Chemotherapy-resistant Cancer Cells Research Articles

An updated review of the pharmacological effects and potential mechanisms of hederagenin and its derivatives.

Hederagenin (HG) is a natural pentacyclic triterpenoid that can be isolated from various medicinal herbs. By modifying the structure of HG, multiple derivatives with superior biological activities and safety profiles have been designed and synthesized. Accumulating evidence has demonstrated that HG and its derivatives display multiple pharmacological activities against cancers, inflammatory diseases, infectious diseases, metabolic diseases, fibrotic diseases, cerebrovascular and neurodegenerative diseases, and depression. Previous studies have confirmed that HG and its derivatives combat cancer by exerting cytotoxicity, inhibiting proliferation, inducing apoptosis, modulating autophagy, and reversing chemotherapy resistance in cancer cells, and the action targets involved mainly include STAT3, Aurora B, KIF7, PI3K/AKT, NF-κB, Nrf2/ARE, Drp1, and P-gp. In addition, HG and its derivatives antagonize inflammation through inhibiting the production and release of pro-inflammatory cytokines and inflammatory mediators by regulating inflammation-related pathways and targets, such as NF-κB, MAPK, JAK2/STAT3, Keap1-Nrf2/HO-1, and LncRNA A33/Axin2/β-catenin. Moreover, anti-pathogen, anti-metabolic disorder, anti-fibrosis, neuroprotection, and anti-depression mechanisms of HG and its derivatives have been partially elucidated. The diverse pharmacological properties of HG and its derivatives hold significant implications for future research and development of new drugs derived from HG, which can lead to improved effectiveness and safety profiles.

Self-Assembled Micelles Based on Ginsenoside Rg5 for the Targeted Treatment of PTX-Resistant Tumors.

Paclitaxel (PTX) is one of the first-line drugs for prostate cancer (PC) treatment. However, the poor water solubility, inadequate specific targeting ability, multidrug resistance, and severe neurotoxicity are far from being fully resolved, despite diverse PTX formulations in the market, such as the gold-standard PTX albumin nanoparticle (Abraxane) and polymer micelles (Genexol-PM). Some studies attempting to solve the multiple problems of chemotherapy delivery fall into the trap of an extremely complicated formulation design and sacrifice druggability. To better address these issues, this study designed an efficient, toxicity-reduced paclitaxel-ginsenoside polymeric micelle (RPM). With the aid of the inherent amphiphilic molecular structure and pharmacological effects of ginsenoside Rg5, the prepared RPM enhances the water solubility and active targeting of PTX, inhibiting chemotherapy resistance in cancer cells. Moreover, the polymeric micelles demonstrated favorable anti-inflammatory and neuroprotective effects, providing ideas for the development of new clinical anti-PC preparations.

Abstract 3083: Unveiling the role of chemotherapy-resistant, slow-cycling cancer cells in driving cancer cachexia

Abstract Cancer cachexia, a systemic disease marked by substantial, involuntary weight loss, predominantly affecting muscle and adipose tissue, manifests in patients with advanced-stage cancer. The emergence of cancer cachexia poses significant challenges to cancer treatment, diminishing the efficacy of anticancer medications, and the resultant muscle loss adversely impacts the daily lives of patients, diminishing the overall quality of life. Notably, cancer cachexia is implicated in 30-40% of cancer-related deaths, underscoring the pivotal need to comprehend its underlying principles and propose apt interventions to enhance both survival rates and the well-being of cancer patients. Based on our previous findings, cancer cells exhibiting a slow cell cycle, resistant to widely employed clinical chemotherapy agents, exhibit the formation of proinflammatory tumor microenvironment. The escalation of systemic inflammatory cytokines has long been suggested as a potential mechanism contributing to cancer cachexia. Building upon these insights, we hypothesized that slow-cycling cancer cells (SCCs) featuring resistance to chemotherapy are pivotal contributors to cancer cachexia. We investigated the impact of drug-resistant SCCs on the loss of differentiated myotubes and lipid droplets in vitro and muscle and fat loss in in vivo conditions. Interestingly, these cells induced more severe muscle and fat losses. Upon scrutinizing this cell population, we found the overexpression of a particular membrane protein, Protein A. The increase of this Protein A enhanced the release of exosomes from the cell through a novel mechanism distinct from its classical functions, consequently amplifying the release of inflammatory substances originating from cancer cells. In summary, our study elucidates that chemotherapy-resistant cancer cells, characterized by a slow cell cycle, intensify the secretion of inflammatory substances in a manner contingent upon the overexpression of a membrane protein A, which leads to the depletion of muscle and fat located in the distal peripheral regions. Citation Format: Jaebeom Cho, Ho-Young Lee. Unveiling the role of chemotherapy-resistant, slow-cycling cancer cells in driving cancer cachexia [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3083.

