Genetically encoded self-assembling affibody-streptavidin nanoparticles for HER2-positive cancer theranostics.

Ultrasound-Activated Cascade Effect for Synergistic Orthotopic Pancreatic Cancer Therapy.

Antibody-drug conjugates and bicycle toxin conjugates represent a tremendous advance in drug delivery technology and have shown great promise in the treatment of urothelial cancer. Previously approved systemic therapies, including chemotherapy and immunotherapy, are often impractical due to comorbidities, and outcomes for patients with advanced disease remain poor, even when receiving systemic therapy. In this setting, antibody-drug and bicycle toxin conjugates have emerged as novel treatments, dramatically altering the therapeutic landscape. These drugs harness unique designs consisting of antibody or bicycle peptide, linker, and cytotoxic payload with more targeted delivery than conventional chemotherapy, thus eliminating malignant cells while reducing systemic toxicities. Potential targets investigated in urothelial cancer include Nectin-4, TROP2, HER2, and EphA2. Initial clinical trials demonstrated efficacy in treatment of refractory advanced urothelial cancer, as well as improvement in quality of life. These initial studies led to FDA approval of two antibody-drug conjugates, enfortumab vedotin and sacituzumab govitecan. Moreover, antibody-drug and bicycle toxin conjugates are being studied in ongoing clinical trials in frontline treatment of advanced disease as well as for localized cancer. These studies highlight the potential for additional future therapies with novel targets, novel antibodies, cytotoxic and immunomodulatory payloads, and unique structural designs enhancing efficacy and safety. There is increasing evidence that combinations with other cancer therapies, especially immunotherapy, improve treatment outcomes. The combination of enfortumab vedotin and pembrolizumab was recently approved for first-line treatment of advanced urothelial carcinoma. Despite the great promise of these novel drugs, robust predictive biomarkers are needed to determine the patients who would maximally benefit. This review surveys the rationale and current state of the evidence for these new drugs and describes future directions actively being explored.

Background: Cervical cancer ranks fourth in incidence and mortality among female tumors, with over 0.6 million patients diagnosed worldwide in 2020. A large percentage of cervical cancer patients will develop advanced diseases (stage IV or recurrent disease) and the 5-year survival rate of these patients was only 15-20% due to the lack of efficacy and adverse effects of first line chemodrugs. Recently, antibody drug conjugates (ADCs) emerge as a novel class of targeted therapeutics that selectively ablate tumor cells without damaging normal organs and tissues, which has shown promising efficacy in treating solid tumors. In this study, we sought to develop a cervical cancer-targeted ADC using a novel molecular target. Methods: Firstly, human cervical cancer and paracancerous tissue were stained by immunohistochemistry (IHC) in order to determine the difference of the expression level of a molecular target. Three human cervical cancer cell lines (C-33A, Si-Ha and Ca-Ski) and one normal cervical cell line (HCerEpiC) were used to determine the subcellular location of an ADC target on the cells by immunofluorescence (IF) staining, flow cytometry and single photon confocal microscopy. Secondly, endocytosis capability of target antibodies was visualized by confocal imaging and was quantified by flow cytometric analysis. Furthermore, a panel of ADC formulations with different chemical linkers and payloads were designed, constructed and characterized as ADC candidates for cervical cancer therapy. We next evaluated in vitro inhibitory activity of these ADC candidates on three cervical cancer cell lines by CCK8 assay. Finally, orthotopic and lung metastasis animal models of cervical cancer were established to determine the in vivo inhibitory activity and biosafety of constructed ADC candidates. Results: IHC staining of clinical specimens showed that the expression level of CD54 in cervical tumor tissues was significantly higher than those of paracancerous tissues. Fluorescence imaging and flow cytometry further determined the overexpression level, plasma membrane location and antigen-mediated endocytosis activity of CD54 in multiple human cervical cancer cell lines. Among several ADC formulations, an optimized CD54-targeted ADC linker and warhead combination was identified by comparing its IC50 with other ADC candidates and standard-of-care chemodrugs for cervical cancer. The optimized CD54 ADC exhibited promising anti-tumor activity on primary and lung metastatic cervical tumor models. Conclusion: In our study, we identified CD54 as a novel ADC target for cervical cancer. An optimized CD54-targeted ADC formulation was determined and its anti-tumor activity and biosafety profile were determined in multiple human cervical cancer cell lines in vitro and orthotopic and lung metastatic cervical tumor models in vivo, providing a promising targeted therapeutic candidate for the targeted treatment of cervical cancer. Citation Format: Shili Yao, Bing Zhu, Xinyan Wang, Rui Xu, Tong Yang, Peng Guo, Huarong Tang, Tao Zhu. Developing a novel antibody drug conjugate against cervical cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 6174.

