Next-generation Antibody-drug Conjugates Research Articles

Mechanisms and optimization strategies for the occurrence of antibody-drug conjugate toxicity

Antibody-drug conjugates (ADCs) have emerged as a promising class of targeted cancer therapies, with 13 ADCs approved by the FDA and over 140 undergoing clinical research. ADCs are designed to enhance the selective delivery of cytotoxic drugs to tumor cells through high-affinity monoclonal antibodies (mAbs), improving the therapeutic index of the drug. Despite their potential, ADCs face significant challenges related to toxicity, including hematologic, ocular, skin, and neurological side effects, which limit their clinical application. This review provides an in-depth analysis of the mechanisms underlying ADC toxicity, which includes both targeted and off-target effects that can damage normal cells and tissues. Strategies for optimizing ADC safety, such as improving antibody structure, effector molecules, linkers, and combination therapies, are discussed. The review emphasizes the need for continued research to address these challenges and reduce off-target toxicity while enhancing the efficacy of ADCs, paving the way for the development of safer and more effective next-generation ADC drugs.

Development and validation of a radiomics-based nomogram for predicting two subtypes of HER2-negative breast cancer.

Breast cancer is the most common malignant tumor among women, with an increasing incidence each year. The subtypes of human epidermal growth factor receptor 2 (HER2)-negative breast cancer, classified as HER2-low and HER2-zero based on HER2 receptor expression, show differences in clinical characteristics, therapeutic approaches, and prognoses. Distinguishing between these subtypes is clinically valuable as it can impact treatment strategies, including the use of next-generation antibody-drug conjugates (ADCs) targeting HER2-low tumors. This study aimed to develop a nomogram based on dynamic magnetic resonance imaging (MRI) and clinical indicators to differentiate between HER2-low and HER2-zero subtypes in HER2-negative breast cancer patients. This study included 214 breast cancer patients from two centers, Hospital A (Affiliated Hospital of Jining Medical University, n=178) and Hospital B (Ningyang No. 1 People's Hospital, n=36). HER2 status was determined by immunohistochemistry (IHC) and fluorescence in situ hybridization (FISH). Among the participants, 112 cases were identified as HER2-low and 102 as HER2-zero. Patients from Hospital A were split into a training set and an internal test set in an 8:2 ratio, while the 36 patients from Hospital B were used as an external test set. Regions of interest (ROI) were delineated on phase 2 enhanced scans and diffusion weighted imaging (DWI) images, with features selected via Pearson correlation coefficients and least absolute shrinkage and selection operator (LASSO) regression. A K-Nearest Neighbor (KNN) model was employed to calculate the rad score, and clinical predictors (tumor maximum diameter and CA153) were identified through logistic regression analysis. These predictors, combined with the rad score, were incorporated into the final nomogram model. The model's accuracy was evaluated using area under curve (AUC) values in both the internal and external validation sets. The nomogram achieved AUC values of 0.873 and 0.859 in the internal and external validation sets, respectively, demonstrating superior performance over single-feature models. Decision curve analysis (DCA) indicated substantial net clinical benefits, and calibration curves displayed strong alignment between the model's predictions and actual outcomes in both sets. This nomogram shows high accuracy and stability in differentiating HER2-low and HER2-zero subtypes among HER2-negative breast cancer patients, suggesting potential clinical utility in refining treatment decisions and identifying candidates for ADC therapy in HER2-low cases.

Site-Specific Quadruple-Functionalised Antibodies.

Antibody-drug conjugates (ADCs) are a growing class of chemotherapeutic agents that have yielded striking clinical successes. However, the efficacy of ADCs often suffers from issues associated with tumor heterogeneity and resistance. To overcome these problems, a new generation of ADCs comprising a single monoclonal antibody with multiple different payloads attached, termed multi-payload ADCs, have been developed. Here we deploy multiple orthogonal site-specific protein modification strategies to generate highly homogeneous multi-functionalised antibody conjugates comprising up to four different functionalities installed at four unique sites on the antibody. This work, which includes the use of a site-specific cyclopropenone (CPO)-based reagent, represents the first example of a homogeneous multi-payload ADC with a payload count greater than two, and thereby facilitates the development of the next generation of ADCs.