Abstract 152: Exploring the impact of heat shock protein 90 on the antitumor efficacy and stemness in medulloblastoma cells

Abstract Background: Medulloblastoma (MB) is the most common children’s malignant brain tumor. Although MB can be treated with conventional chemotherapies, chemoresistance has been frequently developed and thereby hampers their therapeutic effectiveness. The preliminary reports of our research showed that some heat shock proteins have higher levels in MB cells. Among these heat shock proteins, HSP70 and HSP90 expressions are particularly high in MB CD133+ cancer stem cells (CSCs), which arouses our attention and interest. Numerous studies demonstrated that heat shock proteins are prominent in some chemotherapy-resistant cancer cells and CSCs. As known HSP90 and its client proteins are required for the maintenance of self-renewal/stemness and regulation of cell cycle, synergistic killing of cancer cells by simultaneous inhibition of HSP90 expression and activity may be a considerable good therapeutic strategy for treating MB. The purpose of this study is to verify the regulatory function of HSP90 in chemoresistance and stemness of MB. Materials and Methods: The levels of HSP90, its client proteins (P13K/pAKT, Erk, NFκB, and c-Myc), and stemness makers were detected using immunoblotting and immunoprecipitation after the treatment of HSP90 inhibitor (17-AAG) or knockdown of HSP90 by siRNA in MB and MB CSCs. The antitumor effects of HSPs inhibitors or knockdown of HSPs on these cells were evaluated by MTT assay, Annexin V/PI analysis and cell cycle analysis. Results: Our findings revealed elevated expression of HSP90 and its target client proteins in MB cells compared to normal astrocytes. Silencing or inhibiting HSP90 expression induced cell cycle arrest in MB cells. Notably, CD133+ MB CSCs exhibited higher HSP90 levels than parental MB cells. Inhibition of HSP90 led to reduced sphere formation and expression of stemness markers in CD133+ MB CSCs. Treatment with 17-AAG demonstrated antitumor activity in CD133+ MB CSCs, and a synergistic effect was observed when combining conventional chemotherapeutic agents with 17-AAG in MB cells Conclusion: These results illustrated that inhibition of HSP90 could eliminate CSCs, and thereby overcome resistance to chemotherapeutic agents in MB. Chemotherapies in combination with HSP90 blockade may be an emerging MB therapy in the future. Citation Format: Shu-Mei Chen, Hsieng-Yun Huang, Chien-Kai Wang, Yi-Yuan Yang, Sy-Jye Leu, Ying-Ying Li. Exploring the impact of heat shock protein 90 on the antitumor efficacy and stemness in medulloblastoma cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 152.

Bimodal effects on lipid droplets induced in cancer and non-cancer cells by chemotherapy drugs as revealed with a green-emitting BODIPY fluorescent probe.

Lipid droplets (LDs) are cytoplasmic lipid-rich organelles with important roles in lipid storage and metabolism, cell signaling and membrane biosynthesis. Additionally, multiple diseases, such as obesity, fatty liver, cardiovascular diseases and cancer, are related to the metabolic disorders of LDs. In various cancer cells, LD accumulation is associated with resistance to cell death, reduced effectiveness of chemotherapeutic drugs, and increased proliferation and aggressiveness. In this work, we present a new viscosity-sensitive, green-emitting BODIPY probe capable of distinguishing between ordered and disordered lipid phases and selectively internalising into LDs of live cells. Through the use of fluorescence lifetime imaging microscopy (FLIM), we demonstrate that LDs in live cancer (A549) and non-cancer (HEK 293T) cells have vastly different microviscosities. Additionally, we quantify the microviscosity changes in LDs under the influence of DNA-damaging chemotherapy drugs doxorubicin and etoposide. Finally, we show that doxorubicin and etoposide have different effects on the microviscosities of LDs in chemotherapy-resistant A549 cancer cells.

Establishing and Characterizing Chemotherapy-Induced Immunological Dormant Tumor Cell Lines.

Chemotherapy, together with radiotherapy, targeted therapies, and immunotherapy, is the main option to treat cancer patients in neoadjuvant/adjuvant setting to reduce the risk of disease progression and metastasis formation from disseminated tumor cells. Cancer cells that survived chemotherapy treatment may emerge with novel characteristics, one of which is the ability to stimulate the native and adaptive immune systems. Models allowing the characterization of chemotherapy-induced tumor cell plasticity and induction of immune response or adaptation are needed to identify novel mechanisms and devise novel therapeutic strategies to prevent relapses. Here we describe a protocol for selecting chemotherapy-resistant cancer cells and testing the in vivo effects on the local and systemic immune responses. While originally developed to characterize the effects of methotrexate and doxorubicin on murine 4T1 breast cancer cells and the relative immune response, the method can be broadened to other chemotherapies and syngeneic cancer models.

