Introduction

The past two decades have witnessed rapid advances in the treatment of metastatic renal cell carcinoma (RCC) (Fig. 1). The introduction of targeted therapies directed at vascular endothelial growth factor (VEGF) receptors and mTOR in 2005 (refs1,2,3,4) was predicated on the observation that the von Hippel–Lindau (VHL) gene was frequently altered in sporadic metastatic RCC. Loss of VHL heterozygosity is observed in >90% of cases and VHL mutations are present in 50–65% of cases5,6,7. However, an interesting paradox emerged — although these drugs were developed with the intent of addressing altered VHL biology, response to treatment was not dependent on the presence or absence of VHL alterations5,8. First-generation VEGF-directed therapies were followed by new inhibitors with increased specificity for VEGF, and multi-kinase inhibitors directed at other putative targets in RCC pathogenesis9 (Fig. 1).

Fig. 1: Evolution of therapies in the metastatic RCC armamentarium.
figure 1

Since the 2000s, cytokine therapies that were used to treat metastatic clear cell renal cell carcinoma (RCC), such as IFNα and IL-2, have been largely supplanted by targeted therapies directed at vascular endothelium growth factor receptors (VEGFRs) and mTOR1,2,3,4. New inhibitors with improved specificity for VEGFR (for example, axitinib) were subsequently developed, as well as multi-kinase inhibitors9. Cabozantinib, for example, targets the MET receptor tyrosine kinase encoded by MET and the tyrosine protein kinase receptor UFO encoded by AXL, whereas lenvatinib targets include VEGFRs and fibroblast growth factor receptors204,205. Nivolumab, which prevents signalling through programmed cell death 1 (PD-1), was the first immunotherapy agent approved for RCC10. The combination of nivolumab and ipilimumab, which inhibits cytotoxic T lymphocyte protein 4 (CTLA4), was subsequently shown to improve patient survival and became the standard of care first-line treatment for patients with intermediate and high-risk RCC11,12. The latest combination therapy regimens include agents that target VEGF and immune checkpoints13,14 such as axitinib–avelumab and axitinib–pembrolizumab, which target VEGFR in combination with PD-1 ligand 1 (PD-L1) and PD-1, respectively.

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The use of nivolumab, an antibody against programmed cell death 1 (PD-1), marked the beginning of the immunotherapy era in RCC treatment10 (Fig. 1). Since then, several immunotherapy combinations have been shown to improve survival in patients with RCC11,12. Combination regimens that simultaneously target VEGF and immune checkpoints were approved in 2019. These drug combinations have profound activity, are well tolerated and prolong patient survival13,14.

As the treatment landscape of RCC grows in complexity and an increasing armamentarium of choices becomes available, defining biological strategies for selecting the right therapy for the right individual becomes crucial. For example, combination strategies might lead to overtreatment of patients who would have garnered equivalent benefit from monotherapy. These patients could be spared the unnecessary toxicity and financial encumbrance. Fortunately, the use of genomic research in clinical trials has begun to clarify the predictive or prognostic roles of certain genomic alterations in guiding RCC outcomes.

In this Review, we detail existing evidence of the prognostic and predictive value of genomics in RCC. We examine, for each histological RCC type, the associations between clinical outcomes and alterations in various genes, including VHL, chromatin remodelling genes such as PBRM1, BAP1, SETD2 and EZH2, as well as the DNA damage repair (DDR) genes, MET, TERT, NF2, SMARCB1 and TFE3. We also discuss gene expression signatures with prognostic and predictive value in localized and metastatic RCC, and conclude with a proposal for future directions in the analysis of clinical, pathological, genomic and patient-specific parameters that could be used to direct systemic therapy.

Relevant genomic alterations

The majority of genomics research in RCC has focused on identifying the clinical relevance of individual genes. Advances in whole-genome sequencing techniques enabled researchers to examine a larger number of genes and their prognostic and predictive relevance in large cohorts of localized and metastatic RCC. These studies, discussed below, revealed that the genomic profile and clinical relevance of individual genes differed based on the histological subtype of RCC.

Clear cell RCC

Clear cell RCC (ccRCC) is the major histological subtype of RCC and is observed in 75–80% of cases. Therefore, the majority of information on RCC genomics has been obtained from studies of this RCC subtype. The most commonly studied genes include VHL, owing to its historical association with VHL syndrome, as well as chromatin remodelling genes residing in close proximity to VHL in the short arm of chromosome 3. The list of genes with suggested clinical relevance in patients with ccRCC has expanded in line with our increasing understanding of the links between genomics and ccRCC outcomes.

VHL

VHL syndrome is a rare autosomal dominant disease caused by germline VHL mutations that presents with susceptibility to several cancer types, including RCC5. Further investigations demonstrated a link between somatic VHL mutations and the pathophysiology of sporadic ccRCC cases6,15. Proximal tubule epithelial cells have long been considered to be the cell of origin in ccRCC (Fig. 2), but some reports suggest that these tumours can also originate in the Bowman’s capsule16,17,18,19. ccRCC pathogenesis begins with a chromothripsis event, which occurs decades before clinical diagnosis20 and causes gene rearrangements, including the acquisition of one extra copy of chromosome 5q and the loss of one copy of chromosome 3p (Fig. 3). VHL is located at chromosome 3p (Fig. 3a) and loses its function, owing to point mutations or epigenetic silencing, years after the initial chromothripsis event20. Functional loss of VHL results in impaired ubiquitylation and accumulation of hypoxia-inducible factors (HIFs) within cell nuclei (Fig. 3b). Accumulated HIFs, in turn, increase the production of several growth factors that have key roles in the progression of RCC21,22,23 (Fig. 3b). A study from The Cancer Genome Atlas (TCGA) included a comprehensive genomic analysis of 478 patients with ccRCC and revealed that 52% of the cohort had genomic alterations in VHL. However, correlations between clinical outcomes and the presence of VHL mutations were weak in this cohort8. The prognostic value of VHL mutations in ccRCC has also been examined in several other studies, with little evidence demonstrating a relationship between VHL mutations or hypermethylation and clinicopathological characteristics or outcomes24,25,26,27,28. Current evidence suggests that, overall, VHL alterations are not associated with clinical outcomes29,30,31,32,33,34.

Fig. 2: Location and cell of origin of RCC histological subtypes.
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Epithelial cells from the proximal convoluted tubule have been proposed as the cells of origin for the two most common subtypes of renal cell carcinoma (RCC) — clear cell RCC and papillary RCC. Rare RCC subtypes such as Xp11 translocation RCC have also been suggested to originate from the proximal tubule epithelium. Chromophobe RCC originates from intercalated cells in the distal convoluted tubule, whereas collecting duct RCC and medullary RCC originate from distal collecting duct principal cells and calyceal epithelium, respectively. Importantly, the cells of origin for these RCC subtypes are currently under debate; for example, some evidence suggests that ccRCC might also originate from the Bowman’s capsule16,17,18,19.

