bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 2328: BiTE-T cell: A novel design for solid tumor
Authors: Luo Feifei , Wang Yuedi , Liu Jie , Chu Yiwei
Published: August 2019

Genetically engineered T cells therapy is a promising strategy in cancer immunotherapy. Chimeric antigen receptor (CAR)-T cells therapy, as one successful strategy, has achieved the breakthrough and been approved in non-solid tumor but not solid tumor, which might be limited partially by CARs-restricted on cell surface and side effects of retrovirus. Meanwhile, bispecific T cell engager (BiTE), as another successful strategy, has been approved in non-solid tumor, the therapeutic limitation of which in solid tumor might be the short half-life of antibodies, the lack of endogenous effector T cells in patients with advanced cancer, and severe adverse effects. In this study, we developed a novel glypican-3 (GPC3)-targeting BiTE, αGPC3/CD3. These GPC3-BiTE engineered human T cells persistently produced GPC3-BiTE fusion proteins, which were released extracellularly to help both engineered T cell and bystander T cells to exhibit GPC3-specific activity. As a result, GPC3-BiTE-T cells could possess the capability of both CAR-T cells and BiTE, and even aid more than conventional CART cells in the CART-like function. In addition, GPC3+ tumor-bearing mice treated with the secretable GPC3-BiTE-T cells got a significant tumor growth inhibition and prolonged survival without observed adverse effect. Compared with control groups, more GPC3-BiTE-T cells with less PD-1 expression were detected at tumor site, which was similar to the GPC3-CART cell therapy. And the accumulation of GPC3-BiTE-T cells was positively related with the inhibition of tumor growth. Moreover, GPC3-BiTE-T cells assisted bystander T cells accumulated into tumor site in vivo, which was lack in GPC3-CART cell therapy. Thus, the novel GPC3-BiTE-T cells have the characteristics of high potency, long term and low toxicity, which might supply an attractive GPC3-targeting immunotherapy for solid tumors.

Citation Format: Feifei Luo, Yuedi Wang, Jie Liu, Yiwei Chu. BiTE-T cell: A novel design for solid tumor [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 2328.
Abstract 2358: Porphyromonas gingivalis promotes colorectal cancer development by regulating NLRP3 inflammasome signaling
Authors: Wang Zhi , Wang Xi , Jia Yiqun
Published: August 2019

Metagenomic analyses indicate that Porphyromonas spp. is associated with human colorectal carcinoma (CRC), however, whether this is an indirect or causal link remains unclear. We found that Porphyromonas gingivalis (P. gingivalis) were enriched in human stool or FFPE samples from colorectal carcinoma patients compared to colorectal adenoma or healthy subjects. Two colorectal cohort studies demonstrated that P. gingivalis infection was associated with poor prognosis of CRC. Moreover, in the ApcMin/+ mouse model of colorectal tumorigenesis, P. gingivalis increases tumor counts and in the orthotopic rectal MC38 carcinoma model increases tumor growth. Furthermore, tumors from orthotopic MC38 model exposed to P. gingivalis exhibited a tumor-infiltrating myeloid cells recruiting signature that was confirmed in human P.gingivalis-positive colorectal carcinomas. We next evaluated the role of NLRP3 in P. gingivalis promoting on CRC progression. When comparing with wild type mice, the increased tumor growth was subtracted in Nlrp3-/- mice by the same orthotopic MC38 model exposed to P. gingivalis. To determine relevant cell compartment responsible for NLRP3 function, we generated Nlrp3-/- chimeric mice with adoptive bone marrow transplantation by using the same orthotopic tumor model. The results showed the effect of NLRP3 on P. gingivalis pathogenesis was from both hematopoietic and non-hematopoietic sources. Additionally, we performed expression profile to show that NLRP3 attenuates multiple pathways associated with P. gingivalis promoting colorectal cancer development. Collectively, these data suggest that, through recruitment of tumor-infiltrating immune cells by regulating NLRP3 inflammasome activity, P. gingivalis generates a proinflammatory microenvironment that is conductive for colorectal neoplasia progression.

Note: This abstract was not presented at the meeting.

Citation Format: Zhi Wang, Xi Wang, Yiqun Jia. Porphyromonas gingivalis promotes colorectal cancer development by regulating NLRP3 inflammasome signaling [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 2358.
Abstract 2367: Enhancing the efficacy of liver cancer immunotherapy by specific inhibition of histone deacetylase 8
Authors: Yang Wei-Qin , Zhou Jingying , Feng Yu , Cheung Ka Wing , Cheng Alfred Sze-Lok
Published: August 2019

