Failure of Immunotherapy—The Molecular and Immunological Origin of Immunotherapy Resistance in Lung Cancer

Immune checkpoint inhibitors (ICIs) have a huge impact on clinical treatment results in non-small cell lung cancer (NSCLC). Blocking antibodies targeting programmed cell death protein 1 (PD-1), programmed cell death protein ligand 1 (PD-L1) or CTLA-4 (cytotoxic T cell antigen 4) have been developed and approved for the treatment of NSCLC patients. However, a large number of patients develop resistance to this type of treatment. Primary and secondary immunotherapy resistance are distinguished. No solid biomarkers are available that are appropriate to predict the unique sensitivity to immunotherapy. Knowledge of predictive markers involved in treatment resistance is fundamental for planning of new treatment combinations. Scientists focused research on the use of immunotherapy as an essential treatment in combination with other therapy strategies, which could increase cancer immunogenicity by generating tumor cells death and new antigen release as well as by targeting other immune checkpoints and tumor microenvironment. In the present review, we summarize the current knowledge of molecular bases underlying immunotherapy resistance and discuss the capabilities and the reason of different therapeutic combinations.     

Current Immunotherapeutic Strategies Targeting the PD-1/PD-L1 Axis in Non-Small Cell Lung Cancer with Oncogenic Driver Mutations

Treatment strategies targeting programed cell death 1 (PD-1) or its ligand, PD-L1, have been developed as immunotherapy against tumor progression for various cancer types including non-small cell lung cancer (NSCLC). The recent pivotal clinical trials of immune-checkpoint inhibiters (ICIs) combined with cytotoxic chemotherapy have reshaped therapeutic strategies and established various first-line standard treatments. The therapeutic effects of ICIs in these clinical trials were analyzed according to PD-L1 tumor proportion scores or tumor mutational burden; however, these indicators are insufficient to predict the clinical outcome. Consequently, molecular biological approaches, including multi-omics analyses, have addressed other mechanisms of cancer immune escape and have revealed an association of NSCLC containing specific driver mutations with distinct immune phenotypes. NSCLC has been characterized by driver mutation-defined molecular subsets and the effect of driver mutations on the regulatory mechanism of PD-L1 expression on the tumor itself. In this review, we summarize the results of recent clinical trials of ICIs in advanced NSCLC and the association between driver alterations and distinct immune phenotypes. We further discuss the current clinical issues with a future perspective for the role of precision medicine in NSCLC.     

Multi-Omics Immune Interaction Networks in Lung Cancer Tumorigenesis,Proliferation, and Survival

There are currently no effective biomarkers for prognosis and optimal treatment selection to improve non-small cell lung cancer (NSCLC) survival outcomes. This study further validated a seven-gene panel for diagnosis and prognosis of NSCLC using RNA sequencing and proteomic profiles of patient tumors. Within the seven-gene panel, ZNF71 expression combined with dendritic cell activities defined NSCLC patient subgroups (n = 966) with distinct survival outcomes (p = 0.04, Kaplan–Meier analysis). ZNF71 expression was significantly associated with the activities of natural killer cells (p = 0.014) and natural killer T cells (p = 0.003) in NSCLC patient tumors (n = 1016) using Chi-squared tests. Overexpression of ZNF71 resulted in decreased expression of multiple components of the intracellular intrinsic and innate immune systems, including dsRNA and dsDNA sensors. Multi-omics networks of ZNF71 and the intracellular intrinsic and innate immune systems were computed as relevant to NSCLC tumorigenesis, proliferation, and survival using patient clinical information and in-vitro CRISPR-Cas9/RNAi screening data. From these networks, pan-sensitive and pan-resistant genes to 21 NCCN-recommended drugs for treating NSCLC were selected. Based on the gene associations with patient survival and in-vitro CRISPR-Cas9, RNAi, and drug screening data, MEK1/2 inhibitors PD-198306 and U-0126, VEGFR inhibitor ZM-306416, and IGF-1R inhibitor PQ-401 were discovered as potential targeted therapy that may also induce an immune response for treating NSCLC.     

