Ovarian Tumor Model Research Articles

Enzyme-Activatable Interferon-Poly(α-amino acid) Conjugates for Tumor Microenvironment Potentiation.

Protein-polymer conjugation is a clinically validated approach to enhanced pharmacokinetic properties. However, the permanent attachment of polymers often leads to irreversibly reduced protein bioactivity and poor tissue penetration. As such, the use of protein-polymer conjugates for solid tumors remains elusive. Herein, we report a simple strategy using enzyme-activatable and size-shrinkable protein-polypeptide conjugates to overcome this clinical challenge. Briefly, a matrix metalloproteinase (MMP)-responsive peptide sequence is introduced between a therapeutic protein interferon (IFN) and a synthetic polypeptide P(EG3Glu)20. The resulting site-specific MMP-responsive conjugate, denoted as PEP20-M-IFN, can, therefore, release the attached P(EG3Glu)20 to achieve both protein activation and deep penetration into the tumor microenvironment (TME). Compared to a similarly produced nonresponsive analogue conjugate PEP20-IFN, our results find PEP20-M-IFN to show higher bioactivity in vitro, improved tumor retention, and deeper penetration in a MMP2-dependent manner. Moreover, systemic administration of PEP20-M-IFN shows outstanding antitumor efficacy in both OVCAR3 and SKOV3 ovarian tumor models in mice. This work highlights the releasable PEPylation strategy for protein drug potentiation at the TME and opens up new opportunities in clinics for the treatment of malignant solid tumors.

Recruitment of Intratumoral CD103+ Dendritic Cells by a CXCR4 Antagonist-Armed Virotherapy Enhances Antitumor Immunity

Intratumoral dendritic cells play an important role in stimulating cytotoxic T cells and driving antitumor immunity. Using a metastatic ovarian tumor model in syngeneic mice, we explored whether therapy with a CXCR4 antagonist-armed oncolytic vaccinia virus activates endogenous CD103+ dendritic cell responses associated with the induction of adaptive immunity against viral and tumor antigens. The overall goal of this study was to determine whether expansion of CD103+ dendritic cells by the virally delivered CXCR4 antagonist augments overall survival and in situ boosting with a tumor antigen peptide-based vaccine. We found that locoregional delivery of the CXCR4-A-armed virus reduced the tumor load and the immunosuppressive network in the tumor microenvironment, leading to infiltration of CD103+ dendritic cells that were capable of phagocytic clearance of cellular material from virally infected cancer cells. Further expansion of tumor-resident CD103+ DCs by injecting the FMS-related tyrosine kinase 3 ligand, the formative cytokine for CD103+ DCs, provided a platform for a booster immunization with the Wilms tumor antigen 1 peptide-based vaccine delivered intraperitoneally with polyriboinosinic:polyribocytidylic acid as an adjuvant. The vaccine-induced antitumor responses inhibited tumor growth and increased overall survival, indicating that expansion of intratumoral CD103+ dendritic cells by CXCR4-A-armed oncovirotherapy treatment can potentiate in situ cancer vaccine boosting.

Abstract 1661: Molecular modeling of an emerging chemopreventive agent wedelolactone against multiple targets in oncogenetic pathway

Abstract Targeting multiple pathways involved either in drug resistance or/and in intact cancer cell survival are promising strategies. Epigenetics is being immensely recognized as exciting frontier in cancer research. Kinases have direct and indirect epigenetic impacts. Kinases are among the popular discovery targets and kinase inhibitors represent one of the key classes of drugs in the last two decades of oncology research. Braf kinases and cyclin dependent protein kinases exert their role as key mediators of signal transduction, apoptosis and overall intercellular communications. Topoisomerase II inhibitors including etoposide, tinoposide are important drugs used in breast cancer, sarcomas and lymphomas while inhibitors of dihydrofolatereductase, a key enzyme in the synthesis of DNA, are also clinically useful antagonists/inhibitors of ERα and aromatases have the potential to be therapeutically significant towards estrogen mediated breast, ovarian and uterine (endometrium) cancers. Wedelolactone is emerging as a small molecule with cytotoxic potential. Auto Dock Vina program was used for docking study .Affinity of best docked pose of ligand and protein target complex was determined by E-value (Kcal/mol).3D-structures of target proteins were taken from http://www.rcsb.org/pdb/home/home.do. The target proteins include dihydrofolatereductase, braf kinase, cyclin dependent protein kinase, nuclear factor kappa b, topoisomerase II, aromatase, estrogen receptor alpha and alkaline phosphatase (PDB ID: 2C2T, PDB ID: 3Q4C, PDB ID: 1HCL, PDB ID: 1NFK, PDB ID: 5B58, PDB ID: 3S7S, PDB ID: 3ERT, PDB ID: 1ANI). The structures (wedelolactone and standard drugs) were downloaded in.xml format and converted to PDB format via Open Babel JUI software. PDB form of wedelolactone, EGCG, reference drugs and target proteins were converted to PDBQT via AutoDock Tools (Version1.5.6 Sep_17_14). Discovery studio visualizer was used for post docking interactions for number of Hydrogen bonds (classical and non-classical), π-π bonds and other hydrophobic interactions by binding amino acid residues. The cell survival fraction was determined using the MTT reduction assay. The order of binding affinity of wedelolactone was found as: estrogen receptor alpha >dihydrofolate reductase>braf kinase > topoisomerase II > CDPK > aromatase > alkaline phosphatase >Nfkb. We identified wedelolactone as a cytotoxic agent in breast and ovarian tumor models with the potential to inhibit multiple targets in oncogenic as depicted through its In-silico study. Further research should be encouraged to investigate different aspects of wedelolactone. Note: This abstract was not presented at the meeting. Citation Format: sadia sarwar, Tauqeer Ahmed, Jun Q. Yu, Fazlul Huq. Molecular modeling of an emerging chemopreventive agent wedelolactone against multiple targets in oncogenetic pathway [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1661.