The heterogeneous responses to immune-checkpoint blockade (ICB) therapy e.g. anti-programmed death-ligand 1 (PD-L1) antibody are attributable to the complex interplay between a range of cancer-cell-autonomous cues and the immunosuppressive tumor microenvironment. Two recent phase I/II trials of checkpoint inhibitors in patients with advanced hepatocellular carcinoma (HCC) have produced promising results, yet the objective response rates were relatively low (17-20%) as compared to other solid tumors such as melanoma and lung cancer (~40-60%). Accumulating evidence is underscoring the fundamental importance of epigenetic regulation in tumor immune evasion. We have previously elucidated a critical role of histone deacetylase 8 (HDAC8) in hepatic carcinogenesis (Cancer Research 2015;75:4803-16). Here, we aim to investigate the therapeutic potential of a HDAC8-specific inhibitor PCI-34051 in preclinical hepatocellular carcinoma (HCC) models. PCI-34051 significantly reduced HCC tumorigenicity in immunocompetent but not immunodeficient mice. Immune profiling revealed specific reduction in tumor-infiltrating regulatory T cells, which was associated with significant increase in CD8+ T cells. Notably, combined PCI-34051 and anti-PD-L1 treatment resulted in complete tumor eradication in all of the co-treated mice. Moreover, the combination therapy promoted long-term survival, which was associated with elevated CD8+ T effector and central memory cells. Our data suggest that selective chromatin modifications by HDAC8 alter the tumor immune surveillance program and demonstrate the potential of rational combinatorial epigenetic immunotherapy to fully unleash T-cell responses, leading to long-term remission of HCC. Acknowledgement: This work is supported by the RGC CRF (C4017-14G) and the Focused Innovations Scheme-Scheme B from the Chinese University of Hong Kong (1907309).

Citation Format: Wei-Qin Yang, Jingying Zhou, Yu Feng, Ka Wing Cheung, Alfred Sze-Lok Cheng. Enhancing the efficacy of liver cancer immunotherapy by specific inhibition of histone deacetylase 8 [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 2367.
Abstract 2368: Domatinostat increases apoptosis, G2M cell-cycle arrest and immunogenicity of Merkel cell carcinoma
Authors: Song Lina , Bretz Anne Catherine , Gravemeyer Jan , Spassova Ivelina , Hamm Svetlana , Muminova Shakhlo , Bartz Rene , Becker Juergen C.
Published: August 2019

Merkel cell carcinoma (MCC) is a rare, highly aggressive skin cancer prevalent in elderly and immunocompromised patients. MCC is highly immunogenic as it is either associated with Merkel cell polyoma virus (MCPyV) integration or UV-associated mutations in non-viral cases. Indeed, immune checkpoint inhibitors (CPI) like PD-1/PD-L1 blocking antibodies exert a strong clinical activity; however, primary or secondary resistance often occurs. Domatinostat (4SC-202) is an orally available small molecule inhibitor targeting histone deacetylases (HDAC) class I currently in clinical evaluation to improve response to CPI (SENSITIZE, NCT03278665). In syngeneic tumor mouse models domatinostat treatment demonstrated immune-modulatory effects by increasing the intra-tumoral infiltration of cytotoxic CD8+ T cells (CTLs) and enhancing gene expression of a CPI response signature.

Immune escape mechanisms described for MCC include low intra-tumoral levels of CTLs and reduced expression of antigen-presenting MHC class I molecules. Our previous data suggest that low MHC-I levels are reversible and involve epigenetic silencing of genes encoding the antigen-processing machinery (APM). Here, we present novel preclinical data about the efficacy and mode of action of domatinostat in MCC. Global gene expression by single cell RNA-seq revealed regulation of genes involved, among others, in apoptosis and antigen presentation. Importantly, domatinostat additionally inhibited proliferation of MCC cell lines by induction of a G2M cell-cycle arrest and apoptosis. Moreover, expression of MCPyV-encoded transforming early genes, particularly during the G1-phase, was inhibited by domatinostat. Thus, it exerts also a direct anti-tumoral effect. In viable cells, domatinostat increases their susceptibility to immune responses, as qPCR and immunoblot analysis confirmed the induction of APM and MHC-I expression by domatinostat. APM and MHC-I expression has been reported to be restored by an epigenetic drug combination (vorinostat plus mithramycin). In contrast, we now provide evidence that the single agent treatment with domatinostat alone is sufficient for increasing immunogenicity of MCC cells.

In summary, domatinostat counteracts immune escape of MCC in many aspects suggesting a combination treatment of the HDACi domatinostat with CPI as a promising therapeutic strategy. Prospective clinical trials are needed to confirm this hypothesis.

Citation Format: Lina Song, Anne Catherine Bretz, Jan Gravemeyer, Ivelina Spassova, Svetlana Hamm, Shakhlo Muminova, Rene Bartz, Juergen C. Becker. Domatinostat increases apoptosis, G2M cell-cycle arrest and immunogenicity of Merkel cell carcinoma [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 2368.
Abstract 2390: FcgR co-engagement by anti-TIGIT monoclonal antibodies enhances T cell functionality and antitumor immune responses
Authors: Chand Dhan , Waight Jeremy D. , Paltrinieri Elena , Dietrich Sylvia , Bushell Mark , Costa Mathew , Gombos Randi , Wilson Nicholas S. , Buell Jennifer S. , Stein Robert B. , Duncan Alexander , Savitsky David A.
Published: August 2019