Circular RNA hsa_circ_0000190 Facilitates the Tumorigenesis and Immune Evasion by Upregulating the Expression of Soluble PD-L1 in Non-Small-Cell Lung Cancer

Lung cancer is the leading cause of death from cancer in Taiwan and throughout the world. Immunotherapy has revealed promising and significant efficacy in NSCLC, through immune checkpoint inhibition by blocking programmed cell death protein (PD)-1/PD-1 ligand (PD-L1) signaling pathway to restore patients’ T-cell immunity. One novel type of long, non-coding RNAs, circular RNAs (circRNAs), are endogenous, stable, and widely expressed in tissues, saliva, blood, urine, and exosomes. Our previous results revealed that the plasma level of hsa_circ_0000190 can be monitored by liquid-biopsy-based droplet digital PCR and may serve as a valuable blood-based biomarker to monitor the disease progression and the efficacy of immunotherapy. In this study, hsa_circ_0000190 was shown to increase the PD-L1 mRNA-mediated soluble PD-L1 (sPD-L1) expression, consequently interfering with the efficacy of anti-PD-L1 antibody and T-cell activation, which may result in immunotherapy resistance and poor outcome. Our results unraveled that hsa_circ_0000190 facilitated the tumorigenesis and immune evasion of NSCLC by upregulating sPD-L1 expression, potentially developing a different aspect in elucidating the molecular immunopathogenesis of NSCLC. Hsa_circ_0000190 upregulation can be an effective indicator for the progression of NSCLC, and hsa_circ_0000190 downregulation may possess a potential therapeutic value for the treatment of NSCLC in combination with immunotherapy.     

Ferroptosis in Non-Small Cell Lung Cancer: Progression and Therapeutic Potential on It

As a main subtype of lung cancer, the current situation of non-small cell lung cancer (NSCLC) remains severe worldwide with a 19% survival rate at 5 years. As the conventional therapy approaches, such as chemotherapy, radiotherapy, targeted therapy, and immunotherapy, gradually develop into therapy resistance, searching for a novel therapeutic strategy for NSCLC is urgent. Ferroptosis, an iron-dependent programmed necrosis, has now been widely considered as a key factor affecting the tumorigenesis and progression in various cancers. Focusing on its effect in NSCLC, in different situations, ferroptosis can be triggered or restrained. When ferroptosis was induced in NSCLC, it was available to inhibit the tumor progression both in vitro and in vivo. The dominating mechanism was due to a regulation of the classic ferroptosis-repressed GSH-dependent GPX4 signaling pathway instead of other fractional regulating signal axes that regulated ferroptosis via impacting on the ROS, cellular iron levels, etc. In terms of the prevention of ferroptosis in NSCLC, an GSH-independent mechanism was also discovered, interestingly exhibiting the same upstream as the GPX4 signaling. In addition, this review summarizes the progression of ferroptosis in NSCLC and elaborates their association and specific mechanisms through bioinformatics analysis with multiple experimental evidence from different cascades. Finally, this review also points out the possibility of ferroptosis working as a novel strategy for therapy resistance in NSCLC, emphasizing its therapeutic potential.     

Understanding LAG-3 Signaling

Lymphocyte activation gene 3 (LAG-3) is a cell surface inhibitory receptor with multiple biological activities over T cell activation and effector functions. LAG-3 plays a regulatory role in immunity and emerged some time ago as an inhibitory immune checkpoint molecule comparable to PD-1 and CTLA-4 and a potential target for enhancing anti-cancer immune responses. LAG-3 is the third inhibitory receptor to be exploited in human anti-cancer immunotherapies, and it is considered a potential next-generation cancer immunotherapy target in human therapy, right next to PD-1 and CTLA-4. Unlike PD-1 and CTLA-4, the exact mechanisms of action of LAG-3 and its relationship with other immune checkpoint molecules remain poorly understood. This is partly caused by the presence of non-conventional signaling motifs in its intracellular domain that are different from other conventional immunoregulatory signaling motifs but with similar inhibitory activities. Here we summarize the current understanding of LAG-3 signaling and its role in LAG-3 functions, from its mechanisms of action to clinical applications.     