OS Therapies to Develop ‘Tunable’ ADCs

OS Therapies is advancing a next-generation antibody-drug conjugate (ADC) technology platform, known as tunable ADCs (tADCs), which features tunable, tailored antibody-linker-payload candidates. This platform leverages the company’s proprietary silicone linker technology, enabling the delivery of multiple payloads per linker.

Single-Domain Antibodies as Antibody-Drug Conjugates: From Promise to Practice-A Systematic Review.

Antibody-drug conjugates (ADCs) represent potent cancer therapies that deliver highly toxic drugs to tumor cells precisely, thus allowing for targeted treatment and significantly reducing off-target effects. Despite their effectiveness, ADCs can face limitations due to acquired resistance and potential side effects. This study focuses on advances in various ADC components to improve both the efficacy and safety of these agents, and includes the analysis of several novel ADC formats. This work assesses whether the unique features of VHHs-such as their small size, enhanced tissue penetration, stability, and cost-effectiveness-make them a viable alternative to conventional antibodies for ADCs and reviews their current status in ADC development. Following PRISMA guidelines, this study focused on VHHs as components of ADCs, examining advancements and prospects from 1 January 2014 to 30 June 2024. Searches were conducted in PubMed, Cochrane Library, ScienceDirect and LILACS using specific terms related to ADCs and single-domain antibodies. Retrieved articles were rigorously evaluated, excluding duplicates and non-qualifying studies. The selected peer-reviewed articles were analyzed for quality and synthesized to highlight advancements, methods, payloads, and future directions in ADC research. VHHs offer significant advantages for drug conjugation over conventional antibodies due to their smaller size and structure, which enhance tissue penetration and enable access to previously inaccessible epitopes. Their superior stability, solubility, and manufacturability facilitate cost-effective production and expand the range of targetable antigens. Additionally, some VHHs can naturally cross the blood-brain barrier or be easily modified to favor their penetration, making them promising for targeting brain tumors and metastases. Although no VHH-drug conjugates (nADC or nanoADC) are currently in the clinical arena, preclinical studies have explored various conjugation methods and linkers. While ADCs are transforming cancer treatment, their unique mechanisms and associated toxicities challenge traditional views on bioavailability and vary with different tumor types. Severe toxicities, often linked to compound instability, off-target effects, and nonspecific blood cell interactions, highlight the need for better understanding. Conversely, the rapid distribution, tumor penetration, and clearance of VHHs could be advantageous, potentially reducing toxicity by minimizing prolonged exposure. These attributes make single-domain antibodies strong candidates for the next generation of ADCs, potentially enhancing both efficacy and safety.

Abstract PO1-04-02: ACE-Breast-03: A phase 2 study of ARX788, a novel next-generation anti-HER antibody-drug conjugate in HER2-positive metastatic breast cancer patients previously treated with trastuzumab deruxtecan

Abstract Background: HER2 overexpression and/or amplification confers aggressive tumor behavior and decreased survival in patients with breast cancer. Targeting of HER2 is a validated therapeutic strategy, and trastuzumab deruxtecan (T-DXd) has emerged as a preferred treatment in the second-line setting. However, effective treatment following progression on T-DXd remains a major unmet medical need. ARX788 is a next-generation antibody-drug conjugate (ADC) using a technology platform whereby a HER2-specific monoclonal antibody is conjugated to amberstatin 269 (AS269), a potent cytotoxic tubulin inhibitor. Site-specificity, high homogeneity, and stable covalent conjugation of ARX788 leads to the slow release and prolonged peak of serum pAF-AS269, which may contribute to the lower systemic toxicity and increased targeted delivery of payload to tumor cells, at a lower effective dose compared with other HER2 ADCs. Methods: ACE-Breast-03 (NCT04829604) is a global, phase 2 study designed to assess anticancer activity and safety in approximately 40 patients with unresectable or metastatic HER2 positive breast cancer who have been previously treated with T-DXd. Eligibility criteria include measurable disease by RECIST v1.1, radiographically stable brain metastases, no more than three prior lines of treatment in the metastatic setting, and HER2 positivity as determined by central review. If recently progressed on T-DXd or trastuzumab emtansine (T-DM1), a new biopsy for HER2 status is required. Patients will be administered 1.5 mg/kg of ARX788 every three weeks. Efficacy will be assessed using RECIST v1.1 at nine-week intervals after starting ARX788 until progression or start of new cancer treatment. Objective response rate is the primary endpoint, with disease control rate, progression-free survival, overall survival, best overall response, duration of response, and time to response as additional endpoints. The safety and tolerability profile will also be evaluated. PK analysis will determine serum concentrations of ARX788, total antibody, and pAF-AS269. Potential predictive and/or prognostic biomarkers will be analyzed for exploratory purposes. Descriptive statistics will be used to evaluate anticancer activity, safety, and tolerability. The study is currently recruiting patients. Citation Format: Kashif Ali, Laila Agrawal, Anu Thummala, Kevin Kalinsky, Sami Ali, Sibel Blau, Margaret Block, Michael Danso, Denise Yardley, Jay Andersen, Adrienne Gropper-Waks, Priya Jayachandran, Igor Makhlin, Petros Nikolinakos, Joyce O'Shaughnessy, Sandra Aung, Colin Hessel, Hope Rugo, William Gradishar, Debu Tripathy. ACE-Breast-03: A phase 2 study of ARX788, a novel next-generation anti-HER antibody-drug conjugate in HER2-positive metastatic breast cancer patients previously treated with trastuzumab deruxtecan [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO1-04-02.

