Drug-resistant Ovarian Cancer Research Articles

Yolk-Shell-Type Gold Nanoaggregates for Chemo- and Photothermal Combination Therapy for Drug-Resistant Cancers.

Epithelial ovarian cancer is a gynecological cancer with the highest mortality rate, and it exhibits resistance to conventional drugs. Gold nanospheres have gained increasing attention over the years as photothermal therapeutic nanoparticles, owing to their excellent biocompatibility, chemical stability, and ease of synthesis; however, their practical application has been hampered by their low colloidal stability and photothermal effects. In the present study, we developed a yolk-shell-structured silica nanocapsule encapsulating aggregated gold nanospheres (aAuYSs) and examined the photothermal effects of aAuYSs on cell death in drug-resistant ovarian cancers both in vitro and in vivo. The aAuYSs were synthesized using stepwise silica seed synthesis, surface amino functionalization, gold nanosphere decoration, mesoporous organosilica coating, and selective etching of the silica template. Gold nanospheres were agglomerated in the confined silica interior of aAuYSs, resulting in the red-shifting of absorbance and enhancement of the photothermal effect under 808 nm laser irradiation. The efficiency of photothermal therapy was first evaluated by inducing aAuYS-mediated cell death in A2780 ovarian cancer cells, which were cultured in a two-dimensional culture and a three-dimensional spheroid culture. We observed that photothermal therapy using aAuYSs together with doxorubicin treatment synergistically induced the cell death of doxorubicin-resistant A2780 cancer cells in vitro. Furthermore, this type of combinatorial treatment with photothermal therapy and doxorubicin synergistically inhibited the in vivo tumor growth of doxorubicin-resistant A2780 cancer cells in a xenograft transplantation model. These results suggest that photothermal therapy using aAuYSs is highly effective in the treatment of drug-resistant cancers.

Emerging targeted drug delivery strategies toward ovarian cancer.

Ovarian cancer is a high-mortality malignancy in women. The contemporary clinical chemotherapy with classic cytotoxic drugs, targeted molecular inhibitors would mostly fail when ovarian cancer cells become drug-resistant or metastasize through the body or when patients bare no more toleration because of strong adverse effects. The past decade has spotted varying targeted delivery systems including antibody-drug conjugates (ADCs), peptide/folate/aptamer-drug conjugates, polymer-drug conjugates, ligand-functionalized nanomedicines, and dual-targeted nanomedicines that upgrade ovarian cancer chemo- and molecular therapy effectively in preclinical/clinical settings via endowing therapeutic agents selectivity and bypassing drug resistance as well as lessening systemic toxicity. The targeted delivery approaches further provide means to potentiate emergent treatment modalities such as molecular therapy, gene therapy, protein therapy, photodynamic therapy, dual-targeting therapy and combination therapy for ovarian cancer. This review highlights up-to-date development of targeted drug delivery strategies toward advanced, metastatic, relapsed, and drug resistant ovarian cancers.

Low Expression of SLC7A11 Confers Drug Resistance and Worse Survival in Ovarian Cancer via Inhibition of Cell Autophagy as a Competing Endogenous RNA.

Drug resistance is the main cause of chemotherapy failure in ovarian cancer (OC), and identifying potential druggable targets of autophagy is a novel and promising approach to overcoming drug resistance. In this study, 131 genes associated with autophagy were identified from three autophagy-related databases, and of these, 14 were differentially expressed in 90 drug-resistant OC tissues versus 197 sensitive tissues according to the Cancer Genome Atlas ovarian cancer cohort. Among these 14 genes, SLC7A11 was significantly decreased in two paclitaxel-resistant OC cells (HeyA8-R and SKOV3-R) and in 90 drug-resistant tissues compared with their controls. In vitro overexpression of SLC7A11 significantly increased the sensitivity of HeyA8-R cells to paclitaxel, inhibited colony formation, induced apoptosis, and arrested cell cycle. Further, low SLC7A11 expression was correlated with poor overall survival (OS), progression-free survival (PFS), and post-progression survival (PPS) in 1815 OC patients. Mechanistically, SLC7A11 strongly regulated cell autophagy as a competing endogenous RNA (ceRNA) based on pan-cancer analyses of 32 tumor types. Specifically, as a ceRNA for autophagy genes STX17, RAB33B, and UVRAG, SLC7A11 was strongly and positively co-expressed with these three genes in 20, 12, and 12 different tumors, respectively, in 379 OC tissues and in 90 drug-resistant OC tissues, and the former two were significantly upregulated in SLC7A11-overexpressed HeyA8-R cells. Further, SLC7A11 induced the protein expression of other autophagy genes, such as LC3, Atg16L1, and Atg7, and the expression of the respective proteins was further increased when the cells were treated with paclitaxel. The results strongly suggest that SLC7A11 regulates autophagy via ceRNA interactions with the three abovementioned genes in pan-cancer and in drug-resistant OC. Moreover, low expression of STX17 and UVRAG also significantly predicted low OS, PFS, and PPS. The combination of SLC7A11 with STX17 was more predictive of OS and PFS than either individually, and the combination of SLC7A11 with UVRAG was highly predictive of OS and PPS. The above results indicated that decreased SLC7A11 resulted in drug resistance and effected low rates of survival in OC patients, probably via ceRNA interactions with autophagy genes, and thus the gene could serve as a therapeutic target and potential biomarker in OC.

