Immune checkpoint inhibitors produce durable long-term survival in some patients with advanced melanoma and lung cancer. Better immune targets and combination strategies can harness the immune system by supporting the three elements of a successful T-cell antitumor response: (A) generation of sufficient numbers of antitumor T cells within the lymphoid compartment; (B) effective T-cell trafficking and extravasation out of the lymphoid compartment, through the bloodstream, and into the tumor microenvironment; and (C) T-cell effector function within the tumor microenvironment that is characterized by the ability to bypass immune checkpoints, soluble and metabolic inhibitory factors, and inhibitory cells. Strategies that hold promise include dual immune checkpoint blockade, as well as the combination of immune checkpoint blockade with costimulatory receptor agonists, enhancers of innate immunity, inhibition of indoleamine 2,3-dioxygenase, adoptive T-cell transfer/T-cell engineering, therapeutic vaccines, small-molecule inhibitors, and radiation therapy. Novel, rational clinical trial designs seek to combine targeted agents and one or more immune checkpoint inhibitors, with the goal of producing deep and durable antitumor responses, which thus far have been observed in only a minority of patients.
Introduction
Novel immunotherapy combinations raise the prospect of improving both overall survival (OS) and quality-of-life outcomes for cancer patients. Immune checkpoint blockade targeting programmed death ligand 1 (PD-L1); its receptor, programmed death 1 (PD-1); and/or cytotoxic T-lymphocyte–associated antigen 4 (CTLA-4) causes durable tumor regression, most notably in melanoma and lung cancers, and has demonstrated favorable activity in bladder cancer, head and neck cancer, renal cell carcinoma (RCC), ovarian cancer, and hematologic malignancies.[1-8] This review summarizes promising new targets and immunotherapy combination strategies currently under clinical development.
A Historical Perspective
A century ago, in the first attempts at immunotherapy, Dr. William Coley inoculated cancer patients with dead bacteria and observed several spontaneous remissions.[9] For decades, cytokines such as interleukin (IL)-2 (aldesleukin) and interferon alfa have benefited small subsets of otherwise healthy patients with RCC and melanoma.[10-13] Sipuleucel-T, an autologous active therapeutic vaccine that induces T-cell immune responses against prostatic acid phosphate antigen, improved survival and was approved in 2010 by the US Food and Drug Administration (FDA) for patients with asymptomatic or minimally symptomatic metastatic castration-resistant prostate cancer.[14] Ipilimumab, a monoclonal antibody targeting the CTLA-4 immune checkpoint, improved survival and was approved by the FDA in 2011 for treatment of melanoma.[6,15,16] In 2014, pembrolizumab and nivolumab became the first PD-1 checkpoint inhibitors to be approved for use in melanoma among patients with ipilimumab and BRAF inhibitor–refractory disease.[17,18] Nivolumab was approved in March 2015 for platinum-refractory, metastatic, squamous-cell non–small-cell lung cancer (NSCLC), based upon improved survival when compared with conventional chemotherapy with docetaxel; it was approved for nonsquamous NSCLC in October.[19,20] On September 30, 2015, the FDA granted accelerated approval of nivolumab combined with ipilimumab for BRAF V600 wild-type unresectable or metastatic melanoma, based on randomized controlled data showing improved response rate and progression-free survival (PFS) as well as prolonged duration of response with the combination vs ipilimumab alone (as described in the nivolumab package insert). Nivolumab plus ipilimumab is the first checkpoint inhibitor combination to be approved by the FDA. Based on a favorable response rate, pembrolizumab was approved in October 2015 for platinum-refractory and, if applicable, tyrosine kinase inhibitor (TKI)-refractory, metastatic NSCLC that is PD-L1–positive based on an FDA-approved test (as described in the pembrolizumab package insert).[1] Figure 1 outlines cancer immunotherapy milestones of the last two decades.
The Cancer Immunity Cycle Offers Therapeutic Targets
An emerging hallmark of cancer is immunoevasion-the cancer cell’s ability to avoid destruction by the immune system.[21] The three general categories of immunoevasive mechanisms include: (A) an insufficient number of T cells generated within the lymphoid compartment; (B) an insufficient number of T cells extravasating into the tumor; and (C) inhibition of T cells in the tumor microenvironment (Figure 2). The tumor microenvironment, in turn, offers three main immunoevasive tools: (1) surface membrane proteins that function as immune checkpoints, including PD-1, CTLA-4, lymphocyte-activation gene 3 (LAG-3) protein, T-cell immunoglobulin and mucin domain–containing protein 3 (TIM-3), B- and T-lymphocyte attenuator (BTLA), and the adenosine A2a receptor (A2aR); (2) the relationship between selected soluble factors and metabolic alterations, such as IL-10, transforming growth factor beta, adenosine, indoleamine 2,3-dioxygenase (IDO), and arginase; and (3) inhibitory cells, including cancer-associated fibroblasts (CAFs), regulatory T cells, myeloid-derived suppressor cells (MDSCs), and tumor-associated macrophages.