Precision Medicine in Rheumatic Diseases: Unlocking the Potential of Antibody-Drug Conjugates.

In the era of precision medicine, antibody-drug conjugates (ADCs) have emerged as a cutting-edge therapeutic strategy. These innovative compounds combine the precision of monoclonal antibodies with the potent cell-killing or immune-modulating abilities of attached drug payloads. This unique strategy not only reduces off-target toxicity but also enhances the therapeutic effectiveness of drugs. Beyond their well established role in oncology, ADCs are now showing promising potential in addressing the unmet needs in the therapeutics of rheumatic diseases. Rheumatic diseases, a diverse group of chronic autoimmune diseases with varying etiologies, clinical presentations, and prognoses, often demand prolonged pharmacological interventions, creating a pressing need for novel, efficient, and low-risk treatment options. ADCs, with their ability to precisely target the immune components, have emerged as a novel therapeutic strategy in this context. This review will provide an overview of the core components and mechanisms behind ADCs, a summary of the latest clinical trials of ADCs for the treatment of rheumatic diseases, and a discussion of the challenges and future prospects faced by the development of next-generation ADCs. SIGNIFICANCE STATEMENT: There is a lack of efficient and low-risk targeted therapeutics for rheumatic diseases. Antibody-drug conjugates, a class of cutting-edge therapeutic drugs, have emerged as a promising targeted therapeutic strategy for rheumatic disease. Although there is limited literature summarizing the progress of antibody-drug conjugates in the field of rheumatic disease, updating the advancements in this area provides novel insights into the development of novel antirheumatic drugs.

Radiotherapy activates picolinium prodrugs in tumours.

Radiotherapy-induced prodrug activation provides an ideal solution to reduce the systemic toxicity of chemotherapy in cancer therapy, but the scope of the radiation-activated protecting groups is limited. Here we present that the well-established photoinduced electron transfer chemistry may pave the way for developing versatile radiation-removable protecting groups. Using a functional reporter assay, N-alkyl-4-picolinium (NAP) was identified as a caging group that efficiently responds to radiation by releasing a client molecule. When evaluated in a competition experiment, the NAP moiety is more efficient than other radiation-removable protecting groups discovered so far. Leveraging this property, we developed a NAP-derived carbamate linker that releases fluorophores and toxins on radiation, which we incorporated into antibody-drug conjugates (ADCs). These designed ADCs were active in living cells and tumour-bearing mice, highlighting the potential to use such a radiation-removable protecting group for the development of next-generation ADCs with improved stability and therapeutic effects.

Azobenzene-Based Linker Strategy for Selective Activation of Antibody-Drug Conjugates.