BackgroundTrastuzumab emtansine (T-DM1) is an antibody-drug conjugate that carries a cytotoxic drug (DM1) to HER2-positive cancer. The target of T-DM1 (HER2) is present also on cancer-derived exosomes. We hypothesized that exosome-bound T-DM1 may contribute to the activity of T-DM1.MethodsExosomes were isolated from the cell culture medium of HER2-positive SKBR-3 and EFM-192A breast cancer cells, HER2-positive SNU-216 gastric cancer cells, and HER2-negative MCF-7 breast cancer cells by serial centrifugations including two ultracentrifugations, and treated with T-DM1. T-DM1 not bound to exosomes was removed using HER2-coated magnetic beads. Exosome samples were analyzed by electron microscopy, flow cytometry and Western blotting. Binding of T-DM1-containing exosomes to cancer cells and T-DM1 internalization were investigated with confocal microscopy. Effects of T-DM1-containg exosomes on cancer cells were investigated with the AlamarBlue cell proliferation assay and the Caspase-Glo 3/7 caspase activation assay.ResultsT-DM1 binds to exosomes derived from HER2-positive cancer cells, but not to exosomes derived from HER2-negative MCF-7 cells. HER2-positive SKBR-3 cells accumulated T-DM1 after being treated with T-DM1-containg exosomes, and treatment of SKBR-3 and EFM-192A cells with T-DM1-containing exosomes resulted in growth inhibition and activation of caspases 3 and/or 7.ConclusionT-DM1 binds to exosomes derived from HER2-positive cancer cells, and T-DM1 may be carried to other cancer cells via exosomes leading to reduced viability of the recipient cells. The results suggest a new mechanism of action for T-DM1, mediated by exosomes derived from HER2-positive cancer.