Immunotherapeutic strategies are under development to disrupt steps in the above-described immunoinvasive mechanisms A to C and generate a tumor-specific immune response (Table 1 and Figure 2). For example, CTLA-4 blockade (immune checkpoint inhibition), agonism of T-cell costimulatory receptors, adoptive T-cell transfer (ACT) of tumor-infiltrating lymphocytes (TILs), and T-cell engineering with chimeric antigen receptor (CAR) T-cell therapy or T-cell receptor gene therapy all increase the antitumor T-cell number in the lymphoid compartment.[22] Radiation, chemotherapy, cytokines, and certain molecularly targeted therapies all increase the influx of T cells into tumors.[22] Additional approaches to boost tumor-targeted immune responses include immune checkpoint inhibition (anti–PD-1, anti–PD-L1, anti–LAG-3, anti-A2aR); suppression of T regulatory cells (Tregs) and MDSCs; and stimulation of innate immune cells, such as natural killer (NK) cells, macrophages, and dendritic cells.
Combining immune checkpoint inhibitors with other immunotherapeutics
Either combining or sequencing immunotherapies with distinct targets is a rational approach for improving efficacy. Combination approaches currently under clinical development include dual checkpoint inhibition, checkpoint inhibition plus agonism of T-cell costimulatory receptors, and checkpoint inhibition plus TIL ACT-strategies that may simultaneously exploit expansion of T cells in the lymphoid compartment (anti–CTLA-4) and antitumor T-cell function at the tumor site (see Figure 2, parts A and C). Immune checkpoint blockade is also being combined with radiation, chemotherapy, and small-molecule inhibitors, acting at the level of trafficking and effector function (see Figure 2, parts B and C). Other combinations focus entirely on the tumor microenvironment. Examples include use of checkpoint blockade plus suppressors of Tregs and MDSCs; and checkpoint blockade plus stimulation of the innate immune cells, such as NK cells, macrophages, and dendritic cells (see Figure 2, part C).
Dual T-cell checkpoint blockade
CTLA-4 (also known as CD152) is expressed on the surface of cytotoxic T cells and competes with the costimulatory receptor CD28 to bind to B7-1 (CD80) or B7-2 (CD86) on the antigen-presenting cell. Signaling through CTLA-4 produces an inhibitory signal in T cells. Blocking CTLA-4 with an antibody prevents this negative signaling in tumor cell–specific T cells within lymph nodes, expanding the number of antitumor T cells as well as the breadth of their repertoire.[23-25] If the T cell is primed and migrates to the site of effector function, then it must still go on to evade the tumor’s immune checkpoint ligands.[26] One such checkpoint ligand is PD-L1, which binds to the T-cell surface checkpoint protein PD-1. When PD-1 on the effector T cell interacts with PD-L1 on either the tumor cell or the myeloid cells, immune tolerance develops.[27] When this interaction is blocked, the T cell is freed to perform its function.[26,28,29] Dual blockade with both CTLA-4 inhibition and PD-1/PD-L1 inhibition targets immune evasion at the site of both priming/activation (lymph nodes) and effector function (tumor parenchyma).
Single-agent blockades of CTLA-4, PD-1, and PD-L1 have been active across several tumor types.[7,8,16,18] In melanoma, the antitumoral activity of checkpoint inhibition has translated into improved long-term survival. A phase III study of ipilimumab in patients with metastatic melanoma showed an OS rate of 25% at 2 and 3 years.[30] A pooled analysis of 12 ipilimumab trials showed that the OS plateau of 22% began at 3 years and extended up to 10 years in some patients.[31] Immunotherapy combinations offer the prospect of long-term survival for increased numbers of patients with melanoma and other cancers.
The first clinical data to support dual checkpoint blockade came from a phase I study of ipilimumab (1 mg/kg or 3 mg/kg) and nivolumab (0.3 mg/kg, 1 mg/kg, or 3 mg/kg) either concurrently or sequentially in patients with stage III or IV melanoma.[6] In the combination group, the objective response rate (ORR) was 40%, with grade 3/4 treatment-related adverse events occurring in 53% of patients. At the maximum tolerated dose (MTD) (nivolumab at 1 mg/kg and ipilimumab at 3 mg/kg), the ORR was 53% and tumor volume reductions of 80% or more were observed among the responders. The responses appeared to be more rapid and deeper than those reported previously. In the sequential therapy group, the ORR was 20%, and 18% of patients had grade 3/4 adverse events related to therapy.
