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Eur J Cardiothorac Surg. 2016 May; 49(5): 1324–1333.
Published online 2015 October 29. doi:  10.1093/ejcts/ezv371
PMCID: PMC4851162

Immunotherapy for non-small cell lung cancer: current concepts and clinical trials

Abstract

Recent successes in immunotherapeutic strategies are being investigated to combat cancers that have less than ideal responses to standard of care treatment, such as non-small-cell lung cancer. In this paper, we summarize concepts and the current status of immunotherapy for non-small cell lung cancer, including salient features of the major categories of immunotherapy—monoclonal antibody therapy, immune checkpoint blockade, immunotoxins, anticancer vaccines, and adoptive cell therapy.

Keywords: Lung cancer, Antibody therapy, PD-1/PD-L1 blockade, Chimeric antigen receptor, Adoptive cell therapy

INTRODUCTION

Recent insights into the immunological milieu of solid tumours have paved the way for novel therapeutic strategies now being applied in the clinic. Application of these strategies to non-small cell lung cancer (NSCLC) is of particular interest as current therapeutic strategies for this prevalent form of cancer have been unable to significantly improve patient prognosis.

Advances in immunotherapeutic approaches can be categorized into 5 main strategies; 3 fall under the broad category of antibody-based therapies (Fig. (Fig.1).1). The first antibody-based approach consists of targeted monoclonal antibodies constructed from either human/murine chimeric or fully human antibody components that bind to specific tumour-associated antigens, resulting in antibody-dependent cellular cytotoxicity (ADCC). The second antibody-based immunotherapy is immune checkpoint regulation. Following an antigen-specific immune response, tumour-infiltrating immune cells up-regulate inhibitory receptors. Then, solid tumours up-regulate inhibitory ligands that bind to these inhibitory receptors and render immune cells ineffective. Checkpoint blockade immunotherapy strategy emerged from the concept that a patient's own immune response to the tumour can be augmented using antibodies to block the immunosuppressive effect of inhibitory signalling on T cells. Immunotoxin therapy, the third antibody-based immunotherapy, utilizes antibodies to chaperone potent toxins to the cytosol of the cancer cell, thereby resulting in cell death. The fourth category of immunotherapy is cancer vaccines. This concept stems from the known beneficial effects of viral and bacterial immunization. Cancer vaccines are administered to the patient as biologically active antitumour antigen preparations with the purpose of activating the patient's immune system against that specific tumour antigen. The final and rapidly developing category of adoptive cell therapy (ACT) is a novel methodology wherein a patient's own immune cells are modified to target cancer cells and then re-administered to the patient. The most recent development in this category has been in adoptive T-cell therapy, wherein T cells of the patient are genetically engineered ex vivo to target a specific cancer antigen, then expanded in cell engineering facilities and subsequently re-administered to the patient via infusion, thus effectively providing the patient with an antitumour immune response. This review will give a brief overview of the 5 aforementioned immunotherapeutic strategies, with a focus on their potential applications in the treatment of NSCLC.

Figure 1:
Current immunotherapeutic strategies for non-small cell lung cancer. MHC: major histocompatibility complex; TCR: T-cell receptor; APC: antigen presenting cell.

MONOCLONAL ANTIBODIES

The concept of monoclonal antibody (mAb) therapy was pioneered following identification of tumour cell-specific mutations in NSCLC. Antibodies are generated by fully human or chimeric (mouse/human) fragments. They function in a direct manner by binding to the target and blocking its function, thereby inhibiting the effect of the mutation-induced signalling pathway as well as having indirect effects by initiating ADCC (Fig. (Fig.1,1, Section 1).

Epidermal growth factor receptor-targeted antibodies

Several mAbs are used in clinical practice and act via competitive inhibition of epidermal growth factor receptor (EGFR), blocking its site of interaction with epidermal growth factor. The receptor is then internalized—which down-regulates surface expression of EGFR and effectively inhibits aberrant growth signals—and halts progression of the cancer. Cetuximab (ERBITUX), an anti-EGFR antibody, has demonstrated marginal benefits when used in combination with chemotherapy as a first-line treatment for advanced NSCLC, as demonstrated in Phase III clinical trials [1] and meta-analyses [2]. Necitumumab (IMC-11 F8), a fully human antibody similar in structure to cetuximab but without murine components, is being evaluated in 2 ongoing Phase III clinical trials for NSCLC (NCT00981058, NCT00982111). Results from 1 trial evaluating necitumumab in combination with chemotherapy as first-line treatment for Stage IV squamous NSCLC demonstrated a significant improvement in overall survival (OS) from 9.9 to 11.5 months (n = 1093) compared with chemotherapy alone [3], even though this trial was limited to squamous NSCLC and application to non-squamous NSCLC has not been promising [4].

