Datopotamab deruxtecan: rational strategies, novel advancements, challenges, and future perspectives.

TPS3162 Background: c-Met (MET protein) is commonly overexpressed in a number of tumors, including hepatocellular carcinoma (HCC), pancreatic ductal adenocarcinoma (PDAC), biliary tract cancers (BTC), esophageal squamous cell carcinoma (ESCC), breast cancer (BC), and head and neck squamous cell carcinoma (HNSCC). Patients with c-Met–overexpressing tumors represent an underserved population, with the need for more effective c-Met–targeting therapies to become available. ABBV-400 is an antibody-drug conjugate, consisting of the c-Met–targeting antibody telisotuzumab conjugated to a potent topoisomerase 1 inhibitor payload. Initial results from the ongoing first-in-human study (NCT05029882) of ABBV-400 in patients with advanced solid tumors indicate a tolerable safety profile, with a maximum tolerated dose of 3 mg/kg once every 3 weeks (Q3W), and promising antitumor activity, with an overall response rate of 24.4% (1). Herein, we describe a signal-seeking study evaluating ABBV-400 treatment in patients with select solid tumors. Methods: Multicenter, open-label, phase 1 signal-seeking study(NCT06084481). Eligible patients (≥18 years) have confirmed locally advanced/metastatic disease measurable per RECIST v1.1 and Eastern Cooperative Oncology Group performance status ≤1. Approximately 220 patients are planned for enrollment across 7 cohorts (HCC, n=40; PDAC, n=40; BTC, n=20; ESCC, n=40; triple-negative BC, n=20; hormone receptor-positive/HER2-negative BC, n=20; HNSCC, n=40). The primary objectives are to assess efficacy and safety/tolerability of ABBV-400 in each tumor indication. Secondary objectives include the evaluation of pharmacokinetics (PK) and immunogenicity of ABBV-400. Pharmacodynamic (PD) and biomarker analyses are exploratory endpoints. Patients receive intravenous ABBV-400 at 3 mg/kg Q3W until disease progression, intolerable toxicity, or any other per-protocol discontinuation criteria. c-Met expression will be assessed retrospectively by immunohistochemistry. The maximum treatment duration is 2 years. Tumor assessments are performed at screening and every 6 weeks from the first dose of study drug, with objective response rate as primary efficacy endpoint and duration of response, clinical benefit rate, progression-free survival, and overall survival as secondary efficacy endpoints. Safety evaluations include adverse events monitoring (graded according to the NCI CTCAE, v5.0), physical examinations, vital sign measurements, ECG variables, and clinical laboratory testing. Blood samples for PK, PD, and biomarker analysis are collected at designated time points throughout the study. Enrollment started in November 2023. As of 19 January 2024, 24 patients have been enrolled. 1. Sharma et al. JCO 2023;41[16 suppl]:3015. Clinical trial information: NCT06084481 .

Background: HER3 is overexpressed in 30-50% of breast cancers and has been associated with poor prognosis. Patritumab deruxtecan (HER3-DXd; U3-1402) is a HER3-directed ADC with a potent topoisomerase I (TOP1) inhibitor payload. A phase 1/2 study of HER3-DXd (NCT02980341) demonstrated promising antitumor activity in hormone receptor positive (HR+) metastatic BC patients with clinical activity observed across baseline levels of HER3 protein or mRNA expression. A window of opportunity clinical trial is currently ongoing to evaluate the biological activity of HER3-DXd in patients with treatment naïve BC according to HER3 mRNA/protein expression levels (NCT04610528). Here, we aimed to describe the activity of HER3-DXd in PDX models to identify robust biomarkers of response. Methods: The antitumor activity of HER3-DXd was assessed in 21 BC PDX models (14 HR+ and 7 triple negative). HER3-DXd sensitivity was established as a complete response that lasted longer than 120 days, following 4 weekly doses of 10 mg/kg. HER3-DXd antitumor activity was compared to the antitumor activity of irinotecan (50 mg/kg dosed once weekly), which was evaluated according to modified RECIST criteria in a subset of 14 BC PDX models. PDX models of acquired resistance to HER3-DXd were generated by repeated treatment cycles. Pharmacodynamic (PD) experiments were conducted by collecting tumor samples from PDXs after a single dose of HER3-DXd or irinotecan. Baseline HER3 expression was assessed by immunohistochemistry (IHC) and Western blot. mRNA expression of 72 genes including ERBB3 and genes from the PAM50 signature were measured using the nCounter platform. Two-class unpaired significance analysis of microarrays (SAM), using a false-discovery rate<5%, identified differential gene expression across response groups. Genetic alterations harbored by PDX models were