A phase III trial (CheckMate 067) is evaluating concurrent ipilimumab and nivolumab vs monotherapy with either drug in patients with untreated, unresectable stage III or IV melanoma. The median PFS was superior in both the combination arm (11.5 months [95% confidence interval (CI), 8.9–16.7]; hazard ratio [HR] for disease progression or death, 0.42 [99.5% CI, 0.31–0.57; P < .001]) and the nivolumab arm (6.9 months [95% CI, 4.3–9.5]; HR, 0.57 [99.5% CI, 0.43–0.76; P < .001]), when compared with the ipilimumab arm (2.9 months [95% CI, 2.8–3.4]). The study was not designed to detect statistical differences between the nivolumab-containing arms. In patients with PD-L1–positive tumors, the PFS was 14 months in both the combination and nivolumab groups. However, in patients whose tumors did not express PD-L1, the combination group had an improved PFS compared with patients treated with nivolumab alone: 11.2 months (95% CI, 8.0 to not reached) vs 5.3 months (95% CI, 2.8–7.1). Grade 3/4 adverse effects related to therapy occurred in 16.3% of patients in the nivolumab group, 27.3% of patients in the ipilimumab group, and 55.0% of patients in the combination group. Longer follow-up is needed to determine the impact on OS.
A phase I, open-label, dose-escalation and expansion study evaluating durvalumab and tremelimumab in advanced solid tumors showed a 27% response rate (95% CI, 13–46) in PD-L1–negative patients, with a disease control rate of 48% (95% CI, 31–66) at ≥ 16 weeks after therapy.[32] Forty percent of patients experienced grade 3/4 toxicities, most frequently colitis, and 20% of patients had to discontinue treatment due to drug-related adverse events. Increasing doses of tremelimumab were associated with higher rates of serious adverse events. Durvalumab at 20 mg/kg every 4 weeks plus tremelimumab at 1 mg/kg every 4 weeks was the dose level chosen for phase III development. The phase I study showed that at this dose level, toxicity leading to discontinuation was < 10%, but lower tremelimumab dosing did not erode clinical efficacy. Notably, anti–PD-1/PD-L1 monotherapy produces an approximately 5% to 10% response rate in PD-L1–negative patients; thus, the addition of low-dose anti–CTLA-4 therapy may benefit these patients.[1]
Although small-cell lung cancer (SCLC) is chemosensitive, responses to both first- and second-line chemotherapy are short-lived and outcomes remain poor.[33,34] CheckMate 032 (ClinicalTrials.gov identifier: NCT01928394) is an ongoing, open-label, phase I/II study of nivolumab with or without concurrent ipilimumab in patients with recurrent SCLC. Patients were randomized to two arms: nivolumab at 3 mg/kg every 2 weeks or nivolumab plus ipilimumab in three dose cohorts (1 mg/kg + 1 mg/kg, 1 mg/kg + 3 mg/kg, or 3 mg/kg + 1 mg/kg) every 3 weeks for 4 cycles, followed by maintenance nivolumab (3 mg/kg every 2 weeks). The primary endpoint was ORR by Response Evaluation Criteria In Solid Tumors (RECIST) v1.1. Grade 3/4 drug-related adverse events occurred in at least 5% of patients and included diarrhea and rash (6% each in the combination arm). Pneumonitis occurred in one patient per arm, and one patient died as a result of drug-related myasthenia gravis.
Updated interim data for 40 patients in the nivolumab arm and 46 patients in the combination arm were presented in June 2015, and showed an ORR of 32.6% in the nivolumab-plus-ipilimumab arm vs 18% in the nivolumab arm. One patient in the nivolumab plus ipilimumab arm had a complete response. Responses occurred in patients regardless of PD-L1 expression, and there was a trend towards increased OS in the nivolumab-plus-ipilimumab arm (8.2 months [95% CI, 3.7 to not reached]) vs the nivolumab-alone arm (4.4 months [95% CI, 2.9–9.4]).[35] The combination of nivolumab and ipilimumab is also being studied in other cancers (Table 2), including a phase I study in metastatic RCC (ClinicalTrials.gov identifier: NCT01472081), with interim results showing an ORR of 29% (6 of 21 patients) in the group receiving nivolumab at 3 mg/kg plus ipilimumab at 1 mg/kg, and an an ORR of 39% (9 of 23 patients) in the group treated with nivolumab at 1 mg/kg plus ipilimumab at 3 mg/kg.[36]
In September 2015, updated results from CheckMate 012 evaluating the safety and efficacy of first-line nivolumab monotherapy vs nivolumab-based combinations in advanced NSCLC showed that nivolumab at 3 mg/kg every 2 weeks plus ipilimumab at 1 mg/kg every 12 weeks resulted in a response rate of 39% (n = 38) and a disease control rate of 74% (95% CI, 57–87).[37] There was a very low frequency of treatment-related grade 3/4 adverse events leading to discontinuation in this dose cohort, again indicating that low-dose, less frequent anti–CTLA-4 therapy combined with anti–PD-1 therapy may produce optimal clinical results, even in the first-line setting.
