Natalya Bodyak

A Polymer-Based Antibody-Vinca Drug Conjugate Platform: Characterization and Preclinical Efficacy

Antibody-drug conjugates (ADC) are an emerging drug class that uses antibodies to improve cytotoxic drug targeting for cancer treatment. ADCs in current clinical trials achieve a compromise between potency and physicochemical/pharmacokinetic properties by conjugating potent cytotoxins directly to an antibody at a 4:1 or less stoichiometric ratio. Herein, we report a novel, polyacetal polymer-based platform for creating ADC that use poly-1-hydroxymethylethylene hydroxymethyl-formal (PHF), also known as Fleximer. The high hydrophilicity and polyvalency properties of the Fleximer polymer can be used to produce ADC with high drug loading without compromising physicochemical and pharmacokinetic properties. Using trastuzumab and a vinca drug derivative to demonstrate the utility of this platform, a novel Fleximer-based ADC was prepared and characterized in vivo. The ADC prepared had a vinca-antibody ratio of 20:1. It exhibited a high antigen-binding affinity, an excellent pharmacokinetic profile and antigen-dependent efficacy, and tumor accumulation in multiple tumor xenograft models. Our findings illustrate the robust utility of the Fleximer platform as a highly differentiated alternative to the conjugation platforms used to create ADC currently in clinical development.

Abstract LB-231: A novel, highly potent HER2-targeted antibody-drug conjugate (ADC) for the treatment of low HER2-expressing tumors and combination with trastuzumab-based regimens in HER2-driven tumors

Antibody-drug conjugates are effective in the treatment of HER2-amplified breast cancer and Hodgkin9s lymphoma, but current ADC technologies have faced limitations expanding the addressable patient population and target space. Ado-trastuzumab emtansine (T-DM1) is an ADC with 3-4 cytotoxic drugs per antibody that was recently approved for HER2 IHC 3+ or HER2-amplified breast cancer. Even within this high HER2-expressing population, several studies have now shown greater T-DM1 benefit in patients with HER2 mRNA expression above the median. These data suggest the need for more potent anti-HER2 ADCs to maximize benefit for HER2 IHC 3+ or amplified patients, and to extend HER2 ADC therapy to low HER2-expressing patients (HER2 IHC 1+/2+). XMT-1522 is an anti-HER2 ADC that uses a novel, human anti-HER2 antibody optimized for cytotoxic payload delivery, and is non-competitive with trastuzumab or pertuzumab for HER2 binding. Each antibody is conjugated to ∼15 proprietary auristatin molecules using Fleximer, a biodegradable hydrophilic polymer. XMT-1522 shows nanomolar potency in cultured tumor cells with HER2 receptor densities as low as 10,000 per cell, and is typically 1-3 logs more potent than T-DM1 across a panel of 25 tumor cell lines. In mouse xenograft studies XMT-1522 has excellent pharmacokinetic properties and achieves complete tumor regressions at well-tolerated doses. In the high HER2-expressing N87 gastric cancer model (800,000 HER2 receptors/cell), complete regressions are achieved with a single 1 mg/kg dose of XMT-1522, while 10 mg/kg T-DM1 is required for comparable activity. In the same model, the XMT-1522/trastuzumab/pertuzumab triple combination results in tumor regressions where the same doses of XMT-1522 alone or the trastuzumab/pertuzumab doublet result in tumor stasis. In the low HER2-expressing JIMT-1 breast cancer (79,000 HER2/cell) and SNU5 gastric cancer (22,000 HER2/cell) models, complete regressions are achieved with single 1 mg/kg or 0.67 mg/kg doses of XMT-1522, respectively, while T-DM1 is inactive at doses ≥10 mg/kg. In non-human primates XMT-1522 demonstrates good stability of drug conjugate in plasma with t1/2 ∼5 days (comparable to antibody t1/2) and minimal exposure to free payload. Despite the high potency of XMT-1522 in low HER2 tumor models, there is no XMT-1522-related toxicity observed in critical HER2-expressing tissues including heart and lung. The preclinical data support testing XMT-1522 as a single agent in tumors with low HER2 expression where current HER2-directed therapies are not indicated. Furthermore, combination of XMT-1522 with trastuzumab and/or pertuzumab achieves efficient cytotoxic payload delivery while retaining the potential for full inhibition of HER2 signaling, which may be necessary to improve on current regimens in HER2-driven tumors. Citation Format: Donald A. Bergstrom, Natalya Bodyak, Alex Yurkovetskiy, Peter U. Park, Michael DeVit, Mao Yin, Laura Poling, Joshua D. Thomas, Dmitry Gumerov, Dongmei Xiao, Elena Ter-Ovanesyan, LiuLiang Qin, Alex Uttard, Alex Johnson, Timothy B. Lowinger. A novel, highly potent HER2-targeted antibody-drug conjugate (ADC) for the treatment of low HER2-expressing tumors and combination with trastuzumab-based regimens in HER2-driven tumors. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr LB-231. doi:10.1158/1538-7445.AM2015-LB-231

