Renal cell carcinoma (RCC) represents 2% of all

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1 Predictors of Response to Targeted Therapy in Renal Cell Carcinoma Laurie J. Eisengart, MD; Gary R. MacVicar, MD; Ximing J. Yang, MD, PhD N Context. The prognosis for patients with metastatic renal cell carcinoma is poor, with an average 5-year survival of approximately 10%. Use of traditional cytokine therapy, specifically high-dose interleukin 2, is limited by significant toxicity. Better understanding of the molecular pathogenesis of renal cell carcinoma has led to the development of targeted therapies to inhibit specific cellular pathways leading to tumorigenesis. These drugs provide improved survival with a more favorable toxicity profile. There is ongoing investigation of markers that predict response of an individual patient to different targeted therapies. Objective. To explain the molecular basis for vascular endothelial growth factor inhibitor (antiangiogenic) and mammalian target of rapamycin inhibitor therapies for renal cell carcinoma, summarize the clinical trials demonstrating the effectiveness of these drugs, and describe the biomarkers shown to correlate with outcome in patients treated with targeted therapy. Data Sources. All included sources are from peerreviewed journals in PubMed (US National Library of Medicine). Conclusion. Emerging evidence shows promise that biomarkers will be useful for predicting an individual patient s response to targeted therapy, leading to a more personalized approach to treating renal cell carcinoma. (Arch Pathol Lab Med. 2012;136: ; doi: / arpa ra) Renal cell carcinoma (RCC) represents 2% of all malignancies worldwide, and the incidence has risen in the United States in the last 2 decades. 1,2 This increase includes RCC at all stages, suggesting a true rise rather than an artifact of improved imaging for detecting asymptomatic, small tumors. 1 Surgical resection is curative for most patients with localized RCC, but for patients who present with advanced disease or whose disease recurs after surgery, the prognosis is poor. While overall survival rates have improved during the last 6 decades, for patients with metastatic disease, the 5-year survival rate averages 10%. 3 Fortunately, survival may be improved with new therapies for RCC, including molecularly targeted therapies. Traditional cytokine immunotherapy for advanced RCC includes interferon a (IFN-a), low-dose interleukin 2 (IL-2), and high-dose IL-2. In particular, high-dose IL-2 therapy can provoke an immune response against RCC, resulting in lasting, complete remission in approximately 10% of patients. 4 However, use of high-dose IL-2 is limited by its severe toxicity, including hypotension, cardiac arrhythmias, metabolic acidosis, renal failure, neurotoxicity, and dermatologic complications. Patients must receive Accepted for publication July 29, From the Departments of Pathology (Drs Eisengart and Yang) and Medicine, Division of Hematology/Oncology (Dr MacVicar), Northwestern University Feinberg School of Medicine, Chicago, Illinois; and the Robert H. Lurie Comprehensive Cancer Center of Northwestern University, Chicago, Illinois (Drs MacVicar and Yang). The authors have no relevant financial interest in the products or companies described in this article. Reprints: Ximing J. Yang, MD, PhD, Feinberg Pavilion 7-338, Surgical Pathology, Northwestern Memorial Hospital, Northwestern University Feinberg School of Medicine, 251 E Huron St, Chicago, IL ( xyang@northwestern.edu). high-dose IL-2 in a facility that can provide blood pressure support, and only patients with good or intermediate prognosis by Memorial Sloan-Kettering Cancer Center (MSKCC, New York, New York) criteria and excellent organ function are candidates for therapy. 3 As the understanding of the molecular pathogenesis of RCC has improved, new drugs have been developed that target the specific pathways driving tumor growth, specifically, vascular endothelial growth factor (VEGF) driven angiogenesis and the mammalian target of rapamycin (mtor) pathways. Clinical trials have shown that these targeted therapies are effective at improving survival and have a more favorable toxicity profile than traditional cytokine therapy These trials provide some guidance as to which therapy may be most appropriate in a given clinical situation, but discovery and validation of biomarkers that can predict an individual patient s response to a given therapy is an area of active investigation, and many of these predictors are still in the research and investigation phase. However, some trials have shown that the histologic subtype and morphologic features of renal cell carcinoma can be important predictors of response to targeted therapy. This article will discuss the molecular basis of targeted therapies, summarize the clinical trials demonstrating their effectiveness, and review the predictors of response, including histologic and immunohistochemical features that can be used by the practicing pathologist and those that are currently in the investigative phase. VEGF PATHWAY INHIBITORS (ANTIANGIOGENIC AGENTS) Molecular Basis The von Hippel-Lindau (VHL) syndrome is an autosomal dominant disorder characterized by loss of the VHL 490 Arch Pathol Lab Med Vol 136, May 2012 Response to Targeted Agents for Kidney Cancer Eisengart et al