TCP1 increases drug resistance in acute myeloid leukemia by suppressing autophagy via activating AKT/mTOR signaling

T-complex protein 1 (TCP1) is one of the subunits of chaperonin-containing T complex (CCT), which is involved in protein folding, cell proliferation, apoptosis, cell cycle regulation, and drug resistance. Investigations have demonstrated that TCP1 is a factor being responsible for drug resistance in breast and ovarian cancer. However, the TCP1 role in acute myeloid leukemia (AML) remains elusive. In the present study, we discovered that the TCP1 expression was elevated in AML patients and high TCP1 expression was associated with low complete response rate along with poor overall survival. TCP1 showed higher expression in the adriamycin-resistant leukemia cell line HL60/A and K562/A, comparing to their respective parent cells HL60 and K562 cells. TCP1 inhibition suppressed drug resistance in HL60/A and K562/A cells, whereas TCP1 overexpression in HL60 cells incremented drug resistance, both in vitro and in vivo. Mechanistic investigations revealed that TCP1 inhibited autophagy and adriamycin-induced cell apoptosis, and TCP1-mediated autophagy inhibition conferred resistance to adriamycin-induced cell apoptosis. Furthermore, TCP1 interacted with AKT and mTOR to activate AKT/mTOR signaling, which negatively regulates apoptosis and autophagy. Pharmacological inhibition of AKT/mTOR signal particularly activated autophagy and resensitized TCP1-overexpressing HL60 cells to adriamycin. These findings identify a novel role of TCP1 regarding drug resistance in AML, which advise a new strategy for overcoming drug resistance in AML through targeting TCP1/AKT/mTOR signaling pathway.

The Non-Coding RNAs Inducing Drug Resistance in Ovarian Cancer: A New Perspective for Understanding Drug Resistance.

Ovarian cancer, a common malignant tumor, is one of the primary causes of cancer-related deaths in women. Systemic chemotherapy with platinum-based compounds or taxanes is the first-line treatment for ovarian cancer. However, resistance to these chemotherapeutic drugs worsens the prognosis. The underlying mechanism of chemotherapeutic resistance in ovarian cancer remains unclear. Non-coding RNAs, including long non-coding RNAs, microRNAs, and circular RNAs, have been implicated in the development of drug resistance. Abnormally expressed non-coding RNAs can promote ovarian cancer resistance by inducing apoptosis inhibition, protective autophagy, abnormal tumor cell proliferation, epithelial-mesenchymal transition, abnormal glycolysis, drug efflux, and cancer cell stemness. This review summarizes the role of non-coding RNAs in the development of chemotherapeutic resistance in ovarian cancer, including their mechanisms, targets, and potential signaling pathways. This will facilitate the development of novel chemotherapeutic agents that can target these non-coding RNAs and improve ovarian cancer treatment.

Mechanisms of drug resistance in ovarian cancer

Ovarian cancer is the fifth most common cancer affecting women and the most lethal cancer among the gynecological cancers. Because of the lack of specific symptoms and no special screening tools, it is recognised in an advanced stage. In addition, drug resistance in ovarian cancer is so frequent, that genes and cross-talks between some important pathways are still analysed. In this review, the major and recently identified molecular mechanisms of drug resistance, including platinum, taxane, bevacizumab and PARPi resistance mechanisms in ovarian cancer from relevant literature have been investigated.

