Lewis Lung Carcinoma Research Articles

Abstract 3480: Modulating DNA and histone methylation as a therapeutic target for non-small cell lung cancer by shifting the composition of immune cells

Abstract Lung cancer is one of the most common malignancies worldwide with non-small cell lung cancer (NSCLC) being the dominant subtype. Surgical resection remains the best treatment for NSCLC, but late diagnosis with associated metastasis precludes this as a viable modality for most patients. Furthermore, other treatments such as targeted therapies, cytotoxics, and immunotherapy only benefit a minority of patients. Therefore, new therapeutic strategies are needed to benefit more patients presenting with this devastating disease. Recent studies reveal that DNMT inhibitor (DNMTi) promotes tumor-infiltrative cytotoxic CD8+ T cells, shifts myeloid-derived suppressor cells (MDSCs) to anti-tumor macrophage-like phenotype, and enhances immunotherapy efficacy. However, some immune-regulatory genes are silenced by H3K27me3, which is facilitated by the transcriptional repressor EZH2. Additionally, conditional ablation of Ezh2 in T cells boosts effector functions of CD8+ T cells and prevents the development of regulatory T cells. Given the above, the application of combining DNMTi and EZH2i might be a viable path to facilitate tumor immune microenvironment remodeling. In this study, we aimed to define the therapeutic potential of combining DNMT and EZH2 inhibition to facilitate an anti-tumor immune response. First, we explored the potential of combined DNMTi and EZH2i in a clinically relevant murine model of NSCLC, Lewis Lung Carcinoma 1 (LLC1). Therapeutic application in this model led to a significant reduction in tumor volume. Our transcriptomic data revealed DNMTi facilitated transcriptional perturbation of both coding and transposable element-related genes, which is driven further by EZH2i combination. Immune deconvolution analysis uncovers combination therapy-induced shifts in tumor immune-cellularity, which is most profound for myeloid-associated populations. Orthogonal validation by flow cytometry defines significant reductions of MDSCs and regulatory T cells occurring within the tumor and associated microenvironment. Given the perturbations of T and myeloid cells observed in-vivo, we sought to define the drug effects on critical cell populations contained in these lineages. These data revealed that combining DNMTi and EZH2i promotes effector CD8+ T cells with more IFN-g secretion upon activation but halts the cells from exhaustion after chronic stimulation. Moreover, co-treatment promoted maturation, cross-presentation, and antigen-specific CD8+ T cell priming in dendritic cells (DCs). These data indicate that CD8+ T cells and DCs subjected to co-treatment might facilitate more robust tumor immune responses in the tumor microenvironment (TME). In summary, our data define an effective therapeutic paradigm for the treatment of NSCLC, which potentiates significant shifts in TME resident immune populations and dendritic cell phenotypes. Citation Format: Ying-Yu Chen. Modulating DNA and histone methylation as a therapeutic target for non-small cell lung cancer by shifting the composition of immune cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 3480.

Abstract 607: CD8+ T cell clones maintain a pool of memory-like Ly108+ cells in the tumor during their contraction phase

Abstract The immune response against cancer is effectuated by the recruitment of expanding CD8+ T cell clones. However, the fate of these clonal populations that have migrated into the tumor environment - whether they persist, contract, and/or produce memory cells - is unknown. An improved understanding of clonal durability and potency will be essential for the development of novel cancer immunotherapies. Here, we applied our previously developed bilateral tumor mouse model to track the fate of endogenous T cell clones responding to tumors in the same individual. This approach relied on the fact that the clonal composition of the two tumors is identical, allowing the sampling and comparison of tumor-infiltrating cells from the same clonal populations on different days in the same individual. Mice were inoculated bilaterally with Lewis lung carcinoma (LLC), and tumors were resected, one after the other, on days 14 and 21 post-inoculation. At each timepoint, sampled CD8+ T cells were gated on PD1, Ly108, and TIM-3 expression and subjected to T-cell receptor (TCR) sequencing to identify clones. We utilized this data to quantify changes in the fraction of memory-like (Ly108+TIM-3-) and exhausted (Ly108-TIM-3+, Ly108+TIM-3+) cells of individual clonal populations over time. The majority (75%) of tumor-infiltrating CD8+ T cells on day 14 belonged to clonal populations that persisted on day 21. These clonal populations were classed as ‘expanding’ or ‘contracting’ based on changes in their clone size. Despite the polyclonal response to the tumor, almost all expanding clones contained a high fraction of memory-like (Ly108+TIM-3-) cells, whereas almost all ‘contracting’ clones contained a high fraction of exhausted (TIM-3+) cells. These characteristics were sufficient to predict whether a clonal population at day 14 would later expand or contract. We further quantified how the makeup of ‘contracting’ clonal populations changed over time. While all ‘contracting’ clonal populations lost a considerable number of exhausted (TIM3+) cells, half of the clonal populations maintained a pool of memory-like (Ly108+TIM-3-) cells in the tumor. These results demonstrate that clonal dynamics can be predicted from static measurements, facilitating the development of novel methods to infer patient immune activity. The results also demonstrate the possibility of altering clonal fate. Reinvigoration of the pool of memory-like cells maintained in the tumor could re-expand contracting clonal populations, and thereby enhance the immune response against the tumor. Citation Format: Munetomo Takahashi, Mikiya Tsunoda, Hiroyasu Aoki, Haru Ogiwara, Shigeyuki Shichino, Kouji Matsushima, Satoshi Ueha. CD8+ T cell clones maintain a pool of memory-like Ly108+ cells in the tumor during their contraction phase [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 607.