2 tumor suppressor gene located on chromosome 3p25. Clinical manifestations include central nervous system hemangioblastomas, retinal angiomas, and solid tumors, including clear cell RCC. In VHL syndrome, a second hit results in the loss of both copies of the VHL gene in a given cell and leads to tumorigenesis. In sporadic clear cell RCC, biallelic VHL gene loss through either deletion of the short arm of chromosome 3 (3p) or promoter hypermethylation is observed in 70% to 80% of cases. 16,17 The VHL gene encodes a 213 amino acid protein (pvhl) that functions as the substrate recognition component of a ubiquitin ligase targeting the transcription factor hypoxia-inducible factor (HIF) for degradation in the presence of oxygen. 18,19 pvhl recognizes the a subunits HIF-1a and HIF-2a. 19 In hypoxic conditions, or when VHL is lost, HIF is not degraded and travels to the nucleus where it induces transcription of genes to restore normoxia. 19 Of the proteins transcribed, the most important in terms of tumor angiogenesis is VEGF, a member of the platelet-derived growth factor family. Vascular endothelial growth factor binds transmembrane receptors with protein kinase activity that activate cellular pathways to induce proangiogenic changes. 19,20 Targeted Therapies Two approaches have been developed to target the VEGF pathway. The first is the use of small-molecule tyrosine kinase inhibitors such as sunitinib or sorafenib to block the intracellular protein kinase activity of the VEGF receptor. The second is a monoclonal antibody (bevacizumab) that binds circulating VEGF and prevents its binding to and activation of the receptor. Sunitinib. Sunitinib is a tyrosine kinase inhibitor that blocks the VEGF receptor, as well as platelet-derived growth factor receptor, FLT-3, and c-kit. 21 Efficacy of sunitinib was demonstrated in 2 phase II trials of patients with metastatic RCC whose disease had progressed during cytokine therapy, with response rates of 34% to 40% and a median time to progression of 8 months. 13,14 Compared with IFN-a in a phase III trial, sunitinib had an increased response rate (39% versus 8%) and prolonged progression-free survival (PFS) (11 versus 5 months) and overall survival (OS) (26.4 versus 21.8 months). 11,12 As a result, sunitinib is often selected as first-line treatment for patients with advanced RCC, particularly when immunotherapy is not a consideration. Sorafenib. Sorafenib is a tyrosine kinase inhibitor that blocks the VEGF receptor, as well as FLT-3, c-kit, plateletderived growth factor receptor, fibroblast growth factor receptor, C-raf, and B-raf. 22 The phase III Treatment Approaches in Renal Cancer Global Evaluation Trial (TARGET), comparing sorafenib to placebo in patients for whom prior cytokine therapy had failed, showed improved OS for sorafenib when excluding those patients who received sorafenib only after the disease had progressed with placebo. 6 A phase II trial comparing sorafenib to IFN-a as first-line therapy showed greater tumor shrinkage and quality of life for patients receiving sorafenib, but no increase in PFS. Sorafenib dose escalation after disease progression was well tolerated and led to an additional 4 months of PFS. 8 Bevacizumab. Bevacizumab is a monoclonal antibody that binds and neutralizes all circulating isoforms of VEGF with high affinity. 23 A phase II trial of patients with disease progression after cytokine therapy showed a significant increase in PFS for high-dose bevacizumab compared to placebo, but no difference in OS. 15 The phase III AVOREN trial compared bevacizumab plus IFN-a to IFN-a alone as first-line therapy for patients with metastatic RCC. Bevacizumab plus IFN-a produced significantly increased PFS (10.2 versus 5.5 months) and response rate (31% versus 13%), 7 and final analysis showed a nonsignificant trend toward improved survival (23.3 versus 21.3 months). 24 The authors 24 note that poststudy treatment of patients with tyrosine kinase inhibitors had most likely increased their survival, especially for those who had received placebo on trial, confounding the OS results. Predictors of Response Clinical Features. In the drug trials, some pretreatment clinical features have had a statistically significant relationship to outcome in patients treated with VEGF inhibitors. In the phase II trial of sunitinib for patients with unsuccessful cytokine therapy, longer progression freesurvival was observed for patients with normal serum hemoglobin (than those with baseline hemoglobin below the lower limit of normal), with favorable Eastern Cooperative Oncology Group (ECOG) performance status, lack of liver metastasis, and fewer disease sites (0 or 1 versus 2 or more). A trend toward improved OS (P 5.06) was seen for alkaline phosphatase levels lower than 120 U/L. 14 In the phase III trial of sunitinib as first-line therapy, improved OS was seen for patients with favorable ECOG performance status, normal hemoglobin levels, time from diagnosis to treatment of more than 1 year, corrected calcium levels lower than 10 mg/dl, lactate dehydrogenase (LDH) levels less than 1.5 times the upper limit of normal, alkaline phosphatase levels below the upper limit of normal, and 1 (versus 2 or more) metastatic site. 11,12 In the phase III TARGET trial of sorafenib, ECOG performance status and MSKCC score were independent prognostic factors for PFS and OS. In addition, patients older than 70 years had significantly improved PFS without increased toxicity. 