Up-regulation of GSTT1 in serous ovarian cancer associated with resistance to TAXOL / carboplatin

Serous ovarian cancer (SOC) is the most common women cancer and the leading cause of cancer-related mortality among the gynaecological malignancies. Although effective chemotherapeutics combined with surgery are developed for the treatment, the five-year survival rate is unsatisfactory due to chemoresistance. To overcome this shortcoming of chemotherapy, we established taxol and carboplatin resistant SOC cell lines for the understandings of the molecular and cellular mechanisms of chemoresistance. Here, we found that these chemoresistant cell lines showed less viability and proliferation, due to more cells arrested at G0/G1 phase. Glutathione-S-transferases-theta1 (GSTT1) was significantly upregulated in these chemoresistant cells, along with other chemoresistant genes. Meanwhile, GSTT1 expression was also significantly upregulated in the SOC patient tissues after taxol treatment, indicating this upregulation was physiologically relevant to chemotherapy. Further, suppression of GSTT1 expression by shRNA in SOC cell lines led to more sensitivity to drug treatment, through increasing divided cells and promoting cell death. Moreover, the expression of DNA topoisomerase 1 (Topo I) was in synergy with that of GSTT1 in the chemoresistant cells, and GSTT1 can bind to Topo I in vitro, which suggested GSTT1 could function through DNA repair mechanism during chemoresistance. In summary, our data imply that GSTT1 may be a potential biomarker or indicator of drug resistance in serous ovarian cancer.

The Emerging Role of Non-coding RNAs in Drug Resistance of Ovarian Cancer.

Ovarian cancer is one of the most common gynecological malignancies with highest mortality rate among all gynecological malignant tumors. Advanced ovarian cancer patients can obtain a survival benefit from chemotherapy, including platinum drugs and paclitaxel. In more recent years, the administration of poly-ADP ribose polymerase inhibitor to patients with BRCA mutations has significantly improved the progression-free survival of ovarian cancer patients. Nevertheless, primary drug resistance or the acquisition of drug resistance eventually leads to treatment failure and poor outcomes for ovarian cancer patients. The mechanism underlying drug resistance in ovarian cancer is complex and has not been fully elucidated. Interestingly, different non-coding RNAs (ncRNAs), such as circular RNAs, long non-coding RNAs and microRNAs, play a critical role in the development of ovarian cancer. Accumulating evidence has indicated that ncRNAs have important regulatory roles in ovarian cancer resistance to chemotherapy reagents and targeted therapy drugs. In this review, we systematically highlight the emerging roles and the regulatory mechanisms by which ncRNAs affect ovarian cancer chemoresistance. Additionally, we suggest that ncRNAs can be considered as potential diagnostic and prognostic biomarkers as well as novel therapeutic targets for ovarian cancer.

Predicting response to platinum and non-platinum drugs through bioluminescence resonance energy transfer (BRET) based bio-molecular interactions in platinum resistant epithelial ovarian cancer

Therapy induced rewiring of signalling networks often lead to acquirement of platinum-resistance, thereby necessitating the use of non-platinum agents as second-line treatment particularly for epithelial ovarian cancer (EOC). A prior subject-specific assessment can guide the choice of optimal non-platinum agent/s and possible targeted therapeutic/s. Assessment of protein-protein interactions are superior to simple cytotoxicity assays to determine therapeutic efficacy and associated molecular responses. Utilizing improved PIP3-AKT and ERK1/2 activation Bioluminescence Resonance Energy Transfer (BRET) sensors, we report chemotherapy-induced ERK1/2 activation predominantly in cisplatin-paclitaxel resistant EOC cells and increased activation of both ERK1/2 and AKT in malignant ascites derived cancer cells from platinum-resistant patients but not from treatment-naive or platinum-sensitive relapse patients. Further, majority of the non-platinum drugs except irinotecan increased ERK1/2 activation in platinum-taxol resistant cells as observed by live-cell BRET assessment which were associated with p90RSK1/2 and BAD activation along with upregulation of multidrug transporter gene ABCC1 and cell survival genes like cyclin D1 and Bcl2. Interestingly, only irinotecan was able to sensitize these resistant cells. Altogether, this first report of BRET based sensing of molecular pathway activations in platinum resistant cell lines and patient's derived cancer cells highlight the clinical potential of BRET sensors in management of therapy resistant cancer.