Abstract 1194: Discovery and preclinical development of a highly potent NaPi2b-targeted antibody-drug conjugate (ADC) with significant activity in patient-derived non-small cell lung cancer (NSCLC) xenograft models

The type II sodium-dependent potassium transporter NaPi2b (SLC34A2) is highly expressed in non-squamous NSCLC and non-mucinous ovarian cancer (OC) with restricted normal tissue expression, suggesting it may be a suitable ADC target for these indications. XMT-1536 is a novel, highly potent anti-NaPi2b ADC comprised of an average of 15 auristatin molecules conjugated to XMT-1535, a novel humanized anti-NaPi2b antibody, via the Dolaflexin ADC platform. The auristatin payload is enzymatically cleaved upon ADC trafficking to the endosome/lysosome compartment, releasing a cytotoxic auristatin derivative that is capable of bystander effect killing. In cell binding assays, XMT-1535 antibody binds to OC cells with low nanomolar affinity, which is unaffected by conjugation of the Dolaflexin drug conjugate. In vitro cytotoxicity assays show picomolar potency of XMT-1536 in OVCAR3 (OC; 32,000 NaPi2b molecules/cell; IC50 2 pM), TOV21G (OC; 10,000 NaPi2b molecules/cell; IC50 40 pM), and HCC-4006 (NSCLC; 52,000 NaPi2b molecules/cell; IC50 130 pM). In each cell line, XMT-1536 is 1-2 logs more potent than a non-binding Dolaflexin ADC control, consistent with target-dependent cytotoxic effect. XMT-1536 was tested in mouse xenograft models of OC and NSCLC. In the OVCAR3 OC model, XMT-1536 induced partial tumor regressions after a single dose of 3 mg/kg (0.21 mg/kg payload equivalent dose), and complete tumor regressions after a single dose of 5 mg/kg (0.36 mg/kg payload dose) or 3 weekly doses of 3 mg/kg. In contrast, a non-binding Dolaflexin ADC with comparable drug loading was inactive after 3 weekly administrations of 3 mg/kg, consistent with the anti-tumor activity of XMT-1536 being mediated through binding to the NaPi2b target. XMT-1536 was also tested in a patient-derived model of KRAS mutant NSCLC, where 3 weekly doses of 3 mg/kg led to significant tumor growth delay and regressions in some animals. Evaluation of XMT-1536 in additional patient derived xenograft models is on-going and will be updated at the meeting. XMT-1535 is cross-reactive with cynomolgous monkey NaPi2b, allowing an informative evaluation of whether XMT-1536 retains good tolerability in non-human primate. XMT-1536 was administered to cynomolgous monkeys in an exploratory single dose study up to 5 mg/kg ADC (4294 μg/m2 auristatin payload equivalents), with no observed target-mediated toxicity and limited adverse findings. Of note, there was no evidence of bone marrow toxicity, which has been observed generally for cleavable auristatin ADCs, and specifically for a recently published auristatin-based NaPi2b ADC (Lin et al., Clinical Cancer Research, 2015). Based on these data XMT-1536 is advancing to early clinical development for the treatment of NaPi2b-expressing tumors. Citation Format: Natalya Bodyak, Alex Yurkovetskiy, Mao Yin, Dmitry Gumerov, Reddy Bollu, Patrick Conlon, Venu R. Gurijala, Dennis McGillicuddy, Cheri Stevenson, Elena Ter-Ovanesyan, Peter U. Park, Laura Poling, Winnie Lee, Michael DeVit, Dongmei Xiao, LiuLiang Qin, Timothy B. Lowinger, Donald A. Bergstrom. Discovery and preclinical development of a highly potent NaPi2b-targeted antibody-drug conjugate (ADC) with significant activity in patient-derived non-small cell lung cancer (NSCLC) xenograft models. [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 1194.

