Berkeley Pharma Tech Journal of Medicine Correspondence: sanchithabpt1@gmail.com Keywords: Tumor microenvironment Immunotherapy Cytotoxic t lymphocytes PD-1 CTLA-4 Lactic acid Non-small cell lung cancer (NSCLC) Submitted August 11, 2023 Accepted May 23, 2024 Published June 28, 2024 Full Open Access Creative Commons Attribution License 4.0 Abstract The tumor microenvironment (TME) surrounds the tumor and includes blood vessels, immune cells, fibroblasts, signaling molecules, and the extracellular matrix. This review examines the cellular components and pathways within the TME, highlighting their potential as targets for immunotherapy. It also covers recent advances from the past several years in TME, immunotherapy, and combination therapy. The review emphasizes the role of CD8+ T cells in the TME and their relevance to immunotherapy. It discusses various T cell-targeted treatments, including PD-1/PD- L1, CTLA-4, VEGF, Interferon-ɣ, LAG-3, and ER stress-XBP1. Recognizing the complexity and uniqueness of each tumor's immunotherapy network, the review aims to understand and compare the TMEs of different cancers and the respective immunotherapies. Cancers covered include non-small cell lung Cancer (NSCLC), pancreatic ductal adenocarcinoma (PDAC), cervical cancer, chronic lymphocytic leukemia, and gastric cancers. The review also addresses future research directions and applications of immunotherapies, aspiring to advance TME understanding and research. Decoding the Interplay: Exploring Immunotherapy Resistance in the Tumor Microenvironment By: Sanchitha Kannabran, Akshaya Ajan, Leanne Lui, Zaden Yet 1. Introduction The tumor microenvironment (TME) constitutes the immediate surroundings of a tumor, forming a complex ecosystem inclusive of nearby blood vessels, immune cells, �broblasts, signaling molecules, and the extracellular matrix (ECM).1 Within the TME, there exist hypoxic and acidic conditions, alongside altered expression patterns of ECM proteins. These conditions foster the presence of resident and in�ltrating immunosuppressive cells, trigger the expression of immune checkpoint proteins, and promote the exclusion and exhaustion of cytotoxic T lymphocytes (CTLs).2 Such circumstances serve as the foundation for immunotherapy targeting the TME. While contemporary interest in TME immunotherapy appears recent, investigations into utilizing immune cells for combating tumors date back to the 1900s.3 New studies are directed towards pinpointing cellular pathways within the TME with the use of immunotherapy components to hinder growth. Among the primary pathways are lactic acid accumulation, metabolic processes, and diverse signaling routes within the TME. 5,6,7 All these pathways represent potential targets for immunotherapy interventions. Moreover, treatments are geared towards enhancing the immune system's ability to eradicate cancer cells and impede their proliferation. To achieve this goal, two primary strategies are employed: immune checkpoint inhibition and adoptive cellular therapy (ACT).4 Immune checkpoint inhibition involves obstructing certain proteins, known as checkpoints, to enhance the responsiveness of immune cells.4 Adoptive cell therapy (ACT) entails administering T cells to patients to bolster their ability to combat tumor cells.4 In this discussion, our focus lies on T cell-based immunotherapies, including anti-PD-1/PD-L1, anti-CTLA-4, VEGF, Interferon-ɣ (IFN-ɣ), LAG-3, and ER stress-XBP1. However, the complexity of immunotherapy arises from the variable responses of individual cancers to di�erent immunotherapeutic approaches. This review delves into the distinctive attributes of cancers such as non-small cell lung cancer (NSCLC), Berkeley Pharma Tech Journal of Medicine | 114 melanoma, pancreatic cancer, cervical cancer, and chronic lymphocytic leukemia. 2. Immune Cell Overview The extracellular matrix (ECM) is comprised of collagen, �bronectin, elastin, and laminin, serving as the structural framework within the tumor microenvironment (TME). Within this environment, various immune cells including T cells, B cells, Natural Killer cells (NK), macrophages (M1 and M2), stromal cells, endothelial cells, and cancer-associated �broblasts (CAFs) play pivotal roles in either promoting or suppressing tumor growth.8 T cells, equipped with T-cell receptors, target speci�c antigens on tumor cells, halting angiogenesis—the formation of new blood vessels—and leading to tumor cell destruction. However, T cells can become in�ltrated and dysfunctional within the immune system.8,9 B cells contribute to antibody production, antigen presentation, and cytokine secretion. 8,9 In the context of lymphocytes, both T and B cells, particularly tumor-in�ltrating lymphocytes (TILs), are crucial in research. While TILs e�ectively eliminate tumor cells, they can be recruited by the TME, compromising immune response e�cacy.4 Regulatory T cells (Tregs) maintain immune response balance, but when recruited to the TME, they hinder T cell formation, posing a danger to immune function. Consequently, immune mechanisms originally aimed at protecting the body now facilitate TME growth.10 NK cells surveil the bloodstream for tumor cells, inhibiting metastasis.8,9 Macrophages regulate immune responses by phagocytosing pathogens and presenting antigens. Notably, increased macrophage in�ltration in tumors, particularly the M2 phenotype, and cytokine secretion, support tumor growth.8,9 Berkeley Pharma Tech Journal of Medicine | 115 Figure 1: This figure illustrates the distinct contributions of various immune cells to the Tumor Microenvironment (TME). It delineates the overall immune response attributed to each mentioned immune cell type: T cells, B cells, Macrophages (M1 and M2), Natural Killer cells (NK), Tregs, and CD8+ cells. The upper side represents the positive immune response, while the lower side depicts the autoimmune suppressive response. Stromal cells, including endothelial cells, �broblasts, adipocytes, and stellate cells, play crucial roles in tumor development. Tumor cells recruit these supporting cells from nearby tissue to aid in tumor formation. Stromal cells secrete various factors that in�uence processes like angiogenesis, proliferation, invasion, and metastasis.8,9 Endothelial cells, for instance, are pivotal in orchestrating blood vessel formation and signi�cantly contribute to cancer progression by promoting cancer cell migration, invasion, angiogenesis, and metastasis. They transition into cancer-associated �broblasts (CAFs), facilitating communication between tumor cells and the tumor microenvironment (TME).1 This interaction often leads to the disruption of cell connections and detachment. CAFs can derive from various immune cells, but within the TME, they become Berkeley Pharma Tech Journal of Medicine | 116 signi�cant producers of extracellular matrix (ECM), tumor growth factors, cytokines, and other essential components. Each stromal cell type contributes uniquely to the TME, and targeting their speci�c functions presents an opportunity for novel immunotherapeutic approaches.1 The immune response involves numerous intricate processes, among which angiogenesis stands out—a phenomenon where endothelial cells proliferate and migrate, forming new blood vessels.11 This process is pivotal as it grants tumors the capability to metastasize, spreading to other locations within the body. Conversely, inhibiting angiogenesis can e�ectively curtail tumor growth and metastasis.11 For instance, research conducted by Chen et al. demonstrates that the administration of anti-malarial drugs like dihydroartemisinin and artesunate resulted in a reduction in the size of cell lines. These drugs were found to hinder the growth factors crucial for cancer cell proliferation, thereby impeding angiogenesis.11 Anti-angiogenic (AA) therapy plays a signi�cant role in remodeling the extracellular matrix (ECM), altering the distribution of cell types and populations.12 This therapy redistributes pericyte proteins