Randomized Phase III Trial of Gemcitabine and Cisplatin With Bevacizumab or Placebo in Patients With Advanced Urothelial Carcinoma: Results of CALGB 90601 (Alliance).

PURPOSE: The combination of gemcitabine and cisplatin (GC) is a standard therapy for metastatic urothelial carcinoma. Based on data that angiogenesis plays a role in urothelial carcinoma growth and progression, a randomized placebo-controlled trial was performed with the primary objective of testing whether patients treated with GC and bevacizumab (GCB) have superior overall survival (OS) than patients treated with GC and placebo (GCP). PATIENTS AND METHODS: Between July 2009 and December 2014, 506 patients with metastatic urothelial carcinoma without prior chemotherapy for metastatic disease and no neoadjuvant or adjuvant chemotherapy within 12 months were randomly assigned to receive either GCB or GCP. The primary end point was OS, with secondary end points of progression-free survival, objective response, and toxicity. RESULTS: With a median follow-up of 76.3 months among alive patients, the median OS was 14.5 months for patients treated with GCB and 14.3 months for patients treated with GCP (hazard ratio for death = 0.87; 95% CI, 0.72 to 1.05; two-sided stratified log-rank P = .14). The median progression-free survival was 8.0 months for GCB and 6.7 months for GCP (hazard ratio = 0.77; 95% CI, 0.63 to 0.95; P = .016). The proportion of patients with grade 3 or greater adverse events did not differ significantly between both arms, although increased bevacizumab-related toxicities such as hypertension and proteinuria occurred in the bevacizumab-treated arm. CONCLUSION: The addition of bevacizumab to GC did not result in improved OS. The observed median OS of about 14 months is consistent with prior phase III trials of cisplatin-based chemotherapy.

Developments in Vascular-Targeted Photodynamic Therapy for Urologic Malignancies.

With improved understanding of cancer biology and technical advancements in non-invasive management of urological malignancies, there is renewed interest in photodynamic therapy (PDT) as a means of focal cancer treatment. The application of PDT has also broadened as a result of development of better-tolerated and more effective photosensitizers. Vascular-targeted PDT (VTP) using padeliporfin, which is a water-soluble chlorophyll derivative, allows for tumor-specific cytotoxicity and has demonstrated efficacy in the management of urologic malignancies. Herein, we describe the evolution of photodynamic therapy in urologic oncology and the role of VTP in emerging treatment paradigms.

Hydronephrotic urine in the obstructed kidney promotes urothelial carcinoma cell proliferation, migration, invasion through the activation of mTORC2-AKT and ERK signaling pathways.

Obstructive nephropathy is the most common presentation of urothelial carcinoma. The role of the urine in the obstructed kidney namely "hydronephrotic urine" in urothelial carcinoma has not been extensively explored. This study aims to evaluate whether hydronephrotic urine in the obstructed kidney could promote urothelial carcinoma. The hydronephrotic urine was collected from the obstructed kidneys of Sprague-Dawley rats induced by different periods of unilateral ureteral obstruction (UUO). By the inhibition of LY294002 and PD184352, we confirm that hydronephrotic urine promotes urothelial carcinoma cell (T24) and immortalized normal urothelial cells (E6) proliferation, migration and invasion in a dose-dependent manner through the activation of the mTORC2-AKT and ERK signaling pathways. Hydronephrotic urine also increases the expression of cyclin-D2, cyclin-B and CDK2. It also decreases the expression of p27 and p21 in both urothelial carcinoma cells and normal urothelial cells. By the protein array study, we demonstrate that many growth factors which promote tumor cell survival and metastasis are over-expressed in a time-dependent manner in the hydronephrotic urine, including beta-FGF, IFN-γ, PDGF-BB, PIGF, TGF-ß, VEGF-A, VEGF-D and EGF. These results suggest that hydronephrotic urine promotes normal and malignant urothelial cells proliferation, migration and invasion, through the activation of the mTORC2-AKT and ERK signaling pathways. Further investigation using live animal models of tumor growth may be needed to clarify aspects of these statements.

CD105 is a more appropriate marker for evaluating angiogenesis in urothelial cancer of the upper urinary tract than CD31 or CD34.

