Drug-resistant Lung Cancer Research Articles

APOBEC Mutagenesis Activity Confers Drug Resistance in Lung Cancer.

Tyrosine kinase inhibitors induce APOBEC3A, which promotes the emergence of drug-tolerant persister clones.

Cytosolic malic enzyme and glucose-6-phosphate dehydrogenase modulate redox balance in NSCLC with acquired drug resistance.

Lung cancer cells often show elevated levels of reactive oxygen species (ROS) and nicotinamide adenine dinucleotide phosphate (NADPH). However, the connections between deregulated redox homeostasis in different subtypes of lung cancer and acquired drug resistance in lung cancer have not yet been fully established. Herein, we analyzed different subtypes of lung cancer data reported in the Cancer Cell Line Encyclopedia (CCLE) database, the Cancer Genome Atlas program (TCGA), and the sequencing data obtained from a gefitinib-resistant non-small-cell lung cancer (NSCLC) cell line (H1975GR). Using flux balance analysis (FBA) model integrated with multiomics data and gene expression profiles, we identified cytosolic malic enzyme 1 (ME1) and glucose-6-phosphate dehydrogenase as the major contributors to the significantly upregulated NADPH flux in NSCLC tissues as compared with normal lung tissues, and gefitinib-resistant NSCLC cell line as compared with the parental cell line. Silencing the gene expression of either of these two enzymes in two osimertinib-resistant NSCLC cell lines (H1975OR and HCC827OR) exhibited strong antiproliferative effects. Our findings not only underscored the pivotal roles of cytosolic ME1 and glucose-6-phosphate dehydrogenase in regulating redox states in NSCLC cells but also provided novel insights into their potential roles in drug-resistant NSCLC cells with disturbed redox states.

The role of alternative splicing in lung cancer.

Aberrant alternative splicing (AS) events are frequently observed in lung cancer, which can be attributed to aberrant gene AS, alterations in splicing regulatory factors, or changes in splicing regulatory mechanisms. Consequently, the dysregulation of alternative RNA splicing is the fundamental cause of lung cancer. In this review, we have summarized the pivotal role of AS in the development, progression, invasion, metastasis, angiogenesis, and drug resistance of lung cancer. Ultimately, this review emphasizes the potential of AS as biomarkers in lung cancer prognosis and diagnosis, and introduces some applications of AS isoform in the treatment of lung cancer. The comprehension of the AS may provide a glimmer of hope for the eradication of lung cancer.

Isoorientin reverses lung cancer drug resistance by promoting ferroptosis via the SIRT6/Nrf2/GPX4 signaling pathway

Cisplatin, or DDP, is a highly successful and well-known chemotherapy drug used to treat cancer. Acquired resistance to chemotherapy is a major clinical concern, yet the mechanisms of this resistance are still unknown. Ferroptosis is a type of cell death distinct from other forms, fueled by a buildup of iron-associated lipid reactive oxygen species (ROS). Gaining insight into the process of ferroptosis could lead to novel treatments for overcoming cancer resistance. In this study, the combination of isoorientin (IO) and DDP treatment resulted in a significant decrease in the viability of drug-resistant cells, a substantial increase in intracellular iron, malondialdehyde (MDA) and ROS concentrations, a notable decrease in glutathione concentration, and the occurrence of ferroptosis in cells, as revealed by in vitro and in vivo experiments. Additionally, there was a decrease in the expression of nuclear factor-erythroid factor 2-related factor 2 (Nrf2), glutathione peroxidase 4 (GPX4), and sirtuin 6 (SIRT6) proteins, and an increase in cellular ferroptosis. Isoorientin acts as a mediator to regulate cellular ferroptosis and reverse drug resistance in lung cancer cells by controlling the SIRT6/Nrf2/GPX4 signaling pathway. The findings of this study suggest that IO can promote ferroptosis and reverse drug resistance in lung cancer through the SIRT6/Nrf2/GPX4 signaling pathway, thus offering a theoretical basis for its potential clinical application.