along blood vessels, increasing coverage and eliciting a pro-aggressive tumor response. Notably, glycosylation of these proteins is speculated to contribute to malignant resistance to AA therapy.12 However, a complication arises as resistance can develop post-AA treatment, fostering tumor cell migration and invasion. Tumor cell receptors possess mechanisms to detect AA-induced alterations in their environment, prompting a remodeling of the entire tumor microenvironment (TME) to facilitate tumor growth once more.12 3. T Cell Based Immunotherapy This section of the review will delve into several key surface cell receptors found on endothelial cells, tumor cells, and T-cells. It will explore their roles in promoting tumor growth, strategies to overcome resistance, and the Berkeley Pharma Tech Journal of Medicine | 117 underlying mechanisms of resistance. Emphasis will also be placed on the critical functions of in�ltrating tumor lymphocytes and cytotoxic T cells. Within the tumor microenvironment (TME), various surface cell receptors recognize speci�c antigens, initiating signaling cascades that ultimately impact downstream events. These cascades often lead to the inactivation or impairment of e�ective T-cells, which are typically responsible for targeting and eliminating tumor cells. The TME, being highly adaptable, fosters conditions conducive to tumor cell growth, proliferation, and survival.13 Several immune response cells contribute to immune resistance, including myeloid-derived suppressor cells (MDSCs), regulatory T cells (Tregs), tumor-associated macrophages (TAMs), tumor-associated neutrophils (TANs), and endothelial cells. The secretion of negative regulatory factors by these cells can induce T-cell exhaustion, dysfunction, and apoptosis.13 Figure 2: A. Vascular Endothelial Growth Factor-A (VEGF-A) binds to Vascular Endothelial Growth Factor Receptors (VEGFRs) situated on endothelial cell surfaces, initiating a series of reactions that stimulate angiogenesis. B. The interplay between proteins located on T cell surfaces and their corresponding ligand surface cell receptors on tumor cells contributes to T cell exhaustion. Berkeley Pharma Tech Journal of Medicine | 118 One e�ective therapeutic approach, immune checkpoint inhibition, has demonstrated success in rejuvenating the cytotoxic function of CTLs and T-cells. This therapy works by alleviating the suppression of antitumor immunity, freeing CTLs from exhaustion.13 The FDA has approved antibodies targeting Programmed Cell Death 1 (PD-1/PD-L1) and Cytotoxic T Lymphocyte-Associated Protein 4 (CTLA-4) for treating various cancers. Additionally, ongoing clinical research is targeting other immune checkpoints like LAG3, CD39, CD73, and CD47, all showing promising potential in advancing cancer immunotherapy.14 3.1 PD-1/PD-L1 Immune checkpoint inhibition stands out as an e�ective therapeutic approach in reviving the cytotoxic capabilities of CTLs and T-cells. By alleviating the exhaustion-induced suppression of antitumor immunity, this therapy unleashes CTLs to combat tumors.13 The FDA has approved the use of antibodies targeting Programmed cell death 1 (PD-1/PD-L1) and cytotoxic T lymphocyte-associated protein 4 (CTLA-4) as treatments across various cancer types.13 Numerous other immune checkpoints are currently under investigation in clinical research, including LAG3, CD39, CD73, and CD47, all of which show promising potential in advancing the landscape of cancer immunotherapy.14 Among these, PD-1 is a pivotal protein implicated in T cell apoptosis, or programmed cell death. It is predominantly expressed on the surface of activated immune cells such as macrophages, dendritic cells, B cells, and T cells.14 Notably, PD-1 expression is particularly elevated on exhausted T cells, where it plays a role in inhibiting their normal immune function. When PD-1 interacts with its ligand receptor, PD-L1, on tumor cells, it initiates a series of signaling events that directly impede the response of activated cells by enhancing local evasion mechanisms.14,15 These immune checkpoint inhibitors (ICIs) interfere with T cell functionality, leading to Berkeley Pharma Tech Journal of Medicine | 119 an increase in exhausted T cell populations, both of which contribute to adaptive resistance against immune checkpoint blockade therapy.16 Researchers conducted clinical trials investigating the interaction between PD-1 and PD-L1 on cytotoxic T cells in mice. They observed accelerated tumor growth, which was mitigated when treated with an anti-PD-L1 antibody or through PD-1 knockout, resulting in reduced tumor growth. Similar �ndings were noted in human trials.17 For instance, Qian et al. demonstrated in a study focusing on glioma cancer that tumor-in�ltrating T cells become activated and can increase PD-1 expression, leading to T cell dysfunction mediated by Immune Checkpoint Inhibitors (ICIs). Consequently, the researchers suggested that anti-PD therapy could impede glioma cancer progression.17 Another instance illustrating the e�ectiveness of immune checkpoint inhibition therapy in microsatellite instability/de�cient mismatch repair (MSI/dMMR) tumors comes from the ongoing trial conducted by IMHOTEP, which showcases a group of 120 patients bene�ting from anti-PD therapy.18 While anti-PD therapy has shown success in certain patients, others have exhibited poor responses due to both primary and acquired resistance mechanisms working against the treatment. E�ective anti-PD therapy hinges on the precise blockade of the PD-1 and PD-L1 pathways. However, if either protein lacks expression, the treatment proves ine�ective against the tumor cells—a direct primary resistance mechanism observed in some cases.15 Furthermore, certain cancers lack tumor-in�ltrating T cells or PD-L1, rendering them unresponsive to anti-PD therapy.15 To combat this adaptive resistance, combination therapy involving both immuno- and chemo-therapies has been proposed as a more e�ective approach to treating cancer compared to immunotherapy alone. 3.2 CTLA-4 CTLA-4, a protein associated with cytotoxic T lymphocytes, plays a role in dampening the anti-tumor immune response by regulating the activation of CTLs, thus hindering an e�cient immune reaction.14 Unlike PD-1, which manages programmed cell death, CTLA-4 dictates whether a T cell Berkeley Pharma Tech Journal of Medicine | 120 undergoes activation or remains in a state of rest termed exhaustion. It is present on the surface of stimulated functional T cells, and activation occurs upon stimulation from a T cell receptor.14 CTLA-4 competes with CD28 for binding to the T cell ligand, blocking the crucial costimulation signals required for activation.14 Due to its higher a�nity for the ligand, CTLA-4 binds more e�ectively, leading to T cell exhaustion and inhibiting activation. Using an antibody that selectively targets and binds to CTLA-4 can restore T cell activation by eliminating the competition for T cell ligand binding with costimulation signals.14 Additionally, CTLA-4 is prominently expressed on the surface of Tregs, which play a role in immune response suppression.14 Research indicates that CTLA-4 plays a pivotal role in regulating the function and production of Tregs,14 which in turn directly suppress the activation of target immune cells by increasing CTLA-4 expression.19 Utilizing CTLA-4 blockade therapy has emerged as an e�ective strategy for enhancing anti-tumor immune responses.14 The FDA-approved monoclonal anti-CTLA-4 antibody, ipilimumab, has signi�cantly enhanced overall survival rates among patients with malignant melanoma.4 It's worth noting that while anti-CTLA-4 antibodies show e�cacy in late-stage melanoma, their e�ectiveness in other tumor types is limited. Consequently, clinical trials are exploring combination therapies to address this limitation.4 3.3 LAG-3 Lymphocyte Activation Gene 3 (LAG-3) serves as an immune checkpoint receptor, capable of dampening responses orchestrated by T and NK cells, thereby