Existing antibody-drug conjugate (ADC) linkers, whether cleavable or non-cleavable, are designed to release highly toxic payloads or payload derivatives upon internalisation of the ADCs into cells. However, clinical studies have shown that only <1 % of the dosed ADCs accumulate in tumour cells. The remaining >99 % of ADCs are nonspecifically distributed in healthy tissue cells, thus inevitably leading to off-target toxicity. Herein, we describe an intelligent tumour-specific linker strategy to address these limitations. A tumour-specific linker is constructed by introducing a hypoxia-activated azobenzene group as a toxicity controller. We show that this azobenzene-based linker is non-cleavable in healthy tissues (O2 >10 %), and the corresponding payload derivative, cysteine-appended azobenzene-linker-monomethyl auristatin E (MMAE), can serve as a safe prodrug to mask the toxicity of MMAE (switched off). Upon exposure to the hypoxic tumour microenvironment (O2<1 %), this linker is cleaved to release MMAE and fully restores the high cytotoxicity of the ADC (switched on). Notably, the azobenzene linker-containing ADC exhibits satisfactory antitumour efficacy in vivo and a larger therapeutic window compared with ADCs containing traditional cleavable or non-cleavable linkers. Thus, our azobenzene-based linker sheds new light on the development of next-generation ADC linkers.

Exploring the next generation of antibody-drug conjugates.

Antibody-drug conjugates (ADCs) are a promising cancer treatment modality that enables the selective delivery of highly cytotoxic payloads to tumours. However, realizing the full potential of this platform necessitates innovative molecular designs to tackle several clinical challenges such as drug resistance, tumour heterogeneity and treatment-related adverse effects. Several emerging ADC formats exist, including bispecific ADCs, conditionally active ADCs(also known asprobody-drug conjugates), immune-stimulating ADCs, protein-degrader ADCs and dual-drug ADCs, and each offers unique capabilities for tackling these various challenges. For example, probody-drug conjugates can enhance tumour specificity, whereas bispecific ADCs and dual-drug ADCs can address resistance and heterogeneity with enhanced activity. The incorporation of immune-stimulating and protein-degrader ADCs, which have distinct mechanisms of action, into existing treatment strategies could enable multimodal cancer treatment. Despite the promising outlook, the importance of patient stratification and biomarker identification cannot be overstated for these emerging ADCs, as these factors are crucial to identify patients who are most likely to derive benefit. As we continue to deepen our understanding of tumour biology and refine ADC design, we will edge closer to developing truly effective and safe ADCs for patients with treatment-refractory cancers. In this Review, we highlight advances in each ADC component (the monoclonal antibody, payload, linker and conjugation chemistry) and provide more-detailed discussions on selected examples of emerging novel ADCs of each format, enabled by engineering of one or more of these components.

Development of antibody-drug conjugates in cancer: Overview and prospects.

In recent years, remarkable breakthroughs have been reported on antibody-drug conjugates (ADCs), with 15 ADCs successfully entering the market over the past decade. This substantial development has positioned ADCs as one of the fastest-growing domains in the realm of anticancer drugs, demonstrating their efficacy in treating a wide array of malignancies. Nonetheless, there is still an unmet clinical need for wider application, better efficacy, and fewer side effects of ADCs. An ADC generally comprises an antibody, a linker and a payload, and the combination has profound effects on drug structure, pharmacokinetic profile and efficacy. Hence, optimization of the key components provides an opportunity to develop ADCs with higher potency and fewer side effects. In this review, we comprehensively reviewed the current development and the prospects of ADC, provided an analysis of marketed ADCs and the ongoing pipelines globally as well as in China, highlighted several ADC platforms and technologies specific to different pharmaceutical enterprises and biotech companies, and also discussed the new related technologies, possibility of next-generation ADCs and the directions of clinical research.

Abstract B128: Preclinical characterization of ETx-22, a next-generation antibody drug conjugate (ADC) targeting nectin-4

