Related to figure 1. (A) SPR sensorgrams of WT WRN treated with AC1 and AC2. (B) DNA unwinding activity of wildtype WRN in the presence of AC1 and AC2. (C) Molecular docking of AC1 bound to the WRN helicase region (D) Viability of WRN dependent and independent cell lines treated with AC1 (left) or AC2 (right) after 3 days inhibitor treatment measured with CTG n=2 (E) Fold change γH2A.X immunofluorescence staining normalized to DMSO control in WRN dependent and independent cells treated with AC1 (left) or AC2 (right) for 3 days. n=2 for all lines except SW620 n=1

Abstract The Werner syndrome helicase (WRN) is a promising target for cancers with microsatellite instability (MSI), leading to the initiation of at least five phase I clinical trials. Acquired resistance is a substantial obstacle to obtaining lasting benefits from targeted therapies in oncology and may be particularly acute in the setting of mismatch repair–deficient (dMMR) tumors, which can sample increased fitness landscapes owing to a higher mutational burden. In this study, we characterized resistance mechanisms using the clinical candidate HRO761 and two novel inhibitors in MSI cell lines and xenograft models. We observed the rapid emergence of resistance both in vitro and in vivo, with sequencing revealing clustered mutations within the WRN helicase domain. Computational structural analyses indicated that these mutations either directly interfere with inhibitor binding or alter the protein conformation required for inhibitor engagement. Notably, although most mutations conferred broad resistance across all three compounds, we identified specific alterations (L528S, C727R, and F730L) that exhibited selectivity between chemical scaffolds. This chemotype-specific resistance profile suggests opportunities for developing next-generation inhibitors that retain activity against resistant variants and for implementing rational treatment strategies with existing inhibitors. Overall, our findings demonstrate that on-target resistance to WRN inhibitors emerges rapidly in dMMR backgrounds but also highlight potential approaches to overcome resistance, supporting the continued development of WRN-targeted therapies for MSI cancers.

Related to figure 3. (A) Mean tumor volume measurements (± SEM) in a HRO71-resistant SW48 xenograft model treated with vehicle or 120 mg/kg HRO761 (n = 10 mice per treatment group). (B) Mean body weight of mice corresponding to figure 3A (C) Mean body weight of mice corresponding to figure S3A (D) Heatmap of WES results for tumors treated with HRO761 in Fig 3A, B (E) Experiment level mutation results of HRO761 resistance experiments (F) Sanger sequencing results of HRO761 dose ramp-up resistance experiments (G) SPR sensorgram of WRN WT and G729D treated with HRO761 (H) As in G with C727R (I) As in G with F730L (J) As in G with D526_P530del

DNA unwinding activity of WRN mutant proteins

Abstract The expression of ILT (LILRB) family members and their major ligand HLA-G within solid tumors are associated with immune suppression and poor patient survival. Blocking multiple ILT family members might be more potent than targeting individual members by overcoming compensatory resistances within tumor microenvironment. Based on this hypothesis, we developed a monoclonal antibody (mAb) LITCHI 7 which reacts to three major ILT family members, ILT2, ILT4 and ILT5, and blocks ligand binding to these receptors. LITCHI 7 exhibits a broad binding capability to a wide range of human myeloid and T cell subsets that express ILT2, ILT4, and/or ILT5. Binding to monocyte-derived dendritic cells (moDC), LITCHI 7 released the inhibition on activating Fcγ receptor. On LPS-treated macrophages, LITCHI 7 enhanced inflammatory macrophage differentiation. The effect is primarily mediated by blocking ILT4 since ILT4 mono-specific mAb is as potent as cross-reactive LITHI 7 in modulating myeloid differentiation although both ILT2 and ILT5 are highly expressed by different myeloid subsets. The result was further confirmed by CRISPR Cas9 knock down of ILT2 or ILT5. In the absence of either ILT2 or ILT5, the accumulation of CD14+ monocytes and inflammatory macrophages were not changed during differentiation. While different ILT family members are generally enriched on myeloid cells, ILT2 is also expressed on NK cells and effector memory CD45RA+ T cells (Temra). Blocking ILT2 by LITCHI 7 enhanced TCR signaling when co-culturing the MART-1 specific T cell reporter line with MART-1/HLA-G positive melanoma cells. The effect of LITCHI 7 is mediated by ILT2 blockade since knocking out ILT2, but not ILT4 and ILT5, reduced accumulation of Temra in activated PBMC. To assess whether immune modulation of LITCHI 7 can be translated into anti-tumor activity by blocking different ILT family members, tumor killing assays were performed using innate and adaptive immune effector cells. LITCHI7 enhanced NK cell-mediated cytotoxicity on HLA-G+ tumor cells by blocking ILT2. Both ILT2 and ILT4 blockade play a role in macrophage phagocytosis on lymphoma cells since LITCHI 7 as well as individual anti-ILT2 or ILT-4 mAbs increased macrophage phagocytosis to a similar level. Finally, LITCHI 7 could boost in vitro T cell-mediated killing of CD19+ lymphoma by CD19 specific CAR-T cells in the presence of CD33+ myeloid cells. In summary, LITCHI 7 can enhance both innate and adaptive anti-tumor immunity through ILT2 or ILT4 blockade directly on effector cells or indirectly through myeloid cell reprogramming. As clinical trials are ongoing by targeting ILT2 and ILT4, blocking multiple ILT family members through a single cross-reactive mAb may provide broad anti-tumor effects and advantages when combining with additional oncology treatments. Citation Format: Shiming Ye, Dong Zhang, Brenal Singh, Jaishree Bankoti, Diane Cohen, Siusze Tan, Donghee Choi, Jonathan Hickson, Min-Zu Wu, Urvi Kolhatkar, Mikhail Binnewies, Wei Jin, Jason Hall, John Engelhardt, Alexander Shoemaker. Distinct functional effects of ILT family members in mediating the activity of an ILT2/ILT4/ILT5 cross-reactive monoclonal antibody in releasing the suppression on innate and adaptive anti-tumor immunity [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 741.