Salinomycin and IR780-loaded upconversion nanoparticles influence biological behavior of liver cancer stem cells by persistently activating the MAPK signaling pathway

The combination of chemotherapy and phototherapy has emerged as a promising therapeutic approach for enhancing the efficacy of cancer treatment and mitigating drug resistance. Salinomycin (SAL), a polyether antibiotic, exhibits potent cytotoxicity against chemotherapy-resistant cancer cells. IR780 iodide, a novel photosensitive reagent with excellent near-infrared (NIR) light absorption and photothermal conversion abilities, is suitable for use in photothermal therapy for cancers. However, both SAL and IR780 exhibit hydrophobic properties that limit their clinical applicability. Upconversion nanoparticles (UCNPs) are an emerging class of fluorescent probe materials capable of emitting high-energy photons upon excitation by low-energy NIR light. The UCNPs not only function as nanocarriers for drug delivery but also serve as light transducers to activate photosensitizers for deep-tissue photodynamic therapy. Here, to enhance the targeting and bioavailability of hydrophobic drugs in liver cancer stem cells (LCSCs), we employ distearoyl phosphorethanolamine-polyethylene glycol (DSPE-PEG) to encapsulate SAL and IR780 on the surface of UCNPs. Cell viability was evaluated using the CCK-8 assay. Cell migration was assessed by the Transwell Boyden Chamber. The activation of the mitogen-activated protein kinase (MAPK) signaling pathway was measured via western blot. The results demonstrated successful loading of both IR780 and SAL onto the UCNPs, and the SAL and IR780-loaded UCNPs (UISP) exhibited a robust photothermal effect under NIR light irradiation. The UISP effectively inhibited the viability of HCCLM3 and LCSCs. Under NIR light irradiation, the UISP further suppressed HCCLM3 viability but had no impact on LCSC viability; however, it could further inhibit LCSC migration. Meanwhile, under NIR light irradiation, the UISP persistently activated the MAPK pathway more significantly in LCSCs. These findings suggest that exposure to NIR light results in persistent activation of the MAPK pathway by UISP, thereby influencing the biological behavior of LCSCs and enhancing their therapeutic efficacy against liver cancer.

How gallic acid regulates molecular signaling: role in cancer drug resistance.

Cancer is one of the deadliest and most heterogeneous diseases. Cancers often develop drug resistance, which can lead to treatment failure or recurrence. Accordingly, anticancer compounds are essential for chemotherapy-resistant cancer cells. Phenolic compounds are of interest in the development of cancer drugs due to their medicinal properties and ability to target different molecular pathways. Gallic acid (GA), as one of the main components of phenol, which is abundantly present in plant compounds such as walnut, sumac, grapes, tea leaves, oak bark, and other plant compounds, has antitumor properties. GA can prevent cancer progression, cell invasion, and metastasis by targeting molecular pathways and is an effective complement to chemotherapy drugs and combating multidrug resistance (MDR). In this review, we discuss various mechanisms related to cancer, the therapeutic potential of GA, the antitumor properties of GA in various cancers, and the targeted delivery of GA with nanocarriers.

Current insights into the oncogenic roles of lncRNA LINC00355.

Long noncoding RNAs (lncRNAs) are a class of nonprotein-coding transcripts that are longer than 200 nucleotides. LINC00355 is a lncRNA located on chromosome 13q21.31 and is consistently upregulated in various cancers. It regulates the expression of downstream genes at both transcriptional and posttranscriptional levels, including eight microRNAs (miR-15a-5p, miR-34b-5p, miR-424-5p, miR-1225, miR-217-5p, miR-6777-3p, miR-195, and miR-466) and three protein-coding genes (ITGA2, RAD18, and UBE3C). LINC00355 plays a role in regulating various biological processes such as cell cycle progression, proliferation, apoptosis, epithelial-mesenchymal transition, invasion, and metastasis of cancer cells. It is involved in the regulation of the Wnt/β-catenin signaling pathway and p53 signaling pathway. Upregulation of LINC00355 has been identified as a high-risk factor in cancer patients and its increased expression is associated with poorer overall survival, recurrence-free survival, and disease-free survival. LINC00355 upregulation has been linked to several unfavorable clinical characteristics, including advanced tumor node metastasis and World Health Organization stages, reduced Karnofsky Performance Scale scores, increased tumor size, greater depth of invasion, and more extensive lymph node metastasis. LINC00355 induces chemotherapy resistance in cancer cells by regulating five downstream genes, namely HMGA2, ABCB1, ITGA2, WNT10B, and CCNE1 genes. In summary, LINC00355 is a potential oncogene with great potential as a diagnostic marker and therapeutic target for cancer.