Abstract Nectin-4 is overexpressed in a variety of cancers including urothelial, breast, cervix, lung and ovarian, with low level of expression in normal tissues. Enfortumab vedotin (EV), an anti-Nectin-4 ADC with a monomethyl auristatin (MMAE) payload, has been a significant advancement in the treatment of urothelial cancer. Yet, it is associated with important skin toxicity owing to the expression of nectin-4 in the skin, and peripheral neuropathy secondary to its toxin. We have developed a next-generation anti-nectin-4 ADC, ETx-22, which consists of a humanized IgG1, Fcg-silent antibody that is bound via maleimide-ß-glucuronide poly-sarcosine linkers to 8 molecules of the topoisomerase-I inhibitor, exatecan. Here, we describe the key pharmacological experiments, in-vivo efficacy and results from the good laboratory practice (GLP) toxicology studies. In-vitro studies have shown that the ETx-22 antibody exhibits a high affinity for tumor-expressed Nectin-4, but 10-fold lower affinity for nectin-4 expressed on skin, with significantly higher internalization in tumor cells. In-vivo efficacy was evaluated in various patient-derived xenograft models. ETx-22 was shown to be consistently superior to EV as well as chemotherapy, in a broad array of tumors, including urothelial, ovarian, cervical and triple negative breast cancer irrespective of the degree of nectin-4 expression. Antitumor activity was also observed in MMAE-resistant models. GLP studies evaluated repeated doses of ETx-22 in cynomolgus monkeys at two doses, 10 and 20 mg/kg. The treatment was administered as 30-minute intravenous infusions once every 2 weeks for a total of three cycles. The ETx-22 antibody alone was also evaluated at 20 mg/kg. Both ETx-22 doses were well tolerated and all animals survived up to scheduled euthanasia. The common observation at both doses was mild multifocal skin hyperpigmentation, which was reversible. No skin rash or pruritus was reported. Mild transient decrease in red blood cells was observed in between cycles. No gastrointestinal toxicities were observed. No signs of interstitial lung disease or eye toxicity were detected. Based on these findings, the highly non- severely toxic dose (HNSTD) is at least 20 mg/kg. The half-life of ETx-22 ranged from 3.36 – 3.6 days. In conclusion, preclinical and pharmacological shows that ETx-22 has better tolerance, longer half-life, and higher efficacy to EV. No skin rash was reported in toxicology studies. A first in human study (EXCEED) is planned to start in 2023. Citation Format: Hatem Azim, Florence Lhospice, Xavier Preville, Joanna Fares, Marc Lopez, Emerence Crompot, Emmanuelle Josselin, Remy Castellano, Maria Wehbeh, Yves Collette, Jack Elands, Daniel Olive. Preclinical characterization of ETx-22, a next-generation antibody drug conjugate (ADC) targeting nectin-4 [abstract]. In: Proceedings of the AACR-NCI-EORTC Virtual International Conference on Molecular Targets and Cancer Therapeutics; 2023 Oct 11-15; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2023;22(12 Suppl):Abstract nr B128.

A novel human single-domain antibody-drug conjugate targeting CEACAM5 exhibits potent in vitro and in vivo antitumor activity.

Leveraging the specificity of antibody to deliver cytotoxic agent into tumor, antibody-drug conjugates (ADCs) have become one of the hotspots in the development of anticancer therapies. Although significant progress has been achieved, there remain challenges to overcome, including limited penetration into solid tumors and potential immunogenicity. Fully human single-domain antibodies (UdAbs), with their small size and human nature, represent a promising approach for addressing these challenges. Carcinoembryonic antigen-related cell adhesion molecule 5 (CEACAM5) is a glycosylated cell surface protein that rarely expressed in normal adult tissues but overexpressed in diverse cancers, taking part in tumorigenesis, progression, and metastasis. In this study, we investigated the therapeutic potential of UdADC targeting CEACAM5. We performed biopanning in our library and obtained an antibody candidate B9, which bound potently and specifically to CEACAM5 protein (KD = 4.84 nM) and possessed excellent biophysical properties (low aggregation tendency, high homogeneity, and thermal stability). The conjugation of B9 with a potent cytotoxic agent, monomethyl auristatin E (MMAE), exhibited superior antitumor efficacy against CEACAM5-expressing human gastric cancer cell line MKN-45, human pancreatic carcinoma cell line BxPC-3 and human colorectal cancer cell line LS174T with IC50 values of 38.14, 25.60, and 101.4 nM, respectively. In BxPC-3 and MKN-45 xenograft mice, administration of UdADC B9-MMAE (5 mg/kg, i.v.) every 2 days for 4 times markedly inhibited the tumor growth without significant change in body weight. This study may have significant implications for the design of next-generation ADCs.

Synthesis and Optimization of 1-Substituted Imidazo[4,5-c]quinoline TLR7 Agonists.

