Analysis of the Efficacy of a Sublingual Bacterial Vaccine in the Prophylaxis of Recurrent Urinary Tract Infection.

OBJECTIVES: Recurrent urinary tract infections (R-UTIs) are very common amongst women, and alternatives to antibacterial prophylaxis are necessary. This study evaluates the effectiveness of a sublingual bacterial vaccine for the prophylaxis of R-UTIs. METHODS: We conducted a quasi-experimental pretest-posttest study of 166 women diagnosed with R-UTIs. Both before and after the start of treatment with the vaccine, we analysed the total number of R-UTI episodes, the urine culture results, and the type and number of antibiotic packages consumed. Symptoms and urine cultures were evaluated 3, 6, 9, 12, 18, and 24 months after initiating treatment with the vaccine. RESULTS: The mean time of follow-up after vaccination was 1.7 years. After vaccination, there was a 54.6% reduction in episodes of UTI, and a 56.2% reduction in positive urine cultures. At 3 months, 74.4% of the patients had no R-UTI, the rate falling to 68.1% at 6 months, 52.4% at 12 months, and 44.5% at 24 months. The cumulative probability of maintaining negative urine cultures was 76% at 3 months, 37% at 12 months, and 18% at 2 years. CONCLUSIONS: The use of a sublingual bacterial vaccine for the prophylaxis of R-UTIs in women is an effective treatment that contributes to a reduction in the number of UTI episodes.

Role of CYP1B1 gene polymorphisms in bladder cancer susceptibility.

PURPOSE: CYP1B1 activates procarcinogens in some human tissues, including the urinary tract. Changes related to genetic polymorphisms are a known risk factor for cancer. We analyzed the association between CYP1B1 sequence variations and bladder cancer. MATERIALS AND METHODS: Sequence variations in the coding region (exons 2 and 3) and the neighboring introns of CYP1B1 were analyzed by direct polymerase chain reaction and DNA sequencing in 208 unrelated patients with bladder cancer and 208 healthy controls. RESULTS: We identified 6 known single nucleotide polymorphisms organized into 2 linkage disequilibrium blocks. The Ala/Ala and Leu/Val genotypes of the Ala119Ser and Leu432Val polymorphisms were significantly more common in patients than in controls (55.3% vs 42.8% and 54.8% vs 42.3%, respectively). The strongest individual single nucleotide polymorphism risk was found under an over dominant model for Leu432Val (OR 1.65, CI 95% 1.12-2.44). The 2 susceptibility single nucleotide polymorphisms were predicted to be structured into 4 haplotypes and more than 10 diplotypes. No individual haplotype was associated with bladder cancer but the diplotype Ala-Leu/Ala-Val was significantly overrepresented in cases compared to controls (31.73% vs 17.31%, OR 2.22, 95% CI 1.36-3.62, p = 0.001). The OR was approximately 1.6 for the individual genotypes Ala/Ala and Leu/Val, which increased to 2.2 for the Ala-Leu/Ala-Val diplotype. A risk occupation had a modifying effect, increasing the crude OR of the combined genotype Ala/Ala + Leu/Val from 2.2 to 8.3. CONCLUSIONS: This study provides strong evidence for the role of common CYP1B1 variants as risk factors for bladder cancer, which increases with occupational exposure.

Carbonic anhydrase IX as a specific biomarker for clear cell renal cell carcinoma: comparative study of Western blot and immunohistochemistry and implications for diagnosis.

