bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract A003: Anti-angiogenic activity of the CXCR4 antagonist balixafortide
Authors: Zimmermann Johann , Obrecht Daniel , Remus Tobias
Published: March 2020

Background: Balixafortide is a potent and selective antagonist of the chemokine receptor CXCR4, and is in PhIII for metastatic HER2-negative breast cancer in combination with eribulin (NCT03786094). Clinical proof-of concept of the combination with comparable tolerability to that of eribulin monotherapy was demonstrated in a recent PhI single arm dose-escalation trial (NCT01837095). Balixafortide can inhibit tumor growth through several mechanisms including chemosensitization in combination with chemotherapy (e.g. eribulin), suppression of metastasis, and activation of immune cell response in the tumor microenvironment. In addition, data regarding the co-regulation of the angiogenic factor VEGF and CXCR4 in colorectal cancer (CRC) tissue suggest a possible anti-angiogenic activity of Balixafortide. Methods: VEGF alpha-induced migration of endothelial cells and vascular permeabilization were assessed in vitro. Changes in VEGF alpha levels were determined in CXF260 CRC PDX tumor samples by Nanostring and in CXF260 cells in vitro by qPCR. Results: Balixafortide potently inhibited VEGF alpha-induced migration and permeabilization of an endothelial monolayer in vitro and reduced VEGF alpha levels in both, cultured CXF260 tumor cells in vitro and CXF260 tumors in vivo. Conclusions: Balixafortide modulates angiogenic mechanisms in vitro and in vivo and suggests further investigation of balixafortide in anti-angiogenic therapies.

Citation Format: Johann Zimmermann, Daniel Obrecht, Tobias Remus. Anti-angiogenic activity of the CXCR4 antagonist balixafortide [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference on Molecular Targets and Cancer Therapeutics; 2019 Oct 26-30; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2019;18(12 Suppl):Abstract nr A003. doi:10.1158/1535-7163.TARG-19-A003
Abstract A006: Preclinical models from rare aggressive tumors: Establishment, drug sensitivity and genomic analyses of mesothelioma PDX as models for oncology drug development
Authors: Becker Michael , Brzezicha Bernadette , Rolff Jana , Conrad Theresia , Rau Beate , Walther Wolfgang , Hoffmann Jens
Published: March 2020

Malignant mesothelioma is a rare but aggressive disease with few therapeutic options and a poor prognosis. The main cause of the disease is the exposition to asbestos, even after 50 years of latency. Intended therapeutic goals, such as increase of patient survival and improvement of quality of life have only been achieved to a limited extent. Curative treatment is currently still not possible. Therefore, there is a strong clinical need for translational models. For this, we have characterized patient-derived xenograft models (PDX) of mesothelioma to evaluate their suitability for preclinical research. For PDX establishment, tumor tissue from surgically resected pleura or peritoneum mesotheliomas, was implanted subcutaneously into immunodeficient mice and serially passaged for at least 3 in vivo-passages. Established PDX models were characterized towards sensitivity to standard of care drugs. PDX tissue was collected for RNA sequencing and immunohistochemistry. The generated molecular profiles (comparative gene expression and sequence variations) of the models were analyzed comprehensively. A panel of 6 mesothelioma PDX has been established. In these PDX models, characteristic histological structure of the original patient tumor was preserved. The PDX models responded individually to standard of care drugs like pemetrexed, cisplatin, and gemcitabine as well as combinations thereof. The combination of pemetrexed or gemcitabine with cisplatin showed a stronger tumor growth inhibition than the respective monotherapies. The analyzed mutational and gene expression landscape resembles the known pattern of mesotheliomas. In summary, our established mesothelioma PDX will serve as a preclinical tool to investigate cellular and molecular mechanisms of drug sensitivity and resistance. They will provide a valuable preclinical model system to identify and validate biomarkers and to develop new therapeutic approaches.

Citation Format: Michael Becker, Bernadette Brzezicha, Jana Rolff, Theresia Conrad, Beate Rau, Wolfgang Walther, Jens Hoffmann. Preclinical models from rare aggressive tumors: Establishment, drug sensitivity and genomic analyses of mesothelioma PDX as models for oncology drug development [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference on Molecular Targets and Cancer Therapeutics; 2019 Oct 26-30; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2019;18(12 Suppl):Abstract nr A006. doi:10.1158/1535-7163.TARG-19-A006
Abstract A012: Mammary fat pad injections: An alternative implantation method for syngeneic tumor models
Authors: Obodozie Cynthia , Ruf Susanne , Bijelic Gojko , Moor Sandra , Giesen Bianca , Leisegang Ulrike , Dempe Sebastian , Weber Holger
Published: March 2020

Syngeneic tumor models used for discovery of immune therapeutics should have several features such as a long study duration, responsiveness to checkpoint inhibitors, high immune cell infiltration and a high homogeneity in tumor growth. Moreover, models should consider the ethical rules (3R reduce, refine, replace). At present, the standard implantation method for syngeneic tumor models is subcutaneous tumor cell inoculation. We have developed an alternative implantation method for syngeneic tumor models: inoculation into the mammary fat pad. Both implantation sides are heterotopic related to the original tumor entity except for syngeneic breast tumor cells. In addition, both tumor inoculation methods can easily be applied and monitored by calipering reducing the costs. We compared the two implantation methods with models MC38-CEA, Ct26wt, Hepa1-6, RENCA, LL-2, AB12, CloneM3, B16.F10, 4T1 and EMT-6 tumor cells in respect to growth characteristics and immune response. Intra-mammary tumor growth showed more homogeneity with higher final tumor volumes compared to the subcutaneous tumor growth. Moreover, in all tested syngeneic models, tumor ulceration was prevented by almost 100% when injecting the tumor cells into the mammary fat pad. In contrast, animals of the subcutaneous tumors were mainly euthanized due to tumor ulceration. Both findings favor the mammary fat pad injection with regard to the 3R rules by strongly reducing tumor ulceration (refinement) and animal number due to a more homogenous growth (reduction). In addition, the immune checkpoint inhibitor treatment was tested and found comparable between the intra-mammary and subcutaneous models. The presence of immune cell populations was investigated in the Ct26wt colon tumor model time-dependently by flow cytometry using a 17 marker-staining panel . The number of isolated cells per gram tumor mass was more than doubled in the intra-mammary tumors. In conclusion, tumor models of the heterotopic intra-mammary implantation side are found to be superior compared to the traditional subcutaneous tumor models: a higher tumor homogeneity with no tumor ulceration, combined with a high number of immune cells and connective stroma tissue, making the mammary fat pad implantation of syngeneic tumor cells the implantation side of choice.

