Elsevier

Seminars in Oncology

Volume 42, Issue 3, June 2015, Pages 363-377
Seminars in Oncology

Immune Checkpoint Protein Inhibition for Cancer: Preclinical Justification for CTLA-4 and PD-1 Blockade and New Combinations

https://doi.org/10.1053/j.seminoncol.2015.02.015Get rights and content

Over the last two decades, our understanding of the molecular basis of immunity has revealed the complexity of regulatory pathways involved in immune responses to cancer. A significant body of data support the critical importance of immune checkpoints in the control of the adaptive immune response to malignancy, and suggest that inhibitors of those checkpoints might have significant utility in treating cancer. This has been borne out by the recent US Food and Drug Administration (FDA) approvals of two different antibodies, one against cytotoxic T-lymphocyte antigen-4 (CTLA-4) and one against programmed death-1 (PD-1). Here, we provide a comprehensive review of the literature regarding the preclinical justification for the use of CTLA-4 and PD-1 blockade as monotherapy, and as combination therapy in the treatment of cancer. The animal data strongly supported the use of these drugs in patients, and in many cases suggested strategies that directly led to successful registration trials. In contrast, many of the toxicities, and some of the unusual response patterns seen in patients with these drugs, were not predicted by the preclinical work that we cite, highlighting the importance of early-phase trials with patients to inform future drug development. In addition, we review herein the preclinical data surrounding emerging immune checkpoint proteins, including BTLA, VISTA, CD160, LAG3, TIM3, and CD244 as potential targets for inhibition. The current comprehensive review of the literature regarding CTLA-4 and PD-1, as well as a number of novel checkpoint proteins demonstrates a strong preclinical basis for the use of these antibodies singly and in combination to overcome checkpoint inhibition in the treatment of cancer. We also suggest that the use of these antibodies may augment the efficacy of other activating immune antibodies, cytokines, radiation, and adoptive cell therapy in human cancer.

Section snippets

CTLA-4: Preclinical Background

The enormous progress made in the last two decades in our understanding of the molecular basis of immunity has revealed the complexity of regulatory pathways involved in immune responses to cancer. Key pathways that are altered in T cells derived from patients with cancer have been identified, and investigators have developed new reagents, models, and tools to augment T-cell immunity in experimental systems. T cells of the adaptive immune system recognize antigen in association with major

Mechanism of Action: Anti-Tumor Effects

Early experiments in animal model systems indicated that CTLA-4 abrogation employing an anti-murine antibody had few anti-tumor effects when established non-immunogenic tumors were treated. However, the addition of a GM-CSF–transduced cell vaccine in melanoma, breast and prostate cancer animal models revealed potent anti-tumor activity for CTLA-4 antibody, with the simultaneous onset of autoimmune vitiligo in the B16 model. No other autoimmune phenomena were observed in these animal models.24,

Preclinical Background

PD-1 is an inhibitory checkpoint molecule expressed on activated T and B cells and monocytes, and like CTLA-4 is involved in regulating the balance between immune activation and tolerance.55, 56, 57 The induction and maintenance of T-cell tolerance requires PD-1, and the presence of its principal ligand PD-L1 on non-hematopoietic cells can limit effector T-cell responses and protect against immune-mediated tissue damage. Its ligands PD-L1 and PD-L2, or B7-H1 and B7-H2, are expressed on APCs, a

Anti-Tumor Mechanism of Action

PD-L1, the principal ligand of PD-1, is constitutively expressed on murine and human T and B cells, dendritic cells, endothelial cells, macrophages, mesenchymal stem cells, and bone marrow–derived mast cells. PD-L1 is found to be aberrantly expressed on tumors, which is likely to result in tumor-induced immune suppression via signaling to PD-1. PD-L1 expression has been found in situ by immunohistochemical analysis on a wide variety of solid tumors, including breast, lung, colon, ovarian,

Combination Therapies

Combination PD-1 and CTLA-4 blockade increased effector T-cell infiltration into B16 melanoma in mice, resulting in an increased T-effector-to-Treg ratio within the tumor. The fraction of tumor-infiltrating T-effector cells expressing CTLA-4 and PD-1 increased, reflecting the proliferation and accumulation of cells that would otherwise have been “exhausted” and anergized.40 In mice, a triple treatment therapy of anti–PD-L1 blockade, depletion of CD4 T cells (primarily Tregs), and irradiated

