bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 4765: MERTK is a potential therapeutic target in osimertinib-resistant non-small cell lung cancer
Authors: Yan Dan , Huelse Justus , Parker Rebecca , Wang Xiaodong , Frye Stephen V. , Earp H. Shelton , DeRyckere Deborah , Graham Douglas
Published: August 2019

Non-small cell lung cancer (NSCLC) remains a global problem causing more deaths in both men and women than any other cancer worldwide with an urgent need for more efficacious treatments. During the last decade the therapeutic landscape for NSCLC has been profoundly changed with the discovery of activating mutations in Epidermal Growth Factor Receptor (EGFR) (EGFRMT). Osimertinib, a 3rd generation EGFR TKI, was recently FDA-approved as a front-line agent for newly diagnosed EGFRMT NSCLCs due to its superior efficacy relative to earlier-generation EGFR TKIs in the international FLAURA trial. However, unmet clinical needs have arisen in conjunction with osimertinib use, including understanding mechanisms of osimertinib resistance and developing novel approaches to prevent or reverse resistance and/or enhance osimertinib efficacy in responsive patients. Our group previously identified MERTK receptor tyrosine kinase as a potential therapeutic target in NSCLC and developed MRX-2843, a novel small molecule inhibitor with dual MERTK and FLT3 activity, which is currently in Phase I clinical trials. Here, we report upregulation of MERTK and its ligand PROS1 in xenograft tumors derived from the EGFRMT H4006 NSCLC cell line and treated with osimertinib relative to vehicle-treated tumors. MERTK expression was also increased in osimertinib-resistant derivatives of the H4006 and H4011 cell lines generated by culture in escalating doses of osimertinib. However, overexpression of exogenous MERTK in H4006 cells was not sufficient to confer osimertinib resistance. In contrast, stimulation with TAM kinase ligands GAS6 or PROS1 protected H4006 cells from osimertinib treatment, as indicated by restoration of AKT, ERK, and ribosomal S6 phosphorylation in the presence of osimertinib. Together these data implicate MERTK as a mediator of resistance to osimertinib. Indeed, combined treatment with osimertinib and MRX-2843 effectively blocked PI3K-AKT and MAPK-ERK signaling and mediated synergistic inhibition of colony formation in osimertinib-resistant H4006 cells. Thus, MERTK inhibition may be an effective therapeutic strategy to re-sensitize osimertinib-resistant NSCLCs to EGFR TKI treatment.

Citation Format: Dan Yan, Justus Huelse, Rebecca Parker, Xiaodong Wang, Stephen V. Frye, H. Shelton Earp, Deborah DeRyckere, Douglas Graham. MERTK is a potential therapeutic target in osimertinib-resistant non-small cell lung 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 4765.
Abstract 4769: Gene expression profiling of response to short- and long-term treatment of LSZ102 in MCF7 cells
Authors: Yip Choi Lai Tiong , Korn Joshua , Ruddy David , Rakiec Daniel , Stuart Darrin , Gaither Alex
Published: August 2019

Breast cancer is the most common cancer in women worldwide, and the second most common cause of cancer death in woman in the US. Close to 80% of breast cancer are estrogen receptor (ER) positive and can benefit from antiestrogen treatment. LSZ102 is a novel oral selective estrogen receptor degrader (SERD) in clinical development, which exhibits potent activity in inducing ER degradation, modulating ER-target genes transcription and inhibits the proliferation of breast cancer cells both in-vitro and in-vivo. In this study, we have investigated the effects of short term and prolonged LSZ102 treatment on global gene expression in ER positive MCF-7 breast cancer cells. MCF-7 cells were treated with LSZ102 for 24 hours, 8 weeks and 14 weeks, followed by transcriptome profiling by RNA sequencing. Our findings show LSZ102 effectively inhibited ER transcriptional activity and lead to a sustained suppression of estrogen responsive genes expression in LSZ102-treated cells of all time points. We also observed a cumulative up-regulation of EPAS1, a hypoxia-inducible transcription factor. In addition, we found up-regulation of EGFR and ERBB2 genes and regulated genes in cells with prolonged treatment of LSZ102. In summary, our results suggest a possible role of EPAS1 in eliciting a compensatory growth promoting signals in breast cancer cells adapted to ER inhibition.

Citation Format: Choi Lai Tiong Yip, Joshua Korn, David Ruddy, Daniel Rakiec, Darrin Stuart, Alex Gaither. Gene expression profiling of response to short- and long-term treatment of LSZ102 in MCF7 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 4769.
Abstract 4771: The addition of toca 511 and 5-FC to temozolomide improves response in a temozolomide-resistant murine glioblastoma model and correlates with toca 511 dose
Authors: Rodriguez-Aguirre Maria E. , Viaud Sophie , Mendoza Daniel , Montellano Tiffany , Ostertag Derek G. , Gruber Harry , Jolly Douglas , Bentley Cornelia
Published: August 2019