determined using the MSK-IMPACTTM targeted exome panel. Quantification of proliferation (% of Ki67-positive cells) and of DNA damage during the S-phase of the cell cycle (γH2AX nuclear foci in geminin-positive cells) was evaluated in untreated/treated PD samples by IHC or immunofluorescence (IF), respectively. Western blot was used to assess HER3-pathway downmodulation and induction of apoptosis. Results: Eight out of 21 (38%) PDXs were highly sensitive to HER3-DXd, and in 5/14 (36%) models HER3-DXd showed a superior antitumor activity when compared to irinotecan. We observed an enrichment of basal-like models amongst the non-relapsed PDXs, compared to the relapsed ones (6/8 (75%) vs. 3/13 (20%), p=0.0195). Baseline levels of HER3/ERBB3 were not associated with treatment response and a model of acquired-resistance did not exhibit a reduction in baseline HER3 expression. Interestingly, relapsed models showed increased expression of genes related with chemotherapy-resistance (MDM2, NAT1, MAPT, GRB7, BCL-2). Mechanistically, treatment with HER3-DXd did not reduce the level of Ki67-proliferating cells but resulted in a significantly higher induction of S-phase DNA damage measured as γH2AX nuclear foci in non-relapsed models, compared to relapsed ones. This was accompanied by a reduction of HER3 protein levels and downmodulation of pERK1/2 T202/Y204, along with activation of PARP cleavage. Conclusions: HER3-DXd exerts a potent antitumor response in BC PDXs, independently of baseline HER3/ERBB3 levels, in line with the clinical data of NCT02980341. Basal-like tumors were more sensitive to HER3-DXd than luminal B models. Mechanistically, our data suggests that treatment with HER3-DXd results in parallel HER3/ERK signaling downmodulation and induction of S-phase DNA damage, resulting in tumor cell death. Citation Format: Andreu Òdena, Laia Monserrat, Fara Brasó-Maristany, Marta Guzmán, Judit Grueso, Olga Rodríguez, Maurizio Scaltriti, Sarat Chandarlapaty, Yang Qiu, Kumiko Koyama, Mafalda Oliveira, Aleix Prat, Violeta Serra. Antitumor activity of patritumab deruxtecan (HER3-DXd), a HER3-directed antibody drug conjugate (ADC) across a diverse panel of breast cancer (BC) patient-derived xenografts (PDXs) [abstract]. In: Proceedings of the 2021 San Antonio Breast Cancer Symposium; 2021 Dec 7-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2022;82(4 Suppl):Abstract nr P5-13-14.

Datopotamab deruxtecan (Dato-DXd) is an antibody-drug conjugate (ADC) consisting of a humanized anti-TROP2 monoclonal antibody linked to a potent DNA topoisomerase I (TOP1) inhibitor payload via a plasma-stable, tumor-selective, tetrapeptide-based cleavable linker. Dato-DXd is currently under clinical investigation for the treatment of patients with solid tumors including non-small cell lung cancer (NSCLC), hormone receptor positive (HR+ BC) and triple negative breast cancer (TNBC). Pharmacotherapy of brain tumors can be limiting due to restricted drug delivery across blood brain and blood tumor barrier. Enhertu®, that uses the same DXd ADC technology, has reported clinical activity in patients with brain metastases from HER2+ breast cancer, but very little information is available on what drives ADC biodistribution and activity in CNS-involved cancers. Here, we investigated whether systemically administered ADCs can penetrate the brain microenvironment and mediate anti-tumor activity in a preclinical model. Luciferase-tagged H1373 (TROP2-expressing NSCLC) tumor cells were intracranially implanted into NSG mice. Tumor-bearing mice were dosed with Dato-DXd or matched isotype Control IgG-ADC (DAR4) at 10mpk 7 days or 14 days post intracranial tumor implant. Dato-DXd inhibited intracranial tumor growth better than Control ADC (Day 7: 105% TGI vs 38% TGI; Day 14: 65% TGI vs <10% TGI, respectively, compared to Vehicle). Immunohistochemistry analysis of brain tissue validated localization of Dato-DXd in the tumor, suggesting Dato-DXd can distribute into the local tumor microenvironment in this preclinical tumor model. Additionally, treatment with Dato-DXd provided a significant survival benefit over Control ADC (Median survival of 63 days vs 43 days, P=0.0002). This preclinical study supports the inclusion of Dato-DXd in treatment of patients with CNS-involved tumors. Understanding the pharmacological determinants of Dato-DXd activity in the CNS will help outline strategies to implement Dato-DXd-based treatment of patients with CNS-involved tumors. Citation Format: Kristen Jones Jones, Montira Suksomboon, SaraAnn rosenthal, Jacob Gordon, Corinne Reimer, Matthew Sung, Chetan Rane. Dato-DXd mediates anti-tumor activity in preclinical TROP2-expressing intracranial tumor model [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 1911.

