Malignant pleural mesothelioma (MPM) is an inexorably progressive and almost universally fatal malignancy.

 The vast majority of cases are associated with exposure to fibrous minerals, specifically asbestos or erionite fibers. In addition, prior thoracic radiation therapy

 have also been linked to the development of MPM. With the continued use of asbestos products in many countries, the incidence of MPM continues to rise worldwide, causing an estimated 43,000 deaths annually.

Effective treatment options are largely lacking. The role of surgery and standard multimodality therapy remains controversial and may only benefit highly selected patients.

 Folate antimetabolite-based chemotherapy, the current standard of care, has modest benefits and has been shown to improve overall survival by approximately 3 months.

 Chemotherapy provides even less of a benefit for patients with sarcomatoid subtypes of MPM.

Recently, immunotherapies have been shown to be effective for the treatment of metastatic melanoma and prostate cancer and are now approved by the Food and Drug Administration (FDA) for these diseases. In MPM, a number of isolated cases demonstrate that immune-mediated mechanisms can effectively target this tumor. Consequently, immunotherapeutic strategies are felt to represent an attractive alternative for the treatment of MPM. The complexity of the immune system allows for many ways to therapeutically modulate anti-tumor immunity. A comprehensive review of the immunologic therapies tested in MPM is beyond the scope this focused review, but can be found elsewhere.

This review specifically focuses on the therapeutic modulation of immune checkpoints for the treatment of MPM. Despite the ability of the immune system to recognize tumors, effective anti-tumor immune responses are blunted by many immunosuppressive mechanisms.

 In addition to successful T-cell receptor and major histocompatibility complex binding (signal 1), effective T-cell activation against self, tumor, and foreign antigens also depends on activating costimulatory signals (signal 2), and the presence of effector cytokines (signal 3). Immune activation is controlled by negative costimulatory signals, immune checkpoints. Immune checkpoints are critical to maintain the delicate balance between effective antimicrobial responses, self-tolerance, and prevention of autoimmunity. The two most characterized immune checkpoints are the cytotoxic T-lymphocyte antigen-4 (CTLA-4) - CD80/CD86 (B7-1/B7-2) checkpoint, and programmed cell death 1 (PD-1) - programmed cell death ligand 1 (PD-L1, B7-H1) - programmed cell death ligand 2 (PD-L2, B7-DC). Both of these pathways have been implicated in tumor-mediated suppression of anti-tumor immunity.

CTLA-4 is a surface receptor expressed on T lymphocytes. CTLA-4 inhibits the costimulatory interaction between CD28 on T cells and CD80/CD86 on antigen-presenting cells by sequestering CD80/CD86 because of its higher binding affinity. Two human monoclonal antibodies targeting CTLA-4, ipilimumab (Yervoy, IgG1) and tremelimumab (IgG2), have been developed to disrupt CTLA-4 - CD80/CD86 binding and have been or are being studied for the treatment of solid tumors. The FDA approved ipilimumab for metastatic melanoma after an objective response rate of 10.9% and a median overall survival of 10.1 months were demonstrated in a subsequent clinical trial.

 Tremelimumab, by comparison, in the first-line setting for metastatic melanoma, demonstrated an objective response rate of 10.7% and a median overall survival of 12.6 months.

 Tremelimumab has demonstrated promising activity in MPM.

 In a phase II trial, including 29 pretreated patients with pleural and peritoneal mesothelioma, treatment with tremelimumab resulted in 3 objective responses, 2 up front and 1 after initial progression. In addition, disease stabilization occurred in 9 patients. Overall survival was very encouraging in this population, 48.3% and 36.7% at 1 and 2 years, respectively. Consequently, tremelimumab is being studied in ongoing clinical trials (NCT01655888, NCT01843374). Unfortunately, there are no predictive biomarkers to identify responders to CTLA-4 blockade. In some studies, patients with low levels of vascular endothelial growth factor were more likely to benefit from ipilimumab for the treatment of metastatic melanoma,

 but this relationship remains to be explored in MPM. Similar to the retrospective analysis of patients with metastatic melanoma who were treated with ipilimumab,

 T lymphocytes after the initiation of therapy was associated with improved survival in MPM patients treated with tremelimumab. However, baseline levels of CD4

 T lymphocytes were not predictive of treatment benefit.

