistance to various anticancer treatments. Our objectives are to investigate the synergistic effects of GSK-3ß in combination with autophagy inhibitors to evade GSK-3ß drug resistance. Small molecule GSK-3ß inhibitors and GSK-3ß knockdown using siRNA promote the expression of autophagy-related proteins. We further investigated that GSK-3ß inhibition induced the nucleus translocation of transcription factor EB (TFEB). Compared to the GSK-3ß inhibition alone, its combination with chloroquine (an autophagy inhibitor) significantly reduced BC cell growth. These results suggest that targeting autophagy potentiates GSK-3ß inhibition-induced apoptosis and retarded proliferation in BC cells.

Tumor-infiltrating immune cell status predicts successful response to immune checkpoint inhibitors in renal cell carcinoma.

Immune checkpoint inhibitors (ICI) have dramatically changed the treatment of metastatic renal cell carcinoma (mRCC). Although many studies have reported biomarkers as predicting the efficacy of ICI in mRCC, they remain controversial and have challenges to apply in real-world practice. We evaluated prognostic significance of multiple molecules associated with tumor immunity in patients treated with ICI. The molecules were detected in tumor tissues by immunohistochemical staining. We identified CD8-positive T cells and CD68-positive macrophages infiltrating into the tumor tissue as significant favorable prognostic factors for ICI treatment. Conversely, high expression of CD4-positive T cells was associated with poor response to ICI. Furthermore, we demonstrated that scoring for the expression status of these three molecules provides a remarkably accurate biomarker in patients with mRCC. Even the classical approach of immunohistochemistry could predict the outcome of ICI treatment by assessing the combined status of tumor-infiltrating immune cells.

The Association of Tumor Immune Microenvironment of the Primary Lesion with Time to Metastasis in Patients with Renal Cell Carcinoma: A Retrospective Analysis.

Biological or immunological differences in primary lesions between synchronous and metachronous metastatic renal cell carcinoma (mRCC) have been reported. However, the association between the tumor immune microenvironment (TIME) of primary lesions and time to metastasis remains unknown. We investigated the differences in the TIME of primary lesions based on time intervals to metastasis, mainly between the synchronous group (SG; metastasis within 3 months) and metachronous group (MG; metastasis after 3 months), and its association with clinicopathological parameters in patients with mRCC. Overall, 568 patients treated first-line with vascular endothelial growth factor receptor inhibitors comprised the analysis population (SG: N = 307 [54.0%]; MG: N = 261 [46.0%]). SG had a higher proportion of patients with poor prognostic pathological feature tumors: WHO/ISUP grade 4, necrosis, lymphovascular invasion, infiltrative growth pattern, and sarcomatoid differentiation. Regarding the TIME, more immunogenic features were seen in SG than MG, with a higher PD-L1 positivity and a lower proportion of the desert phenotype. This is the first study to examine the differences in the TIME of primary lesions in patients with mRCC based on the time intervals to metastasis. The TIME of primary lesions could affect the time to metastasis.

Development of patient­derived tumor organoids and a drug testing model for renal cell carcinoma.

The selection of effective therapeutic agents is critical for improving the survival of patients with renal cell carcinoma (RCC). The aim of the present study was to develop an ex vivo drug testing assay using patient­derived tumor organoid (TO) cultures. For this purpose, surgical tumor specimens were obtained from 20 patients with RCC. TOs were developed ex vivo from freshly resected RCC tumors, and their histopathological and molecular characteristics were evaluated using histological staining and whole­exome sequencing (WES). Using a cell viability assay, the therapeutic efficacy of standard of care tyrosine kinase inhibitors in RCC TOs was determined. It was found that TOs recapitulated the histological features of primary RCC tumors. Using WES, a strong concordance was identified at the genetic level between the primary tumors and their corresponding TOs. Using patient­derived TO models, a prototype of an ex vivo drug testing assay was developed, and it was found that RCC TOs exhibited differential responses to sunitinib, pazopanib, cabozantinib, axitinib and sorafenib treatment. On the whole, although the predictive value of the current assay has to be tested and validated in future clinical studies, the findings of the present study demonstrate a novel approach for ex vivo drug testing in patient­derived TO models, which may have potential for use in the personalized treatment of cancer patients.

