bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 2156: Targeting estrogen receptor mutations for treatment of endocrine therapy resistance in breast cancer
Authors: Alluri Prasanna G. , Larios Jose , Malik Rohit , Rae James , Chiinaiyan Arul M.
Published: July 2016

Approximately 70% of all breast cancers express the Estrogen Receptor (ER) and inhibition of the ER signaling remains the mainstay of systemic therapy in such cancers. The advent of endocrine therapies (ET) have contributed significantly to reduction in the incidence, recurrence and mortality associated with breast cancer. However, approximately one-third of patients with ER-positive breast cancer do not benefit from adjuvant ET, and nearly all patients with ER-positive metastatic breast cancer (MBC) ultimately become refractory to all known ETs. While several mechanisms of resistance to ET have been proposed, there are currently no clinically effective treatments for ET resistance. Thus, ET resistance remains a major obstacle to the effective treatment of a significant subset of ER-positive breast cancers. We have recently reported recurrent somatic mutations in the ligand binding domain (LBD) of the Estrogen Receptor gene (ESR1) in metastatic breast cancer patients with a history of prior treatment with ET. The ESR1 mutations result in constitutive activation of the ER and confer ET resistance when ectopically expressed in ER-positive cell lines. In an attempt to develop novel therapeutic strategies to treat ET-resistant breast cancer, we have evaluated OTX015, a clinical bromodomain and extraterminal (BET) domain inhibitor, in pre-clinical models of ET resistance due to ESR1 mutations. Our studies suggest that BET inhibition may serve as an attractive strategy for treatment of ET resistance due to ESR1 mutations.

Citation Format: Prasanna G. Alluri, Jose Larios, Rohit Malik, James Rae, Arul M. Chiinaiyan. Targeting estrogen receptor mutations for treatment of endocrine therapy resistance in breast cancer. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 2156.
Abstract 2171: Beta-hairpin peptide hydrogels as scaffolds for 3D high-throughput cancer drug discovery
Authors: Worthington Peter , Drake Katherine M. , Napper Andrew D. , Pochan Darrin J. , Langhans Sigrid A.
Published: July 2016

High-throughput screening (HTS) remains a promising initial step in discovering novel lead compounds for cancer therapy, but its value is limited by the poor predictability of clinical effectiveness of candidate compounds. One compelling reason for this lack of reliability to predict in vivo efficacy is the fact that most HTS is done using traditional two-dimensional (2D) cell cultures. While 2D cultures are convenient and can easily be automated, compelling evidence suggests that cells cultured in these non-physiological conditions differ from cells grown in the more in vivo like three-dimensional (3D) systems that better mimic microenvironments. Thus, a 3D culture model is expected to be a better platform for cancer drug discovery. Here we demonstrate the feasibility of a beta-hairpin hydrogel as a novel platform for 3D HTS. MAX8 forms a bonded fibrillar hydrogel network under physiological conditions and moreover, due to its unique shear-thinning abilities can be incorporated into standard HTS equipment. MAX8 is biocompatible with a variety of cancer cell lines, can be tuned for porosity, permeability and stiffness, and can be functionalized with common and cell-type specific ligands to more closely resemble an in vivo-like tumor environment. Using medulloblastoma cells that are known to display more immature, tumor-like features in 3D and that were automatically dispensed onto 384-well plates we have developed a robust assay to measure cell viability on a HTS platform that will have broad applicability for other cancers for which 3D HTS is desirable.

Citation Format: Peter Worthington, Katherine M. Drake, Andrew D. Napper, Darrin J. Pochan, Sigrid A. Langhans. Beta-hairpin peptide hydrogels as scaffolds for 3D high-throughput cancer drug discovery. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 2171.
Abstract 2176: Targeting eEF-2Kinase by thymoquinone in triple negative breast cancer
Authors: Kabil Nashwa N. , Bayraktar Recep , Kahraman Nermin , Ozpolat Bulent
Published: July 2016

Triple-negative breast cancers (TNBCs) constitute a heterogeneous subtype of breast cancers that present a significant challenge for the treatment and management, and have a poor clinical outcome. Eukaryotic elongation factor-2 kinase (eEF-2K) is an atypical calcium/calmodulin (Ca2++/CaM)-dependent Ser/Thr-kinase that regulates peptide chain elongation by phosphorylating its substrate elongation factor 2 (eEF2), resulting in decreased protein translation. Recently, we have reported that eEF-2K expression is significantly upregulated in TNBC cell lines and is associated with poor survival and worse prognosis, indicating that eEF-2K is a potential therapeutic target in TNBC. Moreover, we demonstrated that eEF-2K promotes TNBC cell proliferation, invasion/migration, and tumorigenesis. Its inhibition by systemically administered liposomal siRNA resulted in marked inhibition of tumor growth and enhancement in the efficacy of chemotherapy in orthotopic TNBC mouse models (Tekedereli 2012). Currently, there are no reported inhibitors of eEF-2K. Natural dietary poly phenolic compounds have been shown to inhibit various cancers by regulating various kinases and molecular targets, with both in-vitro and in-vivo activity. Thymoquinone (TQ) is an active ingredient isolated from Nigella sativa and has been investigated for its antioxidant, anti-inflammatory, and anticancer activities in several in-vitro and in-vivo models. However, the mechanism by which TQ mediates its effects not well understood. Here, we investigated if TQ inhibits eEF-2K in TNBC cells. We demonstrated, for the first time, that treatment with TQ decreases the expression of eEF-2K in a dose dependent manner as wells as its downstream targets, resulting in decreased proliferation, colony formation, migration, and invasion of TNBC cells. Furthermore, eEF-2K knockdown by siRNA recapitulates the effects of TQ, including cell proliferation, migration/invasion, and correlated events. Additionally, TQ treatment inhibits Src activity and other molecular targets, and induces significant apoptosis in TNBC cells in a caspase independent manner. Currently, we are investigating the downstream effects of TQ by RPPA analysis and its in-vivo efficacy in orthotopic xenograft mouse models of TNBC. Overall, our studies suggest that TQ could present a novel therapeutic strategy in targeting eEF-2K and inhibition of TNBCs.

