Lung Cancer Mouse Research Articles

COX-2/PTGS2-targeted herbal-derived oligonucleotide drug HQi-sRNA-2 was effective in spontaneous mouse lung cancer model.

In 2020, the number of deaths caused by lung cancer worldwide reached 1,796,144, making it the leading cause of cancer-related deaths. Cyclooxygenase-2/prostaglandin endoperoxide synthase 2 (COX-2/PTGS2) is overexpressed in lung cancer, which promotes tumor proliferation, invasion, angiogenesis, and resistance to apoptosis. Here, we report that the oligonucleotide drug HQi-sRNA-2 from Traditional Chinese Medicine Huangqin targeting COX-2/PTGS2 significantly inhibited proliferation, migration, and invasion and induced apoptosis in the human lung cancer cell line NCI-H460. Oral delivery of HQi-sRNA-2 bencaosomes prolonged survival, reduced tumor burden, and maintained weight in a spontaneous mouse lung cancer model. Compared with paclitaxel, HQi-sRNA-2 may be less toxic and have approximately equal efficacy in reducing tumor burden. Our previous studies reported that herbal small RNAs (sRNAs) are functional medical components. Our data suggest that sphingosine (d18:1)-HQi-sRNA-2 bencaosomes, targeting COX-2/PTGS2 and downregulating the PI3K and AKT signaling pathways, may provide novel therapeutics for lung cancer.

Antitumor Study of Neoantigen-reactive T Cells Co-expressing IL-7 and CCL19 in Mouse Lung Cancer

Neoantigen reactive T cell (NRT) has the ability to inhibit the growth of tumors expressing specific neoantigens. However, due to the difficult immune infiltration and the inhibition of tumor microenvironment, the therapeutic effect of NRT in solid tumors is limited. In this study, we designed NRT cells (7×19 NRT) that can express both interleukin-7 (IL-7) and chemokine C-C motif ligand 19 (CCL19) in mouse lung cancer cells, and evaluated the difference in anti-tumor effect between 7×19 NRT cells and conventional NRT cells. We performed next-generation sequencing and neoantigen prediction for mouse Lewis lung carcinoma (LLC), prepared RNA vaccine, cultured NRT cells, constructed retroviral vectors encoding IL-7 and CCL19, transduced NRT cells and IL-7 and CCL19 were successfully expressed, and 7×19 NRT was successfully obtained. The anti-tumor effect was evaluated in vivo and in vitro in mice. The 7×19 NRT cells significantly enhanced the proliferation and invasion ability of T cells by secreting IL-7 and CCL19, achieved significant tumor inhibition in the mouse lung cancer and extended the survival period of mice. The T cell infiltration into tumor tissue and the necrosis of tumor tissue increased significantly after 7×19 NRT treatment. In addition, both 7×19 NRT treatment and conventional NRT treatment were safe. The anti-solid tumor ability of NRT cells is significantly enhanced by the arming of IL-7 and CCL19, which is a safe and effective genetic modification of NRT.

Glucose-6-phosphate dehydrogenase maintains redox homeostasis and biosynthesis in LKB1-deficient KRAS-driven lung cancer

Cancer cells depend on nicotinamide adenine dinucleotide phosphate (NADPH) to combat oxidative stress and support reductive biosynthesis. One major NADPH production route is the oxidative pentose phosphate pathway (committed step: glucose-6-phosphate dehydrogenase, G6PD). Alternatives exist and can compensate in some tumors. Here, using genetically-engineered lung cancer mouse models, we show that G6PD ablation significantly suppresses KrasG12D/+;Lkb1-/- (KL) but not KrasG12D/+;P53-/- (KP) lung tumorigenesis. In vivo isotope tracing and metabolomics reveal that G6PD ablation significantly impairs NADPH generation, redox balance, and de novo lipogenesis in KL but not KP lung tumors. Mechanistically, in KL tumors, G6PD ablation activates p53, suppressing tumor growth. As tumors progress, G6PD-deficient KL tumors increase an alternative NADPH source from serine-driven one carbon metabolism, rendering associated tumor-derived cell lines sensitive to serine/glycine depletion. Thus, oncogenic driver mutations determine lung cancer dependence on G6PD, whose targeting is a potential therapeutic strategy for tumors harboring KRAS and LKB1 co-mutations.