 Further clinical trials, including correlative studies to develop predictive biomarkers, are needed to define the role of CTLA-4 blockade in the treatment of MPM.

Programmed cell death 1 (PD-1) is another immune checkpoint receptor expressed on T-cell lymphocytes. Both of its ligands, PD-L1/(B7-H1) and PD-L2/(B7-DC), have homology with the B7 family of costimulatory molecules. Upon its discovery in peripheral blood monocytes, PD-L1 was shown to negatively regulate T cells through IL-10 production.

 Subsequently, it became clear that PD-L1 (B7-H1) inhibits T-cell proliferation by engagement of PD-1,

 and consequently it is now mostly referred to as PD-L1. PD-L1 is normally expressed by antigen-presenting cells where it controls immune activation. Tumor cells can aberrantly express PD-L1, and the engagement of PD-L1 with PD-1 within the tumor microenvironment induces apoptosis of tumor-specific T-cells.

 Another ligand for PD-1, PD-L2 (B7-DC), also inhibits T-cell receptor mediated proliferation.

 Expression of PD-L1 by tumor cells has been correlated with worse survival in many malignancies, including various forms of renal cell carcinoma,

 In MPM, PD-L1 (as defined by 5% or greater expression by tumor cells) was detected in 40% of cases, including almost all sarcomatoid mesotheliomas except for 1 desmoplastic tumor. PD-L1 expression remained associated with poor survival following multivariate analysis adjusting for subtype, extent of disease, and age. 

Some antibodies have been developed to inhibit the PD-1/PD-L1 axis and have been tested primarily in melanoma, renal cell carcinoma, and non-small cell lung cancer (NSCLC),

 but some early-phase clinical trials have included patients with MPM (eg, NCT01772004 with the PD-L1 inhibitor MSB0010718C), and a phase II clinical trial is planned in MPM.

 Ongoing clinical trials and other translational studies foreshadow the potential difficulties in targeting the PD-1/PD-L1 axis in MPM. The most mature data for the use of anti-PD-1 therapy are available for metastatic melanoma.

 In fact, the FDA recently approved the fully humanized IgG4 anti-PD-1 antibody pembrolizumab for the treatment of melanoma in the US. A separate anti-PD-1 antibody, nivolumab, was approved in Japan also for the treatment of metastatic melanoma, and received clearance from the FDA in December 2014. One study suggests that response to therapy is enriched in patients whose tumors express PD-L1,

 however, several issues obfuscate our understanding of these data.

First, while there are numerous antibodies marketed for the detection of PD-L1 by immunohistochemistry, their diagnostic performance has been highly inconsistent across laboratories. Available antibodies detect different epitopes of PD-L1. Recently, discordant patterns of PD-L1 expression in NSCLC were attributed to different affinities, cross reactivity, or variable expression of distinct target epitopes.

Second, tumor heterogeneity and sampling error may also affect detection of PD-L1. For example, although there are cases of diffuse PD-L1 expression, it can be very focal (ie, limited to the periphery of the tumor). Furthermore, the expression of PD-L1 is dynamic. While interferon-a is well known to upregulate PD-L1 expression,

 many other therapies may also affect PD-L1 expression. For example, doxorubicin was found to downregulate cell surface expression of PD-L1 in  breast cancer cell lines and xenografts,

 whereas paclitaxel and etoposide increased PD-L1 cell surface expression in breast cancer cell lines.

 PD-L1 expression was upregulated by MAP-kinase activation in melanoma cells resistant to BRAF inhibition,

 and in an epidermal growth factor receptor (EGFR) mutant model of NSCLC, where EGFR inhibition downregulated expression.