Abstract Immunoglobulin-like transcript 3 (ILT3) is an inhibitory receptor that is expressed by mature monocytes, dendritic cells (DC), plasma blasts, and the malignant cells arising thereof including acute myeloid leukemia (AML), B cell lymphoma and multiple myeloma (MM). We developed an anti-ILT3 blocking monoclonal antibody (mAb) 17G8 to test the hypothesis that blockade of ILT3 on hematopoietic malignant cells might drive anti-cancer responses through several mechanisms such as cell migration, ADCC, and T cell driven immune responses. Blockade of ILT3 using 17G8 without Fc effector function impaired AML cell line THP1 dissemination into distant sites in vivo and using 17G8 with enhanced Fc function through afucosylation further reduced the tumor burden. This effect of afucosylated 17G8 appears to be mediated by a mechanism other than ADCC, since afucosylated 17G8 showed modest tumor inhibition in ICR mice carrying ILT3 positive AML and MM xenografts. In immune competent mice carrying murine syngeneic AML cell C1498 expressing human ILT3, 17G8 also showed modest anti-tumor effects as a single agent, however, when combined with anti-PD1, afucosylated 17G8 increased CD8+ effector memory T cells, reduced proportion of cells with an exhausted phenotype and significantly improved survival when compared to different controls. Since T cells don’t express ILT3, the effects of 17G8 on T cells are hypothesized to be mediated by blocking ILT3 expressed on myeloid cells. The expression of ILT3 on immature human monocyte-derived DC (moDC) is higher than on mature moDC. Blocking ILT3 with afucosylated 17G8 on immature moDC potentiated maturation of moDC when compared to the controls in vitro. Surprisingly, afucosylated 17G8 stimulated IFNγ production from moDC during maturation. The enhanced DC function induced by afucosylated 17G8 was subsequently found to be translated into enhanced T cell function in an allogeneic mixed lymphocyte reaction (allo-MLR) assay, in which afucosylated 17G8 significantly increased IFNγ production when combined with anti-PD1. The activation of effector T cells was also monitored through a cytotoxicity assay by mixing THP-1 cells with human PBMC in culture. Afucosylated 17G8 induced strong cytotoxicity on THP-1 cells with minimal effects on normal ILT3 positive cells such as monocytes or moDC, suggesting that blockade of ILT3 with afucosylated 17G8 induced more potent T cell mediated allogeneic cytotoxicity than innate cell mediated ADCC killing. In summary, ILT3 is a highly selective M4/M5 AML target. Developing a blocking antibody with enhanced Fc function could add an additional therapeutic strategy of inducing host anti-tumor immunity to current AML treatments which induce direct tumor killing, such as chemotherapy, targeted therapy, and antibody drug conjugate. Citation Format: Shiming Ye, Dong Zhang, Min-Zu Wu, Harini Raghu, Diane Cohen, Mally Romero, Siu-Sze Tan, Donghee Choi, Jonathan Hickson, John Engelhardt, Alex Shoemaker. Blocking inhibitory receptor ILT3 by an antibody with enhanced Fc function promoted adaptive immunity against hematopoietic malignancy independent of ADCC [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1363.