Subgroup analysis of double-blind, placebo-controlled Ph. 2 study of nanvuranlat in treatment of pre-treated, advanced, refractory biliary tract cancer (BTC): Patients with high LAT1 expression and response to nanvuranlat.

4011 Background: L-type amino acid transporter 1 (LAT1, SLC7A5) is overexpressed in cancer cells leading to aggressive proliferation and lymphatic metastases. LAT1 sustains energy resources by supplying essential amino acids to the TCA cycle in chemotherapy-resistant cancer cells. LAT1 is a documented marker of poor prognosis. In a placebo-controlled, randomized trial involving patients with pre-treated, advanced, refractory biliary tract cancer (BTC), monotherapy with nanvuranlat (JPH203), a selective LAT1 inhibitor, demonstrated a significant improvement in progression free survival (PFS) compared to placebo. Subgroup analysis was conducted to determine whether the level of expression of LAT1 affects nanvuranlat efficacy. Methods: Patients with four different subtypes of advanced BTC were enrolled: intrahepatic and extrahepatic cholangiocarcinoma, and gallbladder and ampulla of Vater cancers. All were refractory to or intolerant of standard chemotherapy and other investigational medicines. Our analysis compared efficacy and safety in the subgroup with high LAT1 expression and all patients group. LAT1 expression was immunohistochemically evaluated in tumor specimens of BTC patients at baseline, as defined by Yanagisawa N, et al. (Cancer Med 2014). Results: At data cut-off (Feb 28, 2022), 211 BTC patients consented at 14 centers in Japan, and 104 patients were randomized (2:1) to nanvuranlat (n = 69) or placebo (n = 35) as the full analysis set (FAS) population. Among the samples immunohistochemically evaluable for LAT1, 62.5% had high LAT1 expression. There was no significant difference in background demography between all patient group and the high-LAT1 subgroup. Nanvuranlat met its primary endpoint (FAS by the blinded independent central review (BICR)), demonstrating a statistically significant improvement in PFS by BICR in comparison with the placebo group. The hazard ratios were further improved with nanvuranlat versus placebo, on analysis of the high-LAT1 subgroup of patients in both PFS and OS. Safety was comparable between nanvuranlat and placebo, on analysis of the high-LAT1 subgroup. Conclusions: This subgroup analysis indicates that the efficacy of nanvuranlat in PFS and OS is enhanced in BTC patients with high LAT1 expression, when compared with placebo. Safety profiles were similar for nanvuranlat- and placebo-treated patients in this subgroup analysis. Clinical trial information: UMIN000034080 . [Table: see text]

Prognostic evaluation of pancreatic cancer based on the model of chemo-radiotherapy resistance-related genes.

The incidence and mortality of pancreatic cancer are almost the same, and the 5-year survival rate is less than 10%. The high mortality of pancreatic cancer is related to chemo-radiotherapy. The present study aimed to establish a prognostic signature of pancreatic cancer based on chemo-radiotherapy resistant-related genes (CRRGs). In this study, we explored the radiation-resistant and chemotherapy-resistant pancreatic cancer cell lines by colony formation and a subcutaneous tumor model in nude mice. Next, we obtained CRRGs from radiation- and gemcitabine-resistant pancreatic cancer cell lines in the Gene Expression Omnibus (GEO) database. Based on univariate Cox and least absolute shrinkage and selection operator (LASSO) Cox regression analyses, a prognostic model of the pancreatic adenocarcinoma (PAAD) cohort in The Cancer Genome Atlas (TCGA) database (N=177) was established and verified using the GEO cohort data set (N=112). Finally, the functions of candidate target genes were verified by a methyl thiazolyl tetrazolium (MTT) assay, a colony formation assay, and a subcutaneous tumor model in nude mice. Through the in vitro and in vivo experiments, we found that radiotherapy- and chemotherapy-resistant pancreatic cancer cells were cross-resistant to chemotherapy and radiotherapy. We constructed a risk model consisting of nine CRRGs (SNAP25, GPR87, DLL1, LAD1, WASF3, ARHGAP29, ZBED2, GAD1, and JAG1) by using public databases. According to the Kaplan-Meier curve analysis, the survival of the high-risk group was worse than that of the low-risk group. We then used nomograms to predict the 1/3/5-year overall survival (OS) in pancreatic cancer patients. We chose JAG1 as a candidate target since it has been proven to be involved in the stemness maintenance of cancer cells, and found that JAG1 silencing inhibited the proliferation and chemo-radiotherapy tolerance of pancreatic cancer cells. This study established and validated a prognostic signature of pancreatic cancer using nine CRRGs. The in vitro and in vivo experiments showed that JAG1 could promote the proliferation and chemoradiotherapy tolerance of pancreatic cancer cell lines. These findings may offer new insights into the role of CRRGs in pancreatic cancer and provide novel prognostic biomarkers for the treatment of pancreatic cancer.