IKS04 is an antibody drug conjugate (ADC) targeting human CA242, also known as CanAg, a tumor-associated carbohydrate antigen. This glycotope is highly expressed in most colorectal, pancreatic, gallbladder, and gastric cancers, with limited expression in normal tissues. While prior experience demonstrated that CA242 can be safely targeted with an ADC, the limited clinical efficacy observed with microtubule inhibitor payloads in gastrointestinal indications suggests that alternate payloads such as DNA crosslinkers could provide an improved therapeutic to target CA242+ tumors more effectively. The ADC was generated by site-specific bioconjugation of a pyrrolobenzodiazepine (PBD) prodrug payload to the antibody light chain at a drug-to-antibody ratio of 2. The linker-prodrug incorporates two glucuronide-trigger moieties that require lysosomal cleavage by beta-glucuronidase to both release and activate the PBD dimer payload. In vitro antiproliferative and bystander activity was evaluated by CellTiterGlo assay. In vivo efficacy studies for ADC alone and antibody co-administration were performed with subcutaneous human xenografts in CB17-SCID mice. CA242 expression was determined in formalin fixed tumor samples by immunohistochemistry and on cell lines by quantitative flow cytometry. In vitro, IKS04 results in potent and specific cell killing of panel of CA242+ target cells and potency correlated with antigen expression determined by flow cytometry. The PBD payload confers strong bystander activity in co-culture studies in vitro, an attribute that may allow treating tumors with heterogeneous antigen expression. The ADC was active against gastric and colorectal cancer xenograft models at single doses of 0.5 to 1 mg/kg, while the maytansinoid-based ADC benchmark cantuzumab ravtansine was inactive even at 2 mg/kg. Contrary to in vitro results, we observed a trend towards greater in vivo activity in models with lower antigen expression. This is suggestive of an antigen barrier effect where high target expression results in ADC accumulation at the tumor surface, precluding deeper penetration into tumor tissue. As the maximum tolerated ADC dose can be limited by target-independent payload toxicities, we increased the overall in vivo dose by addition of antibody to improve delivery. Consistent with this concept, co-administration of the unconjugated antibody with the ADC at a 5 to 50-fold excess improved the minimally effective dose in the CA242-high SNU16 gastric and HT29 colon cancer models compared to ADC alone while the antibody itself had no anti-tumor activity. IKS04 is an ADC with a DNA crosslinker payload that shows potent in vitro and in vivo efficacy against gastric and colorectal cancer tumor models. Co-administration of the unconjugated antibody can improve ADC delivery which drives increased efficacy in preclinical models without increasing toxicity. Citation Format: Jutta Deckert, Justyna Mysliwy, Adam Lodge, James Stephenson, Sarah Robinson, Robert J. Lutz. IKS04, an antibody drug conjugate with a highly potent DNA crosslinker payload for the treatment of gastrointestinal cancers [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 2885.

Background: Interferon beta (IFN-β), a promising potent cytokine, has been attracting attention for treatment of cancer. The pleiotropic antitumor effects of IFN-β have been studied, with specific reference to their direct role on cancer cells and indirect action through the immune effector cells. However, its systemic toxicities and poor biophysical properties prevent IFN-β from being widely used for cancer therapy. Since the potential of cytokine therapies are impaired due to dose-limiting systemic toxicities, novel treatment methods should be developed to safely deliver effective drug quantities at the tumor sites. Like antibody-drug conjugates (ADCs), immunocytokine can be attempted to induce cytokine's organ-targeting and alleviate its systemic side effects. Here, we designed recombinant IFN-β mutein immunocytokines that comprise a HER2-targeting antibody and IFN-β mutein, and evaluate the antitumor properties against HER2-positive cancers.Method: A panel of human gastric cancer cell lines was treated with trastuzumab-IFN-β mutein to evaluate direct antitumor effect. In addition, to test the immune cell-mediated antitumor effect, cancer cells were co-cultured with effector cells (e.g. PBMC) in the absence or drug. The antitumor efficacy of trastuzumab-IFN-β mutein in vivo was tested in HER2-positive cancer xenograft models using nude mice or humanized mice. Result: Trastuzumab-IFN-β mutein directly inhibited the growth of HER2-positive gastric cancer cell lines and was more effective than trastuzumab or IFN-β mutein alone. Trastuzumab-IFN-β mutein also displayed enhanced immune cell-mediated cytotoxicity. Collectively, trastuzumab-IFN-β mutein may have indirect immune cell-mediated antitumor effects and direct cell growth inhibitory effects. Moreover, trastuzumab-IFN-β mutein significantly suppress tumor growth in HER2-positive cancer xenograft models. Tumor-infiltration of lymphocytes was enhanced by trastuzumab-IFN-β mutein, implying that the tumor-targeting IFN-β may have an antitumor effect through increased immune response. Conclusion: We have characterized and evaluated the antitumor properties of trastuzumab-IFN-β mutein in HER2-expressing cancers. Since IFN-β activates antitumor immune responses, it is expected to be administered in combination with immunotherapeutic drugs. Therefore, the study suggests that trastuzumab-IFN-β mutein is a promising candidate for the treatment of HER2-positive carcinoma. Citation Format: Chan Gyu Lee, Tae Eun Kim, Sungyoul Hong, Jongwan Chu, Ju Eun Kang, Hae Min Jeong, Young Kee Shin. HER2-targeted interferon-beta-1a mutein, a potent immunocytokine for the treatment of HER2-positive cancers [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr LB170.

