>Magister Scientiae - MSc / Kidney cancers, of which clear cell renal cell carcinoma comprises an estimated 70%, have been placed amongst the top ten most common cancers in both males and females. With a mortality rate that exceeds 40%, kidney cancer is considered the most lethal cancer of the genitourinary system. Despite advances in its treatment, the mortality- and incidence rates across all stages of the disease have continued to climb. Since the release of the Human Genome Project in the early 2000’s, most genetics studies have focused on the protein coding region of the human genome, which accounts for a mere 2% of the entire genome. It has been suggested that diverting our focus to the other 98% of the genome, which was previously dismissed as non-functional “junk DNA”, could possibly contribute significantly to our understanding of the underlying mechanisms of complex diseases.In this study a whole genome sequencing somatic mutation data set from the International Cancer Genome Consortium was used. The non-coding somatic mutations within the promoter, intronic, 5-prime untranslated and 3-prime untranslated regions of clear cell renal cell carcinoma-implicated genes were extracted and submitted to RegulomDB for their functional annotation.As expected, most of the variants were located within the intronic regions and only a small subset of identified variants was predicted to be deleterious. Although the variants all belonged to a selected subset of kidney cancer-associated genes, the genes frequently mutated in the non-coding regions were not the same genes that were frequently mutated in the whole exome studies (where the focus is on the coding sequences). This indicates that with whole genome sequencing studies a new set of genes/variants previously unassociated with the clear cell renal cell carcinoma could be identified. In addition, most of the non-coding somatic variants fell within multiple transcriptions factor binding sites. Since many of these variants were also deleterious (as predicted by RegulomDB), this suggests that mutations in the non-coding regions could contribute to disease due to their role in transcription factor binding site disruptions and their subsequent impact on transcriptional regulation. The substantial overlap between the genes with the most aberrantly methylated variants and the genes with the most transcription factor binding site disruptions signifies a potential link between differential methylation and transcription factor binding site affinities. In contrast to the upregulated DNA methylation generally seen in promoter methylation studies, all of the significant hits in this study were hypomethylated, with the subsequent up-regulation of the genes of interest, suggesting that in the clear cell renal cell carcinoma, aberrant methylation may play a role in activating proto-oncogenes, rather than the silencing of genes. When a cross-analysis was carried out between the gene expression patterns and the transcription factor binding site disruptions, the non-coding somatic variants and differential methylation profiles, the genes affected again showed a clear overlap. Interestingly, most of the variants were not present in the 1000genomes data and thus represent novel mutations, which possibly occurred as a result of genomic instability. However, identifying novel variants are always promising, since they epitomise the possibility of developing pioneering ways to target diseases. The numerous detrimental effects a single non-coding mutation can have on other genomic processes have been demonstrated in this study and therefore validate the inclusion of non-coding regions of the genome in genetic studies in order to study complex multifactorial diseases. / National Research Foundation (NRF) and DAAD

Development of novel ill vitro and ill vivo models for determining primary events in HLRCC tumourigenesis Linda O'Flaherty, Mansfield College Thesis submitted for degree of Doctor of Philosophy Nuffield Department of Clinical Medicine, University of Oxford Hilary Term 2012 Germline mutations of fumarate hydratase (FR), encoding an enzyme of the tricarboxylic acid (TCA) cycle, predispose affected individuals to hereditary leiomyomatosis and renal cell cancer (HLRCC). FH-deficient cells and tissues have been shown to accumulate fumarate, exhibit S-(2-succinyl) cysteine (2SC) protein modifications and to constitutively express hypoxia-inducible factor alpha (HIF -1 a and -20.), under nonnoxic conditions. This thesis presents a phenotypic characterisation of FhI-I- mouse embryonic fibroblasts (MEFs), generated from previously reported conditional Fhl knockout mice, as a new in vitro system for investigating and identifying biochemical and metabolic pathways that are dysregulated as a result of FhI inactivation. These cell lines reproduced the aforementioned phenotypes, in addition to an observed shift from oxidative phosphorylation (OXPHOS) to glycolytic metabolism. Re-expression of either full length, mitochondrial-targeted FH (FhI-I- +FH) or cytoplasmic FH (Fhrl- +FHl'1MTS) in FhI-deficient MEFs was sufficient to reduce intracellular fumarate and to correct for the dysregulation of the Hif pathway. These results were of particular interest as they demonstrated that nonnoxic stabilisation of Hif-Ia occurs independently of the persistent mitochondrial defect observed in Fhrl- +FHl'1MTS MEFs. These findings were corroborated in vivo following the development of transgenic mouse models, ubiquitously expressing either FH or FHl'1MTS in mice with targeted inactivation of FhI in renal tubular cells. Surprisingly, the cytoplasmic-restricted FH (FHl'1MTS) transgene was just as efficient as the transgenic mice expressing mitochondrial- targeted FH at rescuing the cystic phenotype associated with Fh I-deficiency in the kidneys. As the function of cytoplasmic FH has remained poorly understood, these results go some way to extricating a role for this isofonn of FH. The results of this thesis demonstrate that these novel in vitro and in vivo models, used either alone or in combination, are a versatile and robust paradigm for studying altered cell metabolism in not only HLRCC but other diseases associated with metabolic dysregulation.

