Molecular Protein and Expression Profile in the Primary Tumors of Clear Cell Renal Carcinoma and Metastases

Abstract

Metastasis involves the spread of cancer cells from the primary tumor to surrounding tissues and distant organs and is the primary cause of cancer morbidity and mortality. The aim of the study was the determination of change in molecular factors expression in primary kidney cancers (ccRCC) and metastatic sites. In total, 62 patients with RCC were enrolled in the study. The mRNA levels of molecular markers were studied by real-time PCR, and the content of the studied parameters was determined by Western blotting and ELISA. The features in the intracellular signal metabolites in the series of normal renal parenchyma, tumor tissue of localized, disseminated kidney cancer and metastatic tissue were studied. A decrease in some indicators in the tissue of the metastatic lesion was noted. Protein products of transcription factors HIF-1, CAIX, PTEN and activated AKT kinase, as well as expression of the VEGFR2 receptor and m-TOR protein kinase were revealed to be reduced in the metastatic sites. In addition, some indicators increased in metastasis: the protein levels of NF-κB p 50, NF-κB p 65, HIF-2, VEGF, VEGFR2, m-TOR and mRNA of HIF-1, CAIX, PTEN and PDK. There were indicators with multidirectional changes. HIF-1, CAIX, PTEN, VEGFR2 and m-TOR mRNA: VEGFR2, m-TOR, HIF-1, CAIX, PTEN and PDK had an opposite change in protein content and mRNA level. PTEN loss resulted in the downstream activation of AKT/mTOR signaling in secondary cancer lesions and determined the overall ccRCC patient's survival. The AKT/mTOR signaling cascade activation was found in the primary kidney tumors. The PTEN content and mRNA level were correlated with total AKT, GSK-3β, the 70S 6 kinases and AKT expression.

Keywords: AKT/mTOR components; PTEN; growth factors; kidney cancers; metastatic sites; primary tumors; transcription factors.

Figure 1

Scatterplot of PTEN and total AKT protein level in kidney cancers. Note: We found the associations between the protein level of PTEN with a total AKT (AKT pan) level in the primary tumors (r = 0.54; p = 0.0005), indicating the phosphatase influence on downstream of intracellular signaling.

Figure 2

Change in the profile of proteins and mRNAs of molecular factors in the primary tumor of the kidney and its metastatic sites. Note: Arrows indicate changes in the studied factors. We found a decrease in HIF-1, CAIX, PTEN, phospho-AKT content and RNA levels of VEGFR2 and m-TOR in metastases compared to the primary tumors. NF-κB p50, NF-κB p65, HIF-2, VEGF VEGFR2 and m-TOR protein contents and HIF-1, CAIX, PTEN and PDK RNA levels were enhanced in secondary cancer lesions. Indicators with change in opposite direction are highlighted in bold. Proteins level and mRNA alterations show the specific trend in biological features of primary tumors and metastases. Despite the reduction in mRNA levels of VEGFR2 and m-TOR, the protein contents of these molecular factors increased. The HIF-1 and PTEN protein levels showed the opposite trend. The kidney cancer aggressiveness is associated with the overactivation of AKT/mTOR signaling pathway and excess of tyrosine kinase receptors.