TPL Inhibits the Invasion and Migration of Drug-Resistant Ovarian Cancer by Targeting the PI3K/AKT/NF-κB-Signaling Pathway to Inhibit the Polarization of M2 TAMs.

Chemoresistance is the primary reason for the poor prognosis of patients with ovarian cancer, and the search for a novel drug treatment or adjuvant chemotherapy drug is an urgent need. The tumor microenvironment plays key role in the incidence and development of tumors. As one of the most important components of the tumor microenvironment, M2 tumor-associated macrophages are closely related to tumor migration, invasion, immunosuppressive phenotype and drug resistance. Many studies have confirmed that triptolide (TPL), one of the principal components of Tripterygium wilfordii, possesses broad-spectrum anti-tumor activity. The aims of this study were to determine whether TPL could inhibit the migration and invasion of A2780/DDP cells in vitro and in vivo by inhibiting the polarization of M2 tumor-associated macrophages (TAMs); to explore the mechanism(s) underlying TPL effects; and to investigate the influence of TPL on murine intestinal symbiotic microbiota. In vitro results showed that M2 macrophage supernatant slightly promoted the proliferation, invasion, and migration of A2780/DDP cells, which was reversed by TPL in a dose-dependent manner. Animal experiments showed that TPL, particularly TPL + cisplatin (DDP), significantly reduced the tumor burden, prolonged the life span of mice by inhibiting M2 macrophage polarization, and downregulated the levels of CD31 and CD206 (CD31 is the vascular marker and CD206 is the macrophage marker), the mechanism of which may be related to the inhibition of the PI3K/Akt/NF-κB signaling pathway. High-throughput sequencing results of the intestinal microbiota in nude mice illustrated that Akkermansia and Clostridium were upregulated by DDP and TPL respective. We also found that Lactobacillus and Akkermansia were downregulated by DDP combined with TPL. Our results highlight the importance of M2 TAMs in Epithelial Ovarian Cancer (EOC) migration ability, invasiveness, and resistance to DDP. We also preliminarily explored the mechanism governing the reversal of the polarization of M2 macrophages by TPL.

Abstract 197: MONE: A construction for interpreting deep learning features in pathology slides

Abstract Deep learning has become a popular tool for analyzing hematoxylin and eosin (H&E) stained whole slide images (WSIs) and has been utilized to study conserved spatial behaviors across cancers [1]. Deep learning models are black-boxes and difficult to interpret. We propose the concept of an abstract morphological gene, hereafter called mone, defined as the features encoding the morphology at each region of a WSI. Mones averaged over WSIs share distributional similarities with bulk-level expressions. Such similarities allow using tools originally developed for studying gene expressions. We study >22000 H&E slides of 19 cancers of the cancer genome atlas (TCGA) using the inceptionV3 network. TSNE plots suggest mones detect tumors and distinguish tissues of origin. We obtained AUCs above 95% for one-versus-all predictions of mone-based classifiers detecting tumors and tissue of origin. We also obtained cross-classification accuracies comparable to [1] using a mone-based logistic regression model. Differential mone analysis identifies pan-cancer and cancer-specific mones differentiating tumor and normal slides. Mone 893 is a pan-cancer feature and an indicator of dense cellular regions. We identified this mone in breast cancer WSIs and validated its consistent behavior in ovarian and lung cancers. Differential mone analysis comparing formalin-fixed paraffin-embedded (FFPE) and fresh frozen slides identifies deep learning features which may be affected by frozen tissue artifacts. Removing such features is essential for developing models not confounded by tissue artifacts. While recent deep learning models predict expressions from WSIs (see [2] for examples), here, we use them to identify the morphological features mones encode. Integrative mone-gene co-expression analysis suggests mone 893 heavily correlates with genes in integrin signaling and inflammation mediated by cytokines and chemokines pathways in breast cancer. Mone 869 heavily correlates with expression of COL8A1 in ovarian cancer. Expression of collagen genes is associated with poor prognosis and drug resistance in ovarian cancer [3,4]. These findings indicate the significant associations between individual deep learning-defined features and both genetic and prognostic quantifications.