MET receptor blocking antibodies

Overexpression of MET receptor (c-MET), a tyrosine kinase involved in cell proliferation, survival, and invasion, has been demonstrated in up to 40% of NSCLC [5]. Monoclonal antibodies that target this pathway, including onartuzumab (MetMAb) and ficlatuzumab are currently being developed. Results from a Phase III trial of 499 patients that was designed to evaluate onartuzumab plus erlotinib in MET-positive NSCLC were stopped due to futility after demonstrating that addition of onartuzumab to erlotinib did not improve OS [hazard ratio (HR) 1.27, P = 0.068] [6]. Whereas onartuzumab targets cMET receptors to block activation, ficlatuzumab, a humanized mAb, targets cMET ligand HGF to block the signalling pathway [7]. Results of a Phase IB trial evaluating ficlatuzumab in combination with gefitinib in patients with NSCLC demonstrated promising results, with partial remission achieved in 5 of 12 patients [8]. A Phase II trial (NCT02318368) evaluating ficlatuzumab plus erlotinib versus placebo versus erlotinib alone in previously untreated metastatic EGFR mutant NSCLC is open and currently recruiting patients.

Vascular endothelial growth factor-targeted antibodies

Another mechanism involved in tumour growth is vascular endothelial growth factor (VEGF). Along with its receptor (VEGFR), VEGF stimulates angiogenesis-favouring tumour invasion and metastasis, thus making it a potential target for mAb therapy [9].

The most well-studied mAb currently in use is bevacizumab (Avastin). The Eastern Cooperative Oncology Group 4599 trial demonstrated a significant survival benefit of paclitaxel/carboplatin/bevacizumab versus paclitaxel/carboplatin in patients with recurrent or advanced NSCLC [median survival 12.3 vs 10.3 months, HR =0.79, P = 0.003, median progression-free survival (PFS) 6.2 vs 4.5 months, HR = 0.66, P < 0.001] [10]. Based on these results, bevacizumab has been approved for first-line treatment of select patients. In another Phase III trial called AVAiL (AVAstin in Lung study), which is the addition of bevacizumab in both low and high doses to cisplatin/gemcitabine chemotherapeutic regimens, resulted in significant improvement in PFS (6.5 vs 6.1 months, P = 0.03; 6.7 vs 6.1 months, P = 0.003 for high dose and low dose versus chemotherapy alone, respectively) [11]. Although the increased risk of pulmonary haemorrhage was reported in patients with squamous NSCLC, a panel of experts concluded that an independent risk-benefit assessment should be undertaken, as there were no clear biological reasons confirmed for treatment-associated increased haemorrhage [12].

Ramucirumab (IMC-1121B, CYRAMZA) is a fully human mAb that binds the extracellular portion of VEGFR, thereby blocking its interaction with VEGF. A multicentre, Phase III trial that investigated ramucirumab plus docetaxel versus placebo plus docetaxel for second-line treatment of progressive stage IV NSCLC demonstrated an increase in median OS (10.5 vs 9.1 months, HR = 0.86, P = 0.023) as well as PFS (4.5 vs 3.0 months, P < 0.0001) [13].

In summary, mAb immunotherapy has demonstrated promising, yet modest, results; some, as first-line therapy for NSCLC in combination with chemotherapeutic regimens (i.e. cetuximab, necitumumab and bevacizumab) and others with limited efficacy as second-line treatment (i.e. ramucirumab and bevacizumab) (Table (Table1).1). However, this therapy is limited to NSCLC that expresses mutant EGFR, cMET, or VEGF. This strongly suggests that immunotherapies that have broader applications are needed.

Table 1:
Monoclonal antibodies: ongoing clinical trials

IMMUNE CHECKPOINT BLOCKADE

In contrast to mAb therapy, immune checkpoint blockade utilizes antibodies to prevent cytotoxic T-cell anergy that results from up-regulation of inhibitory signals (Fig. (Fig.1,1, Section 2). An important step in the progression of malignancies is evasion of the immune system, an effect that is achieved by expression of inhibitory signals, such as cytotoxic T-lymphocyte antigen-4 (CTLA-4) and programmed death receptor ligand 1 (PD-L1), described below.