Background: Toca 511 (vocimagene amiretrorepvec), an amphotropic retroviral replicating vector (RRV), can successfully and safely deliver a functional, optimized yeast cytosine deaminase (CD) gene to tumors. Within infected cells, CD converts 5-fluorocytosine (5-FC) to the anti-cancer drug 5-fluorouracil (5-FU). The combination of Toca 511 with oral extended release 5-FC (Toca FC), is currently being evaluated in a randomized phase III clinical trial for recurrent high grade glioma (glioblastoma (GBM) and anaplastic astrocytoma) (NCT02414165, Toca 5). Temozolomide (TMZ), in combination with radiation therapy, is the standard of care used for first-line chemotherapy treatment of patients with GBM, the most common and aggressive form of primary brain cancer. Previously, we have shown that: (1) Toca 511/5-FC treatment provides durable response in a syngeneic murine glioma model and supports anti-tumor immune memory; (2) The combination of TMZ and Toca 511/5-FC had synergistic efficacy in a TMZ-sensitive human glioma nude mouse model; (3) TMZ did not inhibit the efficacy of Toca 511/5-FC in a TMZ resistant murine glioma syngeneic model; (4) Toca 511/5-FC caused significant radiosensitization in a radioresistant murine glioma model.

Results: To assess the interaction of TMZ with escalating doses of Toca 511 (as defined by percent of tumor transduction by RRV), an orthotopic TMZ-resistant murine glioma model, Tu-2449, was utilized. These results show that moderate levels of tumor transduction of Toca 511 (30% - 50%) with 5-FC treatment prolongs survival in the presence of TMZ compared with lower transduction rates (10%). Additionally, mice treated with Toca 511/5-FC and TMZ are being tested for the establishment of anti-tumor immune memory.

Conclusion: These results demonstrate that (1) survival correlates with the transduction levels of Toca 511 when combined with temozolomide in the Tu-2449 orthotopic glioma model and (2) that this combination may support anti-tumor immune memory. These studies along with prior work support evaluation of the combination of Toca 511/5-FC with temozolomide in patients with newly diagnosed GBM (NRG-BN006).

Citation Format: Maria E. Rodriguez-Aguirre, Sophie Viaud, Daniel Mendoza, Tiffany Montellano, Derek G. Ostertag, Harry Gruber, Douglas Jolly, Cornelia Bentley. The addition of toca 511 and 5-FC to temozolomide improves response in a temozolomide-resistant murine glioblastoma model and correlates with toca 511 dose [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 4771.
Abstract 4778: Identification of promoters for targeted gene expression in tumors
Authors: Sajib Abdul Mohin , Sandey Maninder , Morici Samantha , Schuler Bradley , Agarwal Payal , Smith Bruce
Published: August 2019

A variety of novel therapeutic approaches to cancer have been proposed in the past several decades. One of the most compelling is the use of gene therapy to treat tumors. Restricting the expression of these genes to tumor avoids toxicity in normal cells and the concurrent side effects that this activity engenders. Promoter sequences can be used as a tool for transcriptional targeting, by restricting the expression of the therapeutic genes, or, in the case of oncolytic virotherapy, the replication of the viral agent. Several tumor-upregulated promoters have been identified in a wide variety of human tumors. Given the recognition of the validity of canine tumors as comparative models of human disease, it is critical to determine the activities of these promoters in the canine system. To achieve this goal, endogenous cSurvivin, cTERT, and cCXCR4 promoter activity was evaluated in a panel of canine normal and tumor cell lines/tissues and primary tumors by RT-Q-PCR. These three promoters, along with an E2F modified Canine Adenovirus 2 E1a promoter (E2F-E1a), were also assessed for their activity when supplied exogenously to tumor cells in an in vitro GFP reporter transfection assay. Endogenous expression of cTERT showed negligible differences between normal canine cells/tissues and tumor cell/tissues. cSurvivin showed elevated endogenous activity in non-hematopoietic tumor cells and moderately elevated activity in some normal tissues and hematopoietic tumors. cCXCR4 activity was elevated exclusively in a T-lymphoma cell line and a primary T-cell lymphoma. Exogenous expression in most cancer cell lines showed enhanced activity with most of the promoters tested. The percentage of cells expressing GFP was elevated in tumor compared to normal with all four tested promoters, when normalized to CMV-GFP expression. However, the relative fluorescence intensity varied significantly between cell lines, while the fluorescence intensity varied less between promoters within a specific cell line. Exogenous expression results did not correlate well with endogenous expression in tumor cells, but they did correlate well in normal cells. This disparity implies that promoter selection for transcriptional targeting of tumors should account for both endogenous and exogenous expression patterns. Additionally, these findings indicate that identification of a pan-cancer promoter is complex and that expression targeting may need to rely on selecting patient-specific promoters to drive the activity of therapeutic genes.

Citation Format: Abdul Mohin Sajib, Maninder Sandey, Samantha Morici, Bradley Schuler, Payal Agarwal, Bruce Smith. Identification of promoters for targeted gene expression in tumors [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 4778.
Abstract 4779: Coxsackievirus A11 as a novel oncolytic viral therapy for human colorectal cancer
Authors: Ogata Hisanobu , Wang Beibei , Miyamoto Shoei , Takishima Yuto , Sagara Miyako , Murahashi Mutsunori , Onishi Hideya , Tani Kenzaburo
Published: August 2019