Citation Format: Nashwa N. Kabil, Recep Bayraktar, Nermin Kahraman, Bulent Ozpolat. Targeting eEF-2Kinase by thymoquinone in triple negative breast cancer. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 2176.
Abstract 2179: Macrolide toxin Mycalolide B is a potent inhibitor of HER2 cancer cell invasion and is the basis of actin targeted therapy for metastatic cancers
Authors: Williams Rodette N. , Craig Andrew W. , Allingham John S.
Published: July 2016

Breast and ovarian cancers are among the leading causes of cancer related deaths in women worldwide. Human epidermal growth factor receptor 2 (HER2) is highly expressed in a subset of these cancers that show high rates of progression to metastatic disease. Metastasis is driven by filamentous actin (F-actin) based protrusions that penetrate and degrade extracellular matrix (ECM) to facilitate tumor invasion. In this study, we tested an actin-depolymerizing macrolide toxin Mycalolide B, as a potential suppressor of highly metastatic HER2+ breast and ovarian cancer cell models. Dose responses with Mycalolide B in SKBR3 breast cancer and SKOV3 ovarian cancer cells resulted in similar cytotoxicity (∼65 nM IC50) profiles. In addition, Mycalolide B doses well below the IC50 showed potent suppression of leading edge protrusions, and motility in SKBR3 and SKOV3 cancer cells. This also correlated with reduced ECM degradation and Transwell invasion at low nanomolar doses of Mycalolide B. In contrast, other F-actin based processes such as endocytosis of HER2 and EGFR, were less sensitive to Mycalolide B treatment. However, Mycalolide B treatment did skew the size of endocytic vesicles, which may reflect defects in F-actin based vesicle motility or maturation. Given that HER2 cancers have been effectively targeted by Trastuzumab and Trastuzumab-based antibody-drug conjugates, we are currently testing the compatibility of combined treatments with Mycalolide B and Trastuzumab, for their effects on F-actin based invasion of HER2+ cancers. This strategy may yield a novel class of antibody-drug conjugate, and lead to new HER2 targeted therapies that suppress tumor metastasis by disrupting early steps that are dependent on efficient F-actin polymerization.

Citation Format: Rodette N. Williams, Andrew W. Craig, John S. Allingham. Macrolide toxin Mycalolide B is a potent inhibitor of HER2 cancer cell invasion and is the basis of actin targeted therapy for metastatic cancers. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 2179.
Abstract 218: Ewing sarcoma gene, EWS, role in mitochondrial homeostasis
Authors: Park Jun Hong
Published: July 2016

Peroxisome proliferator-activated receptor γ coactivator (PGC-1α) is a central regulator of mitochondria biogenesis, function and cellular energy metabolism. PGC-1α protein is intrinsically unstable but the mechanism by which PGC-1α stability is regulated is not well understood. Here, we report that a multifunctional protein EWS (Ewing sarcoma) is critical for PGC-1α protein stability and mitochondrial homeostasis. EWS inactivation results in a rapid degradation of PGC-1α via ubiquitin-dependent proteasome pathway and reduces mitochondrial abundance. Re-introduction of EWS in Ews-deficient cells restores PGC-1α expression and mitochondrial anomalies. In the absence of Ews, expression of E3 ubiquitin ligase, FBXW7 (F-box/WD40 domain protein 7), is increased and depletion of Fbxw7 in Ews-null cells restores PGC-1α expression and mitochondrial density. Consistent with these findings, mitochondrial abundance and activity are significantly reduced in brown fat and skeletal muscles of Ews-deficient mice and expression of slow myosin heavy chain is significantly decreased in skeletal muscles of Ews-/- mice compared to controls. Furthermore, Ews-null mice display reduced expression of mitochondrial biogenesis, respiration and fatty acid β-oxidation genes in the liver and abnormally high serum lactate levels. These results demonstrate a novel role of EWS in mitochondrial and cellular energy homeostasis by controlling PGC-1α protein stability.