Targeted nutritional intervention attenuates experimental lung cancer cachexia.

Cachexia, a syndrome with high prevalence in non-small cell lung cancer patients, impairs quality of life and reduces tolerance and responsiveness to cancer therapy resulting in decreased survival. Optimal nutritional care is pivotal in the treatment of cachexia and a recommended cornerstone of multimodal therapy. Here, we investigated the therapeutic effect of an intervention diet consisting of a specific combination of high protein, leucine, fish oil, vitamin D, galacto-oligosaccharides, and fructo-oligosaccharides on the development and progression of cachexia in an orthotopic lung cancer mouse model. Eleven-week-old male 129S2/Sv mice were orthotopically implanted with 344P lung epithelial tumour cells or vehicle (control). Seven days post-implantation tumour-bearing (TB) mice were allocated to either intervention- or isocaloric control diet. Cachexia was defined as 5days of consecutive body weight loss, after which mice were euthanized for tissue analyses. TB mice developed cachexia accompanied by significant loss of skeletal muscle mass and epididymal fat mass compared with sham operated mice. The cachectic endpoint was significantly delayed (46.0±15.2 vs. 34.7±11.4days), and the amount (-1.57±0.62 vs. -2.13±0.57g) and progression (-0.26±0.14 vs. -0.39±0.11g/day) of body weight loss were significantly reduced by the intervention compared with control diet. Moreover, systemic inflammation (pentraxin-2 plasma levels) and alterations in molecular markers for proteolysis and protein synthesis, indicative of muscle atrophy signalling in TB-mice, were suppressed in skeletal muscle by the intervention diet. Together, these data demonstrate the potential of this multinutrient intervention, targeting multiple components of cachexia, as integral part of lung cancer management.

Nonviral CRISPR/Cas9 mutagenesis for streamlined generation of mouse lung cancer models

Functional analysis in mouse models is necessary to establish the involvement of a set of genetic variations in tumor development. A modeling platform to facilitate and cost-effectively analyze the role of multiple genes in carcinogenesis would be valuable. Here, we present an innovative strategy for lung mutagenesis using CRISPR/Cas9 ribonucleoproteins delivered via cationic polymers. This approach allows the simultaneous inactivation of multiple genes. We validate the effectiveness of this system by targeting a group of tumor suppressor genes, specifically Rb1, Rbl1, Pten, and Trp53, which were chosen for their potential to cause lung tumors, namely small cell lung carcinoma (SCLC). Tumors with histologic and transcriptomic features of human SCLC emerged after intratracheal administration of CRISPR/polymer nanoparticles. These tumors carried loss-of-function mutations in all four tumor suppressor genes at the targeted positions. These findings were reproduced in two different pure genetic backgrounds. We provide a proof of principle for simplified modeling of lung tumorigenesis to facilitate functional testing of potential cancer-related genes.

Mitigating Effect of Matricin Against Benzo(a)pyrene-induced Lung Carcinogenesis in Experimental Mice Model.

Lung cancer is a life-threatening disease that is still prevalent worldwide. This study aims to evaluate the effects of matricin, a sesquiterpene, on the carcinogenic agent benzo(a)pyrene [B(a)P]-induced lung cancer in Swiss albino mice. Lung cancer was induced by oral administration of B(a)P at 50 mg/kg b. wt. in model Swiss-albino mice (group II) as well in experimental group III, and treated with matricin (100 mg/kg b. wt.) in group III. Upon completion of treatment for 18 weeks, the changes in body weight, tumor formation, enzymatic and non-enzymatic antioxidant levels (GSH, SOD, GPx, GR, QR, CAT), lipid peroxidation (LPO) level, pro-inflammatory cytokines (TNF-α, IL-6, IL-1β), immunoglobulin levels (IgG, IgM), apoptosis markers (Bax, Bcl-xL), tumor markers (carcinoembryogenic antigen (CEA), neuron-specific enolase (NSE)), and histopathological (H&E) alterations were determined. The results indicate that B(a)P caused a significant increase of tumor formation in the lungs, increased tumor markers and inflammatory cytokines in serum, and depletion of enzymatic/ non-enzymatic antioxidants and immunoglobulins, compared to the untreated control group. Matricin treatment significantly reversed the changes caused by B(a)P as evidenced by the biochemical and histopathological assays. The changes caused by matricin clearly indicate the cancer-preventive effects of matricin against B(a)P-induced lung cancer in animal models, which can be attributed to the antioxidant activity, immunomodulation, and mitigation of the NF-kβ pathway.

