Breast Cancer PDX Models Research Articles

Abstract LB-022: Real-time intraoperative tumor imaging in a breast cancer PDX model

Abstract Breast cancer is one of the most common types of cancer diagnosis in women of all ages. The primary treatment of breast cancer is surgical excision of the tumor with the margin of normal breast tissue surrounding the tumor. However, current clinical goals include complete resection of the tumor at the initial surgery, there is often a need for further surgery and removal of more tissue due to the difficulties inherent in getting negative margins. Therefore, there is a clinical need to optimize the excision by precisely identifying tumor margins significantly benefitting patients with breast cancer. Image-guided surgery with fluorescent agents provides surgeons numerous advantages such as real-time detection with a high-resolution image and relatively flexible instrument. We have developed a unique tumor-targeting fluorescence imaging agent. We identified a cell-penetrating peptide (CPP) p28, a fragment of azurin isolated from opportunistic pathogen Pseudomonas pathogen. p28 was chemically conjugated with Indocyanine green (ICG), a near-infrared red (NIR) fluorescent agent. ICG has been approved by the FDA for clinical applications with an excellent safety record. When p28 was conjugated to ICG, there was no significant alternation of fluorescence quantum yield (ΦF) of ICG-p28 compared to ICG alone. Triple-negative (ER, PR, HER2) human breast cancer PDX mouse model in NSG mice was used to evaluate the image-guided surgical procedure with ICG-p28. After 24h of ICG-p28 injection i.v. opportunistic 0.5 mg/kg b.w., NIR-fluorescence positive mammary tumors with a 2-mm safe margin were resected under real-time guidance of the PDE imaging unit (Mitaka USA, Hamamatsu Photonics). NIR-fluorescence negative surrounding tissues of at least two different sites (superior and inferior) were also collected separately. To validate the accuracy of tumor margin identification by the image guidance, Alu-based real-time PCR analysis was used for the quantitative detection of human cancer cells. Five ng of genomic DNA obtained from tumors and surrounding tissues were subjected to RT-PCR. Ct values were normalized by the housekeeping gene, mouse GAPDH. Our results showed that preclinical intraoperative imaging with our imaging agent, ICG-p28 at 0.5 mg/kg, significantly reduced positive margin as compared to control groups (e.g. ICG alone, p<0.01). In conclusion, nontoxic p28 is a potentially ideal CPP that can serve as a tumor-targeting carrier for an intraoperative imaging agent. Results suggest that our imaging approach will potentially provide a significant impact on resection procedure benefitting patients with breast cancer. Citation Format: Masahide Goto, Ingeun Ryoo, Samer Naffouje, Konstantin Christov, Jing Wang, Albert Green, Anne Shilkaitis, Tapas K. Das Gupta, Tohru Yamada. Real-time intraoperative tumor imaging in a breast cancer PDX model [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr LB-022.

Abstract 5059: Breast cancer patient-derived xenograft models for pre- and co-clinical investigation of immune based therapies

Abstract Breast cancer is a highly heterogeneous disease, with a heterogeneous prognosis. Patient-derived xenograft models (PDX) can reflect this heterogeneity and can be used to support the development of novel therapeutic strategies against breast cancer. Preclinical experiments with larger cohorts of breast cancer PDX can be used to model a clinical phase II study with new drugs or drug combinations. The predictive value of these preclinical trials is currently evaluated in co-clinical trials, where treatment effects are compared between patients and their corresponding PDX. We present results from our translational pre- and co-clinical studies in breast cancer immune therapy. As part of the EFRE-POP Project, we established PDX by collecting breast cancer tissue samples from surgery and engrafting them on immunodeficient mice. The tissues were obtained from patients with disease progression after chemotherapy with three to four drugs. PDXs have been successfully engrafted, phenotypically characterized and drug-screened. The PDXs were tested for response to the standard of care therapies. The staining for estrogen/progesterone/androgen/Her2 receptors, Ki-67, and CK5/6 of the original tumor and the PDX were comparable. The patient tumors and the breast cancer PDX models were further analyzed for the expression of PD-L1. We identified four patients that had received PD-1 or PD-L1 antibodies after a sample (used to establish a corresponding PDX) was obtained. To evaluate the response of our preclinical PDX models to immune checkpoint inhibitors, we engrafted mice with a human immune system either by transplanting allogeneic hematopoietic stem cells (HSC) or allogenic PBMC's from healthy donors. Besides, we were able to obtain PBMC's from the patients to set up an autologous model. The patient-derived tumors were transplanted to these humanized mice and after successful engraftment treated with the PD-1 or PD-L1 antibodies corresponding with the clinical treatment of the patient. In summary, humanized PDX of refractory tumors have been established and the first pre-clinical evaluation of response to immune therapy provides evidence that xenografts respond in a qualitatively similar manner as the patient. This suggests a translational relevance of the described pre-clinical models. Citation Format: Annika Wulf-Goldenberg, Verena Kiver, Maria Stecklum, Bernadette Brzezicha, Philipp Jurmeister, Caroline Schweiger, Jens-Uwe Blohmer, Ulrich Keilholz, Jens Hoffmann. Breast cancer patient-derived xenograft models for pre- and co-clinical investigation of immune based therapies [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 5059.