Abstract 6384: ATG-034, an LILRB4 antagonist antibody, reinvigorates dendritic cells and prevents tumor progression

Abstract Background: Dendritic cells (DCs) are vital for initiating antigen-specific T cell-dependent antitumor immune responses. However, emerging evidence reveals that local DCs within the tumor microenvironment (TME) are tolerized to facilitate immune evasion. Reprogramming of DCs is recognized as a promising strategy for tumor immunotherapy. Leukocyte immunoglobulin-like receptor B4 (LILRB4) is an inhibitory receptor belonging to the LILR family which is mainly expressed on normal myeloid cells and myeloid-derived malignant cells. It is upregulated on tolerogenic DCs (tolDCs), which exhibit low expression levels of costimulatory molecules and resistance to DC maturation. Targeting LILRB4 to reprogram tolDCs has been reported to be a promising strategy for cancer treatment. Here we report the preclinical development of an LILRB4 antagonist antibody, ATG-034. Method: The protein-based and cell-based binding affinity of ATG-034 were measured using SPR, ELISA and FACS analysis. Fc receptor (FcR) stimulation assay was used to evaluate the ability of ATG-034 to block the interaction of LILRB4 with its ligand fibronectin. ATG-034-mediated enhancement of antigen presentation and co-stimulation ability of DCs was determined by FACS analysis for the change of surface expression of HLA-DR, HLA-ABC, CD86, and CD206. The mixed lymphocyte reaction (MLR) assay was used to assess the immunomodulatory potential of ATG-034. The in vivo antitumor efficacy of ATG-034 was evaluated in a radiation therapy-resistant murine Lewis lung carcinoma (LLC) syngeneic model. Results: ATG-034 binds to LILRB4 protein with a single-digit nM affinity. It potently reversed the fibronectin-mediated inhibition of FcR-driven TNF-α cytokine production. TolDCs were reprogrammed by ATG-034 to immunogenic DCs with significant upregulation of the expression of HLA-DR, HLA-ABC, CD86 and downregulation of the expression of CD206. Furthermore, ATG-034 remarkably reinvigorated tolDCs for priming T cell activation with noticeable upregulation of the expression of CD25 and the secretion of IFN-γ, enhancing antitumor immunity. In addition, 10 mg/kg ATG-034 significantly inhibited the LLC tumor growth in vivo with a TGI of 40.28%, while a benchmark antibody only demonstrated a 24.43% TGI. Conclusion: Our data show that ATG-034 reprograms tolDC, enhancing anti-tumor immunity, and demonstrates potent in vivo anti-tumor efficacy. Therefore, ATG-034 may be a promising strategy for the treatment of cancer. Citation Format: Ao Sun, Enlin Zheng, Huili Cao, Mengshi Cao, Suya Bai, Peng Chen, Linjie Tian, Jay Mei, Bo Shan, Bing Hou. ATG-034, an LILRB4 antagonist antibody, reinvigorates dendritic cells and prevents tumor progression [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 6384.

Abstract 2002: Intravenously administered STING agonist-loaded positively charged liposomes selectively target tumor microenvironments and reprogram tumor vasculature