Citation Format: Jun Hong Park. Ewing sarcoma gene, EWS, role in mitochondrial homeostasis. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 218.
Abstract 2183: Baicalein and meformin decrease small cell lung cancer growth by inhibiting the mTOR pathway in itro
Authors: Marcinkowski Emily F. , Raz Dan , Shen Bing , Xing Quanhua , Yan Jin , Wen Wei , Han Ernest , Yim John
Published: July 2016

Background: Metformin use has been associated with decreased lung cancer risk in several observational studies and is currently in clinical trial in conjunction with standard chemotherapeutics. One possible antitumor mechanism is the negative regulation of the mTOR pathway. We previously reported that the natural product Baicalein has antitumor effects through targeting mTOR pathway. We hypothesized that Baicalein to have antitumor effects in small cell lung cancer. Methods: H1417 small cell lung cancer cells were cultured and treated with increasing doses of Baicalein or Metformin. The cells were harvested at 24 hours and subjected to Western blotting staining for the downstream products of the mTOR pathway. Cell proliferation was determined by MTT assay at 24, 48, and 72 hours. MTT conversion to formazan dye correlates with the number of living cells. Results: We found a dose dependent decrease in the downstream mTOR1 targets pS6K1 and pS6 using both Baicalein and Metformin. There was also an increase in the expression of the mTOR inhibitors DDIT4 and IRF-1. Using the MTT assay, we were able to demonstrate a marked dose dependent decrease in cell proliferation that was sustained over 72 hours from treatment. Interestingly, Baicalein has a markedly higher potency working at micromolar doses verses Metformin which required millimolar doses. Conclusions: Both Baicalein and Metformin effectively decrease cell proliferation in small cell lung cancer cells in vitro. We have shown these drugs to target the mTOR pathway. Cell proliferation is inhibited at a markedly smaller dose by Baicalein compared with Metformin. Baicalein may be useful in cancer chemotherapy and chemoprevention.

Citation Format: Emily F. Marcinkowski, Dan Raz, Bing Shen, Quanhua Xing, Jin Yan, Wei Wen, Ernest Han, John Yim. Baicalein and meformin decrease small cell lung cancer growth by inhibiting the mTOR pathway in itro. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 2183.
Abstract 2184: The effects of specific heterocyclic compounds on angiogenesis and apoptosis factors in cancer cells
Authors: Kosova Funda , Temiz-Arpacı Özlem , TUĞLU İbrahim , ÖLMEZ Ercüment , ÖZDAL KURT Feyzan , KISMALI Gorkem , Ari Zeki , ARISOY Mustafa
Published: July 2016

Introduction: Benzoxazole have received considerable attention during last few decades as they are endowed with variety of biological activities and have wide range of therapeutic properties. The present study aimed to evaluate cytotoxic and antiapoptotic activities of new synthesize benzoxazole derivative in breast cancer cell lines.

Material and Method: In this study, the breast cancer cell lines MDA-MB and MCF-7 were used. Cytotoxic activities of novel benzoxazole-derived compound were analyzed with MTT assay. Additionally, the level of its effects on NF-κB and apoptosis-related proteins were examined by the western blot. Immunohistochemical stainings were done for VEGF, eNOS proteins and Tunnel assay was performed to show DNA damage.

Results: The structure of the compound synthesized in our study 5-amino-2-(p-bromophenyl)-benzoxazole was proved by elemental analysis, IR, 1H NMR and mass

spectroscopy analysis methods. Novel benzoxazole analogue detected cytotoxic on breast cancer cells dose dependent manner, MCF-7 cells were found more sensitive by MTT assay. When the protein was examined in immunohistochemistry with regard to VEGF, eNOS and TUNEL, it was observed that it caused a reduction in VEGF and an increase in eNOS and TUNEL. We observed that it wasn't between groups in Apaf-1 and BCL-2 levels, but down regulation was observed in caspase 3 and Nfkβ levels compared to control group. Novel benzoxazole compound increased Cytochrome C level in treated cells compared to the control group in MDA-MB cells but not in the MCF-7 cells.

Conclusion: In summary, It is felt that this newly synthesized heterocyclic compound increases apoptozis by reducing the activation of Nfkβ, and in this way has shown an effect of inhibiting tumor growth in cancer treatment.

Key Words: Heterocyclic Compounds, NF-κB, APAF-1, Cytochrome C, Caspase-3, bcl-2

Citation Format: Funda Kosova, Özlem Temiz-Arpacı, İbrahim TUĞLU, Ercüment ÖLMEZ, Feyzan ÖZDAL KURT, Gorkem KISMALI, Zeki Ari, Mustafa ARISOY. The effects of specific heterocyclic compounds on angiogenesis and apoptosis factors in cancer cells. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 2184.
Abstract 2185: Sann-Joong-Kuey-Jian-Tang can inhibit human breast cancer cells through dural- blocking both the Ras/Raf/MEK/ERK and Ras/PI3K/Akt/mTOR pathways
Authors: SU. CHIN CHENG
Published: July 2016