Dynamic Changes in the Immune Microenvironment in Tumor-Draining Lymph Nodes of a Lewis Lung Cancer Mouse Model After Microwave Ablation.

Microwave ablation (MWA) is a minimally invasive technique for treating lung cancer. It can induce immune response; however, its effect on the immune microenvironment in tumor-draining lymph nodes (TdLN) is not well understood. This study aims to identify changes in the immune microenvironment in TdLN following MWA in a Lewis lung cancer (LLC) mouse model. LLC mouse model was established and followed by MWA. TdLN were collected at various time points, including pre-MWA and days 1, 2, 4, and 8 post-MWA. Flow cytometry was used to determine the frequencies of CD4+ T cells, CD8+ T cells, regulatory T (Treg) cells, natural killer (NK) cells, dendritic cells (DCs) and other immune cells in the TdLN. Certain cytokines were also detected. Compared with pre-MWA, the frequency of CD4+ T cells significantly increased from day 1 to day 8 post-MWA. The frequency of CD8+ T cells decreased significantly on days 2 and 4, but no significant changes occurred on days 1 and 8. Significant decreases in the frequencies of Treg cells and Klrg1+ Treg cells were observed from day 1 to day 4. On days 4 and 8, there was a significant increase in the frequency of NK cells. The frequency of resident cDC2 significantly increased on day 4, whereas CD11b+ migratory cDCs increased on day 1. Additionally, on day 4, a notable rise was observed in the frequency of NK cells secreting IFN-γ, while on day 8, there was a significant increase in the frequency of CD8+ T cells secreting both IFN-γ and TNF-α. MWA of lung cancer can alter the immune microenvironment in the TdLN, triggering immune responses. These changes are particularly evident and intricate within the initial 4 days post-MWA. Treatment combined with MWA within a certain period may significantly enhance anti-tumor immunity.

Antitumor efficacy of atorvastatin in lung cancer mice model

Antitumor efficacy of atorvastatin in lung cancer mice model

S-(N,N-diethyldithiocarbamoyl)-N-acetyl-l-cysteine for the treatment of non-small cell lung cancer through regulating NF-κB signalling pathway without neurotoxicity

The discovery of novel targeted agents for non-small cell lung cancer (NSCLC) remains an important research landscape due to the limited efficacy, side effects and drug resistance of current treatment options. Among many repurposed drugs, disulphiram (DSF) has shown the potential to target tumours. However, its unpleasant neurotoxicity greatly limits its use. A DSF derivative, S-(N,N-diethyldithiocarbamoyl)-N-acetyl-l-cysteine (DS-NAC), was synthesised against NSCLC. The therapeutic effects, mechanism and toxicities of DS-NAC were evaluated in A549 and H460 cells and the mouse model of in situ lung cancer. The in vitro results exhibited that DS-NAC had potent anti-proliferation, apoptotic, anti-metastasis and epithelial–mesenchymal transition (EMT) inhibition effects. In the orthotopic lung cancer mouse model, therapeutic effects of DS-NAC were better than those of DSF and were similar to docetaxel (DTX). Also, results from western blot and immunohistochemistry showed that DS-NAC in combination with copper exerted therapeutic effects via regulating NF-κB signalling pathway and ROS-related proteins such as HIF-1α, Nrf2 and PKC-δ rather than regulating ROS level directly. Moreover, the safety evaluation study showed that DS-NAC had low haematologic and hepatic toxicities in comparison with DTX as well as low neurological toxicity compared with DSF. DS-NAC could be a promising anti-lung cancer agent with a favourable safety profile.

Liraglutide enhances the effect of checkpoint blockade in lung and liver cancers through the inhibition of neutrophil extracellular traps.