Abstract Introduction: The stimulator of interferon genes (STING) signaling pathway activates type I interferon (IFN) in antigen-presenting cells to promote T cell priming and activation. STING agonists have shown great potential in preclinical studies, but they are not applicable for most patients due to limited systemic delivery. Therefore, we designed a positively charged liposome loaded with STING agonist (PoSTING) for systemic delivery of STING agonists and to selectively target tumor immunity and tumor vasculature. Method: We designed PoSTING using 1,2-dioleoyl-3-trimethylammonium-propane (DOTAP), 18:0 PEG2000 PE (PEG-PE), and 1,2-Dioleoyl-sn-glycero-3-phosphoethanolamine (DOPE). The properties and stability of PoSTING were evaluated by dynamic light scattering, transmission electron microscopy, and high-performance liquid chromatography. PoSTING (1 mg/kg) was administered intravenously to Lewis lung carcinoma (LLC)-bearing mouse models every three days. Tumor tissues were comprehensively analyzed by histologic, flow cytometric, and Nanostring immune profiling assays. Results: PoSTING efficiently loaded 88.5% of STING agonists. The average particle size was 198.4 nm, which avoided clearance by the mononuclear phagocyte system. PoSTING could fuse with cell membrane of target cells due to a positive surface charge (+44.53 mV). Stability tests found that PoSTING could be stored at 4°C for 12 weeks and remained stable in serum for a week. Intravenously administered PoSTING selectively accumulated in LLC tumor tissues and suppressed tumor growth. In particular, PoSTING was preferentially accumulated in tumor endothelial cells and perivascular cells, and suppressed tumor angiogenesis and normalized abnormal tumor vasculatures. PoSTING also increased expression levels of P-selectin, which is involved in endothelial-lymphocyte interactions and prepares tumor vessels for lymphocyte trafficking. PoSTING induces type I IFN responses in antigen-presenting cells, tumor ECs, and cancer cells, and promotes intratumoral infiltration of CD8+ T cells by remodeling tumor microenvironment. In addition, PoSTING significantly upregulated genes related to type I/II IFNs, Th1/Th2 responses, innate/adaptive immunity, and chemokines. Conclusion: PoSTING is a novel and stable systemic delivery platform that effectively and selectively delivers STING agonists to tumor tissue, triggering vascular normalization and immune reprogramming. Citation Format: Eun-Jin Go, Hannah Yang, Wooram Park, Won suk Lee, So Jung Kong, Seung Joon Lee, Dong Keun Han, Hong Jae Chon, Chan Kim. Intravenously administered STING agonist-loaded positively charged liposomes selectively target tumor microenvironments and reprogram tumor vasculature [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2002.

Abstract 1088: Acquired radiation resistance induces thiol-dependent cisplatin cross-resistance

Abstract Purpose. Resistance to radiation, both intrinsic and acquired, remains a significant clinical challenge. It is therefore import to identify the underlying molecular and cellular features involved in this resistance. Materials and Methods. We generated genetically-matched counterparts of various degrees of radiation sensitivity in non-small cell lung carcinoma (NSCLC) tumor models to define characteristics of acquired resistance through repeated exposures to conventional X-ray radiation. Results. As assessed by cell viability and clonogenic assays murine Lewis lung carcinoma (LLC) and human A549 cell lines acquired an approximate 1.5 - 2-fold increase in radiation resistance (RR) as compared to its parental counterpart. Morphologically, we found that A549-RR exhibited a greater nucleus-to-cytosol (N/C) ratio as compared to its respective counterpart. Since the N/C ratio has been linked to the differentiation state, we next investigated the epithelial-to-mesenchymal transition (EMT) phenotype and cellular plasticity. We found that A549 have a greater radiation-induced plasticity, as measured by E-cadherin, vimentin and double positive (DP) modulation, as compared to LLC. Additionally, migration was suppressed in A549-RR, as compared to A549. Subsequently, we confirmed that the LLC-RR and A549-RR are also more resistant to radiotherapy than their isogenic-matched counterpart in vivo. Remarkably, we found that the acquired radiation resistance also induced resistance to first-line chemotherapeutic cisplatin, but not carboplatin or oxaliplatin. This cross-resistance was attributed to induced elevated thiol levels. Decreasing the amount of thiols with buthionine sulfoximine (BSO), a gamma-glutamylcysteine synthetase inhibitor, sensitized the resistant cells to cisplatin, to levels similar as parental cells. Conclusions. By generating genetically-matched radiation resistant NSCLC tumor models we were able to identify and overcome cisplatin cross-resistance. This is an important finding arguing for multi-modality treatment regimens with the potential of improving clinical outcomes in the future. Citation Format: Samir V. Jenkins, Shruti Shah, Azemat Jamshidi-Parsian, Amir Mortazavi, Gunnar Boysen, Kieng B. Vang, Robert J. Griffin, Narasimhan Rajaram, Ruud P. Dings. Acquired radiation resistance induces thiol-dependent cisplatin cross-resistance [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1088.

Combined Effects of Programmed Cell Death-1 Blockade and Endostar on Brain Metastases of Lung Cancer.

The blockade of programmed cell death-1 (PD-1) and recombinant human endostatin can be used for the treatment of non-small cell lung cancer (NSCLC) and its metastasis. This study aims to explore the therapeutically potential of PD-1 blockade plus Endostar in brain metastasis of NSCLC. The mouse brain metastases model was established using Lewis lung carcinoma luciferase (LLC-Luc) and PC-9-Luc cells. Tumor metastasis in the brain and tumor burden were analyzed by using bioluminescence imaging (BLI), qRT-PCR and ELISA which were used to determine the mRNA and protein levels of biomarkers in tumor tissues. Immunohistochemical staining was used to determine the expression and location of CD31 in tumor tissues in the brain. Treatment with anti-PD-1 and Endostar suppressed tumor metastasis in the brain and prolonged overall survival rate in LLC-Luc and PC-9-Luc brain metastases mouse model. In addition, treatment with anti-PD-1 and Endostar inhibited the expressions of CD31 and VEGF in tumor tissues in the brain. Furthermore, treatment with anti-PD- 1 and Endostar significantly suppressed the levels of IL1β, IFNγ, and TGFβ in the tumor tissues. The combination of PD-1 blockade and endostar suppressed brain metastases of NSCLC.