Citation Format: Cynthia Obodozie, Susanne Ruf, Gojko Bijelic, Sandra Moor, Bianca Giesen, Ulrike Leisegang, Sebastian Dempe, Holger Weber. Mammary fat pad injections: An alternative implantation method for syngeneic tumor models [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference on Molecular Targets and Cancer Therapeutics; 2019 Oct 26-30; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2019;18(12 Suppl):Abstract nr A012. doi:10.1158/1535-7163.TARG-19-A012
Abstract A027: Combination of TROP2 and SLFN11 as the sensitivity biomarkers for DS-1062a, a novel TROP2-targeting antibody-drug conjugate with a novel DNA topoisomerase I inhibitor DXd
Authors: Yasuda Satoru , Okajima Daisuke , Toki Tadashi , Yamaguchi Junko , Karibe Tsuyoshi , Fujitani Tomomichi , Nakada Takashi , Noguchi Yutaka , Murakami Masato
Published: March 2020

Background: DS-1062a is a TROP2-targeting antibody-drug conjugate with Daiichi-Sankyo DXd technology. DS-1062a is expected to deliver DXd, a derivative of Exatecan, to TROP2 highly expressed tumors such as epithelial cancers with stronger anti-tumor effect and safer profile than the systemic chemotherapy. SLFN11 has been reported as a sensitivity biomarkers of DNA topoisomerase I inhibitors in several cancers. In this research, we elucidated the potency of combination of TROP2 and SLFN11 as a sensitive biomarkers of DS-1062a sensitivity using in vitro and in vivo preclinical models. Materials and Methods: Growth inhibition effect of DS-1062a and DXd were tested using 39 cell lines of various cancer types and correlation between DS-1062a efficacy and both TROP2 and SLFN11 mRNA level was elucidated. To confirm the contribution of SLFN11 expression to DS-1062a sensitivity, SLFN11 was knocked down in 5 of TROP2 and SLFN11 expressed cell lines and changes of growth inhibition effects of DS-1062a and DXd were evaluated. DS-1062a efficacy were also evaluated in 25 PDX (Patient-derived xenograft) models which express various levels of TROP2 and SLFN11 and relationship of sensitivity of each tumor to DS-1062a and TROP2 and SLFN11 mRNA levels was analyzed. Results: In vitro tumor cell growth inhibition effect of DS-1062a was observed in the most of TROP2 highly expressed cell lines. However, DS-1062a showed weak efficacy in some cell lines with high TROP2 expression, which have relatively low sensitivity to DXd. Correlation analysis between DXd efficacy and gene expression suggested cell lines with low sensitivity tend to express limited level of SLFN11. Knock down of SLFN11 weakened in vitro tumor cell growth inhibition effect of both DS-1062a and DXd in cell lines with high expression of SLFN11 level. Furthermore, in vivo study with 25 PDX models, strong drug efficacy of DS-1062a was frequently observed in both TROP2 and SLFN11 expressed tumor. Conclusion: We revealed that SLFN11 is one of the sensitivity biomarkers other than TROP2 expression levels against DS-1062a. This finding suggests combination of TROP2 and SLFN11 could be a promising biomarker to predict antitumor effect of DS-1062a.

Citation Format: Satoru Yasuda, Daisuke Okajima, Tadashi Toki, Junko Yamaguchi, Tsuyoshi Karibe, Tomomichi Fujitani, Takashi Nakada, Yutaka Noguchi, Masato Murakami. Combination of TROP2 and SLFN11 as the sensitivity biomarkers for DS-1062a, a novel TROP2-targeting antibody-drug conjugate with a novel DNA topoisomerase I inhibitor DXd [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference on Molecular Targets and Cancer Therapeutics; 2019 Oct 26-30; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2019;18(12 Suppl):Abstract nr A027. doi:10.1158/1535-7163.TARG-19-A027
Abstract A038: Assessment of angiogenesis markers in advanced cervical carcinoma patients that received chemoradiation therapy plus/minus neoadjuvant chemotherapy
Authors: ARAI ROBERTO JUN , COSTA THIAGO RODRIGUES DA , FERREIRA SILVANEIDE , SICHERO LAURA , DIZ MARIA PILAR ESTEVEZ
Published: March 2020

