Antibody-drug Conjugates Technology Research Articles

Abstract CT155: Trial in progress: A phase Ia/Ib, an open-label, multicenter study of ZL-1310 to evaluate the safety, tolerability, and pharmacokinetics in subjects with small cell lung cancer

Abstract Background: ZL-1310, a novel antibody-drug conjugate (ADC) targeting delta-like protein 3 (DLL3) with a camptothecin derivative as its payload via a protease-cleavable linker, employs TMALIN® (Tumor Microenvironment Activable LINker-payload) ADC technology platform. DLL3, an inhibitory Notch pathway ligand and a validated target for directed therapy, is highly expressed in more than 80% of patients with small cell lung cancer (SCLC) and also has variable expression in other neuroendocrine (NE) tumors. There remains an unmet medical need for 2nd-line SCLC, as the standard of care chemotherapy has a modest response rate of ~25%. ZL-1310 is designed to minimize payload toxicity and improve anti-tumor effectiveness. Methods: This is an open-label, multiple-dose, phase 1 study of ZL-1310 administered to subjects with relapsed/refractory (r/r) metastatic SCLC who have progressed after at least one platinum-based chemotherapy regimen. The study consists of two parts: Part 1 (dose escalation) and Part 2 (dose expansion). In Part 1, the Bayesian optimal interval (BOIN) design is employed for dose escalation, while in Part 2, patients will be randomized into two cohorts with different doses of ZL-1310 to further define safety and preliminary antitumor activity. The major inclusion criteria are as follows: (1) Subjects must have histologically or cytologically confirmed metastatic or extensive-stage SCLC with no more than 3 prior regimens in the r/r setting. (2) Subjects must have at least one measurable target lesion as defined by RECIST v1.1. (3) Eastern Cooperative Oncology Group (ECOG) performance status of 0 or 1. The major exclusion criteria include: (1) Prior exposure to DLL3-targeted therapy. (2) Subjects with symptomatic or uncontrolled brain metastasis requiring concurrent treatment, impaired major organ functions, active autoimmune disease, or active infections.Part 1 Dose Escalation is recruiting subjects in the US and China. Citation Format: Afshin Dowlati, Alexander Spira, Xiaodong Shen, Yinjia Fu, Yiyuan Pan, Linda Liu, Renke Zhou, Yi-Long Wu. Trial in progress: A phase Ia/Ib, an open-label, multicenter study of ZL-1310 to evaluate the safety, tolerability, and pharmacokinetics in subjects with small cell lung cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 2 (Late-Breaking, Clinical Trial, and Invited Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(7_Suppl):Abstract nr CT155.

Abstract 4689: 4HF CancerDataMiner platform accelerates target discovery and evaluation

Abstract For the management of metastasized cancer, tumor-targeted chemotherapies with improved therapeutic index are urgently needed. Over the last 25 years, antibody drug conjugates (ADCs) targeting cancer-associated proteins have emerged as a promising new treatment modality. While the first ADCs were registered for the treatment of acute leukemias and lymphomas, more recently at least six ADCs were approved by the FDA for solid tumors. Despite these successes, clinical development of ADCs remains a risky exercise as evident from the failure of more than 20 pivotal clinical phase III trials. A narrow therapeutic index due to excessive toxicity of potent payloads, little therapeutic activity, and difficulties in chemistry, manufacturing, and controls were among the limiting factors. Meanwhile, ADC technology and production have advanced substantially. More than 100 ADCs are currently in clinical development in approximately 600 clinical trials, with some of the investigated ADCs being close to approval. While issues related to ADC linkers and payloads have been extensively investigated and many problems have been solved, the choice of molecular targets for ADCs remains challenging. This is, in part, due to a lack of information about candidate targets: while knowledge about generally accepted cancer targets such as ERBB2 is abundant, data about many novel targets is still scarce. In particular, reliable information about expression in normal tissues, subcellular localization and protein function is often missing. 4HF Biotec has established the CancerDataMiner, an in-silico platform dedicated to cancer data mining by integrating large OMICS datasets that are connected and aligned. It allows molecular analyses of all human genes in >12,000 tumor samples and >14,000 normal tissues, using proprietary tools and widgets for visualization and statistics. Target evaluations are performed at the DNA, RNA and protein levels and employ complementary tools such as pathway analysis and literature search to obtain a comprehensive picture of candidate targets. Using this approach, we recently reviewed the molecular targets of all ADCs that either have already received approval or are in clinical development. As proof-of-concept for the CancerDataMiner, here we analyze targets of ADCs and compare our findings with the clinical data obtained for those ADCs. The profiles of selected ADC targets will be discussed, among them CD79B and CD33 for hematologic malignancies and FOLR1 and Claudin 18.2 for solid cancers. We will demonstrate that proteins with very diverse profiles can make good ADC targets. In conclusion, the 4HF CancerDataMiner is a platform which has proven its value for the identification and characterization of cancer-associated targets for ADC development for all tumor types. Citation Format: Heinz-Herbert Fiebig, Anne-Lise Peille, Mirko Schmitz, Vincent Vuaroqueaux, Thomas Metz. 4HF CancerDataMiner platform accelerates target discovery and evaluation [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 4689.