Exosomes secreted from cardiomyocytes suppress the sensitivity of tumor ferroptosis in ischemic heart failure

Heart failure (HF) patients in general have a higher risk of developing cancer. Several animal studies have indicated that cardiac remodeling and HF remarkably accelerate tumor progression, highlighting a cause-and-effect relationship between these two disease entities. Targeting ferroptosis, a prevailing form of non-apoptotic cell death, has been considered a promising therapeutic strategy for human cancers. Exosomes critically contribute to proximal and distant organ-organ communications and play crucial roles in regulating diseases in a paracrine manner. However, whether exosomes control the sensitivity of cancer to ferroptosis via regulating the cardiomyocyte-tumor cell crosstalk in ischemic HF has not yet been explored. Here, we demonstrate that myocardial infarction (MI) decreased the sensitivity of cancer cells to the canonical ferroptosis activator erastin or imidazole ketone erastin in a mouse model of xenograft tumor. Post-MI plasma exosomes potently blunted the sensitivity of tumor cells to ferroptosis inducers both in vitro in mouse Lewis lung carcinoma cell line LLC and osteosarcoma cell line K7M2 and in vivo with xenograft tumorigenesis model. The expression of miR-22-3p in cardiomyocytes and plasma-exosomes was significantly upregulated in the failing hearts of mice with chronic MI and of HF patients as well. Incubation of tumor cells with the exosomes isolated from post-MI mouse plasma or overexpression of miR-22-3p alone abrogated erastin-induced ferroptotic cell death in vitro. Cardiomyocyte-enriched miR-22-3p was packaged in exosomes and transferred into tumor cells. Inhibition of cardiomyocyte-specific miR-22-3p by AAV9 sponge increased the sensitivity of cancer cells to ferroptosis. ACSL4, a pro-ferroptotic gene, was experimentally established as a target of miR-22-3p in tumor cells. Taken together, our findings uncovered for the first time that MI suppresses erastin-induced ferroptosis through releasing miR-22-3p-enriched exosomes derived from cardiomyocytes. Therefore, targeting exosome-mediated cardiomyocyte/tumor pathological communication may offer a novel approach for the ferroptosis-based antitumor therapy.

The Voltage-Gated Hv1 H+ Channel Is Expressed in Tumor-Infiltrating Myeloid-Derived Suppressor Cells.

Myeloid-derived suppressor cells (MDSCs) are key determinants of the immunosuppressive microenvironment in tumors. As ion channels play key roles in the physiology/pathophysiology of immune cells, we aimed at studying the ion channel repertoire in tumor-derived polymorphonuclear (PMN-MDSC) and monocytic (Mo-MDSC) MDSCs. Subcutaneous tumors in mice were induced by the Lewis lung carcinoma cell line (LLC). The presence of PMN-MDSC (CD11b+/Ly6G+) and Mo-MDSCs (CD11b+/Ly6C+) in the tumor tissue was confirmed using immunofluorescence microscopy and cells were identified as CD11b+/Ly6G+ PMN-MDSCs and CD11b+/Ly6C+/F4/80-/MHCII- Mo-MDSCs using flow cytometry and sorting. The majority of the myeloid cells infiltrating the LLC tumors were PMN-MDSC (~60%) as compared to ~10% being Mo-MDSCs. We showed that PMN- and Mo-MDSCs express the Hv1 H+ channel both at the mRNA and at the protein level and that the biophysical and pharmacological properties of the whole-cell currents recapitulate the hallmarks of Hv1 currents: ~40 mV shift in the activation threshold of the current per unit change in the extracellular pH, high H+ selectivity, and sensitivity to the Hv1 inhibitor ClGBI. As MDSCs exert immunosuppression mainly by producing reactive oxygen species which is coupled to Hv1-mediated H+ currents, Hv1 might be an attractive target for inhibition of MDSCs in tumors.

EXT418, a novel long-acting ghrelin, mitigates Lewis lung carcinoma induced cachexia in mice.