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Fig. 3: Functional consequences of chromosome 3p loss.
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a | The short arm of chromosome 3 is particularly important in clear cell renal cell carcinoma (ccRCC) as loss of chromosome 3p is a key pathogenetic event. Accordingly, genes at this chromosomal location, which include VHL and the chromatin remodelling genes SETD2, PBRM1 and BAP1, have been implicated in the pathogenesis of ccRCC. b | Functional loss of VHL leads to a pseudohypoxia state due to impaired ubiquitylation and degradation of hypoxia-inducible factor 1α (HIF1α) and HIF2α. Accumulated HIF1α and HIF2α dimerize with HIF1β and activate transcription of HIF target genes, which include vascular endothelial growth factors (VEGFs), erythropoietin, platelet-derived growth factors (PDGFs), glucose transporter type 1 (GLUT1) and transforming growth factor-α (TGFα). c | In addition to the functional loss of VHL, loss of heterozygosity in chromosome 3p can also affect the expression of neighbouring chromatin remodelling genes. Genomic alterations in the component of the polycomb repressive deubiquitinase complex BAP1, the methyltransferase SETD2, or the component of SWI/SNF complex PBMR1, lead to impaired chromatin remodelling, which enables cancer cells to maintain their oncogenic activity through genome reprogramming.

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A 2017 meta-analysis that included 633 patients from six studies assessed the potential link between survival and VEGF-directed treatment; overall, 61.8% of patients had VHL alterations35. Among the studies included in the meta-analysis to determine the predictive value of VHL alterations were two retrospective studies of sunitinib, sorafenib, bevacizumab and axitinib, and one prospective study of pazopanib. The researchers determined that VHL alterations were not associated with overall response rate (ORR), progression-free survival (PFS) and overall survival. These findings suggest that VHL alterations do not have a predictive or prognostic role in ccRCC.

PBRM1

Polybromo 1 (also known as BAF180) is encoded by PBRM1 and is a major component of the PBAF complex, a mammalian SWI/SNF chromatin remodelling complex that has a key tumour suppressor role by regulating cellular proliferation and differentiation36,37,38,39 (Fig. 3c). PBRM1 mutations are the second most common somatic alteration in ccRCC and are detected in 30–40% of patients8,40. PBRM1 is located in the short arm of chromosome 3 in close proximity to VHL, SETD2 and BAP1 (ref.7) (Fig. 3a). Accordingly, loss of heterozygosity in chromosome 3p, which is the key tumour driver event in RCC pathogenesis, is also linked to loss of PBRM1 function7,41,42. Given that VHL alterations seem to have minimal clinical relevance, altered PBRM1 represents an alternative potential clinicopathological and prognostic correlate. Once again, however, contrasting results have emerged in the literature (Table 1).

Table 1 Studies of PBRM1 alterations and clinical outcomes
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Initial studies in this field investigated a link between PBRM1 mutations and pathological tumour characteristics. One study, which reported whole-genome and exome sequencing data from 176 ccRCC tumours, suggested that PBRM1 mutation status was unrelated to tumour grade43. By contrast, another retrospective study that included 185 patients who underwent nephrectomy reported that patients with PBRM1 mutations had higher odds of presenting with stage III disease44. However, this study failed to demonstrate a relationship between PBRM1 status and cancer-specific survival.

Investigators sought to validate the above findings in a TCGA cohort of 446 patients with ccRCC and found that PBRM1 status did not correlate with pathological stage or cancer-specific survival45. Interestingly, PBRM1 mutation frequency was similar across all pathological stages, suggesting that PBRM1 mutations occur early in the pathogenesis of ccRCC. Another study examined the potential association between PBRM1 mutation and recurrence-free survival in patients with localized ccRCC. PBRM1 mutations were associated with longer recurrence-free survival compared with tumours without these mutations (hazard ratio = 0.5; P = 0.073)46. The studies noted above pre-dated the global introduction of targeted therapies and immune checkpoint inhibitors. In parallel with these advances in the treatment of metastatic RCC, further studies examined the prognostic and predictive role of PBRM1 in the context of novel therapies.

In 2016, one study reported a genomic analysis of 31 patients with metastatic ccRCC who received targeted therapies, including bevacizumab, pazopanib, sunitinib, axitinib and others47 (Table 2). The frequency of PBRM1 mutations was 47.7% in this cohort, which was higher than that reported in previous studies that predominantly included patients with localized RCC. Importantly, the presence of PBRM1 mutations was associated with exceptional responses to targeted therapies — patients with a response duration >21 months had a higher frequency of PBRM1 mutations than those with shorter responses. Investigators also analysed patient responses according to the mechanism of targeted therapy. PBRM1 mutations were associated with a better response to treatment with VEGF-directed therapies, but no link was observed between PBRM1 status and the response to mTOR inhibitors. This study47 was the first to suggest a predictive value of PBRM1 alterations for VEGF-directed therapies. Another study similarly reported that the frequency of exceptional treatment responses (defined by partial or complete response >3 years) was highest among patients with PBRM1 mutations48.

Table 2 Mechanism of action and targets of therapeutics used in mRCC treatment
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The RECORD-3 phase III clinical trial compared sunitinib and everolimus as first-line treatments in patients with metastatic ccRCC49. Genomic analysis in this study showed that, for patients treated with everolimus, PFS was prolonged in patients with mutated PBRM1 compared with those without mutations; this survival advantage was not observed in patients who received sunitinib49. In another study, investigators studied the effect of mutations in PBRM1, BAP1 and TP53 on overall survival in a dataset that included not only the RECORD-3 cohort but also patients from the COMPARZ clinical trial, which compared sunitinib with pazopanib50. The analysis of the COMPARZ cohort (n = 377) showed that overall survival was significantly better in patients with mutated PBRM1 than in those without PBRM1 mutations. In order to generate a more effective risk stratification system, investigators combined the genomic findings regarding BAP1, PBRM1 and TP53 status with the Memorial Sloan Kettering Cancer Centre (MSKCC) risk stratification nomogram — a tool that had been developed to predict overall survival in patients with metastatic RCC based on clinical (that is, time from diagnosis to first-line therapy and performance status) and laboratory (that is, serum calcium, serum lactate dehydrogenase and haemoglobin) findings. This combined model was evaluated in the RECORD-3 cohort and was shown to improve prediction of PFS and overall survival compared with the overall survival prediction of the MSKCC nomogram alone.

In the IMmotion150 trial, 305 patients with metastatic RCC were treated with atezolizumab alone, or in combination with bevacizumab, or sunitinib alone51. Tumours with PBRM1 mutations expressed high levels of genes associated with angiogenesis. Accordingly, in patients with PBRM1 mutations, PFS was significantly longer in both the sunitinib and atezolizumab–bevacizumab combination groups compared with the atezolizumab alone group. Among those patients who received sunitinib, patients with PBRM1 mutations had 62% lower odds of progression compared with those with non-mutated PBRM1. PFS did not differ based on PBRM1 status within the other two treatment groups. Overall, these findings suggest a beneficial effect of PBRM1 mutations when patients are treated with angiogenesis inhibitors, but not with immunotherapies.