Cell-in-cell promotes lung cancer malignancy by enhancing glucose metabolism through mitochondria transfer

Heterotypic cell-in-cell structure (CICs) is the definition of the entry of one type of living cells into another type of cell. CICs between immune cells and tumor cells have been found to correlate with malignancy in many cancers. Since tumor immune microenvironment promotes non-small cell lung cancer (NSCLC) progression and drug resistance, we wondered the potential significance of heterotypic CICs in NSCLC. Heterotypic CICs was analyzed by histochemistry in an expanded spectrum of clinical lung cancer tissue specimens. In vitro study was performed using the mouse lung cancer cell line LLC and splenocytes. Our results revealed that CICs formed by lung cancer cells and infiltrated lymphocytes were correlated with malignancy of NSCLC. In addition, we found CICs mediated the transfer of lymphocyte mitochondria to tumor cells, and promoted cancer cell proliferation and anti-cytotoxicity by activating MAPK pathway and up-regulating PD-L1 expression. Furthermore, CICs induces glucose metabolism reprogramming of lung cancer cells by upregulating glucose intake and glycolytic enzyme. Our findings suggest that CICs formed by lung cancer cell and lymphocyte contribute to NSCLC progression and reprogramming of glucose metabolism, and might represent a previously undescribed pathway for drug resistance of NSCLC.

Phosphorylation of Bcl-2 by JNK confers gemcitabine resistance in lung cancer cells by reducing autophagy-mediated cell death.

The most common cancer-related death in the world is non-small cell lung cancer (NSCLC). Gemcitabine (GEM) is a common and effective first-line chemotherapeutic drug for the treatment of NSCLC. However, the long-term use of chemotherapeutic drugs in patients usually induces cancer cell drug resistance, leading to poor survival, and prognosis. In this study, to observe and explore the key targets and potential mechanisms of NSCLC resistance to GEM, we first cultured lung cancer CL1-0 cells in a GEM-containing medium to induce CL1-0 cells to develop GEM resistance. Next, we compared protein expression between the parental and GEM-R CL1-0 cell groups. We observed significantly lower expression of autophagy-related proteins in GEM-R CL1-0 cells than in parental CL1-0 cells, indicating that autophagy is associated with GEM resistance in CL1-0 cells. Furthermore, a series of autophagy experiments revealed that GEM-R CL1-0 cells had significantly reduced GEM-induced c-Jun N-terminal kinase phosphorylation, which further affected the phosphorylation of Bcl-2, thereby reducing the dissociation of Bcl-2 and Beclin-1 and ultimately reducing the generation of GEM-induced autophagy-dependent cell death. Our findings suggest that altering the expression of autophagy is a promising therapeutic option for drug-resistant lung cancer.

3-Deoxysappanchalcone Inhibits Cell Growth of Gefitinib-Resistant Lung Cancer Cells by Simultaneous Targeting of EGFR and MET Kinases.

The mechanistic functions of 3-deoxysappanchalcone (3-DSC), a chalcone compound known to have many pharmacological effects on lung cancer, have not yet been elucidated. In this study, we identified the comprehensive anti-cancer mechanism of 3-DSC, which targets EGFR and MET kinase in drug-resistant lung cancer cells. 3-DSC directly targets both EGFR and MET, thereby inhibiting the growth of drug-resistant lung cancer cells. Mechanistically, 3-DSC induced cell cycle arrest by modulating cell cycle regulatory proteins, including cyclin B1, cdc2, and p27. In addition, concomitant EGFR downstream signaling proteins such as MET, AKT, and ERK were affected by 3-DSC and contributed to the inhibition of cancer cell growth. Furthermore, our results show that 3-DSC increased redox homeostasis disruption, ER stress, mitochondrial depolarization, and caspase activation in gefitinib-resistant lung cancer cells, thereby abrogating cancer cell growth. 3-DSC induced apoptotic cell death which is regulated by Mcl-1, Bax, Apaf-1, and PARP in gefitinib-resistant lung cancer cells. 3-DSC also initiated the activation of caspases, and the pan-caspase inhibitor, Z-VAD-FMK, abrogated 3-DSC induced-apoptosis in lung cancer cells. These data imply that 3-DSC mainly increased mitochondria-associated intrinsic apoptosis in lung cancer cells to reduce lung cancer cell growth. Overall, 3-DSC inhibited the growth of drug-resistant lung cancer cells by simultaneously targeting EGFR and MET, which exerted anti-cancer effects through cell cycle arrest, mitochondrial homeostasis collapse, and increased ROS generation, eventually triggering anticancer mechanisms. 3-DSC could potentially be used as an effective anti-cancer strategy to overcome EGFR and MET target drug-resistant lung cancer.

EML4-ALK biology and drug resistance in non-small cell lung cancer: a new phase of discoveries.