Ghrelin is a potential therapy for cachexia due to its orexigenic properties and anabolic effects on muscle and fat. However, its clinical use is limited by the short half-life of active (acylated) ghrelin (~11min in humans). EXT418 is a novel long-acting, constitutively active ghrelin analog created by covalently linking it to a vitamin D derivative. Here, we evaluated the effects and mechanisms of action of EXT418 on Lewis lung carcinoma (LLC)-induced cachexia in mice. Male C57BL/6J mice (5- to 7-month-old) were implanted with 1×106 heat-killed (HK) or live LLC cells. When the tumour was palpable, mice were injected with vehicle (T+V) or EXT418 daily (T+418 Daily, 0.25mg/kg/day) or every other day (T+418 EOD, 0.5mg/kg/EOD) for up to 14days, whereas HK-treated mice were given vehicle (HK+V). Subsets of T+418 Daily or EOD-treated mice were pair-fed to the T+V group. Body composition and grip strength were evaluated before tumour implantation and at the end of the experiment. Molecular markers were probed in muscles upon termination. In tumour-bearing mice, administration of EXT418 daily or EOD partially prevented weight loss (T+V vs. T+418 Daily, P=0.030; and vs. T+418 EOD, P=0.020). Similar effects were observed in whole body fat and lean body mass. Grip strength in tumour-bearing mice was improved by EXT418 daily (P=0.010) or EOD (P=0.008) administration compared with vehicle-treated mice. These effects of EXT418 on weight and grip strength were partially independent of food intake. EXT418 daily administration also improved type IIA (P=0.015), IIB (P=0.037) and IIX (P=0.050) fibre cross-sectional area (CSA) in tibialis anterior (TA) and EXT418 EOD improved CSA of IIB fibres in red gastrocnemius (GAS; P=0.005). In skeletal muscles, tumour-induced increases in atrogenes Fbxo32 and Trim63 were ameliorated by EXT418 treatments (TA and GAS/plantaris, PL), which were independent of food intake. EXT418 administration decreased expression of the mitophagy marker Bnip3 (GAS/PL; P≤0.010). Similar effects of EXT418 EOD were observed in p62 (GAS/PL; P=0.039). In addition, EXT418 treatments ameliorated the tumour-induced elevation in muscle Il6 transcript levels (TA and GAS/PL), independently of food intake. Il-6 transcript levels in adipose tissue and circulating IL-10 were elevated in response to the tumour but these increases were not significant with EXT418 administration. Tumour mass was not altered by EXT418. EXT418 mitigates LLC-induced cachexia by attenuating skeletal muscle inflammation, proteolysis, and mitophagy, without affecting tumour mass and partially independent of food intake.

PCSK9 regulates the efficacy of immune checkpoint therapy in lung cancer.

Proprotein convertase subtilisin/kexin type 9 (PCSK9) secreted by tumors was reported as a deleterious factor that led to the reduction of lymphocyte infiltration and the poorer efficacy of ICIs in vivo. This study aimed to explore whether PCSK9 expression in tumor tissue could predict the response of advanced non-small cell lung cancer (NSCLC) to anti-PD-1 immunotherapy and the synergistic antitumor effect of the combination of the PCSK9 inhibitor with the anti-CD137 agonist. One hundred fifteen advanced NSCLC patients who received anti-PD-1 immunotherapy were retrospectively studied with PCSK9 expression in baseline NSCLC tissues detected by immunohistochemistry (IHC). The mPFS of the PCSK9lo group was significantly longer than that of the PCSK9hi group [8.1 vs. 3.6 months, hazard ratio (HR): 3.450; 95% confidence interval (CI), 2.166-5.496]. A higher objective response rate (ORR) and a higher disease control rate (DCR) were observed in the PCSK9lo group than in the PCSK9hi group (54.4% vs. 34.5%, 94.7% vs. 65.5%). Reduction and marginal distribution of CD8+ T cells were observed in PCSK9hi NSCLC tissues. Tumor growth was retarded by the PCSK9 inhibitor and the anti-CD137 agonist alone in the Lewis lung carcinoma (LLC) mice model and further retarded by the PCSK9 inhibitor in combination with the CD137 agonist with long-term survival of the host mice with noticeable increases of CD8+ and GzmB+ CD8+ T cells and reduction of Tregs. Together, these results suggested that high PCSK9 expression in baseline tumor tissue was a deleterious factor for the efficacy of anti-PD-1 immunotherapy in advanced NSCLC patients. The PCSK9 inhibitor in combination with the anti-CD137 agonist could not only enhance the recruitment of CD8+ and GzmB+ CD8+ T cells but also deplete Tregs, which may be a novel therapeutic strategy for future research and clinical practice.

Cancer-Cell-Biomimetic Carbazole-Based AIE Nanoplatform for Targeted Phototheranostics of Lung Cancer

Lung cancer is one of the most diagnosed cancers and is the leading cause of cancer death. Photodynamic therapy (PDT) has been considered as a promising strategy due to its strong efficacy and negligible side effects. The development of potent PDT agents with an excellent tumor targeting capability is highly desirable but still not satisfied. In this work, we report a highly efficacious organic nanoplatform based on an aggregation-induced emission luminogen (AIEgen) and biomimetic modification for precise phototheranostics of lung cancer. An AIEgen with strong light absorption ability and bright deep red/near-infrared emission is designed and synthesized that possesses both type I and type II PDT processes. The AIEgen is encapsulated into nanoparticles (NPs) and further camouflaged with a Lewis lung carcinoma cell membrane to build a biomimetic nanoplatform. Both in vitro and in vivo experiments indicate that the cancer-cell-biomimetic NPs could significantly increase the tumor cell targeting ability and sensitively delineate the tumor site. Moreover, the cell-membrane-camouflaged NPs also show excellent antitumor efficacy in lung-cancer-bearing mice. This work demonstrates that the integration of highly efficient AIEgen and biomimetic cell membranes is able to boost the phototheranostic efficacy, representing a promising strategy for precise cancer diagnosis and treatment.