OBJECTIVE: This study aimed to evaluate the usefulness of carbonic anhydrase IX (CA-IX) expression in clear cell renal cell carcinoma (CCRCC) using two different techniques to detect protein expression. MATERIAL AND METHODS: An experimental, cross-sectional, analytical study was conducted to analyse proteins in renal tumour and healthy tissue specimens from 38 consecutive patients who underwent nephrectomy for renal cancer. CA-IX protein expression was measured by immunohistochemistry and Western blot analysis and quantified. Statistical analysis was performed with the positive and negative specific agreements and kappa coefficient. The sensitivity and specificity of both techniques were assessed. Statistical tests were conducted to analyse the association between CA-IX expression quantitation and normal prognosis factors (TNM stage and Fuhrman nuclear grade), only in CCRCC. RESULTS: The mean patient age was 65 years, 78.9% of patients were men and 57.9% of tumours were CCRCC. CA-IX protein expression was positive in 63.2% of tumours by immunohistochemistry and in 60.5% by Western blot. Both techniques detected CA-IX expression only in CCRCC and unclassifiable tumours. High concordance indices were observed for CCRCC diagnosis. Western blot and immunohistochemistry had a sensitivity of 95.5% and 100%, respectively; the specificity was 100% in both techniques. CA-IX expression quantitation did not correlate with tumour stage or Fuhrman nuclear grade. CONCLUSIONS: Immunochemistry and Western blot techniques can be used to detect abnormal CA-IX protein expression in CCRCC and to support morphology-based diagnostic techniques.

Supernumerary testicle: case report and bibliographic review.

OBJECTIVE: We report a case of supernumerary testis, a rare anomaly with only around 100 cases reported in the literature. METHODS: We describe the case of a 26-year-old man who consulted for a left paratesticular tumor. Physical examination and ultrasound showed a 2-cm nodular lesion over the left epididymis. The lesion was confirmed as supernumerary testis by surgical examination and biopsy and was subsequently excised, given the malignancy potential. RESULTS: The supernumerary testis was evaluated using two standard classifications, one assessing function and embryological development, and the other assessing topography, anatomy, and reproductive potential. CONCLUSIONS: The differential diagnosis for an intrascrotal mass should include the possibility of a supernumerary testis; hence, surgical examination and biopsy are necessary. Supernumerary testes should be excised in the case of pain, dysplasia, or in situ carcinoma, or whenever the biopsy is inconclusive.

Neuroendocrine carcinoma of the urinary bladder. 15-year retrospective analysis.

OBJECTIVES: Neuroendocrine tumors (NET) are cancers found in the bronchopulmonary tract, where they were first described in 1926. The tumors are associated with poor prognosis due to their high metastatic potency even after radical treatments associated with other neo- or adjuvant therapies. NET of the urinary bladder is a rare tumor and accounts for 0.5% of bladder tumors. METHODS: The study was designed as an observational, descriptive and retrospective study of 13 patients diagnosed, treated, and followed for NET of the urinary bladder at the Hospital and University Complex of Albacete, Albacete, Spain between 1995 and 2010. RESULTS: The sample was composed of 11 men and 2 women. Mean patient age at the time of diagnosis was 71 (range, 57-88; SD, 6.98) years. T4 (6 patients) was the most common T tumor stage, followed by T2 (5 patients) and T3 (2 patients). In the case of N tumor stage, the most common was N2. In the remaining 3 patients, the degree of lymph node involvement could not be assessed. Six presented distant metastasis at the time of diagnosis. Eleven patients presented small-cell NET on histology. Seven underwent radical surgery (radical cystectomy). All other patients were treated by deep transurethral resection of bladder tumor, except for 1 patient treated by partial cystectomy. Adjuvant chemo-therapy (usually an association of carboplatin/cisplatin and etoposide) was administered to 4 patients. Of these 4 patients, 2 were also treated by pelvic radiotherapy. Two patients survived more than 5 years following diagnosis. In fact, at the time the study data was collected only 4 patients had survived and 2 presented tumor recurrence. Of the 9 deaths, 8 occurred within 6 months of diagnosis and 1 at 24 months. All of them were the result of the disease itself. CONCLUSIONS: Neuroendocrine tumor of the urinary bladder is a rare, aggressive tumor with high metastatic potential that should be considered in the differential diagnosis of urinary bladder neoplasms. Despite various multimodality treatments have been used prognosis is poor.

[Biomarkers in renal cancer.] / Biomarcadores en cáncer renal.