The expression profile of angiogenesis associated proteins was assessed in patients with locally advanced cervical cancer included in a prospective, randomized phase II study. The majority of patients had squamous cell carcinoma (87.8%). Group A individuals were treated with neoadjuvant chemotherapy (cisplatin and gemcitabine) (NAC), followed by chemoradiation with cisplatin (CRT). Group B patients received CRT. Serum samples from patients in group A were collected at 03 different time points (at baseline (n=34), after NAC (n=24), and after CRT (n=27)). Samples of group B were collected in 02 different time points (at baseline (n=40) and after CRT (n=34)). Samples were grouped and evaluated with a non-parametric Mann-Whitney test where we considered the points as independent groups. The Friedman test (ANOVA) was used for analysis of group A (with three evaluation points). The levels of different proteins associated with angiogenesis (EGF, angiopoietin, G-CSF, endothelin-1, FGF-1, follistatin, IL-8, HGF, HB-EGF, PLGF, VEGF- D, VEGF-A and FGF-2) were assessed by a Multiplex Assay using the Luminex® technology. HB-EGF, IL-8 and VEGF-C were significantly decreased in patients from group A, whereas all others biomarkers did not present intra or intergroup differences. NAC was associated with an inferior PFS (progression free survival), with 3-year PFS rates of 40.9% compared with 60.4% in the CRT arm (HR, 1.84; 95% CI, 1.04 – 3.26; P = .033). NAC was also associated with a lower OS (overall survival) (3-year OS, 60.7% vs 86.8%; HR, 2.79; 95% CI, 1.29-6.01; P=0.006). After treatment completion, complete response rates were 56.3% in the NAC arm and 80.3% in the CRT arm (P=0.008). Toxicities were similar in both arms, with the exception of hypomagnesemia and neuropathy that were more commonly observed within NAC individual. We can conclude, within the limitations of presented study, that the NAC in group A results in significant alterations in specific markers associated with angiogenesis.

Citation Format: ROBERTO JUN ARAI, THIAGO RODRIGUES DA COSTA, SILVANEIDE FERREIRA, LAURA SICHERO, MARIA PILAR ESTEVEZ DIZ. Assessment of angiogenesis markers in advanced cervical carcinoma patients that received chemoradiation therapy plus/minus neoadjuvant chemotherapy [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference on Molecular Targets and Cancer Therapeutics; 2019 Oct 26-30; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2019;18(12 Suppl):Abstract nr A038. doi:10.1158/1535-7163.TARG-19-A038
Abstract A090: A novel THO complex member LENG8 connects mRNP export, genome integrity and tumorigenesis
Authors: Zhang Nan , Zhao Yongxu , Chen Jianxia , Ding Qiurong , Liu Feng
Published: March 2020

THO/TREX complex is evolutionarily conserved from yeast to human and required for both of mRNP export and genome integrity, but the underlying mechanism remains unknown. Here we demonstrate that DNA damage represses mRNA biogenesis and export, and recruits mRNP to the DNA damage foci. LENG8, an evolutionarily conserved nuclear protein, is a novel member of THO/TREX complex and connects the mRNP machinery with poly(ADP-ribosyl)transferase PARP1. The expression of THOC1 and LENG8 are lower in various human tumors including gastric tumors as well as lung cancers, and their abundancy correlates with poor prognosis of patients. We show that knockdown of THOCs or LENG8 promotes proteolytic cleavage of PARP1, suppresses DNA repair and therefore increases tumor growth both in vitro and in vivo. Our findings suggests THO/TREX complex as a novel regulator of genome integrity and a potential therapeutic target in cancer prevention and treatment.

Citation Format: Nan Zhang, Yongxu Zhao, Jianxia Chen, Qiurong Ding, Feng Liu. A novel THO complex member LENG8 connects mRNP export, genome integrity and tumorigenesis [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference on Molecular Targets and Cancer Therapeutics; 2019 Oct 26-30; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2019;18(12 Suppl):Abstract nr A090. doi:10.1158/1535-7163.TARG-19-A090
Abstract B010: SOX2-mediated 5hmC dysregulation in GBM Stem Cells
Authors: Lopez-Bertoni Hernando
Published: March 2020

Primary brain tumors are among the most devastating forms of cancer and glioblastoma (GBM) represents the most aggressive and lethal form of the disease. We now know that GBM contain small subsets of cells that display tumor-propagating stem-like phenotypes (i.e. glioma stem cells or GSCs) that act as critical determinants of GBM resistance to current treatments and tumor recurrence for which there is no proven therapy. Altered patterns of DNA methylation are widely reported in human GBM. However, substantial knowledge gaps remain in our understanding of the molecular mechanisms responsible for this epigenetic dysregulation, its downstream consequences and role in the GBM tumor propagating phenotype. Understanding and ultimately targeting the epigenetic mechanisms that induce and maintain these tumor-propagating cell subsets is critical to improving GBM therapy and patient outcomes. DNA methylation generally occurs in cytosine-guanine (CpG) sequences and is established by DNMTs, which catalyze the conversion of cytosine to 5-methylcytosine (5mC). DNA methylation is a reversible process and is partially mediated by the ten-eleven translocation (TET) family of enzymes which function as deoxygenases to catalyze the conversion of 5mC to 5-hydroxymethylcytosine (5hmC). Levels of 5hmC closely track with the differentiation state of cells with terminally differentiated cells containing high levels of 5hmC and less differentiated cells having very low levels. Multiple studies found negative correlations between 5hmC levels and glioma grade and loss of 5hmC correlates with poor prognosis of GBM patients, strongly suggesting that these enzymes activate tumor suppressing mechanisms. Our preliminary data shows that exogenous SOX2 represses the TET2 demethylase and decreases 5hmC, the enzymatic product catalyzed by TET proteins, in GSCs. TET2 repression using 2 independent shRNA hairpins efficiently decreases 5hmc levels and significantly enhanced self-renewal capacity and tumor growth capacity of low-passage GSC isolates. We also show that SOX2 induces expression of miR-10b-5p, a known onco-miR predicted to target TET2, and inhibiting this miRNA partially rescues the reduction in 5hmC expression observed in GSCs expressing transgenic SOX2, thus implicating miR-10b-5p as a critical mediator of SOX2-induced onco-methylation, GSC induction and glioma malignancy. Our preliminary results support a mechanism in which SOX2 represses TET2 leading to 5hmC loss by directly activating miR-10b-5p transcription and predicts that targeting this novel mechanism of epigenetic dysregulation by inhibiting miR-10b-5p in vivo will can lead to pre-clinical GBM therapeutics.