Le carcinome rénal représente 5% des tumeurs de l’adulte et se développe au niveau des tubules rénaux. Le sous-type histologique majeur des cancers du rein est le carcinome rénal à cellules claires (cRCC). 90% des cRCC présentent une inactivation biallélique du gène suppresseur de tumeur de Von Hippel Lindau (VHL) induisant une activation constitutive de la voie de l’hypoxie via le facteur de transcription HIF1-α (Hypoxia Inducible Factor) qui contribue à la physiologie des tumeurs. Les cRCC sont des tumeurs à la fois radio- et chimiorésistantes, rendant la prise en charge thérapeutique des patients très difficile.Nos recherches consistaient en l’étude des rôles de la mucine membranaire MUC1, dont la queue cytoplasmique (MUC1 CT) peut interagir avec différentes voies de signalisation et agir en tant que co-activateur transcriptionnel de nombreux gènes impliqués dans la progression tumorale et la diffusion métastatique. Des travaux antérieurs réalisés au laboratoire montraient que la surexpression de MUC1 observée dans les cRCC était associée au statut métastatique des patients et marquait un mauvais pronostic. Cette surexpression de MUC1 est également impliquée dans la voie de l’hypoxie, voie majeure de la carcinogenèse rénale. Le premier objectif de l’étude était donc de déterminer les effets de la surexpression de MUC1 sur les propriétés des cellules de cRCC. Nous montrons ainsi que le domaine extracellulaire de MUC1 ainsi que sa partie cytoplasmique sont impliqués dans l’augmentation des capacités migratoires et de la viabilité des cellules cancéreuses rénales et qu’elle leur confère une résistance à l’anoïkis, programme de mort cellulaire déclenché lorsque la cellule perd ses contacts avec les cellules voisines ou avec la matrice extra-cellulaire et diminuent les propriétés d’agrégation des cellules tumorales. Nous montrons également que MUC1 est impliquée dans la chimiorésistance des cRCC en induisant l’expression de genes de chimiorésistance comme ABCG2 et GSTO2. Nous montrons par ailleurs que les propriétés invasives des cellules de cRCC sont exclusivement liées à MUC1 CT. Le deuxième objectif de l’étude était d’identifier les mécanismes moléculaires à l’origine du clivage de MUC1 CT. En utilisant différentes stratégies (siARN, inhibiteurs pharmacologiques et peptides), nous montrons pour la première fois que deux sheddases, ADAM10 et ADAM17 et la gamma secrétase sont nécessaires au clivage de MUC1 C, permettant ainsi sa délocalisation nucléaire et l’augmentation des propriétés invasives des cellules de cRCC. Enfin, nous montrons que la surexpression de MUC1 augmente l’expression protéique d’ADAM10/17, suggérant une boucle de régulation positive existant en conditions pathologiques.En conclusion, notre étude souligne le rôle de MUC1 dans la progression tumorale rénale et montre que la localisation nucléaire de MUC1-C est à l’origine de l’acquisition d’un phénotype invasif et chimiorésistant via l’action des sheddases ADAM10/17 et de la gamma secrétase. MUC1 apparait alors comme une cible thérapeutique potentielle intéressante dans la prise en charge des cRCC. / Renal cell carcinoma corresponds to 5% of all adult malignancies and originates from renal tubules. The main histologic subtype is represented by clear renal cell carcinoma. Ninety percent of cRCC present a biallelic inactivation of the von Hippel Lindau (VHL) tumor suppressor gene resulting in constitutive activation of hypoxia signaling pathway via the Hypoxia Inducible Factor (HIF) -1 transcription factor that contributes to the physiology of tumours. cRCC is typically highly resistant to conventional systemic therapies. MUC1 is a membrane-anchored mucin and its cytoplasmic tail (CT) can interact with many signaling pathways and act as a co-transcription factor to activate genes involved in tumor progression and metastasis. Previous studies have shown that MUC1 is diffusely overexpressed in cRCC and MUC1 overexpression has been found to be associated with metastatic disease and a worse prognosis.MUC1 is overexpressed in renal cell carcinoma with correlation to prognosis and has been implicated in the hypoxic pathway, the main renal carcinogenetic pathway. In this context, we assessed the effects of MUC1 overexpression on renal cancer cells properties. Using shRNA strategy and/or different MUC1 constructs, we found that MUC1-extracellular domain and MUC1 CT are both involved in