Colorectal cancer (CRC) is a major cause of morbidity and mortality throughout the world. It is the third most common cancer worldwide and the fourth most common cause of death. FOLFOX is known as the effective chemotherapy for stage III and stage IV CRC. Oxaliplatin (L-OHP) is a third-generation platinum agent used among FOLFOX in CRC treatment. Patients with L-OHP-resistant CRC has a worse prognosis. Coxsackievirus is a virus that belongs to a family of nonenveloped, linear, positive-sense ssRNA viruses, Picornaviridae and the genus Enterovirus. Enteroviruses are among the most common and important human pathogens, and ordinarily its members are transmitted by the fecal-oral route. Coxsackieviruses share many characteristics with poliovirus. Coxsackieviruses are among the leading causes of aseptic meningitis. Recently, coxsackievirus A21, which is also called Cavatak, has already been reported to have the oncolytic activity. In this study, we tested whether coxsackievirus A11 (CVA11) has an oncolytic activity or not in CRC. Through our first screening assay, we found that CVA11 displayed the potent oncolytic activities in 4 of 5 various human CRC cell lines including two L-OHP-resistant CRC cells. Mechanistically, the CVA11-mediated cytotoxicity was significantly impaired when PI3K or Wnt inhibitor was added to the CVA11 infection. CVA11 had little effect on the L-OHP-resistant CRC. The pretreatment with L-OHP in L-OHP-resistant CRC cells displayed enhanced oncolytic activity of CVA11 in L-OHP-resistant CRC cells, whereas either L-OHP or CVA11 treatment alone caused slight oncolytic activity. These results suggest that L-OHP pretreatment could sensitize the susceptibility of CRC cells to CVA11 infection. Furthermore, our results of in vivo mouse study showed that intratumoral CVA11 administrations with pretreatment of L-OHP significantly suppressed the outgrowth of L-OHP-resistant xenografts. This combination therapy was more effective than the treatment with only CVA11. Altogether, our findings showed that the combination therapy with L-OHP and CVA11 displayed a remarkable oncolytic activity against L-OHP-resistant human CRC both in vitro and in vivo. Our study also provides the therapeutic potential for L-OHP-resistant CRC by Coxsackieviruse.

Note: This abstract was not presented at the meeting.

Citation Format: Hisanobu Ogata, Beibei Wang, Shoei Miyamoto, Yuto Takishima, Miyako Sagara, Mutsunori Murahashi, Hideya Onishi, Kenzaburo Tani. Coxsackievirus A11 as a novel oncolytic viral therapy for human colorectal 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 4779.
Abstract 4783: Highly efficient, non-viral precision genome engineering for the generation of personalized neoepitope-specific adoptive T cell therapies
Authors: Jacoby Kyle , Moot Robert , Lu William , Nguyen Diana , Sennino Barbara , Conroy Andrew , Purandare Bhamini , Litterman Adam J. , Urbinati Fabrizia , Foy Susan P. , Hunter Theresa , Tai Albert , Bethune Michael T. , Peng Songming , Dalmas Olivier , Franzusoff Alex , Mandl Stefanie J.
Published: August 2019

Methods used to engineer cells for adoptive cell therapies (ACT) utilizing receptors that are constant across many patients (CAR or shared Ag TCRs) typically rely on Lenti-, retro-, or adeno-associated virus to deliver specificity-altering sequences to T cells. However, for personalized therapies such as the generation of neoepitope-specific TCR T cell therapies, use of viral vectors is not feasible due to long manufacturing timelines and prohibitive per-patient costs. PACT Pharma has developed a highly efficient, DNA-mediated (non-viral) proprietary precision genome engineering approach to engineer neoepitope-specific primary human T cells. This method can be widely utilized to generate T cells at research scale, as well as for ex vivo manufacturing.

Briefly, genomes of individual primary human CD8 and CD4 T cells are engineered with site-specific nucleases in a single-step transfection process to yield efficient, targeted replacement of the endogenous TCR with the therapeutic neoTCR sequences. In this way, the expression of the endogenous TCR is abolished ensuring natural expression and regulation of the inserted neoTCR. The precision of neoTCR-T cell genome engineering was evaluated by Targeted Locus Amplification (TLA) for off-target integration hot spots or translocations, and by next generation sequencing based off-target cleavage assays and found to lack evidence of unintended outcomes.

Engineered neoepitope-specific T cells are highly functional as demonstrated by antigen-specific proliferation, killing and cytokine production. Phenotype and detailed functional characterization of PACTs neoTCR-P1 T cells were performed and are described in a separate abstract.

PACT’s precision genome engineering approach enables highly efficient generation of bespoke NeoTCR T cells for personalized adoptive cell therapy for patients with solid tumors. Furthermore, PACT precision genome engineering method is not restricted to the use in T cells and has also been applied successfully to other primary cell types, including natural killer and hematopoietic stem cells.

Citation Format: Kyle Jacoby, Robert Moot, William Lu, Diana Nguyen, Barbara Sennino, Andrew Conroy, Bhamini Purandare, Adam J. Litterman, Fabrizia Urbinati, Susan P. Foy, Theresa Hunter, Albert Tai, Michael T. Bethune, Songming Peng, Olivier Dalmas, Alex Franzusoff, Stefanie J. Mandl. Highly efficient, non-viral precision genome engineering for the generation of personalized neoepitope-specific adoptive T cell therapies [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 4783.
Abstract 4788: Novel hybrid benzothiazole screening in U87-MG glioblastoma cell line
Authors: Kyi Priscilla , Murray Desmond H. , Smith Denise L.
Published: August 2019