Anaplastic lymphoma kinase (ALK) can be driven to oncogenic activity by different types of mutational events such as point-mutations, for example F1174L in neuroblastoma, and gene fusions, for example with echinoderm microtubule-associated protein-like 4 (EML4) in non-small cell lung cancer (NSCLC). EML4-ALK variants result from different breakpoints, generating fusions of different sizes and properties. The most common variants (Variant 1 and Variant 3) form cellular compartments with distinct physical properties. The presence of a partial, probably misfolded beta-propeller domain in variant 1 confers solid-like properties to the compartments it forms, greater dependence on Hsp90 for protein stability and higher cell sensitivity to ALK tyrosine kinase inhibitors (TKIs). These differences translate to the clinic because variant 3, on average, worsens patient prognosis and increases metastatic risk. Latest generation ALK-TKIs are beneficial for most patients with EML4-ALK fusions. However, resistance to ALK inhibitors can occur via point-mutations within the kinase domain of the EML4-ALK fusion, for example G1202R, reducing inhibitor effectiveness. Here, we discuss the biology of EML4-ALK variants, their impact on treatment response, ALK-TKI drug resistance mechanisms and potential combination therapies.

Genome-directed discovery of antiproliferative bafilomycins from a deepsea-derived Streptomyces samsunensis

Genomic bioinformatics analysis identified a bafilomycin biosynthetic gene cluster (named bfl) in the deepsea-derived S. samsunensis OUCT16-12, from which two new (1 and 2, named bafilomycins R and S) along with four known (3–6) bafilomycins were targetly obtained. The structure of 3 was clearly identified for the first time, thus named bafilomycin T herein. Differ from the fumarate substitution at C-21 of known bafilomycins, its location on C-23 is a unique feature of 1 and 2. The stereochemistry of the compounds was established based on NOE correlations, ketoreductase (KR)-types in PKS modules of bfl, and ECD calculations. Moreover, a detailed biosynthetic model of 1–6 in S. samsunensis OUCT16-12 was provided based on the gene function prediction and sequence identity. Compared with the positive control doxorubicin, 1–6 showed more potent antiproliferative activities against drug-resistant lung cancer cell line A549-Taxol, with IC50 values ranging from 0.07 μM to 1.79 μM, which arrested cell cycle in G0/G1 phase to hinder proliferation.

N6-methyladenine-mediated aberrant activation of the lncRNA SOX2OT-GLI1 loop promotes non-small-cell lung cancer stemness

Despite the advent of precision medicine and immunotherapy, mortality due to lung cancer remains high. The sonic hedgehog (SHH) cascade and its key terminal factor, glioma-associated oncogene homolog 1 (GLI1), play a pivotal role in the stemness and drug resistance of lung cancer. Here, we investigated the molecular mechanism of non-canonical aberrant GLI1 upregulation. The SHH cascade was upregulated in stem spheres and chemo-resistant lung cancer cells and was accountable for drug resistance against multiple chemotherapy regimens. GLI1 and the long non-coding RNA SOX2OT were positively regulated, and the GLI1-SOX2OT loop mediated the proliferation of parental and stem-like lung cancer cells. Further mechanistic investigation revealed that SOX2OT facilitated METTL3/14/IGF2BP2-mediated m6A modification and stabilization of the GLI1 mRNA. Additionally, SOX2OT upregulated METTL3/14/IGF2BP2 by sponging miR-186-5p. Functional analysis corroborated that GLI1 acted as a downstream target of METTL3/14/IGF2BP2, and GLI1 silencing could block the oncogenicity of lung cancer stem-like cells. Pharmacological inhibition of the loop remarkably inhibited the oncogenesis of lung cancer cells in vivo. Compared with paired adjacent normal tissues, lung cancer specimens exhibited consistently upregulated GLI1/SOX2OT/METTL3/14/IGF2BP2. The m6A-modified GLI1-SOX2OT loop may serve as a potential therapeutic target and prognostic predictor for lung cancer therapy and diagnosis in the clinic.

M6A-mediated upregulation of lncRNA-AC026356.1 promotes cancer stem cell maintenance in lung adenocarcinoma via activating Wnt signaling pathway.