Tumor-derived microparticles-based nanomaterial as platform for delivery of tumor antigens to enhance immunogenicity

BackgroundTumor‐derived microparticles (TMPs) are not only carriers for tumor antigens but also can be used as good delivery platform. Therefore, for such an excellent delivery platform, we considered modifying TMPs with lipopolysaccharide (LPS) and polyethyleneimine (PEI) to synthesize TMPs-PEI-LPS for the delivery of tumor antigen and investigated its delivery efficiency, histocompatibility and anti-tumor effect. ResultWe investigated the bioactivity of TMPs-PEI-LPS with dendritic cells and their potential for therapeutic application to treat lung cancer in mice models bearing Lewis lung carcinoma cells. Both in vitro and in vivo assays demonstrated successful TMPs-PEI-LPS induced the uptake and maturity of dendritic cells. TMPs-PEI-LPS exhibited certain immunostimulation activity and anti-tumor effects on inhibiting tumor growth in vivo. Notably, TMPs-PEI-LPS were safe and non-toxic in vivo study. ConclusionThese results indicated that TMPs-PEI-LPS are promising platforms for delivering tumor antigens to enhance immunogenicity.

Cisplatin and Albumin-Based Gold–Cisplatin Nanoparticles Enhance Ablative Radiation Therapy–Induced Antitumor Immunity in Local and Distant Tumor Microenvironment

Ablative radiation therapy (RT) is an important strategy to eliminate primary tumor and can potentially induce the abscopal effect. Human serum albumin nanoparticle (NP) was used for controlled release of cisplatin to decrease cisplatin's systemic toxicity, and gold (Au) was added to increase RT-induced immunogenic cell death and potentiate the abscopal antitumor immunity. The designed albumin-based cisplatin-conjugated AuNPs were administered concurrently with ablative RT. C57BL/6 mice implanted with syngeneic murine Lewis lung carcinoma or murine MB49 tumor models were treated with ablative RT (12 Gy per fraction for 2 fractions, total 24 Gy), cisplatin, or Au-cisplatin NPs. Combining ablative RT with cisplatin or Au-cisplatin NPs both destroyed the primary tumor effectively and elicited immunogenic cell death accompanied by release of danger-associated molecular patterns. This enhanced recruitment of effector tumor-infiltrating immune cells, including natural killer T cells and CD8+ T cells, and elicited an increased percentage of professional antigen-presenting CD11c+ dendritic cells. Transient weight loss, accompanying hepatotoxicity, nephrotoxicity, and hematopoietic suppression, was observed as a systemic adverse event in the cisplatin but not the Au-cisplatin NPs group. Cisplatin and Au-cisplatin NPs both showed equivalent ability to reduce metastatic potential when combined with ablative RT, confirmed by suppressed unirradiated flank tumor growth and decreased metastatic lung tumor burden, which translated to improved survival. Mobilization and abundance of effector tumor-infiltrating immune cells including CD8+ T cells and dendritic cells were observed in the distant lung tumor microenvironment after ablative RT with cisplatin or Au-cisplatin NPs, demonstrating increased antitumor immunotherapeutic activity as an abscopal effect. Compared with cisplatin, the albumin-based Au-cisplatin NPs exhibited equivalent but no superior antitumor immunotherapeutic activity while reducing systemic adverse events and can be safely administered concurrently with ablative RT. Alternative NP formulations may be designed to further improve anticancer outcomes.

Cancer aggravation due to persistent pain signals with the increased expression of pain-related mediators in sensory neurons of tumor-bearing mice

A growing body of evidence suggests that intractable pain reduces both the quality of life and survival in cancer patients. In the present study, we evaluated whether chronic pain stimuli could directly affect cancer pathology using tumor-bearing mice. For this purpose, we used two different models of chronic pain in mice, neuropathic pain and persistent postsurgical pain, with Lewis lung carcinoma (LLC) as tumor cells. We found that tumor growth was dramatically promoted in these pain models. As well as these pain models, tumor growth of LLC, severe osteosarcoma (AXT) and B16 melanoma cells was significantly promoted by concomitant activation of sensory neurons in AAV6-hM3Dq-injected mice treated with the designer drug clozapine-N-oxide (CNO). Significant increases in mRNA levels of vascular endothelial growth factor-A (Vegfa), tachykinin precursor 1 (Tac1) and calcitonin-related polypeptide alpha (Calca) in the ipsilateral side of dorsal root ganglion of AAV6-hM3Dq-injected mice were observed by concomitant activation of sensory neurons due to CNO administration. Moreover, in a model of bone cancer pain in which mice were implanted with AXT cells into the right femoral bone marrow cavity, the survival period was significantly prolonged by repeated inhibition of sensory neurons of AAV6-hM4Di-injected mice by CNO administration. These findings suggest that persistent pain signals may promote tumor growth by the increased expression of sensory-located peptides and growth factors, and controlling cancer pain may prolong cancer survival.

