Abstract SY26-03: Immune effects of targeted therapy and implications for combination strategies
Authors: Wargo Jennifer A. 
Published: August 2015

Abstract
There have been significant advances in cancer treatment over the past several years, particularly with the use of molecularly targeted therapy and immunotherapy. However with these advances, we are posed with therapeutic dilemmas with regard to timing and sequence of therapy. Namely, there is significant debate as to whether to begin treatment with targeted therapy versus immunotherapy upfront, and at which point to change treatment strategy. This is highly relevant, as each of these treatments as mono-therapy have significant limitations. An example of this is in targeted therapy for melanoma, where treatment with BRAF-targeted agents results in responses in the majority of patients - though responses are not durable. This is also evident in melanoma immunotherapy (such as immune checkpoint blockade) where treatment results in responses in a smaller proportion of patients, though these responses tend to be more durable.
There a strong clinical rationale for using these strategies in combination, and this concept is being empirically tested in clinical trials. In addition, there is a strong scientific rationale for combining molecularly targeted therapy and immunotherapy. This is based on growing evidence that oncogenic mutations may affect anti-tumor immunity, and that blocking these mutations can make tumors more immunogenic. Thus, treatment with targeted therapy has significant effects on anti-tumor immunity. This has been best studied in melanoma, where over 50% of tumors harbor an activating mutation in the BRAF gene. Oncogenic mutations in the BRAF gene lead to several deleterious effects including immune evasion, and pharmacologic inhibition of BRAF (via selective inhibitors of BRAF-V600E or MEK inhibitors) in vitro leads to enhanced immunogenicity of melanoma tumors. Inhibition of BRAF also leads to immune effects in vivo, as treatment with BRAF-targeted therapy in patients with metastatic melanoma is associated with increased immunogenicity of melanoma tumors within 2 weeks of therapy - with increased melanoma antigens and CD8+ T cell infiltrate, and decreased immunosuppressive cytokines and VEGF. However concurrently, there is an increase in expression of immunomodulatory molecules on the surface of T cells (such as PD-1 and Tim-3) and in the tumor microenvironment (such as PD-L1) in the microenvironment. Together, these likely contribute to immune-based adaptive resistance to this form of therapy.
There are several important considerations in combining targeted therapy with immunotherapy for cancer. The kinetics of the immune response to targeted therapy is an important issue, and preliminary evidence suggests that the favorable effects on the tumor microenvironment are early and transient, with infiltrating CD8+ T cells increased at 2 weeks on treatment but essentially absent several weeks into the course of therapy. Another important consideration is the effect of the molecularly targeted therapy on immune cells themselves. This is critical, as T cell function is dependent on many of the same signaling pathways that are upregulated (and targeted) in transformed cells. This is particularly evident in the MAPK pathway. Interestingly, selective inhibitors of BRAF-V600E do not have deleterious effects on T cell function and actually augment T cell signaling through paradoxical activation of non-mutant RAS. However MEK inhibitors have a profound deleterious effect on T cell function in vitro, with decreased proliferation and an abrogated response. Studies in human clinical trials demonstrate no significant decrease in T cell infiltrate in melanoma tumors in the setting of combined BRAF and MEK inhibition, though the function of the infiltrating T cells has not been well studied. The effect of MEK inhibitor monotherapy on tumor-infiltrating T cells in patients has also not been well evaluated, and is a critical point as MEK inhibitors may be more widely applicable in combinations of targeted therapy and immunotherapy for NRAS-mutant melanoma and for other cancer types. There is ample literature suggesting a deleterious effect of MEK inhibitors on T cells and other immune subsets in vitro, however the effects of MEK inhibitors on peripheral blood lymphocytes or tumor infiltrating lymphocytes in patients on therapy have not been well studied. Recent in vivo murine studies suggest that “timing is everything,” and that MEK inhibitors may demonstrate synergy with immune checkpoint blockade in the treatment of RAS-mutant cancer if both are given concurrently or if MEK inhibitors were given prior to treatment with PD-1 blockade (but was not seen if PD-1 was given prior to MEK inhibitors). There is data to suggest that MEK inhibition has different effects depending on the stage of T cell memory differentiation (with MEK inhibitors adversely affecting naïve T cells and having little effect on effector memory T cells).
Another issue in combining molecularly targeted therapy and immunotherapy for cancer is the type of immunotherapy used in the regimen and the potential for toxicity. Murine studies have shown synergy with targeted therapy and various forms of immunotherapy (including immune checkpoint blockade and adoptive cell transfer). However not all forms of immunotherapy are created equal, and differences in responses are likely based on mechanistic differences between the strategies. Clinical trials are currently underway combining targeted therapy (against BRAF or MEK) and immune checkpoint inhibitors (against CTLA-4, PD-1, and PD-L1), and are well-founded given the data demonstrating an increased expression of immunomodulatory molecules in tumors of patients on BRAF inhibitors. Trials are also underway combining BRAF-targeted therapy with adoptive cell transfer using autologous tumor infiltrating lymphocytes. Response data from these trials is not mature, and it is not yet clear that synergy will be seen when these regimens are combined. Nonetheless there are some early hints that this strategy may enhance immune responses above either therapy alone. However despite the enthusiasm for this approach, there have been issues with unexpected toxicity in these some of these combination trials, and this needs to be taken into careful consideration.
As we move forward, there is clearly some evidence for synergy between molecularly targeted therapy and immunotherapy, though a great deal of research lies ahead. In addition to combining approved strategies, novel targeted agents (e.g. MEK, CDK4, PI3K, MDM2, FGFR, c-MET) and immunotherapeutic approaches (e.g. against TIM-3, Lag-3, GITR, OX40, CD27) are now available and are in clinical trials, and could be used in combination. In addition, “triplet” strategies (such as BRAF inhibitor + MEK inhibitor + immune checkpoint inhibitor) now exist, thus these combination strategies are becoming increasingly complex, with a large number of potential combinations / schedules. A better understanding of mechanisms of response and resistance to each of these forms of therapy is critical, and will help guide us on how best to combine these strategies. Central to this understanding is translational research conducted on longitudinal patient samples during the course of therapy, which may inform (and be informed by) parallel in vitro and murine studies. Overall, this represents a changing paradigm in cancer treatment, with the opportunity to capitalize on the complex interplay between genomics and anti-tumor immunity to improve responses to therapy.
Citation Format: Jennifer A. Wargo. Immune effects of targeted therapy and implications for combination strategies. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr SY26-03. doi:10.1158/1538-7445.AM2015-SY26-03