Another T-cell surface checkpoint protein is the A2aR. The Warburg effect, a metabolic abnormality in which cancer cells exhibit increased rates of glycolysis and lactic acid fermentation; hypoxia in the tumor microenvironment; and CD73 on both tumor cells and CAFs all lead to elevated adenosine levels in the tumor milieu.[38] Adenosine signaling via the A2aR directly inhibits T cells in addition to supporting the growth of CAFs, which directly inhibit killing of tumor cells by CD8+ T cells (cytotoxic T cells). PBF-509, an A2aR antagonist entering phase I testing in NSCLC (ClinicalTrials.gov identifier: NCT02403193), will be studied in combination with other checkpoint inhibitors in the near future.
Phase I studies are currently evaluating a host of dual checkpoint blockade combinations, including ipilimumab plus nivolumab, ipilimumab plus pembrolizumab (anti–PD-1), tremelimumab (anti–CTLA-4) plus durvalumab (anti–PD-L1), and the anti–LAG-3 monoclonal antibody BMS-986016 plus nivolumab (see Table 2).[39,40]
T-cell checkpoint blockade plus costimulatory receptor agonists
Checkpoint inhibition combined with costimulatory receptor agonists may be an opportunity for synergy, but to date this remains theoretical. Selection of costimulatory agonists must be undertaken with cautious optimism, as evidenced by the excessive toxic effects of the CD28 agonist TGN1412 in a first-in-human phase I trial.[41] Six healthy volunteers, 18 to 30 years old, were treated with 0.1 mg/kg of TGN1412, and within 90 minutes each experienced a cytokine storm leading to multi-organ failure and prolonged intensive care. Given that CD28 is expressed on all mature T cells, this antibody agonist was thought to have a “super-agonist” effect.[42] Other costimulatory molecules are expressed on subgroups of T cells and have shown less propensity to cause such dramatic adverse events.
Clinical studies targeting costimulatory receptors with agonistic antibodies are currently in development. CD137 (also called 4-1BB and TNFRSF9 [tumor necrosis factor receptor superfamily, member 9]), glucocorticoid-induced TNFR family-related protein (GITR, also known as TNFRSF18 [TNFR superfamily, member 18]), and CD134 (also called OX40 and TNFRSF4 [TNFR superfamily, member 4]) are costimulatory receptors that promote T-cell proliferation and survival. CD40 (also known as TNFRSF5 [TNFR superfamily, member 5]) stimulates the activation of antigen-presenting cells. CD27 is a costimulatory receptor implicated in multiple functions of T cells, NK cells, and B cells, including long-term memory.[43]
A single-arm, open-label, phase I trial evaluated combined treatment with the CD40 agonist antibody CP-870,893 plus tremelimumab in 24 patients with metastatic melanoma.[44] The ORR was 27.3% and median OS was 26.1 months. The MTD level was CP-870,893 at 0.2 mg/kg every 3 weeks plus tremelimumab at 10 mg/kg every 12 weeks. Two patients (9.1%) had complete responses. Grades 1 and 2 cytokine release syndrome occurred in 19 patients (79.2%) immediately after the administration of CP-870,893, but symptoms resolved within 24 hours with standard supportive care. A host of early-phase checkpoint blockade plus costimulatory receptor agonist clinical trials are underway (see Table 2).
T-cell checkpoint blockade to improve innate immune cell function
Killer cell immunoglobulin-like receptors (KIRs) are cell surface proteins found on NK cells that function to downregulate NK cell activity. In immune physiology, KIRs bind to major histocompatibility complex (MHC) class I molecules expressed on all normal cells to maintain self-tolerance. However, tumor cells also express MHC class I antigens and thereby evade immune surveillance by NK cells. Anti-KIR antibodies can block this immunoevasion, thereby potentiating NK cell effector function and thus innate immunity.[45-48] Lirilumab is an inhibitory anti-KIR antibody that is being studied in combination with ipilimumab in solid tumors and with nivolumab in hematologic malignancies.[49] Given the close interplay between the adaptive and innate immune systems, strategies that enhance cellular components of both response systems may prove efficacious.