T-cell immunoreceptor with Ig and ITIM domains (TIGIT) has emerged as an important regulator of the cancer-immunity cycle. Preclinical studies have demonstrated that TIGIT antibodies can enhance T and NK cell anti-tumor immunity, and therapeutic antibodies targeting TIGIT are advancing in the clinic. Nonetheless, questions on the optimal format for TIGIT therapeutics remain unresolved. Preclinical studies have demonstrated that anti-TIGIT antibodies enhance anti-tumor immunity by (1) blocking inhibitory signaling downstream of TIGIT/PVR and TIGIT/PVRL2 receptor-ligand interactions and (2) redirecting PVR/PVRL2 ligand binding to the co-stimulatory receptor CD226. Here we describe a novel and unanticipated mechanism of action of anti-TIGIT antibodies in which interactions between the antibody Fc domain and select FcγRs, particularly FcγRIIIA, dramatically improve T cell activation and effector function. In mouse tumor models, TIGIT antibodies that promote FcγR interactions enhance anti-tumor responses, whereas Fc-inert TIGIT antibodies show no such benefit. Moreover, TIGIT antibodies that enhance binding to human FcγRIIIA or mouse FcγRIV promote superior single agent activity and combination activity with other checkpoint modulators in antigen-stimulation assays and mouse tumor models respectively. Notably, this mechanism of action is independent of regulatory T cell depletion. Our findings support a dependence on Fc-FcγR interaction for promoting T cell responsiveness and effector function upon TIGIT antagonism. We further demonstrate that this novel mechanism also extends to anti-CTLA-4, but not anti-PD-1 or anti-LAG-3 antibodies. Altogether, our data describe a novel FcγR-dependent mechanism of action that may enhance the therapeutic activity of anti-TIGIT antibodies, deepen our understanding of this class of therapies, and inform on the optimal design for a new class of Fc-engineered antibodies that could be leveraged to potentially enhance antitumor immune responses.

Citation Format: Dhan Chand, Jeremy D. Waight, Elena Paltrinieri, Sylvia Dietrich, Mark Bushell, Mathew Costa, Randi Gombos, Nicholas S. Wilson, Jennifer S. Buell, Robert B. Stein, Alexander Duncan, David A. Savitsky. FcgR co-engagement by anti-TIGIT monoclonal antibodies enhances T cell functionality and antitumor immune responses [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 2390.
Abstract 2397: Pre-clinical evaluation of PE0116: A potent cross-linking depednet 4-1BB agonist antibody
Authors: Song Ningning , Li Zhengwu , He Shan , Yang Teddy , Duan Qing
Published: August 2019

4-1BB is a member of the tumor necrosis factor superfamily and plays a critical role in activation of cytotoxic T cells to promote anti-tumor immunity. As a co-stimulatory molecule, it is ideally suited for activation via agonist therapeutic antibodies in order to reactivate the immune system and promote cytolytic activity through CD8+ T cells. Clinical trials of two agonist antibodies (Urelumab and Utomilumab) are ongoing. Urelumab has shown clinical efficacy but been hampered by inflammatory liver toxicity at doses above 1 mg/kg, suggesting potential safety concerns at higher doses. Utomilumab has a superior safety profile, but is a less potent 4-1BB agonist. Hence, it may be necessary to achieve potent immune activation while avoiding limiting immune-related adverse events. PE0116 is a fully human monoclonal antibody generated from immunization of Harbour H2L2 human transgenic mice with recombinant human 4-1BB protein. It blocks the binding of 4-1BB to its ligand 4-1BBL and its activity in T cell activation is crosslinking dependent. After crosslinking, PE0116 strongly increases IFNg secretion from T cells in the presence of anti-CD3 antibody and its activity is superior to Utomilumab analog. More importantly, PE0116 has demonstrated robust anti-tumor activity and better potency than both benchmark antibodies in MC38 tumor model with human 4-1BB KI mice. PE0116 has also demonstrated good safety profile in a non-GLP safety study in cyno monkey. Currently the antibody is under late stage IND-enabling study. Its safety and efficacy will be further tested in patients.

Citation Format: Ningning Song, Zhengwu Li, Shan He, Teddy Yang, Qing Duan. Pre-clinical evaluation of PE0116: A potent cross-linking depednet 4-1BB agonist antibody [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 2397.
Abstract 2756: A high-resolution G-quadruplex structure involved in cmyc oncogene regulation
Authors: Dickerhoff Jonathan , Chen Luying , Wu Guanhui , Onel Buket , Lin Clement , Yang Danzhou
Published: August 2019

