[The present state and the future perspective of immunotherapy of renal cell carcinoma].

In extremely rare cases, the metastatic lesions of renal cell carcinoma (RCC) spontaneously regress after removal of the primary disease, and thus RCC has been considered to be immunogenic. Thus far, several immunotherapies including cytokine therapy, peptide vaccine and recent immune checkpoints inhibitors have been employed to treat patients with metastatic RCC. In this review, the recent progress in immunotherapy against RCC is surveyed and its future perspective is discussed. Our experimental research on galectin 9 and PINCH as promising immunotherapy targets is also mentioned.

Present and future perspectives on immunotherapy for advanced renal cell carcinoma: Going to the core or beating around the bush?

Metastatic lesions of renal cell carcinoma (RCC) occasionally regress spontaneously after surgical removal of the primary tumor. Although this is an exceptionally rare occurrence, RCC has thus been postulated to be immunogenic. Immunotherapies, including cytokine therapy, peptide-based vaccines, and immune checkpoint inhibitors have therefore been used to treat patients with advanced, metastatic RCC. We review the history, trends, and recent progress in immunotherapy for advanced RCC and discuss future perspectives, with consideration of our experimental work on galectin 9 and PINCH as promising specific immunotherapy targets.

Induction of HLA-A*33-restricted cytotoxic lymphocytes against renal cell carcinoma targeting galectin 9 and PINCH.

Galectin 9, a ligand of T cell immunoglobulin and mucin domain 3 (TIM-3), and PINCH, an epithelial-to-mesenchymal transition (EMT)-promoting molecule, are expressed at much higher levels in cancerous lesions of clear cell type renal cell carcinoma (RCC) compared to normal renal tissues, and their expression levels are extremely low in normal tissues, except for galectin 9 in the spleen. Galectin 9- and PINCH-derived peptides have previously been shown to induce human leukocyte antigen (HLA)-A*2402-restricted and HLA-A*0201-restricted cytotoxic lymphocytes (CTLs) with specific and highly cytotoxic activities toward RCC cells. The present study aimed to identify the peptides that induced HLA-A*33-restricted CTLs that exhibited specific and highly cytotoxic activities toward RCC cells. Specific CTLs were induced significantly, as shown by cluster of differentiation 107a degranulation stimulated with VMRC-RCW renal carcinoma cells. Therefore, peptide vaccines targeting galectin 9 and PINCH appear to be promising for clinical application.

Galectin 9 and PINCH, novel immunotherapy targets of renal cell carcinoma: a rationale to find potential tumour antigens and the resulting cytotoxic T lymphocytes induced by the derived peptides.

OBJECTIVE: To analyse and then generalize the mechanism by which partial or complete response is achieved among a limited number of patients with metastatic renal cell carcinoma (RCC) treated with interferon or interleukin-2. MATERIALS AND METHODS: An expression library of RCC (clear-cell carcinoma) was screened using the sera of patients with metastatic RCC who benefited from partial or complete response to cytokine therapy, the postulation being that those remarkable responders obtained specific cellular immunity against RCC with the antibodies to react with the cancer antigen. Peripheral blood mononuclear-cells (PBMCs) from healthy volunteers were stimulated with the antigen-derived peptides to induce specific cytotoxic T lymphocytes (CTLs). Specific activities of CTLs were measured by 5¹Cr-releasing assay. RESULTS: Among 15 positive clones isolated, two novel genes, galectin 9 and PINCH, were expressed at much higher levels in cancerous lesions than in normal tissues in all the patients with clear-cell carcinoma who were examined. Both HLA-A*2402-restricted and HLA-A*0201-restricted CTLs were induced by each antigen-derived peptide to exhibit specific and highly cytotoxic activities towards RCC cells. Specific CTLs were induced abundantly, as shown by flow cytometry analysis of the CTLs labelled with fluorescein isothiocyanate anti-CD107a and APC anti-CD8. The clonal expansion of the CTLs was shown by the clonality of T-cell receptor Vß repertoires. CONCLUSION: A novel approach based on clinical observations yielded promising tumour antigens as immunotherapy targets of RCC.