Table S1: ABT-165 binding affinity; Table S2: ABT-165 in vitro potency; Table S3: Effect of VEGF on anti-DLL4 cellular potency of ABT-165; Table S4: Summary of in vivo efficacy; Table S5: Key safety findings of ABT-487 and ABT-165; Figure S1: Serum concentration-time profiles of ABT-165 in cynomolgus monkeys; Figure S2: Effect of antibody or antibody fragment valency on anti-DLL4 cellular potency; Figure S3: Effect of VEGF on the activity of DLL4 mAb and ABT-165 to downregulate DLL4 protein; Figure S4: Plasma levels of total soluble DLL4 and VEGF in cynomolgus monkeys after ABT-165 treatment.

Figures S1-S7 show LRRC15 RNA and protein expression in cancer, and demonstrate the anti-tumor efficacy of the LRRC15-targeted ADC ABBV-085 in vitro and in vivo as a monotherapy and in combination with standard-of-care therapies.

Supplementary Figure Legends

Compounds that inhibit glutathione peroxidase 4 (GPX4) hold promise as cancer therapeutics in their ability to induce a form of nonapoptotic cell death called ferroptosis. Our research identified 24, a structural analog of the potent GPX4 inhibitor RSL3, that has much better plasma stability (t1/2 > 5 h in mouse plasma). The bioavailability of 24 provided efficacious plasma drug concentrations with IP dosing, thus enabling in vivo studies to assess tolerability and efficacy. An efficacy study in mouse using a GPX4-sensitive tumor model found that doses of 24 up to 50 mg/kg were tolerated for 20 days but had no effect on tumor growth, although partial target engagement was observed in tumor homogenate.

Cross species binding and tolerability of ABBV-085.

Bruton's tyrosine kinase (BTK) is a key component of B cell receptor (BCR) signaling, and as such a critical regulator of cell proliferation and survival. Aberrant BCR signaling is important in the pathogenesis of various B cell malignancies and autoimmune disorders. Here, we describe the development of a novel positron emission tomography (PET) tracer for imaging BTK expression and/or occupancy by small molecule therapeutics. Radiochemistry was carried out by reacting the precursor with [18F]fluoride on a GE FX-FN TracerLab synthesis module to produce [18F]BTK-1 with a 6

In the development of cancer therapeutics, no suitable replacements for the use of animals that are capable of modeling such complex disease processes are currently available. In orthotopic models, surgery is often required to access the target organ for tumor cell inoculation. Historically analgesics have been withheld in such models in light of potential effects on tumor development. The current study evaluated the effect of the opioid buprenorphine on tumor growth of a human ovarian cancer cell line (OVCAR5 OT luc2 mCherry). Female CB17 SCID mice (n = 150) underwent surgery for orthotopic inoculation and were assigned to 1 of 3 treatment groups: vehicle control, 1 dose of buprenorphine, or 2 doses of buprenorphine administered perioperatively. Bioluminescence imaging revealed no significant difference on tumor engraftment rate or growth between control and analgesia-treated groups. These data demonstrate that acute, perioperative analgesia with buprenorphine did not alter tumor growth. Although further research is needed to evaluate potential effects of buprenorphine in other cell lines and mouse strains, the justification for withholding analgesia and the potential influence of pain and stress due to insufficient analgesia in these models should be considered thoroughly.