fostering a hyporesponsive condition, which aids tumors in escaping immune surveillance. Typically found on activated and exhausted T and NK cells, B cells, dendritic cells, and Tregs, LAG-3 signaling inhibits T cell proliferation, cytokine generation, and cytolytic activity.20 Additionally, its presence on Tregs contributes to immunosuppression.20 An illustrative instance of this phenomenon is evident in individuals a�icted with chronic lymphocytic leukemia (CLL), where there is signi�cant dysregulation of LAG-3. Both NK cells and T cells exhibit Berkeley Pharma Tech Journal of Medicine | 121 heightened LAG-3 expression, and elevated levels of LAG-3 or soluble LAG-3 (sLAG-3) are associated with adverse cytogenetics and unfavorable outcomes among CLL patients. Consequently, treatment involving the application of an anti-LAG-3 blocking antibody called relatlimab to peripheral blood mononuclear cells (PBMCs) has demonstrated e�cacy in reducing leukemic cell counts and restoring NK cell and T cell mediated responses.20 As a result, interventions aimed at reinstating T and NK cell mediated responses through LAG-3 blockade have emerged as promising therapies for hematological malignancies such as CLL, follicular lymphoma (FL), di�use large B cell lymphoma (DLBCL), and acute myeloid leukemia (AML).20 3.4 ER Stress/IRE1a-XBP1 A fresh angle in cancer therapy involves focusing on the ER stress pathway, with a particular emphasis on the XBP1 gene.21 In a 2022 study conducted by Zundell et al., investigating ovarian clear cell carcinomas (OCCC), it was discovered that the ARID1A gene, responsible for epigenetically regulating gene expression via the SWI/SNF chromatin remodeling complex, is mutated in over 50% of OCCCs. This mutation results in inadequate expression of ARID1A protein, leading to advanced-stage disease and early recurrence.21 This element plays a vital role by transcriptionally inhibiting the IRE1𝛂-XBP1 pathway of the ER stress response. Their research using mouse models indicates that disabling XBP1 and inhibiting the IRE1𝛂/XBP1 pathways signi�cantly enhances the survival rate of mice with OCCC, as it suppresses the growth of ARID1A mutant OCCC cells.21 In the face of ER stress, the IRE1-𝛂 component, a type of unfolded protein response (UPR), undergoes a structural change, splicing the mRNA encoding the XBP1 transcription factor. This splicing facilitates the translation of XBP1, which in turn aids cancer cell survival by resolving ER stress.21 ER stress itself fosters the survival of cancer cells by activating adaptive programs through the unfolded protein response (UPR) once detected. Berkeley Pharma Tech Journal of Medicine | 122 Consequently, inhibiting the UPR is emerging as a therapeutic strategy for cancers characterized by heightened ER stress response.21 In this study, the e�cacy of a therapeutic intervention targeting the IRE1𝛂 RNase, B-I09, was examined. By inhibiting IRE1𝛂 RNase activity, B-I09 e�ectively suppressed the growth of ARID1A-activated cells in ovarian clear cell carcinomas (OCCCs), indicating the potential of targeting the IRE1𝛂-XBP1 axis of the ER stress response as a promising approach for treating ARID1A mutant OCCCs.21 A study by Ma et al. illustrated the role of cholesterol, the primary sterol distributed throughout the human body, in the ER stress-XBP1 pathway.22 Through staining techniques to observe cholesterol levels in tumor-in�ltrating T cells in mice, researchers found that elevated cholesterol content triggers CD8+ T cell exhaustion via the ER stress-XBP1 pathway.22 Inhibiting the XBP1 pathway or reducing cholesterol levels in CD8+ T cells restored normal antitumor function, while increased cholesterol was consistently linked to elevated PD-1 expression on tumor-in�ltrating T cells.22 Similar patterns were observed in human colon cancer and myeloma samples, underscoring the signi�cance of cholesterol content in T cell exhaustion.22 3.5 VEGF/VEGF-A: Vascular endothelial growth factors (VEGFs) are signaling proteins crucially involved in the microvasculature of tumors, the suppression of immune cells, and the promotion of immunosuppression, both locally and systemically, in cancer.19 These proteins are predominantly secreted by the endothelial cells lining blood vessels, with additional production occurring in immune cells within the tumor microenvironment (TME).19 VEGFs play a pivotal role in fostering tumor development and progression by interacting with receptors on tumor cells through autocrine and paracrine signaling pathways. Among them, VEGF-A primarily binds to VEGFR-2, triggering the proliferation and migration of endothelial cells, thereby facilitating angiogenesis.19 Berkeley Pharma Tech Journal of Medicine | 123 VEGF-A boosts the expression of both PD-1 and CTLA-4, dampening the body's ability to �ght tumors and reducing the activity of CD8+ T cells by transitioning them from an active state to an exhausted one, impairing their normal function. Blocking VEGF with anti-angiogenic agents can reverse the immune checkpoints' inhibitory e�ects.19 In a phase I trial led by Daniel Chiu et al., a combination of anti-PD-1 therapy and antiangiogenic agents was used to treat glioblastoma, a challenging form of brain cancer known for its resistance to ICI therapy. The �ndings revealed that this combination treatment was well tolerated, safe, and did not lead to any previously unreported adverse events. Furthermore, the results showed decreased levels of VEGF-A and other angiogenic factors in the tumor microenvironment, underscoring the relationship between VEGF and PD-1 expression.16 4. Overcoming Resistance to Immune Checkpoint Blockade Although the immune checkpoint blockade strategy has shown e�cacy in reviving the antitumor response within tumor-in�ltrating lymphocytes (TILs) and cytotoxic T cells, the tumor microenvironment (TME) employs adaptive resistance mechanisms against this therapy. Speci�cally, the upregulation of immune checkpoints such as PD-1 and CTLA-4 in response to targeted antibodies can foster this adaptive resistance.14 Moreover, the TME utilizes evasion tactics, such as enhancing PD-1 expression in response to T cell attacks, leading to the functional impairment of TILs.13,17 Additionally, the di�erentiation of CD8+ T cells poses a signi�cant challenge, as TILs develop resistance to antitumor treatments, resulting in a diverse CD8+ T cell subset.17 Researchers are exploring combination therapies, pairing immunotherapy with chemotherapy or radiation therapy, to address malignant tumors. In a study by Feng Y. et al., a dual gene therapy approach was investigated to counter adaptive resistance to immune checkpoint blockade therapy in both CD4+ and CD8+ T cells. The scientists developed a dual gene delivery system aimed at eliminating tumor adaptive resistance and restoring T cell function by modulating the expression of VEGF-A and PD-L1 proteins.14 Gene-silencing techniques were employed to decrease VEGF-A expression in the TME, reducing angiogenic activity and blocking PD-L1 immune Berkeley Pharma Tech Journal of Medicine | 124 checkpoint. The �ndings demonstrated an increase in the number of CD8+ T cells expressing granzymes B, leading to inhibited tumor growth.14 Furthermore, increasing VEGF-A gene silencing e�ectively alleviated tumor hypoxia. However, it's crucial to regulate the dosage of each treatment, as excessive pShVEGF-A antibody treatment can exacerbate tumor hypoxia by overly depleting tumor vessels. Upon treating tumor-infected mice with the dual gene therapy, scientists observed substantial inhibition of tumor growth and prolonged overall survival rates compared to mice treated with monotherapy.14 This dual gene therapy not only countered immune checkpoint blockade-induced adaptive resistance but also reversed the immunosuppressive tumor microenvironment simultaneously. This dual gene delivery system represents a promising avenue for immunotherapy across various tumor types, though further research is