Abstract 626: Mechanistic differentiation of targeted DDR agent combinations

Abstract The DNA damage response (DDR) safeguards genome stability and promotes cellular survival by counteracting deleterious consequences of DNA damage. Deficiencies in DDR pathways contribute to genome instability, a hallmark of cancer development, and targeting the DDR is a promising anti-cancer therapy strategy. We describe here in vitro and in vivo studies allowing mechanistic understanding of the benefit of combining our DDR kinase inhibitors (DDRi) targeting ATM (AZD0156), ATR (AZD6738) and DNA-PK (AZD7648) with the PARP inhibitor, olaparib, in homologous recombination-deficient backgrounds. Using the break-induced replication (BIR) assay, which measures homologous-recombination repair (HRR) of DNA double-strand breaks resembling broken replication forks, we observed that ATRi decreases this type of homologous recombination repair by up to 50%, while inhibiting DNA-PK or ATM showed no effect. This provides a mechanistic rationale for combining ATRi with compounds that cause replication-fork collapse, such as PARPi. Indeed, combination of olaparib with the ATRi in BRCA1 mutant UWB1.289 cells enhanced olaparib sensitivity a thousand-fold (GI50 from 590 nM to 0.6 nM). This correlates with the complete tumour growth inhibitions (TGI) observed for the ATRi + olaparib combination in the BRCA2 mutant HBCx-10 triple negative breast cancer PDX model and is consistent with a synergistic increase of genome instability measured by metaphase spread analysis (total aberrations: olaparib; 11, ATRi; 8, combination; 35). DNA-PKi, however, did not synergise with olaparib, showing little enhancement of olaparib single-agent treatment (GI50 from 0.59 μM to 0.33 μM) and no significant impact on the total amount of chromosomal aberrations (total aberrations: olaparib; 11, DNA-PKi ; 3, combination; 14), suggesting that DNA-PKi alone or in combination with olaparib have little impact on BRCA1 mutated tumours. In ATM KO FaDu xenografts, ATRi or DNA-PKi single-agent treatment resulted in TGI, with complete tumour regressions when combined. Focusing on the olaparib combination, metaphase spread analysis of ATRi-treated FaDu ATM KO cells showed an enhancement of DNA replication-dependent chromatid breaks (1.3 vs 0.2 breaks/spread in ATM WT), while DNA-PKi treatment predominantly caused chromosome breaks (1.7 vs 0.2 breaks/spread in ATM WT) that are not dependent on replication. When combined with olaparib, ATRi acted synergistically and in a manner similar to that observed in the BRCA1 mutant cells, while the DNA-PKi + olaparib combination was additive. These data suggest that enhanced olaparib sensitisation of ATM KO cells by ATRi or DNA-PKi combinations occurs through different mechanisms. Together, these findings provide mechanistic differentiation of our DDRi in specific genetic backgrounds. This work informs how to clinically position these agents to benefit the right patients not only as single-agent treatments, but also in combination. Citation Format: Josep V. Forment, Paul Wijnhoven, Antonio Ramos-Montoya, Jacqueline Fok, Valeria Follia, Mercedes Vazquez-Chantada, Rajesh Oderda, Zena Wilson, Alan Lau, Elisabetta Leo, Stephen Durant, Elaine Cadogan, Mark J. O'Connor. Mechanistic differentiation of targeted DDR agent combinations [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 626.

Abstract 5281: Development of a panel of breast cancer patient-derived xenograft models (PDX) with estrogen independence and/or acquired resistance to endocrine treatment

Abstract Eighty five percent of preclinical agents entering oncology clinical trials fail to demonstrate sufficient safety or efficacy to gain regulatory approval (1). This failure rate shows a weak understanding of the complexity of human cancer, the continued limitations of the predictive value of existing preclinical models and the scale at which cancer models are interrogated in the preclinical setting (2). Consequently, there is a need of new experimental models that better replicate the diversity of human tumor biology in a preclinical setting. It is now evidenced that PDXs models recapitulate human tumor biology and predict patient drug response (3) by directly comparing drug responses in patients and their corresponding xenografts. To extend such observations to a greater number of human cancers, we have generated in collaboration with EISAI an extensive collection of breast cancer PDXs. Similar to what is observed in the clinic, 60% of our breast PDX panel is classified as estrogen receptor (ER) positive. Tested compounds which interfere with the ER pathway as anti-estrogens (fulvestrant) or estrogen deprivation (without estrogen supplementation, or ovariectomy), are able to decrease the tumor growth in PDX-bearing mice. However, in clinic, endocrine treatment efficacy is limited by intrinsic and acquired resistance (4). The main mechanisms of resistance to these therapies are lack of expression of ER, deregulation of ER-associated transcription factors, coactivators, activation of receptor tyrosine kinase signaling, and aberrant expression of cell-cycle regulators (5). In order to reconcile this need for relevant preclinical models and to study acquired resistance we developed a panel of breast cancer PDX models either resistant to fulvestrant or able to grow in the complete absence of estrogen (ovariectomy or without estrogen supplementation). Each generated tumor was then analyzed by IHC for their ER/PR expression. Whole exome mutation and gene expression were also analyzed and compared to the parental tumor. 1: Arrowsmith, J. & Miller, P. Trial watch: phase II and phase III attrition rates 2011-2012. Nat. Rev. Drug Discov. 12, 569 (2013). 2: Paul, S.M. et al. How to improve R&D productivity: the pharmaceutical industry's grand challenge. Nat. Rev. Drug Discov. 9, 203-214 (2010). 3: Hidalgo, M. et al. Patient-derived xenograft models: an emerging platform for translational cancer research. Cancer Discov. 4, 998-1013 (2014). 4. Ring, A. & Dowsett, M. Mechanisms of tamoxifen resistance. Endocr Relat Cancer 11:643-58 (2004). 5. Hanny, A. & Abdel-Hafiz. Epigenetic mechanisms of tamoxifen resistance in luminal breast cancer. Diseases Jul 6;5(3) (2017). Citation Format: Marc Hillairet de Boisferon, Ismahene Benzaid, Elodie Marie Dit Chatel, Nicolas Hoffmann, Bruce Littlefield. Development of a panel of breast cancer patient-derived xenograft models (PDX) with estrogen independence and/or acquired resistance to endocrine treatment [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 5281.

Abstract A008: PDX-based screen to evaluate and compare approved CDK4/6 inhibitors in breast cancer and determine palbociclib efficacy in additional tumor types