The natural history of renal cell canceris unpredictable and despite the increased knowledgeof this disease, the incidence has been increasing in recent years. Renal cancer represents a tumor withsignificant mortality and efforts to understand its behaviorhavenot yet translated into a decrease in mortality.In the study of renal cell cancer, the knowledge ofmolecular pathways is very important, since they arethe basis for the development of new therapies. Thisknowledge has made it possible to reclassify these tumorsand the current challenge is the search for biomarkersthat allow to establish an adequate diagnosisand prognosis and predict the response to a certaintype of treatment.In the present manuscript we carry out a review ofthe main markers studied and their potential value inrenal cell cancer.

La historia natural del cáncer renal esimpredecible y a pesar del aumento del conocimientode esta enfermedad, la incidencia ha ido en aumentoen los últimos años. Representa un tumor con unamortalidad importante y los esfuerzos por conocer sucomportamiento todavía no se traducen en una disminuciónde su mortalidad.Dentro del estudio del cáncer renal es cada vezmás importante el conocimiento de su funcionamientomolecular, que por otra parte ha sido fundamentalpara la aparición de las nuevas terapias. Dicho conocimientoha permitido reclasificar estos tumores, siendoel desafío actual la búsqueda de biomarcadores quenos permitan establecer un adecuado diagnóstico ypronóstico y predecir la respuesta a un determinadotipo de tratamiento.En el presente trabajo realizamos una revisión delos principales marcadores estudiados y su potencialvalor en el cáncer renal.

Prognostic Value of the Neutrophil/Lymphocyte Ratio in Squamous Cell Carcinoma of the Penis.

INTRODUCTION: To determine the predictive role of the neutrophil/lymphocyte ratio (NLR) in the prognosis and survival of patients with squamous cell carcinoma of the penis. MATERIALS AND METHODS: A retrospective cohort study of 96 patients with squamous cell carcinoma of the penis (SCCP). Clinical and histological data, bloodwork and disease evolution information were collected. We determined neutrophil/lymphocyte ratios and analyzed their relationship to prognosis and survival. RESULTS: The mean age of patients was 72,1 years. The average follow-up time for the sample was 3,8 years (CI 95%: 3,0-4,6). Compared to patients with NLR <3, those with NLR >3 presented a higher proportion of cancers in stages cN1 (29.7% vs 6.8%; p = 0,004), stages pT3 and pT4 (24,3% vs 9,5%; p = 0,05), and stages TNM III and IV (32,4% vs 10,2%; p = 0,01); additionally, there were more cases of perineural invasion in the NLR >3 patients (29,7% vs 10,2%; p = 0,03). The mean overall survival (OS) was 7,9 years (CI 95%: 6,2-9,6) and the cancer-specific survival (CSS), 1,3 years (CI 95%: 0,7-1,9). There were no differences in OS, CSS or in progression-free survival (PFS) in patients with NLR >3 compared to those with NLR <3 . However, in the Cox regression analysis, a higher NLR was independently associated (along with metastasis and need for adjuvant treatment) with lower PFS, with an HR: 1,27 (CI 95%: 1,02-1,57; p = 0,02). CONCLUSIONS: The utilization of NLR in clinical practice can be considered an additional tool to aid in the diagnosis and prognosis of patients with squamous cell carcinoma of the penis.

Polymorphic deletions of the GSTT1 and GSTM1 genes and susceptibility to bladder cancer.