Abstract 2623: Enabling 5T4 for targeted cancer therapy: TUB-030, a novel ADC built with ethynylphosphonamidate conjugation chemistry, shows long-lasting anti-tumor activity via Topoisomerase-I inhibition with an optimized therapeutic index

Abstract TUB-030 is a novel Antibody-Drug Conjugate (ADC) targeting the oncofetal 5T4 antigen, a glycoprotein, that is functionally involved in regulation of the Wnt signalling pathway and promoting cellular motility and adhesion. Immunohistochemical analysis identified 5T4 overexpressed in many types of cancers, including bladder, lung, breast, stomach, esophagus, head and neck, colon, and ovarian. Analysis in fresh frozen healthy human tissue with the therapeutic mAb candidate of TUB-030 revealed limited cross reactivity. Ethynylphosphonamidate conjugation chemistry was used to build TUB-030 with a homogenous DAR (drug-antibody-ratio) of 8 and the Topoisomerase I inhibitor Exatecan as payload, which is connected to a humanized, Fc-silenced IgG1 antibody targeting 5T4 using a cleavable linker system. Overall, TUB-030 contrasts previous ADC designs targeting 5T4, implementing several layers of differentiation in terms of antibody, payload and conjugation technology. In vitro, TUB-030 is characterized by sub-nanomolar affinity to 5T4, efficient target-mediated internalization and strong cytotoxicity towards cancer cells from different tumor indications with a broad range of 5T4 expression levels. Differently to previous 5T4-targeting ADCs, TUB-030 has efficient bystander activity against co-cultured target-negative cells, which facilitates targeting of tumors with heterogenous 5T4 expression. Pharmacokinetic analysis showed that TUB-030 is highly stable - and does not lose or transfer its linker-payload to serum proteins during blood circulation and antibody-like clearance profile contrasting maleimide-conjugated ADCs and enabling most efficient and continued delivery of Exatecan to the tumor site. Single-dose treatment with TUB-030 results in long-lasting tumor regression with high complete remission rates across a variety of both cell-line derived xenograft (CDX) and patient-derived xenograft (PDX) cancer models of multiple tumor indications in vivo, including models with low target expression. Preliminary repeat-dose toxicological assessment of TUB-030 in a pharmacologically relevant non-human primate species demonstrated that TUB-030 is well tolerated. The preclinical results underline that TUB-030 with novel ADC technology has the potential to enable 5T4 as a therapeutic target and provide treatment options in numerous solid cancer indications with high unmet medical need supporting further development towards the clinic. Citation Format: Saskia Schmitt, Isabelle Mai, Paul Machui, Sarah Herterich, Danila Hauswald, Philipp Ochtrop, Philipp Cyprys, Izabela Kozlowska, Annabel Kitowski, Marcus Gerlach, Olivier Marcq, Pamela A. Trail, Dominik Schumacher, Marc-André Kasper, Günter Fingerle-Rowson, Björn Hock, Jonas Helma-Smets, Annette M. Vogl. Enabling 5T4 for targeted cancer therapy: TUB-030, a novel ADC built with ethynylphosphonamidate conjugation chemistry, shows long-lasting anti-tumor activity via Topoisomerase-I inhibition with an optimized therapeutic index [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 2623.