Gene expression needs strict regulation in human cells to prevent tumorigenesis. Among other control mechanisms, guanine-rich sequences which are clustered in promoter regions of oncogenes can form four stranded nucleic acid structures, called G-quadruplexes (G4), and regulate their transcription. The cmyc protein is the most important cellular regulator for proliferation and is often overexpressed in cancer cells. The DNA G-quadruplex formed in the cmyc promoter is a transcriptional silencer element and stabilization of this G4 by small molecules suppresses cmyc transcription, making it a promising target for cancer therapy. High-resolution structures of the highly diverse G4s provide the opportunity to understand their molecular interactions with transcription factors and to design small molecules that specifically bind a particular G4 conformer. Characteristically, multiple G4s can be formed in gene promoter regions because their G-rich sequence generally contains more than the minimum of four G-stretches. While all reported cmyc G4s share the same core with parallel oriented strands, their overall shape is individualized by distinct combinations of varying flanking and loop motifs. In this study, we present a new high-resolution NMR structure of a G4 found to form in the cmyc promoter region. The determined cmyc G4 has a 1:6:1 loop-size arrangement and its conformation is largely defined by its long central loop of 6 nt. We show that this G4 structure is the preferred target of nucleolin protein in comparison to other G4s formed in the cmyc promoter, demonstrating how the polymorphism of a G-rich sequence can modulate the interaction with transcription factors and other regulators. Our NMR structure shows that, despite the length of the 6-nt loop motif and the associated increased flexibility, the folding converges well due to a base pairing between the central loop and the 5’-flanking. Thus, the long loop is pulled towards the 5’-end while analogous loop-flanking interactions are prevented at the opposite 3’-site. In conclusion, the presented new addition to the ensemble of possible cmyc promoter G4 conformations extends the folding landscape for this promoter region, in particular due to the long central loop. This may shed light on how proteins such as nucleolin can target specific G4 and shift the equilibrium of folding to fine-tune gene expression. Finally, this new structure is another promising target for cancer drug development.

Citation Format: Jonathan Dickerhoff, Luying Chen, Guanhui Wu, Buket Onel, Clement Lin, Danzhou Yang. A high-resolution G-quadruplex structure involved in cmyc oncogene regulation [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 2756.
Abstract 2928: New insight into signaling contexts for the administration of the imipridone ONC201 to prostate cancer cells
Authors: Amoroso Francesca , Pickard Adam , Glass Kimberly , Tarapore Rohinton , Allen Josh , Mills Ian G.
Published: August 2019

Prostate cancer is the most common non-cutaneous cancer in men and a notable cause of cancer mortality. In the majority of the cases localised disease radical treatment consists of surgery or radiotherapy. Upon progression androgen deprivation therapy can extend survival however metastatic disease is incurable. The prostate is a specialized accessory gland with a high secretory capacity. As prostate cancer develops and treatment resistance emerges, the unfolded protein response (UPR) is an important adaptive biology which co-amplifies with key cancer drivers1. The UPR can be cytoprotective but when acutely activated can lead to cell death. In this study we sought to enhance the acute activation of the UPR using radiation and ONC201, previously reported to be a UPR activator2. We found that treating prostate cancer cells with ONC201 induced increases in the expression of components in all arms of the UPR - ATF4, ATF6 and IRE1-XBP1 - at protein and transcript levels at a 24-hour time point and this was followed at 72 hours by cell death. This delay in the induction of cell death provided a time window for the short-term administration of ONC201 to prime an enhanced cytotoxic response to radiation. Pre-treatment with ONC201 for 24 hours followed by withdrawal and subsequent irradiation led to enhanced cytotoxicity assessed by cell viability and clonogenic assays. In order to decipher the impact of ONC201 and radiation we undertook RNA-seq under a range of treatment conditions. We noted a durable suppression of transcripts encoding cell cycle regulatory genes when ONC201 pre-treatment was used to enhance radiation response and this translated into cell cycle arrest and induction of both necrotic and apoptotic cell death as assessed by flow cytometry. Pathway analysis of the RNA-seq suggests alternative targets that could replicate this priming effect to enhance radiation response.

1.Storm, M. et al., Oncotarget 7(33): 54051-54066 (2016).

2.Allen, J. et al., Oncotarget (2016). doi:10.18632/oncotarget.11814.

Citation Format: Francesca Amoroso, Adam Pickard, Kimberly Glass, Rohinton Tarapore, Josh Allen, Ian G. Mills. New insight into signaling contexts for the administration of the imipridone ONC201 to prostate cancer cells [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 2928.
Abstract 2933: Application of a mouse model humanized for the major pathways of drug disposition in anticancer drug development and use
Authors: Henderson Colin J. , Scheer Nico , Kapelyukh Yury , McLaren Aileen , MacLeod Kenneth , Rode Anja , Lin De , Wolf C Roland
Published: August 2019

A vast number of targeted anticancer drugs are being developed in the pharmaceutical industry whose efficacy will only become fully realized through their combination with other established or novel anti-tumor agents. This is particularly the case because combination chemotherapy is a major approach being taken to overcome the rapid onset of drug resistance which current dosing regimens can cause. This has raised the conundrum of which of the large number of possible drug combinations, which may involve combinations of more than two drugs, have the greatest chance of success. It is not possible to test all the possible combinations by clinical trial alone so more informative preclinical models are badly needed. Nearly all targeted anti-tumor agents are substrates for the cytochrome P450-dependent monooxygenase system. The testing of novel drug regimens in mice is severely compromised by the major species differences in this enzyme system both in catalytic function, in the pattern of metabolite formation and the regulation by transcription factors such as CAR and PXR. In order to circumvent this problem we have created a mouse model where thirty four murine P450’s have been deleted from the mouse genome and substituted for the major enzymes involved in drug disposition in man ie CYP1A1, CYP1A2, CYP2C9, CYP2D6, CYP3A4 and CYP3A7. The mice have also been humanized for the transcription factors CAR and PXR. CYP3A4 and CYP2D6 are expressed off the human promoters. We report the validation the utility of this model by studying the in vivo metabolism and disposition of model drugs and the metabolism and disposition of the EGFR inhibitor, osimertinib and the BRAF inhibitor, dabrafenib. We show that the use of this model allows accurate prediction of clinically observed drug exposures, in the generation of human metabolites and drug/drug interactions. This model has therefore great potential for the development of combination therapies involving complex drug regimens and in the design of clinical trials targeted at overcoming drug resistance.