Sann-Joong-Kuey-Jian-Tang (SJKJT) consists of 17 species of medicinal herbs, is a traditional Chinese medicine prescription, and has been used as complementary medication for a number of types of solid cancer in Taiwan. Transmembrane tyrosine kinase has been strongly implicated in the growth, survival, and metastasis of a wide variety of human tumors. The phosphoinositide-3-kinase (PI3K)/AKT/ mammalian target of rapamycin (mTOR) and RAS/RAF/MEK/ERK pathways are two of the most frequently dysregulated kinase cascades in human cancer were well documented. Both pathways represent important signal transduction mechanisms that facilitate the proliferation and survival of cancers driven by growth factor receptors, such as vascular endothelial growth factor receptor (VEGFR), insulin-like growth factor-I receptor (IGF-IR), human epidermal growth factor receptor 2 (HER2) or epidermal growth factor receptor (EGFR).The individual downstream components of these signaling cascades either through somatic mutation or epigenetic modification, are also known to be frequently altered in cancer, thus contributing to tumorigenesis and resistance to anticancer therapies. Targeting both the Ras/Raf/MEK/ERK and Ras/PI3K/Akt/mTOR Pathways for suppressing Inhibitor Resistant Cells is necessary. In the present study, the human breast cancer BT-20 and MCF-7 cells were treated with SJKJT in vitro. The cytotoxicity of SJKJT were evaluated by 3–(4,5–dimethylthiazol–2–y1)–2,5–diphenyltetrazolium bromide (MTT) assay. The effects of SJKJT on the protein expressions of VEGFR, IGF-IR, PI3K, AKT, mTOR, Ras, Raf, MEK and ERK and β-actin in the BT-20 and MCF-7 cells were examined by western blot analysis. The results showed that SJKJT can induce the proliferation inhibition with time and dose dependent. As a potential mechanism, it was noted that SJKJT treatment significantly inhibited the activity of the Ras/Raf/MEK/ERK and Ras/PI3K/Akt/mTOR pathways, which played a protective role against the cytotoxicity. In addition, it was demonstrated that SJKJT treatment inhibited the protein expressions of VEGFR, EGFR and IGF-IR significantly. These results suggest that one of the molecular mechanisms for SJKJT to inhibit breast cancer BT-20 and MCF-7 cells maybe through inhibiting the protein expressions of VEGFR, EGFR, IGF-IR and both Ras/Raf/MEK/ERK and Ras/PI3K/Akt/mTOR pathways. The use of traditional chinese medicine prescription SJKJT may become a feasible novel therapy option. Further studies are warranted to fully elucidate its mechanisms of action.

Citation Format: CHIN CHENG SU. Sann-Joong-Kuey-Jian-Tang can inhibit human breast cancer cells through dural- blocking both the Ras/Raf/MEK/ERK and Ras/PI3K/Akt/mTOR pathways. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 2185.
Abstract 2187: Garcinone inhibits nasopharyngeal carcinoma cell growth and arrests cell cycle via modulating the expression of ATR/Stat3/4E-BP1
Authors: Xia Yudui , Liu Xia , Zou Chunllin , Guo Hongwei , Yang Yeguo , Lei Yong , Zhang Jian , Lu Yi
Published: July 2016

Nasopharyngeal carcinoma (NPC) is one of the common head and neck malignancies. Radiotherapy and chemotherapy are the standard treatments for NPC. Garcinone, a natural compound isolated from Garcinia oblongifolia Champ., is a xanthone derivative and has been shown to have potential cytotoxic effects on certain cancer types. However, there have been limited studies performed on its effects on NPC cells and the mechanisms of these effects remain unknown. In this study, we firstly found that garcinone significantly inhibited cell viability of human NPC cell lines CNE1, CNE2, HK1, and HONE1. This inhibition was in a time- and dose-dependent manner. Then, flow cytometry analysis was performed on cell cycle analysis. We observed that garcinone arrested cell cycle at S phase. With 10μM of high-dose garcinone treatment, the cells exhibited necrotic morphology changes including cell swelling, rough endoplasmic reticulum (ER) degranulation, ER dilatation, mitochondria swelling, and vacuolar degeneration. Finally, we found that garcinone stimulated the expression levels of ATR and 4E-BP1, while efficiently inhibited the expression levels of Cyclin B1, Cyclin D1, Cyclin E2, cdc2, CDK7, and Stat3. Collectively, the ability of garcinone to inhibit NPC in vitro suggested that garcinone might be a novel agent for the management of NPC. Supported by NSFC Key Project 81130046; NSFC Project 81171993 and NSFC81272415; Guangxi Key Projects 2013GXNSFEA053004 and 2012GXNSFCB053004; Guangxi Ministry of Education 201202ZD022 and 201201ZD004.

Citation Format: Yudui Xia, Xia Liu, Chunllin Zou, Hongwei Guo, Yeguo Yang, Yong Lei, Jian Zhang, Yi Lu. Garcinone inhibits nasopharyngeal carcinoma cell growth and arrests cell cycle via modulating the expression of ATR/Stat3/4E-BP1. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 2187.
Abstract 219: PHF20 induced necrotic like cell death mediated by ROS via enhanced mitochondrial biogenesis
Authors: Park Jisoo , Li Yuwen , Kong Gyeyeong , Mun Kisun , Lee Hyunji , Kim Dohoon , Tran Quangdon , Wang Ming , Peng Bingjie , Hong Youngeun , Hur Gang Min , Park Jongsun
Published: July 2016