Implementation of antibody-drug conjugates in HER2-positive solid cancers: Recent advances and future directions

As a rare malignancy, cholangiocarcinoma (CCA) is a lethal disease with a 5-year overall survival rate of merely 5%. Highly invasive and asymptomatic characteristics of CCA cause most patients diagnosed at advanced stages, severely deteriorating their clinical outcomes. The drug treatment options for CCA are limited and surgical resection of CCA frequently suffers from a high recurrence rate of 66%. Albeit Pemigatinib, a FGFR2 inhibitor, has recently been approved by FDA as the first CCA-targeted therapeutic, only less than 10% of patients with CCA carry FGFR2 fusion mutations that can benefit from this drug. Ivosidenib, another targeted drug against IDH1 mutation, only works for 13% CCA patients carrying this mutation. Therefore, discovering new molecular targets and developing associated targeted drugs remain a significant and unmet medical need in CCA therapy. In this study, we first screened potential molecular targets for CCA by comparing the expression levels of a panel of cancer-related cell membrane antigens between human CCA cells and normal human intrahepatic biliary epithelial cells through flow cytometry. CD54 was identified as a remarkable molecular target of CCA with strong binding force to its antibodies and high endocytosis efficiency by flow cytometry and confocal fluorescent microscopy. Meanwhile, we confirmed that CD54 was absent in most normal human tissues but was highly upregulated in CCA tumor tissues through immunohistochemical (IHC) staining. Based on this newly discovered CCA target, we constructed a series of CD54 antibody-drug conjugates (ADCs) using different chemical linkers and payloads. We next evaluated and compared the half maximum inhibitory concentrations (IC50s) of each ADC in ablating human CCA cells in vitro in order to identify an optimized ADC formulation. Furthermore, we validated its anti-tumor efficacy against primary and peritoneal metastatic CCA tumors in vivo in comparison with conventional chemotherapy. Eventually, we determined the biodistribution and safety profile of this CD54 ADC in order to ensure that it is well-tolerated under the effective dose. In general, we identified CD54 as a novel molecular target for CCA through antibody library screening, and explored its potential in the development of CCA-targeted ADCs, providing a promising targeted therapeutic for clinical treatment of CCA. Citation Format: Bing Zhu, Shi-Li Yao, Xin-Yan Wang, Rui Xu, Peng Guo, Jie-Er Ying. Developing antibody-drug conjugates against cholangiocarcinoma using a novel molecular target [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 6175.

Antibody–drug conjugates (ADCs) take the advantage of monoclonal antibodies to selectively deliver highly potent cytotoxic drugs to tumor cells, which have become a powerful measure for cancer treatment in recent years. To develop a more effective therapy for human epidermal growth factor receptor 2 (HER2)-positive cancer, we explored a novel ADCs composed of anti-HER2 scFv–HSA fusion antibodies conjugates with a potent cytotoxic drug DM1. The resulting ADCs, T-SA1–DM1 and T-SA2–DM1 (drug-to-antibody ratio in the range of 3.2–3.5) displayed efficient inhibition in the growth of HER2-positive tumor cell lines and the half-maximal inhibitory concentration on SKBR-3 and SKOV3 cells were both at the nanomolar levels in vitro. In HER2-positive human ovarian cancer xenograft models, T-SA1–DM1 and T-SA2–DM1 also showed remarkable antitumor activity. Importantly, three out of six mice exhibited complete remission without regrowth in the high-dose group of T-SA1–DM1. On the basis of the analysis of luminescence imaging, anti-HER2 scFv–HSA fusion antibodies, especially T-SA1, showed strong and rapid tumor tissue penetrability and distribution compared with trastuzumab. Collectively, the novel type of ADCs is effective and selective targeting to HER2-positive cancer, and may be a promising antitumor drug candidate for further studies.