Citation Format: Colin J. Henderson, Nico Scheer, Yury Kapelyukh, Aileen McLaren, Kenneth MacLeod, Anja Rode, De Lin, C Roland Wolf. Application of a mouse model humanized for the major pathways of drug disposition in anticancer drug development and use [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 2933.
Abstract 2941: Combination of hemiasterlin derivative (R)(S)(S)-BF65 and PI3K/mTOR dual inhibitor NVP-BEZ235 as a new strategy to inhibit ovarian cancer growth and metastasis
Authors: Su Po-Chia , Huang Tzu-En , Hsu Lih-Ching
Published: August 2019

Metastasis is a big issue to cancer therapy and is the major cause of therapeutic failure and cancer death. Therefore, inhibiting cancer metastasis is a key challenge in cancer treatment. Ovarian cancer is the most deadly cancer among women and combination treatment strategy is usually used to fight against this disease not only to reduce the toxicity and side effects, but also to enhance the anticancer effect.

(R)(S)(S)-BF65 is a hemiasterlin derivative and potent anti-tubulin agent with IC50 values in the low nanomolar range against various cancer cells. Previous studies indicated that (R)(S)(S)-BF65 in combination with an allosteric Akt inhibitor MK-2206 synergistically inhibited the growth of SKOV3 human ovarian cancer cells. Using the MTT assay, we demonstrated that (R)(S)(S)-BF65 could also enhanced the growth inhibitory effect of NVP-BEZ235, a PI3K/mTOR dual inhibitor in SKOV3 cells. The IC50 was 11.6 μM for NVP-BEZ235 and 20.6 nM for (R)(S)(S)-BF65 after 48 h-treatment. With a molar ratio of 1:1000 or 1:500, the combination of (R)(S)(S)-BF65 and NVP-BEZ235 displayed an additive to mild synergistic cytotoxic effect with CI50 values of 0.96 and 0.91, respectively.

Interestingly, the combination of (R)(S)(S)-BF65 and NVP-BEZ235, at concentrations much lower than those required for growth inhibition, significantly inhibited cell migration and invasion as demonstrated by wound healing migration, transwell migration and invasion assays. The combinatorial effect was much more effective than single treatments.

Epithelial-mesenchymal transition (EMT) plays an important role in cancer metastasis. It has been reported that activation of the PI3K/Akt pathway leads to phosphorylation and inactivation of GSK3β, which may in turn induce EMT. Western blot analysis of EMT markers revealed that NVP-BEZ235 downregulated mesenchymal markers β-catenin and Slug, and upregulated the epithelial marker, E-cadherin. (R)(S)(S)-BF65 also downregulated mesenchymal markers β-catenin and Snail, but had no effect on E-cadherin. The combination of both led to downregulation of more mesenchymal markers including β-catenin, Slug, Snail and ZEB1, and upregulation of E-cadherin, which may at least in part explain why the combination was more effective than single treatments to suppress cell migration and invasion.

Taken together, the combination of (R)(S)(S)-BF65 and NVP-BEZ235 may be a promising strategy to inhibit ovarian cancer growth and metastasis.

Citation Format: Po-Chia Su, Tzu-En Huang, Lih-Ching Hsu. Combination of hemiasterlin derivative (R)(S)(S)-BF65 and PI3K/mTOR dual inhibitor NVP-BEZ235 as a new strategy to inhibit ovarian cancer growth and metastasis [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 2941.
Abstract 2955: Development of non-apoptotic, caspase-independent cell death inducers against cancer cells
Authors: Pasternak Mariah A. , Hussein Noor , Chandrabose Karthikeyan , Erhardt Paul W. , Tiwari Amit K.
Published: August 2019

Since most cancer therapeutics currently exert their effects using apoptotic pathways to which cancer cells can become resistant, it is important to develop new drug molecules that work through non-apoptotic pathways. A novel, non-apoptotic cell death mechanism called “methuophagy,” which combines features of both autophagy and methuosis, is being examined using structure-activity relationship (SAR) studies of synthesized compounds. Methuosis is a non-apoptotic cell death that includes macropinocytosis and the fusing of smaller vesicles into larger vesicles that is seen along with cell death. BAPT compounds that induce methuophagy have been synthesized, and minor changes on a primary scaffold are being used to determine the SAR for the series. The series retains the 4-pyridinyl moiety that induced methuosis with the compound MOMIPP. Screening experiments with HCT-116 colorectal cancer cells (CRC), BT-20 triple-negative breast cancer cells (TNBC), and U-251 glioblastoma cancer cells (GBM) have shown that compounds having a rigid, completely aromatic structure with three aromatic centers induce methuophagy more potently than compounds with a “hydrazone-like” moiety or a non-aromatic linker. The cell death caused by methuophagy can be seen as early as 6 hours, with small vacuoles that coalesce together to form larger vacuoles. From these screening experiments, it has been concluded that the completely aromatic compounds are the most promising candidates for methuophagy drug development and will be used in experiments to determine protein targets and in vivo to further determine efficacy.