UNLABELLED: What's known on the subject? and What does the study add? Bladder cancer susceptibility may be determined by genetic differences in the activity of glutathione S-transferases, enzymes that regulate the conversion of exogenous carcinogens to excretable hydrophilic metabolites by glutathione conjugation. The discrepancy of results regarding the association of common genetic polymorphisms and complex diseases such as cancer has raised scepticism in this area of research. Although the evidence generally supports the implication of GSTM1 and GSTT1 polymorphisms in bladder cancer, there is still some debate, with some studies in favour and some against. This study shows a greater risk of bladder cancer in individuals with GSTM1 null genotype, particularly women. This relationship is less evident with GSTT1 null genotypes. Null genotypes in both genes appear to be synergistic, particularly among smokers, and to increase the predisposition to more aggressive tumours. Nevertheless, the role of GSTM1 and GSTT1 polymorphisms in predisposition to bladder cancer should be viewed with caution, due to the multifactorial genetic origin of this condition and the need for long-term longitudinal studies to confirm these results. OBJECTIVE: To estimate the prevalence and importance of GSTT1 and GSTM1 genotypes (implicated in glutathione S-transferase activity) in bladder cancer, to determine whether smoking and occupational factors influence this relationship, and to identify the value of GSTT1 and GSTM1 genotypes as prognostic factors. PATIENTS AND METHODS: A cross-sectional study was conducted with a group of patients with bladder carcinoma and a control group with benign conditions and no history of tumours. The controls were selected and paired as subjects were recruited. Sociodemographic variables, smoking, professional occupation, histological features and the presence of GSTT1 and GSTM1 polymorphisms by multiplex PCR techniques were assessed. RESULTS: GSTM1 genotypes were investigated in 201 patients and 193 controls and GSTT1 genotypes in 190 patients and 163 controls. In the patients group, GSTT1 null genotype was observed in 22.1% (not significant) and GSTM1 null genotype in 54.2% (P=0.008) (odds ratio, OR, 1.7); when considered together, 15.5% (P<0.05; OR, 3.5) of patients had both null genotypes. In the multivariate analysis, the presence of GSTM1 null genotype remained in the model (OR, 2.1) in addition to smoking and age. Subjects with bladder tumour and GSTM1 null genotype were younger than patients without gene deletion (P=0.049). Women with GSTM1 null genotype presented a higher OR than men (P=0.024). When stratified by smoking habit, smokers with both null genotypes showed an OR of 4.7. The percentage of patients with G3 tumours was higher in patients with GSTT1 null genotype (P=0.013) and in patients with both null genotypes (P=0.002). A higher percentage of infiltrating tumours was also observed in patients with both null genotypes (P=0.035). CONCLUSIONS: The data obtained in the present study suggest a higher risk of bladder cancer in individuals with the GSTM1 null genotype. This risk is twofold higher when GSTM1 and GSTT1 null genotypes are both present and is also higher in smokers. A greater predisposition for more aggressive tumours appears to exist, particularly when both null genotypes are combined. Longer-term longitudinal studies are needed to confirm these results.

Clinical value of perioperative levels of DNA and mRNA in plasma of patients with renal cell carcinoma.

INTRODUCTION: The current challenge on renal cell carcinoma (RCC) is to finding a non-invasive biomarker for improving their diagnostic and therapeutic management. In the present study, we analyzed the clinical value of plasma levels of cell-free DNA (cfDNA) and RNA (cfRNA) of two genes: glyceraldehyde 3-phosphate-dehydrogenase (GAPDH) and human telomerase reverse transcriptase (hTERT). MATERIALS AND METHODS: We recruited 82 patients with RCC, and 20 healthy subjects. Using RT-PCR techniques, plasma levels of cfDNA and cfRNA from hTERT and GAPDH genes were quantified pre- and post-operatively, and one year after surgery. Relationships between such plasma levels and clinicopathological features and evolution of disease were analyzed. RESULTS: Levels of GAPDH cfDNA and cfRNA were significantly higher in patients than in healthy subjects. hTERT cfDNA was detected in plasma from 35% of RCC patients and in none healthy subject. At diagnosis, plasma levels of GAPDH cfDNA were higher in advanced pT and TNM stages, and hTERT cfDNA in patients with 3-4 Fuhrman grade and affected lymph nodes. Levels of cfNAs were not related to the presence of metastasis. Following nephrectomy, GAPDH cfDNA levels dropped, and patients with higher levels before and after nephrectomy, showed lower overall survival (OS). However, Cox's multivariate model did not prove any association of the cfNA levels with progression. CONCLUSION: Plasma levels of cfDNA from GADPH and hTERT genes were correlated to tumor diagnosis and progression and, thus, such analyses might help to diagnosis and prognosis of RCC patients.

Diagnostic and prognostic value of the detection of hTERT mRNA in renal tumors.