LncRNA plays a pivotal role in the stemness and drug resistance of lung cancer. Here, we found that lncRNA-AC026356.1 was upregulated in stem spheres and chemo-resistant lung cancer cells. Our fish assay also shows that AC026356.1 was predominantly located in the cytoplasm of lung cancer cells and does not have protein-coding potential. Silencing AC026356.1 significantly inhibited proliferation and migration but increased apoptosis in A549-cisplatin (DDP) cells. Additionally, IGF2BP2 and the lncRNA-AC026356.1 positively regulated the proliferation and stemness of stem-like lung cancer cells. Further mechanistic investigation revealed that METTL14/IGF2BP2-mediated m6A modification and stabilization of the AC026356.1 RNA. Functional analysis corroborated that AC026356.1 acted as a downstream target of METTL14/IGF2BP2 and AC026356.1 silencing could block the oncogenicity of lung cancer stem-like cells. AC026356.1 expression was correlated with immune cell infiltration and T cell exhaustion. Compared with paired adjacent normal tissues, lung cancer specimens exhibited consistently upregulated METTL14/IGF2BP2/AC026356.1. M6A-modified METTL14/IGF2BP2/AC026356.1 loop may serve as a potential therapeutic target and prognostic predictor for lung cancer therapy and diagnosis in the clinic.

In vitro differences analysis in proliferation and drug sensitivity of tumor-associated fibroblasts from different regions of FDG metabolism to non-small-cell lung carcinoma

Objective: 18F-FDG PET can directly reflect the glucose metabolism level of cells from the molecular level. Tumor-associated fibroblasts, as the natural physiological barrier of tumors, support and protect the survival of non-small cell lung cancer cells. This study will further observe the differences in the supporting effects of CAFs from different regions of 18F-FDG PET metabolism on cancer cells. Methods: Biopsy tissues were collected from PET/CT patients diagnosed with NSCLC for the first time in the Department of Nuclear Medicine, The First Hospital of Shanxi Medical University. The locations of puncture were all selected in areas with high metabolism of tumor. Primary CAFs were cultured by tissue adhesion method. A549 acts as EGFR (-) tumor cells, while PC9 acts as EGFR (+) tumor cells. CAFs conditional medium was collected to induce the culture of A549 and PC9 cells, labeled as A549high/low and PC9high/low, respectively. CCK was used to measure cancer cell proliferation rate, Transwell was used to measure cancer cell invasiveness, and scratch assay was used to measure cancer cell migration. The cells were treated with cisplatin and gefitinib respectively to observe the resistance of the cells. Results: Tissues of two NSCLC patients with SUVmax were 1.25 and 11.15, respectively. The conditioned medium induced cancer cells for 48 hours. CCK (48h) results showed that SUVhigh-derived CAFs could promote the proliferation of two types of cancer cells more than SUVlow-derived CAFs. The wound healing experiment showed that A549high had completed the healing at 72h, followed by low metabolism area, and A549 was the worst. The results for PC9 cells were similar to those for A549. Transwell experiment showed that PC9high/ low showed obvious difference in invasiveness at 24 hours, and the difference was more significant at 48 hours. A549 also showed difference in invasiveness, but the difference was relatively weak. When the corresponding cells were further treated with cisplatin or gefitinib, the proliferation rate of A549high/low cells was significantly different in the gradient range of high cisplatin concentration, while the proliferation rate of PC9high/low cells was significantly different in the low-medium gefitinib concentration. Conclusion: The regions of FDG metabolism difference shown in PET support the reverse warburg effect. As a physiological barrier to tumors, CAFs is one of the key factors that promote the occurrence, metastasis and drug resistance of lung cancer. National Natural Science Foundation of China (Grant number:81971655) This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Wee1 epigenetically modulates H2B mono-ubiquitination at K120 lysine and DNA double-strand break repair through phosphorylation of H2BY37-dependent manner in small-cell lung cancer.