Abstract 1693: ASN004, a novel 5T4-targeted Dolaflexin™ antibody drug conjugate, causes complete regression in multiple solid tumor models

ASN004 is an Antibody Drug Conjugate (ADC) that targets the 5T4 oncofetal antigen (trophoblast glycoprotein), which is highly expressed in a wide range of malignant tumors, while having very limited expression in normal tissues. ASN004 incorporates a novel single-chain homo-dimer antibody, Fleximer® linker technology (Mersana Therapeutics), and several cytotoxic dolastatin (auristatin) analog warheads per ADC molecule (drug/antibody ratio ∼15). ASN004 shows high affinity for the 5T4 antigen and for 5T4-expressing tumor cells. As well, ASN004 shows potent cytotoxicity that is selective for 5T4-expressing tumor cells. ASN004 provides strong tumor regression and tumor-free survivors in multiple tumor xenograft models, at well-tolerated doses as low as 0.3 mg/kg iv. Furthermore, ASN004 causes tumor regression when administered to xenografts bearing more advanced (500 mm3) tumors. Robust, potent efficacy for ASN004 has also been demonstrated in head-to-head comparison studies with relevant reference ADCs. A favorable pharmacokinetics profile was observed in rodents and primates. Dose-range finding exploratory toxicology studies have been completed in both pharmacological and non-pharmacological non-clinical species. Efforts toward IND-enabling safety studies with this promising new agent are in progress. Citation Format: Roger A. Smith, Nitin K. Damle, Sanjeeva P. Reddy, Alex Yurkovetskiy, Natalya Bodyak, Mao Yin, Dmitry Gumerov, Elena Ter-Ovanesyan, Liu Qin, Peter U. Park, Timothy B. Lowinger, Sandeep Gupta. ASN004, a novel 5T4-targeted Dolaflexin™ antibody drug conjugate, causes complete regression in multiple solid tumor models. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 1693. doi:10.1158/1538-7445.AM2015-1693

Abstract 48: Non-clinical pharmacokinetics of XMT-1522, a HER2 targeting auristatin-based antibody drug conjugate