Benzothiazole is a heteroaromatic compound known for its wide range of bioactivities including anticancer, antiviral, antimicrobial, anti-inflammatory, anticonvulsant, antidiabetic, antihelminthic, and antitubercular activities. Research has shown that derivatives of benzothiazole exhibit inhibition of proliferation via apoptosis in various human cancer cell lines, such as liver cancer (Wang, et. al., 2010). In this study, a series of novel hybrid benzothiazole α-cyanostilbene derivatives and styrylbenzothiazole derivatives containing boronic acid and non-boronic acid pharmacophores were synthesized and their anti-cancer properties on U87-MG glioblastoma cells were investigated in vitro. U87-MG cells were incubated with synthesized novel hybrid compounds at varying concentration to determine the inhibitory concentration 50 (IC50) of the compounds. Most hybrid compounds displayed inhibitory effects on cell growth and the IC50 of the compounds varied depending on the nature of the pharmacophores. Moreover, boronic acid-containing compounds exhibit lower IC50 than nonboronic acid-containing compounds. Cell mobility has been investigated and we have found that treated cells were less mobile than untreated cells. In the future, we will be further screening for mobility, invasiveness, and mode of death of these novel hybrid benzothiazole treated glioblastoma cells.

Citation Format: Priscilla Kyi, Desmond H. Murray, Denise L. Smith. Novel hybrid benzothiazole screening in U87-MG glioblastoma cell line [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 4788.
Abstract 4792: Azetidine functionalized small-molecules potently inhibit STAT3 signaling and block tumor growth in human breast cancer xenografts
Authors: Yue Peibin , Lopez-Tapia Francisco , Fu Wenzhen , Brotherton-Pleiss Christine , Tius Marcus , Turkson James
Published: August 2019

The development of inhibitors of Signal transducer and activator of transcription (STAT) 3 for clinical application has faced significant challenges and to date there are no suitable STAT3 small molecule inhibitors in the clinic. We report a novel class of azetidine-based small molecules that potently and preferentially inhibit STAT3 DNA-binding activity in vitro, with IC50 of 300-800 nM, compared to IC50 of 17 μM or higher against STAT1 DNA-binding activity. In the human breast cancer MDA-MB-231 and MDA-MB-468 cells treated with the azetidine compounds, including H182, constitutive STAT3 DNA-binding activity and tyrosine phosphorylation were strongly inhibited in both time- and dose-dependent manner. By contrast, the azetidine compounds showed little effects on EGF-induced STAT1 activity, Ras-Raf-MEK-ERK or other STAT3-independent signaling pathways, or on other SH2 domain-containing proteins. Furthermore, H182 and other azetidine compounds strongly inhibited viability, anchorage-dependent and -independent growth, and colony survival of the human breast cancer cells, with IC50 of 1.0 - 1.9 μM, compared to the IC50 of 3.8 - 8.1 μM against the viability and growth of the immortalized breast epithelial MCF-10A cells. The migration of breast cancer cells in vitrowere similarly inhibited by treatment with low concentrations of H182 for 18 h, without changes in viable cell number. Analysis of STAT3 downstream genes show that the expression of VEGF and survivin were significantly suppressed in breast cancer cells treated with the azetidine compounds. In vivo evaluation of antitumor efficacy shows H182 inhibited growth of human breast tumor xenografts. Studies together identify the azetidine-functionalized small molecules as potential candidate compounds for clinical development as novel therapeutic agents for human breast and other cancers that harbor aberrantly-active STAT3.

Citation Format: Peibin Yue, Francisco Lopez-Tapia, Wenzhen Fu, Christine Brotherton-Pleiss, Marcus Tius, James Turkson. Azetidine functionalized small-molecules potently inhibit STAT3 signaling and block tumor growth in human breast cancer xenografts [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 4792.
Abstract 4801: 2’-Hydroxyflavanone, a diet-derived citrus bioflavonoid, inhibits in vitro and in vivo growth of breast cancer cells by targeting RLIP
Authors: Singhal Sharad S. , Chikara Shireen , Singhal Jyotsana , Horne David , Salgia Ravi , Awasthi Sanjay
Published: August 2019

Breast cancer remains one of the major causes of cancer deaths in women. Consumption of citrus-fruits is associated with reduced incidence of breast cancer, the most common cancer diagnosed in women across the globe. In this study, we investigated the anticancer potential of 2-Hydroxyflavanone (2HF) in breast cancer. 2HF, a citrus-bioflavonoid, has demonstrated anticancer properties in various cancers, but its anticancer role in breast cancer has not been well studied. We investigated the in-vitro and in-vivo growth inhibitory effects of 2HF in an array of BC lines and in xenograft mouse models of ER-positive and HER2-positive breast cancer cells. Compared to control, 2HF treatment reduced cell viability of breast cancer cells, while, no growth inhibitory effects were observed in non-tumorigenic breast epithelial cells. Further, 2HF inhibited the expression of RLIP, a stress-defensive and anti-apoptotic protein, which is over-expressed in breast cancer cells and simultaneously reduced proliferation of breast cancer cells. Nude mice bearing MCF7 or SKBR3 breast cancer cells xenografts treated with either 2HF or targeting RLIP by RLIP-antisense or RLIP-antibody treatment had significantly lower tumor-weight as compared to corresponding controls. In addition, Western-blotting and immunohistochemical analysis of tumor tissue from control and treatment group mice showed that 2HF decreased protein expression levels of RLIP, and the decrease was similar to those seen following RLIP-antisense treatment. Furthermore, 2HF decreased expression of Ki67, CD31, vimentin, inhibited phosphorylation of AKT and expression of survivin and Bcl2, and increased levels of Bax, E-cadherin, and cleaved-PARP. Therefore, our results indicate that 2HF may suppress breast cancer growth in-vitro and in-vivo by targeting RLIP, and may serve as a potential adjuvant treatment in breast cancer patients. (This work was supported in part by the Department of Defense grant W81XWH-16-1-0641. Funding from the Beckman Research Institute of City of Hope is also acknowledged).