TLR7 agonists have significant therapeutic potential in a variety of oncology and autoimmune applications. We recently reported a potent TLR7 selective agonist 1 that could be delivered by antibody-drug conjugate (ADC) technology to elicit potent anticancer activity. Herein we report synthetic chemistry and structure-activity relationship studies to develop TLR7 agonists with improved potency for next-generation ADC efforts. We found that the addition of hydrophobic acyl tails to parent compound 1 generally resulted in retained or improved TLR7 agonist activity without sacrificing the permeability or the selectivity over TLR8. In contrast, the addition of a simple alkyl tail at the same position resulted in a dramatic loss in potency. Molecular modeling was performed to provide a rationale for this dramatic loss in potency. We ultimately identified compounds 17b, 16b, and 16d as highly potent TLR7 agonists that potently induced the activation of mouse macrophages and hPBMCs at low-nanomolar concentrations.

Review of Antibody-Drug Conjugates

Antibody-drug conjugates (ADCs) are target-specific conjugates that consist of a monoclonal antibody connected to a cytotoxic payload using a stable linker. These conjugates combine the highly specific targeting ability and potent killing effect to accurately and efficiently eliminate cancer cells, which has ushered in a new era of targeted therapy. Since the first approval by the United States Food and Drug Administration in 2000, ADCs have experienced rapid development, with 14 of them receiving market approval. In this review, we examine the history, molecular structure, and pharmacological principles of ADCs, describing clinical trials and discussing the current challenges and future perspectives for the development of next-generation ADCs.

Antibody drug conjugates as targeted cancer therapy: past development, present challenges and future opportunities.

Antibody drug conjugates (ADCs) are promising cancer therapeutics with minimal toxicity as compared to small cytotoxic molecules alone and have shown the evidence to overcome resistance against tumor and prevent relapse of cancer. The ADC has a potential to change the paradigm of cancer chemotherapeutic treatment. At present, 13 ADCs have been approved by USFDA for the treatment of various types of solid tumor and haematological malignancies. This review covers the three structural components of an ADC-antibody, linker, and cytotoxic payload-along with their respective structure, chemistry, mechanism of action, and influence on the activity of ADCs. It covers comprehensive insight on structural role of linker towards efficacy, stability & toxicity of ADCs, different types of linkers & various conjugation techniques. A brief overview of various analytical techniques used for the qualitative and quantitative analysis of ADC is summarized. The current challenges of ADCs, such as heterogeneity, bystander effect, protein aggregation, inefficient internalization or poor penetration into tumor cells, narrow therapeutic index, emergence of resistance, etc., are outlined along with recent advances and future opportunities for the development of more promising next-generation ADCs.

Abstract 1538: Revisiting the dogma of antibody drug conjugates (ADCs): Emerging data challenge the benefit of linker stability and the primacy of payload delivery

Abstract Antibody drug conjugates (ADCs) are an effective class of cancer therapeutics which have become more prominent after eight FDA approvals in the past three years. ADCs are envisioned to enable the selective delivery of a potent drug to cells that express the antibody target while sparing normal tissues, and thus to improve the therapeutic window by both increasing the maximum tolerated dose (MTD) and decreasing the minimum efficacious dose (MED) of the conjugated drug. Mounting clinical evidence does not support this simplified paradigm [1]. Herein we focus on three prominent issues in ADC development: 1. Alternative mechanisms over and above direct tumor targeting are likely to contribute to ADC therapeutic benefits in the clinic [2]. The responses to certain ADCs in patients with different levels of target antigen expression reveal some target-independent antitumor effects, suggesting the possibility that circulating payload may contribute to efficacy. 2. The preclinical optimization of ADCs is based on efficacy and tolerability in non-human animal studies. Fundamental differences in biology and translatability between species potentially overemphasize the importance of linker stability. 3. Clinical data, although limited, does not support the idea that increased ADC stability drives substantial therapeutic benefit. None of the 13 approved ADCs are completely ‘stable’ in circulation, and more stable drug-linkers have been associated with alternative toxicity profiles and no significant improvement in MTD. Novel meta-analyses and case studies assembled and integrated from &amp;gt;40 approved or clinically active ADCs and &amp;gt;70 discontinued ADCs will be presented in support of these three perspectives. The ADCs included in the analysis differ in terms of payload class, linker type, conjugation chemistry, drug-to-antibody ratio, and antibody target. Gaining a better understanding of the mechanism(s) of action of effective ADCs and the interrelationship between ADC structural components and their pharmacokinetics and pharmacodynamics is critical to inform the next generation of ADCs.