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: Antibody-drug conjugates (ADCs) have improved survival for patients with metastatic breast cancer (MBC). In patients with HER2 negative MBC, two ADCs have FDA approval: sacituzumab govitecan (SG) and trastuzumab deruxtecan (T-DXd), both with topoisomerase-I (topo-I) inhibitor payloads. Given the many other ADCs in development, there is great interest in using ADC after ADC to maximize treatment benefit for patients. However, there is limited understanding regarding resistance mechanisms to ADCs and impact on ADC sequencing. We conducted a translational study to address this, and here we report the incidence of TOP1 mutations and clinical impact on ADC sequencing. Methods: All patients with MBC treated with ADCs at a large academic medical institution (Massachusetts General Hospital) who had comprehensive plasma-based genotyping (500 gene GuardantOMNI panel) were included. Since both SG and T-DXd have topo-I inhibitor-based payloads, we particularly focused on TOP1 mutations, variant allele frequency (VAF), and germline/somatic characterization. Incidence of TOP1 mutations in the ADC cohort was compared to The Cancer Genome Atlas (TCGA). Clinical “cross-resistance” was defined as progressive disease (PD) as best response to second ADC (ADC2) or treatment time on ADC2 of less than 60 days. Results: Based on comprehensive plasma-based genotyping we identified 4 distinct TOP1 mutations: S57C, R364H, W401C, G359E, at a frequency of 6.0% (4/67) at the time of disease progression on ADC compared to a frequency of 0.5% described in primary breast cancer in TCGA. Two of the amino acids found to be mutated are known to form direct interactions with the DNA backbone (G359) or the topoisomerase inhibitor itself (R364). For clinical resistance, one patient was briefly on ADC2 but stopped after 1 dose for toxicity; among the other 3 patients, two had cross-resistance to ADC after ADC, with both ADCs containing topo-I inhibitor payloads. Median duration on first ADC was 455 days compared to a median of 52 days for ADC2. Finally, one patient treated sequentially with 3 ADCs (all with topo-I inhibitor payloads) was found to have rising TOP1 VAF with progressive ADC treatments. Conclusion: This is the first report describing emergence of TOP1 mutations under selective pressure from ADCs and the impact on mediating cross-resistance to ADC after ADC with topo-I inhibitor payloads. Novel ADCs with alternative payloads may potentially be more effective when used sequentially after an ADC with a topo-I inhibitor. Further biomarker research is needed to optimize ADC sequencing for patients with TOP1 mutant MBC. Citation Format: Rachel Occhiogrosso Abelman, Haley Barnes, Arielle J. Medford, Annika Putur, Bogang Wu, Caroline Weipert, Geoffrey Fell, Laura M. Spring, Seth A. Wander, Beverly Moy, Andreas Varkaris, Dejan Juric, Leif Ellisen, Ryan Corcoran, Aditya Bardia. TOP1 mutations mediate cross resistance to ADCs in metastatic breast cancer [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 3888.

Introduction: Folate receptor alpha (FOLR1) is a member of the folate transporter family expressed on normal tissues and overexpressed in multiple types of tumors, such as ovarian cancer, uterine cancer, non-small cell lung cancer, gastric cancer, breast cancer and kidney cancer. Currently, several clinical trials of FOLR1-targeting drugs [conventional IgG1 antibodies, which exhibit antibody-dependent cellular cytotoxicity/complement-dependent cytotoxicity (ADCC/CDC) activities, folic acid or antibody-drug conjugates and vaccines] have been conducted for ovarian and lung cancer. Therefore, FOLR1 is a remarkable target for cancer therapy under ongoing investigation. AccretaMab® technology involves combining both the POTELLIGENT®, a clinically validated ADCC-enhanced technology, and COMPLEGENT®, a new CDC-enhanced technology, systems to result in a superior technology for enhancing the killing activity of antibodies. KHK2805 is a novel humanized and CDR-altered anti-FOLR1 antibody developed with AccretaMab® technology. In this study, we evaluated the anti-cancer activity of KHK2805 in preclinical ovarian cancer models, both in vitro and in vivo, and confirmed the safety profile of KHK2805 in cynomolgus monkeys, since KHK2805 cross-reacts to cynomolgus monkey FOLR1. Materials and Methods: The binding kinetics of KHK2805 against recombinant FOLR1 (rFOLR1) were measured using the Biacore system. The epitope was determined with an ELISA against rFOLR1s. The in vitro ADCC and CDC activities against FOLR1-positive ovarian cancer cells were evaluated using PBMCs and serum from healthy volunteers. The in vivo anti-tumor activity of KHK2805 was examined using a SCID mouse model. The safety profile of KHK2805 was evaluated in cynomolgus monkeys. Results: KHK2805 induced potent ADCC and CDC activities against FOLR1-positive ovarian cancer cells. The ADCC activity of KHK2805 was significantly higher than that of the conventional anti-FOLR1 antibody. Furthermore, KHK2805 showed a potent ADCC activity against ovarian cancer cells with a low FOLR1 expression or low folic acid-uptake activity, which may be difficult to target with current FOLR1-targeting drugs. The results also showed that the markedly higher ADCC activity of KHK2805 was caused by its super-high affinity, unique epitope and use of AccretaMab® technology. In addition, the CDC activity of KHK2805 was also clearly higher than that of the conventional anti-FOLR1 antibody. This indicates that the higher CDC activity of KHK2805 is due to the application of protein engineering of CDR alterations and AccretaMab® technology. Moreover, the potent anti-tumor activity of KHK2805 was observed in a peritoneal dissemination model in SCID mice. Finally, we completed preliminary safety experiments with KHK2805. A repeated-dose toxicity study of KHK2805 (weekly 100 mg/kg for 4 weeks, intravenously) showed an acceptable tolerability profile in cynomolgus monkeys. Conclusions: KHK2805 may be a promising novel anti-FOLR1 therapeutic agent with a potent anti-tumor activity and tolerable safety profile for patients with the FOLR1 expression. Citation Format: Munetoshi Ando, Keiko Nagata, Hiroshi Ando, Mariko Nakano, Naoya Kameyama, Tsuguo Kubota, Maiko Adachi, Yui Suzuki, Kazuyasu Nakamura, Toshihiko Ishii, Ryuichiro Nakai, Takeshi Takahashi. A novel anti-FOLR1 antibody developed with AccretaMab® technology, KHK2805, exhibits markedly high ADCC/CDC activity and a tolerable safety profile in preclinical models. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2015 Nov 5-9; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(12 Suppl 2):Abstract nr C123.