warranted 4.1 Lactic Acid in the Tumor Microenvironment One of the key areas of exploration for scientists has been the manipulation of lactic acid within the tumor microenvironment (TME). Lactic acid, a byproduct of altered metabolism, plays a signi�cant role in shaping the TME. Its acidic nature fosters various processes including angiogenesis, metastasis, and drug resistance. A 2020 study highlighted that heightened lactic acid production can detrimentally a�ect anticancer immunity by suppressing immune responses due to the low pH environment it creates.7 Lactic acid impedes the di�erentiation of monocytes into dendritic cells, leading to a reduction in antigen-presenting functions. Additionally, it hampers the anti-tumoral activities of immune cells like natural killer cells and cytotoxic T cells.7 Consequently, the targeting of lactate and lactic acid has emerged as a compelling area of focus within contemporary immunotherapy research. Scientists have been investigating the neutralization of lactic acid as a potential method. By bu�ering the tumor microenvironment using oral bicarbonate and coupling it with anti-PD-1 immunotherapy and adoptive T-cell transfer, there's potential for enhancing survival rates.5 Furthermore, research has shown that V-domain Ig suppressor of T cell activation (VISTA) can inhibit T cells in an acidic pH environment. Consequently, Berkeley Pharma Tech Journal of Medicine | 125 blocking VISTA in conjunction with PD-1 blockade, and the development of pH-speci�c antibodies against VISTA, have demonstrated e�cacy in tumor rejection.5 Another signi�cant focus lies in targeting the lactate-producing enzyme LDH, which shows promising anti-cancer e�ects. However, its impact on immune cells remains largely unexplored, as LDH inhibition can decrease T cell levels5. An alternative approach involves inhibiting lactate transporters. Notably, researchers identi�ed SLC4A4 as the predominant bicarbonate transporter in pancreatic ductal adenocarcinoma (PDAC). Inhibiting SLC4A4 in PDAC cells reduces TME acidi�cation by accumulating bicarbonate in the extracellular space and lowering lactate production. Combining SLC4A4 targeting with immune checkpoint blockades has been found to overcome immunotherapy resistance.6 Targeting lactic acid holds signi�cant promise for advancing immunotherapy in the tumor microenvironment, as it plays a pivotal role in TME acidi�cation and subsequent immune response suppression. 4.2 Targeting Metabolism in the Tumor Microenvironment Focusing on metabolism to enhance the tumor microenvironment for cancer immunotherapy holds signi�cant promise. Cancer cells exhibit heightened metabolic activity, fueling their rapid proliferation. By zeroing in on metabolism, scientists can slow down cancer cell spread and impede their function. The realm of immune metabolism o�ers potential metabolic targets to bolster anti-cancer immunity. The dynamic metabolism of immune cells signi�cantly in�uences their functions as well.25 A primary approach involves targeting amino acid metabolism, where speci�c amino acids such as glutamine, tryptophan, and arginine have emerged as key targets for inhibiting tumor progression and enhancing immunity. Glutamine, in particular, plays a vital role as a nutrient for cancer cells. Enzymes like glutaminase (GLS) facilitate the conversion of glutamine to glutamate, making GLS a prime target to curb cancer cell metabolism and glucose utilization. However, targeting GLS poses challenges as cancer cells employ alternative methods to boost glucose metabolism. Additionally, directing GLS inhibition towards immune cells, particularly T cells, a�ects Berkeley Pharma Tech Journal of Medicine | 126 their di�erentiation, survival, proliferation, and e�ector functions.5 This presents a signi�cant hurdle, as it necessitates reducing glutamate levels in cancer cells while ensuring adequate glutamine for proper T cell function. Consequently, a strategy has been devised to broadly block glutamine metabolism alongside anti-PD-1 immunotherapy. This combined approach enhances anti-tumor e�ects by dampening tumor metabolism while maintaining robust glucose metabolism in T cells.5 Furthermore, researchers have shown keen interest in targeting arginine and nitric oxide, delving into amino acid metabolism. Arginine plays a pivotal role in cancer and immune cell functions, particularly in proliferation, survival, and protein synthesis. The metabolic pathways of arginine heavily rely on enzymes like arginase (ARG) and nitric oxide synthase (NOS). However, given arginine's signi�cance to both cancer and immune cells, its potential as a therapeutic target presents complexities. While arginine depletion proves e�ective in certain scenarios, it can hinder anti-tumor T cell responses and bolster the population of myeloid-derived suppressor cells (MDSCs), thus exacerbating arginine depletion.5 Yet, inhibiting ARG o�ers promise by restoring arginine levels, leading to tumor regression and enhancing T cell functionalities.5 Targeting arginine and nitric oxide in myeloid cells could potentially enhance immunotherapy, but further research is needed to understand its impact on other immune cells. Tryptophan is another signi�cant amino acid being investigated for immunotherapy due to its role in cell growth and maintenance. Adaptive immune cell subsets such as Tregs, tolerogenic dendritic cells (DCs), and myeloid-derived suppressor cells (MDSCs) exhibit high levels of indoleamine 2,3-dioxygenase (IDO), the enzyme that converts tryptophan to kynurenine.5 Kynurenine has immunosuppressive e�ects on T cells, so targeting IDO activity could not only mitigate these immunosuppressive e�ects on T cells but also inhibit these adaptive immune cell subsets.5 Lipid metabolism has emerged as a recent target in immunotherapy. Cancer cells exploit lipid metabolism to support their rapid proliferation, survival, migration, invasion, and metastasis.26 Reprogramming lipid metabolism can Berkeley Pharma Tech Journal of Medicine | 127 prevent e�ector T cell senescence, suggesting that lipid metabolism plays a role in regulating senescence development in T cells.27 Fatty acid metabolism is crucial for the di�erentiation of immune cells, and the upregulation of fatty acid oxidation (FAO) can enhance the functions of CD8+ T cells. Reprogramming fatty acid metabolism in immune cells could promote an inhibitory immune microenvironment.28 It is often used in combination therapies with other immunotherapeutic treatments, such as immune checkpoint blockades (ICBs).28 Recent studies have shown that targeting fatty acid metabolism in T cells can improve the e�cacy of ICBs. For instance, a study found that combining beza�brate with PD-L1 antibody treatment yielded better results in LLC xenograft mouse models. Beza�brate treatments increased the expression of FAO-related genes in cytotoxic T lymphocytes (CTLs), such as PGC-1α, CPT1a, and LCAD, and maintained the survival and function of CTLs.29 Scientists have explored manipulating iron metabolism as a strategy for treating tumors, given iron's crucial role in tumor proliferation. Iron metabolic dysfunction directly impacts cancer pathophysiology, with the availability of iron regulating the aggressive phenotypes of tumors.30 Many tumor microenvironment (TME) cells rely on iron to thrive and function properly. For instance, activated M2 tumor-associated macrophages (TAMs) can disrupt iron homeostasis within tumor cells by exporting high amounts of iron and increasing the production of other iron-related proteins.30 It has been observed that iron chelation, the bonding of ionic molecules to metal ions, can reverse the iron-exporting phenotype of M2 TAMs.30 Blocking transferrin receptor 1 (TFR1), a receptor protein that recognizes transferrin-bound iron, has proven e�ective in suppressing tumor growth, as TFR1 is often upregulated in cancer cells to enhance their iron supply.30 Thus, cutting o� a tumor's iron supply appears to be an e�ective