Abstract Background: FDA-approved CDK 4/6 inhibitors (CDK4/6i) including palbociclib, abemaciclib, and ribociclib have demonstrated clinical benefit in treating hormone receptor-positive (HR+) breast cancer. However, whether there is differential sensitivity to each therapy in vivo is unknown. In addition, whether these inhibitors demonstrate activity towards other tumor types is unclear. To better understand potential differences in CDK4/6i activity, we compared activity of each in a panel of 100 HR+/- breast cancer PDX models. To identify additional tumor types sensitive to CDK4/6i, we screened single agent palbociclib in an additional panel of 350 PDX models representing solid and hematologic malignancies. FFPE samples were collected from control and treatment groups in all studies at endpoint, and %T/C values in each model were calculated and in breast studies compared. Methods: START PDX models were established and validated as previously described and further characterized using IHC, WES, RNAseq, and drug sensitivity studies. For each CDK4/6i study, models were implanted SC in immune-deficient mice and studies initiated at 200-300 mm3 (n=1-3/grp). CDK4/6i agents were administered orally once daily at 50-75 mg/kg for a minimum of twenty-eight days or until study completion. Models tested with palbociclib included breast, lung, ovary, pancreas, head/neck, renal, gastric, uterine, colorectal, melanoma, and various hematologic malignancies. Study endpoints included tumor volume and time from treatment initiation with %T/C values and tumor regression reported at study completion; a %T/C of ≤ 20% versus control was considered sensitive. Partial tumor regression (PR) (%PR=1-99%) and complete tumor regression (CR) (%CR=100%) values versus Day 0 tumor volume of treated groups were also reported. Results: HR+/- breast models demonstrated variable sensitivity to the three CDK4/6i with HR+ models established from chemo-naïve patients reporting greatest tumor growth inhibition (TGI). Two of four HR+ ESR1 mutant models reported modest sensitivity to CDK4/6i (ST941Y537S and ST2535D538G), while ST2177Y537S was sensitive and ST2056Y537S refractory to the three therapies. Evaluation of palbociclib in additional indications reported notable TGI versus control in several indications with the following percentage of tested models reporting sensitivity: Lung: 10%, Ovary: 15%, Pancreas: 30%, Head/Neck: 35%, Renal: 45%, Gastric: 60%, Uterine: 20%, Colorectal: 45%, Melanoma: 40% and Hematologic: 15%. In addition, several indications reported tumor regressions including one CR (lung: ST1748) and partial responses in ovary, pancreas, head/neck, renal, colorectal, and melanoma. Conclusion: We evaluated and compared three FDA-approved CDK4/6i therapies in a panel of HR+/- breast PDX models and identified models with differential responses. In addition, we screened single agent palbociclib in a panel of 350 PDX models representing solid and hematologic malignancies and identified several sensitive models in multiple indications, including some with partial or complete tumor regressions. Citation Format: Lizette Gamez, Tomoyuki Mashimo, Alyssa Moriarty, Kyriakos Papadopoulos, Drew Rasco, Amita Patnaik, Amy Lang, Ronald Drengler, Lon Smith, Michael J Wick. PDX-based screen to evaluate and compare approved CDK4/6 inhibitors in breast cancer and determine palbociclib efficacy in additional tumor types [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 A008. doi:10.1158/1535-7163.TARG-19-A008

Abstract 4815: Novel Globo H targeting antibody-drug conjugate with binding specificity and anti-tumor efficacy in multiple cancer types

Abstract Background: Globo H, a hexasaccharide, has been reported to be highly expressed in multiple cancers types, but not express or, if at all, express to a lesser extent in normal tissue. Therefore, Globo H may be a potential target for cancer immunotherapy. OBI-999 is an antibody-drug conjugate (ADC) which consists of a Globo H-specific monoclonal antibody OBI-888, conjugated with monomethyl auristatin E (MMAE), a synthetic antineoplastic agent. The present study investigates antigen specificity, drug internalization, anti-tumor efficacy, and pharmacokinetics profiles of OBI-999. Methods: Globo H expression was surveyed in various human cancer cell lines. The binding specificity and internalization of OBI-999 were determined by flow cytometry and confocal microscopy, respectively. In vivo anti-tumor efficacy was studied in conventional xenograft and patient-derived xenograft models. Pharmacokinetic parameters were evaluated in normal and tumor bearing xenograft mice. Results: We confirmed positive Globo H expression on the cell surface of multiple cancer cell lines ranging from breast, gastric, lung, to pancreatic cancer. From flow cytometry, OBI-999 showed binding selectivity to the aforementioned Globo H expressing cell lines. In contrast, OBI-999 did not bind to non-Globo H expressing cell lines. When bound to the antigen, OBI-999 was internalized and trafficked to endosome and lysosome within 2.5 to 5 hours, suggesting that the drug payload, MMAE, was cleaved in a lysosome dependent manner. OBI-999 showed significant tumor inhibition in breast, gastric, lung, and pancreatic cancer xenograft and PDX models in dose-dependent manner. At 1 mg/kg of OBI-999, tumor growth inhibition in breast, gastric, and lung cancer model were 77%, 89%, and 68% respectively. At 10 mg/kg of OBI-999, complete tumor growth inhibition in pancreatic cancer model was observed. In vitro serum stability studies revealed that OBI-999 has comparable stabilities to Adcetris® in mouse, rat, monkey, and human sera. Tissue distribution study revealed that OBI-999 is accumulated gradually at the tumor site while the level of OBI-999 showed a time dependent decrease in blood-rich organs. Accumulation of OBI-999 at the tumor site reached its maximum level at 168 hour post treatment. Level of MMAE in tumor mass was approximately 25 folds higher than that in other organs and 250 folds higher than that in serum, suggesting that OBI-999 targets and releases its payload at tumor region. Conclusion: The preclinical studies of OBI-999 demonstrated its targeting specificity and anti-tumor efficacy in multiple Globo H positive cancer models. OBI-999 also demonstrated a longer retention at tumor site. The preclinical results provided the fundamental basis for OBI-999’s clinical application as targeted cytotoxicity therapy for solid tumors. Citation Format: Ming-Chen Yang, Yu-Jung Chen, Chi-Sheng Shia, Hui-Wen Chang, Wan-Fen Li, Cheng-Der Tony Yu, I-Ju Chen. Novel Globo H targeting antibody-drug conjugate with binding specificity and anti-tumor efficacy in multiple cancer types [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 4815.

Abstract 2163: Establishment and characterization of an endocrine resistant ESR1D538G -mutant breast cancer PDX model

Abstract Background: Chronic endocrine therapy (ET) to treat luminal type breast cancer can result in acquired mutations in the ligand binding domain (LBD) of ERα (ESR1), primarily at residues Y537 and D538, leading to constitutive ER activation and hormone-independent progressive disease. While ESR1-mutated breast cancers are insensitive to endocrine therapy, agents including selective estrogen receptor degrader (SERD) or modulator (SERM) therapies may be effective. However, few preclinical models of ESR1-mutant breast cancer are available for preclinical evaluation. Previously we reported the establishment and characterization of several ER+ breast PDX models and a cell-derived xenograft representing ESR1Y537S-mutated breast cancer. Methods: We report here the establishment and characterization of ST2535 and ST2535/HI, ER+, ESR1D538G-mutated breast PDX models which are maintained in the presence (ST2535) or absence (ST2535/HI) of exogenous estradiol. We characterized these models using genomic analysis and evaluated both in vivo and compared efficacy results with those from ESR1Y537S-mutated models. Studies were performed evaluating model sensitivity to various test agents at standard dose and schedule regimens. Study endpoints included tumor volume and time from treatment initiation with %T/C values and tumor regression reported at study completion. Results: Genomic analysis confirmed the identified ESR1-D538G mutation which was also reported in the donor patient. In addition, estrogen receptor expression (2+-3+) was confirmed in both models. Growth characteristics for both models were comparable with a doubling time of approximately 15 days after model stabilization in the F3 passage. Slight or no activity was reported in the ST2535/HI model with tamoxifen (T/C=90%) or fulvestrant (T/C>100%). Treatment with everolimus (T/C=92%) or palbociclib (T/C=90%) was ineffective. However, combination of fulvestrant and palbociclib reported a super additive effect in the study with a T/C=46%. Interestingly, all tested agents were notably less responsive in the ST2535/HI versus similar ESR1Y537S/HI models. Conclusion: Overall, we have established and characterized two models of ESR1D538G-mutant breast cancer and compared in vivo activity to previously developed ESR1Y537S-mutant hormone independent models, all of which can be utilized for development of targeted therapies. Citation Format: Megan Groves, Lizette Gamez, Alyssa Moriarty, Amanda Mangold, Melissa Rundle, Kyriakos P. Papadopoulos, Drew W. Rasco, Anthony W. Tolcher, Amita Patnaik, Michael J. Wick. Establishment and characterization of an endocrine resistant ESR1D538G -mutant breast cancer PDX model [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2163.