Citation Format: Hernando Lopez-Bertoni. SOX2-mediated 5hmC dysregulation in GBM Stem Cells [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference on Molecular Targets and Cancer Therapeutics; 2019 Oct 26-30; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2019;18(12 Suppl):Abstract nr B010. doi:10.1158/1535-7163.TARG-19-B010
Abstract B036: Targeting Hsp90-Cdc37 complex in glioma harboring FGFR3-TACC3
Authors: Li Tao , Mehraein-Ghomi Farideh , Namjoshi Sanjeev V. , Song Qian Qian , Ballard Elizabeth A. , Forbes Mary E. , Chou Ping-Chieh , Yang Xuejun , Zhang Wei
Published: March 2020

Glioblastoma is one of the most lethal central nervous system tumors with the poorest prognosis. Fusion genes are common chromosomal aberrations in malignancies that can be used as prognostic markers as well as drug targets in clinical practice. The FGFR3-TACC3 (F3T3) fusion gene was discovered in glioblastoma, with an occurrence rate of up to 8.3% and leading to uncontrolled proliferation and chromosomal instability. Our previous study has shown that F3T3, generated by a rearrangement of chromosome, led to the loss of the 3′-UTR of FGFR3, blocking epigenetic regulation of miR-99a and enhancing expression of the fusion gene. To further explore whether there is a post-translational regulation on F3T3 protein, we immunoprecipitated (IP) the proteins that interact with F3T3 in U251 glioblastoma cells overexpressing the F3T3 or wild-type FGFR3, followed by LC-MS/MS analyses. Quantitative proteomic analysis suggest a stronger interactions of the F3T3 fusion protein with Hsp90 and Cdc37 proteins when compared with wild-type FGFR3. To test inhibition of F3T3 association with the Hsp90-Cdc37 complex, we treated U251 and LN229 cells constitutively expressing F3T3 with either Hsp90 inhibitors or siRNA of Cdc37. Targeting Hsp90-Cdc37 complex significantly suppressed the activation of F3T3 and downstream signaling pathways. Intriguingly, both Hsp90 and Cdc37 inhibition caused a degradation of glycosylation form of F3T3 protein. F3T3 was found highly expressed in both the untreated and matched recurrence glioblastoma, indicating a resistance to the concurrent radiotherapy and temozolomide (TMZ) treatment. Our results provide evidence that the F3T3 fusion gene confers drug resistance to TMZ induced DNA damage and using Hsp90 inhibitor chemosensitizes F3T3 glioma cells to TMZ. In conlusion, our findings establish that F3T3 is an strong Hsp90 client that shows strong addiction to the Hsp90-Cdc37 complex and suggests a novel strategy for targeting F3T3 fusion gene in glioma.

Citation Format: Tao Li, Farideh Mehraein-Ghomi, Sanjeev V. Namjoshi, Qian Qian Song, Elizabeth A. Ballard, Mary E. Forbes, Ping-Chieh Chou, Xuejun Yang, Wei Zhang. Targeting Hsp90-Cdc37 complex in glioma harboring FGFR3-TACC3 [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference on Molecular Targets and Cancer Therapeutics; 2019 Oct 26-30; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2019;18(12 Suppl):Abstract nr B036. doi:10.1158/1535-7163.TARG-19-B036
Abstract B068: The establishment of a large tumor organoid biobank using a well characterized/annotated patient-derived xenograft (PDX) library to enable drug discovery and translational research
Authors: Xu Xiaoxi , Shang Limei , Wang Lili , Li Chunmei , Liu Yan , Han Peng , Sun Zhongman , Qu Yaping , Zhang Likun , Chen Bonnie Xiaobo , Ouyang Davy Xuesong , Huang Yujun , Li Henry Qixiang
Published: March 2020

Background: Patient derived xenograft (PDX), a cancer stem cell (CSC) derived in vivo model, is an accepted model of choice for preclinical and translational research due to its proven predictive power. Patient derived cancer organoids (PDOs), also CSC-derived 3D culture of carcinoma with defined structures, harbor carcinoma’s multicellular components and mimics cancer lesion structures/heterogeneity, both genomicly and histopathologically. PDO was first described by Hans Clevers Lab and proven to be a predictive model for preclinical research, similar to PDX. Method: We have used the Hubrecht organoid technology (HUB approach to systematically create the worlds-first biobank of organoids derived from a well annotated PDX library (the world’s largest with >2,500, covering a variety of carcinomas, with extensive pathology, genomic and treatment information), referred to as PDXOs. We then systematically profiled these PDXOs by WES (whole exome sequencing)/RNAseq (transcriptome sequencing), histopathology and standard of care (SOC) treatment. Results: At present, we have established > 100 PDXOs covering > 9 cancer types, including bladder, breast, colorectal, gastric, liver, lung, ovarian and pancreatic cancer, cholangiocarcinoma, etc. Histopathological analysis showed cellular/structural similarities (ductal, mucous or carcinoid) between PDXO and original PDX, suggesting that tissue specific structural features were maintained in the 3D organoids. A high throughput screening (HTS in 384 well) format was established using the PDXOs and SOC sensitivity testing was conducted. The preliminary results largely correlated to the SOC response seen in vivo for the corresponding PDXs. Conclusion: In summary, we have successfully established a large biobank of the PDXOs that mirror the original PDXs, creating a unique library of matched in vitro/in vivo models with high translational power and enabling HTS, thus likely become an important tool for the future oncology drug discovery and development tools.