Cytotoxic T-lymphocyte antigen-4 inhibition

CTLA-4 is a homodimeric glycoprotein receptor that is expressed by activated T cells and regulatory T cells (Tregs), and it is a strong negative regulator of T cells [14]. Binding of CTLA-4 to co-stimulatory B7 molecules (CD80/CD86) expressed on antigen presenting cells (APCs) both inhibits interaction with T-cell CD28-activating receptor and initiates a direct inhibitory signal that decreases T-cell effector function. It also induces inhibitory pathways in APCs [15]. Antitumour effects of CTLA-4 inhibitors are caused by increases in CD28/B7 T-cell activation and depletion of Tregs [16]. CTLA-4 blockade has also been shown to expand a subpopulation of tumour-infiltrating CD4 + T cells [17]. Anti-CTLA-4 antibodies were among the first checkpoint blockade agents to be tested in the clinic; one such antibody, ipilimumab, demonstrated significant improvement in OS of patients with advanced melanoma [18].

Tremelimumab and ipilimumab are currently being tested in lung cancer and mesothelioma. Tremelimumab (CP-675 206) is a human IgG2 mAb that binds to CTLA-4. A Phase II trial evaluating tremelimumab following first-line, platinum-based therapy in patients with advanced NSCLC demonstrated an objective response rate of only 4.8% (n = 2) with no increase in PFS compared with best supportive care [19]. Multiple additional trials (NCT02352948, NCT02000947) evaluating tremelimumab in combination with other immune checkpoint blockade agents, such as MEDI4736, are ongoing. Ipilimumab (MDX-010, YERVOY) is a human IgG1 mAb that stimulates antitumour immune response by blocking CTLA-4 [20]. A Phase II trial demonstrated improved immune-related PFS, an assessment developed from the World Health Organization to capture regression of index lesions in the face of new lesions as well as tumour stabilization or decrease in tumour burden after initial progression. Improvement seen in immune-related PFS was only seen with phased, but not combination, ipilimumab with paclitaxel and carboplatin when compared with chemotherapy alone (5.7 vs 4.6 months, HR = 0.72; P = 0.05) [21]. These have led to numerous clinical trials that have evaluated the combination of ipilimumab with EGFR or ALK tyrosine kinase inhibitors in lung cancer (Table (Table2).2). However, the major drawback to anti-CTLA-4 therapy is generation of autoimmune toxicities resulting from overactivated T cells [22]. Thus, the challenge for anti-CTLA-4 therapy is defining a favourable therapeutic index that can strike a balance between antitumour immunity and autoimmunity.

Table 2:
Immune checkpoint blockade: ongoing clinical trials

PD-1 inhibition

The surface receptor programmed cell death protein 1 (PD-1) is a member of the B7-CD28 superfamily—similar to CTLA-4—and is a key immune checkpoint receptor. It is expressed on natural killer (NK) cells and activated T and B cells, and it binds to either PD-L1 (B7-H1) or PD-L2 (B7-DC), both of which are expressed on tumour cells, APCs, T cells, and B cells [34]. Binding of PD-1 with its ligand inhibits downstream NF-κB transcription and proliferation with resultant induction of T-cell tolerance. A study of PD-1 expression in 21 NSCLC patients showed that increased PD-1 expression on tumour-infiltrating CD8+ T cells correlated with impaired T-cell function [23]. PD-1-specific antibodies, Nivolumab (BMS-936558, ONO-4538, OPDIVO) and Pembrolizumab (MK-3475, KEYTRUDA), are currently being evaluated in ongoing clinical trials.

Nivolumab is a human IgG4 PD-1 receptor blocking antibody with no ADCC activity; promising results from ongoing (Phases I, II, and III) and completed (Phase I and II) clinical trials have recently yielded FDA approval for second-line chemotherapy treatment for resistant squamous NSCLC [24]. A single-arm, Phase II trial evaluating the activity and safety of nivolumab in advanced, refractory NSCLC demonstrated a 14.5% objective response rate (17 of 117 patients), as measured by radiographic response, as well as a 26% rate of stable disease (20 patients) [25]. Treatment-related adverse events included fatigue, pneumonitis, and diarrhoea, and occurred in 3–4% of patients. Results from another Phase I trial demonstrated an 18% cumulative response rate in a subset of 76 NSCLC patients [34]. An update from this trial regarding OS and long-term safety of the subset of heavily pretreated, advanced NSCLC patients was recently published; 17% (22 of 129) of treated patients demonstrated an objective response rate with estimated median response duration of 17 months, with similar response rates seen in squamous and non-squamous NSCLC [26].