increase of migration, cell viability, resistance to anoikis and to decrease of cell aggregation in cancer cells. We also showed that MUC1 is involved in cRCC chemoresistance by inducing chemoresistance genes expression like ABCG2 and GSTO2. Invasiveness depends only on MUC1 CT. Then, by using siRNA strategy and/or pharmacological inhibitors or peptides, we showed that sheddases ADAM10, ADAM17 and gamma-secretase are necessary for MUC1 C-terminal subunit (MUC1-C) nuclear location and in increase of invasion property. Finally, MUC1 overexpression increases ADAM10/17 protein expression suggesting a positive regulatory loop. In conclusion, we report that MUC1 acts in renal cancer progression and MUC1-C nuclear localization is driving invasiveness of renal cancer cells through a sheddase/gamma secretase dependent pathway. MUC1 appears as a therapeutic target by blocking MUC1 cleavage or nuclear translocation by using pharmacological approach and peptide strategies.

No description available.

The interplay between cancer cells and the extracellular matrix (ECM) remains a critical regulator of both normal tissue organization and cancer cell invasion. Proteins that function as ECM receptors function to link the cell with the ECM. Abberations in either the structure of the ECM or the expression of ECM receptors leads to disrupted interaction and downstream signaling effects. Dystroglyan (DG) is an ECM receptor that is expressed in a variety of tissue types and functions to mediate sarcolemma stability, epithelial polarity, and is critical in the early formation of basement membranes. However, DG has primarily been studied in muscle where loss of its function is linked to a host of muscular dystrophies. In the epithelium, the role of DG remains enigmatic. While DG has repeatedly been shown to lose function during cancer development and progression, the mechanism and functional consequence of its loss are currently unknown. In order to increase our understanding of DG in cancer development, we analyzed its expression and glycosylation, a functional requirement for DG, in a range of prostate cancer cell lines. Previous work has shown DG to be downregulated in prostate cancer, but the mechanism by which this occurs has remained largely unclear. We found that DG expression is maintained while its glycosylation was heterogeneous in the cell lines. Further investigation revealed that lines with hypoglycosylated DG strongly associated with the loss of expression of the glycosyltransferase LARGE2. Further this enzyme is frequently downregulated in human cancers and appears to serve as a required enzyme in DG glycosylation within prostate epithelium. This is the first work to demonstrate the functional requirement of LARGE2 for DG, and the only work to implicate loss-of-function of LARGE2 in cancer progression. To determine whether loss of LARGE2 is found in other tumor types, we analyzed human clear cell renal cell carcinoma (ccRCC) samples by immunohistochemistry and via in silico analysis with the Cancer Genome Atlas (TCGA). Our work demonstrated a frequent and significant downregulation of LARGE2 expression and its association with DG hypoglycosylation. Additionally, we found the loss of LARGE2 strongly associated with increased mortality. Thus, we again demonstrated a functional requirement of LARGE2 but also found a clinical correlate with increased mortality. Finally, we examined the functional outcome of DG hypoglycosylation or loss of expression in both a mouse model of prostate cancer and a variety of cell lines models. We found that while loss of DG expression does not increase prostate cancer growth or metastasis in one model of cancer, loss of its glycosylation does seem to mediate downstream metabolic changes within cells. The mechanism for this change remains unclear. In summary, these studies have contributed to our understanding of DG glycosylation and function in both prostate and renal carcinoma. Additionally, we have shown a novel mechanism by which DG glycosylation is lost with downregulation of LARGE2 expression. Finally, while we were unable to demonstrate a clear mechanism by which signaling changes arose, we were able to demonstrate a strong correlation between DG hypoglycosylation and increased mortality in ccRCC. These insights could be used to improve treatment of multiple cancer types as our understanding of DG function continues to improve.