Citation Format: Xiaoxi Xu, Limei Shang, Lili Wang, Chunmei Li, Yan Liu, Peng Han, Zhongman Sun, Yaping Qu, Likun Zhang, Bonnie Xiaobo Chen, Davy Xuesong Ouyang, Yujun Huang, Henry Qixiang Li. The establishment of a large tumor organoid biobank using a well characterized/annotated patient-derived xenograft (PDX) library to enable drug discovery and translational research [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference on Molecular Targets and Cancer Therapeutics; 2019 Oct 26-30; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2019;18(12 Suppl):Abstract nr B068. doi:10.1158/1535-7163.TARG-19-B068
Abstract B078: Inhibition of the MAPK pathways enhances the sensitivity of triple negative breast cancer cells to chemotherapeutic drugs
Authors: Bhatt Akshita B. , Wright Thomas D. , Patel Saloni , Chakrabarty Suravi , Barnes Van , Burow Matthew , Flaherty Patrick T. , Cavanaugh Jane
Published: March 2020

Triple negative breast cancer is characterized by the loss of hormone receptors and lack of targeted therapy. Most invasive cancers, including triple negative (TNBC) breast cancer have a mesenchymal phenotype, which is associated with increased chemoresistance. Activation of the MEK1/2 and MEK5 pathways plays a crucial role in the activation of the epithelial to mesenchymal transition program and increases the survival, proliferation, and migration of the cancer cells. Disruption of actin skeleton via ras and src mediated activation of extracellular regulated kinase 1/2 (ERK1/2) and ERK5 is reported, indicating their role in oncogenic transformation. Moreover, inhibition of either pathway results in a compensatory increase in the PI3K/AKT pathway. These crosstalk mechanisms are involved in mediating therapeutic drug resistance. MDA-MB-231, a BRAF and KRAS mutant TNBC cell line has more than 90% of high CD44+/CD24-/low stem cell population, and high ERK5 and ERK1/2 expression; hence it was used as the model for our experiments. Moreover, MDA-MB-231-ERK5-KO cells were utilized to confirm the mechanism of action. To target TNBC, known inhibitors of the MEK1/2 pathway: trametinib, an FDA approved drug for BRAF mutant melanoma, VX-11-e, an ERK2 inhibitor, and XMD-8-92, an ERK5 inhibitor were used in combination with the chemotherapeutic drugs paclitaxel and doxorubicin to examine cell viability. We have shown that dual inhibition of the ERK5 and AKT signaling pathways synergistically reduces TNBC cell viability and enhances sensitivity of the cells to paclitaxel. A series of novel quinazoline derivatives was generated to dually target the ERK5 and the AKT pathway. The effect of novel quinazolines on cell viability in combination with the chemotherapeutic agent paclitaxel was examined in the TNBC cells and some encouraging results were obtained. The overall significance of this research is to enhance the anti-cancer activity of chemotherapeutic agents and reduce off-target toxicity by dose-reduction strategy.

Citation Format: Akshita B. Bhatt, Thomas D. Wright, Saloni Patel, Suravi Chakrabarty, Van Barnes, Matthew Burow, Patrick T. Flaherty, Jane Cavanaugh. Inhibition of the MAPK pathways enhances the sensitivity of triple negative breast cancer cells to chemotherapeutic drugs [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference on Molecular Targets and Cancer Therapeutics; 2019 Oct 26-30; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2019;18(12 Suppl):Abstract nr B078. doi:10.1158/1535-7163.TARG-19-B078
Abstract B104: Combined inhibition of DDR1 and CDK4/6 induces synergistic effects in ER-positive, HER2-negative breast cancer with PIK3CA/AKT1 mutations
Authors: Shariati Maryam , Bristow Christopher A , Carugo Alessandro , Heffernan Tim , Zheng Xiaofeng , Peoples Michael D , Tripathy Debu , Meric-Bernstam Funda
Published: March 2020

Molecular alterations in the PI3K/AKT/mTOR pathway occur frequently in estrogen receptor-positive (ER-positive) breast tumors. Patients with ER-positive, human epidermal growth factor receptor 2-negative (HER2-negative) advanced or metastatic breast cancer (MBC) are often treated with palbociclib, an FDA approved CDK4/6 inhibitor, in combination with endocrine therapy. In clinical studies (NCT01942135), the combination of fulvestrant plus palbociclib was associated with improved progression-free survival compared with fulvestrant plus placebo, irrespective of PIK3CA mutational status. The purpose of this study is to identify synthetic lethality partner that can be targeted in combination with palbociclib to improve its therapeutic response. We demonstrate that the in vitro efficacy of CDK4/6 inhibitor is reduced in the presence of PIK3CA/AKT1 mutations using ER-positive isogenic breast cancer cell lines. Utilizing a shRNA library screen targeting cancer related human kinases, we identified that genomic suppression of discoidin domain receptor 1 (DDR1), a tyrosine kinase active in various cancers, is synthetic lethal with palbociclib. Performing sulforhodamine B (SRB) proliferation assay, DDR1 knockdown by shRNA significantly reduced growth of PIK3CA/AKT1 mutant as well as wild type ER-positive cell lines. In addition, DDR1 pharmacological inhibitor, 7rh benzamide, activated the P53/P21 pathway and enhanced the sensitivity of PIK3CA/AKT1 mutant cells to palbociclib. Likewise, we found that combined treatment of palbociclib and 7rh further induced cell cycle arrest through decreasing G2-M phase population and reducing FoxM1 and Rb phosphorylation in mutant cell lines. Our data indicate that DDR1 inhibition can augment cell cycle suppressive effect of palbociclib and could be an effective rational strategy approach for targeted therapy of ER-positive, HER2-negative breast cancer harboring PIK3CA/AKT1 mutations.