DNA damage repair is a crucial mechanism highly related to therapy resistance for various therapeutic strategies. Our previous results have shown that the degree of drug resistance in small-cell lung cancer (SCLC) cell lines was proportional to both the transcription and expression levels of Wee1, indicating that Wee1, an evolutionarily highly conserved kinase, plays a vital role in the therapeutic resistance of SCLC. In the present study, we aim to determine the nonclassical mechanism of Wee1 on DNA repair regulation. Western blot was conducted to determine the mono-ubiquitination level of H2Bub. Comet assay was used to evaluate the degree of DNA damage. Immunofluorescence was conducted to determine the DNA repair markers. Co-immunoprecipitation was utilized to assess the potential interactions with H2BY37ph. MTT assays were used to evaluate the survival rates of SCLC cells. Overexpression of Wee1 increases the level of H2BK120ub and alleviates ionizing radiation (IR)-induced DNA damage in SCLC cells. Moreover, H2BK120ub is a crucial molecule in Wee1-mediated double-strain break (DSB) repair in SCLC cells. Mechanisms study indicated that H2BY37ph is involved in Wee1-mediated H2BK120ub through interaction with the E3 ubiquitin ligase RNF20-RNF40 complex and upregulates its phosphorylation, mutation of H2BY37 phosphorylation sites attenuated DSB repair and enhanced the sensitivity of IR-induced SCLC cell death. H2BY37ph produces crosstalk with H2BK120ub in an E3 ubiquitin ligase-dependent manner, promoting Wee1-mediated DSB repair in SCLC cells. This study clarifies the nonclassical mechanism of Wee1 regulation of DSB repair, which provides a theoretical basis for the clinical understanding of the regulatory network of Wee1 and its use as a target for overcoming multiple types of therapeutic resistance.

Abstract 5845: Periostin secreted by cancer-associated fibroblasts promotes cancer progression and drug resistance in non-small cell lung cancer

Abstract Background: Cancer-associated fibroblasts (CAFs) are key players in the tumor microenvironment (TME) due to their abundance in most solid tumors and active crosstalk with cancer cells, and promote the initiation of tumor formation, tumor growth, angiogenesis, metastasis, and therapeutic resistance. In this study, we sought to identify effective therapeutic targets in CAFs for non-small cell lung cancer (NSCLC). Experimental Design: We established fibroblast cell lines from the cancerous and non-cancerous parts of surgical lung specimens from patients with NSCLC and evaluated the differences in behaviors towards NSCLC cells using in vitro co-culture models, conditioned medium, and in vivo xenograft mouse models. Next, RNA sequencing analysis was performed to investigate the differentially expressed genes between normal fibroblasts (NFs) and CAFs, and we then verified the biological activity of the identified molecule on NSCLC cells in vitro and in vivo mouse model. Results: Cancer cells showed enhanced cell proliferation, migration, and drug resistance by co-culture with CAFs or CAF-derived conditioned medium. RNA sequencing analysis revealed that CAFs showed higher expressions of POSTN, a matricellular protein, than NFs. Single-cell RNA sequencing data of NSCLC from public database confirmed that POSTN-positive fibroblasts were highly enriched in lung tumor tissues but rarely observed in normal lung tissues, suggesting CAF-specific POSTN expression in lung cancer. Recombinant POSTN increased cell proliferation via NSCLC cells’ ERK pathway activation and induced epithelial-mesenchymal transition (EMT), which improved migration in vitro. In addition, POSTN knockdown in CAFs suppressed these effects, and in vivo experiments demonstrated that the POSTN knockdown improved the sensitivity of EGFR-mutant NSCLC cells for Osimertinib treatment. Conclusion: Our results showed that CAF-derived POSTN is involved in tumor growth, migration, EMT induction, and drug resistance in NSCLC. Targeting CAF-secreted POSTN in TME could be a potential therapeutic strategy for NSCLC. Citation Format: Mao Yoshikawa, Fumiaki Takatsu, Ken Suzawa, Tomohiro Habu, Ohki Masayoshi, Kazuma Iwata, Naoki Matsuda, Yin Min Thu, Kazuhiko Shien, Hiromasa Yamamoto, Shinichi Toyooka. Periostin secreted by cancer-associated fibroblasts promotes cancer progression and drug resistance in non-small cell lung cancer. [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 5845.

Targeting Rab26 to Conquer Cisplatin-Resistant Lung Cancer with Self-Assembled DNA Nanomaterials

Overcoming cisplatin-based drug resistance in lung cancer remains an enormous challenge in clinical tumor therapy worldwide. Recent studies have reported that some Rab GTPases are involved in multiple aspects of tumor progression, including invasion, migration, metabolism, autophagy, exosome secretion, and drug resistance. In particular, Rab26 is essential to vital processes such as vesicle-mediated secretion, cell growth, apoptosis, and autophagy. In this study, we developed a nanosystem based on programmed DNA self-assembly of Rab26 siRNA-loaded nanoparticles (siRNP). We demonstrated that siRNP could be effectively transfected into cisplatin-resistant A549 (A549/DDP) cells. These siRab26-carrying nanoparticles induced apoptosis and inhibited the disruption of autophagy. The combination therapy of siRab26 knockdown with cisplatin could improve the antitumor therapy compared with a single one in vitro. In nude mice, siRNP enhanced the chemosensitivity of cisplatin-resistant cells and inhibited tumor xenograft development. These outcomes suggest that siRNP is an effective platform for lung cancer therapy in cases exhibiting drug resistance.