Citation Format: Mariah A. Pasternak, Noor Hussein, Karthikeyan Chandrabose, Paul W. Erhardt, Amit K. Tiwari. Development of non-apoptotic, caspase-independent cell death inducers against cancer cells [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 2955.
Abstract 2958: Evaluating the therapeutic effects of methylene blue against prostate cancer
Authors: Shanthi Priyadarshini Thiruvalluvan , Foes Abigail , Munirathinam Gnanasekar
Published: August 2019

Prostate cancer (PCa) is the second most commonly occurring cancer among men and the fourth most commonly occurring cancer overall. According to American Cancer Society, in the year 2018, it was estimated that there will be 164,690 new cases and 29,430 deaths from PCa. The treatment options currently available for PCa are found to be ineffective with varied side effects and complications associated with the development of resistance among patients. Therefore, there is an unmet need to find a safe and potent agent to treat PCa. In our study, we focused on studying the anticancer potential of Methylene Blue (MB) which belongs to the class of phenothiazinium salt. MB has been widely used to treat the condition of methemoglobinemia, emerging studies have shown that it has been effectively used as a photosensitizer in the treatment of cancer by means of photodynamic therapy (PDT). Our initial analysis showed that MB effectively reduced the viability of androgen-dependent (LNCaP) and androgen-independent (PC3 and DU145) PCa cells. Further experimental evaluations showed that MB inhibited the colony forming ability of PCa cells in-vitro suggesting its tumor suppressive potential. In addition, our studies showed that MB treatment disrupted the migration potential of PCa cells in a wound healing assay indicating the anti-metastatic function of MB. Moreover, confocal and FACS analysis using Annexin V FITC and propidium iodide staining revealed that MB effectively targeted the PCa cell lines by inducing apoptotic cell death. To delineate the underlying anticancer mechanism of MB, apoptosis protein array was performed employing LNCaP cells, and the results of which showed that key apoptotic molecules such as Bax, TRAIL

R2/D5, and phospho p53 (Serine 15, Serine 46, Serine 392) were robustly upregulated in LNCaP cells following MB treatment. In conclusion, our findings suggest that MB induces apoptosis in PCa cells and thus could serve as a potential anticancer agent for treating both hormone-dependent and -independent PCa.

Citation Format: Priyadarshini Thiruvalluvan Shanthi, Abigail Foes, Gnanasekar Munirathinam. Evaluating the therapeutic effects of methylene blue against prostate cancer [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 2958.
Abstract 2960: NCX4040, a nitric oxide (NO) donating form of aspirin as a potential treatment for prostate cancer
Authors: CHINNAPAKA SOMAIAH , Munirathinam Gnanasekar
Published: August 2019

Nonsteroidal anti-inflammatory drugs (NSAIDs) have been shown to be beneficial in preventing various types of cancers including prostate cancer (PCa). Of all the NSAIDs in use, Aspirin appears to be one of the promising anticancer agents for PCa management. However, experimental and epidemiological studies indicated that larger doses of Aspirin and long term supplementation is required to achieve anticancer effect with accompanied side effects. Therefore, our study focused on evaluating Aspirin derivative, namely NCX4040 which is a NO releasing form of Aspirin as a potent alternative for PCa treatment while minimizing the side effects. In the present study, we assessed the anticancer effects of NCX4040 on PC-3, a bone metastatic PCa cell line employing various in-vitro cellular assays. In our preliminary studies, we compared the effects of NCX4040 with that of Aspirin and NO releasing compound DETA/NO on PC-3 cells using MTT as read out. These results showed that NCX4040 is highly potent in inhibiting the viability of PC-3 cells when compared to Aspirin or DETA/NO treatments. Hence, further studies were focused on characterizing the anticancer properties as well as the underlying anticancer mechanism of NCX4040. Of note, our results showed that NCX4040 robustly inhibited the colony forming ability of PC-3 cells in-vitro implicating its antitumorigenic potential. In addition to inhibiting colony formation of PC-3 cells, NCX4040 also suppressed the Transwell matrigel invasion of PC-3 cells suggesting the antimetastatic effects of NCX4040. The observed antimetastatic effect of NCX4040 in PC-3 PCa cells appears to be via targeting actin cytoskeleton as confirmed by confocal microscopic analysis using phalloidin stain. Following these findings, we performed cell cycle analysis to determine whether NCX4040 enforces cell cycle arrest in PC-3 cells. These results showed that NCX4040 induced robust G0 cell cycle arrest in a dose dependent manner. Moreover, flow cytometry analysis using Annexin V and PI staining revealed that NCX4040 induced potent apoptotic cell death in PC-3 cells. To delineate the underlying anticancer mechanism of NCX4040, PC-3 cells treated with NCX4040 was subjected to apoptotic protein array analysis. Results of this study inferred that NCX4040 down regulated the expression of key anti-apoptotic proteins such as Survivin, XIAP and IAPs. In conclusion, our study suggests that NCX4040 is a promising alternative agent for PCa treatment.