Targeting aldehyde dehydrogenase enzymes in combination with chemotherapy and immunotherapy: An approach to tackle resistance in cancer cells.

Modern cancer chemotherapy originated in the 1940s, and since then, many chemotherapeutic agents have been developed. However, most of these agents show limited response in patients due to innate and acquired resistance to therapy, which leads to the development of multi-drug resistance to different treatment modalities, leading to cancer recurrence and, eventually, patient death. One of the crucial players in inducing chemotherapy resistance is the aldehyde dehydrogenase (ALDH) enzyme. ALDH is overexpressed in chemotherapy-resistant cancer cells, which detoxifies the generated toxic aldehydes from chemotherapy, preventing the formation of reactive oxygen species and, thus, inhibiting the induction of oxidative stress and the stimulation of DNA damage and cell death. This review discusses the mechanisms of chemotherapy resistance in cancer cells promoted by ALDH. In addition, we provide detailed insight into the role of ALDH in cancer stemness, metastasis, metabolism, and cell death. Several studies investigated targeting ALDH in combination with other treatments as a potential therapeutic regimen to overcome resistance. We also highlight novel approaches in ALDH inhibition, including the potential synergistic employment of ALDH inhibitors in combination with chemotherapy or immunotherapy against different cancers, including head and neck, colorectal, breast, lung, and liver.

Abstract 2549: Targeting PI3Kinase AKT pathway and anti-apoptotic protein XIAP in chemotherapy-resistant colorectal cancer cells

Abstract Colorectal Cancer (CRC) ranks among the top three cancers worldwide in terms of incidence and mortality. Treatment of CRC with 5-fluorouracil and oxaliplatin has shown limited efficacy, with resistance often developing to this treatment. Therefore, there is an urgency to identify druggable molecular pathways/proteins that can be efficiently targeted either alone or in combination with chemotherapy to efficiently treat these cancers. In majority of cancers including CRCs, the phosphatidylinositol 3-kinase (PI3K)/AKT/mTOR pathway is frequently found to be constitutively activated leading to poor overall survival. In addition, over expression of the anti-apoptotic protein XIAP has also been shown to induce resistance to chemotherapeutic agents leading to poor prognosis in many epithelial cancers including CRC. Targeting the PI3K/AKT/mTOR pathway and expression of XIAP through specific inhibitors may overcome chemoresistance and altered cell proliferation. The current study examines the effect of oxaliplatin, and inhibitors of PI3K (LY294002) and XIAP (Embelin) in an transiently produced oxaliplatin-resistant colorectal cancer cell line. Transient resistance was induced in CRC cell line HCT116 by intermittent treatment with 25μM oxaliplatin. The anti-proliferative effects of oxaliplatin, Embelin, LY294002 and Torin 2 were determined using MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] assay. Western blots were performed to determine the expression of phosphorylated AKT, XIAP and caspase-3 proteins. Cell cycle and apoptosis assays were performed by flow cytometry to determine the fate of cells following different treatments. Transient resistance was developed in HCT-116 cells (termed HCT-116OXR) after treatment with 25μM oxaliplatin as confirmed by cell viability assays and immunoblotting that showed activation of phospho-AKT and upregulation of XIAP in the transiently resistant cells. HCT116OXR cells responded to treatment with LY294002 and Embelin as compared to parental HCT-116 cells (termed HCT-116WT). Following treatment with LY294002 or Embelin alone or in combination for 48hrs, HCT-116OXR cells underwent a G1 phase cell cycle arrest rather than apoptosis as measured by flow cytometry analysis. The data suggests that PI3K/AKT pathway and XIAP inhibitors cause cell cycle arrest and decreased cell viability in oxaliplatin resistant colorectal cancer cells, which may have implications for treatment of chemotherapy resistant CRC. Citation Format: Rozmeen Akbar, Mati Ur Rehman, Asra Khan, Numair Belgaumi, Iman M. Farooqui, Aafia S. Arain, Azhar Hussain Rajabali, Kulsoom Ghias. Targeting PI3Kinase AKT pathway and anti-apoptotic protein XIAP in chemotherapy-resistant colorectal cancer 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 2549.

Low-dose Pimecrolimus, an FDA-approved Calcineurin Inhibitor, Sensitizes Drug-resistant Cancer Cells via Strong P-gp Inhibition.