Pembrolizumab is a mAb that neutralizes the PD-1 protein, which showed significant antitumour activity in melanoma [27], and has demonstrated an objective response rate of 19.4% in a group of 495 patients with advanced NSCLC [28]. This recently published clinical trial conducted correlative analysis of PD-L1 expression in tumour samples and identified a subset of patients with expression in ≥50% of tumour cells that had enhanced the likelihood of response to PD-1 blockade with pembrolizumab. Additionally, results from an ongoing Phase I/II trial evaluating the safety and clinical efficacy of pembrolizumab in combination with either carboplatin/paclitaxel or carboplatin/pemetrexed were reported at the American Society of Clinical Oncology (ASCO) 2015 Annual Meeting demonstrating a 30–67% preliminary overall response rate (ORR) in 44 treated patients; reported adverse events included reversible transaminase elevation, anaemia, colitis, and rashes [29]. Ongoing clinical trials will evaluate pembrolizumab, in addition to chemotherapeutic agents, for advanced staged NSCLC (Table (Table22).

Programmed death receptor ligand 1 inhibition

PD-L1 (B7-H1) is expressed in T cells, B cells, and surface APCs. It is a ligand of PD-L1 involved in T-cell inhibitory signalling that reduces cytokine production and suppresses T-cell proliferation [34]. It has been reported that PD-L1 up-regulates in a range of tumours and its expression correlates with poor prognosis for several malignancies, including NSCLC [30]. PD-L1 is expressed in 25–43% of NSCLC [35, 36]. PD-L1 and mRNA expression are also associated with tumour-infiltrating lymphocytes (TILs) [36]. Ansen et al. [37] showed that, while PD-L1 expression is associated with distinct genotypes of NSCLC, it was not associated with EGFR/KRAS mutations or ALK rearrangement.

The efficacy of several anti-PD-L1 antibodies (MPDL3280A, BMS-936559 and MEDI4736) are currently being investigated in lung cancer. MPDL3280A is a human anti-PD-L1 mAb that contains an engineered Fc-domain designed to optimize efficacy by minimizing the ADCC effect. Results from a Phase II trial evaluating MPDL3280A in comparison with docetaxel in patients previously treated with PD-L1-positive NSCLC were presented at the ASCO 2015 Annual Meeting. This study stratified patients according to PD-L1 tumour levels; this demonstrated an increase in PFS (9.7 vs 3.9 months) for patients with high levels of PD-L1 expression treated with MPDL3280A [38]. BMS-936559, a human IgG4 mAb that inhibits the binding of PD-L1 to PD-1, was evaluated in a multicentre, Phase I clinical trial of 75 NSCLC patients [31]. Although the results were modest with 5 patients achieving an objective response rate, 6 additional patients demonstrated stable disease lasting 24 weeks. BMS-936559 is currently undergoing an additional Phase I clinical trial (NCT00729664) including a subset of NSCLC. MEDI4736 is a human IgG1 mAb that was engineered to prevent ADCC. It blocks PD-L1 binding to PD-1 and CD-80, and is currently being evaluated for use against locally advanced and metastatic NSCLC (NCT02125461, NCT02273375) and in combination therapy with tremelimumab (NCT02352948, NCT02000947). Preliminary results from a Phase I/II trial of 198 treated patients evaluating MEDI4736 alone demonstrated an ORR of 14–23% in PD-L1-positive patients with a trend towards higher ORR in squamous NSCLC [33].

In checkpoint blockade therapy, endocrine-related adverse events are commonly reported side-effects with reported incidence as high 22% for thyroiditis/hypothyroidism and 9% for hypophysitis [32]. Although the inhibition of PD-1/PD-L1 pathways has shown higher response rates across a wider range of tumours than other immunotherapies, final results from studies targeting NSCLC have yet to be reported and biomarkers predicting the safety and efficacy of PD-1/PD-L1 blockade in patients are lacking (Table (Table2).2). A recent publication has shown that higher non-synonymous mutation burden in NSCLC tumours was associated with improved ORR, durable clinical benefits, and PFS with PD-1 blockade [39]. Further studies that predict the utility of checkpoint blockade in NSCLC are needed.

