A common genetic feature signature of sporadic clear cell RCC is the loss of chromosome 3p, suggesting the presence of one or more RCC tumor suppressor genes at this site. The von Hippel-Lindau tumor suppressor gene (VHL), which resides on chromosome 3p25, is mutated or silenced in greater than 50% of sporadic clear cell RCCs.16,17 Germline VHL mutations give rise to von Hippel-Lindau syndrome, which is characterized by an increased risk of blood vessel tumors (hemangioblastomas), endocrine tumors and RCC. The VHL gene product, pVHL, is the substrate recognition module of an E3 ubiquitin ligase that targets the hypoxia-inducible factor (HIF) α transcription factors (HIF1α, HIF2α, and HIF3α) for destruction in the presence of oxygen. Hypoxic cells, or cells lacking pVHL (“pseudohypoxic”), accumulate high levels of HIF, which activates the transcription of a variety of genes, including vascular endothelial growth factor (VEGF), platelet-derived growth factor (PDGF-3) B, and transforming growth factor alpha (TGF-2). Restoration of pVHL function in VHL−/− mutant renal carcinoma cells suppresses their ability to form tumors experimentally by reducing HIFα levels.^18,19 Inhibition of HIFα is necessary and sufficient for tumor suppression by pVHL in RCC nude mouse xenograft assays. This provides a rationale for treating VHL−/− RCC with inhibitors of HIFα or its downstream targets. While the HIF1α isoform was initially believed to be more important, increasing literature supports HIF2α as the more important HIFα member in mediating tumorigenesis.^18,19 Although most investigation has focused on the role of HIFα isoforms, the pVHL protein also has several other targets in addition to HIFα, postulated by some to contribute to tumorigenesis. Elucidating these targets will lead to further knowledge of how pVHL suppresses tumor growth. Analysis of mutations in exon 3 of the VHL gene may be useful in refining the diagnostic criteria for conventional RCC versus chromophobe RCC with clear cell features.²⁰

A recent work from Dr. Rathmell’s group have generated gene expression microarray data using software that implements iterative unsupervised consensus clustering algorithms to identify the optimal molecular subclasses. A Consensus Cluster analysis identified two distinct subtypes of clear cell RCC within the training set, designated clear cell type A (ccA) and B (ccB). In each subtype the analysis of data defined a small, highly predictive gene set. A validation data set of 177 tumors was analyzed. Tumors designated ccA had improved disease-specific survival compared with ccB (median survival of 8.6 vs. 2.0 years). These preliminary data suggest that a cluster subtype classification is possible in RCC as performed in other diseases such as breast cancer. Prospective clinical studies will be necessary to validate these important findings.²¹

The genetic studies in familial RCC have led to the identification of specific molecular signatures in non–clear cell histotypes as well, such as Met hyperactivation in papillary type I, fumarate hydratase mutation in papillary type II, and c-Kit overexpression in chromophobe RCC (see “Papillary Types I and II Renal Cell Carcinoma”). Preliminary results from ongoing genetic studies have shown the possibility of clustering different histotypes based, for example, on kinase expression (Fig. 82-1A).²²

DNA, histones, and nonhistone proteins are condensed into a highly complex nucleoprotein structure known as chromatin that acts as a prototype for all of the genetic information. Chromatin conformation can be in a heterochromatin form, a highly compact structure, associated primarily with transcriptionally inactive genes, or a euchromatin form, consisting of a more open or relaxed structure, associated with transcriptionally active genes.²³ Epigenetic mechanisms mediated by histone deacetylases (HDACs), histone acetyltransferases, and histone methyltransferases play critical roles in various biological and cellular processes, including cell proliferation, angiogenesis, hypoxia-related effects, and cell-cycle regulation. Histones on the N-terminal are altered by acetylation, methylation, phosphorylation, ubiquitination, sumoylation, deamination, or adenosine diphosphate ribosylation.^24,25 The different histone residues and their modifications result in either transcriptionally active or repressive marks. For example, histone H3 lysine 4 (designated as H3K4), H3K36, and H3K79 are associated with active marks, whereas H3K9, H3K27, and H4K20 are associated with repressive marks.²⁶

Several studies have recently reported SETD2, a histone methyltransferase, as a tumor suppressor gene in RCC.27,28 An initial study consisting of a large-scale next-generation sequencing of primary RCC genomes identified somatic truncating mutations in SETD2 (approximately 3%).²⁸ These mutations are associated with a decrease in H3K36 trimethylation levels in several clear cell renal carcinoma cell lines, VHL mutations and/or hypoxic phenotype (determined by a panel of hypoxia-related genes) in clinical samples, as well as a chromatin remodeling complex protein PBRM1.²⁹ It is also important to note that SETD2 and PBRM1 are located on the short arm of the human chromosome 3 (3p21.31), in close proximity to the VHL gene and the 3p region is frequently altered in clear cell RCCs³⁰ (see Fig. 82-1B). Although mutations in enzymes governing chromatin remodeling are often found in advanced stage disease, these mutations have not been correlated to survival.³¹ The H3K27 methyltransferase EZH2 (which is associated with aggressiveness in breast cancer and overexpressed in metastatic prostate cancer)³² is overexpressed in renal tumor patient samples compared with their normal adjacent.³³ Conversely, histone demethylases governing H3K27 methylation status UTX and JMJD3 are overexpressed in clear cell RCCs compared with the adjacent nontumor tissue, which corresponds to the decreased H3K27 methylation in clear cell RCC.³⁴ These preliminary evidences suggest that alterations in histone-modifying genes may be associated with RCC and could be exploited for therapeutic interventions.

