Several trials have attempted to combine targeted agents with traditional immunotherapies. The combination of bevacizumab and IFN-α is the only approved combination therapy for the treatment of mRCC. Two phase III trials confirmed the activity of the combination of bevacizumab and IFN-α in patients with clear cell mRCC. In the USA, CALGB 90206 was a two-arm open-label study in which patients with clear cell mRCC without prior systemic therapy were randomized to either bevacizumab plus IFN-α or IFN-α monotherapy. The median PFS in the combination group was 8.4 months versus 4.9 months in the monotherapy group (P = 0.0001). OS favored the bevacizumab plus IFN-α arm but was not statistically significant (18.3 months for patients in the combination group vs. 17.4 months in the IFN-α monotherapy group; P = 0.097). Increased fatigue, anorexia, hypertension, and proteinuria were noted in the combination group [23].

In Europe, the AVOREN trial, a blinded and placebo-controlled study, randomized patients with previously untreated mRCC and prior nephrectomy to receive bevacizumab plus IFN-α or IFN-α plus placebo. Median PFS was significantly longer in the bevacizumab plus IFN-α group than in the control group (10.2 months vs. 5.4 months; HR, 0.63; 95 % CI, 0.52–0.75; P = 0.0001). Median OS was 23.3 months with bevacizumab plus IFN-α and 21.3 months with IFN-α plus placebo (HR, 0.91; 95 % CI, 0.76–1.10; P = 0.3360). The results of this study supported the approval of combination bevacizumab plus IFN-α for the treatment for mRCC by both the US FDA and the European Medicines Agency (EMEA) [27, 41].

Given the success of combining bevacizumab and IFN-α, several studies have investigated combining other targeted agents with IFN-α.

The combination of sorafenib and IFN-α has been investigated in several trials. In a phase I trial, Niwakawa et al. combined sorafenib with IFN-α. After 2 weeks of single-agent IFN-α, 18 patients were treated with 28-day cycles of continuous sorafenib 200 mg (cohort 1) or 400 mg (cohorts 2 and 3) twice daily combined with intramuscular IFN-α six million international units (mu) (cohorts 1 and 2) or 9 mu (cohort 3) both three times a week. Five patients had confirmed PR and 11 had SD (response rate = 27.8 %). These researchers noted that five patients had dose-limiting toxicities, most commonly fatigue. All 18 patients treated with this combination experienced at least one treatment-related adverse event, including fatigue, fever, cytopenias, weight loss, and decreased appetite [28]. In a phase II study, Ryan et al. evaluated response to sorafenib plus IFN-α in 62 patients. Response rates in the combination therapy group were higher than expected for either drug alone (19 % of patients achieved a PR and an additional 50 % had an unconfirmed PR or SD). Despite high response rates, higher levels of toxicity necessitated dose reductions and limited therapy. The most common toxicities noted included fatigue, anorexia, anemia, diarrhea, nausea, rigors/chills, leukopenia, fever, and transaminase elevation [24]. A similar trial by Gollob and colleagues found comparable results, with a response rate of 33 % (13 of 40 patients) noted. Five percent of patients achieved a CR, but overall increased toxicities led to dose reductions and breaks between cycles [30]. Two different schedules of IFN combined with sorafenib were compared in patients with previously untreated mRCC by Bracarda et al. in the phase II RAPSODY trial. Patients (n = 101) were randomized to receive sorafenib 400 mg oral twice daily plus subcutaneous IFN, nine million units (MU) three times a week (arm A) or 3 MU five times a week (arm B). The median PFS was 7.9 months in arm A and 8.6 months in arm B (P = 0.049). In arm A, 9 PRs were observed and 3 CRs and 14 PRs were observed in arm B (17.6 % vs. 34.0 %; P = 0.058); 24 and 21 patients (47 % and 42 %), respectively, achieved SD. Common grade 3–4 toxicities were fatigue plus asthenia (28 % vs. 16 %; P = 0.32) and hand-foot skin reactions (20 % vs. 18 %) [42]. Jonasch et al. compared sorafenib versus sorafenib plus IFN-α in 80 patients with mRCC. They reported an ORR of 30 % (95 % CI, 16.6–46.5 %) in the sorafenib arm compared to a 25 % ORR (95 % CI, 12.7–41.2 %) in the combination arm. A median PFS of 7.39 months was observed in the single-agent sorafenib arm (95 % CI, 5.52–9.20 months) versus a PFS of 7.56 months noted in the sorafenib plus IFN-α arm (95 % CI, 5.19–11.07 months). Toxicities were comparable in both arms, leading the researchers to conclude the outcomes among the two study groups were similar [32].

Shek et al. reported the results of a phase II trial in which they combined gefitinib, an EGFR inhibitor, with pegylated IFN-α, in patients with previously treated mRCC. Twenty-one patients received pegylated IFN-α subcutaneously once weekly (initially 6 μg/kg/week and later reduced to 4 μg/kg/week) for 12 weeks and gefitinib 250 mg orally once daily until disease progression or intolerance. A 6-month PFS rate of 29 % and OS of 13.6 months were reported. The toxicities most commonly noted were myelosuppression, rash, and nausea. The study did not meet the prespecified 6-month PFS rate >50 %, although the authors noted that molecular screenings prior to therapy may identify patients who would benefit from this therapy [26].

In a phase III trial, Hudes et al. randomized patients with previously untreated poor-prognosis mRCC to receive 25 mg of intravenous temsirolimus weekly, 3 mu of IFN-α (with an increase to 18 mu) subcutaneously three times weekly, or combination therapy with 15 mg of temsirolimus weekly plus 6 mu of IFN-α three times weekly. Patients randomized to the temsirolimus-only arm had longer OS (HR, 0.73; 95 % confidence interval [CI], 0.58–0.92; P = 0.008) than did patients who received IFN-α alone. OS in the combination therapy arm was similar to that of the IFN-α-only arm (HR, 0.96; 95 % CI, 0.76–1.20; P = 0.70) [33].