IMMUNOTOXIN THERAPY

Antitumour immunotoxin therapy utilizes an antigen-targeted moiety—either antibody or mutated growth factor receptor—linked to a potent toxin, such as Pseudomonas exotoxin A or ricin, a plant toxin (Fig. (Fig.1,1, Section 3). The binding moiety attaches to a tumour surface antigen, thereby initiating endocytosis of the complex including the toxin. Once in the cytosol, the toxin can inhibit protein synthesis and cell-cycle arrest resulting in subsequent cell death (even in quiescent cells) [40].

While immunotoxin therapy has shown promising results in clinical trials for haematological malignancies [41], its application in NSCLC has been limited [42]. An immunotoxin therapy that has shown promise in treating solid tumours is SS1P. SS1P is a recombinant immunotoxin targeted against mesothelin, a cell-surface antigen overexpressed in lung adenocarcinoma and conjugated to a portion of Pseudomonas exotoxin A. This therapy has shown promising results for malignant pleural mesothelioma (MPM) (80% of epithelioid MPM overexpress mesothelin). A Phase II clinical trial (NCT01362790) is currently in recruitment for MPM with plans to include a cohort of lung adenocarcinoma patients based on published results of SS1P's antitumour efficacy in lung cancer cell lines in preclinical studies [43]. These studies, in combination with evidence that mesothelin is highly expressed in NSCLC [44], provide further rationale for targeting mesothelin with immunotoxin therapy for NSCLC in the future.

As immunotoxin therapy for solid tumours continues to develop, some limitations that will need to be addressed in its application to lung cancer include identification of additional tumour targets with limited off-target toxicity and immunosuppressive regimens to improve immunotoxin tolerance and prevention of antitherapeutic immune responses.

ANTICANCER VACCINES

Immunotherapy involving cancer vaccines aims to generate or augment the innate or adaptive antitumour immune response with the use of a biologically active whole-cell or specific protein antigen preparation [45]. Cancer vaccines are administered to patients in combination with an inactive pathogen or non-specific immune stimulant to increase immune reactivity. Once the cancer vaccine is processed by an APC, the APC then presents the tumour-associated antigen to activate B- and T-lymphocyte antitumour effector functions (Fig. (Fig.1,1, Section 4).

Whole-cell vaccines

Belagenpumatucel-L (Lucanix) is a whole-cell vaccine produced from irradiated allogenic NSCLC cell lines transfected with a TGF-β2 antisense gene plasmid; this is based on the knowledge that the level of TGF-β2 correlates with poor prognosis in NSCLC patients and has immunosuppressive effects [46]. A Phase III clinical trial evaluated Belagenpumatucel-L as second-line maintenance therapy following response to front-line chemotherapy in improving OS versus placebo. The study did not meet its primary end-point for OS after randomizing 532 patients with advanced stage NSCLC, finding a median OS ranging from 20.3 to 17.8 months (HR = 0.94, P = 0.594) [47]. However, a subset of 305 advanced stage patients who were randomized within 12 weeks of front-line chemotherapy did not show a statistically significant improvement in OS compared with placebo, with median OS of 20.7 vs 13.3 months (HR = 0.75, P = 0.083).

Tergenpumatucel-L is another whole-cell vaccine that was developed from three allogenic lung tumour cell lines genetically modified to express carbohydrate α-galactosyltransferase, a potent immunogenic enzyme that is responsible for hyperacute rejection. A Phase III clinical trial to evaluate Tergenpumatucel-L versus docetaxel in progressive or relapsed NSCLC is currently recruiting patients [46].

Granulocyte-macrophage colony-stimulating factor (GM-CSF) gene-transfected tumor cell vaccine, a whole-cell vaccine developed from autologous or allogeneic tumour cells genetically modified to secrete granulocyte-macrophage colony-stimulating factor, initially demonstrated evidence of bioactivity, although it did not demonstrate any objective response in NSCLC in a Phase I/II clinical trial [48]. An additional Phase II trial (NCT00074295) was terminated due to lack of vaccine availability.