Citation Format: Sharad S. Singhal, Shireen Chikara, Jyotsana Singhal, David Horne, Ravi Salgia, Sanjay Awasthi. 2’-Hydroxyflavanone, a diet-derived citrus bioflavonoid, inhibits in vitro and in vivo growth of breast cancer cells by targeting RLIP [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 4801.
Abstract 4808: Preclinical studies of the combination of ONC201, radiotherapy and Temozolomide against GBM, DIPG and ATRT cell lines
Authors: Zhou Lanlan , El-Deiry Wafik S.
Published: August 2019

The American Cancer Society predicts, in 2018, there will be ~23,880 new cases and 16,830 deaths caused by primary cancer of the brain and spinal cord, in the US. CNS tumors are the most common cause of cancer-related deaths in adolescents and young adults. First-in-class small-molecule imipridone ONC201 can act as a dual inhibitor of ERK/AKT and can induce an integrated stress response (ISR), pro-apoptotic TRAIL receptor DR5 activation, cancer stem cell depletion, and growth arrest. ONC201 crosses the blood brain barrier and has demonstrated clinical benefits in glioblastoma patients. Radiation therapy is used alone or in combination with surgery and/or chemotherapy such as Temozolomide in the treatment of primary or metastatic brain tumors. We hypothesized that ONC201 may synergize with radiotherapy and Temozolomide in brain tumor treatment. Glioblastoma (GBM), diffuse intrinsic pontine glioma (DIPG) and atypical teratoid rhabdoid tumor (ATRT) cell lines were tested in this study. Cell viability and colony formation assays were performed with ONC201 alone up to 20 μM or in combination with radiotherapy up to 8 Gy and Temozolomide up to 100 μM. Western blots were used to document apoptosis of treated cells. We observed synergy between ONC201 and radiation and between ONC201 and Temozolomide with the best combination indices of 0.51 and 0.21 respectively. Further studies are evaluating the role of dopamine receptors engaged by ONC201, the ISR and TRAIL pathway in the synergistic effect that would support further development of the triple combination therapy in brain tumors.

Citation Format: Lanlan Zhou, Wafik S. El-Deiry. Preclinical studies of the combination of ONC201, radiotherapy and Temozolomide against GBM, DIPG and ATRT cell lines [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 4808.
Abstract 4818: Fc-mediated mechanism of action for the novel EGFR-cMET bispecific antibody (JNJ-61186372) in non-small cell lung cancer
Authors: Vijayaraghavan Smruthi , Bushey Barbara , Lipfert Lorriane , Nanjunda Rupesh , Lacy Eilyn R. , Buckley Peter , Laquerre Sylvie , Lorenzi Matthew V. , Moores Sheri
Published: August 2019

JNJ-61186372 (JNJ-372) is an anti-EGFR and cMet bispecific antibody with an active Fc backbone (IgG1) designed to treat non-small cell lung cancer (NSCLC) disease. A first-in-human study is currently being conducted to assess the safety and preliminary efficacy of JNJ-372 in patients with advanced NSCLC. Early data suggests that JNJ-372 can induce partial responses in subjects with diverse populations of EGFR-mutated NSCLC, including Exon 20ins as well as TKI resistance mutations.

Our previous pre-clinical in vivo studies showed that the Fc inactive version (IgG2sigma) of the EGFR/cMet antibody was significantly impaired in its ability to inhibit tumor growth compared to the Fc active JNJ-372. The IgG2sigma variant also reduced the ability of the bispecific antibody to mediate downregulation of EGFR, cMet and downstream signaling components. This suggested that the interaction of the Fc arm with the Fc receptors on the innate immune cells play a crucial role in the mechanism of action of JNJ-372. While JNJ-372 has demonstrated antibody dependent cellular cytotoxicity (ADCC) and antibody dependent cellular phagocytosis (ADCP) in vitro, the potential role of Fc interactions in downregulation of EGFR and cMet was not well understood.

We explored the different Fc-mediated immune effector functions of JNJ-372 and interrogated how they contribute to EGFR and cMet downregulation and overall efficacy. In NSCLC cell lines, JNJ-372 induced Fc-mediated dose-dependent ADCC and ADCP but not complement-dependent cytotoxicity. Further, the presence of isolated human immune cells (PBMC) significantly enhanced JNJ-372 mediated EGFR and cMet downregulation and dose-dependent tumor cell killing. Studies are in progress to better understand which immune cells and which Fc receptor interactions are essential for the drug efficacy through immune cell depletion and FcR blocking studies; such studies may help guide clinical biomarker development. This work elucidates a novel Fc-dependent mechanism of action for the EGFR-cMet bispecific antibody.