Angiogenesis plays an important role in cancer progression in many types of cancer. Evaluation of angiogenesis is often performed, but the optimal methodology for human cancer has not been agreed upon. As adequate evaluation of angiogenesis in cancer tissues might be important for prediction of prognosis and treatment decisions, we evaluated angiogenesis semiquantitatively by assessing microvessel density (MVD) in urothelial cancer of the upper urinary tract (UC-UUT). We compared the performance of three endothelial cell markers (CD31, CD34, and CD105) on formalin-fixed tissues from 122 patients diagnosed with UC-UUT without metastasis. Vascular endothelial growth factor (VEGF)-A expression was also evaluated immunohistochemically. Correlations between MVD with each marker and pT stage, grade, survival, and VEGF-A expression were investigated. Mean (standard deviation) MVD as estimated by immunohistochemical staining with anti-CD31, anti-CD34, and anti-CD105 were 47.1 (17.9)/high-power field (HPF), 70.9 (19.5)/HPF, and 31.2 (16.7)/HPF, respectively. Although all MVDs were significantly associated with pT stage and grade, CD105-MVD showed the strongest association. Similarly, CD105-MVD showed the strongest correlation with VEGF-A expression (r = 0.530, p < 0.001). Although all MVDs were associated with metastasis-free survival and cause-specific survival on univariate analysis, only CD105-MVD was retained as an independent predictor in multivariate analysis including pT stage and grade. CD105-MVD may be the preferred marker for semiquantitative assessment of angiogenesis in patients with UC-UUT.

[Future perspectives for diagnostic imaging in urology: from anatomic and functional to molecular imaging]. / Scenari futuri della diagnostica per immagini in urologia: passaggio dall'imaging anatomico e funzionale all'imaging molecolare.

The future approach of diagnostic imaging in urology follows the technological progress, which made the visualization of in vivo molecular processes possible. From anatomo-morphological diagnostic imaging and through functional imaging molecular radiology is reached. Based on molecular probes, imaging is aimed at assessing the in vivo molecular processes, their physiology and function at cellular level. The future imaging will investigate the complex tumor functioning as metabolism, aerobic glycolysis in particular, angiogenesis, cell proliferation, metastatic potential, hypoxia, apoptosis and receptors expressed by neoplastic cells. Methods for performing molecular radiology are CT, MRI, PET-CT, PET-MRI, SPECT and optical imaging. Molecular ultrasound combines technological advancement with targeted contrast media based on microbubbles, this allowing the selective registration of microbubble signal while that of stationary tissues is suppressed. An experimental study was carried out where the ultrasound molecular probe BR55 strictly bound to prostate tumor results in strong enhancement in the early phase after contrast, this contrast being maintained in the late phase. This late enhancement is markedly significant for the detection of prostatic cancer foci and to guide the biopsy sampling. The 124I-cG250 molecular antibody which is strictly linked to cellular carbonic anhydrase IX of clear cell renal carcinoma, allows the acquisition of diagnostic PET images of clear cell renal carcinoma without biopsy. This WG-250 (RENCAREX) antibody was used as a therapy in metastatic clear cell renal carcinoma. Future advancements and applications will result in early cancer diagnosis, personalized therapy that will be specific according to the molecular features of cancer and leading to the development of catheter-based multichannel molecular imaging devices for cystoscopy-based molecular imaging diagnosis and intervention.

Thrombospondin-1-derived 4N1K peptide expression is negatively associated with malignant aggressiveness and prognosis in urothelial carcinoma of the upper urinary tract.