TPS321 Background: A significant medical need exists for more effective and tolerable therapeutic options for patients with advanced gastrointestinal cancers (colorectal cancer [CRC], biliary tract cancer [BTC], and hepatocellular carcinoma [HCC]). Human epidermal growth factor receptor (HER3) expression is high in CRC and BTC, and low-to-intermediate in HCC. Patritumab deruxtecan (HER3-DXd; U3-1402/MK-1022) is a HER3-directed antibody-drug conjugate (ADC) comprising a fully human anti-HER3 immunoglobulin G1 monoclonal antibody (patritumab), which includes a plasma-stable, selectively cleavable linker, and a potent topoisomerase I inhibitor payload (DXd; released payload) that leverages the clinically validated deruxtecan (a derivative of exatecan) technology. DXd is cell membrane permeable and enables a bystander antitumor effect after DXd ADC internalization and linker cleavage, resulting in elimination of both target and neighboring tumor cells. Patritumab deruxtecan has shown antitumor activity in phase 1 and 2 studies of EGFR-mutated advanced non–small cell lung cancer and advanced breast cancer and in preclinical studies of CRC. This nonrandomized, open-label, phase 1/2 study (NCT06596694) will examine the safety and efficacy of patritumab deruxtecan in select gastrointestinal cancers. Methods: Approximately 130 adults (aged ≥18 years) will be allocated to 1 of 3 cohorts based on their disease. Patients in cohort 1 (CRC; n = 40) must have previously treated unresectable or metastatic colorectal adenocarcinoma. Patients in cohort 2 (BTC; n = 40) must have previously treated locally advanced or metastatic BTC. Patients in cohort 3 (HCC; n ≤50) must have previously treated advanced HCC. Prior topoisomerase I inhibitor therapy is not allowed. Patients in cohorts 1 and 2 will receive patritumab deruxtecan intravenously (IV) as monotherapy. Cohort 3 includes a dose-escalation phase to determine the recommended phase 2 dose of patritumab deruxtecan monotherapy and an efficacy phase. In the efficacy phase, patients will receive the recommended phase 2 dose of patritumab deruxtecan. The primary objectives are to evaluate safety and tolerability of patritumab deruxtecan and to evaluate the confirmed objective response rate per RECIST v1.1 assessed by blinded independent central review (BICR) for each cohort. Secondary objectives are to evaluate duration of response and progression-free survival per RECIST v1.1 by BICR and overall survival, and to characterize the pharmacokinetics of patritumab deruxtecan. Clinical trial information: NCT06596694 .

Background: HER3, a member of the ErbB family of receptor tyrosine kinases that activate multiple oncogenic signaling pathways, is overexpressed in 50-70% of breast cancers (BC). HER3 mRNA levels are higher in ER+ breast tumors than in other molecular subtypes. Inhibition of ER activity using an antagonist increased HER3 protein expression and activation, which was essential for growth and survival of ER+ BC cells resistant to the ER antagonist. Therefore, therapeutically targeting HER3 with HER3-DXd, an antibody-drug conjugate (ADC) composed of a fully human anti-HER3 monoclonal antibody (patritumab) conjugated to a topoisomerase I inhibitor payload (an exatecan derivative) via a tetrapeptide-based cleavable linker, can be an effective treatment for tamoxifen-resistant (TMR) ER+ BC. After first assessing HER3-DXd as a single agent, we sought to identify a synergistic partner to maximize HER3-DXd’s antitumor activity in HER3+/ER+ TMR BC. Methods: Whole-genome high-throughput siRNA screening was performed to identify targets whose inhibition enhances HER3-DXd’s antitumor efficacy. The antitumor effects of HER3-DXd plus the synergistic partners were assessed using a soft agar colony formation assay and a clonogenic assay in TMR HER3+/ER+ MCF7 and T47D BC cells. Treatment effects on cell cycle distribution, apoptosis, and expression of proteins that regulate cell cycle progression were assessed by flow cytometry, annexin V-PE and 7-AAD staining, and Western blotting, respectively. Results: HER3-DXd inhibited the anchorage-independent growth of HER3+/ER+ cells by >50% at 5 nM and their colony formation at 5-25 nM (P<0.05). To maximize HER3-DXd’s antitumor efficacy, we performed high-throughput siRNA screening and identified ATR, CD247, RAB7A, UPK3A, ROCK2, SLC29A1, and WNT7A as potential synergistic targets. Among these targets, inhibiting ATR with siRNA or BAY1895344 showed the most synergistic effect with HER3-DXd in HER3+/ER+ BC cells. In contrast, no synergistic effect was observed with the combination of