Abstract 3093: Nix mediated mitophagy maintains ovarian cancer cell stemness

Abstract Ovarian cancer is the most frequent cause of death among gynecologic malignancies. Despite the availability of effective therapeutics against cancer, the overall 5-year survival rate of ovarian cancer is still ~30-40%. Most of the lethality are due to cancer relapse and drug resistance, where cancer stem cells (CSCs) are supposed to be the key regulators. CSCs also known as tumor-initiating cells, are a small subgroup of tumor cells that can develop tumors through their capability of self-renewal and differentiation. CSCs can canonically nurture therapeutic resistance, tumor invasion, metastasis, and recurrence, which tremendously attract the scientific community for the unique CSC-based cancer therapeutics. While changing phenotypical features of CSCs complicates their identification and thereby metabolism-based alternate strategy seems to be worthy anti CSC therapy. Though autophagic degradation of mitochondria (mitophagy) based mitochondrial dynamics endowed cancer cells with stemness, but wide informational gaps and missing mechanistic knowledge further necessitates the study on mitophagy dependent CSC regulation. In this study, we investigated the potential involvement of mitophagy in maintenance of cancer cell stemness. In our experimental conditions, FACS sorted CD117/44+/+, ALDHbr and ovarian cancer stem cell like cells (CSCLCs) from OVACR4 and OVCAR3 cells showed higher mitochondrial translocation of cytosolic LC3 that directly indicates the progression of mitophagy in these subpopulations. Reduced mitocontent in CSCLCs as compare to bulk cells also supports the mitochondrial degradation. To analyze the involvement of mitophagy in stemness, mitophagy has been targeted with mDIVI a well-known mitophagy inhibitor, which downregulate the sphere forming ability and protein levels of stemness marker such as SOX2 in ovarian cancer cells. Furthermore, the mechanistic detail of mitophagy involvement in cancer stemness were evaluated. Among different mitophagy proteins, BNIP3L/NIX is highly expressed in ALDHbr and CD117/44+/+ sorted cells as compare to ALDHdim and CD117/44-/- cells, respectively. The functional implication of NIX in ovarian CSCs was verified by gene silencing approach. NIX silencing diminished the sphere forming ability and stem cell marker OCT4 that supported its crucial involvement in ovarian cancer cells stemness. In summary, our findings provide a mechanistic detail of mitophagy underlying maintenance of the CSC population in ovarian cancer and suggest that targeting of NIX based mitophagy could be exploited for the eradication of CSC and thereby tumor recurrence and drug resistance in ovarian cancer. Citation Format: Tejinder Pal Khaket, Qi-En Wang. Nix mediated mitophagy maintains ovarian cancer cell stemness [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 3093.

Abstract CT012: TRX-E-002-1 in treatment-refractory ovarian cancer: Final phase 1 study results from the dose-escalation and dose-expansion cohorts

Abstract Introduction: For women with advanced ovarian cancer survival outcomes with standard cytotoxic chemotherapy are poor and are thought to reflect the existence of drug-resistant ovarian cancer stem cells. TRX-E-002-1 (Cantrixil), a novel third generation benzopyran molecule has been shown to be effective in a mouse model of recurrent chemotherapy-resistant ovarian cancer. We present the final results of a phase I progressive design trial (Part A dose escalation, Part B dose expansion) of Cantrixil (NCT02903771). The objectives were to establish maximum tolerated dose (MTD) when given in combination with chemotherapy, and to evaluate safety, tolerability and anti-tumour activity of intraperitoneal (IP)-administered Cantrixil. Methods: Women who had completed ≥2 prior regimens and whose disease was platinum-refractory, platinum-resistant or who had documented intolerance to platinum therapy, were eligible. Treatment comprised up to eight 3-week cycles. In the first 2 cycles, patients were dosed weekly with Cantrixil as monotherapy after which investigators were allowed to initiate pre-defined standard intravenous chemotherapy regimens (cycles 3-8). All patients were followed up for 3 months after the end of treatment. Results: Of 32 patients enrolled, 25 received ≥1 Cantrixil dose; 6 patients (24%) completed all 8 treatment cycles. Patients (92% Caucasian, mean age 62.5 years) had a median of 2 prior lines of platinum therapy and a median of 3 prior lines of anticancer therapies (including anti-VEGF; n=13, 52% and PARPi; n=6, 24%). Platinum sensitivity: refractory (n=3,12%), resistant (n=17, 68%), sensitive (n=5, 20%). In Part A (n=11), an MTD of 5mg/kg was established on the dose-limiting toxicity of ileus (n=2) and safety signals of bowel obstruction (n=3) and abdominal pain (n=2). The pharmacokinetic profile was multi-exponential, with rapid increases in systemic concentration and distribution and a slower elimination phase. Drug accumulation was minimal and was not influenced by co-administration of chemotherapy. Analysis of weight and dose-normalized (to 1 mg/kg) data found no notable trends. Across Parts A and B, 16 patients received ≥1 Cantrixil dose and had a post-baseline efficacy measurement available. The overall response rate was 18%, including one complete response (platinum-resistant; who remains in remission 37.05 months since starting treatment) and two partial responses (one platinum-resistant, one platinum refractory). The median progression free survival was 3.06 months (95% CI:1.28,∞). Conclusion: IP-administered Cantrixil, a first-in-class, dual acting, anti-cancer therapy has encouraging activity in a cohort of difficult to treat patients with persistent epithelial ovarian, fallopian tube or primary peritoneal cancer who have demonstrated resistance to a range of prior treatments. Disease response compares favorably to a figure of 10% for historical controls. Citation Format: Jermaine Coward, Ganessan Kichenadasse, Paul Harnett, Kathleen Moore, Minal Barve, James Garner, Mary Lopresti, Don S. Dizon. TRX-E-002-1 in treatment-refractory ovarian cancer: Final phase 1 study results from the dose-escalation and dose-expansion cohorts [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr CT012.