INTRODUCTION: Elevated mRNA expression of human telomerase reverse transcriptase (hTERT mRNA) is common in many types of tumors, participating in tumor growth and progression. Such expression has not been sufficiently examined in renal cancer. The goal of the present study was to quantify it and analyze its possible clinical value in the management of this pathology. PATIENTS AND METHODS: The study included 111 patients who underwent surgery for renal cell carcinoma (RCC) between 2015 and 2017. Tumor samples were taken from all patients and, in 94 of them, healthy renal tissue adjacent to the tumor was also sampled. The 2 types of tissue were histologically confirmed, after which mRNA was extracted. Using real-time quantitative PCR, the expression of hTERT and glyceraldehyde-3-phosphate dehydrogenase (as endogenous control) were indirectly quantified using the crossing point (CP), which is inversely correlated with the number of sample replicates yielding positive results. These values were correlated with patient socio-demographic variables and clinical-pathological factors of the RCC. RESULTS: The majority of patients were males, with an average age of 60.5 years (SD: 14.02). Most tumors (69.4%) were clear cell carcinomas. The most frequent stages were pT2 or lower (73%), while 5% were pN1 and 12% pM1. The majority of tumors (58%) were Fuhrman grades 1 or 2 (low grade). All samples of tumor and nontumor tissue expressed glyceraldehyde-3-phosphate dehydrogenase mRNA, with the CP in the tumor sample significantly lower than in the nontumor tissue (P < 0.001). The expression of hTERT mRNA was detected in 68% of tumor tissues and significantly correlated with histopathology: 100% in sarcomatoid RCC and 77.9% in clear cell carcinomas (P < 0.0001). The CP was lower in pN1 (P = 0.018), pM1 (P = 0.046), and TNM IV stages (P = 0,041). A greater number of hTERT mRNA replicas were detected in M1 patients (P = 0.0005) and TNM IV stage (P = 0.017). There was no correlation of hTERT mRNA expression with Fuhrman grade. CONCLUSIONS: The quantitation of hTERT mRNA expression in RCC might be useful as a complementary diagnostic tool as well as for assessing aggressiveness of the tumor.

Increased Serine and One-Carbon Pathway Metabolism by PKCλ/ι Deficiency Promotes Neuroendocrine Prostate Cancer.

Increasingly effective therapies targeting the androgen receptor have paradoxically promoted the incidence of neuroendocrine prostate cancer (NEPC), the most lethal subtype of castration-resistant prostate cancer (PCa), for which there is no effective therapy. Here we report that protein kinase C (PKC)λ/ι is downregulated in de novo and during therapy-induced NEPC, which results in the upregulation of serine biosynthesis through an mTORC1/ATF4-driven pathway. This metabolic reprogramming supports cell proliferation and increases intracellular S-adenosyl methionine (SAM) levels to feed epigenetic changes that favor the development of NEPC characteristics. Altogether, we have uncovered a metabolic vulnerability triggered by PKCλ/ι deficiency in NEPC, which offers potentially actionable targets to prevent therapy resistance in PCa.

HIF1α Suppresses Tumor Cell Proliferation through Inhibition of Aspartate Biosynthesis.

Cellular aspartate drives cancer cell proliferation, but signaling pathways that rewire aspartate biosynthesis to control cell growth remain largely unknown. Hypoxia-inducible factor-1α (HIF1α) can suppress tumor cell proliferation. Here, we discovered that HIF1α acts as a direct repressor of aspartate biosynthesis involving the suppression of several key aspartate-producing proteins, including cytosolic glutamic-oxaloacetic transaminase-1 (GOT1) and mitochondrial GOT2. Accordingly, HIF1α suppresses aspartate production from both glutamine oxidation as well as the glutamine reductive pathway. Strikingly, the addition of aspartate to the culture medium is sufficient to relieve HIF1α-dependent repression of tumor cell proliferation. Furthermore, these key aspartate-producing players are specifically repressed in VHL-deficient human renal carcinomas, a paradigmatic tumor type in which HIF1α acts as a tumor suppressor, highlighting the in vivo relevance of these findings. In conclusion, we show that HIF1α inhibits cytosolic and mitochondrial aspartate biosynthesis and that this mechanism is the molecular basis for HIF1α tumor suppressor activity.