Importantly, contrasting results were reported in a study that employed both clinical and basic science research to examine the link between PBRM1 loss and the clinical benefit of immunotherapies52. The initial analysis involved patients with metastatic ccRCC who received nivolumab. Patients with truncating or loss-of-function mutations of PBRM1 had higher rates of clinical benefit (defined by the sum of complete and partial response rates) than those with non-mutated PBRM1 (82% and 23%, respectively; hazard ratio = 12.93, P = 0.012). Patients with biallelic loss of PBRM1 loss had even longer overall survival and PFS (P = 0.0074) than those with non-mutated PBRM1. These results were confirmed in a validation cohort of 63 patients with metastatic ccRCC who were treated with PD-1 or PD-L1 blocking agents52. Of note, gene sets associated with hypoxia and immune stimulatory activity were upregulated in cell lines deficient for polybromo 1 compared with those with the intact protein; this immune stimulatory effect might contribute to the improved response to immunotherapies observed in patients with PBRM1 mutations.

Another two large-scale studies of treatment with immunotherapies across several solid tumour types, including patients with RCC, reported contrasting results. One study failed to identify a relationship between PBRM1 mutations and overall survival in 143 patients with metastatic ccRCC treated with immunotherapies, alone or in combination with VEGF-directed agents53. By contrast, in another study, patients with RCC tumours harbouring mutations in major SWI/SNF-related genes (that is, PBRM1, ARID2, ARID1A, ARID1B, SMARB4 and SMARCB1) (Fig. 3c) had a longer time to treatment failure and their overall survival was improved compared with patients without alterations in these genes54.

Despite these attempts to investigate the potential association between PBRM1 and clinical outcomes, this issue remains unclear owing to discrepancies in results across different datasets. Notably, most reports are based on retrospective studies; thus, methodologies might differ between them owing to variation in patient characteristics, study assessments, treatment approaches and clinical outcomes. This variation presents challenges for the interpretation of results and the ability to draw broader conclusions. Of note, in contrast to earlier findings, most evidence suggests that PBRM1 mutations do not have adverse effects and several studies have demonstrated positive outcomes in patients with PBRM1 mutations compared with those without PBRM1 mutations (Table 1). Current evidence therefore suggests a favourable prognostic effect of PBRM1 alterations, which potentially also have predictive value, especially with regard to immunotherapies. Nonetheless, further studies that incorporate genomic characteristics into earlier stages of clinical trial design are necessary to clarify the reported discrepancies.

BAP1

BAP1 is a two-hit tumour suppressor gene (that is, inactivation of both alleles is required to induce a change in the phenotype of the cell) with histone deubiquitinase activity that acts as an essential chromatin regulator and suppresses cell proliferation (Fig. 3c). Similarly to PBRM1, BAP1 is located on the short arm of chromosome 3 and can thus also be affected by the most common RCC pathogenesis event — loss of the short arm of chromosome 3 (Fig. 3a). The frequency of BAP1 mutations in ccRCC ranges from 5–16%8,46,50,55. BAP1 mutations have been associated with metastatic disease at diagnosis, high Fuhrman grade and the presence of necrosis43,46,56,57. In a set of 1,479 tumour samples from patients with low-risk localized ccRCC, BAP1 staining, which was performed with a highly specific and sensitive immunohistochemistry assay, revealed that 10.3% of tumours were BAP1-negative, suggesting the loss of BAP1 (ref.57). After a mean follow-up of 8.3 years, the risk of death from RCC was significantly higher in patients with BAP1-negative tumours (hazard ratio = 3.06; 95% confidence interval = 2.28–4.10; and P = 6.77 × 10−14) than in patients with BAP1 positive tumours. This association was still significant after statistical adjustment to control for the effect of the risk groups estimated by the University of California Los Angeles integrated staging system (UISS) — a prognostication tool for patients with localized RCC that uses clinical and pathological characteristics to estimate 5-year disease-free survival after nephrectomy58. Similarly, whole-genome sequencing of tumours from 183 patients from the MSKCC cohort and 421 patients from the TCGA cohort showed that BAP1-mutated tumours were significantly associated with shorter overall survival than tumours with non-mutated BAP1 (ref.59).

Further studies have examined the interplay between BAP1 and PBRM1 mutations in ccRCC43,60. One study examined the effects of simultaneous loss of BAP1 and PBRM1, which was observed in only 3% of patients with ccRCC and was often associated with the presence of rhabdoid pathological features, which characterize a well-known aggressive variant of RCC. Among the entire cohort, researchers investigated polybromo 1 and BAP1 deficiency and found that 55% of tumours were only deficient for polybromo 1, whereas 12% were solely BAP1-deficient43. Gene expression analysis showed that tumours with BAP1 mutations had a gene signature distinct from that of tumours with PBRM1 mutations, with almost no overlap between the two signatures. This finding suggests that these mutations are largely mutually exclusive and might also be associated with distinct mechanisms underlying oncogenesis.

The outcomes associated with BAP1 and PBRM1 mutations were further investigated in studies that examined survival outcomes in cohorts of patients with predominantly localized disease or with metastatic ccRCC. One study compared the overall survival outcomes of patients with BAP1-mutated tumours with those of patients who had PBRM1-mutated tumours in two separate cohorts: University of Texas Southwestern (UTSW) and TCGA56. The analyses revealed that the median overall survival observed in patients with exclusive BAP1 mutations (4.6 years in UTSW and 1.9 years in TCGA) differed significantly from that of patients with exclusive PBRM1 mutations (10.6 years for UTSW and 5.4 years in TCGA). Of note, the differences in survival outcomes between these two cohorts were attributed to discrepancies in the use of targeted therapies between the two sites; however, further analyses revealed similar hazard ratios across the two sites (2.7 for UTSW versus 2.8 for TCGA), which supports the reproducibility and reliability of these data. In addition, tumours with simultaneous BAP1 and PBRM1 mutations appeared to be highly aggressive and were associated, in the TCGA cohort, with a 0.2-year overall survival.

The genomic analysis of tumours from patients in the COMPARZ and RECORD-3 clinical trials50, noted earlier with regard to PBRM1, also provided important insights into the role of BAP1 mutations. In the COMPARZ cohort, BAP1 mutations were associated with reduced PFS and overall survival in treatment-naive patients with metastatic or advanced ccRCC treated with VEGF-directed therapies compared with patients with non-mutated BAP1. BAP1 alterations were also one of the three genomic findings included in the new model that combined the MSKCC nomogram with genomic findings and improved prediction of risk classes50. Given its prospective design and the large number of patients examined, this study provides strong evidence that BAP1 has both a prognostic and a predictive role in treatment planning.