The ADC XMT-1522 consists of a novel human IgG1 anti-HER2 monoclonal antibody and a novel, auristatin-based cytotoxic payload (Auristatin F-hydroxypropylamide, AF-HPA). An average DAR of 12 AF-HPA molecules is achieved via a biodegradable polymer conjugation platform. The non-clinical DMPK properties of XMT-1522 have been characterized in vitro in plasma and microsomal stability studies, and in vivo in plasma and tissue disposition and excretion studies. Sample analysis for total AF-HPA drug payload and released (free) AF-HPA and its metabolites was performed by ESI+ LC/MS/MS; total antibody was determined by ELISA. The half-life for AF-HPA release in plasma was found to be greater than 120 hours in all species tested. Microsomal stability studies showed that AF-HPA was further converted to other metabolites including the carboxylic acid auristatin F (AF), as well as monomethyl auristatin F-HPA (MMAF-HPA) and MMAF. The pharmacokinetic profiles of XMT-1522 were evaluated in mouse, rat and cynomolgus monkey. The antibody of XMT-1522 is cross-reactive with monkey, but not rodent, HER2. In mouse and rat, XMT-1522 exposure was dose-proportional; exposure was slightly greater than dose-proportional in monkey consistent with saturation of target-mediated clearance. All species showed extended exposure to total AF-HPA drug payload, with measured clearance and volume of distribution similar for total AF-HPA and the antibody component of XMT-1522. Exposure to free AF-HPA and AF was less than 1/1000th the exposure of total AF-HPA. These data indicate the vast majority of AF-HPA in plasma is antibody-conjugated, indicating high stability of the ADC in systemic circulation. XMT-1522 tissue disposition was studied in NCI-N87 HER2-positive gastric cancer xenograft tumor bearing mice. After a single 3 mg/kg dose of XMT-1522, free AF-HPA and its metabolite AF were measurable in tumor tissue until the last time point measured (2 weeks). Total AF-HPA and free AF-HPA achieved peak tumor concentrations 48 hours after dosing. In contrast, AF achieved peak tumor concentration 7 days after dosing and showed only a slight decline in tumor concentration at 14 days, consistent with intracellular trapping of this poorly cell-permeable metabolite. Exposure to free AF-HPA or AF in other tissues was at least an order of magnitude lower than in tumor; in tissues with measurable free drug, AF was the predominant species. XMT-1522 excretion studies, conducted in rat, indicated that the AF-HPA payload was mainly excreted by the gastrointestinal route. In the first 96 hours after administration 33% of the AF-HPA dose was excreted in feces, compared to 3% excreted in urine. The major contributing metabolites both in feces and urine were conjugated AF-HPA, AF, and free AF-HPA. In conclusion, the plasma kinetics, tissue distribution and excretion profile of XMT-1522 are acceptable for clinical evaluation in cancer patients. Citation Format: Alex Yurkovetskiy, Dmitry Gumerov, Elena Ter-Ovanesyan, Patrick Conlon, Michael Devit, Charlie Bu, Natalya Bodyak, Timothy Lowinger, Donald Bergstrom. Non-clinical pharmacokinetics of XMT-1522, a HER2 targeting auristatin-based antibody drug conjugate [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 48. doi:10.1158/1538-7445.AM2017-48

Abstract 641: Trastuzumab-dolaflexin, a highly potent Fleximer-based antibody-drug conjugate, demonstrates a favorable therapeutic index in exploratory toxicology studies in multiple species