Implications of mismatch repair genes hMLH1 and hMSH2 in patients with sporadic renal cell carcinoma.

OBJECTIVES: To analyse the implications of DNA mismatch repair genes hMLH1 and hMSH2 in sporadic renal cell carcinoma (RCC). MATERIALS AND METHODS: Specimens of tumour and healthy renal tissue were collected from 89 patients treated for sporadic RCC. Another 95 blood samples taken from individuals with no history of cancer were also analysed. After DNA extraction and PCR amplification, microsatellite instability (MSI) was determined using the Bethesda microsatellite panel, two exonic microsatellites of the TGFbRII and BAX genes, and the microsatellite D3S1611. The promoter methylation status of hMLH1 was investigated using the HpaII and MspI restriction enzymes. In addition, a sequencing analysis of complete coding region of hMLH1 and hMSH2 genes was performed. RESULTS: MSI and promoter hypermethylation of hMLH1 were not detected. Interestingly, loss of heterozygosity (LOH) was common among patients with RCC, particularly in microsatellite D3S1611 (34.9%). Mutations were identified in eight patients: K618A and V716M in gene hMLH1; and I145V, G322D, and the novel mutation P349A, in gene hMSH2. The mutations also appeared in healthy renal tissue and therefore, were considered as germline DNA sequence variations. There were G322D and K618A changes in >1% of the healthy control subjects, suggesting that they are DNA polymorphisms. CONCLUSIONS: Our data show that loss of function of both hMLH1 and hMSH2 is not involved in sporadic RCC, either by promoter methylation or mutation in their exons. However, LOH indicated that chromosomal instability affecting large fragments of DNA was the main genetic alteration we detected associated with RCC.

Implications of p53 gene mutations on patient survival in transitional cell carcinoma of the bladder: a long-term study.

OBJECTIVE: To determine the prognostic value of p53 gene mutations and P53 overexpression for predicting the incidence of recurrence, progression and long-term survival of patients with transitional cell carcinoma (TCC) of the bladder. METHODS: Prospective cohort study with 94 consecutive patients diagnosed and treated for TCC. DNA was obtained from tumor tissue to perform PCR-SSCP of p53 exons 5-9, with automatic sequencing of any mutated samples. Immunohistochemistry using anti-human P53 monoclonal antibody was also performed. Survival was analyzed and the survival curves compared (Mantel-Haenszel). Lastly, a Cox proportional hazards model was constructed. RESULTS: Mutations were found in 46.8% of samples, with 61.8% in infiltrating tumors. Exon 8 was involved in 42.3%. P53 overexpression (cutoff > or =20%) was found in 52.1%. Mean follow-up was 44.1 months; 43.6% had died by the end of this period. Mean survival was lower in patients with exon 8 mutations (38.4 months), compared with patients without this exon mutated (P = 0.016). There were no differences in patient survival based on positive or negative immunohistochemistry (cutoff > or =20%), although survival was lower in patients with a percentage higher than 50% of antibody-stained cells (P = 0.02). In the Cox analysis, tumor stage, pM stage, and interaction between stage > or =pT2 and mutated p53 gene were independent risk factors, with a 6.13-fold risk of death in these patients (P = 0.019). The number of tumors, nuclear grade, pTa stage, and the interaction between GI degree and nonmutated p53 gene remained in the Cox model for superficial tumors, such that these patients had a lower risk of recurrence or progression (P = 0.008). CONCLUSIONS: Alterations in the p53 gene may be indicative of poorer prognosis and greater recurrence in patients with urothelial bladder tumor, in particular, the presence of mutations in exon 8 and a greater percentage of stained cells in the immunohistochemistry. Nevertheless, the classic prognostic factors (primarily, pTNM stage) should still be considered the most useful factors for follow-up of these patients.

Sensitivity and specificity of p53 protein detection by immunohistochemistry in patients with urothelial bladder carcinoma.

