Kazuki Okubo

Ritonavir Interacts With Belinostat to Cause Endoplasmic Reticulum Stress and Histone Acetylation in Renal Cancer Cells.

The histone deacetylase (HDAC) inhibitor belinostat increases the amount of unfolded proteins in cells by promoting the acetylation of heat shock protein 90 (HSP90), thereby disrupting its chaperone function. The human immunodeficiency virus protease inhibitor ritonavir, on the other hand, not only increases unfolded proteins by suppressing HSP90 but also acts as a proteasome inhibitor. We thought that belinostat and ritonavir together would induce endoplasmic reticulum (ER) stress and kill renal cancer cells effectively. The combination of belinostat and ritonavir induced drastic apoptosis and inhibited the growth of renal cancer cells synergistically. Mechanistically, the combination caused ER stress (evidenced by the increased expression of the ER stress markers) and also enhanced histone acetylation by decreasing the expression of HDACs. To our knowledge, this is the first study that showed a beneficial combined effect of belinostat and ritonavir in renal cancer cells, providing a framework for testing the combination in renal cancer patients.

Primary Realignment for Pelvic Fracture Urethral Injury Is Associated With Prolonged Time to Urethroplasty and Increased Stenosis Complexity

OBJECTIVE To compare the clinical courses of patients with pelvic fracture urethral injury (PFUI) according to initial management strategy. METHODS We reviewed the clinical courses of 63 patients with PFUI who were initially treated elsewhere and underwent delayed anastomotic urethroplasty by a single surgeon between 2008 and 2015. Patients were grouped according to their initial treatment: by suprapubic tube placement alone (49 patients, SPT group) or primary realignment (14 patients, PR group). Time to urethroplasty was defined as the period between injury and delayed urethroplasty. Clinical data regarding the status of urethral stenosis, urethroplasty procedure, and treatment outcome were analyzed. RESULTS The mean time to urethroplasty in the PR group was about 3 times than that in the SPT group (133 months vs 47 months, P = .035). Fifty percent of the PR group (7 of 14) had a history of repeated urethrotomy or dilation before referral, a percentage significantly higher than that of the SPT group (20.4%, 10 of 49, P = .027). The percentage of patients having a false passage and iatrogenic scar was significantly higher in the PR group (42.9% vs 16.3%, P = .035), but there was no significant between-group difference in urethral stenosis length, operative time, operative blood loss, or the percentage of patients requiring inferior pubectomy or urethral rerouting. CONCLUSION PR does not facilitate delayed urethroplasty, and patients who undergo PR are at high risk of having a more complicated stenosis and longer time to urethroplasty, presumably because of repeated transurethral procedures.

Recurrent retroperitoneal soft tissue sarcoma showing drastic reduction after pazopanib administration accompanied by severe liver dysfunction

Pazopanib is an orally bioavailable tyrosine kinase inhibitor anticancer drug approved worldwide for the treatment of metastatic renal cell carcinoma and advanced soft tissue sarcoma. Here we report the case of a patient whose recurrent retroperitoneal soft tissue sarcoma showed a drastic reduction immediately after pazopanib administration accompanied by severe liver dysfunction. His liver function was restored conservatively by giving him hepatoprotectors and having him stop taking pazopanib. The recurrent tumor disappeared but by 4 months later had regrown.

Bosniak Category III Renal Cysts Caused by Crizotinib in an Anaplastic Lymphoma Kinase Gene-Rearranged Non–Small Cell Lung Cancer Patient

Bosniak category III renal cystic masses are often treated with surgical resection because of high risk of malignancy. Crizotinib is an anaplastic lymphoma kinase (ALK) inhibitor used to treat ALK gene-rearranged non-small cell lung cancer and reported to be associated with complex renal cyst formation. We herein report a case of Bosniak category III renal cysts occurred in a crizotinib-treated ALK gene-rearranged non-small cell lung cancer patients. The cysts regressed spontaneously after cessation of crizotinib and we could thus avoid unnecessary surgical resection.