Target-specific vaccines

MAGE-A3 immunotherapeutic vaccine was developed to combat melanoma-associated antigen-A3 (MAGE-A3) gene, which is a tumour-specific antigen expressed in 35–50% of NSCLC [49]. Results from a Phase II clinical trial evaluating adjuvant therapy using the MAGE-A3 vaccine in resected NSCLC demonstrated feasibility with minimal toxicity, although there was no statistically significant improvement in disease-free interval or disease-free survival [50]. The subsequent Phase III trial, the MAGRIT (MAGE-A3 as ‘Adjuvant Non-Small Cell Lung Cancer Immunotherapy’) trial, was recently aborted due to futility [51]. A total of 2272 patients were randomized to receive either MAGE-A3 vaccine or placebo. There was no improvement in disease-free survival and no gene signature that identified a subpopulation of responders.

Tecemotide (Stimuvax, L-BLP25) is an antigen-specific vaccine that targets mucin-1 (MUC-1) glycoprotein, which has been shown to elicit a strong immune response against lung cancer in preclinical studies [52]. Results from the ‘Stimulating Targeted Antigenic Response To NSCLC’ (START) clinical trial, a Phase III trial evaluating tecemotide versus placebo after chemoradiotherapy in 1513 advanced stage NSCLC patients, demonstrated no significant difference in OS (median OS of 25.6 vs 22.3 months, HR = 0.88, P = 0.123) [53].

TG4010 is a vaccine generated from a modified vaccinia virus that contains the sequence that is encoded for MUC-1 and IL-2 [54]. Results from a Phase IIB trial demonstrated an increase in 6-month PFS with TG4010 plus cisplatin/gemcitabine when compared with chemotherapy regimens alone in 148 advanced staged NSCLC patients with MUC1 expression via immunohistochemistry (43.2 vs 35.1%) [55]. These promising results have led to a Phase III clinical trial (NCT01383148) that is currently in recruitment.

In summary, vaccine therapies have shown mixed results in Phase II clinical trials without significant evidence of therapeutic efficacy in Phase III trials; however, results from ongoing Phase III clinical trials (i.e. TG4010 and Tergenpumatucel-L) have yet to be reported (Table (Table3).3). Limitations of this therapy are lack of strong expression of target antigens and reliance on an innate immune system to combat the immunosuppressive-dominant tumour microenvironment of NSCLC. Additional efforts to activate the endogenous immune system have included intrapleural administration of cytokines, such as interferon-γ, that have demonstrated eradication of tumour cells in pleural effusions in 2 of 6 patients in one clinical trial [56], or intrapleural adenoviral-mediated interferon-β gene transfer, which has been shown to induce humoral antitumour immune responses in a Phase I clinical trial that included NSCLC patients [57].

Table 3:
Anticancer vaccines: ongoing clinical trials

ADOPTIVE CELL THERAPY

The ACT approach involves: (i) collection of immune cells from the peripheral blood or the tumour itself; (ii) isolation, modification, and ex vivo expansion of tumour-specific immune cells; and (iii) administration of immune cells back to the autologous host [58]. Adoptive cell therapy provides immune cells a defined antigen specificity to enhance antitumour immunity and offers a particular advantage compared with the previously reviewed immunotherapies in that they have the capacity to infiltrate the tumour and generate an antitumour effective population that leads to direct cytotoxic effects. Targeted antigens are overexpressed ideally on cancer cells with limited or no expression on normal cells; this limits graft-versus-host disease. While the traditional ACT approach has focused on reinfusing harvested and ex vivo expanded TILs, recent studies have focused on utilizing genetic engineering approaches to insert antigen-targeted receptors called chimeric antigen receptors (CARs) into T cells (Fig. (Fig.1,1, Section 5). This is the newest branch of immunotherapy for lung cancer with the published literature focused on preclinical evidence that has led to ongoing translation to clinical trials (Table (Table44).

Table 4:
Adoptive cell therapy: ongoing clinical trials

Tumour-infiltrating lymphocyte therapy

Tumour-infiltrating lymphocyte therapy requires isolation of T cells from fresh cancer specimens and progressive expansion of tumour-specific T cells ex vivo using high doses of IL-2. The advantage of TIL therapy is the broad nature of T-cell recognition against both defined and undefined tumour antigens. A Phase II clinical study (NCT02133196) using autologous young TILs derived from patients with NSCLC following a non-myeloablative lymphocyte depleting preparative regimen (i.e. cyclophosphamide and fludarabine) is currently recruiting participants.