The importance of PHD finger protein 20 (PHF20) in the cellular process is recently emerging as a regulator for the p53 and NF-kB signaling as well as the reprograming process of induced pluripotent stem cell. In this study, we have provided the clear evidence that PHF20 exhibit the novel function in tumorigenesis by regulating the mitochondria biogenesis. Overexpression of PHF20 increases mitochondria biogenesis via up-regulation of peroxisome proliferator-activated receptor-γ coactivator 1-α (PGC1-α) in HCT116 cells. PHF20 overexpression enhances mitochondrial respiratory capacity and oxygen consumption, leading to the accumulation of reactive oxygen species (ROS). Accumulated ROS by PHF20 causes the necrotic like cell death in HCT116 cells. The measurement of mitochondrial ROS in the cells expressing PHF20 revealed that excessive production of mitochondrial ROS facilitates the formation of mitochondrial permeability transition pore, resulting in the loss of mitochondrial membrane potential. PHF20-mediated ROS accumulation damages DNA and leads to over-activation of PARP but not caspase. Antioxidants and PARP-1 inhibitor are able to rescue PHF20 induced cell death, whereas z-VAD, pan-caspase inhibitor, had no effects. Immunohistochemical analysis of various human cancer revealed that PHF20 expression is highly elevated in most of tumor compared to normal tissue. Taken together, PHF20 appears to be tumor-specific antigen and PHF20-mediated PGC1-α upregulation is responsible for the elevation of intracellular ROS generation, resulting the necrotic like cell death. Thus, this study provided the new concept for treating glioma patients and novel pharmacological target in various cancers including glioblastoma and hepatocellular carcinoma

Citation Format: Jisoo Park, Yuwen Li, Gyeyeong Kong, Kisun Mun, Hyunji Lee, Dohoon Kim, Quangdon Tran, Ming Wang, Bingjie Peng, Youngeun Hong, Gang Min Hur, Jongsun Park. PHF20 induced necrotic like cell death mediated by ROS via enhanced mitochondrial biogenesis. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 219.
Abstract 220: SR4 uncouples mitochondrial oxidative-phosphorylation, modulates AMPK-mTOR signaling, and inhibits proliferation of HepG2 hepatocarcinoma cells
Authors: Singhal Sharad S. , Figarola James , Singhal Jyotsana , Tompkins Joshua , Horne David , Awasthi Sanjay , Riggs Arthur
Published: July 2016

Mitochondrial oxidative phosphorylation produces most of the energy in aerobic cells by coupling respiration with the production of ATP. Mitochondrial uncouplers, which reduce the proton gradient across the mitochondrial inner membrane, create a futile cycle of nutrient oxidation without generating ATP. Regulation of mitochondrial dysfunction and associated cellular bioenergetics has been recently identified as promising targets for anticancer therapy. Here, we show that SR4 is a novel mitochondrial uncoupler that causes dose-dependent increase in mitochondrial respiration and dissipation of mitochondrial membrane potential (MMP) in HepG2 hepatocarcinoma cells. These effects were reversed by the recoupling agent 6-ketocholestanol, but not cyclosporin A, and were non-existent in mitochondria-DNA depleted HepG2 (po) cells. In isolated mouse liver mitochondria, SR4 similarly increased oxygen consumption independent of adenine nucleotide translocase (ANT) and uncoupling proteins, decreases MMP, and promotes swelling of valinomycin-treated mitochondria in potassium acetate media. Mitochondrial uncoupling in HepG2 cells by SR4 results in the reduction of cellular ATP production, increased ROS production, activation of the energy-sensing enzyme AMPK, and inhibition of acetyl-CoA carboxylase and mammalian target of rapamycin (mTOR) signaling pathways, leading to cell cycle arrest and apoptosis. Global analysis of SR4-associated differential gene expression confirms these observations, including significant induction of apoptotic genes and down regulation of cell cycle, mitochondrial and oxidative-phosphorylation pathway transcripts at 24 h post treatment. Collectively, our studies demonstrate that SR4's previously reported indirect activation of AMPK and in-vitro anticancer properties, as well as its beneficial effects in both animal xenograft and obese mice models could be a direct consequence of its mitochondrial uncoupling activity (Supported by NIH grant CA 77495 and Beckman Research Institute of the City of Hope).

Citation Format: Sharad S. Singhal, James Figarola, Jyotsana Singhal, Joshua Tompkins, David Horne, Sanjay Awasthi, Arthur Riggs. SR4 uncouples mitochondrial oxidative-phosphorylation, modulates AMPK-mTOR signaling, and inhibits proliferation of HepG2 hepatocarcinoma cells. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 220.
Abstract 2201: Tea nanoparticle, a safe and biocompatible nanocarrier, greatly potentiates the anticancer activity of doxorubicin
Authors: Wang Yi-Jun , Huang Yujian , Anreddy Nagaraju , Zhang Guan-Nan , Zhang Yun-Kai , Xie Meina , Lin Derrick , Yang Dong-Hua , Zhang Mingjun , Chen Zhe-Sheng
Published: July 2016

An infusion-dialysis based procedure has been developed as an approach to isolate organic nanoparticles from green tea. Tea nanoparticle (TNP) can effectively load doxorubicin (DOX) via electrostatic and hydrophobic interactions. We established an ABCB1 overexpressing tumor xenograft mouse model to investigate whether TNP can effectively deliver DOX into tumors and bypass the efflux function of the ABCB1 transporter, thereby increasing the intratumoral accumulation of DOX and potentiating the anticancer activity of DOX. MTT assays suggested that DOX-TNP showed higher cytotoxicity toward CCD-18Co, SW620 and SW620/Ad300 cells, as compared to DOX. Animal study revealed that DOX-TNP resulted in a greater inhibitory effect on the growth of SW620 and SW620/Ad300 tumors than DOX alone. The cTnI levels in mice showed that TNP had no cardiotoxicity and DOX-TNP had moderate cardiotoxicity. Blood Smear tests demonstrated that TNP and DOX-TNP did not cause neutropenia or thrombocytopenia in mice. Therefore, TNP is a safe nanocarrier with excellent biocompatibility and minimal toxicity. In pharmacokinetics study, DOX-TNP greatly increased the SW620 and SW620/Ad300 intratumoral concentrations of DOX, as compared to DOX alone. But DOX-TNP had no effect on the plasma concentrations of DOX up to 240 min after administration. Furthermore, ex vivo IHC analysis of SW620 and SW620/Ad300 tumor sections revealed evidence of prominent antitumor activity of DOX-TNP. In conclusion, our findings suggested that natural nanomaterials could be useful in combating multidrug resistance (MDR) in cancer cells and potentiating the anticancer activity of chemotherapeutic agents in cancer treatment.