Checkpoint blockade plus IDO inhibition
IDO is a heme-containing redox enzyme that suppresses the priming and activation of the adaptive immune response through catabolism of tryptophan in the tumor-draining lymph nodes and tumor microenvironment. Depletion of tryptophan leads to impaired activation of helper and cytotoxic T cells, allowing for control of autoimmunity and maintenance of placental pregnancy.[50-52] Preclinical data have shown that this immunosuppressive mechanism can be co-opted by tumors.[53-55] These data have led to the development of multiple IDO pathway inhibitors, including indoximod, epacadostat (INCB24360), and, most recently, GDC-0919. Indoximod differs from the other two compounds in that it does not efficiently directly inhibit IDO, but rather acts downstream of the enzyme to reduce its effects in the tumor microenvironment.[56] Melanoma mouse models have shown that IDO represses the antitumor effect of anti–CTLA-4 therapy. Blockade of IDO and CTLA-4 reversed this effect and increased antitumor activity.[57,58] Preliminary data from a phase I/II study of this combination have been promising, and blockade of IDO and PD-1 has shown significant clinical activity.[59] Other combination therapies include IDO inhibitors administered along with therapeutic cancer vaccines, chemotherapy, and immune checkpoint–blocking drugs (ClinicalTrials.gov identifiers: NCT02327078, NCT02178722, NCT02318277, NCT02298153, NCT01604889, NCT02073123, and NCT02471846) in phase I and II clinical trials (see Table 2). There is work underway to target a related enzyme, tryptophan 2,3-dioxygenase, which is also upregulated in tumors and may act in a similar fashion to IDO.[60]
Checkpoint blockade plus adoptive T-cell transfer/T-cell engineering
ACT is the process by which TILs are grown out of tumors harvested from patients, expanded in vitro, and reintroduced into the patient.[61] Prior to reinfusion into the patient, CAR T-cell therapy can be used to direct the T cell to specific tumor antigens. A third approach is T-cell receptor gene therapy, in which genes encoding tumor-reactive T-cell receptors are introduced into the patient’s T cells.
In a phase II study, ACT of melanoma TILs plus IL-2 led to complete response in 20 of 93 patients (22%) with metastatic, largely pretreated melanoma; 19 patients had durable complete responses lasting at least 3 years after treatment.[62] However, the lymphodepleting preparative regimen (chemotherapy with or without total body irradiation) caused toxicity, including a treatment-related death in a patient who developed sepsis due to an undetected diverticular abscess. A variety of cancer/testis antigens, such as New York–esophageal cancer–1 (NY-ESO-1) protein and melanoma antigen gene family (MAGE)-A10, have been used to generate tumor-specific T-cell receptors and have shown promising results in melanoma and synovial cell sarcoma.[63] CAR T-cell therapy directed at CD19 and mesothelin has shown positive results in multiple myeloma and solid malignancies, respectively.[64,65] Despite promising early data, toxicity reports included a patient with metastatic colorectal cancer who developed respiratory distress within 15 minutes of treatment with CAR T cells targeting epidermal growth factor receptor 2 (EGFR2; also known as human epidermal growth factor receptor 2 [HER2] and erb-b2 receptor tyrosine kinase 2 [ERBB2]).[66] The patient had multi-organ failure and multiple cardiac arrests in the context of a cytokine release storm, which led to death. One patient treated intermittently with CAR T-meso cells, which express a murine antibody to human mesothelin, developed fatal anaphylaxis and cardiac arrest after the third infusion.[67] CARs are being developed with different antibody fragments and more conservative dose-escalation methods to mitigate toxicities.[68] Unexpected toxicity was also encountered after two patients treated with autologous anti–MAGE-A3 T-cell receptor–engineered T cells developed comas and then died of progressive necrotizing leukoencephalopathy due to previously unrecognized expression of MAGE-A3 in the brain.[69] Early-phase studies utilizing ACT combined with immune checkpoint inhibition are underway, and special care must be taken to attenuate the potential toxicities of these combinations (see Table 2; ClinicalTrials.gov identifier: NCT02210104).
Checkpoint Blockade Plus Small Molecules That Create an Immune-Active Microenvironment
Checkpoint blockade plus histone deacetylase inhibition
Increased tumor expression of T-cell chemokines, such as chemokine (C-C motif) ligand 5 (CCL5) and C-X-C motif chemokine 10 (CXCL10), is associated with a better response to immunotherapy, and is strongly and positively associated with increased T-cell infiltration and improved patient survival.[70-72] Therefore, enhancement of expression of T-cell chemokines by use of small-molecule biologic effectors such as histone deacetylase (HDAC) inhibitors may augment response to PD-1 blockade immunotherapy. HDAC inhibition has generally led to disappointing results when used alone and in combination with chemotherapy in solid tumors. However, recent preclinical studies indicate that HDAC inhibition may contribute to an immune-active environment, making tumor cells more susceptible to anti–PD-1 therapy. Mouse tumor models indicate a synergistic interaction between anti–PD-1 and the HDAC inhibitor, which is entirely T-cell dependent; a phase I/II trial has been designed that will combine an HDAC inhibitor with immune checkpoint blockade.[73]
In a phase I/II trial of 5-azacytidine in combination with entinostat in 19 patients with advanced NSCLC, one patient had a complete response and another had a partial response. Four patients had partial responses after subsequent therapy; one of those patients was treated with anti–PD-1.[74] In a phase I study, five patients were treated with a combination of 5-azacytidine and entinostat prior to being treated with anti–PD-1 or anti–PD-L1 therapy. Three patients had partial responses and two had stable disease by RECIST.[75] Rationally designed clinical trials utilizing concurrent HDAC inhibitors and immune therapy with engrained biomarker analysis will be underway soon (see Table 2).