Citation Format: SOMAIAH CHINNAPAKA, Gnanasekar Munirathinam. NCX4040, a nitric oxide (NO) donating form of aspirin as a potential treatment for prostate cancer [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 2960.
Abstract 3219: In vivo efficacy and safety evaluation of anti-human PD-1 and CD40 mAbs using double humanized PD-1/CD40 mouse model
Authors: Guo Yanan
Published: August 2019

In recent years, monoclonal antibodies have been successfully used in clinical trials to block or activate key mediators of immune checkpoint pathways, including CTLA4, PD-1, PD-L1, CD40 and others. Combination therapy can promote antigen release and T cell priming, T cell activation and homing, which helps to overcome tumor immune-evasive mechanisms and maximize efficacy, ultimately benefitting most patients. However, along the IO drug development process, in vivo efficacy models have always been a rate-limiting step, especially to test combination therapy using two IO antibodies.

The transmembrane protein receptor CD40 is a member of the tumor necrosis factor (TNF) receptor super family and is involved in co-stimulation of immune cells. CD40 is expressed by antigen-presenting cells (APCs) including dendritic cells (DCs), B-cells, macrophages, and monocytes. It has the ability to "wake" DCs to prime effective cytotoxic T-cell responses. Thus, effective anti-CD40 antibodies provide an ideal therapy alternative in combination with other IO antibodies.

To evaluate the in vivo efficacy and safety (immune-related adverse events) of CD40 and PD-1 combination antibodies, we developed a double humanized B-hPD-1/hCD40 mouse model. In this model, mouse PD-1 gene exon 2, and CD40 gene exons 2-7 were replaced with their human counterparts. Murine colon cancer MC38 cells were subcutaneously implanted into homozygous B-hPD-1/hCD40 mice. We compared the effect among single hPD-1 antibody Keytruda, single hCD40 antibody Selicrelumab and combination of the two.

Combination treatment of Selicrelumab and Keytruda show higher inhibitory effects than single antibody treatments. The blood chemistry assays showed that the CD40 antibody had no toxicity effects compared with the control group. Based on these findings, we conclude that the B-hPD-1/hCD40 double-humanized mouse model is a powerful tool for in vivo efficacy and safety evaluation of hPD-1 and hCD40 antibodies for combination therapy.

Citation Format: Yanan Guo. In vivo efficacy and safety evaluation of anti-human PD-1 and CD40 mAbs using double humanized PD-1/CD40 mouse model [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 3219.
Abstract 3241: Novel anti-CD40 antibodies demonstrate anti-tumor activity in humanized mouse models
Authors: Guo Chaoshe , Yang Benny(Yi) , Ni Jian , Guo Yanan , Gan Tian
Published: August 2019

CD40 (Cluster of Differentiation 40) is a member of the TNF-receptor superfamily. CD40 is a co-stimulatory protein found primarily on antigen presenting cells, including B cells, macrophages and dendritic cells. Its ligand, CD40L (CD154) is predominantly expressed on T helper cells, whose engagement activates antigen presenting cells and induces a variety of positive downstream effects. Whereas most immune checkpoint receptors are expressed on the surface of T cells, CD40 stands out as it is expressed by antigen presenting cells and controls the proper activation of these cells. In turn, CD40 activation exerts a significant impact on T cell-mediated activity against cancer cells. Several pharmaceutical companies are pursuing anti-CD40 therapies for treating hematological and solid cancers.

Therefore, it is imperative to develop novel CD40 antibodies to boost cancer patients’ own immunity in light of other successful immune checkpoint modulators. We asked whether we could find suitable anti-CD40 reagents that successfully combat developed tumors in preclinical settings. We designed a powerful screening strategy based on Biocytogen’s in vivo drug screening platform. Candidate antibodies were generated by immunizing wild-type mice with recombinant hCD40 protein. Hybridomas were then screened by high-throughput flow cytometry. Instead of examining the immune modulating activity in cell culture, we tested these antibodies in B-hCD40 mice bearing MC38 tumors and monitored the tumor growth. Through this straightforward screening strategy, several clones stood out by their unparalleled benefits. These clones were further selected for humanization. Furthermore, we identified that one clone works in concert with Keytruda in the dual humanized model of h-CD40 and h-PD1. In conclusion, we uncovered CD40 antibody candidates with the most potent anti-tumor activity which are promising as tools for cancer treatment in the preclinical setting.

