MCAM and LAMA4 Are Highly Enriched in Tumor Blood Vessels of Renal Cell Carcinoma and Predict Patient Outcome.

The structure and molecular signature of tumor-associated vasculature are distinct from those of the host tissue, offering an opportunity to selectively target the tumor blood vessels. To identify tumor-specific endothelial markers, we performed a microarray on tumor-associated and nonmalignant endothelium collected from patients with renal cell carcinoma (RCC), colorectal carcinoma, or colorectal liver metastasis. We identified a panel of genes consistently upregulated by tumor blood vessels, of which melanoma cell adhesion molecule (MCAM) and its extracellular matrix interaction partner laminin alpha 4 (LAMA4) emerged as the most consistently expressed genes. This result was subsequently confirmed by immunohistochemical analysis of MCAM and LAMA4 expression in RCC and colorectal carcinoma blood vessels. Strong MCAM and LAMA4 expression was also shown to predict poor survival in RCC, but not in colorectal carcinoma. Notably, MCAM and LAMA4 were enhanced in locally advanced tumors as well as both the primary tumor and secondary metastases. Expression analysis in 18 different cancers and matched healthy tissues revealed vascular MCAM as highly specific in RCC, where it was induced strongly by VEGF, which is highly abundant in this disease. Lastly, MCAM monoclonal antibodies specifically localized to vessels in a murine model of RCC, offering an opportunity for endothelial-specific targeting of anticancer agents. Overall, our findings highlight MCAM and LAMA4 as prime candidates for RCC prognosis and therapeutic targeting. Cancer Res; 76(8); 2314-26. ©2016 AACR.

Long-term proliferation of functional human NK cells, with conversion of CD56(dim) NK cells to a CD56 (bright) phenotype, induced by carcinoma cells co-expressing 4-1BBL and IL-12.

4-1BB ligation co-stimulates T cell activation, and agonistic antibodies have entered clinical trials. Natural killer (NK) cells also express 4-1BB following activation and are implicated in the anti-tumour efficacy of 4-1BB stimulation in mice; however, the response of human NK cells to 4-1BB stimulation is not clearly defined. Stimulation of non-adherent PBMC with OVCAR-3 cells expressing 4-1BB ligand (4-1BBL) or IL-12 resulted in preferential expansion of the NK cell population, while the combination 4-1BBL + IL-12 was superior for the activation and proliferation of functional NK cells from healthy donors and patients with renal cell or ovarian carcinoma, supporting long-term (21 day) NK cell proliferation. The expanded NK cells are predominantly CD56(bright), and we show that isolated CD56(dim)CD16(+) NK cells can switch to a CD56(bright)CD16(-) phenotype and proliferate in response to 4-1BBL + IL-12. Whereas 4-1BB upregulation on NK cells in response to 4-1BBL required 'help' from other PBMC, it could be induced on isolated NK cells by IL-12, but only in the presence of target (OVCAR-3) cells. Following primary stimulation with OVCAR-3 cells expressing 4-1BBL + IL-12 and subsequent resting until day 21, NK cells remained predominantly CD56(bright) and retained both high cytotoxic capability against K562 targets and enhanced ability to produce IFNγ relative to NK cells in PBMC. These data support the concept that NK cells could contribute to anti-tumour activity of 4-1BB agonists in humans and suggest that combining 4-1BB-stimulation with IL-12 could be beneficial for ex vivo or in vivo expansion and activation of NK cells for cancer immunotherapy.

T lymphocyte recruitment into renal cell carcinoma tissue: a role for chemokine receptors CXCR3, CXCR6, CCR5, and CCR6.

BACKGROUND: Evidence suggests that some patients with renal cell carcinoma (RCC) respond to immunomodulatory therapies that activate T lymphocytes. A prerequisite for effective T cell therapy is efficient targeting of effector T cells to the tumour site, yet the molecular basis of T cell recruitment to RCC is unknown. Furthermore, some T cells that naturally infiltrate this cancer are regulatory T cells (Tregs) that may suppress antitumour immune responses. OBJECTIVE: Determine the mechanisms of effector and regulatory T cell recruitment to RCC to allow targeted therapy that promotes local anti-tumour immunity. DESIGN, SETTING, AND PARTICIPANTS: Tumour-infiltrating and peripheral blood T cells were collected from 70 patients undergoing nephrectomy for RCC. MEASUREMENTS: T cells were analysed by multicolour flow cytometry for expression of 19 chemokine receptors and 7 adhesion molecules. Receptors that were expressed at higher levels on tumour-infiltrating lymphocytes (TILs) compared with matched peripheral blood lymphocytes (PBLs) were analysed further for their ability to mediate migration responses in TILs and for expression of corresponding ligands in tumour tissue. RESULTS AND LIMITATIONS: Three chemokine receptors-CCR5, CXCR3, and CXCR6-were significantly overexpressed on TILs compared with matched PBLs (n=16 cases) and were capable of promoting migration in vitro. Their corresponding ligands CCL4-5, CXCL9-11, and CXCL16 were all detected in RCC tissue. However, since they were present in all cases studied, it was not possible to correlate ligand expression with levels of T cell infiltration. Foxp3(+) Tregs were enriched within TILs compared with matched PBLs and expressed high levels of CCR5, CXCR3, and CXCR6, as well as CCR6, the ligand for which (CCL20) was detectable in RCC tissue. CONCLUSIONS: Our data support a role for CCR5, CXCR3, and CXCR6 in the selective recruitment of T cells into RCC tissue and, together with CCR6, in the recruitment of Tregs.