Das NZK ist nach dem Prostata- und Harnblasenkarzinom der dritthäufigste urologische Tumor. Die Prognoseaussichten hängen beim NZK vom Metastasenstatus der Patienten ab. Die Heilungschancen für das lokal begrenzte NZK sind im Vergleich zum metastasierten NZK deutlich besser. Durch den Einsatz von TKI und mTOR-Inhibitoren wurde die Therapie des metastasierten NZK revolutioniert und das Überleben von Patienten signifikant verbessert. Nichtsdestotrotz profitiert ein Teil dieser Patienten aufgrund von Resistenzmechanismen nicht von solch einer anti-VEGF-Therapie. Bisher gibt es keine geeigneten Biomarker, die das Ansprechen auf eine solche Therapie vorhersagen könnten. Daher bestand das primäre Ziel dieser Arbeit darin, molekulare Marker für die Abschätzung der Prognose beim lokal begrenzten und metastasierten NZK sowie die Vorherage für das Ansprechen auf eine Sunitinib-Therapie beim metastasierten NZK zu identifizieren. Für das Prognosemodell am lokal begrenzten NZK konnten auf mRNA-Ebene signifikante Assoziationen der Marker HIF-2α, VEGFR3 und sVEGFR1 mit dem PFS, TSS und OS der Patienten identifiziert werden. Da bisher nur klinische Parameter für Prognosemodelle des lokal begrenzten NZK genutzt werden, könnten die hier untersuchten molekularen Marker nach einer unabhängigen Validierung zur Vorhersage der Prognose herangezogen werden. Patienten mit einem metastasierten NZK wiesen VHL-Mutationen (50%) und -Kopienzahlverluste (60 %) auf, die jedoch nicht mit dem Ansprechen auf Sunitinib assoziiert waren und nur geringfügige Auswirkungen auf die Proteinlevel von VHL und dessen Targetgene HIF-1α, CA9 und VEGFA zeigten. Bei den Untersuchungen zur VHL-Promotormethylierung wurde im tumorfreien Gewebe eine hohe Grundmethylierung festgestellt. Aufgrund der geringen Patientenzahl und der weitgehend unbekannten komplexen Methylierungsstruktur des VHL-Promotors konnten keine Assoziationen mit der Prognose und dem Ansprechen der metastasierten NZK-Patienten auf Sunitinib bestimmt werden. Für die Proteinlevel potenzieller prädiktiver Marker wie CA9, HIF-1α, VEGFR1 und -2, pVEGFR1, pPDGFRα und -β, CD31, pAkt sowie Ki67 wurden signifikante Assoziationen mit dem Ansprechen auf die Sunitinib-Behandlung beobachtet. Die CA9-Membranfärbung und das Ansprechen nach 9 Monaten wurden in der multivariaten Analyse als unabhängige prognostische Marker für das OS bei Patienten mit metastasiertem NZK identifiziert. In anderen Arbeiten wurde CA9 bereits mehrfach als potenzieller Biomarker beschrieben und könnte daher eine Anwendung in der Prognosevorhersage und Patienten-Selektion für eine Target-Therapie finden. Polymorphismen in Angiogenese-assoziierten Genen gelten ebenfalls als potenzielle Marker für das Ansprechen auf eine Therapie mit Sunitinib. Die Überlebensanalysen deckten signifikante Assoziationen zwischen dem VEGFA-SNP -2578 und dem PFS und für die SNPs VEGFR1 B sowie VEGFR2 +1191 mit dem OS auf. Das kombinierte Auftreten der Varianten-Allele der VEGFA-SNPs -2578, -1154 und +405 wirkte sich ebenfalls signifikant auf ein verlängertes OS der mit Sunitinib behandelten Patienten aus. Bei eindeutiger Bestätigung dieser Ergebnisse in prospektiven Studien könnten einfache SNP-Analysen an Blutproben die Therapieentscheidung und das Überleben der NZK-Patienten maßgeblich beeinflussen. Die künstlich erzeugte Sunitinib-Resistenz in den NZK-Zelllinien A498, Caki-1 und KTCTL-26 offenbarte nur begrenzt veränderte Proteinniveaus potenzieller Marker wie HIF-1α, Akt und pAkt zwischen den resistenten und sensitiven Zellen. Ein besseres Verständnis der molekularen Grundlagen der Resistenzentwicklung könnte zusammen mit einem wirkungsvollen, prädiktiven Biomarker für das Ansprechen die Therapie beim metastasierten NZK erheblich verbessern. In dieser Arbeit konnten letztlich verschiedenste Biomarker identifiziert und evaluiert sowie deren Bedeutung für die Prognosevorhersage und Prädiktion von Patienten mit metastasiertem NZK unter Sunitinib-Therapie herausgearbeitet werden. Diese Daten stellen damit einen weiteren Grundstein für mögliche prospektive klinische Studien dar, die den therapeutischen Nutzen der Biomarker eindeutiger definieren könnten.