TO PUT THAT INTO CONTEXT
Michael B. Atkins, MD
Lombardi Comprehensive Cancer Center
Georgetown University
Washington, DC
What Do We Already Know About Immunotherapy Directed at PD-1/PD-L1?
Investigations with high-dose (HD) interleukin-2 (IL-2) have established that activated T cells could produce durable clinical responses and cures in a subset of patients with metastatic melanoma or kidney cancer. Subsequent research also spearheaded by the Surgery Branch of the National Cancer Institute determined that tumors were infiltrated with tumor-infiltrating lymphocytes (TILs) that, when reactivated and expanded ex vivo and readministered following lymphodepleting chemotherapy, could eradicate melanoma in 20% of patients not responsive to HD IL-2. These seminal observations sustained interest in cancer immunotherapy and spawned efforts to reactivate TILs in vivo. These efforts were rewarded by the discovery and targeting of immune checkpoints, such as programmed death 1 (PD-1) and its ligand 1 (PD-L1), which dampen immune responses in the tumor microenvironment.Antibodies against the PD-1/PD-L1 pathway have produced antitumor responses-with little toxicity-not only in patients with advanced melanoma and kidney cancer, but also in at least 18 other tumor types, revolutionizing both immunotherapy and the broader field of cancer therapy. Combining cytotoxic T-lymphocyte–
associated antigen 4 (CTLA-4) and anti–PD-1 antibodies produced antitumor activity superior to anti–PD-1 monotherapy in melanoma, establishing proof of principle that these immunosuppressive mechanisms were not redundant, paving the way for further exploration of this approach and other combination strategies.What Principles Should Guide Rational Development of PD-1 Pathway–Based Combination Therapy? Tchekmedyian et al describe the vast array of PD-1 pathway–based combination strategies under investigation. In prioritizing these options it is important to keep in mind several principles: (1) the greatest impact of immunotherapy is on overall and landmark survival, and use of surrogate endpoints such as progression-free survival will likely underestimate and possibly confound the interpretation of benefit; (2) since T cells recognize neoantigen products of passenger mutations in individual tumors, making each tumor a unique target, efforts to enhance antitumor immunity (eg, vaccination) that do not involve the host tumor will likely be ineffective; (3) TILs are the therapeutic entities, and combination treatments that damage them (eg, chemotherapy, nonselective molecularly targeted therapy) risk sacrificing their curative potential; (4) enhanced toxicity may be acceptable if it is mechanism-related and associated with a boost in frequency of durable responses; (5) subclinical “off-target” organ damage caused by combination treatments will likely be exacerbated by the proinflammatory effects of checkpoint inhibition; (6) bringing more T cells into the tumor through artificial means (eg, destroying natural barriers) may result in accumulation of TILs that lack both tumor specificity and function; and (7) different combination approaches will likely apply to different tumor types and even to different patients with the same tumor type, adding to the complexity of predictive biomarker development.These principles support a focus on rational combination treatment development based on an understanding of tumor immunology and the mechanisms underlying cancer immunotherapy, and favor PD-1 pathway combinations with specific immune activating/modulating agents over those involving other treatment modalities.
Checkpoint blockade plus EGFR TKIs
The relationship between EGFR-sensitizing mutations and response to checkpoint blockade is not clear. Patients with metastatic NSCLC with EGFR-sensitizing mutations treated with pembrolizumab had an ORR of 7.8% (95% CI, 2.9–16.2) vs 21.6% (95% CI, 17.8–25.6) in EGFR wild-type tumors.[76] In addition, a retrospective analysis demonstrated that 0 of 22 patients with EGFR-sensitizing mutations responded to pembrolizumab after prior TKI therapy, whereas patients who were TKI-naive had higher response rates.[77] In vitro experiments showed that PD-L1 expression decreased in TKI-responsive cell lines after treatment with a TKI. A phase I study of the combination of erlotinib and nivolumab (3 mg/kg) in chemotherapy-naive patients with advanced EGFR-mutated NSCLC showed that 3 of 20 patients (15%) with acquired erlotinib resistance had a partial response, with 9 of 20 patients having stable disease.[78] These results may have implications in future clinical trial designs that incorporate checkpoint inhibition in EGFR-mutated NSCLC; clinical studies are ongoing (see Table 2; ClinicalTrials.gov identifiers: NCT02364609 and NCT02039674). Third-generation EGFR TKIs (AZD9291 and rociletinib) have shown promise in tumors harboring the T790M gatekeeper mutation; however, the recent suspension of clinical trials combining AZD9291 and durvalumab due to cases of lung toxicity may portend difficulties combining these agents.[79-81]
Checkpoint blockade plus VEGF inhibition
Tumor cells secrete vascular endothelial growth factor (VEGF) A, which has immunosuppressive effects via support of the formation of MDSCs, decreased T-cell priming, and decreased dendritic cell costimulatory molecule expression. Inhibition of VEGF or its receptor via antibodies such as bevacizumab or small molecules like sunitinib has immunomodulatory effects that include suppression of MDSCs and Tregs and downregulation of immunosuppressive signal-transduction pathways.[43,82] A phase I study demonstrated that concurrent administration of ipilimumab and bevacizumab was safe and feasible in patients with metastatic melanoma, showing a 2-year median OS of 25.1 months.[83] Many ongoing studies are evaluating the combination of immune checkpoint inhibition and VEGF or VEGF receptor inhibition in RCC, gastric cancer, and NSCLC (ClinicalTrials.gov identifiers: NCT01472081, NCT01984242, NCT02210117, NCT02572687, and NCT02443324).