BACKGROUND: Thrombospondin (TSP) is a multi-functional protein that appears to have dual roles in cancer, that is, either as a promoter or a suppressor. 4N1K is a TSP-derived peptide that has been reported to be associated with neovascularity, cell survival, and invasion. There is a little information regarding its pathological roles in human cancer tissues. Our aim was to clarify clinical significance and prognostic value of 4N1K expression in patients with urothelial carcinoma of the upper urinary tract (UC-UUT). METHODS: We investigated 4N1K expression in 97 surgically excised, non-metastasized UC-UUT specimens and five normal tissues via immunohistochemistry. Microvessel density (MVD), lymph vessel density (LVD), cancer cell proliferation (PI), apoptotic index (AI), and matrix metalloproteinase (MMP)-9 expression was also determined. The relationships 4N1K expression and pT stage, grade, and prognosis were analysed. In addition, correlations with these cancer-related and TSP-related factors were also investigated. RESULTS: Strong and moderate 4N1K expression was found in normal urothelial tissues. Of the 97 specimens, 45 patients were positive for 4N1K expression, which was primarily located in the interstitial areas of the cancer tissue. 4N1K expression was negatively associated with pT stage (p = 0.003) and grade (p = 0.002). Survival analyses revealed that 4N1K is a predictor of metastasis-free (p = 0.036) and cause-specific survival (p = 0.009). 4N1K expression was closely associated with malignant behaviour, specifically MVD (p = 0.001), AI (p = 0.013), and MMP-9 expression (p = 0.036), but not PI and LVD, as determined via multivariate analysis models. CONCLUSIONS: 4N1K expression appears to be associated with cancer cell progression and survival in UC-UUT patients via the regulation of angiogenesis, apoptosis, and MMP-9 expression. There is a possibility that the 4N1K-peptide may be a useful marker and novel therapeutic target in patients with UC-UUT.

Phase II study of sunitinib as first-line treatment of urothelial cancer patients ineligible to receive cisplatin-based chemotherapy: baseline interleukin-8 and tumor contrast enhancement as potential predictive factors of activity.

BACKGROUND: A strong rationale supports the role of antiangiogenic drugs in urothelial cancer. This trial was designed to assess the activity of sunitinib as first-line treatment in patients with metastatic urothelial cancer ineligible for cisplatin and to explore molecular and imaging variables predictive of clinical benefit. PATIENTS AND METHODS: This was a multicenter phase II trial with sunitinib 50 mg daily in 4/2-week schedule. Eligibility criteria were as follows: creatinine clearance 30-60 ml/min, Eastern Cooperative Oncology Group Pperformance Sstatus of one or less, and adequate hepatic and hematologic function. Twelve circulating cytokines were evaluated at baseline and sequentially using Luminex xMAP(®) (Austin, TX). Baseline and treatment-related changes in perfusion were evaluated in a patient subgroup using contrast-enhanced computed tomography. RESULTS: On intention-to-treat analysis, 38 patients showed 3 (8%) partial responses (PRs) and 19 (50%) presented with stable disease (SD), 17 (45%) of them ≥3 months. Clinical benefit (PR + SD) was 58%. Median time to progression (TTP) was 4.8 months and median overall survival 8.1 months. Toxicity was consistent with previous reports for sunitinib. Low interleukin-8 (IL-8) baseline levels were significantly associated with increased TTP. Baseline tumor contrast enhancement with >40 Hounsfield units was associated with clinical benefit. CONCLUSIONS: This study highlights the potential role of the angiogenic pathway as a therapy target in urothelial cancer. Baseline IL-8 serum levels and contrast enhancement of lesions warrant further study.

High expression of gastrin-releasing peptide receptors in the vascular bed of urinary tract cancers: promising candidates for vascular targeting applications.

Tumoral gastrin-releasing peptide (GRP) receptors are potential targets for diagnosis and therapy using radiolabeled or cytotoxic GRP analogs. GRP-receptor overexpression has been detected in endocrine-related cancer cells and, more recently, also in the vascular bed of selected tumors. More information on vascular GRP-receptors in cancer is required to asses their potential for vascular targeting applications. Therefore, frequent human cancers (n = 368) were analyzed using in vitro GRP-receptor autoradiography on tissue sections with the (125)I-[Tyr(4)]-bombesin radioligand and/or the universal radioligand (125)I-[d-Tyr(6), beta-Ala(11), Phe(13), Nle(14)]-bombesin(6-14). GRP-receptor expressing vessels were evaluated in each tumor group for prevalence, quantity (vascular score), and GRP-receptor density. Prevalence of vascular GRP-receptors was variable, ranging from 12% (prostate cancer) to 92% (urinary tract cancer). Different tumor types within a given site had divergent prevalence of vascular GRP-receptors (e.g. lung: small cell cancer: 0%; adenocarcinoma: 59%; squamous carcinoma: 83%). Also the vascular score varied widely, with the highest score in urinary tract cancer (1.69), moderate scores in lung (0.91), colon (0.88), kidney (0.84), and biliary tract (0.69) cancers and low scores in breast (0.39) and prostate (0.14) cancers. Vascular GRP-receptors were expressed in the muscular vessel wall in moderate to high densities. Normal non-neoplastic control tissues from these organs lacked vascular GRP-receptors. In conclusion, tumoral vessels in all evaluated sites express GRP-receptors, suggesting a major biological function of GRP-receptors in neovasculature. Vascular GRP-receptor expression varies between the tumor types indicating tumor-specific mechanisms in their regulation. Urinary tract cancers express vascular GRP-receptors so abundantly, that they are promising candidates for vascular targeting applications.