Citation Format: Chaoshe Guo, Benny(Yi) Yang, Jian Ni, Yanan Guo, Tian Gan. Novel anti-CD40 antibodies demonstrate anti-tumor activity in humanized mouse models [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 3241.
Abstract 3277: IACS-9779, a development candidate that inhibits 2,3-dioxygenase (IDO) activity by blocking heme incorporation into IDO apoenzyme
Authors: Mseeh Faika , Hamilton Matthew M. , Marszalek Joseph R. , Rogers Norma E. , Parker Connor A. , Yu Simon S. , Liu Zhen , Reyna Naphtali J. , McAfoos Timothy , Virgin-Downey Brett W. , Leonard Paul G. , Cross Jason B. , Feng Ningping , Harris Angela L. , Zuniga Andy M. , Mikule Keith , Tremblay Martin , Jiang Yongying , Mahendra Mikhila , Pang Jihai , Wu Qi , Xu Quanyun , Heffernan Timothy P. , Jones Philip , Lewis Richard T.
Published: August 2019

Increased expression of IDO1 is believed to create a tumor microenvironment that is immunosuppressive. In the course of our research directed at identifying potent and selective inhibitors of IDO1, we identified a class of compounds that inhibited IDO1 activity in a cellular context, but not in isolated enzymatic assays. We have conducted detailed mechanistic studies and shown that these molecules inhibit IDO1 by binding to the apo-enzyme, thus preventing the incorporation of the heme-cofactor into the active site of the holo-enzyme.

Through an extensive medicinal chemistry campaign, we optimized a series of orally bioavailable, highly potent and selective inhibitors of IDO1 that possess excellent pharmacological properties. For several lead molecules, pharmacokinetic (PK) - pharmacodynamic (PD) relationships were established in whole blood and SKOV3 xenograft assays. The inhibition of IDO1 in a human whole-blood assay correlated well with the suppression of tumor kynurenine (KYN) that was observed in SKOV3 xenografts. At plasma concentrations of 3 µM, IACS-9779 supressed tumor KYN levels by 90%. IACS-9779 was well tolerated with excellent in vivo PK properties across multiple preclinical species, and a human PK prediction consistent with a low daily dose needed for full suppression of KYN production via IDO1.

Note: This abstract was not presented at the meeting.

Citation Format: Faika Mseeh, Matthew M. Hamilton, Joseph R. Marszalek, Norma E. Rogers, Connor A. Parker, Simon S. Yu, Zhen Liu, Naphtali J. Reyna, Timothy McAfoos, Brett W. Virgin-Downey, Paul G. Leonard, Jason B. Cross, Ningping Feng, Angela L. Harris, Andy M. Zuniga, Keith Mikule, Martin Tremblay, Yongying Jiang, Mikhila Mahendra, Jihai Pang, Qi Wu, Quanyun Xu, Timothy P. Heffernan, Philip Jones, Richard T. Lewis. IACS-9779, a development candidate that inhibits 2,3-dioxygenase (IDO) activity by blocking heme incorporation into IDO apoenzyme [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 3277.
Abstract 3304: CNS metastases in prostate cancer: an analysis of SEER database
Authors: Mei Lin , Al-Juhaishi Taha , Poklepovic Andrew , Paul Asit
Published: August 2019

Metastases to central nervous system (CNS) from various solid tumors, such as lung cancer (40-50%), breast cancer (15-25%) and melanoma (5-20%), are relatively common and confer a poor prognosis. CNS metastases from prostate cancer are uncommon. Due to the rarity of this disease and lack of studies, little is known about the clinical and pathological characteristics as well as outcome of these patients. CNS metastases also have therapeutic implications in terms of targeting hypothalamic-pituitary axis for androgen deprivation as well as CNS penetration of systemic chemotherapy agents. The objective of this study is to analyze clinicopathologic features and outcome of prostate cancer patients with CNS metastases in Survival, Epidemiology, and End Result (SEER) database. Between 2010 and 2015, a total of 23,776 patients with metastatic prostate were identified. Among them, 23,468 cases were adenocarcinoma and 308 cases were prostate cancer with neuroendocrine differentiation. A number of variables, including age, race, prostate-specific antigen (PSA) level, Gleason score, and prior radiotherapy, site of metastases were extracted. Primary outcome were overall survival (OS) and disease specific survival (DSS). CNS metastases were identified in 208 (0.89%) and 11 (3.57%) patients with adenocarcinoma and neuroendocrine tumor, respectively. Majority of patients (56.2%) were 65 years or older. Median OS and DSS were 13 months and 14 months, respectively. In comparison to patients with lymph nodes metastases only (n=8,103), the median OS and DSS were 40 months and 58 months, respectively. In patients with bone dominant metastasis (n=14,407), the median OS and DSS were 27 months and 34 months, respectively. Multivariate analysis identified age was an adverse predictive factor in patients with CNS metastasis. Race, high Gleason score (≥9), high PSA level (≥98ng/ml) and prior radiotherapy were not associated with OS and DSS. In conclusion, CNS metastases are more common in neuroendocrine tumor than in adenocarcinoma of prostate, and it confers poor survival outcome irrespective of histological subtypes. OS and DSS were worse in patients with prostate cancer and CNS metastases compared to those with lymph node metastases only or bone-dominant metastases.

Note: This abstract was not presented at the meeting.

Citation Format: Lin Mei, Taha Al-Juhaishi, Andrew Poklepovic, Asit Paul. CNS metastases in prostate cancer: an analysis of SEER database [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 3304.