NEDD9 crucially regulates TGF-ß-triggered epithelial-mesenchymal transition and cell invasion in prostate cancer cells: involvement in cancer progressiveness.

BACKGROUND: NEDD9 is one of the Crk-associated substrate (Cas) family proteins that mediate downstream signaling processes including cytoskeletal organization, cell-cycle and tumorigenesis. While NEDD9 plays a crucial role in epithelial-mesenchymal transition (EMT), the functional mechanism underlying NEDD9-mediated EMT in prostate cancer (PCa) remains uncertain. METHODS: The expression levels of NEDD9 and its downstream molecules in PC-3, LNCaP, and VCaP cells exposed to transforming growth factor-ß (TGF-ß) were determined by western blotting. The invasion of these cells with ectopic overexpression of NEDD9 or silencing of NEDD9 expression was measured by transwell invasion assay. Human tissue samples comprising 45 PCa specimens and ten specimens of normal prostatic tissue were used for immunohistochemical (IHC) analysis of NEDD9 expression. RESULTS: Both NEDD9 and its downstream signaling molecules associated with EMT were strongly induced by TGF-ß in PCa cells. PC-3 cells with stable overexpression of NEDD9 had a mesenchymal phenotype and significantly enhanced cell invasion, despite their decreased cell proliferation. Knockdown of endogenous NEDD9 expression completely diminished TGF-ß-triggered tumor invasion in several PCa cell lines. The IHC data revealed a significant positive correlation between the NEDD9 staining score and tumor aggressiveness (e.g., Gleason grade, serum PSA level). The NEDD9 staining score in primary PCa with bone metastasis was significantly higher than that in PCa without metastasis. CONCLUSIONS: NEDD9 may be a key mediator involved in TGF-ß-mediated EMT and cell motility in PCa cells and a novel target in the treatment of metastatic PCa and prevention of spread of localized PCa cells to other organs.

Conversion of stable ABO-incompatible kidney transplant recipients from mycophenolate mofetil with standard exposure calcineurin inhibitors (CNIs) to everolimus with very low exposure CNIs-a short-term pilot study.

BACKGROUND: A recent report has demonstrated that as with mycophenolate mofetil (MMF), everolimus is capable of inhibiting human B-lymphocyte function and activation including B-lymphocyte proliferation, apoptosis, and immunoglobulin production in vitro. Everolimus may therefore be used as an immunosuppressant in ABO-incompatible kidney transplantation. METHODS: A three-month pilot study was performed to examine the efficacy and safety of conversion of stable ABO-incompatible kidney transplant recipients from MMF with standard exposure calcineurin inhibitors (CNIs) to everolimus with very low exposure CNIs. Sixteen recipients were enrolled in the study. The patients without acute rejection by graft biopsy were switched from MMF to everolimus with CNI minimization. At three months after conversion, graft biopsies were performed to check for acute rejection and C4d deposition. RESULTS: Conversion to everolimus with CNI minimization for three months did not induce acute rejection and C4d deposition in all of the ABO-incompatible kidney transplant recipients. A slight elevation of anti-A/B antibody titer occurred in our present study. Everolimus was associated with hyperlipidemia and edema. CONCLUSIONS: These results demonstrated that short-term conversion from MMF to everolimus after one yr post-transplant may be a safe and effective alternate for ABO-incompatible kidney transplant recipients requiring temporary discontinuation of MMF.

A Case of Pulmonary Pleomorphic Carcinoma With Renal Metastasis.

A 62-year-old man was referred to our hospital for an axillary mass. Computed tomography (CT) revealed a right axillary tumor and a left renal tumor. Needle biopsies of lung tumor and renal tumor were performed, but a definite diagnosis was impossible. Because his performance status worsened and the lung tumor grew day by day, chemotherapy with gemcitabine and cisplatin was started without definite diagnosis. However, the chemotherapy could not be continued because of interstitial pneumonia and the patient died because of the progression of disease. The final histopathologic diagnosis was pulmonary pleomorphic carcinoma based on immunohistochemical staining.