Abstract P4-04-14: Anti-tumor activity of elacestrant (RAD1901) in combination with alpelisib (BYL-719) in patient-derived xenograft models of ER+ breast cancer

Abstract Estrogen-receptor positive (ER+) breast cancers make up approximately 70% of all breast cancers diagnosed. While patients with ER+ breast cancer have a better prognosis than other subtypes of breast cancer, the majority of those with advanced metastatic disease will eventually relapse. This has been attributed, in part, to mutations in the ER gene that result in constitutive activation of ER and contribute to aromatase inhibitor treatment resistance. As a strategy to deliver a more durable response in this setting, the use of selective estrogen receptor degraders (SERDs) that target and inhibit both wild-type and mutant ER has gained widespread attention. Indeed, fulvestrant, the only approved SERD on the market, is currently used as a second-line therapy in the metastatic setting, however, the intramuscular route of administration and pharmacokinetic properties of fulvestrant have fueled the development of orally bioavailable SERDs. We have previously described elacestrant (RAD1901), a novel and orally bioavailable selective estrogen receptor degrader (SERD) as an inhibitor of ER+ breast cancer growth in preclinical models, including those that harbor ER mutations and those that are insensitive to fulvestrant. In addition to ER mutations, the activation of parallel oncogenic pathways can also drive endocrine resistance, with the PI3K/Akt/mTOR pathway chief among those driving growth and treatment resistance to endocrine therapy. Consistent with these above-mentioned findings, recent clinical strategies to treat advanced ER+ disease have involved combining SERDs with PI3K inhibitors. Alpelisib (BYL-719) is a PI3K-alpha specific inhibitor that is being developed in combination with endocrine agents for the treatment of ER+ breast cancer. Here, we examined the effect of elacestrant in combination with alpelisib, in two ER+ breast cancer PDX models (one harboring wild-type ER and one harboring a Y537S mutation in the ER gene). The combination of elacestrant (10mg/kg) and alpelisib (35mg/kg) was well tolerated and resulted in significant tumor growth inhibition in both PDX models. Interestingly, in the mutant ER PDX model, the combination resulted in significantly greater growth inhibition relative to either compound alone. These data suggest the dual inhibition of the ER and PI3K signaling pathways with elacestrant and alpelisib produces significant anti-tumor activity in clinically-relevant PDX models, including those harboring ER mutations. Citation Format: Sankaran B, Garner F, Hattersley G, Purandare D, Bihani T. Anti-tumor activity of elacestrant (RAD1901) in combination with alpelisib (BYL-719) in patient-derived xenograft models of ER+ breast cancer [abstract]. In: Proceedings of the 2017 San Antonio Breast Cancer Symposium; 2017 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2018;78(4 Suppl):Abstract nr P4-04-14.

Abstract PD8-03: ESR1 gene fusions drive endocrine therapy resistance and metastasis in breast cancer

Abstract Background. Dysregulation of the estrogen receptor gene (ESR1) is an established mechanism of inducing endocrine therapy resistance. We previously discovered a chromosomal translocation event generating an estrogen receptor gene fused in-frame to C-terminal sequences of YAP1 (ESR1-YAP1) that contributed to endocrine therapy resistance in estrogen receptor positive (ER+) breast cancer models. This study compares functional, transcriptional, and pharmacological properties of additional ESR1 gene fusion events of both early stage (ESR1-NOP2) late stage (ESR1-YAP1 and ESR1-PCDH11x) breast cancers to gain a better understanding of therapeutic resistance and metastasis. Understanding the role of ESR1 fusions in inducing metastasis is critical, since the primary cause of death in breast cancer patients is through metastasis to distant sites. Methods. RNA-seq screens identified ESR1 fusions from early and late stage, endocrine therapy resistant breast tumor samples. Functional experiments were conducted using ER+ breast cancer cell lines, xenograft, and PDX models to test the ability of ESR1 fusions to induce therapeutic resistance and metastasis. ChIP-seq and RNA-seq were performed to examine transcriptional properties and differential gene expression induced by the fusions which directed subsequent pharmacological experiments with a CDK4/6 inhibitor. Results. ESR1-YAP1 and ESR1-PCDH11x promoted estrogen-independent and fulvestrant-resistant growth in vitro and induced greater tumor growth and increased metastatic capacity to the lungs of xenografted mice. In contrast, the ESR1-NOP2 fusion was sensitive to low estrogen conditions in vitro, and did not promote tumor growth. RNA-seq profiling revealed E2F targets pathway as the most highly enriched pathway induced by the ESR1 fusions. IHC revealed higher levels of pRb in ESR1-YAP1 and ESR1-PCDH11x xenograft tumors and subsequent CDK4/6 inhibition completely blocked tumor growth in an ESR1-YAP1 PDX model. Integrating RNA-seq with ChIP-seq data, we discovered a set of EMT and metastasis genes bound by all ESR1 fusions and WT-ER, but whose expression was strongly and uniquely up-regulated only by the ESR1-YAP1 and ESR1-PCDH11x fusions. These studies also revealed gained sites bound only by the ESR1-YAP1 and ESR1-PCDH11x fusions, not bound by WT-ER nor ESR1-NOP2. Genes mapping to these sites have a role in metastatic biology and were highly up-regulated by the YAP1 and PCDH11x fusions, potentially mediated by long range transcriptional activation. Conclusion. ESR1-YAP1 and ESR1-PCDH11x are driver fusions that occur in drug-resistant, advanced stage breast cancer and are a new class of recurrent somatic mutation that can cause acquired endocrine therapy resistance, yet can be treated with CDK4/6 inhibition. These driver fusions also confer increased metastatic ability through their ability to drive expression of genes that contribute to EMT and metastasis. In contrast, ESR1-NOP2 did not produce functional protein and appears to be a passenger event. These studies may provide pre-clinical rationale for targeting ESR1 translocated breast tumors, since the presence of an ESR1 driver fusion places a patient in a therapeutic category where none of the currently available endocrine therapies are likely to be effective. Citation Format: Lei JT, Shao J, Zhang J, Iglesia M, Chan DW, Cao J, Anurag M, Singh P, Haricharan S, Kavuri SM, Matsunuma R, Schmidt C, Kosaka Y, Crowder R, Hoog J, Phommaly C, Goncalves R, Ramalho S, Rodrigues-Peres RM, Lai W-C, Hampton O, Rogers A, Tobias E, Parikh P, Davies S, Ma C, Suman V, Hunt K, Watson M, Hoadley KA, Thompson A, Perou CM, Creighton CJ, Maher C, Ellis MJ. ESR1 gene fusions drive endocrine therapy resistance and metastasis in breast cancer [abstract]. In: Proceedings of the 2017 San Antonio Breast Cancer Symposium; 2017 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2018;78(4 Suppl):Abstract nr PD8-03.