BAY1895344 plus patritumab or control ADC (IgG-DXd), suggesting its dependence on HER3-DXd-mediated delivery of DXd. The combination of HER3-DXd plus BAY1895344 reprogrammed cell cycle progression from G2/M arrest to G1 arrest by inhibiting both ATR/Chk1/cyclin A2/CDK2 and ATR/Chk1/cyclin E/CDK2 signaling. The combination also reduced expression of H2AX, an ATR substrate that contributes to DNA repair, but increased that of γH2AX, indicating the induction of DNA damage. HER3-DXd and BAY1895344 synergistically inhibited growth of HER3+/ER+ BC cells by inducing apoptosis. Conclusion: The combination of HER3-DXd plus ATR inhibitors has therapeutic potential for overcoming tamoxifen resistance in HER3+/ER+ BC. We are currently validating the synergy in endocrine-resistant ER+ BC xenograft models. Citation Format: Xuemei Xie, Jangsoon Lee, Jon A. Fuson, Huey Liu, Young Jin Gi, Pang-Dian Fan, Kumiko Koyama, Debu Tripathy, Naoto T. Ueno. Targeting ATR enhances the antitumor efficacy of patritumab deruxtecan (HER3-DXd) in tamoxifen-resistant ER+ breast cancer cells by reprogramming cell cycle progression [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 LB088.

Antibody-drug conjugates (ADCs) are at the forefront of the drug development revolution occurring in oncology. Formed from three main components-an antibody, a linker molecule, and a cytotoxic agent ("payload"), ADCs have the unique ability to deliver cytotoxic agents to cells expressing a specific antigen, a great leap forward from traditional chemotherapeutic approaches that cause widespread effects without specificity. A variety of payloads can be used, including most frequently microtubular inhibitors (auristatins and maytansinoids), as well as topoisomerase inhibitors and alkylating agents. Finally, linkers play a critical role in the ADCs' effect, as cleavable moieties that serve as linkers impact site-specific activation as well as bystander killing effects, an upshot that is especially important in solid tumors that often express a variety of antigens. While ADCs were initially used in hematologic malignancies, their utility has been demonstrated in multiple solid tumor malignancies, including breast, gastrointestinal, lung, cervical, ovarian, and urothelial cancers. Currently, six ADCs are FDA-approved for the treatment of solid tumors: ado-trastuzumab emtansine and trastuzumab deruxtecan, both anti-HER2; enfortumab-vedotin, targeting nectin-4; sacituzuzmab govitecan, targeting Trop2; tisotumab vedotin, targeting tissue factor; and mirvetuximab soravtansine, targeting folate receptor-alpha. Although they demonstrate utility and tolerable safety profiles, ADCs may become ineffective as tumor cells undergo evolution to avoid expressing the specific antigen being targeted. Furthermore, the current cost of ADCs can be limiting their reach. Here, we review the structure and functions of ADCs, as well as ongoing clinical investigations into novel ADCs and their potential as treatments of solid malignancies.

Background: Trophoblast cell surface antigen 2 (TROP2) is highly expressed in various epithelial tumors, including non-small cell lung cancer (NSCLC) and triple negative breast cancer (TNBC), and correlates with poor prognosis. Datopotamab deruxtecan (Dato-DXd) is an antibody-drug conjugate (ADC) consisting of a humanized anti-TROP2 IgG1 monoclonal antibody covalently linked to a highly potent topoisomerase I (Topo I) inhibitor payload via a stable, tumor-selective, tetrapeptide-based cleavable linker. Dato-DXd has shown encouraging antitumor activity and a manageable safety profile in patients with NSCLC and TNBC as part of the first-in-human, Phase 1 study (NCT03401385). To enhance clinical responses, clinical trials evaluating combination therapy of Dato-DXd with PD-1/PD-L1 inhibitors in NSCLC and TNBC are currently ongoing (e.g. NCT04526691 and NCT03742102). Here we report preclinical evidence for supporting increased antitumor activity of Dato-DXd in combination with PD-1/PD-L1 inhibitors using a syngeneic mouse tumor model and its impact on tumor immunity. Methods: MC38 mouse colon adenocarcinoma cells stably transfected with human TROP2 (hTROP2_MC38) were inoculated subcutaneously in immunocompetent C57BL6 mice or immunodeficient nude mice and the antitumor activity of Dato-DXd with or without mouse PD-1/PD-L1 inhibitors was evaluated. The tumor infiltrating immune cells in hTROP2_MC38 tumors were analyzed by flow cytometry at 3 days and 10 days after dosing. In addition, the impact of CD8+ T cell depletion on the antitumor activity of Dato-DXd against hTROP2_MC38 tumors in C57BL6 