Citation Format: Yi-Jun Wang, Yujian Huang, Nagaraju Anreddy, Guan-Nan Zhang, Yun-Kai Zhang, Meina Xie, Derrick Lin, Dong-Hua Yang, Mingjun Zhang, Zhe-Sheng Chen. Tea nanoparticle, a safe and biocompatible nanocarrier, greatly potentiates the anticancer activity of doxorubicin. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 2201.
Abstract 2203: Targeting monoclonal antibodies to the tumor microenvironment for cancer immunotherapy
Authors: Hall Frederick L. , Chawla Sant P. , Chawla Neal S. , Gordon Erlinda M.
Published: July 2016

Therapeutic antibodies and immune checkpoint inhibitors can be delivered more efficiently to the tumor microenvironment (TME) by targeting the exposed collagenous (XC) proteins at the site/s of tumor invasion, stroma formation, and neoangiogenesis.Purpose: To assess the ability of bifunctional fusion polypepetides (mAb-Tropins) to selectively deliver monoclonal antibodies (mAbs) to the TME in tumor-bearing nude mice.Materials and Methods: Synthetic peptide probes and polypeptide aptamers (25-45 aa), fluorescein-labeled IgGs, and an anti-VEGF human mAb were tested in vitro in exposed collagen (XC)-agarose binding assays, in HUVEC cultures, and in vivo in a human xenograft model of pancreatic cancer in nude mice.Results: The XC-targeted mAb-Tropins, bound non-covalently to FITC-labeled IgGs [but not to truncated F(Ab’)2 fragments], exhibited selectivity for XC-agarose over control agarose matrices. Importantly, the biological activity of the XC/mAb-Tropin-bound anti-VEGF mAbs was fully preserved, as demonstrated by inhibition of endothelial cell proliferation in HUVEC cultures. In vivo, intense fluorescence was observed in tumors of mice injected with mAb-Tropin targeted IgGs at 15 and 60 minutes after intravenous injection, but not in non-targeted IgG-treated mice (see Figure below).Conclusions: Tumor XC-targeted mAb-Tropins are an effective method of delivering therapeutic mAbs precisely to the tumor compartments, with meaningful implications for cancer immunotherapy.

Figure

Figure Legend: The performance of mAb-Tropin 277 is evaluated in vitro in XC-binding assays and in vivo in a murine xenograft model of metastatic cancer. XC-agarose chromatography using mAb-Tropin 277 and FITC-IgGs, followed by stringent washing and analysis of column eluates and retentates (A,B), demonstrates aptamer-dependent binding of IgGs to XC-proteins (retentates), as well as depletion of the fluorescent IgGs from solution (column pass-through). The mAb-Tropin 277 dependent targeting of fluorescent IgGs (via the systemic circulation) to tumors is demonstrated in a murine metastatic cancer model (C). Taken together with the in vitro binding data—which showed that the bifunctional mAb-targeting onco-aptamers efficiently sequester IgGs (A) and concentrate the bound IgGs on collagen matrices (B)—this in vivo data indicates that mAb-Tropins concentrate and compartmentalize IgGs/mAbs within the TME (C).

Citation Format: Frederick L. Hall, Sant P. Chawla, Neal S. Chawla, Erlinda M. Gordon. Targeting monoclonal antibodies to the tumor microenvironment for cancer immunotherapy. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 2203.
Abstract 2205: Exosomal encapsulation of doxorubicin reduces the cardiac toxicity of mice
Authors: Rizzolio Flavio , Hadla Mohamad , Corona Giuseppe , Caligiuri Isabella , Palazzolo Stefano , Semeraro Sabrina , Gamini Amelia , Canzonieri Vincenzo , Toffoli Giuseppe
Published: July 2016

Exosomes are nanovesicles produced by cells embedded of cellular information and represent a formidable natural cargo for communication. The name “FedExosomes” denotes the general idea that exosomes could deliver cargo that conveniently manipulated could be of help for patient therapy. Up to now, a few papers demonstrated that modified exosomes could deliver drugs specifically to cancer cells and many laboratories are now studying their properties and compositions. Differently from previously published papers that focused on the efficacy of the doxorubicin encapsulated in exosomes (exoDOX), for the first time, we demonstrated that unmodified exosomes loaded with DOX are less toxic than free DOX by altering the biodistribution of the drug. The heart tissue of mice treated with a toxic dosage of DOX exhibited scattered cytoplasmic paranuclear vacuoles and focal intense eosinophilic cytoplasm, devoid of their transversal striations with moderate disarrangement of cardiomyocytes. The heart tissue of exoDOX treated mice at the same dosage appeared normal. Mass spectrometry studies confirmed that although the quantity of exoDOX in the blood was the same compared to DOX, accumulation in the heart was reduced of about forty percent. Maximum tolerated dose of exoDOX treated mice is double than free DOX. Mice treated at double concentration of exoDOX vs DOX have reduced tumor burden compared to free DOX without observable toxicity.