Abstract 2051: Innovations in ADC technology platform with legumain-cleavable KSP-inhibitor payloads adaptable to various aspects of cancer biology

Abstract Despite recent advances in the antibody-drug conjugate (ADC) field it remains challenging to achieve a sufficient therapeutic window. To address key issues of current ADCs such as aggregate formation and payload-related toxicities, we have developed a platform technology with tailored solutions for ADCs and small molecule-drug conjugates (SMDCs). Key features of the ADC technology comprise of kinesin spindle protein inhibitors (KSPi) as a novel payload class, a unique linker selectively cleaved by the tumor-associated protease legumain, and a CellTrapper® modification of the KSPi payload to reduce membrane permeability and provide accumulation in tumor cells beneficial for efficacy and safety1,2. Our advanced ADC VIP943 is targeting CD123 for the treatment of AML and MDS and is currently in Phase 1 clinical trial (NCT06034275). Here we show the extension and applicability of the platform in combination with various antibodies. Two case studies demonstrate how the payload-linker optimization increases the selectivity for tumor over healthy cells and potentially overcomes transporter-mediated drug resistance. Tuning the cleavable linker of cetuximab-ADCs for a higher challenge of legumain-mediated payload release resulted in a 52-fold improved selectivity of cytotoxicity against tumor cells versus HaCaT skin cells. A trastuzumab-ADC using a KSPi payload modified with the CellTrapper, shows low nanomolar potency against JIMT-1 cells resistant to trastuzumab-deruxtecan and thus may overcome resistance. An increased potency was not observed with an ADC releasing a permeable KSPi without CellTrapper. To address heterogenous target expression in solid tumors, novel antibody-2-drug conjugates (A2DCs) were synthesized to combine the advantages of intracellular accumulation of non-permeable payloads with the release of a second, membrane permeable KSPi payload. In vitro studies comparing the A2DCs with their respective mono-drug ADCs on 4 different targets shows the highest cytotoxicity achieved with A2DCs independent of the efficacy driver in mono ADCs. Lastly, synthesis of homogenous KSPi ADCs with a drug to antibody ratio of ~8 is compatible with different antibodies without significant aggregate formation. Furthermore, an easy and efficient process for ring-opening of the thiosuccinimide ring after conjugation was developed and shown to successfully avoid ADC de-conjugation via retro-Michael reaction. Selected examples of each of these subclasses were coupled to trastuzumab; all conjugates were highly efficacious and resulted in tumor regressions in the SKOV-3 model similar to trastuzumab-deruxtecan. Conclusion: Our technology platform offers options for efficient and safe cancer treatments and allows for versatile adaptations to the various aspects of cancer biology. 1. Kirchhoff et al, Cancers (2020). 2. Lerchen et al, Bioconj Chem. (2020). Citation Format: Hans-Georg Lerchen, Anne-Sophie Rebstock, Mareike Wiedmann, Beatrix Stelte-Ludwig, Amy J. Johnson, Raquel Izumi, Ahmed Hamdy. Innovations in ADC technology platform with legumain-cleavable KSP-inhibitor payloads adaptable to various aspects of cancer biology [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 2051.

Abstract 1911: Dato-DXd mediates anti-tumor activity in preclinical TROP2-expressing intracranial tumor model

Abstract 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.

Generation of a Novel SORT1×HER2 Bispecific Antibody-Drug Conjugate Targeting HER2-Low-Expression Tumor.