1025 Introduction: Bruton9s tyrosine kinase (BTK) is a key component of B-cell receptor signaling (BCR), and as such a critical regulator of cell proliferation and survival. Aberrant BCR signaling is important in the pathogenesis of various B-cell malignancies and autoimmune disorders. Here we report for the first time the development of a novel BTK PET tracer with the potential for imaging of target expression and/or target engagement (by small molecule therapeutics). Methods: Radiochemistry was carried out by reacting the precursor with [18F]fluoride on a GE FX-FN TracerLab synthesis module to produce [18F]ABBV-BTK1 with a 6% decay-corrected radiochemical yield, >3000 Ci/mmol specific activity, and a radiochemical purity of 99%.Following i.v. administration of [18F]ABBV-BTK1 (120±17 µCi, 0.084±0.05 µg) 60 minute dynamic images were acquired on an Inveon PET/CT system, in two xenograft models: REC-1 (n=6), an efficacious mantle cell lymphoma model, and U87MG (n=6), a non-efficacious glioblastoma model. Subsequent studies included vehicle, pretreatment (10 min prior to tracer injection) and displacement (30 min post-tracer injection) studies with a reversible BTK inhibitor to examine BTK binding. Regions of interest were manually delineated in the tumor and left ventricle and time activity curves were generated. Estimates of volume of distribution (VT) of the tracer (defined as the ratio of the tracer concentration in tumor (CT, kBq·cm-3) to that in plasma (CP, kBq·mL-1) at equilibrium) were obtained using a metabolite-corrected image-derived input function and kinetic modeling. Results: VT, representing uptake of [18F]ABBV-BTK1, was significantly higher in BTK expressing REC-1 tumors (4.0±1.6) than non-BTK expressing U87MG tumors (0.5±0.13). Administration of BTK inhibitors reduced [18F]ABBV-BTK1 binding in the REC-1 tumor model consistent with [18F]ABBV-BTK1 binding to BTK. Conclusions: [18F]ABBV-BTK1 is a promising PET tracer candidate for PET imaging of BTK. Additional experiments are being conducted to further characterize tracer binding in vivo and explore potential clinical applications including assessment of target engagement. [asterisk]Disclosures: All authors are employees of AbbVie. The design, study conduct, and financial support for this research were provided by AbbVie. AbbVie participated in the interpretation of data, review, and approval of the publication.

Abstract Progress in understanding tumor stromal biology has been constrained in part because cancer-associated fibroblasts (CAF) are a heterogeneous population with limited cell-type–specific protein markers. Using RNA expression profiling, we identified the membrane protein leucine-rich repeat containing 15 (LRRC15) as highly expressed in multiple solid tumor indications with limited normal tissue expression. LRRC15 was expressed on stromal fibroblasts in many solid tumors (e.g., breast, head and neck, lung, pancreatic) as well as directly on a subset of cancer cells of mesenchymal origin (e.g., sarcoma, melanoma, glioblastoma). LRRC15 expression was induced by TGFβ on activated fibroblasts (αSMA+) and on mesenchymal stem cells. These collective findings suggested LRRC15 as a novel CAF and mesenchymal marker with utility as a therapeutic target for the treatment of cancers with LRRC15-positive stromal desmoplasia or cancers of mesenchymal origin. ABBV-085 is a monomethyl auristatin E (MMAE)-containing antibody–drug conjugate (ADC) directed against LRRC15, and it demonstrated robust preclinical efficacy against LRRC15 stromal-positive/cancer-negative, and LRRC15 cancer-positive models as a monotherapy, or in combination with standard-of-care therapies. ABBV-085′s unique mechanism of action relied upon the cell-permeable properties of MMAE to preferentially kill cancer cells over LRRC15-positive CAF while also increasing immune infiltrate (e.g., F4/80+ macrophages) in the tumor microenvironment. In summary, these findings validate LRRC15 as a novel therapeutic target in multiple solid tumor indications and support the ongoing clinical development of the LRRC15-targeted ADC ABBV-085. Significance: These findings identify LRRC15 as a new marker of cancer-associated fibroblasts and cancers of mesenchymal origin and provide preclinical evidence for the efficacy of an antibody-drug conjugate targeting the tumor stroma. Cancer Res; 78(14); 4059–72. ©2018 AACR.