Immunotherapy in NSCLC Patients with Brain Metastases

Approximately 40% of unselected non-small cell lung cancer (NSCLC) patients develop brain metastases (BMs) during their disease, with considerable morbidity and mortality. The management of BMs in patients with NSCLC is a clinical challenge and requires a multidisciplinary approach to gain effective intracranial disease control. Over the last decade, immune checkpoint inhibitors (ICIs) have emerged as a game-changer in the treatment landscape of advanced NSCLC, with significant improvements in survival outcomes, although patients with BMs are mostly underrepresented in randomized clinical trials. Moreover, the safety and activity of ICIs and radiotherapy combinations compared with single-agent or sequential modalities is still under evaluation to establish the optimal management of these patients. The aim of this review is to summarize the state-of-the-art of clinical evidence of ICIs intracranial activity and the main challenges of incorporating these agents in the treatment armamentarium of NSCLC patients with BMs.     

Identification of Predictive Biomarkers of Response to HSP90 Inhibitors in Lung Adenocarcinoma

Heat shock protein 90 (HSP90) plays an essential role in lung adenocarcinoma, acting as a key chaperone involved in the correct functioning of numerous highly relevant protein drivers of this disease. To this end, HSP90 inhibitors have emerged as promising therapeutic strategies, even though responses to them have been limited to date. Given the need to maximize treatment efficacy, the objective of this study was to use isobaric tags for relative and absolute quantitation (iTRAQ)-based proteomic techniques to identify proteins in human lung adenocarcinoma cell lines whose basal abundances were correlated with response to HSP90 inhibitors (geldanamycin and radicicol derivatives). From the protein profiles identified according to response, the relationship between lactate dehydrogenase B (LDHB) and DNA topoisomerase 1 (TOP1) with respect to sensitivity and resistance, respectively, to geldanamycin derivatives is noteworthy. Likewise, rhotekin (RTKN) and decaprenyl diphosphate synthase subunit 2 (PDSS2) were correlated with sensitivity and resistance to radicicol derivatives. We also identified a relationship between resistance to HSP90 inhibition and the p53 pathway by glucose deprivation. In contrast, arginine biosynthesis was correlated with sensitivity to HSP90 inhibitors. Further study of these outcomes could enable the development of strategies to improve the clinical efficacy of HSP90 inhibition in patients with lung adenocarcinoma.     

Research Progress of Heavy Ion Radiotherapy for Non-Small-Cell Lung Cancer

Non-small-cell lung cancer (NSCLC) has a high incidence and poses a serious threat to human health. However, the treatment outcomes of concurrent chemoradiotherapy for non-small-cell lung cancer are still unsatisfactory, especially for high grade lesions. As a new cancer treatment, heavy ion radiotherapy has shown promising efficacy and safety in the treatment of non-small-cell lung cancer. This article discusses the clinical progress of heavy ion radiotherapy in the treatment of non-small-cell lung cancer mainly from the different cancer stages, the different doses of heavy ion beams, and the patient’s individual factors, and explores the deficiency of heavy ion radiotherapy in the treatment of non-small-cell lung cancer and the directions of future research, in order to provide reference for the wider and better application of heavy ion radiotherapy in the future.     