Citation Format: Smruthi Vijayaraghavan, Barbara Bushey, Lorriane Lipfert, Rupesh Nanjunda, Eilyn R. Lacy, Peter Buckley, Sylvie Laquerre, Matthew V. Lorenzi, Sheri Moores. Fc-mediated mechanism of action for the novel EGFR-cMET bispecific antibody (JNJ-61186372) in non-small cell lung 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 4818.
Abstract 4821: Development of a potent Trop-2 antibody-drug conjugate, BAT8003, for the treatment of Trop-2 positive gastric tumors
Authors: Tang Weijia , Mei Xingxing , Ou Ziqiang , Gan Jirong , Li Shengfeng , Yu Jin-Chen
Published: August 2019

Trophoblast cell surface antigen 2 (Trop-2) is overexpressed on many epithelial carcinomas while having low levels of expression on normal tissue. Overexpression of Trop-2 has been correlated with poor prognosis in several solid tumors. Trop-2 is overexpressed in ~60% of gastric cancer, which causes more than 500,000 mortalities every year in China. We have developed a Trop-2 antibody-drug conjugate (ADC), BAT8003, that utilizes an anti-Trop-2 IgG1 antibody, engineered for site-specific conjugation, a novel uncleavable linker, and a potent maytansine derivative as the payload. Site-specific conjugation allows for a more controllable drug-antibody ratio (DAR), resulting in a more homogenous product. BAT8003 also incorporates increased ADCC effects through afucosylation to further enhance potency. Here we show that BAT8003 is internalized in Trop-2 positive cells. It inhibits proliferation of a number of different Trop2-overexpressed tumor cells with IC50s of ~1 nM. In a gastric cancer (NCI-N87) mouse xenograft model, BAT8003 demonstrates strong inhibition activity on tumor growth at doses of 5 mg/kg and 15 mg/kg. A multiple dose toxicity study in monkey reveals that the highest non-severely toxic dose (HNSTD) of BAT8003 is 20 mg/kg when dose once every 3 weeks. The preclinical profile of BAT8003 warrants further clinical development for the treatment of gastric cancer as well as other Trop-2 over-expressing cancers.

Citation Format: Weijia Tang, Xingxing Mei, Ziqiang Ou, Jirong Gan, Shengfeng Li, Jin-Chen Yu. Development of a potent Trop-2 antibody-drug conjugate, BAT8003, for the treatment of Trop-2 positive gastric tumors [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 4821.
Abstract 4830: ABX9xx: A bispecific centyrin that synergizes to attenuate intracellular signaling in Met/EGFR positive tumors
Authors: Addis Russ , Kolakowski Robert , Kulkarni Swapnil , Gorsky Josh , Meyer Rebecca , ONeil Karyn
Published: August 2019

ABX9xx is a bispecific Centyrin targeting the Met-EGFR axis, an emerging cancer treatment paradigm. Centyrins are small, chemically simple proteins based on a human FN3 domain that can be engineered to have high affinity for a selected target and are easily linked to form multi-specific binders. The standard of care for patients with EGFR mutant non-small cell lung cancer (NSCLC) is treatment with small molecule tyrosine kinase inhibitors (TKI) including recently approved third generation molecules (ie. osimertinib). Despite initial promising responses with TKIs, most patients regress within 12-14 months due to resistance. A predominant resistance mechanism dependent on Met amplification and signaling has been described from both in vitro experiments and clinical data. Targeting Met and EGFR with a bispecific ligand blocking inhibitor that attenuates intracellular signaling may provide a significant efficacy advantage and reduced side effect profile compared to small molecule TKI combinations. Exploiting the potential for avidity on tumor tissue, a bispecific ligand blocking inhibitor of Met and EGFRis also anticipated to provide improved selectivity for tumor tissue that overexpress both receptors compared to normal tissue with lower receptor expression. To enable sustained exposure while maintaining small size for good tumor penetration, ABX9xx includes a domain comprising an albumin binding Centyrin. ABX9xx activity on Met, EGFR and downstream signaling proteins was confirmed on several lung cancer cell lines in vitro including those carrying clinically observed mutations in Met and EGFR. ABX9xx inhibited EGFR phosphorylation independent of EGFR mutational status with higher potency than osimertinib. In addition, ABX9xx inhibited Met phosphorylation with higher potency than Crizotinib, an ALK inhibitor with cross reactivity to Met, currently used to treat Met positive patients. The pharmacokinetic properties of ABX9xx in animal models suggest a suitable dosing paradigm in patients. Together the data provides a strong rationale for advancing ABX9xx into clinical development for NSCLC and other cancers where EGFR and MET are drivers of tumor progression.