Influence of induced diabetes mellitus on hormonal profile of Lewis lung carcinoma in BALB/c Nude mice

Purpose of the study. The assessment of diabetes mellitus (DM) effect on levels of sex hormones in tumor and peritumoral tissues in BALB/c Nude mice with Lewis lung carcinoma (LLC).Materials and methods. The study included 42 male and female BALB/c Nude mice aged 8–9 weeks weighing 21–22 g. Alloxan-induced DM was reproduced in mice of the main group, and then LLC was transplanted. Levels of estrone (E1), estradiol (E2), testosterone (T), progesterone (P4) and prolactin (PRL), as well as steroid hormone receptors: estrogens (REα, REβ), androgens (RA), and progesterone (RP4) were measured by RIA and ELISA in samples of tumor and peritumoral tissues. Animals with LLC without DM were used as controls. The statistical analysis was performed using the Statistica 10 program; differences were considered significant at p < 0.05.Results. DM in males was reproduced only after a double injection of alloxan, and was characterized by lower blood glucose levels compared to females. The growth of LLC in animals with alloxan-induced DM was possible only in female BALB/c Nude mice; in BALB/c Nude males, the tumor could not be transplanted either independently or in combination with DM. Females in the main group showed greater average tumor volumes throughout the experiment and reduced survival, compared to the control group. Tumor samples from females with LLC+DM were more saturated with sex steroids, but depleted in steroid hormone receptors, which probably contributed to the ability to avoid the body's regulatory signals.Conclusion. The growth of LLC in presence of induced DM was sex-dependent, since the tumor could not be transplanted to male mice. DM affected the levels of sex steroids and their receptors tumor tissues in female BALB/c Nude mice.

Myeloid-cell-specific role of Gasdermin D in promoting lung cancer progression in mice

The activities of the NLRP3 and AIM2 inflammasomes and Gasdermin D (GsdmD) are implicated in lung cancer pathophysiology but it's not clear if their contributions promote or retard lung cancer progression. Using a metastatic Lewis lung carcinoma (LLC) cell model, we show that GsdmD knockout (GsdmD-/-) mice form significantly fewer cancer foci in lungs, exhibit markedly decreased lung cancer metastasis, and show a significant ∼50% increase in median survival rate. The cleaved forms of GsdmD and IL-1β were detected in lung tumor tissue, indicating inflammasome activity in lung tumor microenvironment (TME). Increased migration and growth of LLC cells was observed upon exposure to the conditioned media derived from inflammasome-induced wild type, but not the GsdmD-/-, macrophages. Using bone marrow transplantations, we show a myeloid-specific contribution of GsdmD in lung cancer metastasis. Taken together, our data show that GsdmD plays a myeloid-specific role in lung cancer progression.

Oral Anticancer Heterobimetallic PtIV−AuI Complexes Show High In Vivo Activity and Low Toxicity

AbstractAuI‐carbene and PtIV−AuI‐carbene prodrugs display low to sub‐μM activity against several cancer cell lines and overcome cisplatin (cisPt) resistance. Linking a cisPt‐derived PtIV(phenylbutyrate) complex to a AuI‐phenylimidazolylidene complex 2, yielded the most potent prodrug. While in vivo tests against Lewis Lung Carcinoma showed that the prodrug PtIV(phenylbutyrate)‐AuI‐carbene (7) and the 1 : 1 : 1 co‐administration of cisPt: phenylbutyrate:2 efficiently inhibited tumor growth (≈95 %), much better than 2 (75 %) or cisPt (84 %), 7 exhibited only 5 % body weight loss compared to 14 % for 2, 20 % for cisPt and >30 % for the co‐administration. 7 was much more efficient than 2 at inhibiting TrxR activity in the isolated enzyme, in cells and in the tumor, even though it was much less efficient than 2 at binding to selenocysteine peptides modeling the active site of TrxR. Organ distribution and laser‐ablation (LA)‐ICP‐TOFMS imaging suggest that 7 arrives intact at the tumor and is activated there.

Oral Anticancer Heterobimetallic PtIV -AuI Complexes Show High In Vivo Activity and Low Toxicity.