9-O-Terpenyl-Substituted Berberrubine Derivatives Suppress Tumor Migration and Increase Anti-Human Non-Small-Cell Lung Cancer Activity

Lung cancer is one of the most common cancers and the leading cause of death in humans worldwide. Non-small-cell lung cancer (NSCLC) accounts for approximately 85% of lung cancer cases and is often diagnosed at a late stage. Among patients with NSCLC, 50% die within 1 year after diagnosis. Even with clinical intervention, the 5-year survival rate is only approximately 20%. Therefore, the development of an advanced therapeutic strategy or novel agent is urgently required for treating NSCLC. Berberine exerts therapeutic activity toward NSCLC; therefore, its activity as an antitumor agent needs to be explored further. In this study, three terpenylated-bromide derivatives of berberrubine were synthesized and their anti-NSCLC activities were evaluated. Each derivative had higher anti-NSCLCs activity than berberrubine and berberine. Among them, 9-O-gernylberberrubine bromide (B4) and 9-O-farnesylberberrubine bromide (B5) showed greater growth inhibition, cell-cycle regulation, in vitro tumorigenesis suppression, and tumor migration reduction. In addition, some degree of apoptosis and autophagic flux blocking was noted in the cells under B4 and B5 treatments. Our study demonstrates that the berberrubine derivatives, B4 and B5, exhibit impressive anti-NSCLC activities and have potential for use as chemotherapeutic agents against NSCLC.     

Immunotherapy as a Therapeutic Strategy for Gastrointestinal Cancer—Current Treatment Options and Future Perspectives

Gastrointestinal (GI) cancer constitutes a highly lethal entity among malignancies in the last decades and is still a major challenge for cancer therapeutic options. Despite the current combinational treatment strategies, including chemotherapy, surgery, radiotherapy, and targeted therapies, the survival rates remain notably low for patients with advanced disease. A better knowledge of the molecular mechanisms that influence tumor progression and the development of optimal therapeutic strategies for GI malignancies are urgently needed. Currently, the development and the assessment of the efficacy of immunotherapeutic agents in GI cancer are in the spotlight of several clinical trials. Thus, several new modalities and combinational treatments with other anti-neoplastic agents have been identified and evaluated for their efficiency in cancer management, including immune checkpoint inhibitors, adoptive cell transfer, chimeric antigen receptor (CAR)-T cell therapy, cancer vaccines, and/or combinations thereof. Understanding the interrelation among the tumor microenvironment, cancer progression, and immune resistance is pivotal for the optimal therapeutic management of all gastrointestinal solid tumors. This review will shed light on the recent advances and future directions of immunotherapy for malignant tumors of the GI system.     

MiR-122 Induces Radiosensitization in Non-Small Cell Lung Cancer Cell Line

MiR-122 is a novel tumor suppresser and its expression induces cell cycle arrest, or apoptosis, and inhibits cell proliferation in multiple cancer cells, including non-small cell lung cancer (NSCLC) cells. Radioresistance of cancer cell leads to the major drawback of radiotherapy for NSCLC and the induction of radiosensitization could be a useful strategy to fix this problem. The present work investigates the function of miR-122 in inducing radiosensitization in A549 cell, a type of NSCLC cells. MiR-122 induces the radiosensitization of A549 cells. MiR-122 also boosts the inhibitory activity of ionizing radiation (IR) on cancer cell anchor-independent growth and invasion. Moreover, miR-122 reduced the expression of its targeted genes related to tumor-survival or cellular stress response. These results indicate that miR-122 would be a novel strategy for NSCLC radiation-therapy.     

JAK2-Mediated Phosphorylation of Stress-Induced Phosphoprotein-1 (STIP1) in Human Cells