Proceedings: AACR 106th Annual Meeting 2015; April 18-22, 2015; Philadelphia, PA Antibody-drug conjugates (ADCs) represent a promising drug class that relies on monoclonal antibody recognition of specific cancer-associated antigens for targeted delivery of chemotherapeutic agents. Fleximer® ADCs utilize a polymer-based conjugation platform to enable high drug-antibody ratios (DAR) and significantly greater anti-tumor potency compared to ADCs with DAR 3-4. We previously showed that trastuzumab-dolaflexin (T-dolaflexin), a HER2-targeted ADC comprised of the antibody trastuzumab conjugated to ∼15 proprietary auristatin molecules via Fleximer, has excellent pharmacokinetics and efficacy in mouse models (AACR 2014 Abstract #2645). T-dolaflexin is efficacious at a single dose of 0.67 mg/kg in mouse xenograft models, and achieves prolonged tumor-free survival after a single 2 mg/kg dose in a low HER2-expressing model that is insensitive to ado-trastuzumab emtansine (T-DM1). We sought to test the tolerability of T-dolaflexin in exploratory mouse, rat and non-human primate toxicology studies. Cohorts of 6 mice were treated with a single dose of vehicle or T-dolaflexin at 20, 30 or 40 mg/kg and followed for 21 days. The 20 and 30 mg/kg doses were well-tolerated based on body weight loss and mortality and achieved a therapeutic index (TI) in mouse >40X. Cohorts of 4 female cynomolgus monkeys were treated with a single dose of vehicle or T-dolaflexin at 0.67, 1.34 or 2.68 mg/kg (payload doses 600, 1200 and 2400 μg/m2, respectively). Day 3 necropsy was performed on 2 animals per cohort, with recovery necropsy on Day 22 in the remaining 2 animals. All animals survived until scheduled necropsy with limited body weight loss. There were no test-article related findings on gross pathology. Most notable clinical pathology findings were transaminase elevations (primarily AST), and decreased platelet counts at Day 8. One high dose animal demonstrated laboratory findings consistent with hepatic injury with full recovery by Day 22 and no findings on gross or microscopic pathology at necropsy. There was no evidence of myelosuppression. Microscopic pathology findings were limited, with no test-article related findings in HER2-expressing organs including heart, lungs and GI tract. All doses were considered well-tolerated. Toxicokinetics demonstrated good stability of drug conjugate in plasma with t1/2 ∼5 days (comparable to antibody t1/2) and minimal exposure to free payload. Plasma exposure at the 2.68 mg/kg dose in cyno was several fold higher than mouse exposure at the minimally efficacious dose of 0.67 mg/kg, and was comparable to the mouse exposure associated with prolonged tumor free survival after a single 2mg/kg dose. Trastuzumab-dolaflexin, a highly potent Fleximer-based ADC, demonstrates a favorable TI in tumor models with low HER2 expression where current HER2-directed therapies are inactive. Citation Format: Natalya Bodyak, Alex Yurkovetskiy, Peter U. Park, Dmitry R. Gumerov, Michael DeVit, Mao Yin, Joshua D. Thomas, LiuLiang Qin, Timothy B. Lowinger, Donald A. Bergstrom. Trastuzumab-dolaflexin, a highly potent Fleximer-based antibody-drug conjugate, demonstrates a favorable therapeutic index in exploratory toxicology studies in multiple species. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 641. doi:10.1158/1538-7445.AM2015-641

Abstract 2645: Advantages of polyacetal polymer-based ADCs: Application to low expression targets

Proceedings: AACR Annual Meeting 2014; April 5-9, 2014; San Diego, CA The application of biodegradable polymers to antibody-drug conjugate (ADC) design can provide numerous advantages, including significantly higher drug-antibody ratios, the use of alternative payloads with potencies considered insufficient for direct conjugation, the improvement of ADC physico-chemical properties, especially for highly hydrophobic payloads, and the potential expansion of protein recognition scaffolds beyond the commonly used IgGs. The basis of the novel polymer-based conjugation system described herein is a hydrophilic, fully biodegradable polyacetal carrier, (poly(1-hydroxymethylethylene hydroxymethylformal) or PHF) modified with chemically orthogonal linkers. A bioconjugation linker is used for efficient covalent attachment of a targeting moiety to the PHF scaffold, while a second, chemically distinct linker is used to attach multiple copies of a drug payload to the polymer to control the mechanism and rate of drug release. Utilizing multiple copies of a proprietary dolastatin derivative chemically conjugated to PHF, we have developed a potent and effective drug conjugation platform for ADC application, which has been named Dolaflexin™. Here, we report the preparation and characterization of a novel trastuzumab DolaflexinTM ADC, employing a maleimide-based bioconjugation approach. The resulting ADC, with a drug-antibody ratio of 20, exhibits enhanced stability and improved pharmacokinetics, with a prolonged plasma half-life and tumor-specific accumulation. Active drug release and accumulation in tumor tissue was also confirmed by LC/MS/MS methods. The activity of this novel trastuzumab-dolaflexin ADC was evaluated in multiple tumor xenograft models with significant variations in target antigen expression levels. Models including BT474 breast cancer, NCI-N87 gastric cancer, and JIMT1 breast cancer models were utilized, and comparisons to a variety of controls and ADC reference standards were made. Significant advantages of the polyacetal polymer-based ADCs in comparison to conventional ADCs, particularly in models with low target antigen expression, were observed. Details of these studies and potential applications for the development of new ADC therapeutics based on this approach will be presented. Citation Format: Alex Yurkovetskiy, Natalya Bodyak, Mao Yin, Joshua D. Thomas, Patrick Conlon, Cheri A. Stevenson, Alex Uttard, LiuLiang Qin, Dmitry R. Gumerov, Elena Ter-Ovaneysan, Venu R. Gurijala, Dennis McGillicuddy, Roberta E. Glynn, Michael DeVit, Laura L. Poling, Peter U. Park, Timothy B. Lowinger. Advantages of polyacetal polymer-based ADCs: Application to low expression targets. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2645. doi:10.1158/1538-7445.AM2014-2645

