(1)
Institute of Pathology, Medical University Graz, Graz, Austria
After being used in the therapy of melanomas, immunotherapy is now also used in NSCLC. A major breakthrough in establishing this type of therapy was the finding of immunomodulation of T-cell response through immune checkpoints inducing immune evasion of cancer cells. Cytotoxic T-lymphocyte antigen-4 (CTLA-4) was found in dendritic cells and regulatory T cells (Tregs) in lymph nodes but also circulating in the blood inducing apoptosis and cell death of cytotoxic T lymphocytes [1]. Programmed cell death 1 (PD1) and its ligand PDL1 were found on tumor cells as well as on lymphocytes inducing immune tolerance [2]. Based on these findings, antibodies for PDL1 and PD1 were tested for their ability to interfere with this mechanism of immune tolerance against tumor antigens. Important efforts are under way and have focused on this new treatment modality [2–10]. At present several pharmaceutical companies have developed checkpoint inhibitors (humanized monoclonal antibodies) for treatment of NSCLC patients, and numerous phase II and III studies have been performed. The FDA and EMA have recently approved three of these drugs for treatment of NSCLC. Whereas some drugs are selectively approved for SCC, others may be used in all NSCLC [9, 11–14].
Like other solid tumors, pulmonary carcinomas have developed several mechanisms by which they escape the attack of cytotoxic immune cells. Some of these mechanisms have recently gained special attention, the programmed death 1 – programmed death ligand 1 and 2 (PDL1/2-PD1) – and the CTLA-4 system. Tumor cells as well as lymphocytes can express ligands for PD1 [4]. By this they interact with surface molecules on CD8+ T cells causing apoptosis and further more influence also the microenvironment via orchestration by cytokines, which all together cause immune tolerance. Therapy using antibodies against PDL1 has shown significant improvement experimentally as well as in clinical studies to restore the cytotoxic attack of T lymphocytes toward tumor cells in several solid malignancies [3, 6, 10]. Parallel to targeting PDL1 by anti-PDL1 antibodies, also antibodies against the receptor PD1 have been created and used experimentally and in clinical trials with success [2, 4, 5, 7, 8]. In some clinical trials, specific immunohistochemistry tests for the expression of PD1 and PDL1 were used to select those patients, who might best respond to the antibody-based therapy. A strong staining in at least 5 % of tumor cells and/or lymphocytes – or 50 % in another trial [15] – was regarded as a positive result and predictive for outcome. Looking up clinical data of these trials, it is evident that the majority of patients are detected by this simple stain. However, there exist also a not to be neglected amount of patients, which do not respond to treatment although being positive for PD1/PDL1 as well as patients who do respond despite being negative or low positive for these molecules [16–18].
Waterfall plot of patient response and immunohistochemical staining for PDL1 according to J. Brahmer (Presentation AACR 2015, Philadelphia)
Although proving PD1-PDL1 as targetable molecules within the immune regulatory system (Fig. 20.1), it is well known from nonneoplastic diseases that there are many more ways the immune system can be shifted toward antigen tolerance or even lymphocyte exhaustion. Several of these regulatory mechanisms are also targetable. Subsequent development of resistance toward the PD1-PDL1 therapy might be due to one of these immunomodulatory alterations. In addition patients might have developed tolerance against their tumor cells not only by expression of PD1-PDL1 but in addition by other tolerance mechanisms – this could be due in patients, who do not respond to anti-PDL1 therapy despite having PD1-PDL1 upregulated.
Fig. 20.1
Immunohistochemistry for PDL1 in a squamous cell (top panel) and an adenocarcinoma (bottom panel). There are differences in the expression of PDL1 between adenocarcinomas and squamous cell carcinomas, but also between the individual tumor cells. Bars 50μm
20.1 Systems Known to Be Able to Induce Immune Tolerance Toward Foreign Antigens
One of the most important cell types in this regard are antigen-presenting cells from the dendritic cell lineage. There are several populations, such as conventional dendritic cells, which promote immune attack by processing foreign antigens and present them to CD8+ T lymphocytes. Others such as plasmacytoid and granulocytic dendritic cells in contrary cause immune tolerance by cooperating either with regulatory T cells or by inducing an inflammatory environment, which promotes tumor cell invasion, spreading, and finally metastasis [19, 20]. The function of Langerhans cells, another dendritic cell type, is entirely unknown, but most probably acts against tumor antigens.
Plasmacytoid dendritic cells can also interact with monocytoid cells promoting the differentiation of macrophages into the tumor-promoting M2 lineage. Macrophages play a major role early on in carcinogenesis. Macrophages can induce angiogenesis and prepare and modulate stroma proteins in favor for invading tumor cells, thus promoting tumor growth, invasion, and metastasis [21, 22]. In contrary M1 macrophages might inhibit tumor progression not only in the early phase but also during metastasis formation [23, 24].
There are additional pathways by which tumor cells can influence the differentiation of M0 macrophages into M2 lineage. Whereas activation of Notch signaling increased M1 macrophages, blocking of Notch induced a M2 response. Wang et al. showed that RBP-J-mediated Notch signaling regulates the M1 versus M2 polarization through SOCS3 [23]. Notch is mutated in several lung carcinomas; however, if this also applies to tumor-associated macrophages has not been studied so far. Tumor cells might use SOCS3 signaling to induce M2 macrophage polarization.
Regulatory T cells (Tregs) can accumulate at the tumor site and block CD8+ T lymphocytes and NK cells in their antitumor action as well as inhibit influx of the cells into the tumor [25, 26].
Autophagy is another mechanism associated with immune cell modulation, although this mechanism is presently not fully understood. Autophagy is seen in many lung cancer types. Increase of autophagy in cancer cells liberates nutrients, decreases the formation of reactive oxygen species, and aids in the clearance of misfolded proteins. This provides a survival advantage for cancer cells in the tumor microenvironment. However, also immune cells infiltrate the tumor environment and encounter hypoxia, resulting in hypoxia-induced autophagy. Due to the fact that autophagy is crucial for immune cell proliferation as well as antigen presentation and T-cell-mediated killing of tumor cells, anticancer treatment strategies based on autophagy modulation will need to consider the impact of autophagy on the immune system [27]. In melanoma it has been shown that hypoxia leads to instability of gap junctional Cx43 and impairs melanoma cell killing by NK cells. Inhibition of autophagy by pharmacological approaches might restore NK-cell-mediated lysis of hypoxic melanoma cells [28].
Since a long time, bronchoalveolar lavage (BAL) is a tool used in the evaluation of inflammatory diseases of the lung. By BAL cells are washed out from lung lobes affected by an inflammatory/immune disease. Typing of lymphocytes allows to evaluate disease activity, impact of therapy, and in few diseases also the diagnosis of the disease [29–33]. In lung tumor BAL is most often used to collect tumor cells from peripheral tumors not otherwise accessible. However, this tool might be used to analyze also the percentages of immune cells in lung lobes bearing lung carcinomas.
References
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Peggs KS, Quezada SA. PD-1 blockade: promoting endogenous anti-tumor immunity. Expert Rev Anticancer Ther. 2012;12:1279–82.CrossRefPubMed
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