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Objective: To develop a predictive model for preoperative differentiation between benign (B) and malignant (M) histology in patients with renal masses (RM) using recursive partitioning. Methods: We analyzed preoperative patient and tumour characteristics in 395 subjects who had surgery for RM suspicious for renal cell carcinoma. Results: The model predicted B vs. M histology with an overall accuracy of 89.6% (95% CI 86.2,92.5). It assigned patients with smaller tumours (<5.67cc) and a predominantly (>45%) exophytic component a high risk of B disease (52.6%). Patients with symptoms, larger tumours (>5.67cc) and larger endophytic component (>35%) have a 0% risk of B disease. Conclusion: B vs. M disease can be predicted accurately. This predictive accuracy is higher than that shown in renal biopsy series. It is hypothesized that for smaller and exophytic RMs, a biopsy is indicated. Symptomatic, larger and endophytic RMs should be removed without further investigations.

No description available.

ABSTRACT There is an increasing incidence of neoplasms in the kidney and a poor prognosis for patients who are diagnosed with advanced or metastatic kidney cancer. Renal cell carcinoma (RCC) constitutes the most prevalent form of kidney neoplasm in the adult population. Although surgery or cryoablation are successful curative treatments for localized RCC, improved diagnostic methods facilitating early detection and characterization of renal tumours may enable more effective use of less invasive treatments, especially for metastases. Currently there are no suitable tumour markers available for diagnostic, prognostic or predictive purposes. By increasing our knowledge of the underlying molecular characteristics of RCC, we may be able to identify molecular pathways involved in tumour growth and metastasis, and this knowledge may expedite the development of targeted therapies, and may identify useful markers of RCC development and progression. This thesis aimed to identify new genes involved in resistance to cancer therapy in RCC and to analyse their incidence and test their function in human RCC tissue, using immunohistochemistry in a large cohort of patients with RCC and paired normal tissue, and in vitro models. The lack of induction of apoptosis in RCCs by conventional cancer therapies such as chemotherapy, immunotherapy or radiation is central to their resistance to treatment. If apoptotic pathways that are activated in successful treatments were identified, they might be used for targets in future therapies. The hypothesis tested in this thesis was that genes or proteins involved in the molecular control of apoptosis, identified from RCC cell culture models and RNA microarray, will be useful for molecular profiling in human RCCs of different subtypes, as markers of those specific subtypes of the RCCs, as indicators of prognosis of the diagnosis, or as targets for future therapy regimens. The broad aims of this project were: To establish a model in which significantly increased apoptosis in RCCs in cell culture could be correlated with alterations in apoptotic pathway genes, to investigate the functional significance of some of those genes; and to maximise information gained from these basic experimental laboratory studies on new apoptosis genes in RCC development and progression by determining their expression patterns in tissue microarrays (TMAs) generated from different subtypes of human RCCs. The specific aims of the project were: 1) To establish an RCC cell culture model with high levels of induced apoptosis for RNA analyses using microarray to identified apoptotic genes that are novel in RCC investigations; 2) To describe the role and functional significance of some of the novel RCC apoptosis pathway genes using molecular investigations, including silencing RNA techniques; and 3) To use TMAs and immunohistochemistry to determine whether RCC subtypes can be distinguished by protein expression profiles of selected new apoptosis pathway genes. The thesis is presented as a literature review (Chapter 1), materials and methods (Chapter 2), four original research segments (Chapters 3 to 6), and finally a segment that summarises the results and presents future directions (Chapter 7). The first of the original research Chapters (Chapter 3) addressed a question “can apoptosis in RCCs be induced in cell culture to such a level that apoptotic pathways may be analysed?” Two RCC cell lines (ACHN and SN12K-1) were treated with IFN-a (500IU/mL), radiotherapy (20 Gy) or dual therapy of these two treatments. Apoptosis