Importantly, a crucial gap exists in our current knowledge of the predictive relevance of BAP1 alterations in the context of immunotherapy. Only a few studies have reported findings for a limited number of cancer types. For example, in peritoneal mesothelioma, in which mutated BAP1 is a frequent driver and a negative prognostic indicator, haploinsufficiency of BAP1 was associated with activation of immune checkpoint receptors61. To our knowledge, such questions have not been addressed in the context of ccRCC. Ongoing and recently completed, large-scale immunotherapy studies in advanced ccRCC have included genomic assessments and might be able to address these clinical questions. Elucidating the molecular pathways downstream of BAP1 alterations will also be crucial to the development of targeted treatments for the aggressive tumours associated with loss of BAP1.

SETD2

The histone-lysine N-methyltransferase SETD2 modulates alternative splicing and active transcription45,62,63,64,65,66,67 (Fig. 3c). Beyond its role in chromatin regulation, SETD2 contributes to DDR — loss of SETD2 and subsequent loss of trimethylated histone 3 lysine 36 (H3K36me3) in ccRCC cell lines resulted in microsatellite instability68,69. SETD2 is also located on chromosome 3p and is therefore prone to alterations in the context of ccRCC (Fig. 3a). Interestingly, a meta-analysis reported that SETD2 mutations were often observed in patients with PBRM1 mutations. Of note, the presence of simultaneous mutations in SETD2 and PBRM1 did not affect overall survival compared with tumours in which PBRM1, but not SETD2, was mutated70.

In a TCGA cohort that included predominantly patients with localized ccRCC tumours, the frequency of SETD2 alterations was 13%, whereas in studies of patients with metastatic ccRCC this frequency increased to >30%47,49. Additionally, the majority of SETD2 mutations were subclonal (that is, the mutations were present in a subpopulation of tumour cells rather than in all cells within a tumour) and SETD2 mutation status might therefore vary across different cancer cells in one specimen71. An analysis of 421 patients from the TCGA cohort also showed that tumours harbouring SETD2 mutations were more likely to recur (hazard ratio = 2.5)59. Notably, SETD2 was the only chromatin regulator gene in chromosome 3p that had a significant association with disease recurrence, suggesting that it might have an important role in oncogenesis. Accordingly, cancer-specific survival was also lower in patients with SETD2 mutations than in those with non-mutated SETD259.

In another study, researchers investigated the SETD2 mutation status in ccRCC tumours using whole-genome and/or whole-exome sequencing (n = 106), as well as targeted deep sequencing (n = 134). Compared with tumours in which SETD2 was not mutated, SETD2 mutations were once again associated with shorter disease-free survival and a higher risk of disease recurrence. However, overall survival was not associated with SETD2 mutation status. The researchers suggested that this effect might be due to the higher frequency of BAP1 mutations, which are associated with a more aggressive disease course, in patients with non-mutated SETD2 than in those with SETD2 mutations7.

In the setting of metastatic disease, next-generation sequencing (NGS) data from the COMPARZ and RECORD-3 trials failed to demonstrate a relationship between overall survival and SETD2 mutation status50. A more specific analysis of the RECORD-3 data revealed no differences in PFS according to SETD2 mutation status in either the sunitinib or the everolimus group. The role of SETD2 mutations in metastatic disease therefore remains to be fully elucidated.

EZH2

The histone methyl transferase EZH2 is a component of the polycomb repressive complex 2 (PRC2) — which is involved in chromatin remodelling — and functions as an important epigenetic modulator in oncogenesis. Activation of EZH2 leads to epigenetic silencing of various genes, including tumour suppressor genes72,73,74. Several reports have demonstrated nuclear overexpression of EZH2 in RCC compared with benign renal parenchymal tissue. EZH2 overexpression has also been linked to tumour aggressiveness in vivo75,76 and transfection of RCC cell lines with a plasmid that encodes EZH2 increased their invasiveness and migration in vitro75. Moreover, findings from nude mice injected with metastatic RCC cells, with or without EZH2 knockdown, and subsequently treated with a methyl transferase inhibitor suggested that inhibition of EZH2 reduced tumour growth and prolonged survival75. Further studies that examined the clinical and pathological correlates of EZH2 overexpression have confirmed its association with a poor prognosis77.

In a study of 257 patients with localized RCC (predominantly ccRCC), high EZH2 expression was associated with higher TNM stage, as well as reduced disease-free survival and overall survival75. Similar associations were reported in several other studies, including a large multicentre study of three cohorts (>2,000 patients), which further supports the association between EZH2 expression and tumour aggressiveness76,77,78,79. In 2017, researchers reported that initial resistance to sunitinib could be overcome by reducing the expression of EZH2 in patient-derived xenograft models80.

The prognostic relevance of EZH2 is clear, but further research is needed to establish whether EZH2 is a potential therapeutic target. Tazemetostat is an experimental, selective and competitive inhibitor of EZH2 — a phase I study in patients with refractory non-Hodgkin lymphoma or with advanced solid tumours confirmed that this drug has a favourable safety profile. Clinical trials of tazemetostat in patients with solid tumours bearing EZH2 gain-of-function mutations, including renal medullary carcinoma, are currently underway81,82.

DNA damage repair genes

Mutational burden increases in cancer cells with alterations in DDR genes, as these cells accumulate mutations that cannot be repaired. These mutations lead to the formation of a diverse set of neoantigens, some of which can trigger an antitumour immune response83. Immune checkpoint inhibitors block the tumour-driven suppression of existing antitumour immune responses and were thus hypothesized to provide greater benefit in patients with defects in DDR, as they might be more likely to mount an antitumour immune response84,85,86. Several studies have suggested that DDR gene alterations have predictive value in the response to immunotherapies, including in colorectal cancer and in bladder cancer84,87.

One study examined the effect of deleterious mutations in DDR genes on the survival of 229 patients with metastatic RCC treated with immunotherapy or a tyrosine kinase inhibitor88. DDR alterations were noted in 17% of the study population. PFS was similar for patients with deleterious mutations in DDR genes and for those without such mutations, regardless of the type of treatment received. However, patients treated with immunotherapies who had tumours with DDR gene alterations had significantly longer overall survival than their counterparts with non-mutated DDR genes88. Another study of patients with metastatic RCC treated with immune checkpoint inhibitors reported that DDR mutations were more frequent in patients with disease control89. These initial results are encouraging and warrant further examination of the potential predictive or prognostic value of DDR defects in metastatic ccRCC.

Papillary RCC

Papillary RCC (pRCC) comprises 15–20% of newly diagnosed RCC cases and represents the largest subgroup of non-ccRCC8. pRCC is proposed to originate from cells of the proximal tubule epithelium (Fig. 2) and is divided into type 1 or type 2 based on histopathological appearance — type 2 presents with more aggressive clinical and pathological features than type 1 and is also associated with poorer survival outcomes18,90,91. In the late 1990s, studies of hereditary pRCC, a rare hereditary cancer syndrome, identified that activating mutations of the MET proto-oncogene have a key role in the development of characteristic multifocal and bilateral pRCC in this familial cancer syndrome92,93. Subsequent genomic profiling of patients with somatic pRCC (that is, non-hereditary pRCC) revealed frequent MET alterations, particularly among patients with type 1 pRCC94. MET is located in the long arm of chromosome 7 and encodes the MET protein (also known as hepatocyte growth factor receptor)95. Under normal conditions, MET induces downstream intracellular pathways that regulate motility, growth and survival in epithelial cells, and tissue remodelling95. Abnormal activation of pathways downstream of MET signalling might result from alterations in the copy number of chromosome 7, MET alterations, transcriptional upregulation of MET or ligand-related mechanisms. This dysregulated activity can lead to oncogenesis by preventing apoptosis, inducing aberrant angiogenesis and promoting cell invasion96,97.