method to reduce tumor growth. Berkeley Pharma Tech Journal of Medicine | 128 Figure 3: The interactions between tryptophan and kynurenine. The enzyme IDO converts tryptophan into kynurenine, which exerts an immunosuppressive effect on T cells. Blocking IDO would not only eliminate the immunosuppressive effect on T cells but also suppress adaptive immune cell subsets, including MDSCs, Tregs, and DCs. 4.3 Targeting Signaling Pathways Di�erent signaling pathways, such as oxygen and nutrient sensing pathways, have signi�cant potential for reducing the glucose metabolism of cancer cells. In low-glucose environments, AMP-activated protein kinase (AMPK) becomes activated, and in high-glucose environments, AMPK is promoted, which can lead to the persistence of immunosuppressive cells.5 High levels of AMPK can inhibit T cells, but disabling AMPK may result in increased glycolysis in cancer cells. Metformin is one drug that targets AMPK by activating its signaling. Studies have shown that metformin reduces cancer risk in both mice models and humans. AMPK activation is believed to reduce tumor burden by slowing tumor growth, supporting the growth of tumor-in�ltrating lymphocytes (TILs) in the tumor microenvironment (TME), and blocking certain glycolytic enzymes necessary for cancer cell Berkeley Pharma Tech Journal of Medicine | 129 proliferation. However, AMPK activation can also increase the number of immunosuppressive cells while promoting tumor regression.5 It is crucial to examine the e�ects on both cancer cells and the phenotypic ratios of immune cells. Another crucial nutrient pathway is the mTOR pathway, which is activated by high levels of glucose and amino acids. In T cells, mTOR supports glycolysis, di�erentiation, and e�ector functions. However, because mTOR enhances memory formation in CD8+ T cells while low mTOR levels boost Treg activity, it is vital to balance mTOR levels to enhance memory formation without promoting Treg activity.5 Like AMPK, the mTOR signaling pathway has a complex nature, necessitating further research to develop nuanced methods that induce bene�cial immune responses without creating a tumor-protecting immune environment.5 Two critical targets of immunotherapy are the immune checkpoint pathways PD-1/PD-L1 and CTLA-4. Anti-CTLA-4 antibodies can deplete Treg cells from the tumor microenvironment (TME), leading to successful tumor rejection in animal studies.31 PD-1/PD-L1 are involved in signaling mediated by antigen recognition through T cell receptors.31 Antibodies that block the PD-1/PD-L1 checkpoint have shown signi�cant therapeutic e�ciency.31 These immune checkpoints mainly function by suppressing the metabolic reprogramming of immune cells, inhibiting glycolysis, and increasing lipolysis. Blocking these pathways promotes anabolic metabolic pathways and glycolysis, thereby restoring the e�ector function of tumor-in�ltrating lymphocytes (TILs). Antibodies against CTLA-4, PD-1, and PD-L1 reverse the glycolysis restrictions on T cells, essentially restoring T-cell glycolysis and interferon production.6 Another signaling pathway that has been extensively studied is the CSF-1/R pathway, which is expressed in myeloid cells. This pathway regulates the in�ltration, phenotypic and functional di�erentiation, and survival of myeloid cells, including tumor-associated macrophages (TAMs). Blocking the CSF-1/R pathway promotes the TME by facilitating the in�ltration and reactivation of cytotoxic T lymphocytes (CTLs).32 Consequently, studying Berkeley Pharma Tech Journal of Medicine | 130 various signaling pathways can pave the way for developing nuanced cancer immunotherapy treatments. As discussed, the TME consists of various components, and understanding these di�erences in TME is crucial for developing e�ective treatments and immunotherapy approaches for di�erent cancers. Identifying and targeting the most relevant aspects of the TME for each speci�c disease is key to creating successful therapies. 5. Non-small Cell Lung Cancer Non-small cell lung cancer (NSCLC) is the leading cause of cancer-related death and has become a primary focus for the development of TME-based immunotherapy. Similar to other cancers, NSCLC features an immunosuppressive tumor microenvironment (TME) and CTL exhaustion. However, it also has distinct characteristics such as variable blood �ow, areas of acidosis and hypoxia, and increased expression of hypoxia markers. These features can lead to metastasis and elevated resistance to treatment.32,33 Consequently, past therapies targeting the TME have focused signi�cantly on addressing hypoxia and angiogenesis.33 A recent study conducted by Zhao et al. adopts a similar strategy focusing on the Tumor Microenvironment (TME). The study delves into a Phase III trial combining bevacizumab, atezolizumab, and chemotherapy for metastatic non-squamous Non-Small Cell Lung Cancer (NSCLC). Bevacizumab, an IgG antibody that obstructs the Vascular Endothelial Growth Factor (VEGF) pathway by binding to VEGF-A, has demonstrated e�cacy in treating NSCLC. The VEGF pathway is a prime target due to its overexpression in NSCLC, correlating with tumor recurrence, low survival rates, metastasis, and mortality. A recent study conducted by Zhao et al. adopts a similar strategy focusing on the Tumor Microenvironment (TME). The study delves into a Phase III trial combining bevacizumab, atezolizumab, and chemotherapy for metastatic non-squamous Non-Small Cell Lung Cancer (NSCLC). Berkeley Pharma Tech Journal of Medicine | 131 Bevacizumab, an IgG antibody that obstructs the Vascular Endothelial Growth Factor (VEGF) pathway by binding to VEGF-A, has demonstrated e�cacy in treating NSCLC.19 The VEGF pathway is a prime target due to its overexpression in NSCLC, correlating with tumor recurrence, low survival rates, metastasis, and mortality.19 Apart from bevacizumab, other antibodies like ramucirumab and nintedanib have demonstrated enhancements in overall survival (OS), along with tyrosine kinase inhibitors (TKIs) capable of inhibiting the Vascular Endothelial Growth Factor (VEGF) pathway.19 However, the VEGF pathway primarily addresses the angiogenic aspect of tumors, prompting its utilization in combination therapies with chemotherapy and immunotherapy, which have proven e�ective. In a similar vein, this Phase III trial combines bevacizumab with atezolizumab, an immune checkpoint inhibitor targeting the PD-L1 ligand (anti-PD-1), alongside chemotherapy. This multifaceted therapeutic approach to Non-Small Cell Lung Cancer (NSCLC) aims to enhance progression-free survival and OS.32 5.1 NSCLC Combination Therapy The primary treatment modality for Non-Small Cell Lung Cancer (NSCLC) typically involves Immune Checkpoint Inhibitors (ICIs) like pembrolizumab, an anti-PD-1 antibody. Pembrolizumab has emerged as the �rst-line treatment for metastatic NSCLC due to its notable response rates and sustained tumor regression.14 However, despite its e�cacy, many patients encounter resistance to these treatments or su�er from immune-related adverse events.14 Consequently, combination therapy has emerged as a pivotal focus for advancing cancer treatments. A recent study conducted in 2022 by Reckamp et al. explored combination therapy involving ICIs and Vascular Endothelial Growth Factor (VEGF) receptor inhibition in NSCLC patients. Patients were administered a combination of either ramucirumab (VEGF inhibitor) and pembrolizumab (ICI), or the investigator’s standard of care, such as docetaxel, gemcitabine, or pemetrexed.34 Among the 136 eligible patients, those who received ramucirumab and pembrolizumab exhibited signi�cantly higher overall Berkeley Pharma Tech Journal of Medicine | 132 survival (OS), suggesting another promising combination therapy avenue for treating NSCLC.34 In September 2022, Casarrubios et al. conducted a study focusing on a speci�c form of combination therapy called neoadjuvant chemoimmunotherapy. This approach utilizes both chemotherapy and immunotherapy to reduce tumor