Delanzomib Interacts with Ritonavir Synergistically to Cause Endoplasmic Reticulum Stress in Renal Cancer Cells

Background/Aim: To investigate the efficacy against renal cancer cells of combining the HIV protease inhibitor ritonavir with the novel proteasome inhibitor delanzomib. Materials and Methods: Renal cancer cell lines 769-P, 786-O, Caki-2 and Renca were treated with ritonavir and delanzomib in vitro and in vivo, and the efficacy of combination was evaluated. Results: The combination of ritonavir and delanzomib synergistically inhibited renal cancer growth and suppressed colony formation. It induced robust apoptosis evidenced by increased cell population in the sub-G1 fraction and increased number of annexin-V-positive cells. A 13-day treatment with the combination was well tolerated in the mouse model and inhibited tumor growth significantly. Mechanistically, the combination synergistically induced endoplasmic reticulum stress and inhibited the mammalian target of rapamycin (mTOR) pathway. Conclusion: The effectiveness of combination of ritonavir and delanzomib appears to be due to the induction of ER stress and inhibition of the mTOR pathway.

Panobinostat and Nelfinavir Inhibit Renal Cancer Growth by Inducing Endoplasmic Reticulum Stress

Background/Aim: There is no curative treatment for patients with advanced renal cancer. We believed that the combination of the histone deacetylase inhibitor panobinostat and the human immunodeficiency virus protease inhibitor nelfinavir would kill renal cancer cells by inducing endoplasmic reticulum (ER) stress. Materials and Methods: Using renal cancer cells (769-P, 786-O, Caki-2), the ability of this combination to induce ER stress and its mechanism of action were investigated. Results: The combination of drugs induced apoptosis and inhibited cancer growth effectively both in vitro and in vivo. Mechanistically, the combination induced ER stress and histone acetylation cooperatively. ER stress induction was shown to play a pivotal role in the anticancer effect of the combination because the protein synthesis inhibitor cycloheximide significantly attenuated combination-induced apoptosis. Nelfinavir was also found to increase the expression of the mammalian target of rapamycin (mTOR) inhibitor AMP-activated protein kinase (AMPK) and inhibited the panobinostat-activated mTOR pathway. Conclusion: Panobinostat and nelfinavir inhibit renal cancer growth by inducing ER stress.

Nelfinavir Induces Endoplasmic Reticulum Stress and Sensitizes Renal Cancer Cells to TRAIL

Background/Aim: Induction of endoplasmic reticulum (ER) stress is a novel strategy for cancer treatment. The human immunodeficiency virus protease inhibitor nelfinavir was recently shown to induce ER stress, but its anti-neoplastic activity has never been investigated in renal cancer, as far as we are aware. Materials and Methods: Using renal cancer cells (769-P, 786-O, Caki-2), the ability of nelfinavir to induce ER stress and sensitize them to tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) was tested. Results: Nelfinavir caused apoptosis and inhibited renal cancer growth in a dose-dependent fashion. It also suppressed colony formation significantly. Nelfinavir induced ER stress and increased the expression of TRAIL death receptor (DR) 4 and DR5, sensitizing the cancer cells to TRAIL. This sensitization was blocked by human recombinant DR4/Fc and DR5/Fc chimeric protein, confirming that the sensitization was due to increased expression of both DR4 and DR5. Conclusion: Nelfinavir induces ER stress in renal cancer cells and sensitizes them to TRAIL.

Evaluation of Therapeutic Potential of Phenoxodiol, a Novel Isoflavone Analog, in Renal Cancer Cells

Background/Aim: In the present study, the antineoplastic activity and mechanism of action of phenoxodiol, a novel isoflavone analog, was investigated in renal cancer cells. Materials and Methods: A panel of renal cancer cells (769-P, 786-O, Caki-2) was treated with phenoxodiol in vitro, and the efficacy of treatment was evaluated. Results: MTS assay results showed that phenoxodiol decreased renal cancer viability in a dose-dependent manner. In addition, it inhibited colony formation significantly and perturbed the cell cycle. Treatment with phenoxodiol increased the number of annexin-V-positive cells as well as the expression of cleaved poly ADP ribose polymerase, demonstrating that phenoxodiol induced apoptosis in renal cancer cells. Phenoxodiol also inhibited Akt pathway via dephosphorylation of Akt. Conclusion: Phenoxodiol inhibited Akt pathway and induced apoptosis of renal cancer cells. The present study provides a theoretical basis for future development of a novel therapy effective against renal cancer.