Human epidermal growth factor receptor 2 (HER2) is considered an ideal antibody-drug conjugate (ADC) target because the gene is overexpressed in many tumors compared to normal tissues. Multiple anti-HER2 ADCs conjugated with different toxic payloads bring benefits to patients with high HER2 expression. However, HER2-targeted ADC technology needs further optimization to improve its effect for the treatment of patients with low HER2 expression. We hypothesized that bispecific antibody-drug conjugate (bsADC) targeting HER2 and Sortilin-1 (SORT1) would overcome this limitation. SORT1 is a suitable target for pairing with HER2 to generate a bispecific antibody (BsAb) since the gene is co-expressed with HER2 in tumors and possesses rapid internalization. We developed a BsAb (bsSORT1×HER2) that exhibited strong binding and internalization activity on HER2-low-expression tumor cells and facilitated higher HER2 degradation. The bsSORT1×HER2 was further conjugated with DXd to generate a bsADC (bsSORT1×HER2-DXd) that showed strong cytotoxicity on HER2-low-expression tumor cells and antitumor efficacy in an MDA-MB-231 xenograft mice model. These results demonstrated that employment of a SORT1×HER2-targeted bsADC may be promising to improve the antitumor efficacy of HER2-targeted ADC for the treatment of tumors with low HER2 expression.

Business Risk Mitigation in the Development Process of New Monoclonal Antibody Drug Conjugates for Cancer Treatment.

Recent developments aim to extend the cytotoxic effect and therapeutic window of mAbs by constructing antibody-drug conjugates (ADCs), in which the targeting moiety is the mAb that is linked to a highly toxic drug. According to a report from mid of last year, the global ADCs market accounted for USD 1387 million in 2016 and was worth USD 7.82 billion in 2022. It is estimated to increase in value to USD 13.15 billion by 2030. One of the critical points is the linkage of any substituent to the functional group of the mAb. Increasing the efficacy against cancer cells' highly cytotoxic molecules (warheads) are connected biologically. The connections are completed by different types of linkers, or there are efforts to add biopolymer-based nanoparticles, including chemotherapeutic agents. Recently, a combination of ADC technology and nanomedicine opened a new pathway. To fulfill the scientific knowledge for this complex development, our aim is to write an overview article that provides a basic introduction to ADC which describes the current and future opportunities in therapeutic areas and markets. Through this approach, we show which development directions are relevant both in terms of therapeutic area and market potential. Opportunities to reduce business risks are presented as new development principles.

Design and Characterization of Immune-Stimulating Imidazo[4,5-c]quinoline Antibody-Drug Conjugates.

Traditional antibody-drug conjugate (ADC) technology has employed tumor-targeting antibodies to selectively deliver ultrapotent cytotoxins to tumor tissue. While this technology has been highly successful, resulting in the FDA approval of over 10 ADCs, the field continues to struggle with modest efficacy and significant off-target toxicity. Concurrent with the struggles of the ADC field, a new generation of immune-activating therapeutics has arisen, most clearly exemplified by the PD-1/PD-L1 inhibitors that are now part of standard-of-care treatment regimens for a variety of cancers. The success of these immuno-oncology therapeutic agents has prompted the investigation of a variety of new immuno-stimulant approaches, including toll-like receptor (TLR) activators. Herein, we describe the optimization of ADC technology for the selective delivery of a potent series of TLR7 agonists. A series of imidazole[4,5-c]quinoline agonists (as exemplified by compound 1) were shown to selectively agonize the human and mouse TLR7 receptor at low nanomolar concentrations, resulting in the release of IFNα from human peripheral blood mononuclear cells (hPBMCs) and the upregulation of CD86 on antigen-presenting cells. Compound 1 was attached to a deglycosylated (Fc-γ null) HER2-targeting antibody via a cleavable linker, resulting in an ADC (anti-HER2_vc-1) that potently and selectively activated the TLR7 pathway in tumor-associated macrophages via a "bystander" mechanism. We demonstrated that this ADC rapidly released the TLR7 agonist into the media when incubated with HER2+ cells. This release was not observed upon incubation with an isotype control ADC and furthermore was suppressed by co-administration of the naked antibody. In co-culture experiments with HER2+ HCC1954 cells, this ADC induced the activation of the NFκB pathway in mouse macrophages and the release of IFNα from hPBMCs, while a corresponding isotype control ADC did not. Finally, we demonstrated that IP administration of anti-HER2_vc-1 induced complete tumor regression in an HCC1954 xenograft study in SCID beige mice. Unlike related ADC technology that has been reported recently, our technology relies on the passive diffusion of the TLR7 agonist into tumor-associated macrophages rather than Fc-γ-mediated uptake. Based on these observations, we believe that this ADC technology holds significant potential for both oncology and infectious disease applications.