TCGA includes samples from 274 patients with pRCC and represents the largest available dataset of genomic features in these tumours8,94. However, similar to the ccRCC cohort of the TCGA, the pRCC cohort is dominated by patients with localized RCC and <5% of patients had proven metastatic disease. A comparison of the genomic features of different histopathological subtypes of RCC91,98 revealed that overall mutation rates were significantly higher in pRCC than in ccRCC; rates of mutation in chromatin regulator genes were comparable. In contrast to ccRCC, PBRM1 mutations were associated with worse survival in pRCC. Chromosomal copy number assessment showed that ~80% of patients with type 1 pRCC had gains of chromosome 7 and chromosome 17, whereas type 2 pRCC was not associated with a particular copy number alteration. MET alterations, including mutations, fusions and an increase in copy number, were noted in 83% of patients with type 1 pRCC and MET expression was significantly higher in type 1 than in type 2 pRCC. By contrast, one quarter of patients with type 2 pRCC had high rates of CDKN2A mutations, which were associated with an overall poor prognosis. Mutations in chromatin regulator genes such as SETD2, PBRM1 and BAP1 were also detected in patients with type 2 pRCC. Another subset of patients with type 2 pRCC had a CpG island methylator phenotype RCC, which was characterized by prominent DNA hypermethylation, low overall mutation rates and high rates of alteration in CDKN2A and FH. Patients with a CpG island methylator phenotype RCC had the poorest survival rates of all RCC subtypes8,99.

We analysed the genomic features of tumours from patients with metastatic RCC using a commercially available comprehensive genomic profiling database and found that 33% of patients with type 1 pRCC had MET alterations98. Moreover, TERT, CDKN2A, CDKN2B and EGFR were among the most commonly mutated genes in this disease subtype. By contrast, only 7% of patients with type 2 pRCC had MET alterations and the most commonly altered genes were CDKN2A, CDKN2B, TERT, NF2 and FH. Patients with type 2 pRCC also had higher rates of alterations in genes that encode components of the SWI/SNF chromatin regulator complex (Fig. 3c) than patients with type 1 pRCC98.

In parallel, another research group analysed MET expression and chromosome 7 copy number gain in patients with pRCC100. Overall, MET expression was significantly higher in pRCC specimens than in matched normal kidney tissue, and was also higher in type 1 pRCC samples than in type 2 samples. Comparative genomic hybridization studies detected chromosome 7 copy gain in 81% and 46% of type 1 and type 2 pRCC samples, respectively. MET expression levels correlated with the alterations in copy number in both disease subtypes. Collectively, these findings support the rationale of targeting MET as a therapeutic strategy across all pRCCs100.

Several multi-kinase inhibitors that have an effect on MET have been investigated in patients with pRCC101,102,103,104,105 (Table 3) — response rates and PFS outcomes varied substantially across different MET inhibitors. For example, PFS was relatively long (9.3 months) in patients treated with foretinib. In this cohort, all patients with germline MET mutations (n = 10) achieved disease control (half of the patients had a partial response and the other half had stable disease)101. In the subgroup of patients with non-germline MET alterations (n = 20), 3 patients had a partial response and 16 patients had stable disease. By contrast, a clinical trial of tivantinib, a non-competitive MET inhibitor, was terminated early because of a lack of efficacy102. Subsequent trials demonstrated the favourable efficacy of crizotinib, savolitinib and cabozantinib in patients with pRCC, irrespective of MET mutation status; of note, patients with MET-altered tumours had better outcomes in response to these drugs. Two ongoing large randomized studies are testing MET-targeted agents in patients with pRCC and NGS of archival tissue is being used to characterize MET alterations. The SAVOIR trial is comparing sunitinib and savolitinib, whereas the PAPMET trial is comparing sunitinib, cabozantinib, savolitinib and cabozantinib106,107. An interim analysis of the PAPMET trial led to closure of the savolitinib and crizotinib arms because their efficacy was shown to be inferior to that of sunitinib and cabozantinib. So far, the scarcity of patients with pRCC has been a major barrier in identifying genomic correlates of clinical outcomes for these patients in prospective studies, but multi-institutional investigations might help to address this limitation.

Table 3 Outcomes in clinical trials of MET-targeting agents in metastatic papillary renal cell carcinoma
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Chromophobe RCC

Chromophobe RCC (chRCC) was initially identified in patients with a rare hereditary disease, Birt–Hogg–Dube syndrome, which is characterized by a high risk of developing benign and malignant skin and kidney tumours108,109. From a clinical standpoint, chRCC tends to have an indolent and localized growth pattern and survival outcomes for patients with this disease are relatively better (5-year overall survival rate >80%) than those for patients with other subtypes of RCC110,111. Data from several studies that assessed DNA methylation, mRNA expression and protein expression through immunohistochemistry suggest that chRCC arises from distal tubule intercalated cells18,112,113 (Fig. 2).

Genomic data from chRCC-specific and RCC-specific TCGA reports, which included 66 and 81 patients with chRCC, respectively8,113, showed that overall mutation frequencies were lower in chRCC than in other RCC subtypes. Accordingly, rates of mutation in chromatin regulator genes were also lower in chRCC and were not significantly associated with clinical outcomes. However, whole-exome sequencing analyses identified a high frequency of mutations in two tumour suppressor genes — TP53 and PTEN — in chRCC samples. PTEN mutations were significantly associated with poor survival in patients with chRCC; a similar association was observed for CDKN2A mutations. Furthermore, the loss of one copy of chromosomes 1, 2, 6, 10, 13 or 17 occurred in a high proportion of chRCC cases; these copy number alterations were less frequent in eosinophilic chRCC, which is histologically characterized by the presence of eosinophilic cytoplasm and densely packed mitochondria. More importantly, novel genomic rearrangements involving the TERT promoter region and consequent TERT overexpression were identified in chRCC. TERT is a telomerase regulator and data from TCGA have suggested that alterations in the TERT promoter might drive chRCC113.

chRCC tumours also overexpress proteins involved in mitochondrial energy production — nearly all genes associated with the Krebs cycle and at least one gene from each unit of the electron transport chain were overexpressed113. Genomic alterations that affected mTOR and its downstream pathways were also identified in 23% of patients with chRCC113. These alterations were significantly associated with poor survival in chRCC; thus, future studies should investigate the potential benefits of mTOR-directed therapeutics in the treatment of chRCC. Another study, which combined a local cohort of patients with two additional chRCC cohorts (TCGA and MSK-IMPACT) showed that alterations in TP53 and PTEN, and unbalanced chromosome duplication (that is, duplication of ≥3 chromosomes) were associated with poor overall survival114. Furthermore, these alterations were more frequently observed in metastatic tissue samples compared with matched primary tumour samples, suggesting a notable role in metastatic progression.