size before more targeted treatment aimed at tumor removal. Through bulk RNA sequencing, the researchers analyzed samples from 41 patients with stage III Non-Small Cell Lung Cancer (NSCLC) who underwent three cycles of nivolumab (an anti-PD-1 drug) and chemotherapy. Their �ndings revealed downregulation of tumor and proliferation markers, along with genes associated with IFNγ signaling.35 Additionally, they observed the regulation of numerous genes such as IFNG and NKG7, distinguishing between complete pathological response (CPR) and non-CPR tumors before and after treatment. This study lays the groundwork for future personalized immunotherapy approaches based on predictive biomarkers of CPR, in conjunction with neoadjuvant immunotherapy.35 5.2 Melanoma Anti-PD-1 therapy represents a standard immunotherapy regimen for melanoma, a type of skin cancer. However, this treatment modality is only e�ective in providing long-term clinical bene�ts to approximately 40% of patients with advanced melanoma.36 To address this challenge, Davar et al. conducted a study involving the administration of pembrolizumab, an anti-PD-1 drug, alongside a fecal microbiota transplant (FMT) in 16 patients with advanced melanoma who did not respond to anti-PD-1 therapy alone.36 This combined approach of FMT and PD-1 blockade resulted in the reprogramming of the Tumor Microenvironment (TME), overcoming resistance to anti-PD-1 therapy. Notably, the treatment led to the upregulation of CD8+ T cells, a decrease in the percentage of naive CD8+ T cells, and an increase in activated and di�erentiated CD8+ T cells. Responders also exhibited downregulation of cytokines and chemokines associated with anti-PD-1 resistance, along with upregulation of biomarkers linked to improved outcomes.36 Berkeley Pharma Tech Journal of Medicine | 133 A recent approach in melanoma treatment involves categorizing the Tumor Microenvironment (TME) to anticipate the cancer's responsiveness to immunotherapy. While combining di�erent Immune Checkpoint Inhibitors (ICIs) like anti-PD-1 and anti-CTLA-4 may appear promising, trials on advanced melanoma patients showed severe immune-related adverse events (irAEs) such as hypophysitis, pneumonitis, and thyroiditis, likely due to the distinct mechanisms of T cell inhibition—CTLA-4 acting on T cell activation, while PD-1 intervenes later.14 A recent study identi�ed 29 functional gene expression signatures (Fges) delineating major functional and cellular components of melanoma tumors, classifying them into four subtypes: immune-enriched/�brotic (elevated angiogenesis and Cancer-Associated Fibroblast activation), immune-enriched/non-�brotic (more immune-active TME), �brotic (low lymphocyte in�ltration, high �broblast activity), and immune-depleted (immune-desert).37 This classi�cation aids in determining treatment e�ectiveness and alternative approaches. For instance, responses to anti-CTLA-4 treatment varied across subtypes, with 82% of immune-enriched subtypes responding compared to only 10% of �brotic subtype, suggesting appropriate utilization of ICI immunotherapy.37 5.3 Other Cancers (Novel Treatments/Combination Therapies) Pancreatic cancer (PDAC) poses a signi�cant challenge for immunotherapy due to its low T cell in�ltration, low tumor mutational burden, and highly suppressive Tumor Microenvironment (TME), making it the fourth leading cause of cancer-related deaths.44 Combination therapy emerges as a promising approach, as demonstrated in a recent study by Padron et al. This study investigated the administration of sotigalimab and/or nivolumab (both anti-PD-1 drugs) alongside chemotherapy.38 The results revealed that combining nivolumab with chemotherapy was associated with a less suppressive TME, increased numbers of activated and circulating T cells, and achieved the primary endpoint of one-year Overall Survival (OS). Although sotigalimab combined with chemotherapy did not meet the OS endpoint, it led to greater in�ltration and di�erentiation of CD4+ cells. Interestingly, administering all three components together did not yield additional improvements.38 Berkeley Pharma Tech Journal of Medicine | 134 Cervical cancer (CC) ranks as the fourth most prevalent cancer, with concurrent chemoradiotherapy (CCRT) serving as the standard treatment.39 However, individuals with advanced-stage disease often experience diminished long-term outcomes, prompting investigation into combination therapies. A recent study by Huang et al. in August 2022 explored the utilization of adoptive cell therapy (ACT), a form of immunotherapy involving the introduction of Tumor-In�ltrating Lymphocytes (TILs) to evoke an immune response, following CCRT and its impact on survival.39 Among the 27 patients receiving ACT alone, 20 saw successful expansion of TILs. Subsequently, 12 patients received TILs post-CCRT, with 75% experiencing complete response and a disease control duration ranging from 9 to 22 months. Treatment-related adverse events were minimal, with only one patient encountering severe toxicity, underscoring the potential of post-CCRT ACT in managing CC.39 E�ective combination therapy entails strategic timing of treatments. Another study focused on determining the optimal timing for immune therapy concerning CCRT revealed the bene�t of administering Immune Checkpoint Inhibitors (ICIs) before CCRT. This strategy capitalizes on maintaining tumor-speci�c immune response, as it was observed to diminish following CCRT. Post-CCRT, reduced numbers of Cytotoxic T Lymphocytes (CTLs), decreased T Cell Receptor (TCR) diversity, and increased Regulatory T cells (Tregs) signi�ed a weakened antitumor immune response.40 Researchers are investigating new treatment avenues for chronic lymphocytic leukemia (CLL) that focus on targeting LAG-3, a protein involved in inhibiting the function of natural killer (NK) cells and CD8+ T cells. In CLL, LAG-3 expression on leukemic cells is associated with poorer outcomes and decreased response to treatment.20 Relatlimab, an IgG4 antibody that blocks LAG-3, has shown promise in preclinical studies. Administering relatlimab increased the proliferation of NK cells and CD8+ T cells without a�ecting the growth of leukemic cells. This suggests that LAG-3 blockade can reverse the inhibition of NK and T cells, potentially restoring their anti-leukemic activity. 20 Berkeley Pharma Tech Journal of Medicine | 135 Moreover, combining relatlimab with lenalidomide, an angiogenesis inhibitor, led to increased T cell numbers and enhanced anti-leukemic activity. This combination therapy approach may o�er a synergistic e�ect in treating CLL by targeting both LAG-3-mediated immune suppression and angiogenesis. Overall, these �ndings provide a promising direction for the development of novel immunotherapies for CLL, particularly for patients who have not responded well to PD-1 or CTLA-4 blockade. 20 Cancer often spreads to the liver, complicating treatment with immunotherapy. Liver metastases attract activated CD8+ T cells, which then undergo apoptosis, resulting in a signi�cant reduction in e�ective T cells. 41 A study comparing patients with metastatic melanoma and NSCLC who received immunotherapy and targeted therapy, or immunotherapy and chemotherapy respectively, revealed that liver metastases were linked to reduced response to immunotherapy but not targeted therapy. 