Nanoengineering of Mesenchymal Stem Cells via Surface Modification for Efficient Cancer Therapy

AbstractMesenchymal stem cells (MSCs) can be used for tumor‐specific delivery of small molecular weight anticancer drugs by using nanoparticle‐encapsulated forms of the drugs. Current approaches to incorporate nanoparticles in MSCs rely on nonspecific endocytosis of nanoparticles or their conjugation to the cell surface via endogenous amines and thiols. These methods result in sub‐optimal drug loading, which hinders the widespread application of MSCs as drug carriers. An advanced nanoengineering strategy is reported here that involves generation of MSCs expressing azide functional groups on their surface and conjugation of dibenzyl cyclooctyne‐functionalized nanoparticles to the azide groups using copper‐free click chemistry. This novel strategy significantly improves the payload capacity of MSCs (≈48 pg of paclitaxel (PTX) per cell) relative to that reported previously (<1–20 pg per cell), without affecting their native phenotype. In vivo, the nanoengineered MSCs significantly inhibit tumor growth (p < 0.05) and improve survival (p < 0.0001) compared to free or nanoparticle encapsulated PTX and Abraxane in an orthotopic ovarian tumor model. In summary, the nanoengineering strategy reported here allows for improved delivery and anticancer efficacy of conventional chemotherapeutic agents using MSCs as drug carriers.

IR780-loaded folate-targeted nanoparticles for near-infrared fluorescence image-guided surgery and photothermal therapy in ovarian cancer.

Background and purpose: Surgery is regarded as the gold standard for patients with advanced ovarian cancer. However, complete surgical removal of tumors remains extremely challenging; fewer than 40% of patients are cured. Here, we developed a new modality of theranostics for ovarian cancer based on a near-infrared light-triggered nanoparticle.Methods: Nanoparticles loading IR780 iodide on base of folate modified liposomes were prepared and used for theranostics of ovarian cancer. Tumor targeting of FA-IR780-NP was evaluated in vitro and in an ovarian xenograft tumor model. A fluorescence stereomicroscope was applied to evaluate the tumor recognition of FA-IR780-NP during surgery. FA-IR780-NP mediated photothermal therapy effect was compared with other treatments in vivo.Results: FA-IR780-NP was demonstrated to specifically accumulate in tumors. IR780 iodide selectively accumulated in tumors; the enhanced permeability and retention effect of the nanoparticles and the active targeting of folate contributed to the excellent tumor targeting of FA-IR780-NP. With the aid of tumor targeting, FA-IR780-NP could be used as an indicator for the real-time delineation of tumor margins during surgery. Furthermore, photothermal therapy mediated by FA-IR780-NP effectively eradicated ovarian cancer tumors compared with other groups.Conclusion: In this study, we present a potential, effective approach for ovarian cancer treatment through near-infrared fluorescence image-guided resection and photothermal therapy to eliminate malignant tissue.

Cowpea Mosaic Virus Promotes Anti-Tumor Activity and Immune Memory in a Mouse Ovarian Tumor Model.