More recently, the BAP1 protein, a nuclear deubiquitinase, has been reported to be inactivated in 15% of clear cell RCCs. BAP1 mediates suppression of cell proliferation, ubiquitinates the histone 2A lysine 119, and its loss sensitizes RCC cells in vitro to genotoxic stress. Initial evidences suggest that BAP1 loss is associated with high tumor grade and poor prognosis. Interestingly, mutations in BAP1 and PBRM1 appear to be mutually exclusive. These preliminary results provide a rationale for future integrated pathological and molecular genetic classification of RCC that will likely lead to subtype specific treatments.³⁵

Intratumor heterogeneity has always been considered a potential major clinical hurdle to develop personalized-medicine strategies that depend on results from single-tumor biopsy samples. A recent report has now shown very elegantly that intratumor heterogeneity is a reality.³⁶ Different techniques, including exome sequencing, chromosome aberration analysis, and ploidy profiling on multiple spatially separated samples obtained from primary renal carcinomas and associated metastatic sites, determined that 63% to 69% of all somatic mutations were not detectable across every tumor region. Mutational intratumor heterogeneity was seen for multiple tumor suppressor genes and chromatin remodeling genes, including SETD2, PTEN, and KDM5C. Gene-expression signatures associated with good and poor prognosis were detected in different regions of the same tumor. These new intriguing findings, combined with the complex genomic landscape, make us rethink the common assumption that a single genomic test might guide therapy. The timing and location of the tumor sample acquisition remain a challenge, but there is still an opportunity to make personalized medicine a goal to achieve in the future for the treatment of cancer, and RCC in particular.

Familial Renal Cell Carcinoma

In a small percentage (5%) of cases, RCC is a feature of one of several hereditary syndromes.15,45 Such syndromes are associated with distinct histologic subtypes of RCC, and in each case patients have increased risk of multifocal tumor development.^15,45 Management is dependent on preservation of renal function. Close surveillance and minimization of surgical procedures constitute the mainstay of treatment.

von Hippel-Lindau syndrome is a disorder of autosomal dominant inheritance that occurs in 1 in 40,000 births. The mean age at onset is in the fourth decade of life. The syndrome is inherited as a result of a germline mutation in a single allele of the VHL gene tumor suppressor gene located on chromosomal band 3p25–26.15,45,46 Sporadic loss of the remaining wild-type VHL allele provides the “second hit” necessary for tumorigenesis, most commonly via chromosome 3p deletion. Multifocal tumorigenesis is observed in multiple organ systems, with each tumor harboring an independent second VHL mutation. Renal manifestations include cysts and clear cell RCC tumors. Both tend to be multifocal and bilateral, and are found in the majority of patients with von Hippel-Lindau disease.^15,45,46 Hundreds of independent clear cell cancers may be present in a single kidney, including dozens of macroscopic tumors. VHL syndrome patients are at high risk for chronic renal insufficiency because of the lifelong risk of multifocal RCC tumor development and need for repeat renal surgeries. As a result, VHL patients should undergo active surveillance until the largest tumor reaches 3 cm, at which time attempts may be made to resect all tumors in that kidney. Resection by enucleation without clamping of the main renal artery is recommended to maximize nephron sparing. Surgical candidates, particularly those with numerous tumors, are counseled as to the high possibility of local recurrence from de novo tumor formation and future ipsilateral surgery. The discovery of the VHL gene in hereditary clear cell RCC enabled the subsequent identification of a VHL mutation in sporadic clear cell RCC tumors (see above).⁴⁷

In addition to renal cysts and cancer, common manifestations of the VHL syndrome include benign vascular tumors of the spinal cord, cerebellum and retina, presenting as neurologic and visual symptoms. The endocrine system may also be affected by adrenal pheochromocytomas and pancreatic neuroendocrine tumors, in addition to pancreatic cysts. Benign papillary cystadenomas of the epididymis or broad ligament also develop in a minority of patients, which, when bilateral are pathognomonic for von Hippel-Lindau disease. Firm epididymal nodules on physical should prompt scrotal ultrasound and with a family history of VHL provides easy method to confirm the diagnostic suspicion. Endolymphatic sac tumors of the inner ear also may be seen.