Double J stent migration as renal penetration.

Transurethral double J (DJ) stent placement is a standard method for drainage during ureteral obstruction caused by a ureteral stone and for the management of complications after transurethral ureterolithotripsy (TUL). This is a safe and minimally invasive technique; however, severe cases of DJ stent migration have been reported, although rarely. Herein, we report the CASE of a 48-year-old man with DJ stent migration as renal penetration, which arose as a complication after an unsuccessful TUL. As transurethral DJ stent placement is one of the basic techniques performed by urologists, possible rare complications of the placement should be taken into consideration.

Membrane fusion of phospholipid bilayers under high pressure: Spherical and irreversible growth of giant vesicles.

Membrane fusion of giant vesicles (GVs) for binary bilayers of unsaturated phospholipids, dioleoylphosphatidyl-ethanolamine (DOPE) having an ability to promote membrane fusion, and its homolog dioleoylphosphatidylcholine (DOPC) having an ability to form GV, was investigated under atmospheric and high pressure. While DOPC formed GVs in the presence of inorganic salts with a multivalent metal ion under atmospheric pressure, an equimolar mixture of DOPE and DOPC formed GVs both in the absence and the presence of LaCl3. We examined the change in size and shape of the GVs of this binary mixture in the absence and presence of LaCl3 as a function of time under atmospheric and high pressure. The size and shape of the GVs in the absence of LaCl3 under atmospheric and high pressure and those in the presence of LaCl3 under atmospheric pressure hardly changed with time. By contrast, the GV in the presence of LaCl3 under high pressure gradually changed in the size and shape with time on a time scale of several hours. Namely, the GV became larger than the original GV due to accelerated membrane fusion and its shape became more spherical. This pressure-induced membrane fusion was completely irreversible, and the growth rate was correlated with the applied pressure. The reason for the GV growth by applying pressure was considered on the basis of thermodynamic phase diagrams. We concluded that the growth is attributable to a closer packing of lipid molecules in the bilayer resulting from their preference of smaller volumes under high pressure. Furthermore, the molecular mechanism of the pressure-induced membrane fusion was explored by observing the fusion of two GVs with almost the same size. From their morphological changes, we revealed that the fusion is caused by the actions of Laplace and osmotic pressure.

Histone deacetylase 6 inhibition in urothelial cancer as a potential new strategy for cancer treatment.

Histone deacetylases (HDACs) are enzymes that remove acetyl groups from histones and have attracted attention as potential targets for cancer therapy. Several small molecule inhibitors have been developed to target HDACs; however, clinical trials of pan-HDAC inhibitors have found these types of inhibitors to be inefficient and to be relatively highly toxic. In the present study, the role of one HDAC isozyme, HDAC6, in urothelial cancer was investigated. Protein expression levels and subcellular localization of HDAC6 was identified in surgically resected bladder tumors using immunohistochemistry. The antitumor effects of 12 small molecule HDAC6 inhibitors were also examined in vitro using cultured urothelial cancer cells. The HDAC6 inhibitors decreased cell viability, with IC50 values in the low µM range, as low as 2.20 µM. HDACi D, E and F had the lowest IC50 values. HDAC6 has been previously reported to regulate programmed death-ligand 1 (PD-L1) and PD-L1 expression was found to be a predictor of decreased overall survival time. There was no association between the protein expression level of HDAC6 and PD-L1 in tumor tissues; however, HDAC6 inhibition by specific small molecule inhibitors resulted in decreased expression levels of membranous PD-L1 in cultured urothelial cancer cell lines. The results suggested that inhibition of HDAC6 could be a promising novel approach for the treatment of urothelial cancer.