Citation Format: Maryam Shariati, Christopher A Bristow, Alessandro Carugo, Tim Heffernan, Xiaofeng Zheng, Michael D Peoples, Debu Tripathy, Funda Meric-Bernstam. Combined inhibition of DDR1 and CDK4/6 induces synergistic effects in ER-positive, HER2-negative breast cancer with PIK3CA/AKT1 mutations [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference on Molecular Targets and Cancer Therapeutics; 2019 Oct 26-30; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2019;18(12 Suppl):Abstract nr B104. doi:10.1158/1535-7163.TARG-19-B104
Abstract B115: Mitotic defects induced by MAT2A inhibitors guides translational drug combination strategies with AG-270 and taxanes
Authors: Kalev Peter , Hyer Marc L , Fletcher Mark , Zhang Peili , Aguado-Fraile Elia , Mandley Everton , Tuncay Yesim , Konteatis Zenon , Travins Jeremy , Marks Kevin M , Marjon Katya
Published: March 2020

Methylthioadenosine phosphorylase (MTAP) is the enzyme in the methionine salvage pathway that metabolizes the byproduct of polyamine biosynthesis, 5′-methylthioadenosine (MTA). The MTAP gene is deleted in approximately 15% of all human cancers and results in accumulation of MTA that partially inhibits protein arginine methyl transferase 5 (PRMT5) function. Recent studies have demonstrated that loss of the MTAP gene sensitizes cancer cells to genetic depletion of PRMT5 and the upstream metabolic enzyme methionine adenosyltransferase 2 alpha (MAT2A). We have developed a first-in-class, highly potent, orally bioavailable MAT2A inhibitor, AG-270, which is currently under investigation in a phase 1 clinical trial (ClinicalTrials.gov NCT03435250). AG-270 selectively inhibits the growth of HCT116 MTAP-/- cells compared to HCT116 MTAP+/+ cells in vitro and in vivo. We utilized RNA sequencing and proteomic approaches to study the underlying mechanism of action for AG-270. The results of these analyses demonstrated that AG-270 treatment induces substantial splicing changes in an MTAP selective manner. The majority of these abnormal splicing events were detained introns, which included genes involved in cell cycle regulation and DNA damage response. Moreover, AG-270 treatment resulted in accumulation of DNA damage and increased mitotic defects in HCT116 MTAP-/- cells. Furthermore, we demonstrated substantially advantageous combination benefits between AG-270 and anti-mitotic taxanes in in vitro cell lines and in in vivo patient-derived xenograft models of non-small cell lung, pancreatic and esophageal cancers. These findings uncover relevant combination strategies targeting MTAP-deleted malignancies, and provide preclinical mechanistic proof-of-concept to explore such combinations in the clinical setting.

Citation Format: Peter Kalev, Marc L Hyer, Mark Fletcher, Peili Zhang, Elia Aguado-Fraile, Everton Mandley, Yesim Tuncay, Zenon Konteatis, Jeremy Travins, Kevin M Marks, Katya Marjon. Mitotic defects induced by MAT2A inhibitors guides translational drug combination strategies with AG-270 and taxanes [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference on Molecular Targets and Cancer Therapeutics; 2019 Oct 26-30; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2019;18(12 Suppl):Abstract nr B115. doi:10.1158/1535-7163.TARG-19-B115
Abstract B134: IMPDH1 and IMPDH2 as therapeutic targets in acute lymphoblastic leukemia
Authors: Dieck Chelsea , Oshima Koichi , Ambesi-Impiombato Alberto , Ferrando Adolfo
Published: March 2020

Acute lymphoblastic leukemia (ALL) tumors are dependent on purine biosynthesis for proliferation and survival and relapsed ALL is particularly associated with transcriptional up-regulation of this pathway. Increased dependency on purine biosynthesis highlights IMPDH (inosine monophosphate dehydrogenase), the rate-limiting enzyme in guanosine production, as a potential therapeutic target for the treatment of ALL. To assess the role of IMPDH1 and IMPDH2 in ALL development and progression, we generated NOTCH1-induced leukemias from Impdh1 knockout and Impdh2 conditional knockout mice. In addition, we have completed whole exome sequencing and RNAseq profiling of 12 pairs of ALL xenografts corresponding to matched diagnostic and relapsed ALL samples and have performed dose response assays with the IMPDH inhibitor mizoribine. Additionally and towards the identification of synthetic lethal genetic vulnerabilities with IMPDH inhibition in ALL we have performed a genome wide CRISPR screen in REH ALL cells with the IMPDH inhibitor mizoribine. Overall, these studies highlight the therapeutic potential for IMPDH inhibitors for the treatment of ALL.