Abstract Background: Leucine rich repeat containing 15 (LRRC15) is a TGFβ-regulated structural protein that is highly expressed on cancer-associated fibroblasts (CAFs) in the stromal microenvironment of many solid tumors, as well as directly on cancer cells of mesenchymal origin. Soft tissue sarcomas (STS) and bone sarcomas (BS) represent a diverse family of mesenchymal malignancies that can develop at any anatomic site and that comprise more than 70 histopathologic subtypes. After screening LRRC15 expression across a variety of sarcoma histologies, we evaluated the antitumor activity of ABBV-085, an MMAE (monomethyl auristatin E) containing antibody-drug conjugate directed against LRRC15 (mouse, cyno, human), in patient-derived xenograft (PDX) models of selected sarcomas with varying levels of LRRC15 expression. Methods: LRRC15 expression/intensity in sarcoma histologies was performed by immunohistochemical (IHC) staining (Leica Bond RX, Bond Polymer Refine Kit) with a human LRRC15 specific mouse IgG2b antibody. Based on LRRC15 expression levels, STS and BS tumor fragments (PDXs) were implanted into NSG mice. Mice with growing tumors were then selected to evaluate the in vivo efficacy of ABBV-085 monotherapy (6 mg/kg). Results: LRRC15 expression was evaluated in 340 human sarcoma tumors representing a variety of STS and BS histologic subtypes. LRRC15 IHC expression was determined by scoring the percentage of positive cells, together with the intensity of staining (score of 0 for negative, 1 for weak, 2 for moderate, and 3 for strong staining), for the cancer cells and stroma, respectively. A stringent cut-off for strong LRRC15 positivity was defined as ≥2+ intensity in ≥50% of the cancer or stromal area. Strong LRRC15 expression was observed in several sarcoma subsets: 67% (14/21) of osteosarcoma (OS) tumor samples, 64% (23/36) of undifferentiated pleomorphic sarcoma (UPS), 18% (8/44) of leiomyosarcomas (LMS) and 17% (6/35) of liposarcomas (LPS). Significant antitumor activity including regressions and cures was induced by ABBV-085 in LRRC15-positive STS and BM PDX models, when compared with isotype-control treated mice. The ABBV-085-induced efficacy in osteosarcoma PDX models was superior to current standard-of-care therapies when dosed maximally in mice. In addition, ABBV-085 demonstrated efficacy in different LRRC15 positive STS subtypes including UPS, LMS and LPS. ABBV-085 was well tolerated with minimal to no body weight loss observed. Conclusions: LRRC15 is highly expressed in the majority of human osteosarcomas and undifferentiated pleomorphic sarcomas, as well as in a range of other STS and BS subtypes. ABBV-085 demonstrates promising preclinical antitumor efficacy in LRRC15-positive PDX models of STS and BS. ABBV-085 is currently being investigated in an ongoing phase 1 study in soft tissue sarcomas (including undifferentiated pleomorphic sarcoma) and osteosarcoma. Citation Format: Eytan Ben-Ami, Ying Huang, Prafulla C. Gokhale, Benjamin Eschle, Lisa Durkin, Jonathan Hickson, Mien Sho, Susan Morgan-Lappe, Kurt Gish, Dominic W. Lai, Randy R. Robinson, Diane Hollenbaugh, Eric D. Hsi, Debra T. Chao, George D. Demetri, James W. Purcell. LRRC15 is a novel antigen in sarcoma and the therapeutic target of the antibody-drug conjugate (ADC) ABBV-085 [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 953.

Abstract Antiangiogenic therapy is a clinically validated modality in cancer treatment. To date, all approved antiangiogenic drugs primarily inhibit the VEGF pathway. Delta-like ligand 4 (DLL4) has been identified as a potential drug target in VEGF-independent angiogenesis and tumor-initiating cell (TIC) survival. A dual-specific biologic targeting both VEGF and DLL4 could be an attractive strategy to improve the effectiveness of anti-VEGF therapy. ABT-165 was uniquely engineered using a proprietary dual-variable domain immunoglobulin (DVD-Ig) technology based on its ability to bind and inhibit both DLL4 and VEGF. In vivo, ABT-165 induced significant tumor growth inhibition compared with either parental antibody treatment alone, due, in part, to the disruption of functional tumor vasculature. In combination with chemotherapy agents, ABT-165 also induced greater antitumor response and outperformed anti-VEGF treatment. ABT-165 displayed nonlinear pharmacokinetic profiles in cynomolgus monkeys, with an apparent terminal half-life > 5 days at a target saturation dose. In a GLP monkey toxicity study, ABT-165 was well-tolerated at doses up to 200 mg/kg with non-adverse treatment–related histopathology findings limited to the liver and thymus. In summary, ABT-165 represents a novel antiangiogenic strategy that potently inhibits both DLL4 and VEGF, demonstrating favorable in vivo efficacy, pharmacokinetic, and safety profiles in preclinical models. Given these preclinical attributes, ABT-165 has progressed to a phase I study. Mol Cancer Ther; 17(5); 1039–50. ©2018 AACR.