Sorafenib induces therapeutic response in a patient with metastatic collecting duct carcinoma of kidney.

BACKGROUND: Collecting duct carcinoma (CDC) is a rare and aggressive variant of renal cell carcinoma (RCC), which has poor response to cytokine therapy and chemotherapy. Introduction of tyrosine kinase inhibitors (TKIs), in particular sorafenib and sunitinib, is changing the treatment paradigm for management of RCC. However, patients with CDC have been excluded from the majority of randomised trials involving the use of TKIs. CASE REPORT: Our patient with metastatic CDC was treated with sorafenib, and demonstrated an excellent response, both clinically and radiologically. She continues on sorafenib treatment with minimal toxicity, and has demonstrated a progression-free survival exceeding 13 months. CONCLUSIONS: This is the first reported case of a patient with CDC responding to sorafenib treatment. Therefore, the role of sorafenib in the management of metastatic CDC needs prospective evaluation.

The natural history of postoperative renal function in patients undergoing ileal conduit diversion for cancer measured using serial isotopic glomerular filtration rate and 99m technetium-mercaptoacetyltriglycine renography.

PURPOSE: There is little consensus regarding long-term followup of renal function in patients who undergo urinary diversion. We established the usefulness of combined serial isotopic glomerular filtration rate measurement and diuresis renography in the early identification of patients at risk for deterioration of renal function following ileal conduit diversion. MATERIALS AND METHODS: A total of 340 patients with ileal conduit diversion who were followed between 1990 and 2000 were identified. We analyzed data on 178 patients who had more than 4 years of followup. Renal function was assessed by serial estimation of serum creatinine, isotopic glomerular filtration rate and diuresis renographic measurements. RESULTS: Of the patients 52 (29%) demonstrated a worsening glomerular filtration rate. Mean followup +/- SEM was 8.2 +/- 0.4 years (range 4 to 30) and 67% of patients had more than 6 years of followup. In this group we found that hypertension, recurrent urinary sepsis and an initial post-diversion glomerular filtration rate of less than 50 ml per minute per 1.73 m were prevalent risk factors. Hypertension was an independent predictor of a decreased glomerular filtration rate in this group. Of 52 patients with a deteriorating glomerular filtration rate 19 had type II or IIIb curves on followup renography, of whom 13 underwent revision surgery. Renal function subsequently stabilized or improved in this group. CONCLUSIONS: Of patients with an ileal conduit 29% have renal function deterioration in the long term. No surgical cause for glomerular filtration rate deterioration was found in 18%. Important predisposing factors in nonobstructed cases were hypertension, recurrent urinary sepsis and a glomerular filtration rate of less than 50 ml per minute per 1.73 m. Hypertension was an independent predictor of a decreased glomerular filtration rate in the group with worsening glomerular filtration rates. In 11% of patients deterioration was due to upper tract obstruction. This was identifiable using renography and the glomerular filtration rate. A type IIIb curve was an early indicator of obstruction.

Novel method for the isolation and characterisation of the putative prostatic stem cell.

BACKGROUND: Prostate stem cells, responsible for the development, maturation, and function of the prostate, have been implicated in the aetiology of both benign prostate hyperplasia (BPH) and prostate cancer (CaP). However, research has been hampered by the lack of a definitive stem cell marker. We have adapted the protocol for differential Hoechst 33342 uptake by hemopoietic stem cells to enable isolation of putative stem cells from the prostate. METHODS: Prostate epithelial cells isolated from prostate tissue obtained from patients with BPH after transurethral resection of the prostate were stained with Hoechst 33342. The Hoechst 33342 Red/Blue flow cytometry profile was then determined. Hoechst 33342 and Pyronin Y staining was used to determined the cell cycle status. RESULTS: A verapamil-sensitive side population (SP) can be isolated from primary prostate tissue accounting for 1.38% +/- 0.07% of prostate epithelial cells. Cell cycle analysis of this SP population revealed that the majority of SP cells are in either G0 (12.38 +/- 0.31%) or G1 (63.19 +/- 2.13%). CONCLUSIONS: The Hoechst 33342 dye efflux protocol can be adapted for the isolation of a SP from primary prostate tissue.