Several trials have also attempted to combine targeted agents with interleukin-2 (IL-2). In the ROSORC trial, Procopio et al. randomized 128 patients with mRCC to receive oral sorafenib 400 mg twice daily plus subcutaneous IL-2, 4.5 mu five times per week for 6 out of every 8 weeks, or single-agent sorafenib. After the enrollment of 40 patients, the dose of IL-2 had to be reduced in order to improve tolerability. A median PFS of 33 weeks with sorafenib plus IL-2 compared to 30 weeks with sorafenib alone (P = 0.109, median follow-up =27 months) was reported. Median PFS for patients receiving the initial higher dose of IL-2 was 43 weeks as compared to 31 weeks for those receiving the lower dose. Common adverse events included hand-foot syndrome, hypertension, and diarrhea. Serious, grade 3–4 adverse events were reported for 38 % of patients receiving combination therapy and 25 % of patients receiving treatment with the single agent. The researchers concluded that combining sorafenib and IL-2 did not significantly improve efficacy, although a trend toward prolonged PFS was associated with the higher dose of IL-2 [25].

A recent phase II study reported by Dandamudi et al. combined bevacizumab with high-dose IL-2 treatment in patients with mRCC. Patients with predominant clear cell histology, measurable disease, Karnofsky performance status of ≥ 80 %, and adequate end-organ function received IL-2 600,000 IU/kg intravenously every 8 h (maximum 28 doses) during two 5-day cycles on days 1 and 15 each 84-day course. In addition, 2 weeks before initiating IL-2, patients began bevacizumab 10 mg/kg infused every 2 weeks. The median PFS reported was 11.2 months (90 % CI, 5.7–17.7 months) with a 2-year PFS of 18 % (90 % CI, 8–27 %). The regimen did not appear to enhance the durable response of single-agent high-dose IL-2. Response rates were at least as high as single-agent therapy, and toxicities did not exceed those expected from each agent alone [43].

Combining targeted therapy with cytotoxic chemotherapy has also been investigated. The combination of gemcitabine and capecitabine has shown efficacy in mRCC, especially in patients previously treated with immunotherapy or targeted agents [22, 44]. This led researchers to attempt combining gemcitabine and capecitabine with targeted therapies. In a phase II trial, 29 patients received the combination of gemcitabine, capecitabine, and bevacizumab; most patients previously failed treatment with VEGF TKI. Seven patients (24 %) had a PR, with a median PFS of 5.3 months (95 % CI, 3.9–9.9) and median OS of 9.8 months (95 % CI, 6.2–14.9). The regimen was well tolerated, but the trial was ended early because of slow accrual [45]. Jonasch et al. conducted a retrospective review of patients treated with the combination of gemcitabine, capecitabine, and bevacizumab in patients with clear and non-clear mRCC. Of 28 patients studied, 9 (32.14 %) had clear cell histology, and 10 (35.71 %) had sarcomatoid features. Initial treatment doses consisted of gemcitabine at a mean treatment dose of 786 mg/m² every 2 weeks, capecitabine at a mean treatment dose of 2.73 g/day, and bevacizumab at a mean dose of 10 mg/kg every 2 weeks. The authors reported a median PFS of 5.9 months and a median OS of 10.4 months. Among 15 patients who had previous TKI therapy, the median PFS was 6.2 months and median OS was 11.7 months. In patients with sarcomatoid features, median PFS and OS were 3.9 months and 9 months, respectively. Three patients discontinued one or more of the drugs because of adverse reactions [35]. A phase II trial of this combination specifically targeted to patients with sarcomatoid mRCC was reported with 18 patients, 9 of which were alive at last follow-up (median follow-up time was 12.1 months). Five remained on treatment with the gemcitabine, capecitabine, and bevacizumab combination. Dose reductions were required in 12 patients, with common toxicities being hand-foot syndrome (5 patients), fatigue (4 patients), and deep vein thrombosis (2 patients). The estimated median time to treatment failure (TTF) was 5.5 months (95 % CI, 3.7–>12 months) and median OS was 12 months (95 % CI, 9.6–>24 months) [46].

Michaelson et al. reported the results of a phase I trial in which they combined sunitinib with gemcitabine in 34 patients with advanced RCC and other solid tumors. They noted activity of this combination in patients with poor-risk mRCC with five of nine patients achieving a PR [36]. This group has designed a phase II study of this combination in patients with sarcomatoid and/or poor-risk mRCC. This study is ongoing, but not recruiting participants (Combination Sunitinib and Gemcitabine in Sarcomatoid and/or Poor-risk Patients With Metastatic Renal Cell Carcinoma. NCT00556049. http://​clinicaltrials.​gov/​ct2/​show/​NCT00556049?​term=​sunitinib+with+g​emcitabine&​rank=​1).

Targeted therapies rarely induce complete responses in patients with mRCC; therefore, sequential use of targeted therapies has become common practice to prolong PFS and OS. Although it is not known how best to overcome resistance to targeted therapies, combination or sequential therapy, it is apparent that sequential therapy allows for optimal dosing of targeted therapies without the increased toxicity that commonly occurs with combination approaches. Targeting different pathways by sequential therapy may help overcome resistance that has developed from prior targeted therapy. The most effective sequence of targeted therapies is yet to be determined. However, accumulating evidence supports this current practice for patients with mRCC.