MP73-13 PHENOXODIOL, A NOVEL SOY ISOFLAVONE ANALOG, INHIBITS AKT PATHWAY AND INDUCES RENAL CANCER APOPTOSIS

INTRODUCTION AND OBJECTIVES: The combination of the human immunodeficiency virus (HIV) protease inhibitors lopinavir and ritonavir has been a standard regimen used to treat HIV infection. Ritonavir acts as a chemical booster to enhance lopinavir’s activity. Lopinavir has recently been shown to act against cancer by inducing endoplasmic reticulum (ER) stress, and we thought that the combination would kill renal cancer cells by inducing robust ER stress. METHODS: The viability and clonogenicity of renal cancer cells (769-P, 786-O, Caki-2) treated with clinically feasible concentrations of lopinavir (10-40 mM) and/or ritonavir (5-10 mM) were assessed by MTS assay and colony formation assay. Apoptosis was evaluated by annexin-V assay. Cell cycle analysis was done using flow cytometry. Induction of ER stress and the expression of cell-cycle regulators, apoptosis-associated proteins, NOXA, Akt, BCL-2, and survivin were evaluated by western blot analysis. Drug synergism was assessed by the Chou-Talalay method. RESULTS: Lopinavir in combination with ritonavir inhibited renal cancer growth synergistically (combination index <1). The combination also inhibited clonogenic survival of cancer cells significantly (p <0.05). It perturbed the cell cycle by inhibiting the expression of cyclin D1 and cyclin-dependent kinase 4, increasing the cells in the sub-G1 fraction. The combination caused apoptosis synergistically: 10-20 mM lopinavir increased the number of annexin-V positive cells and the expression of cleaved poly(ADP-ribose) polymerase slightly but in combination with 10 mM ritonavir increased both drastically. As expected, the combination induced ER stress evidenced by the increased expression of the ER stress markers glucose-regulated protein 78 and endoplasmic reticulum resident protein 44. Furthermore, increased expression of NOXA confirmed that the combination-induced apoptosis was a result of ER stress. We also found that the combination decreased the expression of the antiapoptotic proteins BCL-2 and survivin by inhibiting the Akt signaling pathway. CONCLUSIONS: The combination of lopinavir and ritonavir induces ER stress and causes renal cancer apoptosis synergistically. Inhibition of the Akt pathway is another important mechanism of its action.

MP73-12 LOPINAVIR SYNERGIZES WITH RITONAVIR TO CAUSE RENAL CANCER APOPTOSIS VIA INDUCING ENDOPLASMIC RETICULUM STRESS

INTRODUCTION AND OBJECTIVES: The combination of the human immunodeficiency virus (HIV) protease inhibitors lopinavir and ritonavir has been a standard regimen used to treat HIV infection. Ritonavir acts as a chemical booster to enhance lopinavir’s activity. Lopinavir has recently been shown to act against cancer by inducing endoplasmic reticulum (ER) stress, and we thought that the combination would kill renal cancer cells by inducing robust ER stress. METHODS: The viability and clonogenicity of renal cancer cells (769-P, 786-O, Caki-2) treated with clinically feasible concentrations of lopinavir (10-40 mM) and/or ritonavir (5-10 mM) were assessed by MTS assay and colony formation assay. Apoptosis was evaluated by annexin-V assay. Cell cycle analysis was done using flow cytometry. Induction of ER stress and the expression of cell-cycle regulators, apoptosis-associated proteins, NOXA, Akt, BCL-2, and survivin were evaluated by western blot analysis. Drug synergism was assessed by the Chou-Talalay method. RESULTS: Lopinavir in combination with ritonavir inhibited renal cancer growth synergistically (combination index <1). The combination also inhibited clonogenic survival of cancer cells significantly (p <0.05). It perturbed the cell cycle by inhibiting the expression of cyclin D1 and cyclin-dependent kinase 4, increasing the cells in the sub-G1 fraction. The combination caused apoptosis synergistically: 10-20 mM lopinavir increased the number of annexin-V positive cells and the expression of cleaved poly(ADP-ribose) polymerase slightly but in combination with 10 mM ritonavir increased both drastically. As expected, the combination induced ER stress evidenced by the increased expression of the ER stress markers glucose-regulated protein 78 and endoplasmic reticulum resident protein 44. Furthermore, increased expression of NOXA confirmed that the combination-induced apoptosis was a result of ER stress. We also found that the combination decreased the expression of the antiapoptotic proteins BCL-2 and survivin by inhibiting the Akt signaling pathway. CONCLUSIONS: The combination of lopinavir and ritonavir induces ER stress and causes renal cancer apoptosis synergistically. Inhibition of the Akt pathway is another important mechanism of its action.