Glucagon-like peptide-1 (GLP-1) regulates glycemic excursions by augmenting insulin production and inhibiting glucagon secretion. Liraglutide, a long-acting GLP-1 analog, can improve glycemic control for treating type 2 diabetes and prevent neutrophil extravasation in inflammation. Here, we explored the role of liraglutide in the development and therapy of murine lung and liver cancers. In this study, liraglutide substantially decreased circulating neutrophil extracellular trap (NET) markers myeloperoxidase, elastase, and dsDNA in Lewis lung cancer (LLC) and Hepa1-6 tumor-bearing mice. Furthermore, liraglutide downregulated NETs and reactive oxygen species (ROS) of neutrophils in the tumor microenvironment. Functionally, in vitro experiments showed that liraglutide reduced NET formation by inhibiting ROS. In addition, we showed that liraglutide enhanced the anti-tumoral efficiency of programmed cell death-1 (PD-1) inhibition in LLC and Hepa1-6 tumor-bearing C57BL/6 mice. However, the removal of NETs significantly weakened the antitumor efficiency of liraglutide. We further demonstrated that the long-term antitumor CD8+ T cell responses induced by the combination therapy rejected rechallenges by respective tumor cell lines. Taken together, our findings suggest that liraglutide may promote the anti-tumoral efficiency of PD-1 inhibition by reducing NETs in lung and liver cancers.

Autocrine insulin-like growth factor 2 signaling as a potential target in the associated development of pulmonary emphysema and cancer in smokers

BackgroundTobacco smoking causes pulmonary inflammation, resulting in emphysema, an independent risk factor for lung cancer. Induction of insulin-like growth factor 2 (IGF2) in response to lung injury by tobacco carcinogens, 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol and polycyclic aromatic hydrocarbon benzo[a]pyrene in combination (NB), is critical for the proliferation of alveolar type 2 cells (AT2s) for lung repair. However, persistent IGF2 overexpression during NB-induced severe injury results in hyperproliferation of AT2s without coordinated AT2-to-AT1 differentiation, disrupting alveolar repair, which leads to the concurrent development of emphysema and lung cancer. The current study aims to verify the role of IGF2 signaling in the associated development of emphysema and cancer and develop effective pharmaceuticals for the diseases using animal models that recapitulate the characteristics of these chronic diseases.MethodsThe pathogenesis of pulmonary emphysema and cancer was analyzed by lung function testing, histological evaluation, in situ zymography, dihydroethidium staining, and immunofluorescence and immunohistochemistry analyses utilizing mouse models of emphysema and cancer established by moderate exposure to NB for up to seven months.ResultsModerate NB exposure induced IGF2 expression in AT2s during the development of pulmonary emphysema and lung cancer in mice. Using AT2-specific insulin receptor knockout mice, we verified the causative role of sustained IGF2 signaling activation in AT2s in emphysema development. IGF2-targeting strategies, including voltage-dependent calcium channel blocker (CCB) and a neutralizing antibody, significantly suppressed the NB-induced development of emphysema and lung cancer. A publicly available database revealed an inverse correlation between the use of calcium channel blockers and a COPD diagnosis.ConclusionsOur work confirms sustained IGF2 signaling activation in AT2s couples impaired lung repair to the concurrent development of emphysema and cancer in mice. Additionally, CCB and IGF2-specific neutralizing antibodies are effective pharmaceuticals for the two diseases.

Targeting SIRT3 signaling alleviates lung carcinoma progression by reducing hyperinsulinemia in postmenopausal obese mice: Protective intervention of betaxanthin