γδ T-cell therapy

γδ T cells differ from the majority of T cells (αβ) in that they are not restricted to major histocompatibility complex (MHC) molecules and they recognize a variety of structurally different ligands associated with cellular stress. They can secrete abundant cytokines and exert potent cytotoxicity against cancer cells. A Phase I clinical study is currently ongoing to evaluate the safety and potential antitumour effects of reinfusing γδ T cells ex vivo in patients with advanced or recurrent NSCLC. In this trial, all patients remained alive during the study period with a median survival of 589 days and median PFS of 126 days (UMIN ID: C00000036) [59].

Natural killer cell-based therapy

Natural killer cells are CD3–CD56+ lymphocytes that can kill certain target cells by secreting various effector molecules, release cytoplasmic granules containing perforin and granzymes, and activate death receptor-mediated apoptosis. Natural killer cell-based therapy uses expanded autologous or allogeneic NK cells to treat cancer. In a Phase I clinical trial on the safety and possible clinical efficacy of NK cells, along with chemotherapy, in patients with advanced NSCLC, 15 patients received 2–4 doses of allogeneic-activated NK cells. No local or systemic side-effects were observed, and 56% of patients had 1-year survival rate whereas 19% had 2-year survival rate [64]. A recently published study reported that NK and cytotoxic T lymphocytes mixed effector (NKTm) cells prolonged OS of patients compared with the control group (31.1 vs 18.1 months, P = 0.008, HR = 0.562) and increased the 2-year survival rate [65]. In a Phase IIA trial of the combination of autologous NK cells with docetaxel in patients with advanced NSCLC, PFS was analysed in 19 patients and no improved treatment benefit was found compared with historical controls due to the limited number of enrolled patients [66].

Cytokine-induced killer cell-based therapy

Cytokine-induced killer (CIK) cells are a heterogeneous population of effector CD8 T cells with diverse T-cell receptor (TCR) specificities. Cytokine-induced killer cells are generated by in vitro expansion of peripheral blood lymphocytes (PBLs) with anti-CD3 antibodies and IL-2. Advantages of CIK cells are that they are rapidly proliferating and exhibit high cytotoxicity. A study that investigated the role of dendritic cell (DC)/CIK treatment for patients with advanced NSCLC showed that DC/CIK treatment prolonged PFS compared with the control group (3.2 vs 2.6 months, P < 0.05) and no obvious toxicity was found [60]. Another clinical trial investigated the efficacy of CIK cell therapy following chemotherapy in patients with NSCLC after surgery. The 3-year OS rate and median OS time were significantly higher in the group that was treated with combination therapy with CIK compared with chemotherapy alone (82 vs 66%; P = 0.049, and 73 vs 53 months; P = 0.006, respectively). Chemotherapy/CIK treatment prolonged 3-year PFS compared with the control group (6 vs 3%; P < 0.001) [61].

Engineered T-cell therapy

Engineered T-cell therapy consists in redirecting T-cell specificity and cytotoxic activity against tumour antigens. Two strategies have been developed: (i) genetic insertion of TCR into T cells; and (ii) genetic insertion of a CAR into T cells. These strategies allow recognition of both intracellular and extracellular cancer antigens, thereby resulting in broad applicability for different forms of cancers.

Genetic engineering with T-cell receptors

This method redirects normal T cells to recognize tumour antigens by genetic engineering with tumour antigen-specific TCR genes. As an alternative to TILs, TCR recognizing cancer antigens were reintroduced into autologous T cells and then transferred back into patients. Clinical trials involving TCR immunotherapy (genetically engineered to target NY-ESO-1 or MAGE-A3) are currently under way (NCT01697527, NCT01967823, and NCT02111850).

Chimeric antigen receptor

Although the use of TCR gene-redirected T cells can detect both intracellular and cell-surface tumour-associated antigens, TCRs are restricted by MHC specificity, thereby limiting their applicability in some patients. CAR-modified T cells provide an alternative to TCR gene-redirected T cells and they are not restricted to MHC (Fig. (Fig.2).2). CAR-modified cells endow T-cell populations with a receptor against any given antigen, permitting targeting of T cells towards potentially any tumour. CAR-modified T cells have demonstrated potent clinical efficacy in patients with haematological malignancies [62, 63]. To date, ongoing work is focused on finding the ideal target antigen and manipulating the host-associated factors to support the expansion and persistence of these engineered cells in solid tumour microenvironments.