Abstract 306: Cationic cholesterol liposomes for combination therapy to treat drug-resistant ovarian cancer

Abstract Introduction: Ovarian cancer is the fifth leading cause of cancer mortality in women, with nearly 75% of women who respond to initial platinum-based chemotherapy experiencing relapse due to drug resistance [1,2]. Liposomes are a promising solution to overcoming drug resistance due to their biocompatibility and capacity to encapsulate hydrophilic and hydrophobic drugs as well as complex small interfering RNA (siRNA), which have the potential to downregulate the expression of genes related to drug resistance [3]. We aim to synthesize and characterize a cationic liposomal system to deliver siRNA and paclitaxel (PTX) to ovarian cancer cells. Methods: Cholesterol (CHOL) liposomes were synthesized by the thin-film hydration method. Dynamic light scattering (DLS) was used to determine the size, polydispersity index (PDI), and zeta potential of the liposomes. Uptake of liposomes into OVCAR3 and OVCAR3-T40, a wild-type and a paclitaxel-resistant human adenocarcinoma cell line, was examined using fluorescence microscopy. The cytotoxicity of unloaded CHOL liposomes was evaluated through MTS assay on OVCAR3 and OVCAR3-T40 cells. Results: PTX- and siRNA-loaded CHOL liposomes had an average diameter of 114.9 ± 10.35 nm and a zeta potential of 27.6 ± 1.79 mV. Blank CHOL liposomes had an average diameter of 123.0 ± 2.49 nm and zeta potential of 32.3 ± 2.16 mV. All formulations of liposomes were cationic and formed monodisperse nanoparticles. The encapsulation efficiency of siRNA and PTX was 99.8% and 80.4% respectively. Coumarin 6, a hydrophobic model drug, was loaded into liposomes to verify cellular uptake through fluorescent imaging. Results demonstrated that the liposomal system was efficiently delivered intracellularly. Blank liposomes were used to determine the toxicity of the delivery system. The unloaded liposomes were not cytotoxic to both the wild-type and drug-resistant cell lines at concentrations up to 75 µg/mL, and therefore, cytotoxicity of drug-loaded liposomes can be attributed to paclitaxel, siRNA, or combination treatment. Conclusions: Liposomes were successfully formed with a monodisperse size, exhibited effective drug and siRNA loading, and were internalized into OVCAR3 and OVCAR3-T40 cells. Future work includes investigating the efficacy of the liposomal system in mediating gene silencing. Acknowledgements: This work was supported in part by the National Science Foundation EPSCoR Program under NSF Award # OIA-1655740 and Clemson Creative Inquiry.