Co-treatment with calcineurin inhibitors, such as tacrolimus and cyclosporin A, can sensitize chemotherapy-resistant cancer cells with P-glycoprotein (P-gp)-over-expression. Pimecrolimus (PIME) is a clinically available calcineurin inhibitor with a structure similar to that of tacrolimus. Whether PIME can sensitize P-gp-over-expressing resistant cancer cells remains unclear. Cell viability assay, annexin V analyses, cellular morphology and density observation with a microscope, western-blotting, fluorescence-activated cell sorting (FACS), and analysis for P-gp inhibitory activity were performed to investigate the mechanism of action. PIME exhibited strong cytotoxicity to vincristine (VIC)-treated drug-resistant cell lines (KBV20C and MCF-7/ADR) over-expressing P-gp. Co-treatment with VIC and PIME increased apoptosis and down-regulated the ERK signaling pathway, resulting in G2 arrest. PIME could be co-administered with vinorelbine or eribulin to sensitize resistant KBV20C or MCF-7/ADR cancer cells. Moreover, PIME strongly inhibited the efflux of both rhodamine 123 and calcein-AM substrates through P-gp after 4 h of treatment, indicating that VIC+PIME sensitized cancer cells by inhibiting VIC efflux via direct PIME binding to P-gp. Low doses of PIME, tacrolimus, and cyclosporin A showed similar sensitizing efficiencies in resistant KBV20C cells. These drugs showed similar P-gp inhibitory activities using both rhodamine 123 and calcein-AM substrates, suggesting that calcineurin inhibitors generally have strong P-gp inhibitory activities that sensitize drug-resistant cancer cells with P-gp over-expression. PIME, currently used in clinics, can be repositioned for treating patients with P-gp-over-expressing resistant cancer (stem) cells.

RUNX1 Is Regulated by Androgen Receptor to Promote Cancer Stem Markers and Chemotherapy Resistance in Triple Negative Breast Cancer.

Triple negative breast cancer (TNBC) is an aggressive breast cancer subtype for which no effective targeted therapies are available. Growing evidence suggests that chemotherapy-resistant cancer cells with stem-like properties (CSC) may repopulate the tumor. The androgen receptor (AR) is expressed in up to 50% of TNBCs, and AR inhibition decreases CSC and tumor initiation. Runt-related transcription factor 1 (RUNX1) correlates with poor prognosis in TNBC and is regulated by the AR in prostate cancer. Our group has shown that RUNX1 promotes TNBC cell migration and regulates tumor gene expression. We hypothesized that RUNX1 is regulated by the AR and that both may work together in TNBC CSC to promote disease recurrence following chemotherapy. Chromatin immunoprecipitation sequencing (ChIP-seq) experiments in MDA-MB-453 revealed AR binding to RUNX1 regulatory regions. RUNX1 expression is upregulated by dihydrotestosterone (DHT) in MDA-MB-453 and in an AR+-TNBC HCI-009 patient-derived xenograft (PDX) tumors (p < 0.05). RUNX1 is increased in a CSC-like experimental model in MDA-MB-453 and SUM-159PT cells (p < 0.05). Inhibition of RUNX1 transcriptional activity reduced the expression of CSC markers. Interestingly, RUNX1 inhibition reduced cell viability and enhanced paclitaxel and enzalutamide sensitivity. Targeting RUNX1 may be an attractive strategy to potentiate the anti-tumor effects of AR inhibition, specifically in the slow-growing CSC-like populations that resist chemotherapy which lead to metastatic disease.

C9orf16 represents the aberrant genetic programs and drives the progression of PDAC

BackgroundPancreatic ductal adenocarcinoma (PDAC), constituting 90% of pancreatic cancers, is the fourth leading cause of cancer-related deaths in the world. Lack of early detection of PDAC contributes to its poor prognosis as patients are often diagnosed at an advanced stage of disease. This is mostly due to the lack of promising diagnostic and therapeutic targets and corresponding drugs.Methods and resultsHere, by bioinformatic analysis of single cell RNA-sequencing data on normal pancreas tissues, primary and metastatic PDAC tumors, we identified a promising PDAC biomarker, C9orf16. The expression of C9orf16, rarely detectable in normal epithelial cells, was upregulated in primary PDAC cancer cells and was further elevated in metastatic PDAC cancer cells. Gain or loss of function of C9orf16 demonstrated its critical functions in regulating the cell proliferation, invasion and chemotherapy resistance of cancer cells. Pathway analysis and functional studies identified MYC signaling pathways as the most activated pathways in regulating C9orf16 expression and in mediating the development and progression of PDAC.ConclusionsThese data suggested a crucial gene regulation system, MYC-C9orf16, which is actively involved in PDAC development and progression, and targeting this system should be a novel diagnostic and therapeutic target for PDAC.