We demonstrated that exoDOX increased the therapeutic index of DOX by reducing toxic side effects and envisage a similar mechanism for others chemotherapeutic drugs. Since the exosomes could be prepared directly from the patients, our preclinical approach could have a possible immediately application in the clinic.

Citation Format: Flavio Rizzolio, Mohamad Hadla, Giuseppe Corona, Isabella Caligiuri, Stefano Palazzolo, Sabrina Semeraro, Amelia Gamini, Vincenzo Canzonieri, Giuseppe Toffoli. Exosomal encapsulation of doxorubicin reduces the cardiac toxicity of mice. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 2205.
Abstract 2206: Development and characterization of an injectable copper clioquinol formulation: repurposing of a topical antifungal for treatment of cancer
Authors: Wehbe Moe , Anantha Malathi , Backstrom Ian , Malhotra Armaan , Edwards Katarina , Bally Marcel
Published: July 2016

Clioquinol is an FDA approved antifungal and antiprotozoal drug that has recently garnered attention for its anti-cancer activities. In vitro studies with CQ activity suggest that its mechanism of activity may be dependent its ability to bind and transport copper. CQ binds copper to form a stable dimeric complex (2 mol CQ:1 mol Cu ((CuCQ2)). Although therapeutically interesting, CuCQ2 is virtually insoluble in aqueous solutions and therefore it has been difficult to develop formulations suitable for administration. To address this, a liposomal CuCQ2 formulation was developed and characterized for maximum tolerated dose, pharmacokinetics and efficacy in xenograft models. Liposomes (∼100nm) composed of 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC) and cholesterol (CHOL) (55:45 molar ratio) were prepared to contain 300 mM CuSO4. CQ is added to the outside of these pre-formed liposomes; at the appropriate incubation temperature CQ crosses the liposomal membrane where it can interact with the encapsulated Cu to form the insoluble CuCQ2 complex which becomes trapped. The maximum tolerated dose of the resultant formulation was determined in CD-1 mice. The pharmacokinetics of the formulation (liposomal lipid, CQ and Cu) were measured following intravenous administration at a well tolerated dose. Efficacy was measured in two cancer models (sc U251 glioblastoma and i.p A2780 ovarian). The maximum tolerated dose of liposomal CuCQ2 was 30 mg/kg when administered i.v (q2d for two weeks) and 15 mg/kg when administered i.p (QD for two weeks). Pharmacokinetics studies with liposomalCuCQ2 administered iv at 30 mg/kg indicated an terminal elimination half-life (t1/2) of 5.4 hr and an AUC of 994 ug•hr/mL. Results demonstrate that the CuCQ2 complex remains intact upon injection. When used to treat animals bearing established sc U251 glioblastoma tumors showed 30% smaller tumours than those treated with vehicle alone. The activity of liposomal CuCQ2 in the A2780 intraperitoneal cancer model will be presented. This work describes, for the first time, a novel injectable formulation of CuCQ2; this formulation is active against established xenograft models. The activity of this formulation will now be assessed in the context of copper-dependent inhibition of NFκB signalling.

Citation Format: Moe Wehbe, Malathi Anantha, Ian Backstrom, Armaan Malhotra, Katarina Edwards, Marcel Bally. Development and characterization of an injectable copper clioquinol formulation: repurposing of a topical antifungal for treatment of cancer. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 2206.
Abstract 2210: Multiple antigens stimulating cellular therapy increases immune responses in patients with hepatocellular carcinoma
Authors: Han Yanyan , Huang Jing , Tao Ran , Guo Yabing , Liu Li , Wu Dongyun , Li Jin , Zhou Xiangjun , Hou Jinlin
Published: July 2016

Background & Aims: Hepatocellular carcinoma (HCC) is one of the most common tumors in China, and frequently occurs in the patients with chronic hepatitis B virus (HBV) infection. To investigate whether adoptive cell therapy induces tumor specific immune responses in HCC patients and further brings clinical benefits, Multiple Antigens Stimulating Cellular Therapy (MASCT) was applied in addition to conventional therapies.

Methods: Activated T cells that were prepared from the autologous peripheral blood mononuclear cells (PBMCs). Cells were stimulated and amplified with dendritic cells (DCs) pulsed by multiple tumor antigen peptides, including tumor associated antigen peptides and HBV antigen peptides. All patients received infusions of 5-10×107cells/kg of body weight of the resulting T cells every 1-3 months. The immune responses in the patients' PBMCs were examined before and after the treatment.

Results: The resulting amplified cells were almost exclusively CD3+ (95% ± 1%), with a major percentage being CD3+CD8+ cells (70% ± 5%) and a minor proportion being CD3+CD4+(19% ± 3%) and CD3+CD56+ (14% ± 1%) cells, while there were insignificant amounts of CD4+CD25+Foxp3+ regulatory T cells (0.4% ± 0.1%). These T cells have restored immune functions and HLA-restricted specific cytotoxicity against HCC cells. After repeating treatment of MASCT, the frequency of regulatory T cells in the patients’ PBMCs was significantly decreased, and the proliferation and function of tumor antigen specific T cells were enhanced. The specific immune responses against each kind of tumor antigens were also detected in patients. And these immune responses were gradually increased during the treatment of MASCT. Moreover, in retrospective study, the disease control rate (DCR) in one year was found to be raised to 80% in stage B HCC patients receiving MASCT combined with conventional therapies.