mice was evaluated. Results: Dato-DXd showed stronger antitumor activity against hTROP2_MC38 tumors in immunocompetent C57BL6 mice than in immunodeficient nude mice, which suggests that immune cells play an important role in the mechanism of action (MoA) of Dato-DXd. The combination of Dato-DXd and mouse PD-1/PD-L1 inhibitors enhanced antitumor activity compared to monotherapy against hTROP2_MC38 tumors in C57BL6 mice. Activation of the dendritic cells in hTROP2_MC38 tumors in C57BL6 mice was observed after Dato-DXd monotherapy or combination therapy with mouse PD-1/PD-L1 inhibitors, which led to the increase of tumor infiltrating CD8+ T cells. Involvement of CD8+ T cells in the MoA of Dato-DXd was also supported by the result that CD8+ T cell depletion in C57BL6 mice decreased the antitumor activity of Dato-DXd against hTROP2_MC38 tumors. In addition, activation of tumor infiltrating NK cells and macrophages was observed after Dato-DXd monotherapy or combination therapy, suggesting the potential effect on innate immunity. Conclusion: These results suggest that Dato-DXd may stimulate tumor immunity and sensitize tumors to PD-1/PD-L1 inhibitors. The combination of Dato-DXd and PD-1/PD-L1 blockers could be a valuable therapy for patients with TROP2-positive tumors. Citation Format: Daisuke Okajima, Satoru Yasuda, Takami Suzuki, Michiko Kitamura, Junko Yamaguchi, Takanori Maejima, Tsuyoshi Karibe, Tadashi Toki, Penny Phillips, Toshinori Agatsuma. Datopotamab deruxtecan (Dato-DXd) enhances antitumor response to PD-1/PD-L1 inhibitors in TROP2-expressing tumors in mice [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2932.

Overcoming Challenges in the Development of Anticancer ADCs

Folate Receptor α(FRα)exhibits an increased expression on cell surfaces in multiple solid tumors, including ovarian, lung, endometrial cancer, with limited expression in normal tissues. Recently, an antibody drug conjugate (ADC) with a microtubule inhibitor (MTI) payload, was approved by FDA for FRα-expressing platinum-resistant ovarian cancer. However, it can cause severe ocular toxicities and show therapeutic efficacies only in patients with high expression. In order to address the unmet needs of additional patient populations, we sought to develop a next generation FRα-targeting ADC, SYS6041, actively targeting tumors with a broad range of FRα expression, especially moderate or low expression levels. SYS6041 was developed adopting a novel ADC platform with a camptothecin-based topoisomerase 1 inhibitor payload, Exatecan, tethered to a cleavable linker. The drug-to-antibody ratio standed 8.0. Upon binding to FRα, SYS6041 underwent receptor-mediated internalization. Subsequent proteolytic cleavage released the cytotoxic payload Exatecan, leading to DNA strand breaks and consequently disrupting DNA replication and transcription. SYS6041 demonstrated potent cytotoxicities against FRα expressing cancer cell-lines and effective bystander activities towards tumor cells without FRα expression. It demonstrated superior antitumor effects across ovarian endometrial and lung cancer CDX models compared to the MTI-ADC especially in models with low or moderate FRα expression, with a minimum effect dose(MED) at 0.75 mg/kg. In a FRα moderate ovarian PDX model with Olaparib resistance, SYS6041 significantly inhibited tumor growth at 3 mg/kg, whereas the MTI-ADC showed no inhibitory effect at 6 mg/kg. Furthermore, Exatecan demonstrated a significant enrichment in the tumors of mice bearing tumors, achieving a tumor-to-blood ratio of 182 following a dosage of 6 mg/kg of SYS6041.SYS6041 exhibited a stable PK profile with AUC of Exatecan less than 0.01% of that of total Exatecan in ADC. In a GLP toxicity study in cynomolgus monkey, SYS6041 demonstrated an encouraging tolerability with an HNSTD of 50 mg/kg, and favorable PK and DAR stability, and presented a good tolerability profile with therapeutic window (TW)>66 (HNSTD/MED). In conclusion, the distinct mode of action along with the proprietary drug-linker platform resulted in an excellent in-vivo antitumor potency and safety, which supports the potential of SYS6041 as a novel therapeutic agent that may help address unmet needs in patients with moderate and low FRα expressing malignancies. Citation Format: Jieru Liu, Huan Li, Mo Dan, Xiwu Hui, Bin Yao, Yang Zhang, Linlin Xiao, Bin Kang, Can Yuan, Bin Bao, Miaomiao Wei, Chunqing Qu, Yanling Li, Wei Yang. SYS6041, a next-generation Folate Receptor α-targeted antibody-drug conjugates for the treatment of FRα-expressing malignancies [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 6832.