Overexpression of cyclooxygenase-2 in urothelial carcinoma in conjunction with tumor-associated-macrophage infiltration, hypoxia-inducible factor-1alpha expression, and tumor angiogenesis.

This study examines whether the expression of cyclooxgenase-2 (COX-2) in urothelial carcinoma (UC) is associated with macrophage infiltration, hypoxia-inducible factor-1alpha (HIF-1alpha) expression and angiogenesis. We investigated the expression of COX-2 associated with HIF-1alpha and performed double immunohistochemical analysis of 216 UCs for COX-2 expression and the correlation with tumor-associated-macrophage (TAM) density and microvessel density (MVD) in situ. A high expression of COX-2 was positively correlated with tumor invasiveness, histologic grade and HIF-1alpha expression in UC (p<0.0001, p=0.003, p<0.0001, respectively). Quantification of double staining of COX-2/CD34 and COX-2/CD68 showed that a higher MVD and TAM density was found in COX-2 high-expression than in COX-2 low-expression tumor fields (p<0.0001). Adjacent to the principal of COX-2 expression areas, MVD value and TAM density were significantly increased in HIF-1alpha high-expression specimens compared with HIF-1alpha low-expression ones (p<0.0001). Interestingly, our data revealed that high COX-2 expression (p=0.002), high HIF-1alpha expression (p<0.0001) and TAM density (p<0.0001) were all associated with high MVD value. Our results suggest that COX-2 may produce a cooperative effect in promoting tumor progression and may be involved in the process of angiogenesis through increasing TAM infiltration or HIF-1alpha regulation by hypoxia.

[Urological oncology emergencies].

Urologic emergencies in malignancies are difficult to treat for non-urological medical staff because of their anatomical specificity and sensational susceptibility. Urological cancer pain involves somatic and neuropathic problems. In pelvic malignancies, almost always NSAIDs, micro-2-agonisitic opioids and adjuvant analgesics are necessary for pain palliation. The inducements of hemorrhagic cystitis are radiation, chemotherapy and tumor invasion. Although there are some conservative treatments such as hyperbaric oxygen therapy for radiation-induced hemorrhagic cystitis, in emergency circumstances selective embolization of the internal iliac artery may be beneficial with fewer side effects. In uncontrollable tumor bleeding, palliative radiotherapy(maximum dose of 30-36 Gy)is effective. The most important problem in voiding disturbance is bladder outlet obstruction. Sometimes patients are treated with alfa-adrenergic blockers effectively, but eventually they may need an indwelling catheter. Obstructive nephropathy due to malignant ureteral obstruction is an ominous sign of progressive disease, especially in pelvic malignancies. The mean survival rate of 12 months was less than 30%.

Microvessel density (MVD) and cyclooxygenase-2 (COX-2)/ beta-catenin interaction are associated with relapse in patients with transitional carcinoma receiving adjuvant chemotherapy with paclitaxel/carboplatin: a hellenic cooperative oncology group (HECOG) study.