Molecular Templates develops highly potent, specific, next-generation immunotoxins that are proprietarily de-immunized to avoid both innate and adaptive immune recognition. These Engineered Toxin Bodies (ETBs) destroy cancer cells by enzymatic destruction of ribosomes, a mechanism of action (MOA) distinct from that of other therapeutics. Molecular Templates' lead compound, MT-3724, has seen clinical activity in heavily pre-treated lymphoma patients, underscoring the potential for this class of agents to work in the refractory/relapsed setting. Current modalities for the targeted treatment of HER2 positive breast cancer include monoclonal antibodies, antibody-drug conjugates, and tyrosine kinase inhibitors. Although all these modalities have demonstrated clinical benefit, the majority of patients will unfortunately relapse due to a variety of resistance mechanisms. Upon relapse, most of these patients will still express HER2, allowing for the development of new HER2-targeted modalities. MT-5111, a HER2-targeted ETB with picomolar potency against HER2 expressing cells, was designed to overcome mechanisms of resistance to current HER2 modalities such as: escape from antibody dependent cell-mediated cytotoxicity (ADCC), alterations in signal transduction, epitope masking, and enhanced small molecule efflux. Additionally, MT-5111 was genetically engineered to reduce the anti-drug antibody response and signalling through innate receptors, allowing for repeat dosing. MT-5111 binds to an epitope on HER2 distinct from trastuzumab and ado-trastuzumab emtansine (T-DM1), and exhibits effective cytotoxicity on both T-DM1 sensitive and T-DM1 resistant HER2 positive cell lines. The targeting of MT-5111 to a distinct epitope allows for activity of the ETB on cell lines where the trastuzumab epitope is masked, and allows for combination with targeted agents trastuzumab or T-DM1 without competition for binding to HER2. Previous reports have shown that resistance to T-DM1 may be due to increased drug efflux; however, the MT-5111 cytotoxic payload is a large molecule not subject to this mechanism of resistance. Additionally, the novel MOA for MT-5111 should allow for cell kill independent of changes to the tumor microenvironment or alterations in HER2-mediated signal transduction. Pre-clinical data demonstrate the potent activity of MT-5111, including in T-DM1-resistant cell lines, as well as the potential for co-administering MT-5111 and trastuzumab or T-DM1, allowing for concurrent treatment of two HER2-targeted agents with distinct and non-interfering mechanisms of action. In vitro and in vivo data will be presented, highlighting the potential for MT-5111 as a novel agent under development for treatment of breast carcinomas, and other malignancies overexpressing the HER2 receptor. Molecular Templates intends to initiate clinical studies with MT-5111 in 2018. Citation Format: Brigitte Brieschke, Garrett L. Robinson, Sangeetha Rajagopalan, Hilario J. Ramos, Jensing Liu, Jack P. Higgins, Erin K. Willert. Targeted Engineered Toxin Bodies provide a novel mechanism of action against HER2 positive cancers [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 5769.

Arming Antibodies for Cancer Therapy: Transglutaminase-Mediated Toxin Conjugation

HER2 overexpression is associated with aggressive and poor patient outcome. HER2 has become a crucial target in cancer treatment, and the discovery of effective HER2-targeted therapies marked a significant milestone in treating HER2-positive cancers. This led to the approval of trastuzumab, the first HER2-targeted monoclonal antibody. Later, trastuzumab was used to develop antibody-drug conjugates (ADCs) for breast cancer, which have shown promising results. ADCs are combined trastuzumab with a cytotoxic drug to improve effectiveness while reducing side effects. Two ADCs, trastuzumab emtansine (T-DM1) and trastuzumab deruxtecan (T-DXd), have been approved by the FDA for treating HER2-positive breast cancer. However, drug resistance has become a serious issue, reducing the long-term success of these treatments. This review explores key mechanisms of ADCs resistance including alteration in HER2 expression, antibody and payload-related resistance, altered cell signaling, impaired lysosomal and intracellular activity, and tumor microenvironment. By analyzing recent studies in ADCs resistance, this review provides an insight into ADC resistance mechanisms and potential strategies for improving therapeutic outcomes HER2 positive breast cancer.