25 A study of 120 patients with metastatic clear cell RCC treated with any VEGF inhibitor (sunitinib, sorafenib, or bevacizumab) found 5 independent clinical prognostic factors associated with shorter PFS: time from diagnosis to treatment of less than 2 years, platelet count greater than /ml, neutrophil count greater than /ml, corrected serum calcium levels below 8.5 mg/dl or above 10 mg/dl, and ECOG performance status score greater than Histology. While most studies have included only patients with clear cell histology, there are data to suggest that the subtype of RCC is a useful predictor of response to therapy. In particular, papillary or chromophobe RCC subtype may predict superior response to sunitinib over sorafenib. In a retrospective review of 53 patients with papillary or chromophobe RCC treated with sunitinib or sorafenib, there was an overall response rate of 10%, PFS of 8.6 months, and OS of 19.6 months. Progression-free survival was significantly longer for patients treated with sunitinib than sorafenib (11.9 versus 5.1 months). 27 Data also suggest that evaluation of specific histologic features in clear cell RCC may be helpful in predicting response to VEGF pathway inhibitors. A number of histologic features, including Fuhrman grade, growth pattern, degree of clear cytology versus eosinophilia, and Arch Pathol Lab Med Vol 136, May 2012 Response to Targeted Agents for Kidney Cancer Eisengart et al 491

3 presence of rhabdoid and sarcomatoid features, were evaluated in patients with metastatic clear cell RCC receiving first-line VEGF inhibitor therapy. 28 Multivariate modeling showed that greater tumor shrinkage was associated with a higher clear cell component (P 5.02), presence of rhabdoid features (P 5.06), and light eosinophilic cytology of 50% or less (P 5.05). Greater clear cell component was also associated with partial treatment response (P 5.02) and longer treatment duration (P 5.02) but not OS (P 5.57). A retrospective analysis of 43 patients with metastatic clear cell RCC with sarcomatoid features, treated with VEGF-targeted therapy, compared outcomes by underlying RCC subtype and by percentage of sarcomatoid histology. 29 The overall objective partial response rate was 19%; stable disease was the outcome in 49% of cases and progressive disease was the best response in 33%. In regard to RCC subtype, partial response was only achieved in patients with underlying clear cell RCC. Of these patients, 61% had tumor shrinkage, and they had a median PFS of 6 months and OS of 13.1 months. Of those with papillary, chromophobe, or unclassified RCC, only 30% had any tumor shrinkage. Because of the small sample size of patients with non clear cell histology, there were no statistically significant differences in survival outcomes. A larger proportion of sarcomatoid elements negatively affected response: partial response was only achieved in those with less than 20% sarcomatoid elements (P 5.02). Those with less than 20% sarcomatoid elements showed greater tumor reduction (P 5.05), and PFS and OS favored these patients but was not statistically significant. Those with non clear cell histology had a higher percentage of sarcomatoid elements on average. To clarify the effect of the presence of sarcomatoid elements, the patients with underlying clear cell RCC were matched for age, sex, treatment, and prognostic risk group with patients who had clear cell RCC but no sarcomatoid elements. Those without sarcomatoid elements had longer PFS (P,.001), greater tumor shrinkage (P 5.005), and a greater response rate to therapy (P 5.02). Overall, clear cell RCC may be associated with a better response to VEGF-targeted therapy, but papillary and chromophobe RCC may show a better response to sunitinib than to sorafenib. Sarcomatoid elements are a negative predictor of response. Immunohistochemistry: Carbonic Anhydrase IX Expression. Carbonic anhydrase IX (CAIX) is a transmembrane protein involved in cell proliferation that is upregulated by HIF and is expressed in most clear cell RCCs. 30 Carbonic anhydrase IX expression by immunohistochemistry was evaluated in tumors from 118 patients receiving first-line VEGF inhibitor therapy with either sunitinib or sorafenib. 