Metabolic disturbances associated with obesity increase the risk and advancement of various tumor forms, including postmenopausal lung cancer, in humans. Betaxanthin, a type of yellow-orange pigment found in certain plants, particularly in some flowers, fruits, and vegetables, can be utilized to inhibit the growth of tumors and lessen the metabolic dysfunctions brought on by obesity. However, the functional role of betaxanthin (BET) in suppressing lung cancer progression and its potential mechanisms are still not fully understood. In this study, we examined the regulation of BET in obese mice with postmenopausal lung cancer receiving a high-fat diet (HFD). Initially, we found that BET could significantly mitigate the metabolic dysfunction that a high-fat diet causes in mice. Improved histological changes in the mammary fat pad, lower hepatic lipid deposition, and improved glucose tolerance and insulin resistance demonstrated this. Following that, BET treatment inhibited the formation of lung neoplasms in in-situ cancer animal models that were stimulated by HFD. Furthermore, in a postmenopausal mouse model fed HFD, BET significantly reduced the growth of pre-existing lung tumors, as well as attenuating metabolic abnormalities. Notably, in vitro research revealed that BET co-culture significantly reduced the proliferation and migration of mouse lung cancer cells, whereas insulin exposure increased these processes. Research on animals verified that lung cancer progressed due to hyperinsulinemia; however, BET treatment might inhibit this condition in postmenopausal mice on a high-fat diet. BET could up-regulate sirtuin 3 (SIRT3) expression levels in tumor, liver, and mammary fat pad tissues in postmenopausal mice with HFD-induced obesity, according to a bioinformatic and molecular biology study. Crucially, eradicating SIRT3 expression completely eliminated BET’s inhibitory effects on insulin-stimulated cancer cell proliferation and migration. This suggests that increasing SIRT3 expression may be required for BET to perform its anti-tumor function against lung cancer in conditions of hyperinsulinemia. Our findings showed that by modifying SIRT3 signaling, BET consumption is probably effective in the prevention and treatment of obesity-related lung cancer.

Immune profiling of mouse lung adenocarcinoma paraffin tissues using multiplex immunofluorescence panel: a pilot study

BackgroundImmune profiling has become an important tool for identifying predictive, prognostic and response biomarkers for immune checkpoint inhibitors from tumor microenvironment (TME). We aimed to build a multiplex immunofluorescence (mIF) panel to apply to formalin-fixed and paraffin-embedded tissues in mice tumors and to explore the programmed cell death protein 1/ programmed cell death 1 ligand 1 (PD-1/PD-L1) axis.ResultsAn automated eight-color mIF panel was evaluated to study the TME using seven antibodies, including cytokeratin 19, CD3e, CD8a, CD4, PD-1, PD-L1, F4-80 and DAPI, then was applied in six mice lung adenocarcinoma samples. Cell phenotypes were quantified by software to explore the co-localization and spatial distribution between immune cells within the TME. This mice panel was successfully optimized and applied to a small cohort of mice lung adenocarcinoma cases. Image analysis showed a sparse degree of immune cell expression pattern in this cohort. From the spatial analysis we found that T cells and macrophages expressing PD-L1 were close to the malignant cells and other immune cells.ConclusionsComprehensive immune profiling using mIF in translational studies improves our ability to correlate the PD-1/PD-L1 axis and spatial distribution of lymphocytes and macrophages in mouse lung cancer cells to provide new cues for immunotherapy, that can be translated to human tumors for cancer intervention.

High-fat-diet-induced obesity promotes simultaneous progression of lung cancer and atherosclerosis in apolipoprotein E-knockout mice.

Clinical studies have shown that atherosclerotic cardiovascular disease and cancer often co-exist in the same individual. The present study aimed to investigate the role of high-fat-diet (HFD)-induced obesity in the coexistence of the two diseases and the underlying mechanism in apolipoprotein E-knockout (ApoE-/-) mice. Male ApoE-/- mice were fed with a HFD or a normal diet (ND) for 15 weeks. On the first day of Week 13, the mice were inoculated subcutaneously in the right axilla with Lewis lung cancer cells. At Weeks 12 and 15, serumlectin-like oxidized low-density lipoprotein receptor-1 (LOX-1) and vascular endothelial growth factor levels were measured by enzyme-linked immunosorbent assay, and blood monocytes and macrophages were measured by fluorescence-activated cell sorting. At Week 15, the volume and weight of the local subcutaneous lung cancer and metastatic lung cancer and the amount of aortic atherosclerosis were measured. At Week 15, compared with mice in the ND group, those in the HFD group had a larger volume of local subcutaneous cancer (p = 0.0004), heavier tumors (p = 0.0235), more metastatic cancer in the lungs (p < 0.0001), a larger area of lung involved in metastatic cancer (p = 0.0031), and larger areas of atherosclerosis in the aorta (p < 0.0001). At Week 12, serum LOX-1, serum vascular endothelial growth factor, and proportions of blood monocytes and macrophages were significantly higher in the HFD group than those in the ND group (p = 0.0002, p = 0.0029, p = 0.0480, and p = 0.0106, respectively); this trend persisted until Week 15 (p = 0.0014, p = 0.0012, p = 0.0001, and p = 0.0204). In this study, HFD-induced obesity could simultaneously promote progression of lung cancer and atherosclerosis in the same mouse. HFD-induced upregulation of LOX-1 may play an important role in the simultaneous progression of these two conditions via the inflammatory response and VEGF.