Novel development strategies and challenges for anti-Her2 antibody-drug conjugates.

Antibody-drug conjugates (ADCs) combining potent cytotoxicity of small-molecule drugs with the selectivity and excellent pharmacokinetic profile of monoclonal antibody (mAb) are promising therapeutic modalities for a diverse range of cancers. Owing to overexpression in a wide range of tumors, human epidermal growth factor receptor 2 (Her2) is one of the most utilized targeting antigens for ADCs to treat Her2-positive cancers. Owing to the high density of Her2 antigens on the tumor cells and high affinity and high internalization capacity of corresponding antibodies, 56 anti-Her2 ADCs which applied >10 different types of novel payloads had entered preclinical or clinical trials. Seven of 12 Food and Drug Administration (FDA)-approved ADCs including Polivy (2019), Padcev (2019), EnHertu (2019), Trodelvy (2020), Blenrep (2020), Zynlonta (2021), and Tivdak) (2021) have been approved by FDA in the past three years alone, indicating that the maturing of ADC technology brings more productive clinical outcomes. This review, focusing on the anti-Her2 ADCs in clinical trials or on the market, discusses the strategies to select antibody formats, the linkages between linker and mAb, and effective payloads with particular release and action mechanisms for a good clinical outcome.

Optimizing anti–body drug conjugates and radiopharmaceuticals for precision therapy: The next frontier in precision oncology

Drug delivery to select therapeutic targets for precision oncology has always been a major challenge in oncology. Anti–body drug conjugates (ADC) and radiopharmaceuticals attempt to overcome the major limitations of chemotherapy or mono-clonal antibody therapy as they provide a highly specific means of attacking the vulnerability of the cancer cell. Currently there are over 100 clinical trials in various stages of development with these agents. Years of improved understanding of ADC technology and addressing previous failures have yielded the current pipeline of ADCs with better design. The near future of ADC developmental therapeutics includes firstly continuing to improve drug design, secondly picking right targets, thirdly balancing efficacy vs toxicity, fourth understanding innate and acquired resistance mechanisms, fifth development of ADCs across pan-cancers as tissue-agnostic therapeutics, and lastly combination strategies. There is considerable excitement in the ADC developmental therapeutics and ADCs including radiopharmaceuticals are here to stay and offer to expand our pipeline of effective drugs to combat cancer.

Marine Antitumor Peptide Dolastatin 10: Biological Activity, Structural Modification and Synthetic Chemistry.

Dolastatin 10 (Dol-10), a leading marine pentapeptide isolated from the Indian Ocean mollusk Dolabella auricularia, contains three unique amino acid residues. Dol-10 can effectively induce apoptosis of lung cancer cells and other tumor cells at nanomolar concentration, and it has been developed into commercial drugs for treating some specific lymphomas, so it has received wide attention in recent years. In vitro experiments showed that Dol-10 and its derivatives were highly lethal to common tumor cells, such as L1210 leukemia cells (IC50 = 0.03 nM), small cell lung cancer NCI-H69 cells (IC50 = 0.059 nM), and human prostate cancer DU-145 cells (IC50 = 0.5 nM), etc. With the rise of antibody-drug conjugates (ADCs), milestone progress was made in clinical research based on Dol-10. A variety of ADCs constructed by combining MMAE or MMAF (Dol-10 derivatives) with a specific antibody not only ensured the antitumor activity of the drugs themself but also improved their tumor targeting and reduced the systemic toxicity. They are currently undergoing clinical trials or have been approved for marketing, such as Adcetris®, which had been approved for the treatment of anaplastic large T-cell systemic malignant lymphoma and Hodgkin lymphoma. Dol-10, as one of the most medically valuable natural compounds discovered up to now, has brought unprecedented hope for tumor treatment. It is particularly noteworthy that, by modifying the chemical structure of Dol-10 and combining with the application of ADCs technology, Dol-10 as a new drug candidate still has great potential for development. In this review, the biological activity and chemical work of Dol-10 in the advance of antitumor drugs in the last 35 years will be summarized, which will provide the support for pharmaceutical researchers interested in leading exploration of antitumor marine peptides.