28 Level of immunohistochemical expression of CAIX was not predictive of OS. However, high CAIX expression, defined as greater than 85% of tumor cells positive for the marker, was associated with a better response rate in the patients treated with sorafenib, suggesting CAIX expression may be a predictive marker for sorafenib. VEGF Levels. Blood levels of VEGF have been shown to be prognostic for patients receiving VEGF tyrosine kinase inhibitors. In a study of 63 patients with metastatic RCC for whom cytokine therapy was unsuccessful, levels of VEGF and of the soluble VEGF receptors svegfr-2 and svegf-3 were monitored during treatment with sunitinib. Levels of VEGF increased in 44% of patients, while svegfr-2 and svegfr-3 levels decreased in 91% and 87% of patients, respectively. For those with measurable changes in these 3 proteins, significantly larger changes compared to baseline were observed for patients showing objective tumor response than for those with stable disease or disease progression. 31 Additionally, the phase III TARGET trial of sorafenib prospectively studied VEGF levels and found that patients with higher baseline ECOG performance status or MSKCC score both predictors of poor prognosis had significantly higher baseline levels of VEGF. Further, higher baseline levels of VEGF were associated with decreased PFS and OS for patients receiving placebo and with decreased OS for patients treated with sorafenib. The data also suggested that while all patients benefitted from sorafenib, patients with high baseline VEGF levels derived more benefit from sorafenib than those with low levels. 6 In contrast, the phase II trial of bevacizumab did not find any association between detectable pretreatment VEGF levels and clinical response or time to progression. The authors 15 note that the limited sensitivity of their VEGF assay prevents making a definitive conclusion. VHL Gene Mutation Status. The type of VHL mutation in RCC may predict response to VEGF targeted therapy. In a study of 123 patients with metastatic clear cell RCC treated with VEGF inhibitors, patients with VHL inactivation did not have a significantly different response rate than those with wild-type VHL. However, loss-offunction mutations of VHL (frameshift, nonsense, splice and in-frame deletions/insertions) were an independent prognostic factor for improved response versus wild-type VHL. 32 Another study of 43 patients treated with VEGF inhibitors found that VHL mutation status did not affect objective response, but those patients with VHL methylation or with mutations that caused truncation or frameshift mutations appeared to have longer time to tumor progression. 33 Cell Cycle and Proliferation Markers. Gene and protein expression of cell cycle and proliferation markers in posttherapy nephrectomy specimens, assayed with reverse-phase protein array and gene expression microarray, were predictive for PFS and OS in a study of 38 patients treated with bevacizumab. 34 High protein level of AMPK, a tumor suppressor that inhibits the proliferation effects of the phosphoinositide 3 kinase (PI3K)/Akt pathway, was associated with longer PFS and OS. Increased protein expression of PTEN, another tumor suppressor of the PI3K/Akt pathway, correlated with a longer OS. In contrast, increased expression of phosphorylated Akt (pakt) and its downstream product, phospho-s6 (ps6), was associated with shorter OS. Additionally, high protein level of cyclin B1, which promotes mitosis, was associated with shorter PFS and OS. With gene expression microarray, down-regulation of genes in the PI3K pathway and genes involved in cell cycle control were correlated with longer PFS. Radiologic Imaging. Contrast-enhanced imaging of tumors, which can assess tumor microvascularity, has been shown to be predictive of response to antiangiogenic therapy. Contrast-enhanced computed tomography of metastatic lesions in patients treated with tyrosine kinase VEGF inhibitors showed that tumor enhancement and enhancement pattern were predictive of objective response, and tumor enhancement was associated with 492 Arch Pathol Lab Med Vol 136, May 2012 Response to Targeted Agents for Kidney Cancer Eisengart et al

4 tumor size reduction, time to size reduction, and PFS. 35 A study of dynamic contrast-enhanced magnetic resonance imaging (MRI) of metastases in patients treated with sorafenib showed that those with a high baseline parameter of volume transfer constant of contrast agent had better PFS. 36 A study currently ongoing at our institution compares diffusion-weighted MRI of primary renal cell carcinomas and abdominal metastases pre- and post-neoadjuvant treatment with sorafenib. Following surgical resection, MRI changes will be correlated with histologic parameters including tumor necrosis. mtor PATHWAY INHIBITORS Molecular Basis Mammalian target of rapamycin, an intracellular serine/ threonine kinase upregulated in many human cancers, is downstream of the PI3K/Akt pathway regulated by the PTEN tumor suppressor gene. 19,37 When activated, mtor phosphorylates protranslation factors and induces translation of messenger RNAs encoding cell cycle regulators that promote proliferation, as well as the messenger RNA for HIF. Targeted Therapies Temsirolimus. Temsirolimus is a parenterally administered rapamycin analog that inhibits the mtor kinase via competitive inhibition. 38 In a phase II trial of patients who had undergone unsuccessful cytokine therapy, or who were not candidates for cytokine therapy, various doses of temsirolimus were investigated. 