BACKGROUND: Cycloxygenase (COX)-2 has been associated with proliferation, apoptosis and angiogenesis in urothelial cancer. The prognostic significance of COX-2 in patients who received adjuvant chemotherapy for urothelial cancer was examined. PATIENTS AND METHODS: Expression of COX-2, p53, ki67, beta-catenin, vascular endothelial growth factor (VEGF) and microvessel density (MVD) were studied retrospectively in 59 patients with urothelial cancer (pT3, pT4, N+) who had undergone surgery. The patients had subsequently received adjuvant chemotherapy. RESULTS: Thirty-eight out of 59 cases (64%) were positive for COX-2. COX-2 was not associated either with progression-free survival (PFS) or overall survival (OS). MVD levels > or =47 were associated with longer median PFS compared with lower levels (not reached vs. 13 months [95% CI: 8-18], p=0.048). The median PFS for patients with beta-catenin nuclear accumulation and COX-2 expression was 6 months (95% CI: 4-7) compared with 19 months (95% CI: 14-23) for neither or only one of these factors (p=0.018). CONCLUSION: MVD may be a useful indicator of relapse in high-risk urothelial cancer treated with adjuvant chemotherapy.

Targeting vasculature in urologic tumors: mechanistic and therapeutic significance.

Recent advances toward understanding the molecular mechanisms regulating cancer initiation and progression provide new insights into the therapeutic value of targeting tumor vascularity by interfering with angiogenic signaling pathways. The functional contribution of key angiogenic factors toward increased vascularity characterizing metastatic tumors and their therapeutic exploitation is considered in three major urologic malignancies, renal, bladder, and prostate cancer. With the realization that the success of the therapeutic efficacy of the various anti-angiogenic approaches for the treatment of urologic tumors has yet to be proven clinically, the challenge remains to select critical angiogenesis pathways that can be targeted for an individual tumor. Here we discuss the major mechanisms that support formation of vasculature in renal, bladder, and prostate tumors and the current results of targeting of specific molecules/regulators for therapeutic intervention against metastastic disease.

[Molecular imaging of tumor blood vessels]. / Molekulare Bildgebung der Tumorgefässe.

In the past three decades many efforts have been undertaken to understand the mechanisms of tumor angiogenesis. The introduction of the anti-angiogenic drugs in tumor therapy during the last few years necessitates the establishment of new techniques enabling molecular imaging of vascular remodeling. Tumor imaging by X-ray, CT, MRI and ultrasound has to be improved by coupling with molecular markers targeting the tumor vessels. The determination of tumor size as commonly used is not appropriate since the extended necrosis under anti-angiogenic therapy does not result in a reduction of tumor diameter. But remodeling of the tumor vessels under anti-angiogenic therapy obviously occurs at an early stage and seems to be a convincing parameter for tumor imaging. Despite the enormous progress in this field during the last few years the resolution is still not high enough to evaluate the remodeling of the microtumor vessels. Thus, new imaging approaches are needed to overcome this issue.

Mechanisms of disease: angiogenesis in urologic malignancies.

Angiogenesis is critical for growth of tumors and their metastasis. In this article we review the literature on studies of angiogenesis pathways and markers for renal cancer, prostate cancer and bladder cancer. Overall, there is clear evidence that markers of angiogenesis and expression of angiogenic factors are associated with adverse outcomes in each of these tumor types. Relatively few angiogenic pathways have been investigated so far, although over 50 factors are known to be involved, and little has been studied on the antiangiogenic pathways and their suppression. The failing in many of the studies is small size and lack of suitable statistical analysis. Nevertheless, this review demonstrates the importance of these pathways and the need to develop selection criteria for patients who are candidates for antiangiogenic therapies. On the basis of the expression profiles reported so far, therapies that target vascular endothelial growth factor should be considered for the treatment of renal, prostate and bladder cancers. As most tumors express factors that are involved in multiple angiogenic pathways, further research is needed to determine which are coregulated and what the most common patterns are.

Angiogenesis in urothelial tumors of the upper urinary tract.

Angiogenesis is an essential process in the progression of malignant tumors. Tumors of the ureter and renal pelvis account for 5% of all urinary tract neoplasms. Little is known about angiogenesis in upper urinary tract urothelial tumors. We tried to demonstrate angiogenesis by using three endothelial markers CD31, CD34, von Willebrand factor and one pericytes marker (alpha-smooth muscle actin) in 26 cases. The pattern of CD31 immunolabelling was more complex and extensive than the vessel pattern shown by CD34 or factor VIII staining. In non-invasive tumors we observed that angiogenesis process is limited to connective tissue of tumor stroma. In the tumor area, the blood vessels stained with anti-CD31 had large lumen, thin walls and numerous branches, some of them being very thin. Pericyte covered vessels were branching of frequently into smaller, pericyte negative vessels.