Abstract C238: Polyacetal polymer-based anti-HER2 antibody-drug conjugate employing cysteine bioconjugation through thioether linkage allows a high drug loading of dolastatin-derived payload with excellent pharmacokinetics and potent anti-tumor activity.

The application of biodegradable, polyacetal polymers to antibody-drug conjugate (ADC) design can provide numerous advantages, including significantly higher capacity for drug payload (∼20 drugs per antibody), the use of payloads with low potency that are not suitable for direct conjugation, the improvement of physicochemical properties for ADC, especially with highly hydrophobic payloads and the use of protein recognition scaffolds beyond the commonly used IgGs. The basis of this novel polyacetal polymer-based conjugation system is a hydrophilic, fully biodegradable polyacetal carrier (PHF or poly(1-hydroxymethylethylene hydroxymethylformal, or Fleximer®) modified with chemically orthogonal linkers. One linker is used to covalently attach a targeting moiety (mAb or alternative), while a second, chemically distinct linker is used to attach a drug payload and to control the mechanism and rate of drug release. Previously, we have reported potent anti-tumor activity with trastuzumab-s-Dolaflexin™, an anti-HER2 ADC composed of trastuzumab and a proprietary dolastatin derivative coupled to a Fleximer scaffold (Dolaflexin™). In that example, Dolaflexin was conjugated to the antibody through interchain cysteine residues via a hindered disulfide linkage (AACR Annual Meeting 2013 Abstract #4331). Unlike direct drug-cysteine linked ADCs that can result in destabilization of antibody by disruption of interchain disulfide bridges, we have shown that Dolaflexin conjugation via cysteines in the antibody hinge region stabilizes the resulting ADCs through the formation of inter-chain bridge structures. Trastuzumab-s-Dolaflexin ADC exhibited a prolonged plasma half-life, tumor-specific accumulation, and potent anti-tumor activity in vivo. Here, we report a novel trastuzumab-m-Dolaflexin ADC using a maleimide linker that further enhances the pharmacokinetics of the ADC and demonstrates complete regressions of established HER2+ BT-474 xenograft tumors in SCID mice even at a single dose of 2.5 mg/kg. Citation Information: Mol Cancer Ther 2013;12(11 Suppl):C238. Citation Format: Joshua Thomas, Alex Yurkovetskiy, Natalya Bodyak, Mao Yin, Patrick Conlon, Cheri Stevenson, Alex Uttard, Liu Qin, Dmitry Gumerov, Elena Ter-Ovaneysan, Michael DeVit, Laura L. Poling, Peter U. Park, Timothy B. Lowinger. Polyacetal polymer-based anti-HER2 antibody-drug conjugate employing cysteine bioconjugation through thioether linkage allows a high drug loading of dolastatin-derived payload with excellent pharmacokinetics and potent anti-tumor activity. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2013 Oct 19-23; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2013;12(11 Suppl):Abstract nr C238.