Abstract P1-01-08: DPYSL3 modulates proliferation and migration in claudin-low breast cancer

Abstract Cancer is often driven by deregulated protein kinases pathways including breast cancer. To profile the kinome at the protein level we are developing a kinase pulldown (KiP) proteomic platform that uses multiple kinase inhibitors conjugated to sepharose beads for protein kinase profiling. Mass spectrometry-base tryptic peptide sequencing is used to identify and quantify the kinome in an unbiased manner. To date, we have profiled and analyzed the kinomes of more than 50 breast cancer PDX model systems that include all five breast cancer subtypes. In these analyses we have begun to explore the kinome interactome because we have observed that KiP also pulls down breast cancer subtype specific kinase interacting proteins that may represent new biological insights. For example we have consistently identified dihydropyrimidinase-like 3 (DPYSL3) in p21-activated kinase (PAK) inhibitor KiP interactome in the claudin-low WHIM12 PDX model. DPYSL3 is an intracellular phosphoprotein known to play a role in cell migration and metastasis. However it remains to be seen if DPYSL3 functions as a suppressor or a promoter of metastasis. We therefore generated DPYSL3 knock-down cell lines to evaluate function in claudin-low breast cancer. Proliferation levels of DPYSL3 depleted Claudin-low WHIM12 cells (DPYSL3-) were lower than those of control WHIM 12 (DPYSL3+) cells. In contrast, migration levels of DPYSL3- WHIM12 were greater than those of control cells, suggesting a pivotal role for DPYSL3 in several cellular physiologies. Additionally the activity of a PAK inhibitor (FRAX597) on migration appeared to require DPYSL3, as the inhibitory effect of FRAX579 on migration in WHIM12 was lost when DPYSL3 expression was depleted. Additionally we find that only the long isoform of DPYSL3, not a short isoform, interacts with PAK to regulate migration, suggesting that alternative splicing of DPYSL3 may an important to the control of migration. Furthermore Snail is also negatively regulated DPYSL3 with elevated expression in DYPSL3- cells. In conclusion, the study of the kinome interactome in breast cancer PDX identified DPYSL3 as regulator of the PAK family of kinases in a claudin low breast cancer. DPYSL3 is necessary for PAK to promote migration and may play a pivotal role in complex cellular transitions from a proliferative state versus an EMT/cell migratory state. Citation Format: Matsunuma R, Chan DW, Kim B-J, Singh P, Han A, Malovannaya A, Perou CM, Ellis MJ. DPYSL3 modulates proliferation and migration in claudin-low breast cancer [abstract]. In: Proceedings of the 2017 San Antonio Breast Cancer Symposium; 2017 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2018;78(4 Suppl):Abstract nr P1-01-08.

Abstract A106: Development of IDX-1197, a novel, selective, and highly potent PARP inhibitor

Abstract Background: PARP inhibitors have demonstrated clinically meaningful increase in progression-free survival as a single agent in women with recurrent ovarian cancer following a response to platinum-based chemotherapy. However, there still needs more improvement in efficacy because PARP inhibitors have comparatively low efficacy in non-germline BRCA-mutated patients. We aimed to develop a novel PAPR inhibitor that may have potent antitumor efficacy against the non-germline BRCA-mutated patients in addition to the germline BRCA-mutated patients. Here, we describe a novel selective PARP inhibitor IDX-1197. Materials and Methods: We conducted in vitro assays to evaluate IDX-1197 as a PARP inhibitor using PARP enzyme assay and PARP-catalytic inhibition assay in Hela cells. We evaluated in vivo efficacy of IDX-1197 against ovarian and breast tumor growth using patient-derived xenograft models. To investigate PARP inhibition in tumors, tumor PAR levels were measured by ELISA in the PDX models. Results: In enzyme assay, IDX-1197 potently inhibited PARP1 and PARP2 activities with IC50 values of 1.4 nM and 1.0 nM, respectively. By contrast, IDX-1197 was not sensitive to PARP5A (Tankyrase-1), which belongs to the PARP family (IC50>10 μM). In DNA damage-induced Hela cells, IDX-1197 significantly inhibited PARP1-mediated PAR expression (EC50 = 0.5 nM). In the germline BRCA1-mutated ovarian cancer PDX model, oral administration of IDX-1197 exhibited significant PAR inhibition (>90%) in tumor tissues until 24 hr post dose. IDX-1197 also dose-dependently led to potent tumor growth inhibition compared to Olaparib treatment group. Furthermore, IDX-1197 showed improved antitumor activity against breast cancer PDX models, which included both germline BRCA-mutated and non-germline BRCA-mutated models. In these models, IDX-1197 exhibited greater tumor growth inhibition compared with olaparib and/or niraparib. Conclusions: IDX-1197 is a highly potent and selective inhibitor of PARP1/2 with significant in vitro and in vivo activities in multiple cancer models. These preclinical data demonstrate the efficacy of IDX-1197, which has the potential for a best-in-class profile, in non-germline BRCA-mutated patients as well as BRCA-mutated patients. Based on these findings, IDX-1197 has entered clinical phase 1 trial in Korea. Funded by National OncoVenture. Citation Format: Myongjae Lee, Joon-Tae Park, Ji-hun Yang, Dohee Kim, In-Gyu Je, Yoon Suk Lee, Jinah Jeong, Dong Keun Song, Soobong Park, Hong-Sub Lee, Yong-Man Kim, Shin-Wha Lee, Dong Woo Kang, Ha-Young Lee, Jung Yong Kim, Sungsook Lee, Nam Seok Baek, Jae-Hoon Kang. Development of IDX-1197, a novel, selective, and highly potent PARP inhibitor [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2017 Oct 26-30; Philadelphia, PA. Philadelphia (PA): AACR; Mol Cancer Ther 2018;17(1 Suppl):Abstract nr A106.