Chemotherapy and radiotherapy have been considered the first line of antitumor therapies for patients with advanced or metastatic cancer, however these therapies demonstrate numerous drawbacks and acquired resistance is still inevitable in most cases. In order to address these challenges, scientists have discovered a novel class of cancer-targeted drugs referred to as antibody-drug conjugates (ADCs). ADCs are an emerging class of therapies that conjugate monoclonal antibodies with cytotoxic payloads through a chemical linker, allowing precise delivery of drugs to cancer cells. Trastuzumab deruxtecan (T-DXd) is an ADC composed of a humanized IgG antibody that specifically targets HER2, a tetrapeptide-based cleavable linker and a topoisomerase I inhibitor payload. T-DXd is the second HER2-targeting ADC approved by the FDA after trastuzumab emtansine (T-DM1); approved for HER2-positive unresectable or metastatic breast cancer in the second line and as an adjuvant therapy for residual disease in breast cancer patients with early stage disease. Both ADCs have demonstrated promising clinical efficacy in the treatment of HER2-positive solid cancers, which has sparked great interest in the development of the next generation of ADCs. Emerging evidence indicates that the efficacy of a given ADC depends on a number of key factors including the fine tuning between the combination of the antibody, linker and payload components, how the ADC is processed and it’s interaction with the cancer cells. Here we present robust and reproducible data from our comprehensive modular in vitro and in vivo screening platform, that has been developed to facilitate efficient and streamlined approaches for ADC screening and characterization, and have been validated using T-DXd and T-DM1 as proof-of-concept agents. Our platform offers assays that can evaluate ADC binding, internalization, anti-proliferative activity, ADC effects on cell cycle, apoptosis, and bystander effects. Immunological assays, such as antibody-dependent cellular cytotoxicity (ADCC), and antibody-dependent cellular phagocytosis (ADCP) can be employed to assess the induction of Fc effector functions. Additionally, we have validated a number of in vivo models including co-inoculation and intracranial xenograft models. Our data demonstrates that both ADCs exhibit strong binding and internalization, with T-DXd showing a pronounced bystander effect due to its payload release via a cleavable linker. Both agents effectively induced ADCC and ADCP, with minimal complement-dependent cytotoxicity observed. The T-DXd bystander effect was also confirmed used an in vivo co-inoculation xenograft model and efficacy of both T-DXd and T-DM1 against a human intracranial xenograft model was confirmed. This platform provides a comprehensive and customizable tool for ADC preclinical assessment, supporting the optimization of ADC therapeutic efficacy and safety. Citation Format: Zichun Wang, Qiong Wang, Shuyue Wang, Guilan Wang, Qikuan Chen, Wei Xue, Yinfei Yin. A customizable, end-to-end platform for the screening and characterization of ADCs [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 5500.