Delivering More Payload (High DAR ADCs)

Antibody drug conjugates (ADCs) for oncology applications are chemotherapeutic agents designed to selectively deliver cytotoxic drug payloads to neoplastic tissue. This book chapter reviews the latest approaches for high drug loaded ADCs. The primary focus of this review is related to ADC drug payload and antibody-drug bioconjugation linker selection strategies resulting in biotherapeutics with improved physicochemical properties, efficacy, and pharmacokinetics. A separate section of this chapter gives a brief overview of antibody targeted nanotherapeutics, a growing and diverse class of anti-cancer agents specifically designed for delivery of significant amounts of drug payload. New strategies to design the highly potent antibody targeted agents discussed in this chapter provide the opportunity to expand the list of drug payloads suitable for ADC applications and introduce agents with new mechanisms of action, which in turn may potentially lead to improvement in therapeutic index of the ADCs for the treatment of cancer.

Abstract 2670: Combination of anti-HER2 ADC XMT-1522 and checkpoint inhibitor pembrolizumab for treatment of NSCLC in preclinical models

The combination of antibody-drug conjugates (ADCs) and immunomodulatory cancer therapies is emerging as a powerful strategy for cancer treatment. Tumor-targeted delivery of a cytotoxic payload capable of inducing immunogenic cell death (ICD) can trigger both an innate and an adaptive immune response, whereby increased recruitment of effector T-cells to the tumor and formation of tumor specific immunological memory can result in a durable treatment response. The ADC XMT-1522 consists of a novel human IgG1 anti-HER2 monoclonal antibody and a novel, auristatin-based cytotoxic payload (Auristatin F-hydroxypropylamide, AF-HPA). An average DAR of 12 AF-HPA molecules is achieved via a biodegradable polymer conjugation platform. We have characterized the ability of both the free payload AF-HPA and the ADC XMT-1522 to induce ICD in vitro in multiple cell lines (NCI-N87, HT-29, SKBR3), as measured by the cell surface expression of the ICD marker calreticulin (CRT) by microscopy and flow cytometry (FACS). CRT, usually contained in the lumen of the endoplasmic reticulum, translocated to the cell surface within a few hours after treatment with AF-HPA or XMT-1522. XMT-1522 as a single agent induced significant tumor regressions in two patient-derived xenograft (PDX) models of HER2-espressing non-small cell lung cancer (NSCLC) at a dose of 3 mg/kg weekly for 3 doses. The combination of XMT-1522 in combination with the checkpoint inhibitor pembrolizumab was tested in one of these patient-derived HER2-expressing PDX models in a mouse with a humanized immune system. Expression of huPD-L1 in the tumor was confirmed by FACS and immunohistochemistry (IHC). Lymphocyte sub-populations were quantified in whole blood and in tumor by FACS and IHC. XMT-1522 treatment alone induced tumor growth delay after 3 weekly doses of 1 mg/kg. Pembrolizumab as a single agent administered every 5 days for 6 doses (q5dx6) at a dose of 2.5 mg/kg led to less tumor growth delay than XMT-1522 treatment. The combination of these two treatment regimens resulted in a better response than either of the two monotherapies. These data provide a rationale for XMT-1522 to be tested clinically as a single agent in HER2-expressing NSCLC, as well as a rationale for combination of XMT-1522 and immunomodulatory therapies in NSCLC. Citation Format: Natalya Bodyak, Marina Protopopova, Qingxiu Zhang, Alex Yurkovetskiy, Mao Yin, LiuLiang Qin, Laura L. Poling, Rebecca Mosher, Donald A. Bergstrom, Timothy B. Lowinger. Combination of anti-HER2 ADC XMT-1522 and checkpoint inhibitor pembrolizumab for treatment of NSCLC in preclinical models [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 2670. doi:10.1158/1538-7445.AM2017-2670