 Accordingly, the choice of the antibody for PD-L1 detection, the timing of biopsy, sampling error, and prior and concurrent therapies may affect PD-L1 expression and its detection. While the FDA has not required a companion diagnostic test for the use of the PD-1 inhibitor pembrolizumab for the treatment of metastatic melanoma, reliable predictive biomarkers are needed. In addition, as we learn how other therapies affect PD-L1 expression we can begin to explore combinations of immune checkpoint inhibitors and other treatments.

Alternatives to the detection PD-L1 expression in biopsy specimens are also being explored. For example, circulating soluble PD-L1 may identify a subset of tumors expressing PD-L1. Circulating PD-L1 has been identified in the serum of patients with renal cell carcinoma, and is associated with more aggressive disease patterns.

 Additionally, phenotypical changes of circulating T cells may indicate the presence of PD-L1 expression at the tumor site. One group of investigators suggested that the presence of circulating T cells co-expressing CD8, CD11a and PD-1 could be indicative of PD-L1 expression by the tumor. In their study, the level of Bim, a pro-apoptotic molecule, was significantly higher in these CD8

 cells in patients with metastatic melanoma and prostate cancer,

 suggesting that Bim expression in these cells may be indicative of PD-1/PD-L1 interaction.

Immune checkpoint inhibitors also challenge the use of traditional response criteria. Shrinkage of tumors following checkpoint inhibition may be preceded by stable disease or transient pseudo-progression (tumor enlargement) from immune cell infiltration of tumor. This phenomenon is difficult for physicians to separate from progression of disease, and selected clinical trials now allow continuation of therapy as long as there is no symptomatic worsening (eg, NCT01905657). Ongoing research is focused on distinguishing tumor immune infiltration from tumor progression. For example, technetium-linked IL-2 single-photon emission computed tomography is being explored in patients with metastatic melanoma being treated with immunotherapy (NCT01789827). Detection of IL-2 receptor in this setting may help distinguish immune infiltration from tumor progression. Accordingly, this strategy may complement standard imaging modalities in the setting of progression on immune checkpoint inhibitors, providing a rationale to continue with immunotherapy when IL-2 receptor is detected at tumor sites.

Although we are just coming to understand the relevance of immune checkpoint blockade in many malignancies, we will see the exploration of these inhibitors in novel settings. While immune checkpoint blockade has primarily been tested in subsequent lines of therapy, trials for first-line therapy are being developed (NCT02041533) and may be worth exploring in MPM. Other clinical trials are exploring the possibilities of various combinations of immune checkpoint inhibitors. For example, combined anti-CTLA-4 and anti-PD-1 blockade was tested in metastatic melanoma and was associated with a higher response rate than seen with either agent alone.

 In addition, clinical trials are testing PD-1 blockade with immune checkpoint inhibitors against lymphocyte activation gene 3 (LAG-3) (NCT01968109) and killer immunoglobulin receptor (KIR) (NCT01714739). Recent animal models have also shown that PD-1/PD-L1 axis blockade may enhance the effects of oncolytic virotherapy.

 This strategy may be considered in future clinical trials in light of ongoing studies exploring the safety and efficacy of intrapleural viral therapy in MPM (NCT01503177 and NCT01721018).

Immune checkpoint inhibition represents a promising approach to the treatment of MPM. Larger studies to validate the degree of PD-L1 expression in MPM and to identify predictive biomarkers for anti-CTLA-4 and anti-PD1/PD-L1 therapy are needed. Overcoming difficulties with the detection of PD-L1, the modulation of PD-L1 expression, and improved assessment of treatment responses will enhance the utilization ofimmune checkpoint inhibition in MPM.  

Aaron S. Mansfield, MD, assistant professor of medicine and oncology, Department of Oncology, Division of Medical Oncology, Mayo Clinic, and Tobias Peikert, MD, assistant professor of medicine, Department of Internal Medicine and Department of Immunology, Division of Pulmonary and Critical Care Medicine, Mayo Clinic.

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Aaron S. Mansfield MD, Tobias Peikert, MD

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Potential Applications of Immune Checkpoint Blockade for Mesothelioma