Cowpea mosaic virus (CPMV) is a promising platform nanotechnology with applications as a cancer therapeutic. To understand the therapeutic potential of CPMV in more detail, its antitumor mechanisms are investigated using a syngeneic immunocompetent murine orthotopic ovarian cancer model (ID8-Defb29/Vegf-A). CPMV treatment in situ promotes tumor regression and prevents tumor recurrence. Although CPMV does not kill tumor cells directly, it promotes an intra-tumoral cytokine response which induces pre-existing myeloid cells to break immunotolerance and initiate antitumor responses. The upregulation of interleukin-6 and interferon-γ as well as the downregulation of IL-10 and transforming growth factor β are observed, associated with activation and repolarization of tumor-associated macrophages and neutrophils to an anti-tumor phenotype. Furthermore, the in situ administration of CPMV recruits dendritic cells and natural killer cells to the tumor site, and induces the expression of costimulatory molecules on CD11b- myeloid cells. By converting immunosuppressive myeloid cells into potent antigen-presenting cells, in situ CPMV treatment significantly improves effector and memory CD4+ and CD8+ T cell responses and promoted systemic tumor-specific cytotoxic CD8+ T cell activity. CPMV in situ immunotherapy induces significant tumor control in an aggressive ovarian tumor model by coordinating innate and adaptive immune responses involving neutrophils, macrophages, and T cells.

Surface-engineered polyethyleneimine-modified liposomes as novel carrier of siRNA and chemotherapeutics for combination treatment of drug-resistant cancers

Modification of nanoparticle surfaces with PEG has been widely considered the gold standard for many years. However, PEGylation presents controversial and serious challenges including lack of functionality, hindered cellular interaction, allergic reactions, and stimulation of IgM production after repetitive dosing that accelerates blood clearance of the nanoparticles. We report the development of novel liposomal formulations surface-modified with a low molecular weight, branched polyethyleneimine (bPEI)–lipid conjugate for use as an alternative to PEG. The formulations had very good stability characteristics in ion- and protein-rich mediums. Protein adsorption onto the liposomal surface did not interfere with the cellular interaction. bPEI-modified liposomes (PEIPOS) showed enhanced association with three different cell lines by up to 75 times compared to plain or PEGylated liposomes and were without carrier toxicity. They also penetrated the deeper layers of 3D spheroids. Encapsulating paclitaxel (PTX) into PEIPOS did not change its main mechanism of action. PEIPOS complexed and intracellularly delivered siRNAs and downregulated resistance-associated proteins. Finally, tumor growth inhibition was observed in a mouse ovarian xenograft tumor model, without signs of toxicity, in animals treated with the siRNA/PTX co-loaded formulation. These complex-in-nature but simple-in-design novel liposomal formulations constitute viable and promising alternatives with added functionality to their PEGylated counterparts.

CD44-Targeting PLGA Nanoparticles Incorporating Paclitaxel and FAK siRNA Overcome Chemoresistance in Epithelial Ovarian Cancer.

Chemotherapy is commonly used in the treatment of ovarian cancer, yet most ovarian cancers harbor inherent resistance or develop acquired resistance. Therefore, novel therapeutic approaches to overcome chemoresistance are required. In this study, we developed a hyaluronic acid-labeled poly(d,l-lactide-co-glycolide) nanoparticle (HA-PLGA-NP) encapsulating both paclitaxel (PTX) and focal adhesion kinase (FAK) siRNA as a selective delivery system against chemoresistant ovarian cancer. The mean size and zeta potential of the HA-PLGA-NP were 220 nm and -7.3 mV, respectively. Incorporation efficiencies for PTX and FAK siRNA in the HA-PLGA-NPs were 77% and 85%, respectively. HA-PLGA-NP showed higher binding efficiency for CD44-positive tumor cells as compared with CD44-negative cells. HA-PLGA (PTX+FAK siRNA)-NP caused increased cytotoxicity and apoptosis in drug-resistant tumor cells. Treatment of human epithelial ovarian cancer tumor models HeyA8-MDR (P < 0.001) and SKOV3-TR (P < 0.001) with HA-PLGA (PTX+FAK siRNA)-NP resulted in significant inhibition of tumor growth. Moreover, in a drug-resistant, patient-derived xenograft (PDX) model, HA-PLGA (PTX+FAK siRNA)-NP significantly inhibited tumor growth compared with PTX alone (P < 0.002). Taken together, HA-PLGA-NP acts as an effective and selective delivery system for both the chemotherapeutic and the siRNA in order to overcome chemoresistance in ovarian carcinoma.Significance: These findings demonstrate the efficacy of a novel, selective, two-in-one delivery system to overcome chemoresistance in epithelial ovarian cancer. Cancer Res; 78(21); 6247-56. ©2018 AACR.