5 While response rate was low (7%), antitumor activity was shown by the significant fraction of patients with minor responses (26%) and/or stable disease for 6 months or more (17%), and the long time to progression (5.8 months) and median OS of 15 months. 5 A phase III trial of temsirolimus as first-line therapy in poor-prognosis patients compared temsirolimus alone, temsirolimus plus IFN-a, and IFN-a alone. Temsirolimus alone compared to IFN-a alone showed significantly increased OS (10.9 versus 7.3 months) and PFS (3.8 versus 1.9 months or 5.5 versus 3.1 months by site investigator or independent radiologist, respectively). Temsirolimus plus IFN-a was not better than IFN-a alone. As IFN-a alone showed no effect, temsirolimus alone and temsirolimus plus IFN-a were expected to be equivalent. The lack of effect in the temsirolimus plus IFN-a treatment arm may be due to dose reductions of temsirolimus in the combination, necessary because of toxicity. 9 Everolimus. Everolimusisanoralcompetitiveinhibitor of mtor. 38 A phase III trial investigated everolimus in patients whose disease had progressed while taking or within 6 months of stopping treatment with VEGF tyrosine kinase inhibitors; some patients had also received previous VEGF antibodies or cytokine therapy. 10 Compared to placebo, everolimus showed significantly increased PFS (4.0 versus 1.9 months) and stable disease was more common (63% versus 32%). There was no significant difference in OS, but patients were allowed to cross over from placebo to everolimus upon disease progression. 10 Therefore, everolimus is the only agent with phase III data supporting its use in the setting of failed VEGF inhibitor therapy. Predictors of Response Clinical Features. Some clinical features are associated with improved outcomes in patients treated with mtor inhibitors. In the phase III trial of temsirolimus, improved OS was observed for patients younger than 65 years, with Karnofsky performance status score below 70, and with LDH levels more than 1.5 times the upper limit of normal. 9 The phase II trial of temsirolimus found that patients stratified to intermediate and poor prognosis by the clinical criteria for prediction of response to IFN-a had the most benefit from temsirolimus. 5 Histology. As most clinical trials have included only patients with clear cell RCC, or have not specified RCC subtype in their patients, there are limited data available on the effect of RCC subtype and histology on response to mtor pathway inhibitors. However, a subset analysis of the phase III trial of temsirolimus looked at the influence of tumor histology on response to the drug. 39 Most patients included in the trial had clear cell RCC (81%), while 13% of tumors were classified as indeterminate and 6% as non clear cell; of the patients with non clear cell tumor for which the subtype was known, 75% had papillary RCC. For patients with clear cell tumors, median OS and PFS were longer for temsirolimus than IFN, with hazard ratios of 0.82 and 0.76, respectively. For patients with other tumor histologic profiles, median OS and PFS also were longer for temsirolimus, with hazard ratios of 0.49 and 0.38, respectively. These data suggest a good response to temsirolimus even in non clear cell RCC. mtor Pathway Molecule Expression. The levels of mtor pathway products in tissue may predict prognosis. A study using tissue microarrays of primary RCCs, metastatic RCCs, and normal kidney found that levels of phosphorylated mtor, pakt, and p-p70s6k, a downstream mtor signaler, were overexpressed in RCC by both Western blot analysis and immunohistochemistry. 40 An immunohistochemical study using tissue microarrays of nephrectomy specimens evaluated pakt, PTEN, ps6, and p27, a downstream mtor product, and related these to prognosis. 39 High nuclear pakt levels were associated with favorable prognosis, while high cytoplasmic pakt levels were associated with poor prognosis. Expression of both cytoplasmic and nuclear pakt, PTEN, ps6, and cytoplasmic p27 were all predictors of disease-specific survival. In this study, information regarding adjuvant therapy was not available. In another study of patients treated with temsirolimus, immunohistochemical overexpression of ps6 and pakt was associated with response to treatment, and no patient without high ps6 or pakt expression experienced an objective tumor response. 41 However, in this analysis, immunohistochemical expression of CAIX did not correlate with tumor response. 41 Another study 42 used immunohistochemistry to evaluate baseline PTEN and HIF-1a expression in pretreatment tumor specimens and subsequent response to temsirolimus, and found no correlation with PFS or OS. VHL Gene Mutation Status. VHL mutation status was evaluated in patients treated with temsirolimus and was not found to correlate with response to therapy. 41 HIF Subunit Expression. The 2 types of HIF subunits recognized by pvhl, HIF-1a, and HIF-2a show overlapping but distinct characteristics. 43 HIF-1a plays an important role in activation of genes involved in glucose metabolism and inhibits the oncoprotein c-myc. HIF-2a stimulates expression of the stem cell marker Oct-4 and promotes c-myc mediated cell cycle progression. HIF-1a remains susceptible to degradation in the absence of pvhl, whereas HIF-2a does not. 44 Importantly, HIF-1a Arch Pathol Lab Med Vol 136, May 2012 Response to Targeted Agents for Kidney Cancer Eisengart et al 493