Citation Format: Chelsea Dieck, Koichi Oshima, Alberto Ambesi-Impiombato, Adolfo Ferrando. IMPDH1 and IMPDH2 as therapeutic targets in acute lymphoblastic leukemia [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference on Molecular Targets and Cancer Therapeutics; 2019 Oct 26-30; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2019;18(12 Suppl):Abstract nr B134. doi:10.1158/1535-7163.TARG-19-B134
Abstract B142: Co-culture of tumor organoids with immune cells for immuno-oncology drug development
Authors: Li Zhongliang , Zhou Jun , Zhang Rui , Wang Fei , Wang Shuzong , Xu Xiaoxi , Huang Yujun , Li Henry
Published: March 2020

Hubrecht organoid technology (HUB) can in vitro preserve and propagate patient-derived epithelial cancer cells whilst still maintaining tumour heterogeneity and genetic features. Co-culture of tumor organoids with peripheral blood lymphocytes or tumor-infiltrating lymphocytes (TIL) can potentially expand tumor-specific T cells that mediate explicit killing of tumor organoids. Recapitulating the tumor microenvironment (TME) by co-culture of tumor organoids with immune cells is highly attractive for immuno-oncology (IO) drug development, and will provide an efficient and refined approach for assessing the immune modulatory and tumor killing effects of IO drug candidates, such as monoclonal antibodies, CAR-T, CAR-NK and small molecules. In this study, we aimed to establish and validate IO assay platform using co-culture methods of organoids with various immune cells. We used the HUB organoid approach to create panels of patient-derived (PDO), patient-derived-xenograft (PDX)-derived organoids (PDXO) and murine tumor-derived tumor organoids, followed by co-culturing with corresponding cytotoxic T cells. We developed tumor organoid killing assays using both flow cytometry-based methods and real-time imaging by IncuCyte as readouts in 96-well format. Tumor-reactivity of T cells to organoids was analysed by T cell proliferation, IFNg ELISA in the culture medium, and intracellular staining of IFNg in T cells. Co-culture of tumor organoids expressing various antigens was also used to evaluate antibody dependent cell mediated cytotoxicity (ADCC) by NK cells, and complement dependent cytotoxicity (CDC) and antibody dependent phagocytosis by macrophages for monoclonal antibody drugs. We have profiled the expression of immune inhibitory molecules on various tumor organoids and assessing the effect of immune checkpoint blockade (e.g. anti-PD-1/PD-L1) in organoid-immune cell co-culture. Our preliminary data demonstrated the feasibility of in vitro patient-derived model system in the field of IO research using tumor organoid co-culture with immune cells, and their application in IO target and drug discovery.

Citation Format: Zhongliang Li, Jun Zhou, Rui Zhang, Fei Wang, Shuzong Wang, Xiaoxi Xu, Yujun Huang, Henry Li. Co-culture of tumor organoids with immune cells for immuno-oncology drug development [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference on Molecular Targets and Cancer Therapeutics; 2019 Oct 26-30; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2019;18(12 Suppl):Abstract nr B142. doi:10.1158/1535-7163.TARG-19-B142
Abstract C002: Phase 1 study of abemaciclib in children with recurrent and refractory solid tumors including malignant brain tumors
Authors: Cash Thomas , Aguilera Dolly , Macy Margaret E , Hoffman Lindsey , Dorris Kathleen , McCracken Courtney , Hanberry Bradley , Castellino Robert , MacDonald Tobey , Wetmore Cynthia
Published: March 2020

Background: Cyclin-dependent kinases 4 and 6 (CDK4/6) control the transition between G1 and S phases of the cell cycle and are dysregulated in a number of pediatric brain and solid tumors. Abemaciclib, a selective and potent small molecule CDK4/6 inhibitor that crosses the blood brain barrier, limits cell proliferation by suppressing phosphorylation of RB, leading to G1 arrest. Methods: We conducted a phase 1 trial of abemaciclib to determine the maximum tolerated dose (MTD), describe toxicities, and pharmacokinetic (PK) parameters of abemaciclib in pediatric patients with recurrent or refractory solid and CNS tumors. A stratum for newly diagnosed patients with diffuse midline glioma continues to enroll. A CDK mutation was not required, and patients with known RB1 mutations were excluded. Abemaciclib was administered orally twice daily on days 1-28 of a 28-day cycle. Four dose levels were planned (100, 130, 170, and 220 mg/m2/dose) and evaluated using a rolling-six design. Patients were enrolled to a dose expansion cohort at the MTD to ensure adequate PK sampling and safety. PK and pharmacodynamic (PD) analyses are ongoing. Results: The median age of 34 eligible patients was 13.8 years (range: 2.9 - 23.2). Seven (21%) patients had a solid tumor [osteosarcoma (N=2), neuroblastoma (N=1), rhabdomyosarcoma (N=1), desmoplastic small round cell tumor (N=1), clear cell sarcoma (N=1), and MRT of kidney (N=1)] and 27 (79%) patients had a malignant brain tumor [high-grade glioma (N=12), DIPG (N=5), ATRT (N=3), medulloblastoma (N=3), ependymoma (N=2), PNET (N=1), and CNS neuroblastoma (N=1)]. Twenty-three patients were evaluable for DLT. There were 2 cycle 1 DLTs, grade 4 thrombocytopenia and grade 3 vomiting and diarrhea, both occurring at dose level 3. The most common treatment-related cycle 1 adverse events were leukopenia (69%), neutropenia (53%), thrombocytopenia (44%), hypoalbuminemia (41%), anemia (38%), hypophosphatemia (34%) and diarrhea (34%). Conclusions: The pediatric MTD of abemaciclib was 130 mg/m2/dose administered orally twice daily on a 28-day cycle. Abemaciclib was well-tolerated with mainly hematologic toxicity. PK and PD will be used to further inform if 130 mg/m2/dose is the pediatric recommended phase 2 dose.