Simultaneous multimodal imaging and photothermal therapy via renal-clearable manganese-doped copper sulfide nanodots

Ultrasmall sized renal clearable nanoparticles are promising nanomaterials for cancer diagnostic and therapeutic treatments owing to their efficient clearance characteristics and feasible therapeutic effects. However, to integrate multifunction and keep ultrasmall size (<6nm) of the nanoparticles simultaneously is a great challenge for the synthesis of renal clearable nanoparticles. In this manuscript, we engineered an ultrasmall (4.95nm) triple-modal imaging platform by chelate-free doping of manganese2+ (Mn2+) and 68gallium3+ (Ga3+) into the matrix of copper sulfide (CuS) nanodots (NDs) using bovine serum albumin (BSA) as the synthetic template. The resulting Mn/68Ga-CuS@BSA nanodots proved to be an excellent contrast agent for magnetic resonance (MR), positron emission tomography (PET), and photoacoustic (PAT) imaging. All three imaging modalities confirmed that Mn/Ga-CuS@BSA NDs could be efficiently cleared via the renal-urinary route of intravenous injection, therefore effectively minimized its toxicity to non-tumor organs. In addition, Mn/68Ga-CuS@BSA NDs functioned as a potent mediator for the near-infrared (NIR) laser-induced photothermal therapy (PTT), and significantly suppressed tumor growth in the SKOV-3 ovarian tumor model. Our results demonstrated that renal clearable inorganic nanoparticles with multiple imaging and therapeutic capabilities are robust platforms to battle cancer.

Dihydroartemisinin increases gemcitabine therapeutic efficacy in ovarian cancer by inducing reactive oxygen species.

Ovarian cancer is the major cause of death in women gynecological malignancy and gemcitabine (GEM) is commonly used in related chemotherapy. However, more than 90% GEM is catalyzed into an inactive metabolite 2'-deoxy-2',2'-difluorouridine by stromal and cellular cytidine deaminase (CDA). Dihydroartemisinin (DHA), which possesses an intramolecular endoperoxide bridge, could be activated by heme or ferrous iron to produce reactive oxygen species (ROS). The excess ROS generation will excite expression of heme oxygenase-1 and suppress CDA expression. Under low CDA expression, the inactivation of GEM is decreased in turn to exert excellent therapeutic efficiency. Herein, we first studied the ROS generation by DHA in vitro with A2780 cells by means of flow cytometry and confocal laser scanning microscopy. Furthermore, cytotoxicity assay in vitro showed that DHA + GEM had synergistic effect, with molar ratio of DHA and GEM at 10. Eventually, in A2780 ovarian cancer xenograft tumor model, DHA + GEM exhibited significant antitumor efficiency with lower blood toxicity than GEM alone. Noteworthy, the combination treatment group completely eliminated the tumors on day 14.

Abstract A17: Bromodomain and extraterminal proteins regulate PD-L1/PD-1 signaling in breast cancer.

Abstract Introduction: Breast tumors are massively infiltrated immune cells of both lymphoid and myeloid origin that exert an anti-tumor pressure to limit cancer progression. Tumor cells can escape immune clearance by expressing several molecules that constitute the immune checkpoints. Antibodies targeting the PD-L1/PD-1 pathway are being evaluated clinically for several cancers and show early success. However, the realization that not all patients respond well to immunotherapy suggests other modalities could be combined to improve efficacy. The bromodomain and extraterminal (BET) proteins BRD2, BRD3 and BRD4 are epigenetically acting co-regulators of transcription, and are critical for cancer proliferation and metastasis. Little is known about the molecular mechanisms that regulate PD-L1 and PD-1 expression. BRD4 is known to bind directly to the PD-L1 promoter; its targeting suppresses PD-L1 expression while increasing CD8+ T cell activity to limit progression in ovarian tumor models. Hypotheses: BET proteins regulate PD-L1 and PD-L2 expression in breast tumors and PD-1 in CD8+ T cells. BET proteins are effectors of proinflammatory cytokine signaling, regulating PD-L1/PD-1 signaling in the breast tumor microenvironment. Methods: We used different cellular models of breast cancer. We also collected human peripheral blood mononuclear cells matched with mammary adipose tissue from mammoplasty patients at Boston Medical Center to assess expression of immune checkpoint proteins in the breast microenvironment. Pan-BET protein inhibition was performed by JQ1 treatment. Alternatively, BET proteins were individually and specifically depleted by siRNA. PD-1 and PD-L1 expression were determined by qRT-PCR and flow cytometry. Cytokines and chemokines present in the blood or secreted by the mammary adipose tissue were profiled by multiplexed antibody capture assay. Results: BET proteins regulate PD-L1 expression by breast cancer cells. BRD2 and BRD4 expression correlate with PD-1 in activated circulating T cells and BET inhibition ablates activation-induced PD-1 expression. Co-culture of breast cancer cells with activated circulating T cells enhances expression of immune checkpoint proteins in both compartments. BET protein inhibition rescues these phenotypes. Conditioned media from mammary adipose tissue increase PD-1 in PBMCs. Peripheral and mammary adipose tissue inflammatory signatures associate with PD-1 expression in PBMCs, suggesting that local inflammation in the breast tumor microenvironment contributes to diminishing anti-tumor immune responses. Conclusion: BET proteins regulate the PD-L1/PD-1 pathway in breast cancer. Next-generation BET protein inhibitors may combine well with PD-1 or PD-L1-targeted immunotherapies in difficult-to-treat cancers. Personalized BET profiles could inform individual patient responses to BET proteins and immune checkpoint inhibitors. Therapeutic approaches that treat the microenvironment should be leveraged to maximize efficacy of checkpoint inhibitor approaches for specific cancers. Citation Format: Guillaume Andrieu, Gerald V. Denis. Bromodomain and extraterminal proteins regulate PD-L1/PD-1 signaling in breast cancer. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology and Immunotherapy; 2017 Oct 1-4; Boston, MA. Philadelphia (PA): AACR; Cancer Immunol Res 2018;6(9 Suppl):Abstract nr A17.