5 synthesis is more sensitive to mtor inhibitors than is HIF-2a. 45 In a study of 160 clear cell RCCs with VHL inactivation, gene expression microarray found 2 subsets of tumors: those that expressed both HIF-1a and HIF-2a (H1H2), and those that expressed only HIF-2a (H2). 43 The H1H2 tumors showed increased Akt/mTOR activation, while H2 tumors showed increased c-myc expression. 43 These findings suggest possible differences in susceptibility to treatment for H1H2 versus H2 tumors, and future studies will be valuable for correlation with clinical outcomes. ROLE OF THE PATHOLOGIST Precise diagnosis of renal tumors is important in selection of therapy, whether traditional or targeted. While most clinical trials have targeted clear cell RCC, accurate subclassification is important, as some other subtypes have been shown to respond to particular therapies, and for use in future studies of the efficacy of these and other therapies for each subtype of RCC. The World Health Organization classification of renal tumors divides renal epithelial tumors into conventional (clear cell) RCC, papillary RCC, chromophobe RCC, as well as the benign lesion oncocytoma, and other more rare carcinomas. 46 When the morphology is classic, classification of a given tumor can be straightforward. However, renal tumors often do not adhere to classic morphology. Histologic heterogeneity within a lesion is common, including not only variable nuclear grade but also architectural and cytologic features including degree of cytoplasmic eosinophilia. Further, there can be significant overlap in histologic features among these lesions. In a given area of a renal tumor, the differential diagnosis based on morphology can be broad and include both benign and malignant lesions. Familiarity with and judicious use of a basic panel of immunohistochemical stains can clarify the diagnosis in most cases. For example, a panel might include carbonic anhydrase 9 for clear cell RCC, 30 a-methylacyl-coa racemase for papillary RCC, 47 cytokeratin 7 for papillary RCC and chromophobe RCC, 30,39 50 and c-kit for chromophobe RCC and oncocytoma. 51,52 Morphology and immunohistochemistry will resolve the subtype in most RCCs encountered in practice, but newer molecular techniques are also available to aid in subtyping difficult tumors. Gene expression profiles have been able to resolve renal tumors into subtypes that correspond with the histologic classification This technique, which uses clustering analysis to group tumors based on gene expression in complementary DNA microarrays, not only allows a molecular classification of renal tumors but also identifies candidate immunohistochemical markers when a particular gene is uniquely overexpressed in a given tumor subtype The gene expression profile alone may also serve as a predictive marker for outcome in RCC. 57 Accurate classification of tumors will be of increasing importance as more targeted therapies are developed, so that the therapy most effective in a given tumor subtype can be administered for each patient. CONCLUSION As the understanding of the molecular biology of RCC has advanced, therapies targeted to specific pathways of tumorigenesis have offered improved survival, with less toxicity than traditional cytokine therapy, to patients with advanced RCC. Discovery of biomarkers predictive of an individual patient s response to a given targeted therapy is an area of active investigation that promises to better personalize cancer treatment in the future. It is important to note that renal cell carcinoma is composed of a wide spectrum of subtypes based on their morphologic, biochemical, and molecular genetic background. 53,54 Their responses to a specific therapy could be widely different because of their molecular variability. Most current therapies are targeting only the most common type of RCC, clear cell type. 58 The therapeutic experience with other subtypes of RCC, such as papillary RCC, is limited. 27,54,59 In the future, the stratification of subtypes of RCC and evaluation of responses of patients with different subtypes of RCC to targeted therapies will be critical for success of the targeted therapy. References 1. Lipworth L, Tarone RE, McLaughlin JK. The epidemiology of renal cell carcinoma. J Urol. 2006;176(6, pt 1): McLaughlin JK, Lipworth L, Tarone RE. Epidemiologic aspects of renal cell carcinoma. Semin Oncol. 2006;33(5): Motzer RJ, Mazumdar M, Bacik J, Berg W, Amsterdam A, Ferrara J. Survival and prognostic stratification of 670 patients with advanced renal cell carcinoma. J Clin Oncol. 1999;17(8): Belldegrun AS, Klatte T, Shuch B, et al. Cancer-specific survival outcomes among patients treated during the cytokine era of kidney cancer ( ): a benchmark for emerging targeted cancer therapies. Cancer. 2008;113(9): Atkins MB, Hidalgo M, Stadler WM, et al. Randomized phase II study of multiple dose levels of CCI-779, a novel mammalian target of rapamycin kinase inhibitor, in patients with advanced refractory renal cell carcinoma. J Clin Oncol. 