New and Novel Checkpoint Proteins

Increased expression of a variety of newly defined checkpoint proteins including LAG-3, BTLA, and TIM3 have been noted on T cells found within the tumor microenvironment compared with their counterparts in the blood.128 Melan-A specific T cells in melanomas expressing those proteins seemed to exhibit a gene expression profile matching that of exhausted T cells and was associated with impairment of cytokine production and overall T-cell function.129 BTLA and TIM-3 were predominantly expressed on

BTLA

B- and T-cell lymphocyte attenuator (BTLA) is an immunoglobulin-like molecule that is part of the CD28 family and is expressed on B cells, T cells, NK cells, and APCs.133, 134, 135 Although present during all stages of T-cell differentiation, it is predominantly found on naïve T cells.130, 131, 132 Its expression patterns, structure, and key role in early T-cell regulation suggest a role as an early T-cell response gene. The binding of BTLA to its ligand, herpes virus entry mediator (HVEM),

VISTA

V-domain Ig suppresor of T-cell activation (VISTA) is a negative checkpoint ligand that shares homology with PD-L1 (in terms of structure but not expression pattern) and is a potent T-cell suppressor.140, 141, 142 It is a member of the Ig superfamily. Human VISTA is predominantly expressed in hematopoietic tissue (such as spleen, lymph nodes, and blood) or tissues containing significant numbers of infiltrating leukocytes. Immunohistochemical analyses suggested that VISTA expression was greatest

CD160

CD160 is a glycosylphosphatidylinositol (GPI)-anchored protein found on the surface of CD8+ T cells, NK cells, and NK-T cells.143, 144, 145, 146 CD160 also is expressed on CD4+ T cells and its function has not been well elucidated. T-cell proliferation and cytokine production was restricted with blockade of CD160 using a high-affinity antibody.147 While MHC class 1 molecules were felt to be the primary receptor binding CD160, albeit at low binding affinity, CD160 was subsequently defined as a

LAG-3

Lymphocyte activation gene 3 (LAG3) is a transmembrane protein that belongs to the Ig superfamily that binds MHC class II, enhances Treg activity, and negatively regulates T-cell proliferation and differentiation.148 It is expressed on B cells, a subset of T cells, NK cells, and tumor-infiltrating lymphocytes.149 It is overexpressed on CD8+ T cells in a tolerized envionrment, i.e one in which there was chronic exposure to antigen resulting in T cell anergy with low production of effector

TIM3

T-cell immunoglobulin and mucin domain 3 (TIM3) is an inhibitory molecule expressed by IFN-gamma secreting T helper-1 (Th1) cells, dendritic cells, monocytes, CD8+ T cells, and restricted lymphocyte subsets.152, 153 Binding of TIM3 to its ligand, galectin-9, found on many tumors, has resulted in Th1 cell death. Blockade of TIM3 has been shown to increase circulating IFN-gamma–secreting cells.154, 155 In mice, single-agent blocking antibody therapy against TIM3 was well tolerated, as was

CD244

The signaling lymphocyte activation molecule–related receptor (SLAMF4) or 2B4 (CD244) is an important immune regulator that is a member of the immunoglobulin superfamily, mediating potent stimulatory and co-stimulatory signals in both T cells and NK cells.157 2B4 inhibits NK cytotoxicity against CD48-expressing tumor cells, yet 2B4 signaling augments the antigen-specific activation responses induced by TCR engagement. 2B4 cross-linking can lead to both inhibition and activation of T-cell

Conclusion

These data provide a strong preclinical basis for the use of antibodies alone and in combination to overcome checkpoint inhibition, and to augment the efficacy of immune stimulating antibodies, radiation, and adoptive cell therapy in human cancer. In spite of the strong data provided herein, the exact mechanistic basis for the anti-tumor effects in murine and human tumors after abrogation of one or more checkpoint proteins remains unclear. Much work needs to be done to understand how to

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