Inhibition of insulin-like growth factor 2 mRNA-binding protein 1 sensitizes colorectal cancer cells to chemotherapeutics.

Although colorectal cancer (CRC) treatment has seen a remarkable improvement in the recent years, many patients will develop metastasis due to the resistance of cancer cells to chemotherapeutics. Targeting mechanisms driving the resistance of CRC cells to treatment would significantly reduce cases of metastasis and death. Induction of insulin-like growth factor 2 mRNA-binding protein 1 (IGF2BP1), a direct target of the Wnt/β-catenin signaling pathway, might promote resistance of CRC cells to treatment via activation of anti-apoptotic pathways and induction of the multidrug resistance (MDR1) membrane transporter that pumps drugs out of the cells. We hypothesized that inhibition of IGF2BP1 will sensitize CRC cells to chemotherapeutics. We used CRC cell lines with different status of activation of Wnt signaling to show that inhibition of IGF2BP1 potentiates the anti-growth and anti-proliferative effects of chemotherapeutics on CRC cells with activated Wnt/β-catenin signaling pathway. We observed that the inhibition of IGF2BP1 significantly increases apoptosis in the same cells. A remarkable reduction in the migratory capability of those cells was noted as well. We found that inhibition of IGF2BP1 is sufficient to decrease the resistance of chemotherapy-resistant cancer cells with activated Wnt/β-catenin signaling pathway. These findings portray IGF2BP1 as a good candidate for CRC therapy.

GAS6-AXL inhibition by AVB-500 overcomes resistance to paclitaxel in endometrial cancer by decreasing tumor cell glycolysis (276)

<b>Objectives:</b> Chemotherapy is often ineffective in advanced-stage and aggressive histologic subtypes of endometrial cancer. Overexpression of the receptor tyrosine kinase AXL has been found to be associated with therapeutic resistance, metastasis, and poor prognosis. The mechanism of how inhibition of AXL improves response to chemotherapy is still largely unknown. We aimed to determine whether treatment with AVB-500, a selective inhibitor of <i>GAS6</i>-AXL, improves endometrial cancer cell sensitivity to chemotherapy, particularly through metabolic changes. <b>Methods:</b> Immunohistochemistry (IHC) was performed on a tissue microarray containing specimens from patients with primary and metastatic uterine serous carcinoma. Blind scoring was performed by two reviewers. Kaplan-Meier methods were used to generate time-to-event curves. Cell viability was performed with high-grade endometrial, chemo-resistant cell lines, ARK1 and PUC198. Cells were treated with paclitaxel and with AVB-500+paclitaxel. Intraperitoneal (IP) ARK1 or PUC198 tumors were treated with vehicle, AVB-500, paclitaxel, or AVB-500+paclitaxel. A Seahorse Analyzer was used for glycolytic rate assays. Isotope tracing was used for <i>in vivo</i> metabolite abundance quantification. <b>Results:</b> We found that both <i>GAS6</i> and AXL expression were higher by IHC in tumors with a poor response to chemotherapy compared to tumors with a good response to chemotherapy (<i>GAS6</i>: 40.9% vs 30.4%, p=0.012; AXL: 60.7% vs 42.7%, p=0.013). Median PFS and OS were longer in patients with low <i>GAS6</i> expressing tumors compared to high <i>GAS6</i> expressing tumors (PFS: 33.6 mo vs 10.6 mo, p=0.003; OS: 39.5 mo vs 27.7 mo, p=0.003). We showed that chemotherapy-resistant endometrial cancer cell lines, ARK1 and PUC198, had improved sensitivity and synergy with paclitaxel when treated in combination with AVB-500. ARK1 and PUC198 <i>in vivo</i> IP models had significantly fewer tumors (ARK1: 2.8 vs 7.9, p=0.028; PUC198: p=0.029) and decreased tumor weight (ARK1: p<0.001; PUC198: p=0.006) when treated with AVB-500+paclitaxel versus paclitaxel alone. Treatment with AVB-500 + paclitaxel decreased AKT signaling, which resulted in a decrease in basal glycolysis. Finally, treatment with AVB-500 + paclitaxel decreased multiple glycolytic metabolites, including fructose-1,6-bisphosphate (p=0.002), 3-phosphoglyceric acid (p=0.014), phosphoenolpyruvate (p=0.013), and lactate (p=0.024) versus treatment with paclitaxel alone. <b>Conclusions:</b> Our study provides a strong preclinical rationale for combining AVB-500 with paclitaxel in aggressive endometrial cancer models. Additionally, the continued identification of viable biomarkers is critical to the success of targeted therapies, such as AVB-500. Given AXL's role in glucose homeostasis, one or more glycolytic intermediates may prove to be a useful biomarker to help predict sensitivity in patients, thus tailoring who would benefit from this treatment combination.