RCC with sarcomatoid features

Sarcomatoid dedifferentiation in RCC (sr-RCC) is characterized by the presence of malignant spindle cells buried in epithelial malignant cells115. This dedifferentiation is observed in ~5% of all cases of RCC and is most frequently observed in ccRCC and chRCC subtypes116. sr-RCC has long been associated with a worse prognosis in patients with localized or metastatic RCC, and a higher ratio of sarcomatoid cells to subtype-specific epithelial malignant cells is associated with more aggressive disease features116,117. Both the sarcomatoid and epithelial components of RCC originate from the same progenitor cell, but cells in the sarcomatoid component have undergone epithelial-to-mesenchymal transition118,119.

RNA sequencing of nephrectomy specimens from eight patients with sr-ccRCC identified overexpression of aurora kinase A and increased mTOR activity in sarcomatoid cells compared with adjacent ccRCC cells or benign kidney cells120. However, a study of 85 patients with sr-RCC treated with mTOR inhibitors reported an objective response rate of only 7%; 49% of patients had stable disease but PFS was disappointing (2.9 months)121. Two studies, published simultaneously, used NGS to investigate the genomics of sr-RCC122,123. One study compared the sarcomatoid and clear cell components of sr-ccRCC and found that mutational burden and loss of heterozygosity were increased in the sarcomatoid component, which was also the only tissue with mutations in the tumour suppressor genes TP53, BAP1 and ARID1A122. The other study examined sr-RCC sections from a cohort of patients with mixed RCC histology and found that 42% of the specimens carried TP53 alterations; TP53 was the most frequently altered gene in these tumours123.

In 2019, investigators reported that, in a large cohort of patients with metastatic RCC, the presence of sarcomatoid or rhabdoid dedifferentiation was not associated with the frequency of genomic alterations and tumour mutational burden. Notably, alterations of BAP1 were more frequent in patients with sarcomatoid or rhabdoid features and these patients were also more likely to respond to immunotherapies compared with kinase inhibitor therapies. Although time to treatment failure was similar in both treatment arms, the 2-year overall survival rate was significantly higher when patients with sarcomatoid or rhabdoid RCC were treated with immunotherapies than when they were treated with non-immunotherapy agents (hazard ratio for death = 0.52)124.

Despite arising from the same progenitor cell, genomic distinctions between sarcomatoid and primary components of RCC highlight the importance of branched evolution and both intra-tumoural and inter-tumoural heterogeneity125. These findings emphasize the importance of studying the genomic characteristics of sr-RCC in large cohorts of patients in a clinical setting.

Collecting duct RCC

Collecting duct RCC (cdRCC) arises from the principal cells of distal collecting ducts126,127 (Fig. 2). Owing to its rare nature, clinical evidence regarding cdRCC treatment and outcomes is largely derived from studies with small sample sizes and is thus limited. Current treatments largely rely on platinum-based chemotherapies128. A study of five patients with cdRCC demonstrated that a combination of platinum, gemcitabine and bevacizumab had antitumour activity, as three patients had a partial response, one maintained stable disease and another experienced complete clearance of metastatic lesions129. In addition, the overall survival of patients treated with this triple drug combination was 28 months. Overall, cdRCC remains an under-investigated entity with an often aggressive clinical course129. The proximity of origin between cdRCC and upper-tract urothelial carcinomas prompted investigators to study whether these two diseases have common genomic features. Comparative genomic hybridization coupled with transcriptomic assessments confirmed that cdRCCs were more similar to RCCs than to upper-tract urothelial carcinomas130. Furthermore, NGS of previously established cancer-related genes in specimens from 17 patients with cdRCC revealed genomic alterations in SETD2, SMARCB1 and CDKN2A, with further evidence of NF2 alterations in one third of the cohort131. Mechanistically, truncating NF2 mutations result in the loss of merlin — a tumour suppressor protein that regulates receptor tyrosine kinases. In vivo studies demonstrated that loss of merlin activates the mTORC1 protein complex, of which mTOR is a major component132. Furthermore, previous preclinical and clinical studies have demonstrated an antitumour effect of mTOR inhibitors in NF2-altered tumours, especially vestibular schwannoma and urothelial carcinoma133,134,135,136,137. Collectively, the high frequency of NF2 alterations in metastatic cdRCC and the existing translational evidence suggest the potential efficacy of mTOR inhibitors in patients with cdRCC, which warrants further investigation.

Renal medullary carcinoma

Renal medullary carcinoma (RMC) is thought to originate from cells of the distal nephron (Fig. 2) and its clinical features are distinct from other RCC subtypes127,138,139. RMC is almost always seen in individuals with sickle cell trait and remains a tumour of young adults. These tumours are highly aggressive and ~70% of patients are diagnosed with metastatic disease. RMC is almost always lethal — patients often briefly respond to chemotherapy but do not subsequently benefit from therapies that target VEGF or mTOR139,140. Transcriptomic analysis of 11 RMC samples demonstrated that RMC has gene expression features that are similar to chRCC but is distinct from the other subtypes of RCC141. SMARCB1 is a component of SWI/SNF (Fig. 3c) and alterations in this gene are observed in nearly all RMC tumours142,143,144. The assessment of SMARCB1 through NGS at the time of diagnosis is crucial in both supporting the clinical diagnosis and identifying potentially actionable mutations, such as ALK rearrangements, which have also been observed in RMC tumours. Treatment with ALK-targeted therapies has been successful in other solid tumours bearing such rearrangements, including non-small-cell lung cancer145,146. The RMC Working Group recommends debulking nephrectomy combined with chemotherapy as the preferred approach in RMC and encourages clinicians and investigators to engage in collaborative clinical registries, contribute to specimen biorepositories and encourage patients to participate in clinical trials to enhance understanding of this highly aggressive disease145,147,148,149,150.

Translocation RCC

Translocation RCC (tRCC) is another rare and distinctive subtype of RCC predominantly seen in children and young adults151,152,153,154. Similar to ccRCC and pRCC, tRCC is thought to arise from proximal tubule cells155. Despite the absence of prospective studies, retrospective studies have demonstrated the efficacy of VEGF-targeted therapies in this disease subtype. However, overall clinical outcomes in tRCC seem to be worse than those for major RCC subtypes153,154. The unique genomic event in tRCC is a translocation on chromosome Xp11.2, which includes genes that encode members of the microphthalmia-associated transcription factor (MITF) family (that is, TFE3, TFEB and TFEC)156,157,158. Accordingly, tRCCs are associated with overactivation of MITF family target genes, which promotes carcinogenesis159. One study reported that, in addition to the hallmark translocations, tRCC possesses similar genomic features to other major subtypes of RCC, such chromosome 3p loss (ccRCC) or trisomies 7 and 17 (pRCC)160. In another study, RNA sequencing of different non-ccRCC subtypes revealed that tRCC samples shared similar signalling pathways with pRCC161. NGS of seven patients with tRCC confirmed the activation of MITF targets but also revealed novel gene expression patterns, including the increased expression of transforming growth factor β1 and of downstream targets of the PI3K complex162. In the TCGA cohort, TFE3 and TFEB fusions were encountered in tumours that were histologically identified as pRCC or ccRCC45,163, suggesting that tRCC has probably been under-recognized in the past owing to the lack of genomic assessment in clinical practice. The WHO has broadened the definition of tRCCs and the MITF family of tumours now also includes translocation t(6;11) RCC, which is characterized by an Alpha–TFEB gene fusion156. Genomic assessment would be a valuable addition to the histopathological assessment and differential diagnosis of tRCC.