The role of angiogenesis in prostate and other urologic cancers: a review.

Angiogenesis is a process critical to both tumour growth and metastasis. It is a dynamic integrated process involving basement membrane degradation, endothelial cell proliferation and migration, and capillary tubule formation. Under normal circumstances, the microvasculature is maintained in a quiescent state. The acquisition of the angiogenic phenotype depends on the outcome of stimulatory and inhibitory regulation by the tumour and its microenvironment. There are markers of angiogenesis that potentially could provide prognostic information in addition to that gained from conventional clinicopathologic data, and antiangiogenic therapy for urologic cancers has potential advantages over current therapeutic strategies. Promising preclinical studies have led to the initiation of phase I studies of antiangiogenic therapy in combination with chemotherapy, which may lead to novel treatments for urologic malignant tumours and may identify new intermediate markers for the response to therapy.

Advances in angiogenesis research: relevance to urological oncology.

PURPOSE: Important advances in angiogenesis research are reviewed along with recent data implicating angiogenesis in the pathogenesis of urological malignancies. MATERIALS AND METHODS: The current understanding of angiogenesis and its importance in tumor biology is summarized. The rationale for anti-angiogenic therapy is reviewed and the clinical experience with these agents is discussed. An extensive literature search of angiogenesis in urological malignancies was performed. RESULTS: Quantitative immunohistochemistry for endothelial antigens suggests that, as is the case with many other tumors, induction of angiogenesis contributes to the malignant phenotype of prostate and bladder carcinomas. Anti-angiogenic agents have demonstrated efficacy against urological tumors in experimental systems, and recent data suggest that these agents may also be useful for chemoprophylactic purposes. Putative angiogenesis inducers specific for each of the major urological malignancies have been identified. Quantitation of the expression of angiogenesis inducers and estimation of microvessel density have demonstrated prognostic value for urological malignancies. CONCLUSIONS: The available data indicate that angiogenesis has an important role in the progression and metastasis of urological malignancies. Preclinical data coupled with experience in other cancers indicate that combining anti-angiogenic therapy with conventional treatment modalities has the potential to improve dramatically our management of these malignancies. Further research will be needed to define the mechanisms controlling angiogenesis in urological malignancies and to determine if any of the angiogenic correlates will be of genuine clinical use. The rapid pace of research in this field suggests that this aspect of tumor biology will soon have an increasingly important role in the evaluation and treatment of urological cancers.

Expression of platelet-derived growth-factor B-chain mRNA and tumor angiogenesis in invasive transitional cell carcinoma of the upper urinary tract.

In several types of carcinoma, angiogenesis is associated with metastasis and might be a prognostic indicator. Platelet-derived growth factor (PDGF) is one of several growth factors known; it is involved in the regulation of cell migration and proliferation. There have been few reports, however, dealing with the expression of PDGF B-chain mRNA and angiogenesis in transitional cell carcinoma. We evaluated both the expression of PDGF B-chain mRNA and angiogenesis in transitional cell carcinoma of the upper urinary tract and examined their interrelation with metastasis, patient prognosis, and other established clinicopathologic parameters. For this purpose, formalin-fixed, paraffin-embedded tumor tissues from 91 patients with invasive tumors were analyzed with in situ hybridization for PDGF B-chain mRNA and immunohistochemical staining of Factor VIII-related antigen to assess angiogenesis. The expression of PDGF B-chain mRNA was positive in 59 (66%) of the 91 patients. Microvessel density was significantly increased in positive PDGF B-chain mRNA cases (P < .0001). There was no significance, however, in any relationship between the expression of PDGF B-chain mRNA or between the microvessel density and the clinicopathologic findings, metastasis, or prognosis. Assessment of the expression of PDGF B-chain mRNA and microvessel density seems to be of no real value in predicting either the risk of metastasis or the prognosis in transitional cell carcinoma of the upper urinary tract.