Improving the congruity of preclinical models with cancer as it is manifested in humans is a potential way to mitigate the high attrition rate of new cancer therapies in the clinic. In this regard, three-dimensional (3D) tumor cultures in vitro have recently regained interest as they have been acclaimed to have higher similarity to tumors in vivo than to cells grown in monolayers (2D). To identify cancer functions that are active in 3D rather than in 2D cultures, we compared the transcriptional profiles (TPs) of two non-small cell lung carcinoma cell lines, NCI-H1650 and EBC-1 grown in both conditions to the TP of xenografted tumors. Because confluence, diameter or volume can hypothetically alter TPs, we made intra-and inter-culture comparisons using sampleswith defined dimensions. As projected by Ingenuity Pathway Analysis (IPA), a limited number of signal transduction pathways operational in vivowere better represented by 3Dthan by 2D cultures in vitro. Growth of 2Dand 3D cultures as well as xenografts inducedmajor changes in the TPs of these 3 modes of culturing. Alterations of transcriptional network activation that were predicted to evolve similarly during progression of 3D cultures and xenografts involved the following functions: hypoxia, proliferation, cell cycle progression, angiogenesis, cell adhesion, and interleukin activation. Direct comparison of TPs of 3D cultures and xenografts to monolayer cultures yielded up-regulation of networks involved in hypoxia, TGF andWnt signaling aswell as regulation of epithelial mesenchymal transition. Differences in TP of 2D and 3D cancer cell cultures are subject to progression of the cultures. Theemulation of the predicted cell functions in vivo is therefore not only determined by the type of culture in vitro but also by the confluence or diameter of the 2D or 3D cultures, respectively. Consequently, the successful implementation of 3D models will require phenotypic characterization to verify the relevance of applying these models for drug development.

Abstract The first generation anti-angiogenic drugs designed to block the VEGF/VEGFR pathway lend modest clinical benefit for cancer patients. Other than VEGF, DLL4 is the only known angiogenic factor with a haploinsufficiency phenotype, underscoring its essential role in vascular function. Indeed, both the VEGF/VEGFR and the DLL4/Notch signaling axes are known to cooperate during pathological angiogenesis. DLL4 is also implicated in VEGF-independent pathways, cancer stem cell survival, and immune suppression that could collectively contribute to tumor cell resistance. Given both intrinsic and acquired patient resistance mechanisms exist, targeting the DLL4/Notch pathway represents a unique opportunity for a combination strategy to improve upon current VEGF/VEGFR pathway inhibitor therapies. To this end, ABT-165 was developed as a first-in-class dual specific biologic using AbbVie's proprietary dual-variable domain immunoglobulin (DVD-IgTM) technology. ABT-165 is capable of simultaneously binding to DLL4 and VEGF with nanomolar affinities and blocking the cognate ligand-receptor interactions that result in the potent inhibition of DLL4-mediated Notch1 activation and VEGF-stimulated endothelial cell proliferation. ABT-165 is functionally superior in vitro compared to the combination of parental anti-VEGF and anti-DLL4 antibodies. In human tumor xenograft models, ABT-165 induced significant inhibition of tumor growth and survival benefit compared to single anti-DLL4 or anti-VEGF antibody treatments at equivalent doses. Mechanistically, this enhancement of anti-tumor efficacy is due in part to the disruption of new tumor vasculature coupled with blockade of vessel perfusion. Furthermore, ABT-165 in combination with cytotoxic chemotherapy agents induced tumor regression, which outperformed bevacizumab plus chemotherapy in both human breast and colon xenograft models. ABT-165 displays non-linear, dose-dependent pharmacokinetic profiles in mice and cynomolgus monkeys, with an apparent terminal half-life > 5 days in both species at a target saturation dose. In a GLP monkey toxicity study, ABT-165 at doses up to 200 mg/kg was well-tolerated with non-adverse treatment-related histopathology findings limited to the liver and thymus. In contrast, adverse and non-adverse findings were observed in the hearts of rats and monkeys, respectively, with an in-house proprietary anti-DLL4 antibody. Given that coupling of anti-DLL4 with anti-VEGF activities into a DVD-Ig may lend improved safety and/or efficacy profiles compared to antibodies, ABT-165 was advanced into a Phase 1 clinical trial. Disclosures: All authors are employees of AbbVie. The design, study conduct, and financial support for this research were provided by AbbVie. AbbVie participated in the interpretation of data, review, and approval of the publication. Citation Format: Yingchun Li, Jonathan Hickson, Dominic Ambrosi, Deanna Haasch, Kelly Foster-Duke, Lucia Eaton, Fang Jiang, Surekha Akella, Wenqing Gao, Sherry Ralston, Jijie Gu, Susan Morgan-Lappe. ABT-165 is a first-in-class therapeutic Dual Variable Domain Immunoglobulin (DVD-IgTM) that targets DLL4 and VEGF for the treatment of cancer. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 867.