Figure 2:
Chimeric antigen receptor therapy. Chimeric antigen receptor T-cell therapy is performed by isolating T cells from the peripheral blood of the patient, genetically modifying them with a viral vector to express CARs, expanding them ex vivo and reinfusing ...

Our laboratory work is focused on investigating CAR T-cell therapy against thoracic malignancies. Our observations from investigation of large cohorts of patient tumours, which was supported by publications from other groups [67], have led us to target mesothelin as a cancer-associated antigen in mesothelioma [68], NSCLC [44], oesophageal adenocarcinoma [69], and metastatic triple-negative breast cancer [70]. Our preclinical observation that mesothelin promotes an aggressive phenotype is strengthened by our clinical observations that patients with mesothelin-positive tumours have poor prognoses [68]. Furthermore, limited expression of mesothelin in ‘normal’ pleura, peritoneum, and pericardium provides a safety margin for CAR T-cell therapy [71]. A Phase I clinical trial (NCT01355965) of mesothelin-targeted CARs has been initiated for the treatment of patients with pleural disease. Our group has found that regional administration of mesothelin-targeted CAR T cells via intrapleural injection in an orthotopic model of MPM achieved longer term remissions than intravenous injection due to avoidance of T-cell sequestration in lungs and early antigen activation [72]. This significant preclinical observation has led to the initiation of a Phase I clinical trial (NCT02414269) to evaluate the safety of intrapleural administration of mesothelin-targeted CAR T cells in patients with pleural malignancies from mesothelioma, NSCLC, and breast cancer. A Phase I/II study (NCT01218867) of CAR T-cell therapy targeting VEGFR2 for patients with metastatic cancer who have not responded to standard therapy is currently recruiting participants, including those with lung cancer.

IMMUNOTHERAPY AND IMPLICATIONS FOR THORACIC SURGEONS

In early-stage NSCLC, although surgical resection achieves locoregional control, distant recurrences that result in poor survival rates have been a challenge. While newer immunotherapeutic strategies are being investigated for advanced NSCLC, studies that have investigated the tumour immune microenvironment in early-stage NSCLC from our group [73, 74] and others [75] have shed light on potential neoadjuvant and adjuvant immunotherapeutic strategies by differentiating specific immunological milieus with prognostic indication, thereby providing a direction for therapies to shift the balance towards an antitumour immune response. Furthermore, these studies shed light on potential immune biomarkers for NSCLC. Neoadjuvant strategies may help boost innate immune system response to multiple cancer antigens, thereby achieving long-term survival following surgical resection. Analysis of resected tumours for the presence of aggressive antigen-expressing cancer cells, along with defining the tumour immune microenvironment, may provide patient-specific adjuvant immunotherapeutic strategies. Additionally, preclinical evidence for combining radiation with immunotherapy has provided the rationale for clinical trials examining combination therapy, in particular, those with checkpoint blockade [76]. Furthermore, cisplatin immunomodulatory response can be used for combination approaches in the treatment of NSCLC [77].

CONCLUSION

Immunotherapy for solid tumours has only recently emerged as a promising field for development. Insights into the complex tumour microenvironment have allowed progress into treating NSCLC, a solid tumour that was previously thought to be impenetrable to immunotherapeutic strategies. While immunotherapy for lung cancer has thus far been met with significant challenges resulting in less than ideal responses in Phase III clinical trials, combating complexity of the NSCLC tumour microenvironment is an ongoing area of investigation and will likely require a combination of the above strategies. However, the knowledge obtained from these studies, in parallel with understanding of the tumour immune microenvironment of early-stage NSCLC, will eventually lead to novel neoadjuvant and adjuvant translational immunotherapeutic strategies that can prolong survival in surgically treated NSCLC patients.

Funding

This work was supported by grants from the National Cancer Institute at the National Institutes of Health (R21 CA164568-01A1, R21 CA164585-01A1, U54 CA137788, R01 CA136705-06, P30 CA008748, and P50 CA086438-13), the U.S. Department of Defense (LC110202) and the Stand Up To Cancer–Cancer Research Institute Cancer Immunology Translational Cancer Research Grant (SU2C-AACRDT1012). Stand Up To Cancer is a programme of the Entertainment Industry Foundation administered by the American Association for Cancer Research.

Conflict of interest: none declared.

ACKNOWLEDGEMENTS

We thank Alex Torres of the MSK Thoracic Surgery Service for his editorial assistance.

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