Genomic signatures

Despite the research efforts that have led to the discovery of promising associations between clinical outcomes and individual genomic alterations, no single alteration has been shown to directly modulate survival or benefit from RCC therapeutics. As a next step, genomic signatures have been developed, using expression patterns of various genes, to predict disease-free survival, prognosis or response to therapy.

Localized RCC

The treatment of localized RCC comprises surgery, with complete resection of malignant tissue, and subsequent follow-up to monitor for disease recurrence. Trials of adjuvant therapies to prevent tumour recurrence have been largely unsuccessful in both the cytokine era and in the targeted therapy era (Fig. 1). However, the S-TRAC trial demonstrated that 12 months of sunitinib treatment delayed tumour recurrence for ~1 year164. Current ongoing efforts include the use of targeted therapies or immunotherapies in both the adjuvant and neoadjuvant settings165,166,167,168,169.

Several nomograms have utilized clinical and pathological parameters to predict risk of tumour recurrence170,171,172,173. However, whether the genomic characteristics of a tumour can help to predict the risk of disease and whether adjuvant treatment is beneficial in RCC remain unclear. Using two large datasets that included a total of 1,568 patients, researchers developed and validated the ‘16-gene recurrence score’ as a predictor of disease recurrence174,175. An analysis of the expression patterns of 732 genes identified 11 genes whose expression correlated with recurrence-free survival and that corresponded to four major biological pathways: vascular (that is, expression of APOLD1, EDNRB, NOS3 and PPAP2B), cell growth and/or division (that is, expression of EIF4EBP1, TUBB2A and LMNB1), immune response (that is, expression of CEACAM1, CX3CL1 and CCL5) and inflammation (that is, expression of IL6). Five reference genes — AAMP, ARF1, ATP5E, GPX1 and RPLP1 — were also included in the score as indicators of RNA quality. Results suggested that higher expression of vascular and immune response genes was associated with a lower risk of 5-year recurrence, whereas higher expression in the other two categories was associated with an increased risk of recurrence175. This 16-gene signature was later successfully validated using data from the S-TRAC trial175. The authors suggested that the assay can be used to identify patients with localized ccRCC and at a high risk of recurrence (level IB evidence).

The ClearCode34 gene score is also designed to predict disease recurrence in patients with localized RCC and was based on archival tumour samples from 48 patients with ccRCC176,177. These analyses identified two independent clusters of patients, ccA and ccB, with distinct genomic and biological features, and clinical outcomes. Recurrence-free survival and overall survival were better in patients in the ccA subcategory than in those classed as ccB177. The accuracy and reproducibility of this genomic model has been validated in a TCGA cohort and in an additional cohort from the University of North Carolina178. Notably, RCC-specific mortality was more than three-fold higher for patients in the ccB subcategory compared with patients in the ccA group. Overall, the prognostic specificity of this classification was superior to existing clinical risk assessment tools such as the UISS score and the Stage, Size, Grade, Necrosis (SSIGN) score58,179. Moreover, a novel integrated approach that combines ClearCode34 with either the UISS score or the SSIGN score had better prognostic value than each clinical risk nomogram alone178.

The cell-cycle progression (CCP) score, another system based on RNA expression levels, also performed well in stratifying patients based on 5-year recurrence risk and RCC-associated mortality180. The CCP score was shown to predict survival and recurrence independently from clinical characteristics previously implemented in the Karakiewicz nomogram, a scoring system developed to predict RCC-specific survival181. Furthermore, the combination of CCP score and Karakiewicz nomogram was used to stratify a study cohort (n = 565), based on RCC-specific mortality, into low-risk (99% survival at 5 years) and high-risk (84% survival at 5 years) groups.

Despite this high-quality research, the overall lack of benefit from adjuvant therapies in RCC remains the biggest barrier to implementation of genomic signatures that might help predict recurrence-free survival. Specific patient populations might benefit from adjuvant treatment and the implementation of genomic signatures in prospective adjuvant studies might help to guide future management algorithms of patients with localized RCC176,178,182.

Metastatic RCC

Combination therapies represent the latest breakthrough in the treatment of metastatic RCC. Treatment with nivolumab and ipilimumab was the first breakthrough and was associated with compelling overall survival benefit over sunitinib (not reached versus 26 months, respectively; hazard ratio = 0.63, P < 0.001) in patients in the intermediate and poor risk International Metastatic Renal Cell Carcinoma Database Consortium categories11. Subsequently, the phase II clinical trial IMmotion150 compared sunitinib with either single-agent atezolizumab or an atezolizumab–bevacizumab combination51. This was followed by the phase III clinical trial IMmotion151, which compared atezolizumab–bevacizumab with sunitinib. The combination provided PFS benefit over sunitinib but did not meet its co-primary end point and failed to demonstrate an overall survival benefit183. A further two studies reported that combinations of axitinib–pembrolizumab14 and axitinib–avelumab13 offered significant clinical benefits (ORR, PFS and overall survival benefit with axitinib–pembrolizumab and ORR and PFS benefit with axitinib–avelumab) compared with sunitinib alone. Given the increasing number of first-line treatment options, investigators have attempted to develop predictive gene expression signatures to help guide treatment choices.

Exploratory work from the IMmotion150 study stratified patients based on genomic data obtained through whole-exome sequencing51. Three unique gene signatures were identified: angiogenesis (that is, expression of VEGFA, KDR, ESM1, PECAM1, ANGPTL4 and CD34), pre-existing T effector immunity (that is, expression of CD8A, EOMES, PRF1, IFNG and CD274) and myeloid inflammation (that is, expression of IL-6, CXCL1, CXCL2, CXCL3, CXCL8 and PTGS2). Tumours with a high angiogenesis signature had the best response to sunitinib, whereas a combination of atezolizumab and bevacizumab was most effective in patients with a high T effector immunity signature. RNA-based analyses of tumour samples obtained from patients who received sunitinib or pazopanib in the COMPARZ trial have also confirmed the presence of distinct molecular subtypes of RCC184. Patients with a high (that is, over the median value) angiogenesis molecular signature, which was defined by a previously published method, had better PFS and overall survival compared with patients with a low angiogenesis signature185. Notably, for patients treated with sunitinib, response rates were highest in those with tumours that had a high angiogenesis signature184. Overall, these findings might represent exciting opportunities to tailor treatment and improve disease outcomes in patients with metastatic RCC51.