41 Similarly, in NSCLC cases, immunotherapy had limited e�cacy compared to chemotherapy, while radiotherapy showed promising results. These �ndings suggest that liver metastasis may be more e�ectively managed through non- immunotherapeutic methods, informing future treatment strategies.41 6. Practical Applications/Discussion Numerous innovative combination therapies extend beyond traditional chemotherapy and immunotherapy, delving into the modulation of the tumor microenvironment (TME) to bolster immune responses. For instance, one approach involves delivering a fecal microbiota transplant (FMT) via colonoscopy alongside immunotherapeutic anti-PD-1 drugs.36 This strategy e�ectively reshapes the TME, overcoming primary resistance to anti-PD-1 in advanced melanoma patients. Following treatment, there's an increase in the proportion of CD56+CD8+T cells, indicating heightened circulation of active and di�erentiated CD8+ T cells.36 Responders exhibit decreased levels of certain cytokines and chemokines linked to anti-PD-1 resistance, while biomarkers associated with improved survival are upregulated.36 Notably, variations in patient response may occur due to factors like tumor immunogenicity or unsuccessful colonization by Berkeley Pharma Tech Journal of Medicine | 136 FMT, necessitating further investigation for its establishment as a standard treatment.36 Another method, as explored by Ribas et al., involves combining anti-PD-1 therapy (speci�cally pembrolizumab) with intratumoral administration of an oncolytic virus, such as talimogene laherparepvec, in advanced melanoma patients. Initially, tumor size may transiently increase post-injection; however, this approach boosts CD8+ T cell in�ltration and peripheral CD4+ T cell presence.48 Pembrolizumab promotes the proliferation of CD8+ T cells, leading to a comprehensive shift in the TME and an augmented immune response compared to anti-PD-1 therapy alone. Importantly, T cells expressing PD-1 encounter tumor cells expressing PD-L1, a dynamic that, alongside PD-1 blockade, moderates the antitumor e�cacy of the administered virus.48 As previously discussed, extensive research has already been conducted on PD-1, PD-L1, CTLA-4, and VEGF/VEGF-A as targets for treating various cancers. Scientists are now focusing on identifying new potential biomarkers to better predict patient responses to speci�c treatments, given that the existing biomarkers are not perfect indicators of a patient’s response to immune checkpoint inhibitor (ICI) therapy.49 Vanhersecke et al. observed that the tertiary lymphoid structure (TLS) within tumors is associated with the population of CD8+ T cells. Higher TLS densities correspond to increased densities of CD8+ T cells. Moreover, the presence of more mature TLS was linked to improved survival rates, higher objective response rates, and longer progression-free survival. Notably, mature TLS emerged as the most signi�cant predictive factor for an objective response, independent of PD-L1 expression and CD8+ T-cell in�ltration. Patients with tumors containing mature TLS had better outcomes compared to those without mature TLS.49 Studying the TLS in tumors opens a new avenue for identifying biomarkers to predict patient responses to speci�c ICI treatments. Two ongoing clinical trials are exploring recent advances in immunotherapy and tumor microenvironment (TME) mediated treatments, focusing on combination therapies involving LAG-3, anti-PD-1, and VEGF signaling. Berkeley Pharma Tech Journal of Medicine | 137 The �rst trial is investigating the e�ects of anti-LAG-3 (urelumab) alone and in combination with nivolumab (an anti-PD-1 drug) in patients with recurrent glioblastoma. The study aims to determine which treatment regimen is more e�ective at killing tumor cells. Currently, 63 patients are enrolled, and the trial is expected to conclude by November 2023.50 The second trial, entering phase II, is evaluating the e�cacy and safety of a combination of anti-PD-L1/VEGF drugs with traditional chemotherapy in treating non-small cell lung cancer (NSCLC). The primary objectives are to measure the objective response rate to the treatment and progression-free survival, which involves monitoring the time from the start of treatment until the cancer progresses or the patient dies from any cause.51 Other monitored outcomes include overall survival rate, disease control, response duration, treatment-related adverse events, and the correlation between PD-L1 expression and antitumor e�ect. The clinical trial aims to enroll 374 patients and is expected to be completed by December 2025.51 7. Conclusion The tumor microenvironment (TME) is a promising target in immunotherapy. Due to its variability and adaptability, further research is needed to e�ectively target di�erent TME components. Current techniques include immune checkpoint blockades like PD-1 and CTLA-4, and targeting lactic acid and metabolism. Future prospects lie in combination therapies with chemotherapy or neoadjuvant therapy, microbiota fecal transplants to enhance TME for therapy, classifying CPR versus non-CPR tumors, and optimizing the timing of immunotherapy in combination treatments.36 Berkeley Pharma Tech Journal of Medicine | 138 References 1. Jung JG, Le A. Targeting metabolic cross talk between cancer cells and cancer-associated �broblasts. SpringerLink. January 1, 1970. https://link.springer.com/chapter/10.1007/97 8-3-030-65768-0_15. 2. Sadeghi Rad H, Monkman J, Warkiani ME, et al. Understanding the tumor microenvironment for e�ective immunotherapy. Medicinal Research Reviews . 2020;41(3):1474-1498. doi:10.1002/med.21765 3. Dobosz P, Dzieciątkowski T. The intriguing history of cancer immunotherapy. Frontiers. December 3, 2019. https://www.frontiersin.org/articles/10.3389/f immu.2019.02965/full. 4. Yaqi Li, Jing Liu, Long Gao, Yuan Liu, Fang Meng, Xiaoan Li, F. Xiao-Feng Qin, Targeting the tumor microenvironment to overcome immune checkpoint blockade therapy resistance, Immunology Letters, Volume 220, 2020, Pages 88-96, ISSN 0165-2478, https://doi.org/10.1016/j.imlet.2019.03.006. 5. Bader JE, Voss K, Rathmell JC. Targeting metabolism to improve the tumor microenvironment for cancer immunotherapy. Mol Cell . 2020;78(6):1019-1033. doi:10.1016/j.molcel.2020.05.034 6. Cappellesso F, Orban MP, Shirgaonkar N, et al. Targeting the bicarbonate transporter SLC4A4 overcomes immunosuppression and immunotherapy resistance in pancreatic cancer. Nat Cancer . 2022;3(12):1464-1483. doi:10.1038/s43018-022-00470-2 7. Wang JX, Choi SYC, Niu X, et al. Lactic acid and an acidic tumor microenvironment suppress anticancer immunity. Int J Mol Sci . 2020;21(21):8363. doi:10.3390/ijms21218363 8. Anderson NM, Simon MC. The tumor microenvironment. CB. https://pubmed.ncbi.nlm.nih.gov/32810447/. 9. Bindea G, Hahn WC, Rooney MS, et al. Immune cellular components and signaling pathways in the tumor microenvironment. Seminars in Cancer Biology. August 18, 2022. https://www.sciencedirect.com/science/article/abs/ pii/S1044579X22001845?via% 3Dihu. 10. Kazama K, Otake J, Satoyoshi T, et al. Distribution of regulatory T-cells and other phenotypes of T-cells in tumors and regional lymph nodes of colorectal cancer patients. In Vivo. March 1, 2020. https://iv.iiarjournals.org/content/34/2/849. 11. Hien TT, E�erth T, Kawasaki J, et al. Inhibition of human cancer cell line growth and human umbilical vein endothelial cell angiogenesis by artemisinin derivatives in vitro. Pharmacological Research. June 4, 2003. https://www.sciencedirect.com/science/article/abs/ pii/S1043661803001075?via% 3Dihub. 12. Felix Peix, Oriol Casanovas, Promalignant e�ects of antiangiogenics in the tumor microenvironment, Seminars in Cancer Biology, Volume 86, Part 3, 2022, Pages 199-206, ISSN 1044-579X, https://doi.org/10.1016/j.semcancer.2022.03.003 13. Li Y, Liu J, Gao L, et al. Targeting the tumor microenvironment to overcome immune checkpoint blockade therapy resistance. Immunology Letters . 2020;220:88-96. doi:10.1016/j.imlet.2019.03.006 Berkeley Pharma Tech Journal of Medicine | 139 14. Bagchi S, Yuan R, Engleman EG. Immune Checkpoint Inhibitors for the Treatment of Cancer: Clinical Impact and Mechanisms of Response and Resistance. Annu Rev Pathol . 2021;16:223-249. doi:10.1146/annurev-pathol-042020-042741 15. Vesely MD, Zhang T, Chen L. Resistance mechanisms to Anti-PD Cancer immunotherapy. Annual Review of Immunology . 2022;40(1):45-74. doi:10.1146/annurev-immunol-070621-030155 16. Feng Y, Wu J, Chen J, et al. Targeting dual gene delivery nanoparticles overcomes immune checkpoint blockade induced adaptive resistance and regulates tumor microenvironment for improved tumor immunotherapy. Nano Today . 