Abstract B29: Cisplatin is pro-immunogenic and promotes intrinsic and reactive immune suppression in inflamed and noninflamed ovarian cancer mouse models

Abstract Objective: Front-line therapy in ovarian cancer combines platinum/taxane chemotherapy and surgery. We studied here the role of cisplatin on PDL1 biology in vitro and in vivo in new ovarian tumor models. We also explored the in vivo efficacy of PDL1 blockade in combination with cisplatin. Methods: We analyzed in vitro the time and dose dependent effect of cisplatin on PD-L1 expression on human and murine cell lines with varying susceptibility to the drug. Cisplatin-induced molecular changes were detected via unbiased transcriptomics (RNAseq, Illumina). Mechanisms of tumor cell-intrinsic immune resistance were explored in a series of murine ovarian cancer cell lines with defined genetic traits, including conditional mutations in either Kras (activation), Pten (deletion), or both. The effect of cisplatin in combination with PDL1 blockade was tested in vivo in two syngeneic OC models (2F8 and 2F8cis) that mirror platinum sensitive and moderately resistant tumors, respectively. These models have different inflammation at baseline, with 2F8cis tumors being more inflamed (increased tumor-infiltrating lymphocytes [TILs]-increased necrosis) than 2F8 tumors. Results: Exposure of murine and human ovarian cancer cell lines to IC50 cisplatin triggers increased PD-L1 expression in a time- and dose-dependent manner. The effect is partly mediated by Erk/MAPK pathway, as MEK inhibitor AZD6244 inhibits both the baseline and the cisplatin-induced PD-L1 expression. Gene expression analyses (RNASeq) demonstrate that cisplatin upregulates a number of genes that contribute to tumor immunogenicity and increased tumor-reactive CD8 T cells. The cisplatin-induced proinflammatory effect was also observed in vivo, predominantly in platinum-sensitive 2F8 tumors that show increased T-cell infiltration of both CD8 and FoxP3 Tregs. Tumor PDL1 gene expression is also increased upon cisplatin treatment, as detected by qPCR. Mice with platinum-sensitive (2F8) tumors showed increased survival in response to cisplatin alone and anti-PDL1 alone, compared to the combination, which was not significantly different than control treated hosts. In contrast, cisplatin/anti-PDL1 treatment combination triggered significantly longer survival in mice with moderately resistant (2F8cis) tumors. Conclusion: Overall, these results suggest that exposure to cisplatin has the potential to increase tumor immunogenicity and recognition by CD8 TILs while also triggering immune evasion through PD-L1 upregulation either directly (through tumor cell-intrinsic mechanisms) or indirectly via adaptive immune resistance due to IFNγ secreting TILs. Inflamed and noninflamed respond to anti-PD-L1 alone. although the effect is generally modest. Inflamed tumors with moderate platinum resistance respond to cisplatin/anti-PDL1 combination, and the effect is better than for each agent alone. These results provide the rationale for the design of combination therapies that take into consideration tumor inflammation at baseline and drug-induced immunogenic effects. Citation Format: Shannon Grabosch, Mirna Bulatovic, Feitianzhi Zeng, Tianzhou Ma, Lixin Zhang, Malcolm Ross, Joan Brozick, George Tseng, Esther Elishaev, Robert P. Edwards, Anda Vlad. Cisplatin is pro-immunogenic and promotes intrinsic and reactive immune suppression in inflamed and noninflamed ovarian cancer mouse models. [abstract]. In: Proceedings of the AACR Conference: Addressing Critical Questions in Ovarian Cancer Research and Treatment; Oct 1-4, 2017; Pittsburgh, PA. Philadelphia (PA): AACR; Clin Cancer Res 2018;24(15_Suppl):Abstract nr B29.