Birt-Hogg-Dubé syndrome is a disorder of autosomal dominant inheritance. Signs and symptoms usually manifest in the fifth decade of life and include renal tumors and cysts, benign skin tumors (fibrofolliculomas) and pulmonary cysts, which can lead to spontaneous pneumothorax. The renal neoplasms may be multifocal and bilateral tumors and most often have pure chromophobe histology or a “hybrid” mixture of chromophobe and oncocytoma; infrequently, pure oncocytoma tumors may be present. Patients can present with several different tumor types within the same kidney and the presence of benign (oncocytoma) and malignant tumors within the same kidney should immediately prompt the suspicion of Birt-Hogg-Dubé (BHD) syndrome. The BHD gene mutated in this syndrome encodes the protein Folliculin and is located on chromosome 17p11.2.15,45,46,48,49 The BHD gene appears to have the characteristics of a loss-of-function tumor suppressor gene.⁵⁰ Folliculin has unknown function but is found in complexes with adenosine monophosphate-activated protein kinase, the major sensor of cell energy and a negative regulator of the mammalian target of rapamycin (mTOR) pathway. Recently, multiple studies have implicated Folliculin in adherens junction formation and signaling.^51,52

Tuberous sclerosis is a syndrome of autosomal dominant inheritance, with two genes identified, TSC1, located on 9q34, and TSC2, located on 16p13.3. It affects 1 in 6000 people and is usually diagnosed at birth.15,45,46 This syndrome encompasses multiple organ systems, including dermatologic, cardiac, pulmonary, and renal. Skins lesions include facial angiofibromas, periungual fibroma, shagreen patches, and hypopigmented macules. Patients also develop cardiac rhabdomyomas, pulmonary lymphangioleiomyomatosis, retinal hamartomas, subependymal nodules, and giant cell astrocytomas. The renal manifestations include bilateral and multifocal angiomyolipomas (AMLs) and less commonly clear cell renal carcinoma. In contrast with spontaneous AML, AML in this setting can be associated with a low risk of occult RCC (1%). The TSC1 and TSC2 gene products inhibit activation of mTOR signaling, a major promoter of protein synthesis and cell growth.

Hereditary papillary renal cell carcinoma (HPRCC), inherited as an autosomal dominant trait, is caused by mutations in Met protooncogene on chromosomal band 7q31–34.15,46,53 It is characterized primarily by bilateral, multifocal papillary type I RCC. These tumors are not aggressive and rarely metastasize. Age at onset is around the fifth decade. The Met oncogene encodes a membrane tyrosine kinase that, in HPRCC, harbors an activating mutation in the kinase domain. Hyperactivation of the Met oncoprotein leads to upregulation of several intracellular signaling pathways involving Akt, Rac, and MAP kinase.

Hereditary leiomyomatosis and renal cell carcinoma (HLRCC) is a disease of autosomal dominant inheritance. The gene for this disorder localizes to chromosomal band 1q42.3–43 and has been identified as fumarate hydratase (FH).15,45,46 Age at onset is between the third and fourth decades of life. The syndrome consists of cutaneous leiomyomas, uterine leiomyomas (fibroids), and papillary type II RCC tumors with high metastatic potential, even when small in size. Unlike the VHL patients, in whom delay in surgical treatment is usually the rule until the largest tumor reaches 3 cm, there should be no delay in treatment of solid renal lesions in these patients. Cystic lesions in these patients should be followed with close surveillance and early surgical intervention for any radiographic development of a potentially solid component. Thirty percent of patients have solitary and unilateral renal tumors.^45,46

The FH gene functions as a tumor suppressor, with loss of the second allele detected in kidney tumors. The wild-type gene encodes an enzyme in the Krebs cycle catalyzing fumarate conversion into malate. Loss of the FH enzyme leads to accumulation of fumarate, which has been suggested to promote carcinogenesis through indirect stabilization of transcription factors HIFα and NRF2.39,40,54

Distinct from HPRCC and HLRCC, Malchoff and colleagues have described a three-generation family with five cases of papillary thyroid carcinoma and two cases with papillary renal neoplasia.⁵³ With the use of linkage analysis, these investigators demonstrated that the fPTC/PRN phenotype was linked to 1q21.⁵³ They characterized a distinct inherited tumor syndrome that may establish a link between papillary RCC and familial papillary thyroid carcinoma.⁵³

Hereditary renal cell cancer associated with melanoma has been recently described. The TFE3 gene mutated in sporadic RCC (see “Diagnosis of Renal Cell Carcinoma”) is one of four members of the MiT family of transcription factors; although TFE3 mutations have not been identified in hereditary RCC syndromes, the related MiT member, MiTF, was shown to have a specific amino acid substitution associated with hereditary RCC tumors associated with melanoma.⁵⁵ This substitution confers hyperactivation of MiTF transcriptional activity by preventing its sumoylation and degradation. Histologic features of these MiTF renal cancers are yet to be characterized.