Discovery of Novel EGFR Inhibitor Targeting Wild-Type and Mutant Forms of EGFR: In Silico and In Vitro Study.

Targeting L858R/T790M and L858R/T790M/C797S mutant EGFR is a critical challenge in developing EGFR tyrosine kinase inhibitors to overcome drug resistance in non-small cell lung cancer (NSCLC). The discovery of next-generation EGFR tyrosine kinase inhibitors (TKIs) is therefore necessary. To this end, a series of furopyridine derivatives were evaluated for their EGFR-based inhibition and antiproliferative activities using computational and biological approaches. We found that several compounds derived from virtual screening based on a molecular docking and solvated interaction energy (SIE) method showed the potential to suppress wild-type and mutant EGFR. The most promising PD13 displayed strong inhibitory activity against wild-type (IC50 of 11.64 ± 1.30 nM), L858R/T790M (IC50 of 10.51 ± 0.71 nM), which are more significant than known drugs. In addition, PD13 revealed a potent cytotoxic effect on A549 and H1975 cell lines with IC50 values of 18.09 ± 1.57 and 33.87 ± 0.86 µM, respectively. The 500-ns MD simulations indicated that PD13 formed a hydrogen bond with Met793 at the hinge region, thus creating excellent EGFR inhibitory activity. Moreover, the binding of PD13 in the hinge region of EGFR was the major determining factor in stabilizing the interactions via hydrogen bonds and van der Waals (vdW). Altogether, PD13 is a promising novel EGFR inhibitor that could be further clinically developed as fourth-generation EGFR-TKIs.

Ethanolic Extract of Artemisia vulgaris Leaf Promotes Apoptotic Cell Death in Non-Small-Cell Lung Carcinoma A549 Cells through Inhibition of the Wnt Signaling Pathway.

The Wnt signaling pathway is reported to be associated with lung cancer progression, metastasis and drug resistance, and thus it is an important therapeutic target for lung cancer. Plants have been shown as reservoirs of multiple potential anticancer agents. In the present investigation, the ethanolic leaf extract of Artemisia vulgaris (AvL-EtOH) was initially analyzed by means of gas chromatography-mass spectrometry (GC-MS) to identify the important phytochemical constituents. The GC-MS analysis of AvL-EtOH exhibited 48 peaks of various secondary metabolites such as terpenoids, flavonoids, carbohydrates, coumarins, amino acids, steroids, proteins, phytosterols, and diterpenes. It was found that the treatment with increasing doses of AvL-EtOH suppressed the proliferation and migration of lung cancer cells. Furthermore, AvL-EtOH induced prominent nuclear alteration along with a reduction in mitochondrial membrane potential and increased ROS (reactive oxygen species) generation in lung cancer cells. Moreover, AvL-EtOH-treated cells exhibited increased apoptosis, demonstrated by the activation of caspase cascade. AvL-EtOH also induced downregulation of Wnt3 and β-catenin expression along with cell cycle protein cyclin D1. Thus, the results of our study elucidated the potential of bioactive components of Artemisia vulgaris in the therapeutic management of lung cancer cells.

Regorafenib induces NOX5-mediated endoplasmic reticulum stress and potentiates the anti-tumor activity of cisplatin in non-small cell lung cancer cells

Lung cancer is one of the most commonly diagnosed cancers worldwide. Although cisplatin-based chemotherapy regimens serve a pivotal role in non-small cell lung cancer (NSCLC) treatment, drug resistance and serious side effects limited its further clinical application. Regorafenib, a small-molecule multi-kinase inhibitor, was demonstrated to have promising anti-tumor activity in various solid tumors. In the present study, we found that regorafenib markedly enhanced cisplatin-induced cytotoxicity in lung cancer cells by activating reactive oxygen species (ROS)-mediated endoplasmic reticulum stress (ER Stress), c-Jun N-terminal kinase (JNK) and p38 mitogen-activated protein kinase (MAPK) signaling pathways. Regorafenib increased ROS generation by promoting NADPH oxidase 5 (NOX5) expression, and knocking down NOX5 attenuated ROS-mediated cytotoxicity of regorafenib in lung cancer cells. Additionally, mice xenograft model validated that synergistic anti-tumor effects of combined treatment with regorafenib and cisplatin. Our results suggested that combination therapy with regorafenib and cisplatin may serve as a potential therapeutic strategy for some NSCLC patients.