Abstract 3589: Preclinical evaluation of mesothelin-specific T cell receptor (TCR) fusion constructs (TRuC™s) utilizing the signaling power of the complete TCR complex: A new opportunity for solid tumor therapy

Abstract Despite success in treating hematological malignancies, T cells expressing chimeric antigen receptors (CARs) have shown poor efficacy in solid tumor indications. The poor utility of CARs is thought to be due to a number of factors including T cell exhaustion and a lack of persistence. Utilizing only the CD3ζ chain of six distinct T cell receptor subunits in combination with a costimulatory domain, the CARs bypass the natural TCR signaling complex. The failure to initiate and harness a complete TCR response is arguably a primary underlying factor preventing CAR-T cell success in solid tumor indications. Here, we present a novel T cell engineering platform: T Cell Receptor Fusion Constructs (TRuC™s), which target tumors in a MHC non-restricted fashion. Unlike CARs, the constructs integrate into the TCR complex, harnessing the full potential of natural T cell activation, effector function and regulation. Here we demonstrate preclinical evidence underscoring the efficacy of TRuC™-T cells re-programmed to target the solid tumor antigen mesothelin (TC-210). TC-210 showed robust anti-tumor activity in vitro and in vivo in mesothelioma, lung and ovarian tumor models. Surprisingly, this activity was associated with less proinflammatory cytokine production compared to CAR-T cells with the same binder and was supported by strong TRuC™-T cell persistence. Taken together, mesothelin targeting TRuC™-T cells have potent anti-tumor activity and present a promising approach for treating mesothelin-expressing solid tumors. Citation Format: Jian Ding, Ekta Patel, Patrick Tavares, Justin Quinn, Rashmi Choudhary, Bonnie Le, Olga Kiner, Solly Weiler, Daniel R. Getts, Patrick Baeuerle, Robert Hofmeister. Preclinical evaluation of mesothelin-specific T cell receptor (TCR) fusion constructs (TRuC™s) utilizing the signaling power of the complete TCR complex: A new opportunity for solid tumor therapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3589.

Abstract 1716: The PARP inhibitor rucaparib activates the STING pathway and enhances antitumor responses of immune checkpoint inhibitors in BRCA deficient syngeneic models

Abstract Background: The poly(ADP ribose) polymerase (PARP) inhibitor rucaparib effectively kills homologous recombination (HR) deficient cells through impeding DNA repair that leads to DNA damage, apoptosis, and cell death. Detection of cytosolic DNA by the stimulator of interferon genes (STING) pathway mediates type I interferon (IFN) production and activates the immune system. Following rucaparib treatment, the accumulation of damaged DNA in HR impaired tumors may elicit an immune response through STING signaling, and enhance rucaparib activity as a single agent or in combination with immune checkpoint blockade. To test this hypothesis, rucaparib efficacy and mechanism of action were evaluated using BRCA deficient syngeneic ovarian tumor models. Results: Single agent rucaparib showed potent antitumor activity in the BRCAmut BrKras and ID8B3.15 models. In the BrKras model, rucaparib treatment resulted in complete regression and prevented tumor formation upon re-challenge. However, this efficacy was abolished with anti-CD8 but not anti-CD4 depletion. CD8 tumor infiltrating lymphocytes (TILs) but not CD4 TILs increased with rucaparib exposure, and expression profiling of rucaparib treated tumors showed activation of the type I IFN pathway. In vitro PCR assays identified several targets that were upregulated upon rucaparib treatment including Ccl5 and Cxcl10. Notably, increased Ccl5 and Cxcl10 levels were observed in BRCAmut cells but not in BRCAwt cells, and knockdown of the STING pathway genes MB21D1, IRF3, TBK1, and STING (TMEM173) abrogated rucaparib induction of Ccl5 and Cxcl10. Furthermore, rucaparib showed transcriptional activation of IFN type I signaling in BRCAmut reporter cells expressing the IFN stimulated response element consensus sequence driving luciferase expression, but required 6-fold higher concentrations in BRCAwt reporter cells. Similarly, a 10-fold higher rucaparib dose was needed to inhibit proliferation of BRCAwt cells compared to BRCAmut cells in a cell viability assay. Consistent with the in vitro results, rucaparib combined with anti-programmed death 1 or anti-programmed death ligand 1 therapy improved the survival and augmented antitumor responses in BRCA deficient syngeneic tumor models. Conclusions: Rucaparib treatment in HR deficient cells, and at a higher concentration in HR proficient cells, triggers type I IFN signaling through the STING pathway, which participates in the single agent efficacy of rucaparib and enhances the combination of rucaparib and checkpoint inhibitors in syngeneic models. These findings provide further evidence supporting the rationale for combining rucaparib with checkpoint therapy for the treatment of patients with HR defective cancers. Citation Format: Minh Nguyen, Liliane Robillard, Kevin K. Lin, Thomas C. Harding, Andrew D. Simmons. The PARP inhibitor rucaparib activates the STING pathway and enhances antitumor responses of immune checkpoint inhibitors in BRCA deficient syngeneic models [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1716.