OBJECTIVE: To analyze the correlation between the genotypic and phenotypic patterns of p53 in patients with transitional cell carcinoma (TCC) of the urinary bladder. MATERIALS AND METHODS: Cross-sectional study of 73 patients diagnosed with TCC. DNA was obtained from the tumor tissue to perform polymerase chain reaction-single strand conformational polymorphism (PCR-SSCP) of exons 5-9 of the p53 gene, with automatic sequencing done on any mutated samples. Immunohistochemistry (IHC) was also performed using anti-human P53 monoclonal antibody, and the diagnostic performance of this test was analyzed by a ROC curve, using the presence of p53 mutations found by PCR-SSCP as 'gold standard'. RESULTS: The cutoff point for defining immunopositivity was 20%. IHC had a specificity of 62.9%, and a sensitivity of 65.8%. The highest sensitivity values appeared in G3 tumors (75%) and infiltrating tumors (71.4%), and the highest specificity values were observed in G1 (77.7%) and G2 tumors (90%) and superficial tumors (66.6%). Mutations in exon 8 gave a positive result most frequently (73.7%) and were considered most relevant in terms of altering P53 function (60.9%). False negatives were documented in 28.5% of infiltrating tumors, and false positives in 33.4% of superficial tumors. CONCLUSIONS: There is a moderate correlation between p53 mutations and P53 protein overexpression, with this stronger in high-grade, infiltrating tumors, in exon 8 mutations, and when the mutation induces relevant changes in the protein structure. Although IHC is useful in routine clinical practice, the classic prognostic factors should still be considered the most important in the follow-up of these patients.

Prognostic Value of the VHL, HIF-1α, and VEGF Signaling Pathway and Associated MAPK (ERK1/2 and ERK5) Pathways in Clear-Cell Renal Cell Carcinoma. A Long-Term Study.

BACKGROUND: The prognostic value of molecular markers in renal cell carcinoma has been investigated in several studies. Although their value is still not confirmed, various proteins are important. We describe the effect on long-term survival of the status of the von Hippel-Lindau (VHL) hypoxia-inducible factor 1-α (HIF1-α) signaling pathway as well as associated mitogen-activated protein kinase (extracellular signal-regulated kinase [ERK]1/2 and ERK5). PATIENTS AND METHODS: A prospective, longitudinal cohort study was conducted with 50 patients diagnosed with clear-cell renal cell carcinoma to analyze VHL mutations and hypermethylation as well as VHL, HIF1-α, vascular endothelial growth factor (VEGF), ERK1/2, and ERK5 protein expression. Overall survival (OS), disease-specific survival (DSS), and progression- or recurrence-free survival (PFS) were analyzed using the Kaplan-Meier method. Mantel-Haenszel was used for comparisons, and Cox proportional risk models were also constructed. RESULTS: Follow-up was 66.9 months. There were 23 (46.0%) deaths, of which 17 (73.9%) were caused by the tumor. Mean periods were 85.6 months for OS and 94.3 months for DSS. A total of 22 (44.0%) patients showed progression (PFS, 78.1 months). VHL expression (P = .045) and > 10% of HIF1-α expression (P = .034) were associated with greater OS. DSS was greater in patients without VHL methylation (P = .012), with > 10% HIF1-α expression (P = .037), or with ERK5 protein underexpression. Greater PFS was associated with absence of VHL methylation (P = .045), presence of VHL expression (P < .0001), HIF1-α expression > 10% (P = .04), and ERK5 protein underexpression (P = .011). The presence of VHL mutation and/or methylation and VEGF expression had no prognostic value. Fuhrman nuclear grade and Tumor, Node, Metastases (TNM) stage were the only variables that remained in the Cox model. CONCLUSION: The HIF1-α and ERK5 pathway has prognostic value. Patients with no VHL or HIF1-α expression and ERK5 overexpression had a worse course of disease. VHL or VEGF status had no prognostic value. Only TNM stage and Fuhrman nuclear grade remained in the Cox model and, therefore, are still essential in prognostic biomarker panels.

Implication of VHL, ERK5, and HIF-1alpha in clear cell renal cell carcinoma: Molecular basis.