Design of KDM4 Inhibitors with Antiproliferative Effects in Cancer Models.

Histone lysine demethylases (KDMs) play a vital role in the regulation of chromatin-related processes. Herein, we describe our discovery of a series of potent KDM4 inhibitors that are both cell permeable and antiproliferative in cancer models. The modulation of histone H3K9me3 and H3K36me3 upon compound treatment was verified by homogeneous time-resolved fluorescence assay and by mass spectroscopy detection. Optimization of the series using structure-based drug design led to compound 6 (QC6352), a potent KDM4 family inhibitor that is efficacious in breast and colon cancer PDX models.

Abstract 4038: Exploring optimal targeted combination therapies with neratinib for HER2+ breast cancer

Abstract Activation by amplification or overexpression of the proto-oncogene HER2 (also known as ERBB2) is associated with the development and progression of breast cancer. Neratinib is a novel, irreversible, pan-HER tyrosine kinase inhibitor which selectively inhibits EGFR, HER2 and HER4. In this preclinical study, we explored the efficacy of neratinib in combination with other clinically relevant targeted agents for optimal treatment of HER2-positive breast cancer in in vitro and in vivo models. Western blot (WB) analysis of a panel of breast cancer cell lines showed that BT-474, SKBR-3, and HCC-1954 cells displayed elevated HER2 expression levels (here denoted as HER2+), while MDA-MB-361, MDA-MB-453, and CAMA-1 breast cancer cells expressed relatively lower HER2 expression levels. Interestingly, neratinib inhibition positively correlated with HER2 expression levels as assessed by cell survival assays. The IC50 of neratinib in the HER2+ cell lines was in the 3.6 - 113nM range. Consistent with the in vitro data, treatment of a HER2+ breast cancer PDX model with neratinib single agent (p.o. 5 days/week) led to a significant dose-dependent tumor growth inhibition with tumor regressions observed at a low dose of 10 mg/kg (p<0.0001). The activity of neratinib in combination with other targeted agents was then evaluated in vitro in the HER2+ breast tumor cell lines using a Chou-Talalay method. Neratinib showed synergistic anti-proliferative activity in combination with the PI3Kα inhibitor BYL719 in all the HER2+ cell lines tested. By contrast, synergy with mTOR inhibitors (everolimus and TAK228) and with the MEK inhibitor trametinib was only observed in BT-474 and HCC-1954 cells. HCC-1954 cells had the best response to neratinib combinations among all HER2+ cell lines tested, with combination index values <0.5. Immunoblot analysis demonstrated that neratinib treatment displayed complete inhibition of HER2 phosphorylation in these cells. Substantial inhibition of downstream signaling pathways following single agent neratinib treatment was evident by reduced phosphorylation levels of AKT, S6K1, S6, 4EBP1, ERK1/2 and MEK1/2. Consistently with the cell viability data, the combination of neratinib with PI3K/Akt/mTOR and MAPK inhibitors showed a synergy on signaling blockage in these two downstream pathways respectively. Taken together this data shows that neratinib has efficacy as single agent in preclinical models of HER2+ breast tumors and its anti-proliferative activity in vitro was enhanced when combined with PI3K/Akt/mTOR and MAPK pathway inhibitors. Further studies of neratinib-based combination treatments are underway in HER2+ breast cancer xenograft and PDX models to further corroborate these findings. Citation Format: Ming Zhao, Stephen Scott, Kurt Evans, Erkan Yuca, Rashmi Murthy, Francesca Avogadri-Connors, Richard Cutler, Alshad S. Lalani, Sarina Piha-Paul, Funda Meric-Bernstam. Exploring optimal targeted combination therapies with neratinib for HER2+ breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 4038. doi:10.1158/1538-7445.AM2017-4038

Abstract 31: Preclinical development of a novel biparatopic HER2 antibody with activity in low to high HER2 expressing cancers

Abstract HER2-directed therapies have improved clinical outcomes for many patients with HER2 overexpressing breast and gastric cancer. Despite these successes, there remains a need to develop effective HER2-targeted therapies for these and other HER2-expressing tumors, particularly in the setting of recurrent or metastatic disease. One approach is to develop a single multifunctional antibody that has improved capacity and efficiency for binding HER2 compared with available HER2 inhibitors, can elicit ADCC, block ligand induced heterodimerization of HER2 with other HER family receptors, and down-regulate HER2 protein on the cell surface. ZW25 is a novel humanized, IgG1-like bispecific antibody directed against two distinct epitopes on the HER2 receptor. ZW25 has low nM binding affinity (0.9-16 nM) to recombinant HER2 and to cultured cancer cells with a range of HER2 expression, and shows higher maximal binding (1.4-1.9x) than monospecific HER2 antibodies on all tumor cell lines tested. The unique structure of ZW25 favors crosslinking of multiple HER2 receptors which is thought to promote clustering, and improves HER2 internalization compared to monospecific HER2 antibodies. In vitro studies demonstrated that ZW25 alone caused significant inhibition in growth of human cancer cell lines with a wide range of HER2 expression (5-54% inhibition depending on the cell line), and inhibited ligand-mediated tumor cell proliferation. ZW25 elicited concentration-dependent ADCC at nM potency (0.05-64 nM) with maximal lysis up to 52% on TNBC cell lines expressing HER2 at a 0/1+ level. ZW25 also exhibited synergy and additivity with multiple chemotherapeutic agents including platins, taxanes, microtubule inhibitors, and DNA synthesis inhibitors in various HER2 expressing tumor cell lines. In vivo studies demonstrated antitumor activity and/or improved survival against xenografts from a range of tumor types. In a HER2 3+ gastric cancer PDX model ZW25 induced significant tumor regressions, exhibiting 307% tumor growth inhibition relative to hIgG control and a 70% complete regression rate (trastuzumab exhibited 111% tumor growth control with no complete regressions). In a HER2 1+ breast cancer PDX model ZW25 significantly improved median survival (58 vs 28 days) and exhibited 141% tumor growth inhibition relative to hIgG control (trastuzumab exhibited 63% tumor growth inhibition, with median survival 39 days). In a low HER2-expressing NSCLC CDX model ZW25 improved median survival compared to hIgG control or cisplatin (> 66 vs 25 or 26 d respectively) and exhibited 159% tumor growth inhibition relative to hIgG control. In a GLP repeat dose toxicology study, ZW25 was well tolerated with no adverse effects at doses up to 150 mg/kg weekly IV for up to 8 weeks. Based on these findings, a first in human clinical trial of ZW25 has been initiated in patients with recurrent and/or metastatic HER2-expressing cancers. Citation Format: Nina Weisser, Grant Wickman, Rupert Davies, Gerry Rowse. Preclinical development of a novel biparatopic HER2 antibody with activity in low to high HER2 expressing cancers [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 31. doi:10.1158/1538-7445.AM2017-31