Development of an Efficient Dual-Action GST-Inhibiting Anticancer Platinum(IV) Prodrug.

The cytotoxicity of cisplatin (cDDP) is enhanced when co-administered with ethacrynic acid (EA), a glutathione S-transferase (GST) inhibitor. A PtIV -EA conjugate containing a cDDP core and two axial ethacrynate ligands (compound 1) was shown to be an excellent inhibitor of GST, but did not readily release a PtII species to exert a synergistic cytotoxic effect. In this study, a redesigned PtIV construct composed of a cDDP core with one axial ethacrynate ligand and one axial hydroxido ligand (compound 2) was prepared and shown to overcome the limitations of compound 1. The EA ligand in 2 is readily released in vitro together with a cytotoxic PtII species derived from cisplatin, working together to inhibit cell proliferation in cDDP-resistant human ovarian cancer cells. The in vitro activity translates well in vivo with 2, showing effective (∼80 %) inhibition of tumor growth in a human ovarian carcinoma A2780 tumor model, while showing considerably lower toxicity than cisplatin, thus validating the new design strategy.

Biodistribution and Clearance of Stable Superparamagnetic Maghemite Iron Oxide Nanoparticles in Mice Following Intraperitoneal Administration.

Nanomedicine is an emerging field with great potential in disease theranostics. We generated sterically stabilized superparamagnetic iron oxide nanoparticles (s-SPIONs) with average core diameters of 10 and 25 nm and determined the in vivo biodistribution and clearance profiles. Healthy nude mice underwent an intraperitoneal injection of these s-SPIONs at a dose of 90 mg Fe/kg body weight. Tissue iron biodistribution was monitored by atomic absorption spectroscopy and Prussian blue staining. Histopathological examination was performed to assess tissue toxicity. The 10 nm s-SPIONs resulted in higher tissue-iron levels, whereas the 25 nm s-SPIONs peaked earlier and cleared faster. Increased iron levels were detected in all organs and body fluids tested except for the brain, with notable increases in the liver, spleen, and the omentum. The tissue-iron returned to control or near control levels within 7 days post-injection, except in the omentum, which had the largest and most variable accumulation of s-SPIONs. No obvious tissue changes were noted although an influx of macrophages was observed in several tissues suggesting their involvement in s-SPION sequestration and clearance. These results demonstrate that the s-SPIONs do not degrade or aggregate in vivo and intraperitoneal administration is well tolerated, with a broad and transient biodistribution. In an ovarian tumor model, s-SPIONs were shown to accumulate in the tumors, highlighting their potential use as a chemotherapy delivery agent.

Structural characterization and antitumor activity of a novel Se-polysaccharide from selenium-enriched Cordyceps gunnii.

Selenium (Se) has been recognized as an essential element. Animals and humans absorb and retain more organic Se than inorganic Se. Bio-transformation is a major approach to transform inorganic Se to organic Se. Cordyceps gunnii (C. gunnii), well known as a Chinese edible and medicinal fungus, has a variety of biological functions. C. gunnii and selenium were combined through liquid-fermentation to obtain selenium-enriched C. gunnii. A novel Se-polysaccharide (SeCPS-II) was extracted from selenium-enriched C. gunnii. The molecular weight, sugar content and selenium content of SeCPS-II were 4.12 × 103 kDa, 17.89 μg g-1 and 90.75%, respectively. The structure of SeCPS-II was characterized using FT-IR, NMR, GC and GC-MS studies. SeCPS-II was composed of pyranose, which contained α-l-rhamnose, α-d-mannose, α-d-glucose and β-d-galactose at a ratio of 4.33 : 12.62 : 27.50 : 18.99. SeCPS-II contained α-(1 → 4)-d-glucose, α-(1 → 3)-d-glucose, β-(1 → 6)-d-galactose, α-(1 → 6)-d-mannose, and α-(1 → 4)-l-rhamnose, and the main chain was composed of α-(1 → 4)-d-glucose. An MTT assay indicated that SeCPS-II could influence the cell viability of SKOV-3 cells, H1299 cells and HepG2 cells in a dose-dependent and time-dependent manner. Morphologic changes (AO/EB staining and DAPI staining) and an MMP assay (JC-1 staining) of SKOV-3 cells treated with SeCPS-II were performed to research the antitumor activity of SeCPS-II. SeCPS-II could significantly induce apoptosis in SKOV-3 cells with typical apoptotic characteristics. To study the mechanism, the expressions of caspase-3, caspase-9, PARP, p53, Bax and Bcl-2 in SKOV-3 were studied via western blotting. SeCPS-II can stimulate the apoptosis of SKOV-3 cells through the p53-Bax-caspase pathway. Animal experiments revealed that SeCPS-II inhibits tumors within an ovarian tumor model rat, modeled with SKOV-3 cells. All the results indicated that selenium-enriched C. gunnii can be used to develop new selenium-containing additives.

