RESUMEN
BACKGROUND: Housekeeping genes are commonly used as endogenous reference genes for the relative quantification of target genes in gene expression studies. No conclusive systematic study comparing the suitability of different candidate reference genes in clear cell renal cell carcinoma has been published to date. To remedy this situation, 10 housekeeping genes for normalizing purposes of RT-PCR measurements already recommended in various studies were examined with regard to their usefulness as reference genes. RESULTS: The expression of the potential reference genes was examined in matched malignant and non-malignant tissue specimens from 25 patients with clear cell renal cell carcinoma. Quality assessment of isolated RNA performed with a 2100 Agilent Bioanalyzer showed a mean RNA integrity number of 8.7 for all samples. The between-run variations related to the crossing points of PCR reactions of a control material ranged from 0.17% to 0.38%. The expression of all genes did not depend on age, sex, and tumour stage. Except the genes TATA box binding protein (TBP) and peptidylprolyl isomerase A (PPIA), all genes showed significant differences in expression between malignant and non-malignant pairs. The expression stability of the candidate reference genes was additionally controlled using the software programs geNorm and NormFinder. TBP and PPIA were validated as suitable reference genes by normalizing the target gene ADAM9 using these two most stably expressed genes in comparison with up- and down-regulated housekeeping genes of the panel. CONCLUSION: Our study demonstrated the suitability of the two housekeeping genes PPIA and TBP as endogenous reference genes when comparing malignant tissue samples with adjacent normal tissue samples from clear cell renal cell carcinoma. Both genes are recommended as reference genes for relative gene quantification in gene profiling studies either as single gene or preferably in combination.
Asunto(s)
Carcinoma de Células Renales/genética , Regulación Neoplásica de la Expresión Génica/genética , Genes Relacionados con las Neoplasias/genética , Neoplasias Renales/genética , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa/métodos , Proteínas ADAM/genética , Proteínas ADAM/metabolismo , Adulto , Anciano , Anciano de 80 o más Años , Femenino , Perfilación de la Expresión Génica , Humanos , Masculino , Proteínas de la Membrana/genética , Proteínas de la Membrana/metabolismo , Persona de Mediana Edad , Estándares de ReferenciaRESUMEN
OBJECTIVES: Decreased expression of reversion-inducing cysteine-rich protein with Kazal motifs (RECK) was recently shown in several cancer types. To evaluate its potential role for prostate carcinoma, we investigated RECK expression in prostate cancer (pCA) samples. METHODS: RECK messenger RNA levels in 15 microdissected normal/tumor matches were determined by quantitative reverse transcriptase-polymerase chain reaction. Protein expression of RECK was evaluated by immunohistochemical staining in tissue samples of adenomectomies (n=24) and pCA samples after radical prostatectomy (n=247). RECK expression was related to preoperative prostate-specific antigen (PSA), tumor stage and grade, surgical margin status, and PSA relapse-free time after radical prostatectomy. RESULTS: Consistent with lower RECK messenger RNA by 24%, RECK protein expression was decreased in pCA, compared with adjacent normal tissue and prostatic intraepithelial neoplasia. RECK expression in samples of benign prostatic hyperplasia from adenomectomy specimens was higher than in normal adjacent tissue of prostate carcinomas. Decreased RECK expression was associated with higher Gleason score (> or =7) and higher tumor stage. Multivariate analysis using the Cox proportional hazards model revealed that negative RECK expression was an independent prognostic factor for an increased risk of PSA relapse, especially in patients with higher tumor grades (Gleason score > or =7). CONCLUSIONS: Decreased RECK expression correlating with the aggressiveness of pCA and the PSA relapse-free time could become an adjunct tissue biomarker to improve the follow-up and treatment decision for these pCA patients.