Unraveling the Mechanism of Curculiginis Rhizoma in Suppressing Cisplatin Resistance in Non-Small Cell Lung Cancer: An Experimental Study.

Non-small cell lung cancer (NSCLC) stands as one of the most prevalent malignancies, and chemotherapy remains the primary treatment for advanced stages. However, the high expression of ABC binding cassette transporters, including MRP, P-gp, and LRP, along with multidrug resistance proteins, has been identified as a significant factor contributing to decreased chemotherapy drug sensitivity. This study aims to explore the impact and underlying mechanisms of Curculiginis Rhizoma [Hypoxidaceae; Curculigo orchioides Gaertn.] (CR) in combination with cisplatin on improving chemoresistance mediated by ABC binding cassette transporters and multidrug resistance proteins in NSCLC. To unravel the relationship between JNK, MRP, P-gp, and LRP in NSCLC and gain insights into the regulatory mechanism of CR, this study employs an integrated approach encompassing bioinformatics, molecular docking, molecular dynamics, animal and cellular experiments. Bioinformatics analysis revealed a significant increase in the expression levels of JNK, MRP, P-gp, and LRP subtypes in multidrug-resistant non-small cell lung cancer. Subsequent animal experiments have shown that the combination of CR with cisplatin can improve the survival rate of lung cancer mice. Molecular docking and molecular dynamics analyses demonstrated favorable binding interactions between curculigoside and the aforementioned subtypes of JNK, MRP, P-gp, and LRP. In cellular experiments, the combination of cisplatin with both curculigoside and CR extract resulted in a notable decrease in cell viability and downregulation of the expression of JNK1, JNK2, MRP1, MRP2, MRP4, P-gp, and LRP1 in A549/cis cells. Remarkably, curculigoside exerted a significant downregulation effect on the expression levels of JNK1, MRP1, MRP2, MRP4, and LRP1. CR, particularly its main effective metabolite, curculigoside, has the potential to enhance the sensitivity of non-small cell lung cancer to cisplatin by regulating levels of JNK/MRP/LRP/P-gp and mitigating multidrug resistance.

Gene expression correlates of adagrasib response in KRASG12C mutated non-small cell lung cancer (NSCLC).

8529 Background: KRASG12C inhibitors such as adagrasib and sotorasib have shown clinical promise in targeting KRASG12C-mutated lung cancers; however, most patients develop primary or secondary resistance. A biomarker of response to KRASG12C inhibitor is needed for better patient stratification and to understand the resistance mechanism. Methods: We analyzed transcriptional correlates of adagrasib treatment outcome in 68 patients in the KRYSTAL-1 trial, a phase 1/2 clinical trial of adagrasib monotherapy in the second line and beyond treatment for NSCLC. We also treated KRAS-mutated lung cancer mouse models and organoids with a KRAS inhibitor for the long term and characterized the resistant tumors’ transcriptional profile to identify resistance mechanisms. We also performed serial gene expression analysis of KrasG12D mutated lung organoids undergoing squamous transformation to identify transcription factor involved in the resistance process. Results: In patients with lung adenocarcinoma with KRASG12C and STK11/ LKB1 co-mutations, we find an enrichment of the squamous cell carcinoma gene signature in pre-treatment biopsies correlates with a poor response to adagrasib. Studies of Lkb1-deficient KRASG12C and KrasG12D lung cancer mouse models and organoids, treated with KRAS inhibitors adagrasib and MRTX1133, respectively, reveal that tumors invoke a lineage plasticity program, adeno-to-squamous transition (AST), that enables resistance to KRAS inhibition. We identify TP63 to be a transcription factor whose expression correlated with squamous transformation. The analysis of lineage plasticity program, adagrasib resistant tumors, and p63 regulon revealed KRT6A to be a common biomarker whose expression correlated with overall survival in the KRYSTAL-1 cohort. Conclusions: KRASG12C mutated lung adenocarcinoma patients with a higher expression of squamous cell carcinoma gene expression signature respond poorly to adagrasib treatment. Expression of the AST plasticity signature and KRT6A at baseline correlates with poor adagrasib responses. These data indicate the role of AST in KRAS inhibitor resistance and provide predictive biomarkers for KRAS-targeted therapies in lung cancer.