Conclusions: Our study has demonstrated that MASCT is a well tolerate immunotherapy in patients with HCC. Specific T cell responses against tumor antigens can be strongly induced in vivo. Our study also indicated that repeated MASCT treatment improves both the immunologic function and disease control of patients with HCC.

Citation Format: Yanyan Han, Jing Huang, Ran Tao, Yabing Guo, Li Liu, Dongyun Wu, Jin Li, Xiangjun Zhou, Jinlin Hou. Multiple antigens stimulating cellular therapy increases immune responses in patients with hepatocellular carcinoma. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 2210.
Abstract 2223: Membranous expression of programmed cell death-ligand 1 (PDL1) on cancer cells is induced by cisplatin in an ATR dependent manner
Authors: Schaaf Lea , Dong Meng , Heine Simon , van der Kuip Heiko , Aulitzky Walter E.
Published: July 2016

Cancer cells frequently develop strategies allowing to escape potential immune attacks by the host. One central component of this defense mechanism is the PDL1-PD1 axis with PDL1 expression on the surface of antigen presenting cells. DNA damaging chemotherapeutic are known to have suppressive effect on the immune system. Possible effects on immune checkpoints, however, are poorly understood. We investigated if Cisplatin, a widely used chemotherapeutic for treatment of ovarian and lung carcinomas, might influence expression of PDL1 on cancer cells of these pathologies. Treatment of cell lines as well as precision-cut tissue slices from patient tumors with clinically relevant dosages of cisplatin led to a significant induction of PDL1 protein in cancer cells as revealed by Western Blot and IHC analyses. This effect was independent on ATR or ATM activity since pre-treatment with ATR or ATM inhibitors had no significant effect on cisplatin induced total PDL1 protein levels. Interestingly, however, we observed an almost complete inhibition of cisplatin-induced membranous PDL1 upon ATR but not ATM inhibition as revealed by FACS analysis in unfixed cells. The role of the ATR axis was further corroborated by the finding that presentation of PDL1 at the cell surface was paralleled by phosphorylation of CHK1.

Together our data indicate an important role of DNA damage induced ATR/CHK1 activity on the localization of PDL1 at the surface of cancer cells. These data implicate that combination of cisplatin with either anti-PD1 or anti-PDL1 antibodies or ATR/CHK1 inhibitors should reduce immunosuppressive mechanisms and enhance chemotherapy efficacy.

Citation Format: Lea Schaaf, Meng Dong, Simon Heine, Heiko van der Kuip, Walter E. Aulitzky. Membranous expression of programmed cell death-ligand 1 (PDL1) on cancer cells is induced by cisplatin in an ATR dependent manner. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 2223.
Abstract 2224: Discovery, development and characterization of a new class of therapeutic anti-PD-1 antibody
Authors: Wang Lei , Amirina Najmia , Scheuplein Fellix , Spaulding Vikki , Wang Yanyan , Vadde Sri , Chiu M. Isabel , Doty Daniel , McQuade Thomas , Feng Bin , Ranganath Sheila , Nguyen. Anhco “Cokey”
Published: July 2016

Blockade of immune checkpoint (IC), such as PD-1, confers durable clinical benefit to a minority of melanoma and lung cancer patients. Challengingly, the 5-year survival rate is still below 30% in those patients, and most other solid tumors are not particularly responsive to immunotherapy. Therefore, more IC blocking mAbs are under development to meet clinical needs. Using Enumeral's unique single cell immunoprofiling platform, we developed a novel class of anti-PD-1 antibodies, named 244C8, with distinct epitope binding site than currently marketed anti-PD-1 antibodies. To evaluate the function of 244C8 on anti-tumor immunity, we employed ex vivo methods to determine responsiveness of tumor infiltrating lymphocytes (TILs), isolated from human lung cancer biopsies, to various anti-PD-1 antibodies, including our lead anti-PD1 therapeutic candidate 244C8. The results show that all anti-PD-1 antibodies tested reversed T cell exhaustion in tumor microenvironment. Further, 244C8-treated cells produced increased IFN-y compared to nivolumab, indicating a stronger anti-tumor immunity. In the settings of mixed lymphocyte reaction (MLR) and CMV antigen-recall assays, 244C8 treatment also led to higher levels of T cell activation compared to conventional anti-PD-1 antibodies. Interestingly, in both ex vivo and in vitro models, we observed a pattern of cytokine production in 244C8-treated cells that is consistently distinct from that produced by conventional/marketed anti-PD-1 antibodies, suggesting a differentiated mechanism of action.

Citation Format: Lei Wang, Najmia Amirina, Fellix Scheuplein, Vikki Spaulding, Yanyan Wang, Sri Vadde, M. Isabel Chiu, Daniel Doty, Thomas McQuade, Bin Feng, Sheila Ranganath, Anhco “Cokey” Nguyen. Discovery, development and characterization of a new class of therapeutic anti-PD-1 antibody. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 2224.