Citation Format: Russ Addis, Robert Kolakowski, Swapnil Kulkarni, Josh Gorsky, Rebecca Meyer, Karyn ONeil. ABX9xx: A bispecific centyrin that synergizes to attenuate intracellular signaling in Met/EGFR positive tumors [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 4830.
Abstract 4831: Using reporter-based bioassays and engineered TNFalpha and VEGF+ target cells to measure Fc-mediated ADCC and CDC activity of anti-TNFá and anti-VEGF therapeutic antibodies
Authors: Garvin Denise , Grailer Jamison , Moravec Rich , Hartnett Jim , Heid Chris , Fan Frank , Cong Mei , Cheng Jey
Published: August 2019

Fc receptor-mediated effects contribute to the therapeutic response in terms of efficacy and safety of anti-TNF antibodies. Measurement of Fc-mediated antibody-mediated cellular cytotoxicity (ADCC) and complement-dependent cytotoxicity (CDC) during antibody drug discovery and development is not only important for antibodies that harness ADCC and/or CDC as their primary mechanism of action (e.g. rituximab, trastuzumab), but also for antibodies designed to target and block soluble ligands such as TNFα and VEGF. We previously reported the development of a cell-based reporter bioassay platform which has been used to measure ADCC and ADCP mediated through FcgRI, FcgRIIa and FcgRIIIa. These reporter bioassays exhibit the specificity, accuracy, precision, and robustness necessary for qualification according to ICH guidelines and have been used extensively to characterize and measure the potency of antibody-based biologics drugs that target cell surface immune receptors. In the current study, we sought to evaluate Fc-mediated ADCC and CDC activities of therapeutic antibodies designed to target and block soluble ligands including TNFa and VEGF. To measure ADCC activity of anti-TNFa and anti-VEGF blocking antibodies, we developed engineered target cells that express either membrane-bound TNFα or VEGF. When used as target cells with reporter-based effector cells expressing a relevant FcgR, ADCC activity of adalimumab (anti-TNFa) and bevacizumab (anti-VEGF) was detected in a specific and dose-dependent manner. Similarly, when used in a luminescence-based CDC assay, the engineered target cells elicited an appropriate FcgR-mediated response. The assay signals demonstrated IgG isotype specificity as IgG4 variants showed minimal activity in both ADCC and CDC assays. Our results demonstrate that the combined use of cell-based reporter bioassays with target cells engineered to express membrane-bound soluble ligands can provide a simple, specific, and quantitative platform to measure Fc-mediated effector functions of therapeutic antibodies targeting soluble ligands.

Citation Format: Denise Garvin, Jamison Grailer, Rich Moravec, Jim Hartnett, Chris Heid, Frank Fan, Mei Cong, Jey Cheng. Using reporter-based bioassays and engineered TNFalpha and VEGF+ target cells to measure Fc-mediated ADCC and CDC activity of anti-TNFá and anti-VEGF therapeutic antibodies [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 4831.
Abstract 4833: Antibody-drug conjugate targeting glypican-1 shows tumor growth inhibition in cholangiocarcinoma
Authors: Yokota Keiichiro , Serada Satoshi , Tsujii Shigehiro , Hiramatsu Kosuke , Namikawa Tsutomu , Murakami Ichiro , Hanazaki Kazuhiro , Naka Tetsuji
Published: August 2019

Cholangiocarcinoma is one of the most highly malignant cancers. Many patients need systemic chemotherapy for tumor development and recurrence, but their prognosis is poor. Therefore new treatment options are urgently required. We confirmed that the expression of glypican-1 (GPC1) was enhanced in cholangiocarcinoma. GPC1 is a cell surface membrane protein and has been reported as a poor prognostic factor in pancreatic cancer. In this study, we aimed to develop a new therapy for cholangiocarcinoma by the antibody-drug conjugate (ADC) targeting GPC1. By immunohistochemical analysis, enhanced expression of GPC1 was observed in clinical specimens of cholangiocarcinoma. KKU-055 and KKU-100 cells, which are cell lines of cholangiocarcinoma, had high expression of GPC1. In the KKU-055 xenograft model when we administered fluorescently-labeled GPC1 antibody to mice, it accumulated in tumor tissue. We developed a new anti-GPC1 monoclonal antibody (mAb) with highly internalizing activity. The anti-GPC1 mAb was conjugated with the cytotoxic agent monomethyl auristatin F (MMAF). GPC1-ADC showed potent antitumor effect toward KKU-055 and KKU-100 cells compared with the control ADC. In the KKU-055 xenograft model, GPC1-ADC had significant and potent tumor growth inhibition in a dose-dependent manner. In summary, our newly-developed GPC1-ADC showed significant tumor growth inhibition against GPC1-positive cholangiocarcinoma cell lines. Our preclinical data demonstrated that targeting GPC1 by ADC is a promising therapy for GPC1-positive cholangiocarcinoma.

Citation Format: Keiichiro Yokota, Satoshi Serada, Shigehiro Tsujii, Kosuke Hiramatsu, Tsutomu Namikawa, Ichiro Murakami, Kazuhiro Hanazaki, Tetsuji Naka. Antibody-drug conjugate targeting glypican-1 shows tumor growth inhibition in cholangiocarcinoma [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 4833.
Abstract 4838: Leflunomide inhibits the growth of LKB1-inactivated tumors in both xenograft and immunocompetent genetically engineered mouse models
Authors: Jin Rui , Liu Xiuju , Peng Wei , Fan Yijian , Liu Boxuan , Yang Shuanying , Xing Chun-gen , Gilbert-Ross Melissa , Marcus Adam , Zhou Wei
Published: August 2019

Introduction: LKB1 is the third mostly frequently mutated gene in lung adenocarcinoma, and is mutually exclusive with EGFR mutations. Furthermore, a recent clinical trial indicated that lung cancers with LKB1 mutations are unlikely to respond to immune checkpoint therapy. Therefore, there is an urgent need for novel therapies for LKB1-inactivated lung cancer. We previously demonstrated that leflunomide preferentially induces apoptosis in LKB1-null lung cancer cell lines in vitro. Here, we evaluated the effect of leflunomide in both a xenograft model and immune-competent genetically engineered mouse model (GEMM).