[Vesicouterine fistula - Youssef syndrome : a case report].

A 38-year-old woman was referred to our hospital with a chief complaint of cyclic hematuria and amenorrhea after Caesarean section. Magnetic resonance imaging showed vesicouterine fistula. The patient was treated with luteinzing hormone-releasing hormone analog to stop menstruation for six months. We performed transperitoneal closure of the vesicouterine fistula. Normal menstruation resumed after 4 months, and the symptoms disappeared. This case was considered Youssef syndrome (cyclic hematuria without vasinal amenorrhea or urinary incontinence). We discuss the cause of this syndrome.

Urinary levels of hepatocarcinoma-intestine-pancreas/pancreatitis-associated protein as a diagnostic biomarker in patients with bladder cancer.

BACKGROUND: To assess the possibility of hepatocarcinoma-intestine-pancreas/pancreatitis-associated protein (HIP/PAP) as a biological marker for detecting Bladder cancer (BCa), we examined the expression of HIP/PAP in both BCa specimens and BCa cell lines and measured HIP/PAP levels in urine from patients with BCa. METHODS: HIP/PAP expression in BCa samples was evaluated by western blot analysis, and urinary levels of HIP/PAP in patients with BCa were measured by enzyme-linked immunosorbent assay. Urine samples were collected from 10 healthy volunteers and 109 with benign urological disorders as controls, and from 101 patients who were diagnosed with BCa. RESULTS: HIP/PAP was highly expressed in BCa samples as compared with control bladder. Urinary HIP/PAP concentrations were significantly higher in BCa patients than in controls (median value; 3.184 pg/mL vs. 55.200 pg/mL, P <0.0001, by Mann-Whitney U test). Urinary HIP/PAP levels in BCa patients correlated positively with pathological T stages and progression-risk groups among non-muscle invasive BCa (P = 0.0008, by Kruskal-Wallis test). Regarding the recurrence-risk classifications of non-muscle invasive BCa, the urinary levels of HIP/PAP were significantly higher in the intermediate than in the low risk group (P = 0.0002, by Mann-Whitney U test). Based on a cut-off of 8.5 pg/mL, the ability of urinary HIP/PAP levels to detect BCa had a sensitivity of 80.2%, specificity of 78.2%, positive predictive value (PPV) of 75.7%, and negative predictive value (NPV) of 82.3%. CONCLUSIONS: HIP/PAP was abundantly expressed in BCa, and the urinary levels of HIP/PAP could be a novel and potent biomarker for detection of BCa, and also for predicting the risks of recurrence- and progression-risk of non-muscle invasive BCa. A large scale study will be needed to establish the usefulness of this biomarker.

Involvement of estrogen receptors in prostatic diseases.

Accumulating evidence shows that estrogens participate in the pathogenesis and development of benign prostatic hyperplasia and prostate cancer by activating estrogen receptor α. In contrast, estrogen receptor ß is involved in the differentiation and maturation of prostatic epithelial cells, and thus possesses antitumor effects in prostate cancer. However, the natural ligands of estrogen receptor ß are not fully understood, and its mode of action according to its ligands and the binding sites located in the promoter regions of downstream genes remains to be elucidated. Here, we review recent experimental investigations of estrogen receptors and their urological relevance. Estrogen receptor-mediated signaling in the prostate is essential together with the androgen receptor-mediated pathway, providing a new therapeutic target for prostatic diseases.

[Successful combined treatment with maximal androgen blockade (MAB), intra arterial chemotherapy and radiation for advanced prostatic ductal adenocarcinoma : report of two cases].

The first case was in a 73-year-old man with macrohematuria. The second case was in a 59-year-old man with pollakiuria. Their serum prostate specific antigen levels were slightly elevated and urinary cytology was negative. Histological examination by prostatic needle biopsy and biopsy from bladder neck showed prostatic ductal adenocarcinoma. Clinical stage on computed tomography and magnetic resonance imaging was T4N0M0 in both cases. After 10-month maximal androgen blockade(MAB) and arterial chemotherapy using reservoir system, radiation therapy was performed. After that, low dose FP-chemotherapy(5-fluorouracil 600 mg/day, cisplatinum 10 mg/day) was performed for 28 days in the first case. At present, there are no signs of recurrence or metastasis in either case.