Background of the study: [fam-] trastuzumab deruxtecan (DS-8201a) is a HER2-targeting antibody-drug conjugate (ADC) with a humanized anti-HER2 monoclonal antibody attached to a topoisomerase I inhibitor payload by a peptide-based cleavable linker. [fam-] trastuzumab deruxtecan has been shown to be highly effective in preclinical and clinical studies. In addition to payload potency, drug-to-antibody ratio and linker stability, the effectiveness of the ADC is expected to be defined by intracellular molecular dynamics of the ADC/HER2 complex such as internalization activity, translocation to lysosomes and lysosomal enzyme-mediated linker cleavage. However, the machinery of ADC dynamics has not been studied quantitatively. Methods: First, in vitro quantitative assays to study [fam-] trastuzumab deruxtecan intracellular dynamics were established. Second, differentially expressed gene (DEG) analysis among lysosome trafficking-high/low HER2 expressing cell lines was performed to understand intracellular dynamics. Third, functional relevance of the gene extracted from DEG analysis was assessed by siRNA-mediated in vitro knockdown study. Finally, to comprehend contribution of lysosome activity to intracellular dynamics of [fam-] trastuzumab deruxtecan, in vitro lysosome inhibition studies were performed using bafilomycin A1. Then, impact of autophagy activation on the dynamics was studied with mTOR inhibitor everolimus. Results: [Fam-] trastuzumab deruxtecan bound to the cell surface of HER2 expressing cell lines in expression level-dependent manner. Lysosome trafficking study revealed that two cell lines, JIMT1 and MDA-MB-453, showed high trafficking activity for the modest HER2 expression. DEG analysis result exhibited that lysosome-related genes were significantly upregulated in the two cell lines, which suggested involvement of lysosome activity in ADC dynamics. Lysosome trafficking of [fam-] trastuzumab deruxtecan was hampered by knockdown of such genes. Lysosome is supposed to be the endpoint of ADC processing and its activity itself might affect the ADC activity. Inhibition of lysosome activity by bafilomycin A1 resulted in weakening ADC dynamics in the cell lines. In addition, contribution of lysosome activation via autophagy stimulation using everolimus resulted in activation of lysosome trafficking of the ADC in MDA-MB-453. Conclusion: In vitro quantitative analyses of intracellular [fam-] trastuzumab deruxtecan dynamics revealed that lysosome related activity played important roles in the ADC dynamics. As lysosome activity is known to be modulated by autophagy mechanism, influence of autophagy activation to ADC dynamics was elucidated in this study. The results obtained suggest a scientific rationale for combo treatment of ADC and autophagy activator as well as the advantage of ADC on autophagy-active cancers. Citation Format: Manabu Abe, Motoko Nagata, Kumiko Koyama, Satoru Yasuda, Tsuyoshi Hirata, Yusuke Kuwahara, Koichiro Inaki, Naomi Kasanuki, Megumi Minami, Katsunobu Hagihara, Kenichi Wakita, Eric Slosberg, Masato Murakami. Autophagy modulates intracellular dynamics of [fam-] trastuzumab deruxtecan (DS-8201a), a novel HER2 antibody-drug conjugate (HER2-ADC) [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 LB-264.

Introduction: Despite recent treatment advances, the prognosis for patients diagnosed with locally recurrent inoperable or metastatic triple-negative breast cancer (TNBC) remains poor. Single-agent chemotherapy has been the mainstay of treatment for metastatic TNBC for many years, with very limited treatment options for patients who are not candidates for anti-PD-1/PD-L1 therapy; consequently, there remains an urgent unmet need. Trophoblast cell surface protein 2 (TROP2) is a type I transmembrane glycoprotein highly expressed on various solid tumors, including breast. Datopotamab deruxtecan (Dato-DXd) is an antibody-drug conjugate (ADC) composed of a humanized anti-TROP2 IgG1 monoclonal antibody covalently linked to an exatecan derivative (DXd) – a highly potent topoisomerase I inhibitor payload – via a stable, tumor-selective, tetrapeptide-based cleavable linker. Dato-DXd has previously been evaluated in TROPION-PanTumor01 (NCT03401385) – an ongoing Phase I clinical trial across multiple advanced solid tumors, including heavily-pretreated, metastatic TNBC. As of the July 30, 2021 cut-off date, the objective response rate (ORR) by blinded independent central review (BICR) was 34% (15/44) in all patients with TNBC and 52% (14/27) in those patients treatment-naïve to topoisomerase I inhibitor-based ADC therapies, with measurable disease (per RECIST 1.1) at baseline. Furthermore, a manageable safety profile was reported, with no new safety signals identified; low grade nausea and stomatitis were most frequent, with neutropenia and diarrhea being uncommon. The aim of the phase 3 TROPION-Breast02 trial is to evaluate the efficacy and safety of Dato-DXd versus investigator’s choice of chemotherapy (ICC) in patients with TNBC who are not candidates for PD-1/PD-L1 inhibitor therapy. Methods: TROPION-Breast02 (NCT05374512) is an ongoing Phase 3, open-label, randomized study of Dato-DXd versus ICC in first-line treatment of patients with locally recurrent inoperable or metastatic TNBC. Approximately 600 patients will be randomized 1:1 to receive either Dato-DXd 6 mg/kg IV every three weeks or ICC (paclitaxel, nab-paclitaxel, carboplatin, capecitabine, or eribulin mesylate). Patients ≥18 years with histologically/cytologically