Abstract 281: pH-sensitive liposome for siRNA delivery to treat drug-resistant ovarian cancer

Abstract Introduction: With a 5-year survival rate of 47%, ovarian cancer is the 5th leading cause of death amongst women worldwide. Over 75% of patients experience recurrence after initial treatment, indicating a need for improved treatment options. Drug resistance is a major barrier hindering the success of current treatment methods. Our study analyzes the characteristics of a stimuli-sensitive liposomal delivery system for combatting drug resistance. Our delivery system will deliver bioactive siRNAs targeting genes related to drug resistance, cell proliferation, and apoptosis. In this study, we investigate the characteristics of the liposomes to determine particle size, surface charge, and ability to encapsulate/bind both siRNAs. We also begin to investigate the delivery potential of the pH-sensitive liposomal formulation in vitro using ovarian cancer cell lines. Methods: Empty and siRNA loaded cationic, pH-sensitive liposomes (CHEMS-LPs) were synthesized by the thin-film hydration method. Liposome size, zeta potential, and polydispersity index (PDI) were measured by dynamic light scattering (DLS). To measure siRNA encapsulation efficiency, fluorescently labeled siRNA was loaded into CHEMS-LPs and subjected to centrifugation to pellet the LPs. Fluorescence spectroscopy was used to detect siRNA in the supernatant. The toxicity of unloaded CHEMS-LPs was determined by an MTS assay using OVCAR3 (drug-sensitive) and OVCAR3-T40 (drug-resistant) human ovarian cancer cells. Results: The size and zeta potential of blank and siRNA-loaded CHEMS-LPs were 97.88 ± 2.39 nm and 29.0 ± 2.00 mV, and 80.78 ± 0.77​ nm and 13.1 ±1.66​ mV, respectively. The positively charged zeta potential confirms the cationic nature of our liposomes. The PDI demonstrated that the liposomes were unimodal and monodisperse with PDI values of less than 0.300 for each formulation. In addition, siRNA was successfully bound to CHEMS-LPs through electrostatic interaction with the cationic lipid layer, resulting in an encapsulation efficiency of 99.6% Conclusion: CHEMS-LPs are pH-sensitive, cationic, monodisperse liposomes able to encapsulate siRNAs in order to mediate delivery into ovarian cancer cells. Their stable structure, positive charge, and low cytotoxicity is promising for future studies, including delivery of bioactive siRNAs to stimulate downregulation of target genes related to drug resistance. Acknowledgements: This work was supported in part by the National Science Foundation EPSCoR Program under Award # OIA-1655740. We would like to thank George Duran from Stanford University for donating the OVCAR3-T40 cell line. Citation Format: Kharimat Lora Alatise, Samantha Gardner, Angela Alexander-Bryant. pH-sensitive liposome for siRNA delivery to treat drug-resistant ovarian cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 281.

NOX4 Signaling Mediates Cancer Development and Therapeutic Resistance through HER3 in Ovarian Cancer Cells.

Development of resistance to therapy in ovarian cancer is a major hinderance for therapeutic efficacy; however, new mechanisms of the resistance remain to be elucidated. NADPH oxidase 4 (NOX4) is responsible for higher NADPH activity to increase reactive oxygen species (ROS) production. In this study, we showed that higher levels of NOX4 were detected in a large portion of human ovarian cancer samples. To understand the molecular mechanism of the NOX4 upregulation, we showed that NOX4 expression was induced by HIF-1α and growth factor such as IGF-1. Furthermore, our results indicated that NOX4 played a pivotal role in chemotherapy and radiotherapy resistance in ovarian cancer cells. We also demonstrated that NOX4 knockdown increased sensitivity of targeted therapy and radiotherapy through decreased expression of HER3 (ERBB3) and NF-κB p65. Taken together, we identified a new HIF-1α/NOX4 signal pathway which induced drug and radiation resistance in ovarian cancer. The finding may provide a new option to overcome the therapeutic resistance of ovarian cancer in the future.

Chromatin accessibility changes at intergenic regions are associated with ovarian cancer drug resistance

BackgroundResistance to DNA damaging chemotherapies leads to cancer treatment failure and poor patient prognosis. We investigated how genomic distribution of accessible chromatin sites is altered during acquisition of cisplatin resistance using matched ovarian cell lines from high grade serous ovarian cancer (HGSOC) patients before and after becoming clinically resistant to platinum-based chemotherapy.ResultsResistant lines show altered chromatin accessibility at intergenic regions, but less so at gene promoters. Clusters of cis-regulatory elements at these intergenic regions show chromatin changes that are associated with altered expression of linked genes, with enrichment for genes involved in the Fanconi anemia/BRCA DNA damage response pathway. Further, genome-wide distribution of platinum adducts associates with the chromatin changes observed and distinguish sensitive from resistant lines. In the resistant line, we observe fewer adducts around gene promoters and more adducts at intergenic regions.ConclusionsChromatin changes at intergenic regulators of gene expression are associated with in vivo derived drug resistance and Pt-adduct distribution in patient-derived HGSOC drug resistance models.

