Immune exhaustion can be a major clinical problem for patients who have cancer or chronic inflammation. Persistent antigen stimulation drives T cells to express multiple surface markers called immune checkpoints. When these markers bind to their corresponding ligands that are expressed by antigen (e.g. tumor cells), T cells become metabolically impaired and lose several important functions; some cell signaling pathways are inhibited, while other intracellular mediators are re-modulated. Eventually, both CD4+ and CD8+ T cells behave dysfunctionally in ways that may facilitate cancer progression. Immune checkpoints are a major hallmark of immune exhaustion. In addition, natural killer (NK) cells, a critical immune cell subset in the peripheral immune system, also express immune checkpoint molecules, and are responsible for detecting and destroying circulating tumor cells. Yet, little research has investigated immune checkpoints on NK cells. Here, we explored the role of Bromodomain and ExtraTerminal domain (BET) proteins (BRD2, BRD3, BRD4), which are important transcriptional co-regulators, and critical for proliferation and metastasis in many cancer types, in the regulation of immune checkpoint molecules in several immune cell subsets, including CD4+ and CD8+ T cells, and NK cells. Through binding to acetylated histone tails of nucleosomal chromatin, BET proteins assist in transcription of multiple genes. Deregulated expression of BET proteins promotes cancer development or tumor cell metastasis, and new data show the BET proteins contribute to immune exhaustion. Furthermore, Type 2 diabetes mellitus (T2DM) is another worrisome problem related to cancer. T2DM patients show increased risk of developing cancer. Patients with both T2DM and any type of cancer show higher risks for metastasis. Significantly, T2DM patients also show immune exhaustion, suggesting a hypothesis that BET proteins may couple immune system dysfunction, abnormal metabolism and cancer incidence or progression. Specifically, T2DM has been defined to be a metabolic and a chronic inflammatory disease. The 5' Adenosine Monophosphate-activated Protein Kinase (AMPK) signaling pathway is a key pivot of cell metabolism and as well a significant target of drugs that normalize blood glucose, such as metformin. Based on published data, we considered that it is important to explore the mechanism of how immune checkpoints are regulated through metabolic pathways, focusing on immune exhaustion in T2DM patients. Moreover, considering that the expression of BET proteins promotes cancer development and progression, and metastasis and immune exhaustion are characteristic of many cancers as well, we suspected a potential relationship among BET proteins, the AMPK metabolic signaling pathway and immune exhaustion is worth exploring. Here, we measure expression of the immune checkpoint molecules TIM-3, TIGIT, PD-1, and CTLA-4 on normal T cells and NK cells by flow cytometry. We demonstrate different degrees of regulation of immune checkpoints by BET proteins on stimulated T cells and NK cells. Comparing stimulated-only cells with stimulated-plus AMPK inhibitor cells, we found that inhibition of the AMPK signaling pathway causes divergent expression patterns for TIM-3 and TIGIT, PD-1 and CTLA-4. Simultaneous inhibition of both BET proteins and the AMPK signaling pathway, shows that BET proteins regulate TIM-3 and TIGIT through an AMPK-independent metabolic pathway and regulate PD-1 and CTLA-4 through an AMPK-dependent pathway. Overall, we show TIM-3 and TIGIT, PD-1 and CTLA-4 display different expression patterns under regulation of the AMPK signaling pathway, and we show that BET proteins regulate TIM-3, TIGIT, PD-1 and CTLA-4 through both AMPK-dependent and -independent pathways. These findings are important because they reveal novel mechanisms of immune checkpoint regulation, which may be valuable for targeting in cancer patients who are being treated with checkpoint inhibitors.
| Identifer | oai:union.ndltd.org:bu.edu/oai:open.bu.edu:2144/41265 |
| Date | 06 July 2020 |
| Creators | Huang, Kunlin |
| Contributors | Denis, Gerald V. |
| Source Sets | Boston University |
| Language | en_US |
| Detected Language | English |
| Type | Thesis/Dissertation |
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