High-dose-rate brachytherapy and hypofractionated external beam radiotherapy combined with long-term androgen deprivation therapy for very high-risk prostate cancer.

OBJECTIVE: To estimate the outcomes of high-dose-rate brachytherapy combined with hypofractionated external beam radiotherapy in prostate cancer patients classified as very high risk by the National Comprehensive Cancer Network. METHODS: Between June 2009 and September 2015, 66 patients meeting the criteria for very high-risk disease received high-dose-rate brachytherapy (2 fractions of 9 Gy) as a boost of external beam radiotherapy (13 fractions of 3 Gy). Androgen deprivation therapy was administered for approximately 3 years. Biochemical failure was assessed using the Phoenix definition. RESULTS: The median follow-up period was 53 months from the completion of radiotherapy. The 5-year biochemical failure-free, distant metastasis-free, prostate cancer-specific and overall survival rates were 88.7, 89.2, 98.5 and 97.0%, respectively. The independent contribution of each component of the very high-risk criteria was assessed in multivariable models. Primary Gleason pattern 5 was associated with increased risks of biochemical failure (P = 0.017) and distant metastasis (P = 0.049), whereas clinical stage ≥T3b or >4 biopsy cores with Gleason score 8-10 had no significant impact on the two outcomes. Grade 3 genitourinary toxicities were observed in two (3.0%) patients, whereas no grade ≥3 gastrointestinal toxicities occurred. CONCLUSIONS: The present study shows that this multimodal approach provides potentially excellent cancer control and acceptable associated morbidity for very high-risk disease. Patients with primary Gleason pattern 5 are at a higher risk of poor outcomes, indicating the need for more aggressive approaches in these cases.

Clinically relevant GSK­3ß inhibitor 9­ING­41 is active as a single agent and in combination with other antitumor therapies in human renal cancer.

Glycogen synthase kinase­3 (GSK­3), a serine/threonine kinase, is involved in a broad range of pathological processes including cancer. GSK­3 has two isoforms, GSK­3α and GSK­3ß, and GSK­3ß has been recognized as a therapeutic target for the development of new anticancer drugs. The present study aimed to investigate the antitumor effects of 9­ING­41, which is a maleimide­based ATP­competitive small molecule GSK­3ß inhibitor active in patients with advanced cancer. In renal cancer cell lines, treatment with 9­ING­41 alone induced cell cycle arrest and apoptosis, and autophagy inhibitors increased the antitumor effects of 9­ING­41 when used in combination. Treatment with 9­ING­41 potentiated the antitumor effects of targeted therapeutics and increased the cytotoxic effects of cytokine­activated immune cells on renal cancer cell lines. These results provided a compelling rationale for the inclusion of patients with renal cancer in studies of 9­ING­41, both as a single agent and in combination with current standard therapies.

9-ING-41, a small molecule inhibitor of GSK-3beta, potentiates the effects of anticancer therapeutics in bladder cancer.

Glycogen synthase kinase-3 beta (GSK-3ß), a serine/threonine kinase, has been identified as a potential therapeutic target in human bladder cancer. In the present study, we investigated the antitumor effect of a small molecule GSK-3ß inhibitor, 9-ING-41, currently in clinical studies in patients with advanced cancer, in bladder cancer cell lines. We found that treatment with 9-ING-41 leads to cell cycle arrest, autophagy and apoptosis in bladder cancer cells. The autophagy inhibitor chloroquine potentiated the antitumor effects of 9-ING-41 when tested in combination studies. Our findings also demonstrate that 9-ING-41 enhanced the growth inhibitory effects of gemcitabine or cisplatin when used in combination in bladder cancer cells. Finally, we found that 9-ING-41 sensitized bladder cancer cells to the cytotoxic effects of human immune effector cells. Our results provide a rationale for the inclusion of patients with advanced bladder cancer in clinical studies of 9-ING-41.