Hyperbaric oxygen therapy for painful bladder syndrome/interstitial cystitis resistant to conventional treatments: long-term results of a case series in Japan.

BACKGROUND: There is no confirmed strategy for treating painful bladder syndrome/interstitial cystitis (PBS/IC) with unclear etiology. Therefore, a pilot study was carried out to evaluate the efficacy and safety of hyperbaric oxygen (HBO) therapy in treatment-resistant PBS/IC patients. METHODS: HBO treatment (2.0 ATA for 60 minutes/day × 5 days/week for 2 or 4 weeks) was performed on 11 patients with severe symptoms that had not been improved by previous therapy regimens between December 2004 and July 2009. RESULTS: Seven of the 11 patients demonstrated persistent improvement in symptoms during the 12 months after HBO treatment. These responders demonstrated a decrease in the pelvic pain scale and urgency scale from 7.7 ± 1.0 and, 6.6 ± 0.9 to 3.4 ± 2.5 and 4.3 ± 2.4 after 12 months, respectively (p < 0.05). The total score of the interstitial cystitis symptom index and 24-hour urinary frequency demonstrated a significant sustained decrease from the baseline. Two responders, who received an additional course of HBO 12 and 13 months after initial treatment, respectively, did not suffer impairment for more than two years. There was one case of transient eustachian tube dysfunction and three cases of reversible exudative otitis media as a consequence of HBO treatment. CONCLUSIONS: HBO is a potent treatment for PBS/IC patients resistant to conventional therapy. It was well tolerated and provided maintained amelioration of pain, urgency and urinary frequency for at least 12 months.

Combination of a liquid fibrin sealant with sheet-type hemostatic agents: experimental evaluation in partial nephrectomy animal model.

Liquid fibrin sealants, together with sheet-type hemostatic agents, have been used during partial nephrectomies to secure effective hemostasis at the suture site. Using animal kidneys, we investigated which hemostatic agent might adhere most effectively to the renal tissue and serve best as a bolster. Liquid fibrin sealant alone, or in combination with a sheet-type hemostat, such as collagen, gelatin or oxidized-cellulose hemostat, was applied to the cut surface of the kidney of anesthetized rabbits, and the differences in the degree of adherence to the kidney and resultant hemostatic efficacy were evaluated. Histological analyses were also carried out to compare the degree of adherence of each of the aforementioned hemostats to the kidney tissue. Fibrin sealant plus the collagen or gelatin hemostat was found to have a stronger hemostatic effect than fibrin sealant applied alone or fibrin sealant plus oxidized-cellulose hemostat. The histological investigation showed that the fibrin sealant adhered well to kidney tissue when it was applied with the collagen or gelatin hemostat, showing the advantage of combining these two materials for achieving effective hemostasis. Fibrin sealant used in combination with the collagen or gelatin hemostat was the most suitable for obtaining a reinforced hemostatic effect at the suture site in a partial nephrectomy animal model.

Glucose intolerance in renal transplant recipients is associated with increased urinary albumin excretion.

OBJECTIVE: New-onset diabetes mellitus after transplantation (NODAT) is a common and serious complication of renal transplantation, and its incidence is known to be increased by immunosuppressive therapy. It has been reported that urinary albumin excretion is a potent predictor of NODAT. This study was conducted to investigate the relationship between glucose intolerance and urinary albumin excretion in renal transplant recipients. METHODS: A cross-sectional study of 101 renal transplant recipients without prior evidence of diabetes was conducted. All patients underwent an oral glucose tolerance test with 75g of glucose. RESULTS: The patients with glucose intolerance had a significantly greater urinary albumin excretion than those with normal glucose tolerance. Multivariate logistic regression analysis revealed that the increase of urinary albumin excretion correlated significantly with the homeostasis model assessment of insulin resistance and systolic pressure. CONCLUSION: These results indicated that glucose intolerance is associated with increased albuminuria in renal transplant recipients. The rise in insulin resistance and systolic pressure may contribute to the increase of urinary albumin excretion in renal transplant recipients.