Abstract 2978: E2F1 regulates cell competition, increase proliferation and inhibits chemotherapy resistance in cancer cells

Abstract Introduction: Tumor heterogeneity is a key element during the development of chemo-resistance and tumor relapse. It remains unknown how resistant tumor cells influence sensitive cells during conditions of cohabitation. The goal of our study was to determine driving factors mediating interactions between resistant and sensitive cancer cells and the effects of cohabitation on both phenotypes. Materials and methods: We used two isogenic ovarian cancer cell lines pairs, sensitive and resistant to platinum: OVCAR5 (IC50 = 2.05 uM) vs. OVCAR5-CisR (IC50 = 7.26 uM) and PEO1 (IC50 = 5.92 uM) vs. PE04 (IC50 = 14.45 uM) labeled with either GFP or RFP. The cell pairs were seeded together in long direct term culture, separated by fluorescence activated cell sorting (FACS), and used for RNAseq analysis, proliferation assays, Cisplatin IC50 determinations, and cell cycle analysis. Results: Long Term direct co-culture (14 Days) of platinum-sensitive and -resistant cells induced increased proliferation (p &amp;lt; 0.001) and a greater proportions of cells in the S and G2 phases of the cell cycle in sensitive PEO1 and OVCAR5 cells compared with cells cultured by themselves. Resistance to platinum of the sensitive cell lines decreased further by co-culture (IC50 change from 5.92 uM to 2.79 uM in PE01, and from 2.05 uM to 1.51 uM in OVCAR5). No changes in cell proliferation, or resistance to platinum occurred in the platinum resistant cells. Use of conditioned media of resistance cells did not change proliferation, or response to platinum of sensitive cells. RNAseq detected 3840 differentially expressed genes; 1774 upregulated and 2066 downregulated. IPA analysis showed enrichment of the cell cycle pathways (Cell Cycle Control, Cyclins and Cell Cycle Regulation, among others) and upregulation of cell proliferation genes including the E2F1 transcription factor. Western blot and qRT-PCR confirmed upregulation of E2F1 in co-cultured platinum sensitive cells vs cells in monoculture. Furthermore, western blot measurements showed increased levels of other proteins involved in cell cycle regulation such as CDK4 and CDK6. Cell cycle analysis indicated an increase of cells in S and G2 phase in the co-cultured platinum sensitive cells (18.8% up G2 phase; p = 0.031). Treatment with the CDK4/6 inhibitor Palbociclib or with the E2F1 inhibitor HLM006474 reverted the increase in proliferation rate. Conclusion: Long term cohabitation of platinum-sensitive and -resistant cancer cells drives sensitive cells to a higher proliferative state, which is more responsive to platinum. Direct cell-cell contact modulates cell cycle progression in sensitive cells through a mechanism regulated by E2F1. Our data reveal unexpected effects caused by interactions between cancer cells with different proliferative rates and levels of platinum resistance, modelling competition between cells in heterogeneous tumors. Citation Format: Andres Felipe Valdivia, Matthew Cowan, Guangyuan Zhao, Horacio Cardenas, Daniela Matei. E2F1 regulates cell competition, increase proliferation and inhibits chemotherapy resistance in cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 2978.

Anti-cancer potential of cannabis terpenes in a Taxol-resistant model of breast cancer.

Chemotherapeutic resistance can limit breast cancer outcomes; therefore, the exploration of novel therapeutic options is warranted. Isolated compounds found in cannabis have previously been shown to exhibit anti-cancer effects, but little is known about their effects in resistant breast cancer. Our study aimed to evaluate the effects of terpenes found in cannabis in in vitro chemotherapy-resistant model of breast cancer. We aimed to identify whether five terpenes found in cannabis produced anti-cancer effects, and whether their effects were improved upon co-treatment with cannabinoids and flavonoids also found in cannabis. Nerolidol and β-caryophyllene produced the greatest cytotoxic effects, activated the apoptotic cascade, and reduced cellular invasion. Combinations with the flavonoid kaempferol potentiated the cytotoxic effects of ocimene, terpinolene, and β-myrcene. Combinations of nerolidol and Δ9-tetrahydrocannabinol or cannabidiol produced variable responses ranging from antagonism and additivity to synergy, depending on concentrations used. Our results indicate that cannabis terpenes, alone or combined with cannabinoids and flavonoids, produced anti-cancer effects in chemotherapy-resistant breast cancer cell lines. This study is a first step in the identification of compounds that could have therapeutic potential in the treatment of resistant breast cancer.