Whole-exome and RNA sequencing of over 700 tumours from the JAVELIN RENAL 101 study, in which a combination of axitinib–avelumab improved PFS over sunitinib, were also used to identify potential biomarkers of clinical response186. Gene expression signatures in this study included the previously identified angiogenesis, T effector and T cell inflamed signatures, along with a novel immune-related signature developed to predict which patients would benefit from axitinib–avelumab. Once again, patients with a high angiogenesis signature benefited the most from sunitinib, whereas patients with high pre-existing immune activity, defined by high T effector and T cell inflamed signatures, had a greater PFS benefit from the combination immunotherapy than from sunitinib. Among patients treated with axitinib–avelumab, in those with the novel JAVELIN 101 signature, which comprised 26 genes, PFS was 15.2 months, whereas in patients with signature-negative tumours PFS was 9.8 months (hazard ratio = 0.60). A number of additional gene expression signatures have been developed by different research groups in single-centre or collaborative efforts and this remains an active area of research187,188,189,190,191,192.

Future directions

Current scientific knowledge is based largely on studies of genome-wide sequencing data from tissue samples obtained via surgery or biopsies. TCGA, for example, represents an ambitious and highly reliable research effort to characterize cancer genomics that has led to numerous advances in RCC research. The genomic features of primary tumour tissues have been largely detailed and the next step is to track the natural genomic evolution of RCC throughout the course of the disease — the TRACERx Renal study has provided compelling details of these changes over time20,193,194.

In their initial publication, whole-genome sequencing data from the TRACERx Renal and two other cohorts were collected for 575 primary and 335 metastatic site biopsy samples from 100 patients with ccRCC. The collection of multiple biopsies from each tumour site allowed investigators to study intra-tumour and inter-tumour heterogeneity in detail193. An intra-tumour heterogeneity index was generated based on sequencing data from the primary tumour tissue of patients with >15 primary tumour biopsy samples193,194. Patients with a high intra-tumour heterogeneity index had lower odds of rapid metastatic potential than those with low heterogeneity, independently of chromosomal complexity. Patients with tumours characterized by low intra-tumour heterogeneity and high chromosomal complexity had the highest odds of rapid progression following nephrectomy193,194. The TRACERx Renal trial is ongoing195 and the aim is to recruit a total of 350 patients — data from this study promise to reveal further associations between intra-tumour heterogeneity and survival outcomes. Comparison of genomic findings from primary and metastatic sites also provided insight into both inter-tumour heterogeneity and patterns of metastatic progression. The rates of driver alterations were significantly higher in primary tumour sites than in metastatic sites; only 5% of driver events were exclusively present in tumour metastases. These findings were in alignment with previous reports showing that the genomic features of RCC can differ according to tumour location71,196. For example, one study reported 63–69% discordance between different disease sites within the same patient71.

TRACERx Renal investigators also examined tumour clones based on their presence in the primary or metastatic sites193. Clones that were absent in the primary tissue and present only at subclonal level in metastatic tissue, as well as clones that evolved from subclonal in the primary tissue to clonal in the metastatic tissue, had the highest genome instability index, proliferation index and human leukocyte antigen allelic imbalance. Furthermore, loss of chromosome 9p was significantly more frequent in metastasizing clones than in primary tumour clones; loss of chromosome 14q was also substantially more frequent in clones from metastases193. These chromosomal losses were associated with survival outcomes and patients with chromosome 9p loss had reduced survival compared with those without chromosome 9p loss193. Importantly, the loss of both chromosome 9p and 14q was largely subclonal and could only be detected with genomic profiling of different sites within a tumour. Thus, in whole-genome sequencing studies of a single biopsy sample, failure to detect these chromosomal losses might have limited clinical implications owing to intra-tumour heterogeneity. Intra-tumour and inter-tumour heterogeneity might therefore compromise the relevance of data from NGS studies of single tumour biopsy samples.

Liquid biopsies, which involve the isolation and NGS of cell-free circulating tumour DNA (ctDNA) from blood, are less invasive for patients and can be obtained alongside routine blood draws. Liquid biopsies can reveal genomic changes in tumours over time owing to the ease of the longitudinal and repeated assessments and might help to address some of the limitations of single-biopsy tumour sampling. Of note, a comparison of NGS data obtained from single-biopsy tumour tissue and ctDNA in patients with metastatic RCC revealed that both sample types had a similar number of genomic alterations but that the concordance rate was only 8.6%197. These findings suggest that ctDNA assessment might not accurately reflect tumour composition but might be a useful tool to complement tissue-based NGS. ctDNA is generally detectable in the majority of patients with both localized and metastatic RCC198,199.

An analysis of sequential blood samples collected from 220 patients with metastatic RCC using a 73-gene ctDNA platform revealed significant differences between the ctDNA profile of samples collected during the first and further lines of treatment. For example, in the post-first-line treatment samples, the frequencies of genomic alterations in TP53, VHL, NF1, EGFR and PIK3CA were significantly lower than in the samples obtained during first-line treatment, and this decrease in tumour-associated genomic alterations became even more pronounced in samples collected during further lines of treatment199. This study provides concrete evidence of the changing genomic landscape during treatment and underscores the importance of continued surveillance of genomic findings in order to gain insight into mechanisms of treatment resistance.

Patient-specific factors also frequently predict RCC outcomes. In addition to clinical and pathological factors, emerging evidence indicates that the microbiota of patients with RCC can influence disease progression, responses to treatment and the burden of treatment-related side effects200. Microbiota analysis is performed through amplification and sequencing of bacterial nucleic acids obtained from samples such as stool, urine or saliva. One study reported that the relatively high abundance of Akkermansia muciniphila in stool samples from patients with metastatic RCC being treated with immunotherapies was closely linked to treatment response201. In addition, antibiotic use was associated with shorter PFS. Another report indicated that in patients with metastatic RCC treated with VEGF-targeted therapies, high Bacteroides spp. and low Prevotella spp. levels were associated with increased incidence of treatment-related diarrhoea202.

Furthermore, in patients treated with VEGF-targeted therapies, the use of antibiotics that targeted Bacteroides spp. was associated with longer PFS. Several ongoing studies are investigating the interplay between microbiota, treatment effects and RCC outcomes203. These promising preliminary results suggest that bacterial genome analysis might have increasing value in cancer research and might provide important insights into a potential effect modifier that is distinct from the genomics of the patient and of the tumour.

Conclusions

Genomic profiling has substantially enhanced our understanding of the pathological and genomic complexity of RCC. The role of certain alterations in disease progression and treatment has been clearly identified, but, thus far, the identification of these genomic alterations with clinical significance has not resulted in a major breakthrough in the treatment of RCC. The future of response prediction and prognostication in RCC largely depends on easy-to-perform, accurate and reproducible genomic analysis tools, such as liquid biopsies, and the development of nomograms that combine pathological, clinical, genomic and perhaps microbiome information to guide treatment decision-making.