Checkpoint blockade plus BRAF V600E inhibition
Vemurafenib is a potent inhibitor of mutant BRAF; it has immunomodulatory effects, including increasing expression of MHC class I molecules and differentiation antigens of melanocytes (gp100, MART1, tyrosinase, and others), while decreasing secretion of immunosuppressive cytokines.[43,82,84] Combining immune checkpoint inhibitors with BRAF inhibitors has led to unexpected toxicity. Vemurafenib was combined with ipilimumab in a small phase I study of patients with BRAF-mutated metastatic melanoma, which was halted due to grade 3 elevation of liver enzymes in 6 of 10 patients.[85] A report of 13 patients treated sequentially with ipilimumab then vemurafenib showed that the 3 patients who received vemurafenib within 4 weeks of the last dose of ipilimumab rapidly developed a grade 3 rash, which was biopsy-proven to be a drug hypersensitivity reaction.[86] All three patients resumed treatment after the vemurafenib was suspended for up to 11 days and restarted at a lower dose. Two patients treated with vemurafenib after treatment with anti–PD-1 therapy (nivolumab and pembrolizumab) developed hypersensitivity reactions with multi-organ involvement, with one patient also developing acute inflammatory demyelinating polyneuropathy.[87] A phase I study (ClinicalTrials.gov identifier: NCT02027961) evaluating the combination of durvalumab (anti–PD-L1) with dabrafenib (a BRAF inhibitor) and/or trametinib (a MEK inhibitor) in patients with BRAF-mutated and BRAF wild-type advanced melanoma did not reach an MTD and showed clinical activity across all cohorts.[88] Three targeted therapies-vemurafenib, dabrafenib, and trametinib-are FDA-approved for patients with BRAF-mutated advanced melanoma, and studies combining these and others (such as the MEK inhibitor cobimetinib) with immune checkpoint inhibitors are ongoing (ClinicalTrials.gov identifiers: NCT02357732, NCT01656642, and NCT02130466).
Checkpoint Blockade Plus Therapeutic Cancer Vaccines
Sipuleucel-T is the solitary approved therapeutic cancer vaccine. Antigen-specific immunotherapy via cancer vaccines has generally lacked clinical efficacy.[89-96] This may be due to tumoral immunosuppressive mechanisms that inactivate T cells in the tumor microenvironment.[97,98] Immune checkpoint inhibition could theoretically allow vaccine-primed and -activated T cells to accomplish effector functions in the tumor microenvironment. One study showed no benefit of adding a multipeptide vaccine to nivolumab in refractory melanoma.[99] Based on preclinical and clinical data demonstrating possible synergy of anti–CTLA-4 administered in combination with tumor cell vaccines that produce granulocyte-macrophage colony-stimulating factor, ipilimumab plus systemic sargramostim was compared with ipilimumab alone in patients with unresectable stage III or IV melanoma.[100,101] The combination had lower toxicity and yielded better OS (17.5 months vs 12.7 months; P = .01), without any difference in PFS.[102] Several phase I and II combination studies utilizing vaccines and PD-1 inhibitors are ongoing, and the topic is reviewed in detail elsewhere.[103]
Checkpoint Blockade Plus Chemotherapy
Numerous ongoing trials are evaluating the combination of chemotherapy and checkpoint blockade in solid tumors, including melanoma, NSCLC, and SCLC. In untreated metastatic melanoma, a phase III study showed that ipilimumab (at 10 mg/kg) plus dacarbazine improved OS compared with dacarbazine alone (11.2 vs 9.1 months, respectively), but this was at the expense of higher toxicity and there was no ipilimumab-alone comparator arm.[15] A phase II study showed that phased but not simultaneous ipilimumab plus platinum doublet chemotherapy (carboplatin/paclitaxel) improved immune-related PFS in patients with stage IIIB or IV NSCLC and extensive-stage SCLC, when compared with chemotherapy alone.[104,105] The choice of chemotherapy and dosing schedule are thus critical to optimizing outcomes of checkpoint blockade and chemotherapy combinations. With this in mind, a phase I four-cohort study evaluated first-line nivolumab at 10 mg/kg (N10) vs 5 mg/kg (N5) in combination with gemcitabine/cisplatin (N10) in advanced squamous-cell NSCLC, pemetrexed/cisplatin (N10) in advanced nonsquamous NSCLC, and paclitaxel/carboplatin (N5 vs N10) in combined cohorts of squamous and nonsquamous NSCLC.[106] The toxicity profile was additive, representing effects of both nivolumab and chemotherapy. The ORR, PFS, and 1-year OS outcomes were acceptable. In particular, the 1-year OS rate was 85% for the N5 paclitaxel/carboplatin group and 87% for the N10 pemetrexed/cisplatin group, which may reflect a positive signal.
