The phosphoinositide 3-kinase (PI3K)/Akt/mammalian target of rapamycin 1 (mTORC1) signaling pathway is aberrantly activated in renal cell carcinoma (RCC). We previously demonstrated glycogen synthase kinase-3β (GSK-3β) positively regulated RCC proliferation. The aim of this study was to evaluate the role of GSK-3 in the PI3K/Akt/mTORC1 pathway and regulation of the downstream substrates, eukaryotic translation initiation factor 4E-binding protein 1 (4EBP1), ribosomal protein S6 kinase (S6K), and ribosomal protein S6 (S6RP). We used human RCC cell lines (ACHN, Caki1, and A498) and, as normal controls, human renal proximal tubular epithelial cell (HRPTEpC) and non-tumorous kidney tissues that were obtained surgically for treatment of RCC patients. Rapamycin-resistant ACHN (ACHN/RR) cells were generated with chronic exposure of ACHN to rapamycin ranging from 1nM finally to 1 μM. Cell viability, cell cycling and direct interaction between GSK-3β and 4EBP1 were evaluated with MTS assay, flowcytometry and in vitro kinase assay with recombinant GSK-3β and 4EBP1products, respectively. Protein expression and phosphorylation of molecules associated with the PI3K/Akt/mTORC1 pathway were examined by immunoblotting. Effects of drug combination were determined as the combination index with CompuSyn software. Overexpression and phosphorylation of 4EBP1 and S6RP together with GSK-3 activation were observed in RCC cell lines, but not in human normal kidney cells and tissues. Cell proliferation, p4EBP1 and pS6RP were strongly suppressed by GSK-3 inhibition. Rapamycin and LY294002 sufficiently decreased pS6RP, but only moderately p4EBP1. In vitro kinase assays showed that recombinant GSK-3β phosphorylated recombinant 4EBP1, and the effect was blocked by GSK-3 inhibitors. Different from rapamycin, AR- A014418 remarkably inhibited cell proliferation, and rapidly suppressed p4EBP1 and pS6RP in ACHN and ACHN/RR (in 30 min to 1 h). AR- A014418 and rapamycin combination showed additivity at lower concentrations, but antagonism at higher concentrations. GSK-3β could directly phosphorylate 4EBP1 and activate the mTORC1 downstream signaling cascades to enhance protein biosynthesis and cell proliferation in RCC cell lines independent of rapamycin sensitivity. The direct GSK-3β/4EBP1 pathway might be an important subcellular mechanism as an inherent equipment for RCC cells to acquire clinical chemoresistance to mTORC1 inhibitors.

Objectives: Immune checkpoint inhibitors and enfortumab vedotin have opened new avenues for sequential treatment strategies for locally advanced/metastatic urothelial carcinoma (la/mUC). In the pre‐enfortumab vedotin era, many patients could not receive third‐line treatment owing to rapid disease progression and poor general status. This study aimed to analyze real‐world sequential treatment practices for la/mUC in Japan, with a focus on patients who do not receive third‐line treatment.Methods: We analyzed data for 1023 la/mUC patients diagnosed between January 2020 and December 2021 at 54 institutions from a Japanese nationwide cohort.Results: At the median follow‐up of 28.5 months, the median overall survival from first‐line initiation for 905 patients who received systemic anticancer treatment was 19.1 months. Among them, 81% and 32% received second‐ and third‐line treatment. Notably, 52% had their treatment terminated before the opportunity for third‐line treatment. Multivariate logistic regression analysis revealed that low performance status (≥1), elevated neutrophil‐to‐lymphocyte ratio (≥3), and low body mass index (<21 kg/m²) at the start of first‐line treatment were independent risk factors for not proceeding to third‐line treatment (p = 0.0024, 0.0069, and 0.0058, respectively). In this cohort, 33% had one of these factors, 36% had two, and 15% had all three.Conclusions: This study highlights the high frequency of factors associated with poor tolerance to anticancer treatment in la/mUC patients. The findings suggest the need to establish optimal sequential treatment strategies, maximizing efficacy within time and tolerance constraints, while concurrently providing strong supportive care, considering immunological and nutritional aspects.