Preclinical analysis of the efficacy of near-infrared photoimmunotherapy and anti-PD-1 antibody combination therapy for treating CD73-positive lung cancer.

e14521 Background: The CD73/adenosine pathway, which converts extracellular ATP to adenosine, is emerging as a promising target for immunotherapy in various cancers. In the tumor microenvironment, hypoxia triggers upregulation of adenosine-activating molecules, including CD73, a membrane-anchored enzyme expressed on cancer cells. CD73 induces the expression of PD-1 and CTLA4, and all of this leads to immunosuppression. Therefore, we focused utilizing near-infrared photoimmunotherapy (NIR-PIT) to kill CD73-expressing cells and alleviate immunosuppression. NIR-PIT is an innovative cancer treatment approach which involves irradiating NIR light after the administrated antibody-photosensitizer conjugate binds to the target cells. Methods: In vitro, anti-CD73 antibody-IR700 conjugate was prepared and added to the CMT167 and LLC cell lines, which were irradiated with NIR light under a microscope. In vivo, a mouse model was established using the CMT167 cell line. The mice were divided into 6 groups: 1) NIR-PIT group against CD73; 2) anti-PD-1 antibody group; 3) combined treatment group of 1) and 2); 4) control group; 5) anti-CD73 antibody-IR700 conjugate group; and 6) near-infrared light irradiation group. Tumor volume and survival rate were compared among these groups. Additionally, after 2 weeks of treatment, tumors in the four groups 1) to 4) above were pathologically evaluated using two different AI-equipped software programs. Results: Microscope observation revealed that NIR-PIT induced immunogenic cell death of CD73-positive mouse lung cancer cell lines in vitro. In vivo, the effect of NIR-PIT on this mouse model was also demonstrated. Although previous studies have shown the efficacy of treatment with anti-CD73 antibodies, the present study compared the CD73-IR700 conjugate group to the NIR-PIT to CD73 group, and the latter group showed better therapeutic outcomes. Additionally, the combined treatment group displayed the highest anti-tumor efficacy in terms of survival and tumor volume. Pathological evaluation of the tumor interior showed a significant increase in tumor-infiltrating lymphocytes compared to the other groups. Additionally, the percentage of tumor cells was significantly reduced, while the percentage of stroma, including fibrous tissue, was significantly increased. In line with other studies that have already reported the additive effect of combination therapy with anti-CD73 antibodies and immune checkpoint inhibitors, the present study showed that the combination therapy resulted in further activation of immune cells and enhanced the therapeutic effect. Conclusions: CD73 represents a promising target for NIR-PIT, with potential for enhanced effectiveness when used in combination with immune checkpoint inhibitors.

Tumor immunogenicity regulates host immune responses, and conventional dendritic cell type 2 uptakes the majority of tumor antigens in an orthotopic lung cancer model.