Abstract 1079: Preclinical activity of the FGFRinhibitor BAY 1163877 alone or in combination with antihormonal therapy in breast cancer

Abstract BAY 1163877 is an orally available, highly potent and selective pan fibroblast growth factor receptor (FGFR) inhibitor. In an ongoing Phase 1 clinical trial (NCT01976741) BAY 1163877 showed clinical responses at exceptional tolerability in patients suffering from different tumor types including urothelial bladder carcinoma or lung tumors, which were selected based on elevated FGFR1-3 mRNA expression. In the preclinical phase, the compound demonstrated significant single agent anti-tumor activity in various tumor models with different FGFR alterations leading to FGFR overexpression (e.g. FGFR gene amplifications or mutations). Genetic alterations of FGFRs can also be found in breast cancer with 7.5 - 17% of all tumors harboring a FGFR1 gene amplification. Elevated FGFR1 mRNA levels can be found in up to 22% of breast cancer cell lines as well as clinical samples. Other FGFR alterations include FGFR2 or FGFR4 gene amplifications as well as elevated FGFR mRNA levels, which were reported in all breast cancer subtypes. We therefore investigated BAY1163877 monotherapy in various breast cancer models. Due to the favorable clinical safety profile of BAY1163877, we also examined a combination treatment with early line antihormonal therapies in hormone receptor positive breast cancer. In vitro profiling of BAY 1163877 in a number of breast cancer cell lines showed a clear association of efficacy with expression levels of different FGFR isoforms. The efficacy was further investigated in several patient- or cell line-derived breast cancer in vivo models. For instance, BAY 1163877 alone dosed 38mg/kg twice daily induced tumor growth inhibition of greater than 90% in a subcutaneous mouse syngeneic 4T1 breast cancer model expressing elevated levels of FGFR2. Resistance to endocrine therapy appears associated with FGFR1 gene amplification and may explain the poor prognosis of FGFR1 overexpressing tumors treated with adjuvant tamoxifen. We therefore investigated the combination of the panFGFR-inhibitor BAY 1163877 with the clinically used antihormonal compound fulvestrant in selected luminal breast cancer PDX models. Some of these models showed resistance to antihormonal treatment in monotherapy but improved in vivo efficacy in combined treatment using BAY 1163877 and fulvestrant. These data may warrant further clinical investigation of BAY1163877alone or in combination with antihormonal therapy in patients with FGFR overexpressing breast cancer. Citation Format: Oliver Politz, Peter Ellinghaus, Sebastian Bender, Sylvia Gruenewald, Franziska Siegel, Marie-Pierre Collin, Sabine Zitzmann-Kolbe, Dominik Mumberg, Karl Ziegelbauer. Preclinical activity of the FGFRinhibitor BAY 1163877 alone or in combination with antihormonal therapy in breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 1079. doi:10.1158/1538-7445.AM2017-1079

Abstract P4-08-02: Integration of transcriptomic, proteomic and drug response data in triple negative breast cancer cell lines and PDX models

Abstract Drug response screens on panels of cell lines aimed at identifying markers of sensitivity or resistance have been limited in their successes. Unfortunately, the recent release of many such studies has been accompanied by concerns surrounding reproducibility. Since then, several publications have addressed these concerns by pointing out sources of variability and by suggesting better experimental methods as well as more robust analytical approaches. In the presented profiling effort, we integrated the latest advances in drug response measurement and focused on data diversity and quality rather than on breadth. We selected 32 breast cancer cell lines with a strong bias towards triple negative lines as well as 4 cell lines established from relevant patient-derived xenografts. We used high content microscopy to assay the phenotypic responses of the cell lines to a panel of 34 drugs made up largely of kinase inhibitors currently in the clinic along with some standard of care chemotherapeutics. The microscopy based drug response assay allowed us to measure drug potency, and to quantify the efficacy of the drugs in terms of growth inhibition and cell death. For the same cell lines, we used RNAseq to measure basal mRNA expression levels and shotgun mass spectrometry to measure endogenous protein levels. The completeness and controlled conditions under which these data sets were collected provide confidence in their integration. The complementarity of these multi-omics data has allowed us to address questions about the landscape of, particularly triple negative, breast cancer cell lines. Such questions include: where do the patient-derived lines lay among the established cell lines? and how different are the landscapes defined by drug response phenotypes, mRNA expression, and protein levels? We used network-based algorithms to identify eigenstates of signaling pathways related to genomic events, and further explored these states in the TCGA data. At the level of drug response, we have focused on important questions related to the clinical use of kinase inhibitors. In particular, we compared various CDK inhibitors in an effort to identify markers that are informative of response potency and efficacy. We have also looked at variability of the responses of the cell lines studied to multiple PI3K inhibitors that either target specific isoforms or all isoforms. Overall the data set that has been collected is a valuable resource for understanding drug response in triple negative breast cancer, and the transcriptomic and proteomic factors that influence it. Citation Format: Mills CE, Hafner MA, Sorger PK. Integration of transcriptomic, proteomic and drug response data in triple negative breast cancer cell lines and PDX models [abstract]. In: Proceedings of the 2016 San Antonio Breast Cancer Symposium; 2016 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2017;77(4 Suppl):Abstract nr P4-08-02.

164 - Development and characterization of HER2+ T-DM1-resistant breast cancer PDX models

164 - Development and characterization of HER2+ T-DM1-resistant breast cancer PDX models

Abstract 4353: Preclinical evaluation of the PARP inhibitor niraparib and cytotoxic chemotherapy alone or in combination in a panel of 25 triple-negative breast cancer PDX models: Relevance of BRCA mutations, HRD status and other biomarkers