Background: Gastrointestinal cancers (GI) are among the most common cancers worldwide. Current treatment options rely on a chemotherapy backbone combined with targeted antibody therapies, but the prognosis generally remains poor. Iksuda has developed a next-generation Antibody Drug Conjugate (ADC) targeting a cancer-specific carbohydrate antigen (CanAg) which is highly overexpressed in GI cancers but has no expression on normal tissues. This CanAg-targeting ADC is comprised of a proprietary PBD prodrug coupled to an anti-CanAg antibody using PermaLink conjugation technology. The PBD prodrug is designed with a tumor-selective trigger to attenuate its potency before entry of the ADC into the target cancer cells. PermaLink conjugation chemistry ensures conjugate stability. Here, the potential of this anti-CanAg ADC was explored in preclinical models as a proof of concept for targeting GI cancers. Methods: The ADC is composed of a PBD prodrug conjugated to an anti-CanAg antibody through PermaLink to generate conjugate with a drug to antibody ratio of approximately 2. This CanAg-targeting ADC was evaluated in vitro in the CanAg-expressing Colo205 cancer cell line. In vivo efficacy studies were conducted in SCID mice with CanAg-expressing human cancer xenografts representing colorectal (Colo 205), gastric (N87) and pancreatic cancers (BxPC3). An exploratory toxicology study was conducted in cynomolgus monkeys. Results: The CanAg-targeting ADC showed highly potent, specific activity in the Colo205 colorectal cancer cell line with IC50 values in the pM range. A single administration of anti-CanAg ADC in CanAg-positive GI xenograft models showed significant dose-dependent antitumor activity and induced complete tumor regressions with no observable toxicity. In the gastric cancer model, the anti-CanAg ADC gave a complete response by day 36 when dosed at 0.3 mg/kg, and 1 and 3 mg/kg doses exhibited long-lasting tumor regression. In the pancreatic cancer model, administration of anti-CanAg ADC at 0.3 mg/kg induced 76% tumor growth inhibition (TGI) on day 21, and at 1 mg/kg, and 3 mg/kg showed complete tumor regression by day 40 with no subsequent tumor recurrence for the 3mg/kg dose. In colorectal cancer, the anti-CanAg ADC exhibited 58% and 98% TGI with 0.3 mg/kg and 1 mg/kg doses respectively, and with the 3 mg/kg dose regressed tumor with no recurrence. The anti-CanAg ADC was well tolerated in cynomolgus monkeys with 1 mg/kg defined as the highest non-severely toxic dose (HNSTD). Conclusions: This anti-CanAg ADC has shown favorable preclinical anti-tumor activity in gastrointestinal cancers. Our findings support the clinical development of the anti-CanAg ADC for the treatment of GI cancers. Citation Format: Justyna H. Mysliwy, Adam Lodge, Daniel Williamson, Jenny Thirlway, Jutta Deckert, Robert J. Lutz. Developing a next generation antibody drug conjugate for treatment of gastrointestinal cancers [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1751.

<div>Abstract<p>A linker-drug platform was built on the basis of a cleavable linker-duocarmycin payload for the development of new-generation antibody–drug conjugates (ADC). A leading ADC originating from that platform is SYD983, a HER2-targeting ADC based on trastuzumab. HER2-binding, antibody-dependent cell-mediated cytotoxicity and HER2-mediated internalization are similar for SYD983 as compared with trastuzumab. HER2-expressing cells <i>in vitro</i> are very potently killed by SYD983, but SYD983 is inactive in cells that do not express HER2. SYD983 dose dependently reduces tumor growth in a BT-474 mouse xenograft <i>in vivo</i>. The ADC is stable in human and cynomolgus monkey plasma <i>in vitro</i> but shows relatively poor stability in mouse plasma due to mouse-specific carboxylesterase. SYD983 could be dosed up to 30 mg/kg in cynomolgus monkeys with high exposure, excellent stability in blood, and without severe toxic effects. The monkey safety study showed no SYD983-induced thrombocytopenia and no induction of peripheral sensory neuropathy, both commonly observed in trials and studies with ADCs based on tubulin inhibitors. Finally, to improve homogeneity, SYD983 was further purified by hydrophobic interaction chromatography resulting in an ADC (designated SYD985) predominantly containing DAR2 and DAR4 species. SYD985 showed high antitumor activity in two patient-derived xenograft models of HER2-positive metastatic breast cancers. In conclusion, the data obtained indicate great potential for this new HER2-targeting ADC to become an effective drug for patients with HER2-positive cancers with a favorable safety profile. More generally, this new-generation duocarmycin-based linker-drug technology could be used with other mAbs to serve more indications in oncology. <i>Mol Cancer Ther; 13(11); 2618–29. ©2014 AACR</i>.</p></div>