confirmed TNBC who have not received prior chemotherapy or targeted systemic therapy for metastatic or locally recurrent inoperable disease are eligible for inclusion. Patients are also required to have an ECOG PS of 0 or 1 and be eligible for treatment with one of the ICC options per investigator assessment. Patients must have ≥1 measurable lesion per RECIST v1.1 not previously irradiated, along with a formalin-fixed and paraffin-embedded tumor sample available. Those with a previously treated neoplastic spinal cord compression or clinically inactive brain metastases can be included. Key exclusion criteria are history of another primary malignancy; persistent toxicity from previous anti-cancer treatments; uncontrolled infections; current/prior interstitial lung disease/pneumonitis or clinically severe pulmonary function compromise; clinically significant corneal disease; and prior treatment with topoisomerase I inhibitors, TROP2-targeted therapy, or the same chemotherapy agent chosen for on-study ICC. The dual primary endpoints are progression free survival (PFS) per RECIST 1.1 by BICR, and OS. Secondary endpoints include ORR, duration of response, PFS by investigator assessment, time to deterioration for patient-reported outcomes, time to first subsequent therapy, time to second subsequent therapy, time to second progression or death, pharmacokinetics and immunogenicity of Dato-DXd, and safety. Recruitment for this study is ongoing as of June 2022. Citation Format: Rebecca Dent, David W. Cescon, Thomas Bachelot, Kyung Hae Jung, Zhi-Ming Shao, Shigehira Saji, Tiffany A. Traina, Petra Vuković, Darlington Mapiye, Micah Maxwell, Peter Schmid, Javier Cortés. TROPION-Breast02: Phase 3, open-label, randomized study of first-line datopotamab deruxtecan versus chemotherapy in patients with locally recurrent inoperable or metastatic TNBC who are not candidates for anti-PD-(L)1 therapy [abstract]. In: Proceedings of the 2022 San Antonio Breast Cancer Symposium; 2022 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2023;83(5 Suppl):Abstract nr OT1-03-05.

Background: Ado-trastuzumab emtansine (T-DM1), a HER2-targeted antibody drug conjugate (ADC), is approved for patients with HER2-positive metastatic breast cancer (BC) after disease progression on a trastuzumab-based regimen. Approval of T-DM1 was based on the EMILIA trial in which T-DM1 demonstrated an objective response rate (ORR) of 43.6%, a median progression-free survival (PFS) of 9.6 months, and an overall survival (OS) of 30.9 months (Verma S, et al. NEJM. 2012). DS-8201a is a novel HER2-targeted ADC with a humanized HER2 antibody attached to a topoisomerase I inhibitor payload by a cleavable peptide-based linker, and with a high drug-to-antibody ratio of 7 to 8. In an ongoing phase 1 trial, DS-8201a showed a manageable safety profile and promising antitumor activity in HER2-positive BC previously treated with T-DM1 (confirmed ORR of 54.5%; April 2018 data cutoff) (Iwata et al, ASCO 2018). The pivotal, phase 2 DESTINY-BREAST01 trial in this population with HER2-positive BC who received prior T-DM1 is ongoing (Baselga et al, ASCO 2018). Study Description: This multicenter, open-label, phase 3 trial will assess the efficacy and safety of DS-8201a vs T-DM1 in subjects with HER2-positive (IHC 3+ or IHC 2+/ISH+; confirmed by centralized testing) unresectable and/or metastatic BC previously treated with trastuzumab and a taxane (NCT03529110, DESTINY-BREAST03). Subjects who previously received a HER2-targeted ADC are excluded. Approximately 500 eligible subjects will be randomized (1:1) to receive DS-8201a (5.4 mg/kg) or T-DM1 (3.6 mg/kg) IV once every 3 weeks. Randomization will be stratified by hormone receptor status, prior pertuzumab treatment, and history of visceral disease. For subjects randomized to T-DM1, the treatment will be in accordance with the approved label. The primary efficacy endpoint is PFS based on blinded, independent central review using RECIST v1.1 criteria. Secondary efficacy endpoints include OS, ORR, duration of response, clinical benefit rate, and PFS based on investigator assessment. Safety assessments include serious and treatment-emergent adverse events, physical examinations, vital signs, and clinical laboratory parameters. Health related quality of life will also be measured. The primary analysis for PFS will be performed when approximately 331 PFS events have been observed. This will provide 90% power to detect a hazard ratio of 0.70 for PFS with a 1-sided alpha of 0.025, assuming a median PFS with T-DM1 of 9.6 months and that PFS follows an exponential distribution. Long-term follow-up will continue after the primary analysis every 3 months until death, withdrawal of consent, loss to follow-up, or study closure. Efficacy analyses will include all randomized subjects, and safety analyses will include all randomized subjects who received ≥1 dose of study treatment. The study will enroll subjects from approximately 150 sites851468 including in North America, Europe, and Asia. Citation Format: Verma S, Shahidi J, Lee C, Wang K, Cortes J. Trastuzumab deruxtecan (DS-8201a) vs ado-trastuzumab emtansine (T-DM1) for subjects with HER2-positive, unresectable and/or metastatic breast cancer who previously received trastuzumab and a taxane: A phase 3, randomized study [abstract]. In: Proceedings of the 2018 San Antonio Breast Cancer Symposium; 2018 Dec 4-8; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2019;79(4 Suppl):Abstract nr OT2-07-03.