Abstract 5653: LCB97, an antibody drug conjugate (ADC) targeting a novel tumor antigen for the treatment of multiple solid tumors

Abstract LCB97 is an antibody-drug conjugate (ADC) directed against a novel tumor antigen, composed of monomethyl auristatin E (MMAE) as payload (DAR 4) and the humanized IgG1 antibody AFF4 generated by Elthera. AFF4 targets a novel antigen preferentially expressed in various solid tumors including pancreatic cancer, ovarian cancer, colon cancer, breast cancer and melanoma. LCB97 generated by using LCB’s ConjuAllTM technology with AFF4 showed potent cytotoxicity in various cancer cell lines including breast cancer (BC), ovarian cancer (OC), pancreatic cancer (PaC), small cell lung cancer (SCLC), colorectal cancer (CRC) and melanoma. LCB97 also showed excellent anticancer efficacy in several cell line-derived xenograft (CDX) models, including BC, CRC, OC and melanoma. Preliminary toxicity assessment in cynomolgus monkeys (an AFF4 cross reactive species) demonstrated that LCB97 was well tolerated within the estimated therapeutic index of ~20 for repeat dosing. In conclusion, LCB97, generated by combining theAFF4 monoclonal antibody of Elthera and the highly innovative ConjuAllTM platform technology of LegoChem Biosciences is a promising next-generation ADC for the treatment of various solid cancers, including BC, CRC, melanoma, OC and PaC. Citation Format: Yeojin Jeong, Gunther Spohn, Flavio Mehli, Chul-Woong Chung, Changsik Park. LCB97, an antibody drug conjugate (ADC) targeting a novel tumor antigen for the treatment of multiple solid tumors. [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 5653.

Computational pathology–based HER2 quantification to identify novel biomarkers in gastric cancer (GC).

449 Background: Trastuzumab deruxtecan (T-DXd) is an antibody-drug conjugate (ADC) approved for treatment of HER2-positive GC/gastroesophageal junction adenocarcinoma (GEJA) after chemotherapy (Japan) and for advanced/metastatic GC/GEJA after a trastuzumab-containing regimen (US/Israel/Singapore). Conventional HER2 immunohistochemistry (IHC) scoring identifies potential responders to trastuzumab but may not be suitable for next-generation ADCs capable of bystander killing, such as T-DXd [Modi NEJM 2022]. Use of computational pathology–based tools enables precise and accurate quantification of HER2 expression and spatial analysis, which enables design of better scores to reflect the T-DXd mechanism of action. Methods: Quantitative continuous scoring (QCS) was performed to quantify HER2 expression and distribution per cell in digitized images of HER2-stained (IHC, Ventana 4B5) GC. A QCS-derived signature was identified, called the continuous spatial proximity score (cSPS), which considers spatial location of tumor cells in addition to HER2 expression level and can be used as a surrogate for potential bystander activity. We retrospectively stratified patients in DESTINY-Gastric01 (DG-01; NCT03329690; T-DXd arm: n=123a; standard of care [SOC] arm: n=59a) using cSPS to give a binary QCS biomarker positive (BM+) or negative (BM−) status. QCS cSPS signature was independently validated on the DESTINY-Gastric02 cohort (DG-02; NCT04014075; n=71a). Results: In the DG-01 cohort, enrolled per HER2-positive status by IHC using ASCO/CAP GC scoring guidelines, median progression-free survival (mPFS) was 5.5a mo with T-DXd and 2.8a mo with SOC. QCS-derived cSPS scores showed significant survival benefit for BM+ vs BM− in the DG-01 T-DXd arm ( P&lt;0.0001; HR=0.37 [0.24-0.58]) and longer mPFS for BM+ (8.3 mo) vs BM− (3.9 mo). Within the SOC arm, this signature showed the inverse prognosis ( P=0.034; HR=2.17 [1.06-4.44]; mPFS, 2.8 mo for BM+ vs 4.9 mo for BM−), strongly suggesting the signature is predictive of T-DXd response. This signature was validated in the DG-02 cohort (mPFS, 5.5 mo on enrollment by IHC status, 10.1 mo for BM+ vs 3.7 mo for BM−; HR=0.24 [0.13-0.45]; P&lt;0.0001). Conclusions: We established QCS, a novel scoring method for tissue biomarkers. Our validated data and results support possible future use of this quantitative approach against manual scoring for identification of patients with GC who may benefit from T-DXd therapy. Clinical trial information: NCT03329690 , NCT04014075 . [Table: see text]