A phase Ib study enrolled untreated patients with locally advanced or metastatic NSCLC to three treatment arms of atezolizumab plus chemotherapy, including carboplatin/pemetrexed, carboplatin/paclitaxel, and carboplatin/nab-paclitaxel.[107] Atezolizumab at 15 mg/kg every 3 weeks was administered with standard chemotherapy for 4 to 6 cycles followed by atezolizumab maintenance or atezolizumab/pemetrexed maintenance in the carboplatin/pemetrexed arm. An interim analysis showed that the ORR was 67% (95% CI, 48–82) by RECIST, with the carboplatin/pemetrexed arm having the highest response rate at 75% (95% CI, 45–93). The only two complete responses occurred in the carboplatin/nab-paclitaxel arm. The toxicity profile was as expected for chemotherapy, and no pneumonitis was observed. There was one grade 5 adverse event in a patient in the carboplatin/nab-paclitaxel arm who developed candidemia after prolonged neutropenia. Overall, the combination therapy response rates exceeded the 30% traditionally expected with platinum doublet chemotherapy; more mature data are forthcoming.
Checkpoint Blockade Plus Radiation Therapy
Recent studies indicate that at least one component of radiation-induced tumor control involves activation of the adaptive immune system as a result of tumor antigen release following radiation therapy.[108-110] Combining radiation therapy with immune checkpoint blockade may be an effective approach to stimulation of the adaptive immune system, with further amplification of immune response achieved via systemic immune checkpoint blockade. Preclinical studies have shown that combining radiation therapy with immune checkpoint blockade offers an opportunity for synergistic response rates.[111-114] The abscopal effect of radiation therapy is a poorly understood phenomenon that refers to a systemic antitumor response incited by localized radiation. Case reports and series have described the abscopal effect in patients with advanced melanoma treated with ipilimumab and radiation therapy.[115,116] A recent phase I clinical study combining ablative radiation therapy with ipilimumab in patients with metastatic melanoma showed excellent local control at the site of radiation and an 18% partial response rate at sites outside the radiation treatment field.[114] Multiple ongoing and upcoming phase I/II clinical studies that include patients with lung cancer, melanoma, and other solid tumors aim to evaluate whether the combination of radiation therapy with immune checkpoint blockade will be an effective approach for improving response rates in the metastatic setting (ClinicalTrials.gov identifiers: NCT02221739, NCT02239900, NCT02463994, NCT02383212, and NCT02444741) and improving outcomes for patients with potentially curable disease (ClinicalTrials.gov identifiers: NCT02525757 and NCT02434081). The phase III PACIFIC study (ClinicalTrials.gov identifier: NCT02125461) is evaluating consolidation durvalumab vs placebo in patients with unresectable stage III NSCLC treated with definitive chemoradiation (see Table 2).
Conclusion
The elements of a successful antitumor T-cell response include generation of sufficient numbers of antitumor T cells within the lymphoid compartment; successful T-cell trafficking and extravasation out of the lymphoid compartment, through the bloodstream, and into the tumor microenvironment; and successful T-cell effector function within the tumor microenvironment, with the need to bypass immune checkpoints, soluble inhibitory factors, and inhibitory cells. Although the tumor cell can subvert each one of these steps, researchers are developing an array of strategies to overcome such subversion, also at every step. Immune checkpoint inhibition has led to long-term survival exceeding a decade in some patients with metastatic melanoma and NSCLC. Combination strategies with immune checkpoint blockade as a backbone may allow the oncology community to achieve these outstanding outcomes in greater numbers of cancer patients.
Financial Disclosure:Dr. Antonia serves on the advisory boards of Bristol-Myers Squibb (BMS), Genentech, and MedImmune/AstraZeneca (AZ). Dr. Chiappori has made a conference presentation for BMS. Dr. Creelan receives research funding from BMS and Boehringer-Ingelheim; in addition, he receives honoraria and travel grants from AZ and BMS. Dr. Soliman serves as a consultant to Celgene. The other authors have no significant financial relationship with the manufacturer of any product or provider of any service mentioned in this article.
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