Abstract Poly-ADP-ribose polymerase 1 (PARP) inhibitors are therapeutically effective in a subset of breast and high-grade ovarian cancers harboring deleterious BRCA1/2 mutations. Recently, several studies have tried to identify other molecular markers of HRD in BRCA1/2 wild-type (wt) breast and ovarian cancers to identify patients that could benefit from PARP inhibitor treatment. Most clinical studies of single-agent PARP inhibitors are therefore focused on breast or ovarian cancer with BRCA1/2 mutations and/or identified as HRD. Preclinical evidence also indicates that PARP inhibitors have therapeutic potential in combination with genotoxic chemotherapy. However, the toxicity associated with such combinations has generally imposed dose reduction of the chemotherapy, possibly explaining why the clinical activity observed with the combinations has in general not been superior to that of the corresponding single agents given at full dose. In a panel of 25 triple negative breast cancer (TNBC) PDX models, we investigated the antitumor efficacy of the PARP inhibitor niraparib used alone or in combination with cyclophosphamide, a backbone genotoxic drug of TNBC standard chemotherapy. Responses were correlated with both BRCA1/2 mutations and HRD status as defined by the myChoice HRD™ test from Myriad Genetics and with other HRD markers. Responses to cyclophosphamide ranged from progression to partial or complete tumor regression. Responses to niraparib were also variable and occurred only in tumors with BRCA mutations or a high HRD score. High sensitivities to niraparib and cyclophosphamide were tightly associated, suggesting that they may both be a result of a defect in a BRCA1/2-related pathway. We tested if niraparib could potentiate the efficacy of cyclophosphamide in TNBC models that initially regressed on cyclophosphamide but eventually relapsed. We chose a gapped sequential design in which treatment with niraparib was initiated 14 days after a full dose of cyclophosphamide, to avoid toxicity associated with concomitant administration. Potentiation with inhibition of tumor relapse was observed in tumors sensitive to single-agent niraparib, but not in tumors refractory to single-agent niraparib. In responder tumors, the cyclophosphamide-niraparib sequential combination was superior to sequential cycles of cyclophosphamide in preventing tumor relapse. These results show that a gapped sequential combination of full dose cyclophosphamide and niraparib is well tolerated and demonstrates remarkable efficacy in a subset of TNBCs. The data will be discussed with regard to potential adjuvant treatment strategies with PARP inhibitors in TNBC and the value of myChoice HRD™ as a biomarker in predicting response to PARP inhibition alone and in combination with chemotherapy. Citation Format: Olivier Deas, Stefano Cairo, Keith Wilcoxen, Keith Mikule, Truong-An Tran, Kirsten Timms, Jean-Gabriel Judde. Preclinical evaluation of the PARP inhibitor niraparib and cytotoxic chemotherapy alone or in combination in a panel of 25 triple-negative breast cancer PDX models: Relevance of BRCA mutations, HRD status and other biomarkers. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4353.

Abstract 868: Creating a superior, site-specific anti-HER2 antibody-drug conjugate (NG-HER2 ADC) for treatment of solid tumors

Abstract Antibody-drug conjugates (ADCs) have emerged as an important class of cancer therapeutics. The FDA approval of Kadcyla (T-DM1), a single agent for treatment of HER2-positive advanced metastatic breast cancer, was a significant milestone in the field of targeted therapy, as the first and only ADC for treatment of solid tumors. Despite the 3-month improvement over standard of care in the median survival, almost all the patients eventually became refractory to T-DM1. We have identified several possible areas for improvement: 1) The potency of T-DM1 as confirmed by the Phase III clinical data is restricted to high HER2 tumors which leaves moderate or low HER2 expressing patients without access to T-DM1 treatment; 2) The 48% overall response rate is indicative of intrinsic resistance to T-DM1 and all T-DM1 treated patients eventually relapse. 3) The randomized lysine conjugation in T-DM1 generates heterogeneity of the product. We have developed a novel, site-specific anti-HER2 ADC (NG-HER2 ADC) and evaluated it in comparative preclinical studies with T-DM1. The results show that the NG-HER2 ADC is ∼ 10 fold more potent than T-DM1 in HER2 3+ xenograft models of breast and gastric cancers. Our proprietary cleavable and permeable linker-payload can mediate bystander effect and this enables potent anti-tumor activity in non-HER2 amplified breast cancer and heterogeneous low HER2 NSCLC PDX models, where T-DM1 is ineffective. Our ADC can overcome T-DM1 resistance in in vitro and in vivo models.. Our site-specific ADC at HNSTD of 9 mg/kg in cynomolgus monkeys showed high AUC, long half-life and had normal clinical observations with no marked neutropenia. On the contrary, conventional conjugates with cleavable linker payloads typically have severe bone marrow toxicity as DLT above 5 mg/kg. The therapeutic index for NG-HER2 ADC is significantly greater than T-DM1 in all models tested. NG-HER2 ADC has a projected clinical efficacious dose of ∼1 mg/kg, compared to 3-5 mg/kg for T-DM1, based on PK/PD modeling. In addition, the activity of the NG-HER2 ADC shows increased infiltration of CD8 positive effector cells, an essential component for immuno-oncology (IO) efficacy, in a syngeneic HER2 overexpressing model. This property potentially allows the combination of the ADC with IO drugs to improve the long-term, overall survival. Our data provides preclinical proof of concept for NG-HER2 ADC with best-in-class potential and is currently being tested in preparation for clinical trials for treatment of HER2 solid tumors. Citation Format: Dangshe Ma, Bitha Narayanan, Kim Marquette, Edmund Graziani, Frank Loganzo, Manoj Charati, Nadira Prashad, Nathan Tumey, Jon Golas, Christine Hosselet, George Hu, Frank Barletta, Alison Betts, Judy Lucas, Chris O’Donnell, Lioudmila Tchistiakova, Hans-Peter Gerber, Puja Sapra. Creating a superior, site-specific anti-HER2 antibody-drug conjugate (NG-HER2 ADC) for treatment of solid tumors. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 868.

Abstract LB-057: 1-Benzylquinazoline-2,4(1H,3H)-diones as potent PARP inhibitors. SAR of the quinazoline group and in vivo efficacy of clinical candidate IMP4297

Abstract Poly (ADP-ribose) polymerase (PARP) is a critical enzyme for DNA repair and PARP-1 inhibitors have been found to be efficacious against cancers with BRCA mutations. Olaparib (AZD2281) has been approved as the first PARP-1 inhibitor for the treatment of ovarian cancers with BRCA mutations. In this presentation, we will report the SAR studies of 1-benzylquinazoline-2,4(1H,3H)-diones (BQD) as potent PARP inhibitors. We will present data for substituents at the quinazoline group of BQD, as well as in vivo efficacy data of IMP4297 which has been selected as clinical candidate for development. In a human triple negative breast cancer PDX model (BR-05-0028) with BRCA1 mutations, IMP4297 is highly efficacious against tumor growth with potency that is about 20-fold that of AZD2281. An IND for IMP4297 has been filed in China recently. Citation Format: Sui Xiong Cai, Qingbing Xu, Xiuhua Hu, Yangzhen Jiang, Guoxiang Wang, Ed Tian. 1-Benzylquinazoline-2,4(1H,3H)-diones as potent PARP inhibitors. SAR of the quinazoline group and in vivo efficacy of clinical candidate IMP4297. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr LB-057.