Stress-induced phosphoprotein-1 (STIP1)—a heat shock protein (HSP)70/HSP90 adaptor protein—is commonly overexpressed in malignant cells, where it controls proliferation via multiple signaling pathways, including JAK2/STAT3. We have previously shown that STIP1 stabilizes the protein tyrosine kinase JAK2 in cancer cells via HSP90 binding. In this study, we demonstrate that STIP1 may act as a substrate for JAK2 and that phosphorylation of tyrosine residues 134 and 152 promoted STIP1 protein stability, induced its nuclear-cytoplasmic shuttling, and promoted its secretion into the extracellular space. We also found that JAK2-mediated STIP1 phosphorylation enhanced cell viability and increased resistance to cisplatin-induced cell death. Conversely, interference STIP1 with JAK2 interaction—attained either through site-directed mutagenesis or the use of cell-penetrating peptides—decreased JAK2 protein levels, ultimately leading to cell death. On analyzing human ovarian cancer specimens, JAK2 and STIP1 expression levels were found to be positively correlated with each other. Collectively, these results indicate that JAK2-mediated phosphorylation of STIP-1 is critical for sustaining the JAK2/STAT3 signaling pathway in cancer cells.     

L-Type Amino Acid Transporter 1 Regulates Cancer Stemness and the Expression of Programmed Cell Death 1 Ligand 1 in Lung Cancer Cells

The l-type amino acid transporter 1 (LAT1) is a membranous transporter that transports neutral amino acids for cells and is dysregulated in various types of cancer. Here, we first observed increased LAT1 expression in pemetrexed-resistant non-small cell lung cancer (NSCLC) cells with high cancer stem cell (CSC) activity, and its mRNA expression level was associated with shorter overall survival in the lung adenocarcinoma dataset of the Cancer Genome Atlas database. The inhibition of LAT1 by a small molecule inhibitor, JPH203, or by RNA interference led to a significant reduction in tumorsphere formation and the downregulation of several cancer stemness genes in NSCLC cells through decreased AKT serine/threonine kinase (AKT)/mammalian target of rapamycin (mTOR) activation. The treatment of the cell-permeable leucine derivative promoted AKT/mTOR phosphorylation and reversed the inhibitory effect of JPH203 in the reduction of CSC activity in pemetrexed-resistant lung cancer cells. Furthermore, we observed that LAT1 silencing caused the downregulation of programmed cell death 1 ligand 1 (PD-L1) on lung cancer cells. The PD-L1⁺/LAT1⁺ subpopulation of NSCLC cells displayed great CSC activity with increased expression of several cancer stemness genes. These data suggest that LAT1 inhibitors can serve as anti-CSC agents and could be used in combination with immune checkpoint inhibitors in lung cancer therapy.     

The Next-Generation Immune Checkpoint LAG-3 and Its Therapeutic Potential in Oncology: Third Time’s a Charm

The blockade of immune checkpoints (ICPs), such as cytotoxic T lymphocyte associated protein-4 (CTLA-4) and programmed death-1 (PD-1) and its ligand (PD-L1), has propelled the field of immuno-oncology into its current era. Drugs targeting these ICPs have improved clinical outcome in a number of patients with solid and hematological cancers. Nonetheless, some patients have no benefit from these ICP-blocking therapies. This observation has instigated research into alternative pathways that are responsible for the escape of cancer cells from anti-cancer immune responses. From this research, a number of molecules have emerged as promising therapeutic targets, including lymphocyte activating gene-3 (LAG-3), a next-generation ICP. We will review the current knowledge on the biological activity of LAG-3 and linked herewith its expression on activated immune cells. Moreover, we will discuss the prognostic value of LAG-3 and how LAG-3 expression in tumors can be monitored, which is an aspect that is of utmost importance, as the blockade of LAG-3 is actively pursued in clinical trials.     

(20S) Ginsenoside Rh2 Exerts Its Anti-Tumor Effect by Disrupting the HSP90A-Cdc37 System in Human Liver Cancer Cells