3D Clinorotation Affects Drug Sensitivity of Human Ovarian Cancer Cells

Numerous studies have reported that gravity alteration displays a remarkable influence on the biological processes of cancer cells. Therefore, gravity-related experiments have become a promising method of improving knowledge in the field of cancer biology and may be useful to detect remarkable implications for future cancer treatment. Taking this concept further, we used a 3D clinostat (3D-C; 10 rpm of changing direction) to analyse the effect of short-term exposure to simulated microgravity (sμg) on cisplatin sensitivity of drug resistant human ovarian cancer cells SKOV-3. This allowed us to investigate whether altered gravity affects drug susceptibility of cancer cells. Our studies revealed that sμg exposure affects SKOV-3 cells morphology and drug efficiency. We observed the altered cell shape, the presence of membrane blebbing and lamellipodia as well as the lack of filopodia when the cells had been cultured on 3D-C for 2 h. Cytotoxicity, cell death and cell cycle assays showed an increased percentage of apoptotic cells and G0/G1 cell cycle arrest after exposure on the 3D-C with cisplatin in comparison to the static control, non clinorotated cells. Cell proliferation and migration were altered after the exposure to sμg as well. Our studies suggest that the altered gravity conditions affected cellular mechanisms involved in cisplatin resistance, resulting in higher sensitivity of cancer cells to the chemotherapeutic. The investigation and clarification of these results may be a crucial step toward improving our understanding of the relationship between cellular resistance to chemotherapy and the response to altered gravitational conditions.

Dacomitinib improves chemosensitivity of cisplatin-resistant human ovarian cancer cells.

Drug resistance hinders effectiveness of human ovarian cancer (OC) therapies, such as cisplatin or paclitaxel therapy. Although dacomitinib, a novel anticancer agent is used against multiple types of cancers, such as non-small cell lung cancer, head and neck cancer, few studies report its effectiveness in drug-resistant human OC cells. In the present study, would healing, microplate spectrophotometer analysis, flow cytometry analysis, western blotting and Gene Expression Omnibus (GEO) analysis were used to detect the synergistic effect of dacomitinib and cisplatin in human OC SKOV-3 or OV-4 cells. Co-administration of dacomitinib and cisplatin significantly reduced viability and promoted cell apoptosis of drug resistant OC cells. In addition, dacomitinib increased Cadherin 1 (CDH1) levels and decreased P-glycoprotein (P-GP) levels in cisplatin-resistant OC cells. In addition, GEO analysis demonstrated that dacomitinib inhibited the epidermal growth factor receptor (EGFR) signaling pathway. In summary, dacomitinib improves chemosensitivity of cisplatin in human OC by regulating CDH1 and P-GP protein levels and inhibiting the EGFR signaling pathway.

The involvement of small heat shock protein in chemoresistance in ovarian cancer - in vitro study.

Ovarian cancer is the most deadly gynecologic malignancy worldwide. Although the primary response to chemotherapy is high, the majority of patients will develop resistance against applied treatment. In this study, we focused on resistance to cisplatin, a first-line drug used for the treatment of ovarian cancer. The mechanism of the resistance development process is widely described, but there is a lack of information about the involvement of members of small heat shock proteins (HSPs) and their transport via exosomes. In this study, we used two cell lines: A2780 and SKOV3, and their cisplatin-resistance variants: A2780 CDDP and SKOV3 CDDP. We have shown that the expression of three small HSPs (HSPB5, HSPB6, and HSPB8) in cisplatin-resistant cell lines differs from their sensitive counterparts. Further, we isolated exosomes and determined the small HSPs in their cargo. In A2780 WT we observed a low amount of HSPB5 and HSPB6. We did not observe the expression of small HSPs in the SKOV3 cell line in both sensitive and resistant variants. Our data suggest the involvement of small HSPs in drug resistance of ovarian cancer and their presence is not related to exosomal transport. Analysis of the biological consequences of the imbalance of small HSPs expression in cisplatin resistance needs further investigation.