Citation Format: Thomas Cash, Dolly Aguilera, Margaret E Macy, Lindsey Hoffman, Kathleen Dorris, Courtney McCracken, Bradley Hanberry, Robert Castellino, Tobey MacDonald, Cynthia Wetmore. Phase 1 study of abemaciclib in children with recurrent and refractory solid tumors including malignant brain tumors [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference on Molecular Targets and Cancer Therapeutics; 2019 Oct 26-30; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2019;18(12 Suppl):Abstract nr C002. doi:10.1158/1535-7163.TARG-19-C002
Abstract C021: Validation of the interaction between a candidate compound and the intended drug target by a phenotypic rescue approach
Authors: Grandmoursel Laure , Geerts Lieke , Servant Geraldine , Ham Miranda van der , Maier Armin , Aarbiou Jamil , Vlaming Marijn , DeGroot Jeroen , Schuler Julia , Waddell Ian , Zuurmond Anne-Marie
Published: March 2020

New targets for cancer treatment frequently emerge in literature, but the thorough target validation required to consider these targets for a drug discovery program is often lacking. In pharmacological or genetic perturbation studies using complex biological assays, undesired off-target effects cannot be easily distinguished from the intended mode of action at the desired target. This is especially evident in cancer drug development where it is important to discriminate on-target effects on cell viability from off-target effects resulting in non-specific loss of cellular fitness. Neglecting the possibility of being deceived by off-target effects can have tremendous scientific and financial impact on a drug discovery program. Ideally, confidence in a preclinical drug target and a modulating compound is boosted in an early stage by more extensive analysis and validation of the actual drug-target relationship. Rescue of a disease-relevant phenotype by genetic restoration of a target mutation is a gold standard approach in drug discovery by which target validation can be achieved. We aim to follow this approach targeting the BRAF V600E mutation in a number of well described melanoma lines as well as the MAP2K1 Q56P mutation in the non-small cell lung cancer cell line H1437. Target validation for both BRAF and MAP2K1 will be addressed by assessing viability, phenotypic changes and sensitivity to compound modulation upon CRISPR/Cas9 repair of the target mutation or by exogenous re-expression of the wild type variant. Compounds tested will be Vemurafenib for BRAF and Trametinib for MAP2K1. Further investigation into target validity will be done using a physiologically relevant 3D spheroid based co-culture system. Mimicking the tumor microenvironment increases the knowledge about “drug-ability” of a target and sustainability of the target modulation at an early time point in the development process. Such early in-depth validation of the relationship between a compound and the drug target is vital to mitigate the risk of failure at later steps of drug development.

Citation Format: Laure Grandmoursel, Lieke Geerts, Geraldine Servant, Miranda van der Ham, Armin Maier, Jamil Aarbiou, Marijn Vlaming, Jeroen DeGroot, Julia Schuler, Ian Waddell, Anne-Marie Zuurmond. Validation of the interaction between a candidate compound and the intended drug target by a phenotypic rescue approach [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference on Molecular Targets and Cancer Therapeutics; 2019 Oct 26-30; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2019;18(12 Suppl):Abstract nr C021. doi:10.1158/1535-7163.TARG-19-C021
Abstract C042: Inhibition of Dihydroorotate Dehydrogenase (DHODH) by RP7214 attenuates growth and promotes differentiation of AML cell lines
Authors: Viswanadha Srikant , Eleswarapu Satyanarayana , Vakkalanka Swaroop
Published: March 2020

Background: Dihydroorotate dehydrogenase (DHODH), a rate limiting enzyme in pyrimidine synthesis, is overexpressed in certain cancers; blockade of which impedes tumor cell proliferation via cell cycle arrest. RP7214 is a novel and potent DHODH inhibitor that inhibited DHODH activity in enzyme and PHA induced HWB/PBMC proliferation with IC50 and EC50 values of 7.8 & 2.5/0.60 nM respectively. The objective of this study was to evaluate the single-agent activity of RP7214 on AML cell differentiation and growth. Methods: Viability of representative AML cell lines (U937, HL-60, THP-1, and KG-1) following incubation with RP7214 for 72 h as well as rescue in the presence of 300 µM Uridine was determined in an MTT-based assay. For differentiation experiments, THP-1 cells were incubated with RP7214 in complete media for 72 h. RNA was isolated, reverse transcribed to cDNA, and real-time PCR was performed to analyze CD11b gene expression. In addition, cell surface expression of CD11b in THP-1 cells incubated with RP7214 and the effect-reversal in presence of Uridine was evaluated by flow cytometry. Apoptosis and cell cycle were determined following incubation with RP7214 for 72 and 48 h respectively. Results: RP7214 inhibited AML cell proliferation in all the four cell lines tested with GI50 ranging from 2.4 to 7.6 µM. Addition of 300 µM Uridine caused a rightward shift with GI50 >10 µM. RT-PCR analysis indicated an 80-fold increase in mRNA expression of the differentiation marker CD11b in THP-1 cells treated with 3 µM RP7214. Similarly, expression of the CD11b marker on THP-1 cell surface was enhanced by 40% following incubation with 5 µM compound; effect of which was reduced to 15% in the presence of 300 µM Uridine. RP7214 dose-dependently induced apoptosis (30-50% at 3 µM) and arrested cycle progression in the “S-phase” (40-45% at 3 µM) in U937 and THP-1 cells. Conclusions: Results demonstrate the utility of RP7214 as a potent DHODH inhibitor in AML acting via suppression of cellular pyrimidine pools and promoting differentiation to fully mature cells. The compound is currently being evaluated in IND-enabling tolerability studies with Phase-1 trial in AML expected to commence in 2020.

Citation Format: Srikant Viswanadha, Satyanarayana Eleswarapu, Swaroop Vakkalanka. Inhibition of Dihydroorotate Dehydrogenase (DHODH) by RP7214 attenuates growth and promotes differentiation of AML cell lines [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference on Molecular Targets and Cancer Therapeutics; 2019 Oct 26-30; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2019;18(12 Suppl):Abstract nr C042. doi:10.1158/1535-7163.TARG-19-C042