Clinical outcome of ABO-incompatible living unrelated donor kidney transplantation.

BACKGROUND: ABO-incompatible living unrelated donor kidney transplantation is an immunologically high-risk procedure, but few reports have been made on the outcomes of these transplants. PATIENTS AND METHODS: We analyzed 12 kidney transplants using ABO-incompatible living-unrelated donors performed at our institution between January 1999 and December 2007, focusing on the immunosuppressive protocols, complications and graft survivals. RESULTS: Patient and graft survival rates were 100%. One patient experienced antibody-mediated rejection and intractable acute cellular rejection, one had antibody-mediated rejection and one had acute cellular rejection, but their grafts survived after intensive immunosuppressive treatment. There were no severe complications among the recipients. CONCLUSIONS: In ABO-incompatible living unrelated donor kidney transplantation, severe rejections may occur due to ABO incompatibility and poor histocompatibility. Therefore, appropriate desensitization, immunosuppression and recipient care are needed for a successful transplant. Recent significant improvements in outcomes indicate that it has become a viable treatment option, given the lack of available donor organs.

Low-grade albuminuria reduction with angiotensin II type 1 receptor blocker in renal transplant recipients.

BACKGROUND: Microalbuminuria, defined as urine albumin to urine creatinine ratio of 30 to <300 mg/g, is an established risk factor for cardiovascular morbidity and mortality in the general population. Low-grade albuminuria (<30 mg/g) is considered a marker for subclinical vascular damage that predisposes to future cardiovascular diseases and death. Lowering urinary albumin excretion reduces the risk of cardiovascular disease. Our study was designed to evaluate the influence of angiotensin II type 1 receptor blocker (ARB) in normotensive renal transplant recipients with low-grade albuminuria. PATIENTS AND METHODS: Our 6-month prospective observation study used a randomized control and open-label design as we examined the effects of an ARB (valsartan) on blood pressure, urinary albumin excretion, hematocrit, serum potassium and estimated glomerular filtration rate (eGFR) in normotensive recipients with allografts of more than 1 year. A total of 35 renal transplant recipients were enrolled in this study. Patients were randomly assigned to 2 groups: ARB group (n=18), receiving 40-80 mg valsartan daily for 6 months, and the control group (n=17). RESULTS: In the ARB group, urine albumin excretion was significantly reduced from 25.9 ± 19.1 mg/g to 12.0 ± 9.6 mg/g at 6 months after administration. eGFR decreased slightly at 6 months after administration. However, no patients undergoing treatment for adverse effects required discontinuation of ARB. CONCLUSIONS: This study reveals that ARB is safe and reduces low-grade albuminuria in normotensive renal transplant recipients. Thus, early treatment of ARB in recipients with low-grade albuminuria may prevent cardiovascular disease after renal transplantation.

[Primary urothelial carcinoma with sarcomatous transformation of the prostate].

A 64-year-old man visited our hospital presenting with macroscopic hematuria. Right hydronephrosis and hypertrophy of the prostate were shown by DIP and MRI respectively. A small papillary tumor at the prostatic urethra was found by cystourethroscopy. Then, we performed transurethral resection of the tumor and trans-perineal needle biopsy of the prostate, and diagnosed him as primary urothelial carcinoma of the prostate. Following neo-adjuvant chemotherapy(MVAC), the patient was treated with radical cystoprostatectomy. The histopathological examination showed urothelial carcinoma with concomitant sarcomatous transformation. Six months after the surgery, he had a recurrence of the tumor in the pelvic cavity. He was treated with the second-line chemotherapy using paclitaxel and gemcitabin combined with the radiation therapy, resulting in the disappearance of the tumor. No evidence of the recurrence has been observed for 3 years.