Tp53 haploinsufficiency is involved in hotspot mutations and cytoskeletal remodeling in gefitinib-induced drug-resistant EGFRL858R-lung cancer mice

Tumor heterogeneity is the major factor for inducing drug resistance. p53 is the major defender to maintain genomic stability, which is a high proportion mutated in most of the cancer types. In this study, we established in vivo animal models of gefitinib-induced drug-resistant lung cancer containing EGFRL858R and EGFRL858R*Tp53+/− mice to explore the molecular mechanisms of drug resistance by studying the genomic integrity and global gene expression. The cellular morphology of the lung tumors between gefitinib-induced drug-resistant mice and drug-sensitive mice were very different. In addition, in drug-resistant mice, the expression of many cytoskeleton-related genes were changed, accompanied by decreased amounts of actin filaments and increased amounts of microtubule, indicating that significant cytoskeletal remodeling is induced in gefitinib-induced drug-resistant EGFRL858R and EGFRL858R*Tp53+/− lung cancer mice. The gene expression profiles and involved pathways were different in gefitinib-sensitive, gefitinib-resistant and Tp53+/−-mice. Increases in drug resistance and nuclear size (N/C ratio) were found in EGFRL858R*Tp53+/− drug-resistant mice. Mutational hotspot regions for drug resistance via Tp53+/+- and Tp53+/−-mediated pathways are located on chromosome 1 and chromosome 11, respectively, and are related to prognosis of lung cancer cohorts. This study not only builds up a gefitinib-induced drug-resistant EGFRL858R lung cancer animal model, but also provides a novel mutation profile in a Tp53+/+- or Tp53+/−-mediated manner and induced cytoskeleton remodeling during drug resistance, which could contribute to the prevention of drug resistance during cancer therapy.

Abstract P2-24-01: The development of covalent microtubule stabilizers for drug-resistant cancers

Abstract Microtubule targeting agents (MTAs) are some of the most commonly used and effective chemotherapeutic agents used in the treatment of breast cancer and other solid tumors. Clinically approved MTAs include microtubule destabilizers, such as the vinca alkaloids or eribulin, as well as the taxane class of microtubule stabilizers, including paclitaxel. While MTAs are effective chemotherapeutics, the development of resistance due to drug efflux or changes in β-tubulin isotype expression are major clinical limitations to each the currently approved drugs of this class. One strategy for overcoming drug resistance is the development of drugs that covalently bind to their drug target, which has been validated in the clinic with the success of irreversible EGFR inhibitors in drug resistant non-small cell lung cancer. We have identified the C22,23-epoxytaccalonolides (taccas) as a class of plant-derived microtubule stabilizers that covalently bind to β-tubulin with a high degree of specificity and retain efficacy in drug-resistant models in vitro and in vivo. Although the taccas have potent and persistent antitumor efficacy in vivo, they suffer from a narrow therapeutic index and short serum half-life that provides optimal efficacy when targeted directly to the tumor site. We are undertaking a multipronged approach to generate synthetic tacca analogs (taccalogs) that retain the efficacy of the natural product with improved pharmacokinetic properties, including improved serum binding, to increase half-life and provide better in vivo tolerability. We are also identifying strategies to improve tumor targeting of the taccas, which is being guided by determining their in vivo distribution with radiolabeled analogs. In a complementary approach, we tested the hypothesis that non-covalent microtubule stabilizers that have increased binding affinity to tubulin share properties with the taccas, such as the ability to circumvent drug resistance and provide potent and persistent in vivo efficacy with a more tolerable therapeutic window. Our data suggest that while non-covalent microtubule stabilizers with improved binding compared to paclitaxel provide improved cellular persistence after drug washout and potent antitumor efficacy, they are not able to overcome drug resistance to some degree as covalent inhibitors, demonstrating the advantage of continuing to develop irreversible drugs such as the taccalonolides for the treatment of drug resistant disease. Citation Format: Jacob N. Essif, April Risinger. The development of covalent microtubule stabilizers for drug-resistant cancers [abstract]. In: Proceedings of the 2022 San Antonio Breast Cancer Symposium; 2022 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2023;83(5 Suppl):Abstract nr P2-24-01.