was quantified using microscopy and morphological characteristics and verified using enzymatic labelling of cells. The ACHN cell line, treated with the dual therapy and analysed at 24 hours, had a significantly increased level of apoptosis (p<0.05) compared with the non-therapy treated controls, and negligible mitosis. The increased expression or activation of at least some known apoptotic pathway genes (Bcl-2, Bax, caspase-3, 8 and 9, and p53) was verified in this model. The ACHN/dual therapy model was selected for further study. The second of the original research Chapters (Chapter 4) addressed the question “what apoptosis-regulating genes are significantly different in the apoptotic ACHN cells?” An RNA microarray assay (112 apoptosis-related genes) was carried out using RNA extracted from treated and non-treated ACHN cells and analysed for alterations of at least 1.9-fold in transcript levels. 21 genes had upregulated transcript levels in the treated cells, and one had down-regulated transcript levels. A search of the literature revealed three gene families with altered transcript levels in the treated RCCs that were novel: the TNF receptor-associated factor (TRAF), caspase recruitment domain (CARD) and cell death-inducing DFF-45 effector (CIDE) gene families. Representative members of these families were then investigated for protein expression alterations. The results for one particular gene, TRAF1, indicated it might be worthy of further study in modulation experiments (Chapter 6). The next research chapter (Chapter 5) asked the question “since the ACHN cells express TRAF-1 and have increased TRAF-1 with increasing apoptosis, what happens to the levels of mitosis and apoptosis in these cells when TRAF-1 expression is knocked down?” Silencing RNA (siRNA) techniques were used to knock down TRAF-1 in the ACHN RCC cell line using the same model as was described in Chapter 3. Successful knock down was gained after 72 hrs of transfection with a commercially-available siRNA against TRAF-1. These cells were then treated with the radiation, IFN-α or dual therapy treatments. In the cells with TRAF-1 knock down, there was significantly less apoptosis and more mitosis than was seen in the non-transfected cells. These results indicate that TRAF-1 does play a functional role in induction of apoptosis in RCCs and is worth investigating further in targeted therapies. The final of the research Chapters (Chapter 6) looked at the molecular distinctions among ccRCC, papillary, collecting duct, chromophobe and unclassified types of RCC. Molecules that might distinguish one from another would be valuable clinically, and it would be especially valuable if these molecules could also be targeted for therapeutic benefit. By knowing the action of these molecules in the apoptotic pathways used by RCCs when they do regress or die during cancer therapies, we might be able to devise new targeted therapies for RCCs. This research Chapter asked the question “could molecular profiling with an array of apoptotic pathway genes, novel to investigations in RCCs, provide information that would distinguish the subtypes?” TMAs prepared from 121 RCC and paired normal patient samples, where available, were investigated. Most RCC samples were ccRCC. Antibodies against selected members of the TRAF), CARD and CIDE gene families were selected from the RNA microarray data. These genes (TRAF1, TRAF3, TRAF4, inhibitor of caspase-activated DNAase/ICAD and nucleolar protein-3/NOL3) were analysed in TMAs using immunohistochemistry and digital scanning or bright field microscopy and graded scales of intensity and distribution, blinded to sub-type of RCC. After microscopy, scores for subtypes of RCC were compared with their normal tissue. Significant differences were found for TRAF1 and NOL3. The results indicate TRAF1 and NOL3 have potential for improving outcome or diagnosis in RCCs. In summary, there are no effective treatments against metastatic RCC, and no suitable grade or stage-defining biomarker for metastatic RCC subtypes. One of the main reasons for therapy resistance in RCCs is their inability to use or activate apoptotic pathway molecules. By investigating the reasons for RCC resistance to cancer therapies, we may be able to improve both diagnosis and treatment strategies. By defining the mechanisms and pathways of resistance to therapy-induced apoptosis, and developing methods to manipulate the pathways to cell death to defeat therapy resistance, we will have a better chance to develop successful markers of RCC subtypes and also design new and successful RCC therapies.