Asunto(s)
Glicoproteínas de Membrana/metabolismo , Antígeno Prostático Específico/sangre , Próstata/metabolismo , Prostatectomía , Neoplasias de la Próstata/patología , Anciano , Biomarcadores de Tumor/metabolismo , Calgranulina B , Proteínas Ligadas a GPI , Humanos , Inmunohistoquímica , Masculino , Glicoproteínas de Membrana/genética , Persona de Mediana Edad , Pronóstico , Modelos de Riesgos Proporcionales , Hiperplasia Prostática/metabolismo , Neoplasias de la Próstata/metabolismo , Neoplasias de la Próstata/cirugía , RecurrenciaRESUMEN
Allogeneic stem cell transplantation and donor lymphocyte infusions are currently under clinical investigation as an innovative therapeutic option for patients with metastatic renal cell carcinoma. A variety of trials have proven the clinical efficacy of allogeneic stem cell transplantation using reduced-intensity conditioning protocols and donor lymphocyte infusions, as demonstrated by the induction of objective remissions in metastatic renal cell carcinoma patients. However, despite clinical remissions, reduced-intensity conditioning protocols and donor lymphocyte infusions were associated with a high treatment-related mortality rate of approximately 17%. The disproportion between clinical efficacy and treatment-related mortality may mainly be caused by the selection of patients that had often been heavily pretreated, with a large tumor burden and rapidly progressing tumors. The improvement of efficacy with the preservation of a powerful graft-versus-tumor effect while reducing the toxicity, is the major experimental and clinical challenge of allogeneic stem cell transplantation in the treatment of metastatic renal cancer and other solid tumors. Recently, there has been a revolutionary development of molecular-targeted agents in metastatic renal cancer. These inhibitors of angiogenesis and signal-transduction pathways have demonstrated clinical efficacy and significant survival prolongation in the first- and second-line settings, while causing moderate toxicity. Some of these agents have already been approved by the US FDA and will probably replace standard cytokines, such as interferon-alpha2 and interleukin-2, in metastatic renal cancer. In the context of these innovative clinical developments, allogeneic stem cell transplantation clearly has to be regarded an investigational clinical treatment approach. Therefore, patients should only be treated at centers that are experienced in clinical trials, and patient selection remains a critical factor for a successful transplant procedure.
Asunto(s)
Carcinoma de Células Renales/secundario , Carcinoma de Células Renales/cirugía , Neoplasias Renales/cirugía , Trasplante de Células Madre , Carcinoma de Células Renales/patología , Enfermedad Injerto contra Huésped/inmunología , Enfermedad Injerto contra Huésped/patología , Humanos , Neoplasias Renales/inmunología , Neoplasias Renales/patología , Trasplante Homólogo/patologíaRESUMEN
PURPOSE: We assessed the diagnostic accuracy of bone markers in the serum of patients with renal cell carcinoma to detect bone metastases and evaluate the prognostic potential concerning renal cell carcinoma caused mortality. MATERIALS AND METHODS: The bone formation markers total and bone specific alkaline phosphatase, the bone resorption markers cross-linked N-terminal and tartrate-resistant acid phosphatase isoenzyme 5b, and the osteoclastogenesis markers osteoprotegerin and ligand of the receptor activator of nuclear factor-kappaB, were measured in the serum of 72 patients with renal cell carcinoma, including 28 with pN0M0, 8 with pN1M0 and 36 with M1, and in 32 female and 36 male controls by enzyme-linked immunosorbent assay techniques. Data were evaluated by receiver operating characteristics and survival analysis. RESULTS: Bone specific alkaline phosphatase, tartrate-resistant acid phosphatase isoenzyme 5b and ligand of the receptor activator of nuclear factor-kappaB did not significantly differ between patients with renal cell carcinoma and controls. Compared with controls tartrate-resistant acid phosphatase isoenzyme 5b, cross-linked N-terminal and osteoprotegerin showed increased concentrations in patients with nonbone metastases but not in those with bone metastases. No bone turnover marker led to differentiation between patients with nonbone and bone metastases. Increased osteoprotegerin above the upper 95% cutoff limit, tumor stage and distant metastatic spread were associated with renal cell carcinoma related survival on Kaplan-Meier analyses. A multivariate Cox proportional hazards regression model revealed that these 3 variables were independent prognostic factors for cancer related death. CONCLUSIONS: Bone turnover markers are hardly useful to diagnose bone metastases in patients with renal cell carcinoma. However, osteoprotegerin together with clinicopathological characteristics may be helpful as prognosticator of cancer specific death.