AuI -carbene and PtIV -AuI -carbene prodrugs display low to sub-μM activity against several cancer cell lines and overcome cisplatin (cisPt) resistance. Linking a cisPt-derived PtIV (phenylbutyrate) complex to a AuI -phenylimidazolylidene complex 2, yielded the most potent prodrug. While in vivo tests against Lewis Lung Carcinoma showed that the prodrug PtIV (phenylbutyrate)-AuI -carbene (7) and the 1 : 1 : 1 co-administration of cisPt: phenylbutyrate:2 efficiently inhibited tumor growth (≈95 %), much better than 2 (75 %) or cisPt (84 %), 7 exhibited only 5 % body weight loss compared to 14 % for 2, 20 % for cisPt and >30 % for the co-administration. 7 was much more efficient than 2 at inhibiting TrxR activity in the isolated enzyme, in cells and in the tumor, even though it was much less efficient than 2 at binding to selenocysteine peptides modeling the active site of TrxR. Organ distribution and laser-ablation (LA)-ICP-TOFMS imaging suggest that 7 arrives intact at the tumor and is activated there.

Inhibition of STAT6 with Antisense Oligonucleotides Enhances the Systemic Antitumor Effects of Radiotherapy and Anti-PD-1 in Metastatic Non-Small Cell Lung Cancer.

Diverse factors contribute to the limited clinical response to radiotherapy (RT) and immunotherapy in metastatic non-small cell lung cancer (NSCLC), among which is the ability of these tumors to recruit a retinue of suppressive immune cells-such as M2 tumor-associated macrophages (TAM)-thereby establishing an immunosuppressive tumor microenvironment that contributes to tumor progression and radio resistance. M2 TAMs are activated by the STAT6 signaling pathway. Therefore, we targeted STAT6 using an antisense oligonucleotide (ASO) along with hypofractionated RT (hRT; 3 fractions of 12 Gy each) to primary tumors in three bilateral murine NSCLC models (Lewis lung carcinoma, 344SQ-parental, and anti-PD-1-resistant 344SQ lung adenocarcinomas). We found that STAT6 ASO plus hRT slowed growth of both primary and abscopal tumors, decreased lung metastases, and extended survival. Interrogating the mechanism of action showed reduced M2 macrophage tumor infiltration, enhanced TH1 polarization, improved T-cell and macrophage function, and decreased TGFβ levels. The addition of anti-PD-1 further enhanced systemic antitumor responses. These results provide a preclinical rationale for the pursuit of an alternative therapeutic approach for patients with immune-resistant NSCLC.

Feiyiliu Mixture sensitizes EGFRDel19/T790M/C797S mutant non-small cell lung cancer to osimertinib by attenuating the PRC1/Wnt/EGFR pathway.

Introduction: Osimertinib is a potent epidermal growth factor receptor tyrosine kinase inhibitor (EGFR-TKI) for the treatment of patients with EGFR-mutant non-small cell lung cancer (NSCLC). However, the emergence of acquired resistance due to the EGFR-Del19/T790M/C797S mutation limits the clinical application of osimertinib. Feiyiliu Mixture (FYLM), a clinical experience formula of Chinese medicine, was used to treat lung cancer with good clinical efficacy. In this study, we aimed to investigate the mechanism by which Feiyiliu Mixture delays osimertinib resistance in EGFR-mutant cell lines and EGFR-mutant cell tumor-bearing mice. Methods: The osimertinib-resistant cell models were established in mouse Lewis lung carcinoma (LLC) cells transfected with EGFR-Del19/T790M/C797S mutant lentivirus. In cell experiments, after 48h of treatment with Feiyiliu Mixture-containing serum, MTT assay was used to detect the relative cell viability, and western blotting was used to detect EGFR protein phosphorylation expression. In animal experiments, C57BL/6J mice were subcutaneously injected with Lewis lung carcinoma cells stably expressing EGFR-Del19/T790M/C797S mutations to construct a xenograft model. After 2weeks of Feiyiliu Mixture and/or osimertinib treatment, the expression of proliferation-related, apoptosis-related and PRC1/Wnt/EGFR pathway markers was detected by real-time qPCR, western blotting and immunohistochemistry. Results: The results showed that when combined with osimertinib, Feiyiliu Mixture synergistically reduces proliferation and increases apoptosis to improve drug resistance. In vitro, Feiyiliu Mixture-containing serum reduced the EGFR phosphorylation. In vivo, Feiyiliu Mixture downregulated the expression of cyclin B1 and Bcl-2 while upregulating the level of cleaved Caspase-3 protein, indicating that Feiyiliu Mixture promotes apoptosis. Furthermore, Feiyiliu Mixture reduced the expression of p-EGFR, p-Akt, PRC1 and Wnt pathway-related proteins such as β-catenin, c-Myc and c-Jun. Conclusion: The present study identified that Feiyiliu Mixture inhibited PRC1/Wnt/EGFR pathway activation, reduced proliferation, and promoted apoptosis, thereby increasing the sensitivity of EGFR-mutant non-small cell lung cancer to osimertinib. Our study provided a new idea for Chinese medicine to play a role in enhancing efficacy and reducing toxicity in the treatment of non-small cell lung cancer.