2004;22(5): Escudier B, Eisen T, Stadler WM, et al. Sorafenib for treatment of renal cell carcinoma: final efficacy and safety results of the phase III Treatment Approaches in Renal Cancer Global Evaluation Trial. J Clin Oncol. 2009;27(20): Escudier B, Pluzanska A, Koralewski P, et al. Bevacizumab plus interferon alfa-2a for treatment of metastatic renal cell carcinoma: a randomised, doubleblind phase III trial. Lancet. 2007;370(9605): Escudier B, Szczylik C, Hutson TE, et al. Randomized phase II trial of firstline treatment with sorafenib versus interferon Alfa-2a in patients with metastatic renal cell carcinoma. J Clin Oncol. 2009;27(8): Hudes G, Carducci M, Tomczak P, et al. Temsirolimus, interferon alfa, or both for advanced renal-cell carcinoma. N Engl J Med. 2007;356(22): Motzer RJ, Escudier B, Oudard S, et al. Efficacy of everolimus in advanced renal cell carcinoma: a double-blind, randomised, placebo-controlled phase III trial. Lancet. 2008;372(9637): Motzer RJ, Hutson TE, Tomczak P, et al. Overall survival and updated results for sunitinib compared with interferon alfa in patients with metastatic renal cell carcinoma. J Clin Oncol. 2009;27(22): Motzer RJ, Hutson TE, Tomczak P, et al. Sunitinib versus interferon alfa in metastatic renal-cell carcinoma. N Engl J Med. 2007;356(2): Motzer RJ, Michaelson MD, Redman BG, et al. Activity of SU11248, a multitargeted inhibitor of vascular endothelial growth factor receptor and platelet-derived growth factor receptor, in patients with metastatic renal cell carcinoma. J Clin Oncol. 2006;24(1): Motzer RJ, Rini BI, Bukowski RM, et al. Sunitinib in patients with metastatic renal cell carcinoma. JAMA. 2006;295(21): Yang JC, Haworth L, Sherry RM, et al. A randomized trial of bevacizumab, an anti-vascular endothelial growth factor antibody, for metastatic renal cancer. N Engl J Med. 2003;349(5): Herman JG, Latif F, Weng Y, et al. Silencing of the VHL tumor-suppressor gene by DNA methylation in renal carcinoma. Proc Natl Acad Sci U S A. 1994; 91(21): Shuin T, Kondo K, Torigoe S, et al. Frequent somatic mutations and loss of heterozygosity of the von Hippel-Lindau tumor suppressor gene in primary human renal cell carcinomas. Cancer Res. 1994;54(11): Kaelin WG Jr. Molecular basis of the VHL hereditary cancer syndrome. Nat Rev Cancer. 2002;2(9): Suarez C, Morales R, Munoz E, Rodon J, Valverde CM, Carles J. Molecular basis for the treatment of renal cell carcinoma. Clin Transl Oncol. 2010;12(1): Breen EC. VEGF in biological control. J Cell Biochem. 2007;102(6): Mendel DB, Laird AD, Xin X, et al. In vivo antitumor activity of SU11248, a novel tyrosine kinase inhibitor targeting vascular endothelial growth factor and platelet-derived growth factor receptors: determination of a pharmacokinetic/ pharmacodynamic relationship. Clin Cancer Res. 2003;9(1): Wilhelm SM, Carter C, Tang L, et al. BAY exhibits broad spectrum oral antitumor activity and targets the RAF/MEK/ERK pathway and 494 Arch Pathol Lab Med Vol 136, May 2012 Response to Targeted Agents for Kidney Cancer Eisengart et al

6 receptor tyrosine kinases involved in tumor progression and angiogenesis. Cancer Res. 2004;64(19): Ferrara N, Hillan KJ, Gerber HP, Novotny W. Discovery and development of bevacizumab, an anti-vegf antibody for treating cancer. Nat Rev Drug Discov. 2004;3(5): Escudier B, Bellmunt J, Negrier S, et al. Phase III trial of bevacizumab plus interferon alfa-2a in patients with metastatic renal cell carcinoma (AVOREN): final analysis of overall survival. J Clin Oncol. 2010;28(13): Eisen T, Oudard S, Szczylik C, et al. Sorafenib for older patients with renal cell carcinoma: subset analysis from a randomized trial. J Natl Cancer Inst. 2008; 100(20): Choueiri TK, Garcia JA, Elson P, et al. Clinical factors associated with outcome in patients with metastatic clear-cell renal cell carcinoma treated with vascular endothelial growth factor-targeted therapy. Cancer. 2007;110(3): Choueiri TK, Plantade A, Elson P, et al. Efficacy of sunitinib and sorafenib in metastatic papillary and chromophobe renal cell carcinoma. J Clin Oncol. 2008;26(1): Choueiri TK, Regan MM, Rosenberg JE, et al. Carbonic anhydrase IX and pathological features as predictors of outcome in patients with metastatic clearcell renal cell carcinoma receiving vascular endothelial growth factor-targeted therapy. BJU Int. 2010;106(6): Golshayan AR, George S, Heng DY, et al. Metastatic sarcomatoid renal cell carcinoma treated with vascular endothelial growth factor-targeted therapy. J Clin Oncol. 2009;27(2): Liao SY, Aurelio ON, Jan K, Zavada J, Stanbridge EJ. Identification of the MN/CA9 protein as a reliable diagnostic biomarker of clear cell carcinoma of the kidney. Cancer Res. 1997;57(14): Deprimo SE, Bello CL, Smeraglia J, et al. Circulating protein biomarkers of pharmacodynamic activity of sunitinib in patients with metastatic renal cell carcinoma: modulation of VEGF and VEGF-related proteins. J Transl Med. 2007; 5(7): Choueiri TK, Vaziri SA, Jaeger E, et al. von Hippel-Lindau gene status and response to vascular endothelial growth factor targeted therapy for metastatic clear cell renal cell carcinoma [discussion in J Urol. 2008;180(3): ]. J Urol. 2008;180(3): Rini BI, Jaeger E, Weinberg V, et al. Clinical response to therapy targeted at vascular endothelial growth factor in metastatic renal cell carcinoma: impact of patient characteristics and Von Hippel-Lindau gene status. BJU Int. 2006;98(4): Tsavachidou-Fenner D, Tannir N, Tamboli P, et al. Gene and protein expression markers of response to combined antiangiogenic and epidermal growth factor targeted therapy in renal cell carcinoma. Ann Oncol. 