Elevated urinary levels and urothelial expression of hepatocarcinoma-intestine-pancreas/pancreatitis-associated protein in patients with interstitial cystitis.

OBJECTIVES: To investigate the pathophysiology of interstitial cystitis (IC), we examined urinary levels and urothelial expression of human orthologue of pancreatitis-associated protein (PAP) III, namely, hepatocarcinoma-intestine-pancreas (HIP)/PAP, in patients with IC. We have previously shown that PAP III is expressed with increased frequency in the bladder urothelium in a rat cystitis model (Takahara Y, et al. J Urol. 2008;179:1603-1609). METHODS: HIP/PAP levels in the urine from patients with IC were measured by enzyme-linked immunosorbent assay, and HIP/PAP expression in the bladder tissues was assessed by immunohistochemical study. RESULTS: The median concentration of urinary HIP/PAP was significantly higher in patients with IC than in controls (13.67 vs 1.86 pg/mL urine, respectively, P <.0001, with Mann-Whitney U test). HIP/PAP immunoreactivity was observed in the urothelium of 88.2% of patients with IC (15/17) and in 5.9% of bladder tissues of controls (1/17). Urinary HIP/PAP levels in patients with IC were positively correlated with urinary frequency and bladder pain (r = 0.429, 0.443) and inversely correlated with mean voided urine volume (r = -0.488). CONCLUSIONS: Urinary HIP/PAP levels were significantly higher in IC patients and the apparent HIP/PAP expression in the bladder urothelium was more frequently observed among IC patients. The involvement of HIP/PAP in the pathophysiology of IC is suggested.

Indirect antitumor effects of bisphosphonates on prostatic LNCaP cells co-cultured with bone cells.

Bisphosphonates are strong inhibitors of osteoclastic bone resorption in both benign and malignant bone diseases. The nitrogen-containing bisphosphonates (N-BPs) have strong cytotoxicity via inhibition of protein prenylation in the mevalonate pathway, and also demonstrate direct cytostatic and proapoptotic effects on prostate cancer cells. We confirmed the usefulness of a co-culture system comprised of prostatic LNCaP cells, ST2 cells (mouse-derived osteoblasts) and MLC-6 cells (mouse-derived osteoclasts) in vitro. N-BPs (pamidronate and zoledronic acid) inhibited both androgen receptor transactivation and tumor cell proliferation by suppressing the activities of both osteoclasts and osteoblasts with low-dose exposure. This indirect inhibition of prostate cancer cells via bone cells could be beneficial in treating prostate cancer patients with bone metastases.

Overexpression of orotate phosphoribosyl transferase in hormone-refractory prostate cancer.

Orotate phosphoribosyl transferase (OPRT) is the initial enzyme of 5-fluorouracil (5-FU) activation, in which 5-FU is converted to 5-fluorouridinemonophosphate. Dihydropyrimidine dehydrogenase (DPD) is a degrading enzyme that catabolizes 5-FU. In this study, we investigated the expression of these enzymes in normal prostate gland (NP), hormone-sensitive prostate cancer (HSPC) and hormone-refractory prostate cancer (HRPC). Forty-two prostatic tissue specimens were obtained from patients who had undergone prostate needle biopsies without any treatments or with PSA failure after initial androgen deprivation. The tissue samples derived from formalin-fixed, paraffin-embedded sections were made by laser-captured microdissection and from those RNA was extracted. The levels of OPRT and DPD mRNA expression were examined by quantitative reverse transcriptase-polymerase chain reaction (RT-PCR). The level of OPRT mRNA expression in the HSPC or the HRPC specimens was significantly higher than that in the NP specimens. Immunohistochemical staining for OPRT revealed strong expression of OPRT in prostate cancer cells. There was a significant correlation between OPRT mRNA expression levels and the tumor pathological grade. Furthermore, the OPRT/DPD expression ratio, a powerful predictive factor to evaluate 5-FU sensitivity, in the HRPC group was significantly higher than that in the low grade HSPC group. Thus, 5-FU may be an effective option for some HRPC patients.