Methods: LKB1-null H460 xenograft mice were treated with leflunomide (35mg/kg/day) for 23 days through oral gavage and tumor volume was measured every the other day. KrasG12D/Lkb1-/-Luciferase GEMM mice were treated with leflunomide (30mg/kg/day) for 44 days and tumor burden was measured by bioluminescence imaging (BLI) score every week. At the end of experiments, tumors were dissected, weighed, and analyzed by immunohistochemistry.

Results: Although leflunomide is an FDA-approved agent to treat rheumatoid arthritis, its immune suppression function did not facilitate but inhibited the growth of KrasG12DLkb1-/- lung adenocarcinoma. This suppression was also observed in H460 xenografts, and leflunomide treatment did not alter animal weight in either model. Furthermore, distant cancer metastasis was not observed in leflunomide-treated GEMM. In residual tumors from the treated group, cleaved caspase-3 was not detectable but there was a significant decrease in Ki67 staining. This is consistent with our in vitro findings that leflunomide primarily inhibits cell proliferation through the promotion of G1 cell cycle arrest.

Conclusion: Our findings indicate that leflunomide may be a viable reagent for the treatment of LKB1-mutated lung adenocarcinoma.

Citation Format: Rui Jin, Xiuju Liu, Wei Peng, Yijian Fan, Boxuan Liu, Shuanying Yang, Chun-gen Xing, Melissa Gilbert-Ross, Adam Marcus, Wei Zhou. Leflunomide inhibits the growth of LKB1-inactivated tumors in both xenograft and immunocompetent genetically engineered 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 4838.
Abstract 4841: Rapid screening of cyclotide-based libraries for the selection of potent E3 ligase antagonists
Authors: Camarero Julio A.
Published: August 2019

We report novel methods for the biosynthesis of natively-folded MCoTI-based cyclotides inside live E. coli cells using a split protein splicing unit. The cyclotide MCoTI-cylotides are potent trypsin inhibitors recently isolated from the seeds of Momordica cochinchinensis, a plant member of cucurbitaceae family. Biosynthesis of genetically encoded cyclotide-based libraries opens the possibility of using single cells as microfactories where the biosynthesis and screening of a particular inhibitor can take place in a single process within the same cellular cytoplasm. The cyclotide scaffold has a tremendous potential for the development of therapeutic leads based on their extraordinary stability and potential for grafting applications. We will show an example, where a large cyclotide-based genetically-encoded library was used to screen for low nanomolar antagonists for the Hdm2-HdmX RING-mediated E3 ligase activity. We will also present different strategies to improve the cellular uptake and pharmacokinetic profiles of bioactive cyclotides.

Citation Format: Julio A. Camarero. Rapid screening of cyclotide-based libraries for the selection of potent E3 ligase antagonists [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 4841.
Abstract 4847: Investigating the impact of astrocytes on targeted therapy of melanoma brain metastases
Authors: Parris Joshua , Basu Subhasree , Barnoud Thibaut , Zhao Yongkang , Somboonsong Peter , Chen Qing , Murphy Maureen
Published: August 2019

Metastatic melanoma is a devastating disease that leads to the development of brain metastases (MBMs) in 40-60% of patients. Upon diagnosis of brain metastases, patient overall survival is less than 6 months. Poor patient survival is believed to be due to resistance driven by the brain microenvironment. Collectively, these findings underscore the need for novel therapeutic strategies to improve the survival of patients with MBMs. Development of effective therapies relies on an understanding of the interaction between MBMs and different cell types within the brain microenvironment. Astrocytes are the most abundant cell type within the brain and interact with MBMs through direct and indirect mechanisms. Astrocyte interaction with MBMs alters MBM gene expression and promotes metastatic outgrowth and resistance to cytotoxic chemotherapies. Exactly how astrocytes alter MBM response to MAPK inhibition remains unclear. Using conditioned media from primary astrocytes, as well as co-culture of primary astrocytes with either the MBM cell line WM4265-2 or the primary melanoma cell line WM983B, we found that the interaction of melanoma cells with astrocytes increased melanoma sensitivity to MEK inhibition (MEKi). RNA sequencing of WM4265-2 and WM983B cell lines treated with the MEK inhibitor PD0325901 (PD901) showed significant up-regulation of the transcriptional regulator ID3, which is known to play a role in the resistance of melanoma to MAPK inhibition. Interestingly, astrocyte conditioned media prevented the upregulation of ID3. To validate this finding, we silenced ID3 in WM4265-2 and WM983B cell lines, and showed that knockdown of ID3 caused increased sensitivity to MEKi. These combined data support the premise that the increased response to PD901 in MBMs exposed to astrocytes is due to ID3 down-regulation. How ID3 modulates resistance to MAPK inhibition remains unclear. Elucidating the mechanism of ID3 mediated resistance to MEKi may uncover new avenues for improving the treatment of melanoma brain metastases.

Citation Format: Joshua Parris, Subhasree Basu, Thibaut Barnoud, Yongkang Zhao, Peter Somboonsong, Qing Chen, Maureen Murphy. Investigating the impact of astrocytes on targeted therapy of melanoma brain metastases [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 4847.