2010;21(8): Han KS, Jung DC, Choi HJ, et al. Pretreatment assessment of tumor enhancement on contrast-enhanced computed tomography as a potential predictor of treatment outcome in metastatic renal cell carcinoma patients receiving antiangiogenic therapy. Cancer. 2010;116(10): Hahn OM, Yang C, Medved M, et al. Dynamic contrast-enhanced magnetic resonance imaging pharmacodynamic biomarker study of sorafenib in metastatic renal carcinoma. J Clin Oncol. 2008;26(28): Sarbassov DD, Ali SM, Sabatini DM. Growing roles for the mtor pathway. Curr Opin Cell Biol. 2005;17(6): Huang S, Houghton PJ. Inhibitors of mammalian target of rapamycin as novel antitumor agents: from bench to clinic. Curr Opin Investig Drugs. 2002; 3(2): Dutcher J, Szczylik C, Tannir R, et al. Correlation of survival with tumor histology, age, and prognostic risk group for previously untreated patients wtih advanced renal cell carcinoma (adv RCC) receiving temsirolimus (TEMSR) or interferon-alpha (IFN) [abstract]. J Clin Oncol. 2007;25:243s. 40. Lin F, Zhang PL, Yang XJ, Prichard JW, Lun M, Brown RE. Morphoproteomic and molecular concomitants of an overexpressed and activated mtor pathway in renal cell carcinomas. Ann Clin Lab Sci. 2006;36(3): Cho D, Signoretti S, Dabora S, et al. Potential histologic and molecular predictors of response to temsirolimus in patients with advanced renal cell carcinoma. Clin Genitourin Cancer. 2007;5(6): Figlin RA, de Souza P, McDermott D, et al. Analysis of PTEN and HIF- 1alpha and correlation with efficacy in patients with advanced renal cell carcinoma treated with temsirolimus versus interferon-alpha. Cancer. 2009; 115(16): Gordan JD, Lal P, Dondeti VR, et al. HIF-alpha effects on c-myc distinguish two subtypes of sporadic VHL-deficient clear cell renal carcinoma. Cancer Cell. 2008;14(6): Kaelin WG Jr. Kidney cancer: now available in a new flavor. Cancer Cell. 2008;14(6): Toschi A, Lee E, Gadir N, Ohh M, Foster DA. Differential dependence of hypoxia-inducible factors 1 alpha and 2 alpha on mtorc1 and mtorc2. J Biol Chem. 2008;283(50): Eble JN, Sauter G, Epstein JI, Sesterhenn IA. Pathology and Genetics of Tumors of the Urinary System and Male Genital Organs. Lyon, France: IARC Press; World Health Organization Classification of Tumours; vol Tretiakova MS, Sahoo S, Takahashi M, et al. Expression of alphamethylacyl-coa racemase in papillary renal cell carcinoma. Am J Surg Pathol. 2004;28(1): Adley BP, Papavero V, Sugimura J, et al. Diagnostic value of cytokeratin 7 and parvalbumin in differentiating chromophobe renal cell carcinoma from renal oncocytoma. Anal Quant Cytol Histol. 2006;28(4): Memeo L, Jhang J, Assaad AM, et al. Immunohistochemical analysis for cytokeratin 7, KIT, and PAX2: value in the differential diagnosis of chromophobe cell carcinoma. Am J Clin Pathol. 2007;127(2): Skinnider BF, Folpe AL, Hennigar RA, et al. Distribution of cytokeratins and vimentin in adult renal neoplasms and normal renal tissue: potential utility of a cytokeratin antibody panel in the differential diagnosis of renal tumors. Am J Surg Pathol. 2005;29(6): Huo L, Sugimura J, Tretiakova MS, et al. C-kit expression in renal oncocytomas and chromophobe renal cell carcinomas. Hum Pathol. 2005;36(3): Petit A, Castillo M, Santos M, et al. KIT expression in chromophobe renal cell carcinoma: comparative immunohistochemical analysis of KIT expression in different renal cell neoplasms. Am J Surg Pathol. 2004;28(5): Takahashi M, Yang XJ, Sugimura J, et al. Molecular subclassification of kidney tumors and the discovery of new diagnostic markers. Oncogene. 2003; 22(43): Yang XJ, Tan MH, Kim HL, et al. A molecular classification of papillary renal cell carcinoma. Cancer Res. 2005;65(13): Schuetz AN, Yin-Goen Q, Amin MB, et al. Molecular classification of renal tumors by gene expression profiling. J Mol Diagn. 2005;7(2): Young AN, Amin MB, Moreno CS, et al. Expression profiling of renal epithelial neoplasms: a method for tumor classification and discovery of diagnostic molecular markers. Am J Pathol. 2001;158(5): Brannon AR, Reddy A, Seiler M, et al. Molecular stratification of clear cell renal cell carcinoma by consensus clustering reveals distinct subtypes and survival patterns. Genes Cancer. 2010;1(2): Meeks JJ, Kasper KA, Yang X, Kuzel TM, Smith ND. Metastatic renal cell carcinoma with partial response to sunitinib complicated by ureteral obstruction from necrotic tumor. Urology. 2009;73(2):444.e e12. doi: / j.urology Schaeffer EM, Guzzo TJ, Furge KA, et al. Renal medullary carcinoma: molecular, pathological and clinical evidence for treatment with topoisomeraseinhibiting therapy. BJU Int. 2010;106(1): Arch Pathol Lab Med Vol 136, May 2012 Response to Targeted Agents for Kidney Cancer Eisengart et al 495