VIP943 Is a Novel CD123 Antibody Drug Conjugate with in Vitro and In Vivo Efficacy in Acute Myeloid Leukemia (AML) Models

Background: Current treatment options (cytarabine with anthracyclines) for patients with acute myeloid leukemia (AML) are often associated with severe and barely tolerable toxicities. In patients who tolerate induction therapy, the complete remission rate is 60%-80%; however, 50% of patients relapse due to the presence of leukemic stem cells (LSC). Antibody-drug conjugates (ADCs) are putatively designed to deliver potent drugs directly to cancer cells while sparing normal tissue; however, first-generation ADCs are associated with severe bone marrow and liver toxicity. VIP943 is a CD123-targeting ADC which uses a kinesin spindle protein inhibitor (KSPi) as a new payload class and a unique legumain linker designed to overcome liabilities of currently approved ADC therapies. The payload is delivered with a drug-antibody ratio of 6. It consists of the KSPi warhead which is modified with a CellTrapperTM moiety so it accumulates within the CD123-targeted tumor cell and cannot enter normal cells. Non-human primate toxicology studies demonstrated a highly favorable and differentiated safety profile for VIP943 (Kirchhoff et al, Cancers 2020). Here we characterize VIP943 in various patient-derived AML models. Methods: For co-localization experiments, MOLM-13 AML cell line was stained with labelled anti-CD123 antibody or isotype control and CytoPainter LysoGreen indicator reagent and analyzed by imaging flow cytometry. AML primary patient mononuclear cells from blood and bone marrow were treated with VIP943 for 24, 48, and 72 hours. Flow cytometry immunophenotyping for AML blasts and CD34+/CD38- LSCs was used to analyze these populations. An in vivo patient-derived AML PDX mouse model was treated with VIP943 (5 mg/kg IV every 7 days) or in combination with 5-azacytidine (2.5 mg/kg SC days 1-5 x 3) and venetoclax (50 mg/kg PO days 1-5 x 3). Cytokine release syndrome was analyzed comparing VIP943 to isotype control ADC, anti-CD123 antibody, and positive comparator antibodies by in vitro cytokine release assays (CRA) in two different assay formats using whole blood and isolated mononuclear cells from 10 healthy donors. Flow cytometric lymphocyte and leukemic immunophenotyping was performed from blood of non-human primates. A leukemia cell line panel drug screen with VIP943 was evaluated and baseline genomic features were correlated with response. Results: Co-localization studies confirm specific internalization and efficient intracellular trafficking to the lysosome of VIP943. In vitro treatment with VIP943 (66 nM, continuous exposure) delivers blast reductions of 37% after a 24h incubation with VIP943, 45% after 48h incubation and 83% after an 72h incubation. In the CD34+/CD38- cell population, a reduction of 71% was achieved. In a patient-derived AML model, VIP943 with 5-azacytidine and venetoclax led to complete tumor growth inhibition after 21 days showing superiority of this triplet therapy (complete regression: 8/9 mice) with high tolerability (maximum body weight loss -4.8 %). The immunotoxicological evaluation in both CRA formats and with the tested antibody concentrations (100µg, 10µg, 1µg) exerts no induction of cytokines by VIP943 compared to controls. Immunophenotyping demonstrates no impact of VIP943 treatment on lymphocytic subpopulations, whereas the CD123-positive basophils are reduced after 24h followed by an almost complete recovery. VIP943 IC50 ranges from 0.1nM-300nM in a leukemia cell line panel screen and features associated with sensitivity are investigated. Conclusions: VIP943 is a next-generation ADC with a differentiated safety profile from currently approved ADCs, including lack of in vitro cytokine release. In vitro and in vivo studies using patient-derived AML cells show VIP943 has favorable monotherapy and combination efficacy including targeting of leukemic stem cells, which drive relapse. These findings warrant evaluating VIP943 in clinical trials.