2021;38:101194. doi:10.1016/j.nantod.2021.101194 17. Qian J, Wang C, Wang B, et al. The IFN-γ/PD-L1 axis between T cells and tumor microenvironment: hints for glioma anti-PD-1/PD-L1 therapy. J Neuroinflammation . 2018;15:290. doi:10.1186/s12974-018-1330-2 18. Coutzac C, Bibeau F, Ben Abdelghani M, et al. Immunotherapy in MSI/dMMR tumors in the perioperative setting: The IMHOTEP trial. Dig Liver Dis . 2022;54(10):1335-1341. doi:10.1016/j.dld.2022.07.008 19. Zhao Y, Guo S, Deng J, et al. VEGF/VEGFR-targeted therapy and immunotherapy in non-small cell lung cancer: Targeting the tumor microenvironment. International Journal of Biological Sciences . 2022;18(9):3845-3858. doi:10.7150/ijbs.70958 20. Sordo-Bahamonde C, Lorenzo-Herrero S, González-Rodríguez AP, et al. LAG-3 Blockade with Relatlimab (BMS-986016) Restores Anti-Leukemic Responses in Chronic Lymphocytic Leukemia. PubMed Central (PMC). doi:10.3390/cancers13092112 21. Zundell JA, Fukumoto T, Lin J, et al. Targeting the IRE1α/XBP1 endoplasmic reticulum stress response pathway in ARID1A-mutant ovarian cancers. PubMed Central (PMC). doi:10.1158/0008-5472.CAN-21-1545 22. Ma X, Bi E, Lu Y, et al. Cholesterol induces CD8+ T cell exhaustion in the tumor microenvironment. Cell Metabolism . 2019;30(1). doi:10.1016/j.cmet.2019.04.002 23. Chiu D, Qi J, Thin TH, et al. A Phase I Trial of VEGF-A Inhibition Combined with PD-L1 Blockade for Recurrent Glioblastoma. Cancer Res Commun . 2023;3(1):130-139. Published 2023 Jan 25. doi:10.1158/2767-9764.CRC-22-0420 24. Sanmamed MF, Nie X, Desai SS, et al. A burned-out CD8+ T-cell subset expands in the tumor microenvironment and curbs cancer immunotherapy. Cancer Discovery . 2021;11(7):1700-1715. doi:10.1158/2159-8290.cd-20-0962 25. Khodaei T, Inamdar S, Suresh AP, Acharya AP. Drug delivery for metabolism targeted cancer immunotherapy. Adv Drug Deliv Rev . 2022;184(114242):114242. doi:10.1016/j.addr.2022.114242 26. Bian X, Liu R, Meng Y, Xing D, Xu D, Lu Z. Lipid metabolism and cancer. J Exp Med . 2021;218(1). doi:10.1084/jem.20201606 27. Liu X, Hartman CL, Li L, et al. Reprogramming lipid metabolism prevents e�ector T cell senescence and enhances tumor immunotherapy. Sci Transl Med . Berkeley Pharma Tech Journal of Medicine | 140 2021;13(587):eaaz6314. doi:10.1126/scitranslmed.aaz6314 28. Luo Y, Wang H, Liu B, Wei J. Fatty acid metabolism and cancer immunotherapy. Curr Oncol Rep . 2022;24(5):659-670. doi:10.1007/s11912-022-01223-1 29. Wu L, Zhang X, Zheng L, Zhao H, Yan G, Zhang Q, Zhou Y, Lei J, Zhang J, Wang J, Xin R, Jiang L, Peng J, Chen Q, Lam SM, Shui G, Miao H, Li Y. RIPK3 Orchestrates Fatty Acid Metabolism in Tumor-Associated Macrophages and Hepatocarcinogenesis. Cancer Immunol Res. 2020 May 1;8(5):710-721. doi: 10.1158/2326-6066.CIR-19-0261. . 30. Vela D. Iron in the tumor microenvironment. Advances in Experimental Medicine and Biology . Published online June 24, 2020:39-51. doi:10.1007/978-3-030-43093-1_3 31. Pitt JM, Marabelle A, Eggermont A, Soria JC, Kroemer G, Zitvogel L. Targeting the tumor microenvironment: removing obstruction to anticancer immune responses and immunotherapy. Ann Oncol . 2016;27(8):1482-1492. doi:10.1093/annonc/mdw168 32. Datta M, Coussens LM, Nishikawa H, Hodi FS, Jain RK. Reprogramming the Tumor Microenvironment to Improve Immunotherapy: Emerging Strategies andCombination Therapies. PubMed Central (PMC). doi:10.1200/EDBK_237987 33. Graves EE, Maity A, Le QT. The Tumor Microenvironment in Non-Small Cell Lung Cancer. PubMed Central (PMC). doi:10.1016/j.semradonc.2010.01.003 34. Reckamp KL, Redman MW, Dragnev KH, et al. Phase II Randomized Study of Ramucirumab and Pembrolizumab Versus Standard of Care in Advanced Non-Small-Cell Lung Cancer Previously Treated With Immunotherapy-Lung-MAP S1800A [published correction appears in J Clin Oncol. 2022 Sep 1;40(25):3002]. J Clin Oncol. 2022;40(21):2295-2306. doi:10.1200/JCO.22.00912 35. Casarrubios M, Provencio M, Nadal E, et al. Original research: Tumor microenvironment gene expression pro�les associated to complete pathological response and disease progression in resectable NSCLC patients treated with neoadjuvant chemoimmunotherapy. PubMed Central (PMC). doi:10.1136/jitc-2022-005320 36. Davar D, Dzutsev AK, McCulloch JA, et al. Fecal microbiota transplant overcomes resistance to anti–PD-1 therapy in melanoma patients. PubMed Central (PMC). doi:10.1126/science.abf3363 37. Bagaev A, Kotlov N, Nomie K, Svekolkin V, Gafurov A, Isaeva O, Osokin N, Kozlov I, Frenkel F, Gancharova O, Almog N, Tsiper M, Ataullakhanov R, Fowler N. Conserved pan-cancer microenvironment subtypes predict response to immunotherapy. Cancer Cell . 2021;39(6):845-865.e7. doi:10.1016/j.ccell.2021.04.014. 38. Padrón LJ, Maurer DM, O’Hara MH, et al. Sotigalimab and/or nivolumab with chemotherapy in �rst-line metastatic pancreatic cancer: clinical and immunologic analyses from the randomized phase 2 PRINCE trial. PubMed Central (PMC). doi:10.1038/s41591-022-01829-9 39. Huang H, Nie C ping, Liu X feng, et al. Phase I study of adjuvant immunotherapy with autologous tumor-in�ltrating lymphocytes in locally advanced cervical cancer. PubMed Central (PMC). Berkeley Pharma Tech Journal of Medicine | 141 doi:10.1172/JCI157726 40. Li R, Liu Y, Yin R, et al. The Dynamic Alternation of Local and Systemic Tumor Immune Microenvironment During Concurrent Chemoradiotherapy of Cervical Cancer: A Prospective Clinical Trial. Int J Radiat Oncol Biol Phys. 2021;110(5):1432-1441. doi:10.1016/j.ijrobp.2021.03.003 41. Yu J, Green MD, Li S, et al. Liver metastasis restrains immunotherapy e�cacy via macrophage-mediated T cell elimination. PubMed Central (PMC). doi:10.1038/s41591-020-1131-x 42. Graves EE, Maity A, Le QT. The Tumor Microenvironment in Non-Small Cell Lung Cancer. PubMed Central (PMC). doi:10.1016/j.semradonc.2010.01.003 43. Dunn J, McCuaig RD, Y. Tan AH, et al. Selective Targeting of Protein Kinase C (PKC)-θ Nuclear Translocation Reduces Mesenchymal Gene Signatures and Reinvigorates Dysfunctional CD8+ T Cells in Immunotherapy-Resistant and Metastatic Cancers. PubMed Central (PMC). doi:10.3390/cancers14061596 44. Maitra A, Hruban RH. Pancreatic Cancer. PubMed Central (PMC). doi:10.1146/annurev.pathmechdis.3.121806.1 54305 45. Piersma SJ. Immunosuppressive Tumor Microenvironment in Cervical Cancer Patients. PubMed Central (PMC). doi:10.1007/s12307-011-0066-7 46. Ring A, Zenz T. Genetics of “high-risk” chronic lymphocytic leukemia in the times of chemoimmunotherapy. PubMed Central (PMC). doi:10.3324/haematol.2020.246504 47. Hong M, Clubb JD, Chen YY. Engineering CAR-T Cells for Next-Generation Cancer Therapy. Cancer Cell. 2020;38(4):473-488. doi:10.1016/j.ccell.2020.07.005 48. Ribas A, Dummer R, Puzanov I, et al. Oncolytic Virotherapy Promotes Intratumoral T Cell In�ltration and Improves Anti-PD-1 Immunotherapy. PubMed Central (PMC). doi:10.1016/j.cell.2017.08.027 49. Vanhersecke L, Brunet M, Guégan JP, et al. Mature tertiary lymphoid structures predict immune checkpoint inhibitor e�cacy in solid tumors independently of PD-L1 expression. Nat Cancer. 2021;2(8):794-802. doi:10.1038/s43018-021-00232-6 50. Sidney Kimmel Comprehensive Cancer Center at Johns Hopkins. Anti-LAG-3 Alone & in Combination with Nivolumab Treating Patients with Recurrent GBM (Anti-CD137 Arm Closed 10/16/18). ClinicalTrials.gov Identi�er: NCT02658981. Last updated October 6, 2023. Available from: https://clinicaltrials.gov/ct2/show/NCT02658981 51. Biotheus Inc. A Study of PM8002 (Anti-PD-L1/VEGF) in Combination With Chemotherapy in Patients With NSCLC. ClinicalTrials.gov Identi�er: NCT05756972. Last updated July 13, 2023. Accessed July 28, 2023. Available from: https://clinicaltrials.gov/study/NCT05756972 Berkeley Pharma Tech Journal of Medicine | 142 Kannabran et al. _ need page numbers.pdf Kannabran et al. _ need page numbers.pdf 64-Scientific Review Paper-597-1-11-20240625.pdf Kannabran Cover Page Kannabran body.docx (1) Kannabran et al._References - final.pdf