Tumor immunity flounders without FIP200

Tumor Immunology The tumor microenvironment impairs the function of effector and memory T cells, but its effect on naive T cells is not well understood. Xia et al. show that the autophagy component FIP200 is critical to promoting antitumor responses mediated by T cells. Ovarian cancer patients and mouse tumor models had higher rates of apoptosis and impaired autophagy in naive T cells. This was associated with the selective loss of FIP200, which disrupted the balance of pro- and anti-apoptotic factors. Tumor-derived lactate suppressed FIP200 expression, thus causing enhanced apoptosis and attenuated antitumor responses. Sci. Immunol. 2 , eaan4631 (2017).

Tumor Microenvironment and Models of Ovarian Cancer: The 11th Biennial Rivkin Center Ovarian Cancer Research Symposium.

The aim of this study was to review the latest research advances on the topics of the ovarian cancer tumor microenvironment and models of ovarian cancer. In September 2016, a symposium of the leaders in the field of ovarian cancer research was convened to present and discuss current advances and future directions in ovarian cancer research. One session was dedicated to Tumor Microenvironment and Models of Ovarian Cancer, and included a keynote presentation from Anil Sood, MD, and an invited oral presentation from David Huntsman, MD. Eight additional oral presentations were selected from abstract submissions. Twenty-nine abstracts were presented in poster format and can be grouped into the categories of stromal cells in the microenvironment, immune cells in the microenvironment, epithelial-mesenchymal transition and metastasis, metabolomics, and model systems including spheroids, murine models, and other animal models. Rapid advances continue in our understanding of the influence of the tumor microenvironment on ovarian cancer progression and metastasis. Vascular endothelial cells, stromal cells, and immune cells all modulate epithelial tumor cell biology and therefore serve as potential targets for improved treatment responses either in conjunction with or instead of current treatment modalities. Characterization of the underlying genetic alterations in both the tumor cells and surrounding microenvironment cells enhances our understanding of tumor biology. Model systems including both in vitro and in vivo approaches allow novel advances. Technological advances including sequencing strategies, use of mass spectrometry for metabolomics and other studies, and bioengineering approaches all complement conventional methodologies to push forward our understanding and ultimately the treatment of ovarian cancer.

Pre-clinical evaluation of a themosensitive gel containing epothilone B and mTOR/Hsp90 targeted agents in an ovarian tumor model

Despite clinical remission of epithelial ovarian cancer (EOC) after surgical resection and first-line chemotherapy, about 60% of patients will re-develop peritoneal metastasis and about 50% will relapse with chemoresistant disease. Clinical studies suggest that intra-peritoneal (i.p.) chemotherapy effectively treats residual EOC after cyto-reduction by gaining direct access into the peritoneal cavity, enabling elevated drug levels versus intravenous (i.v.) injection. However, chemoresistant disease is still problematic. To overcome resistance against microtubule stabilizing agents such as taxanes, epothilone B (EpoB) has merit, especially in combination with molecular targeted agents that inhibit heat shock protein 90 (Hsp90) and/or mammalian target of rapamycin (mTOR). In this paper, we report on the successful loading and solubilization of EpoB in a poly(d,l-lactic-co-glycolic acid)-block-poly(ethylene glycol)-block-poly(d,l-lactic-co-glycolic acid) (PLGA-b-PEG-b-PLGA) thermosensitive gel (g-E). Further, we report on successful co-loading of 17-AAG (Hsp90) and rapamycin (mTOR) (g-EAR). After i.p. injection in mice, g-EAR showed gelation in the peritoneum and sustained, local-regional release of EpoB, 17-AAG, and rapamycin. In a luciferase-expressing ES-2 (ES-2-luc) ovarian cancer xenograft model, single i.p. injections of g-E and g-EAR delayed bioluminescence from metastasizing ES-2-luc cells for 2 and 3weeks, respectively, despite fast drug release for g-EAR in vivo versus in vitro. In summary, a PLGA-b-PEG-b-PLGA sol-gel has loading and release capacities for EpoB and its combinations with 17-AAG and rapamycin, enabling a platform for i.p. delivery, sustained multi-drug exposure, and potent antitumor efficacy in an ES-2-luc, ovarian cancer i.p. xenograft model.