Objectives: Guidelines for treatment of mRCC recommend nivolumab monotherapy (NIVO) for treated patients, and nivolumab plus ipilimumab combination therapy (NIVO+IPI) for untreated IMDC intermediate and poor-risk mRCC patients. Although molecular-targeted therapies (TTs) such as VEGFR-TKIs and mTORi are recommended as subsequent therapy after NIVO or NIVO+IPI, their efficacy and safety remain unclear. Methods: Outcome of Japanese patients with mRCC who received TT after NIVO (CheckMate 025) or NIVO+IPI (CheckMate 214) were retrospectively analyzed. Primary endpoints were investigator-assessed ORR of the first TT after either NIVO or NIVO+IPI. Secondary endpoints included TFS, PFS, OS and safety of TTs. Results: Twenty six patients in CheckMate 025 and 19 patients in CheckMate 214 from 20 centers in Japan were analyzed. As the first subsequent TT after NIVO or NIVO+IPI, axitinib was the most frequently treated regimen for both CheckMate 025 (54%) and CheckMate 214 (47%) patients. The ORRs of TT after NIVO and NIVO+IPI were 27 and 32% (all risks), and median PFSs were 8.9 and 16.3 months, respectively. During the treatment of first TT after either NIVO or NIVO+IPI, 98% of patients experienced treatment-related adverse events, including grade 3–4 events in 51% of patients, and no treatment-related deaths occurred. Conclusions: TTs have favorable antitumor activity in patients with mRCC after ICI, possibly via changing the mechanism of action. Safety signals of TTs after ICI were similar to previous reports. These results indicate that sequential TTs after ICI may contribute for long survival benefit.

Background: The regulation of the redox balance in the tumor microenvironment is thought to be an adaptive response of tumor cells to hypoxic environments. In recent years, it has been reported that the hemoglobin β-chain (HBB), which is involved in scavenging reactive oxygen species (ROS), is expressed in several carcinomas. However, the relationship between HBB expression and the prognosis of renal cell carcinoma (RCC) remains unclear. Methods: HBB expression was immunohistochemically analyzed in 203 nonmetastatic clear cell RCC (ccRCC) cases. Cell proliferation, invasion, and ROS production were measured in ccRCC cell lines treated with HBB-specific siRNA. Results: The prognosis of HBB-positive patients was worse than that of HBB-negative patients. Cell proliferation and invasion were inhibited, and ROS production was increased by treatment with HBB-specific siRNA. Oxidative stress increased HBB expression in cells exposed to H₂O₂. Conclusions: HBB expression in ccRCC contributes to cancer cell proliferation by suppressing ROS production under hypoxic conditions. Taken together with clinical results and in vitro experiments, HBB expression may serve as a new prognostic biomarker for RCC in the future.

Solitary fibrous tumor (SFT) is a rare neoplasm composed of mesenchymal-derived spindle cells. Although SFT occurs anywhere in the body, they most frequently affects the thoracic region. Here, we reported an extremely rare case of an extrathoracic SFT occurring primarily in the kidney. To our knowledge, little information has been described on the immunohistochemistry (IHC) and genetics of renal SFT. A 41-year old Japanese female came to our hospital for further examination of a left kidney mass detected incidentally with ultrasound. Extensive investigation of the tumor, including physical, laboratory, and image examinations led to a clinical diagnosis of renal cancer (cT1aN0M0), which were in most parts imbedded in the lower polar parenchyma. The patient underwent laparoscopic radical nephrectomy. The mass was diagnosed pathologically as SFT originating from the kidney, but not as renal carcinoma. Microscopically, the tumor was composed of spindle-shape cells distributed variably in dense collagenous stroma and had a focal hemangiopericytomatous staghorn-like vascular pattern. Mitotic figures, atypical structures, necrosis and hemorrhage were not identified. No adjuvant therapies were given postoperatively. The patient has been free of tumor recurrence for 25 months since the surgery. IHC revealed that the tumor diffusely expressed CD34, CD99, Bcl2, PAX8, NAB2, STAT6, and GRIA2. The tumor stained negatively for desmin, S-100, c-Kit, CK-AE1/AE3, CDK4 and MDM2. A NAB2-SATA6 gene fusion was detected in tumor cells by reverse transcription-polymerase chain reaction, direct sequencing, and an in situ proximity ligation brightfield assay. The gene fusion occurred as an 831 bp truncation of exon 2 in NAB2 connected to the beginning of exon 3 in STAT6. We have reported a case of GRIA2 and PAX8-positive SFT occurring primarily in the kidney with such NAB2-STAT6 gene fusion for the first time. Diffuse expression of PAX8 in the tumor might present with a renal origin. Reportedly, benign histology of SFT cannot necessarily predict favorable clinical prognosis. Genetic alterations recently identified in SFT could possibly refer to risk stratification for tumor recurrence. However, malignant preponderance of extrathoracic SFT over thoracic SFT remains unexplained so far. Long-term follow-up after surgery should be performed in the present case.