Human lung cancer carries high genetic alterations, expressing high tumor-specific neoantigens. Although orthotopic murine lung cancer models recapitulate many characteristics of human lung cancers, genetically engineered mouse models have fewer somatic mutations than human lung cancer, resulting in scarce immune cell infiltration and deficient immune responses. The endogenous mouse lung cancer model driven by Kras mutation and Trp53 deletion (KP model) has minimal immune infiltration because of a scarcity of neoantigens. Fine-tuning tumor antigenicity to trigger the appropriate level of antitumor immunity would be key to investigating immune responses against human lung cancer. We engineered the KP model to express antigens of OVA peptides (minOVA) as neoantigens along with ZsGreen, a traceable fluorescent conjugate. The KP model expressing minOVA exhibited stronger immunogenicity with higher immune cell infiltration comprised of CD8+ T cells and CD11c+ dendritic cells (DCs). Consequentially, the KP model expressing minOVA exhibits suppressed tumor growth compared to its origin. We further analyzed tumor-infiltrated DCs. The majority of ZsGreen conjugated with minOVA was observed in the conventional type 2 DCs (cDC2), where cDC1 has minimal. These data indicate that tumor immunogenicity regulates host immune responses, and tumor neoantigen is mostly recognized by cDC2 cells, which may play a critical role in initiating anti-tumor immune responses in an orthotopic murine lung cancer model.

The Triterpenoid CDDO-Methyl Ester Reduces Tumor Burden, Reprograms the Immune Microenvironment, and Protects from Chemotherapy-Induced Toxicity in a Preclinical Mouse Model of Established Lung Cancer.

NRF2 activation protects epithelial cells from malignancy, but cancer cells can upregulate the pathway to promote survival. NRF2 activators including CDDO-Methyl ester (CDDO-Me) inhibit cancer in preclinical models, suggesting NRF2 activation in other cell types may promote anti-tumor activity. However, the immunomodulatory effects of NRF2 activation remain poorly understood in the context of cancer. To test CDDO-Me in a murine model of established lung cancer, tumor-bearing wildtype (WT) and Nrf2 knockout (KO) mice were treated with 50-100 mg CDDO-Me/kg diet, alone or combined with carboplatin/paclitaxel (C/P) for 8-12 weeks. CDDO-Me decreased tumor burden in an Nrf2-dependent manner. The combination of CDDO-Me plus C/P was significantly (p < 0.05) more effective than either drug alone, reducing tumor burden by 84% in WT mice. CDDO-Me reduced the histopathological grade of WT tumors, with a significantly (p < 0.05) higher proportion of low-grade tumors and a lower proportion of high-grade tumors. These changes were augmented by combination with C/P. CDDO-Me also protected WT mice from C/P-induced toxicity and improved macrophage and T cell phenotypes in WT mice, reducing the expression of CD206 and PD-L1 on macrophages, decreasing immunosuppressive FoxP3+ CD4+ T cells, and increasing activation of CD8+ T cells in a Nrf2-dependent manner.

Epithelial−mesenchymal transition induced by tumor cell-intrinsic PD-L1 signaling predicts a poor response to immune checkpoint inhibitors in PD-L1-high lung cancer

BackgroundWe investigated the role of tumor cell-intrinsic PD-L1 signaling in the epithelial−mesenchymal transition (EMT) in non-small-cell lung cancer (NSCLC) and the role of EMT as a predictive biomarker for immune checkpoint inhibitor (ICI) therapy.MethodsPD-L1-overexpressing or PD-L1-knockdown NSCLC cells underwent RNA-seq and EMT phenotype assessment. Mouse lung cancer LLC cells were injected into nude mice. Two cohorts of patients with NSCLC undergoing ICI therapy were analyzed.ResultsRNA-seq showed that EMT pathways were enriched in PD-L1-high NSCLC cells. EMT was enhanced by PD-L1 in NSCLC cells, which was mediated by transforming growth factor-β (TGFβ). PD-L1 promoted the activation of p38-MAPK by binding to and inhibiting the protein phosphatase PPM1B, thereby increasing the TGFβ production. Tumor growth and metastasis increased in nude mice injected with PD-L1-overexpressing LLC cells. In the ICI cohort, EMT signature was higher in patients with progressive disease than in those with responses, and EMT was significantly associated with poor survival in PD-L1-high NSCLC. In PD-L1-high NSCLC, EMT was associated with increased M2-macrophage and regulatory T-cell infiltrations and decreased cytotoxic T-cell infiltration.ConclusionsTumor cell-intrinsic PD-L1 function contributes to NSCLC progression by promoting EMT. EMT may predict an unfavorable outcome after ICI therapy in PD-L1-high NSCLC.