(20S) ginsenoside Rh2 (G-Rh2), a major bioactive metabolite of ginseng, effectively inhibits the survival and proliferation of human liver cancer cells. However, its molecular targets and working mechanism remain largely unknown. Excitingly, we screened out heat shock protein 90 alpha (HSP90A), a key regulatory protein associated with liver cancer, as a potential target of (20S) G-Rh2 by phage display analysis and mass spectrometry. The molecular docking and thermal shift analyses demonstrated that (20S) G-Rh2 directly bound to HSP90A, and this binding was confirmed to inhibit the interaction between HSP90A and its co-chaperone, cell division cycle control protein 37 (Cdc37). It is well-known that the HSP90A-Cdc37 system aids in the folding and maturation of cyclin-dependent kinases (CDKs). As expected, CDK4 and CDK6, the two G₀-G₁ phase promoting kinases as well as CDK2, a key G₁-S phase transition promoting kinase, were significantly downregulated with (20S) G-Rh2 treatment, and these downregulations were mediated by the proteasome pathway. In the same condition, the cell cycle was arrested at the G₀-G₁ phase and cell growth was inhibited significantly by (20S) G-Rh2 treatment. Taken together, this study for the first time reveals that (20S) G-Rh2 exerts its anti-tumor effect by targeting HSP90A and consequently disturbing the HSP90A-Cdc37 chaperone system. HSP90A is frequently overexpressed in human hepatoma cells and the higher expression is closely correlated to the poor prognosis of liver cancer patients. Thus, (20S) G-Rh2 might become a promising alternative drug for liver cancer therapy.     

Association of Irisin/FNDC5 with ERRα and PGC-1α Expression in NSCLC

The rapid growth and division of cancer cells are associated with mitochondrial biogenesis or switching to glycolysis. ERRα, PGC-1α and irisin/FNDC5 are some of the proteins that can influence these processes. The aim of this study was to determine the correlation of these proteins in non-small cell lung cancer (NSCLC) and to investigate their association with clinicopathological parameters. Immunohistochemistry reactions were performed on tissue microarrays (860 NSCLC, 140 non-malignant lung tissue). The normal fibroblast cell line (IMR-90) and lung cancer cell lines (NCI-H1703 and NCI-H522) were used as co-cultures. The mRNA levels of FNDC5 and ESRRA (encoding ERRα) were assessed in IMR-90 cells after co-culture with lung cancer cells. We observed a decreased level of ERRα with an increase in tumor size (T), stages of the disease, and lymph node metastases (N). In the adenocarcinoma (AC) subtype, patients with a higher ERRα expression had significantly longer overall survival. A moderate positive correlation was observed between FNDC5 mRNA and ESRRA mRNA in NSCLCs. The expression of FNDC5 mRNA in IMR-90 cells increased after 24 h, and ESRRA gene expression increased after 48 h of co-culture. The ERRα receptor with PGC-1α participates in the control of FNDC5/irisin expression. Normal fibroblasts revealed an upregulation of the FNDC5 and ESRRA genes under the influence of lung cancer cells.     

Novel Drugs with High Efficacy against Tumor Angiogenesis

Angiogenesis is involved in physiological and pathological processes in the body. Tumor angiogenesis is a key factor associated with tumor growth, progression, and metastasis. Therefore, there is great interest in developing antiangiogenic strategies. Hypoxia is the basic initiating factor of tumor angiogenesis, which leads to the increase of vascular endothelial growth factor (VEGF), angiopoietin (Ang), hypoxia-inducible factor (HIF-1), etc. in hypoxic cells. The pathways of VEGF and Ang are considered to be critical steps in tumor angiogenesis. A number of antiangiogenic drugs targeting VEGF/VEGFR (VEGF receptor) or ANG/Tie2, or both, are currently being used for cancer treatment, or are still in various stages of clinical development or preclinical evaluation. This article aims to review the mechanisms of angiogenesis and tumor angiogenesis and to focus on new drugs and strategies for the treatment of antiangiogenesis. However, antitumor angiogenic drugs alone may not be sufficient to eradicate tumors. The molecular chaperone heat shock protein 90 (HSP90) is considered a promising molecular target. The VEGFR system and its downstream signaling molecules depend on the function of HSP90. This article also briefly introduces the role of HSP90 in angiogenesis and some HSP90 inhibitors.