OBJECTIVES: To determine the expression status of several proteins related to VHL gene function and its relationship with common clinicopathological parameters. MATERIAL AND METHODS: Observational, analytical, cross-sectional study with 50 patients diagnosed with clear cell renal cell carcinoma. The study analyzed VHL mutations and hypermethylation as well as protein expression of VHL, CA-IX, HIF-1alpha, VEGF, ERK1/2, and ERK5, relating them to clinical variables. A bivariate and multivariate descriptive logistical regression analysis was performed, using the presence of metastasis at diagnosis as dependent variable. RESULTS: The study identified 13 (26%) VHL mutations related to nuclear grade (P = 0.036). VHL hypermethylation was found in 20% of cases. VHL expression was associated with the presence of mutations (P = 0.013), and the absence of expression was associated with nuclear grade and the presence of metastasis (P<0.05). HIF-1alpha was negative in only 5 cases. Vascular endothelial growth factor (VEGF) was positive in 31 of 47 cases and was associated with Fuhrman nuclear grade, presence of metastasis, and stage (P<0.05). ERK5 expression was increased in 58% of cases and associated with the presence of metastasis and more advanced stages (P<0.05). In the logistic regression analysis, the only variable remaining in the model was VEGF expression (P = 0.014). CONCLUSIONS: VEGF has prognostic value in clear cell renal cell carcinoma, and ERK5 may be a new prognostic marker in this type of tumor owing to its relationship with metastasis and more advanced stages.

VHL promotes immune response against renal cell carcinoma via NF-κB-dependent regulation of VCAM-1.

Vascular cell adhesion molecule 1 (VCAM-1) is an adhesion molecule assigned to the activated endothelium mediating immune cells adhesion and extravasation. However, its expression in renal carcinomas inversely correlates with tumor malignancy. Our experiments in clear cell renal cell carcinoma (ccRCC) cell lines demonstrated that von Hippel Lindau (VHL) loss, hypoxia, or PHD (for prolyl hydroxylase domain-containing proteins) inactivation decreased VCAM-1 levels through a transcriptional mechanism that was independent of the hypoxia-inducible factor and dependent on the nuclear factor κB signaling pathway. Conversely, VHL expression leads to high VCAM-1 levels in ccRCC, which in turn leads to better outcomes, possibly by favoring antitumor immunity through VCAM-1 interaction with the α4ß1 integrin expressed in immune cells. Remarkably, in ccRCC human samples with VHL nonmissense mutations, we observed a negative correlation between VCAM-1 levels and ccRCC stage, microvascular invasion, and symptom presentation, pointing out the clinical value of VCAM-1 levels as a marker of ccRCC progression.

[Promoter hypermethylation status of the mismatch repair gene hMLH1 in patients with sporadic renal cell carcinoma]. / Hipermetilación del promotor del gen reparador hMLH1 en la patogenia del carcinoma de células renales esporádico.

BACKGROUND AND OBJECTIVE: Epigenetic inactivation is a gene function abnormality that produces no changes in the DNA sequence, with the most frequent epigenetic alteration being hypermethylation of CpG islands in the promoter regions of the genes. Based on recent indications of a potential relationship between mismatch repair genes and renal cell carcinoma (RCC), we were interested in investigating the existence of promoter hypermethylation of the hMLH1 gene in tumor DNA samples from patients with sporadic RCC. MATERIAL AND METHOD: Sixty-five tumor tissue specimens were collected consecutively. The DNA was first obtained and purified, then digested with the restriction enzymes Hpa II and Msp I, followed by polimerase chain reaction amplification of 3 promoter regions of the hMLH1 gene, agarose gel electrophoresis, and densitometric analysis of the images of the amplified bands. RESULTS: Mean patient age was 63.7 years. The most frequent cell type was clear cell carcinoma (67.7%). 73.9% of tumors were diagnosed in stages below pT2, 9.3% had gland involvement and 20%, distant metastasis. No somatic hypermethylation was detected in the promoter region of the hMLH1 gene in any of the patients studied. CONCLUSIONS: Our data indicate that promoter hypermethylation of the hMLH1 gene is not implicated in the pathogenesis of sporadic RCC, and therefore the existence of another type of mutation, microsatellite instability and/or loss of heterozygosity should be examined to determine the possible role of this gene in sporadic RCC.