Dolaflexin: A Novel Antibody-Drug Conjugate Platform Featuring High Drug Loading and a Controlled Bystander Effect.

After significant effort over the last 30 years, antibody-drug conjugates (ADC) have recently gained momentum as a therapeutic modality, and nine ADCs have been approved by the FDA to date, with additional ADCs in late stages of development. Here, we introduce dolaflexin, a novel ADC technology that overcomes key limitations of the most common ADC platforms with two key features: a higher drug-to-antibody ratio and a novel auristatin with a controlled bystander effect. The novel, cell permeable payload, auristatin F-hydroxypropylamide, undergoes metabolic conversion to the highly potent, but less cell permeable auristatin F to balance the bystander effect through drug trapping within target cells. We conducted studies in mice, rats, and cynomolgus monkeys to complement in vitro characterization and contrasted the performance of dolaflexin with regard to antitumor activity, pharmacokinetic properties, and safety in comparison with the ADC platform utilized in the approved ADC ado-trastuzumab emtansine (T-DM1). A HER2-targeted dolaflexin ADC was shown to have a much lower threshold of antigen expression for potent cell killing in vitro, was effective in vivo in tumors with low HER2 expression, and induced tumor regressions in a xenograft model that is resistant to T-DM1.

Overcoming Challenges in the Development of Anticancer ADCs

Overcoming Challenges in the Development of Anticancer ADCs

Antibody-Drug Conjugates: Patient and Treatment Selection

Antibody-drug conjugates (ADCs) are a promising drug platform designed to enhance the therapeutic index and minimize the toxicity of anticancer agents. ADCs have experienced substantial progress and technological growth over the past decades; however, several challenges to patient selection and treatment remain. Methods to optimally capture all patients who may benefit from a particular ADC are still largely unknown. Although target antigen expression remains a biomarker for patient selection, the impact of intratumor heterogeneity on antigen expression, as well as the dynamic changes in expression with treatment and disease progression, are important considerations in patient selection. Better understanding of these factors, as well as minimum levels of target antigen expression required to achieve therapeutic efficacy, will enable further optimization of selection strategies. Other important considerations include understanding mechanisms of primary and acquired resistance to ADCs. Ongoing efforts in the design of its constituent parts to possess the intrinsic ability to overcome these mechanisms, including use of the "bystander effect" to enhance efficacy in heterogeneous or low target antigen-expressing tumors, as well as modulation of the chemical and immunophenotypic properties of antibodies and linker molecules to improve payload sensitivity and therapeutic efficacy, are under way. These strategies may also lead to improved safety profiles. Similarly, combination strategies using ADCs with other cytotoxic or immunomodulatory agents are also under development. Great strides have been made in ADC technology. With further refinements, this therapeutic modality has the potential to make an important clinical impact on a wider range of tumor types.

Proof of site-specificity of antibody-drug conjugates produced by chemical conjugation technology: AJICAP first generation

Antibody-drug conjugates (ADCs) have become major biopharmaceutical drugs in the field of oncology. Traditional ADCs possess a stochastic distribution of cytotoxic payloads linked to several different amino acid residues of the antibody. This heterogeneous nature of stochastic ADCs results in a complex conjugation-site characterization. To improve upon traditional ADC technology, we have developed a chemical conjugation platform, termed AJICAP™, for site-specific modification of native antibodies using a class of IgG Fc-affinity reagents (Yamada et al., 2019). Here, we report further investigation focusing on peptide mapping of the AJICAP™-ADC to confirm the exact conjugation position of the first generation AJICAP™-ADC. Neutral pH pretreatment for peptide mapping prevented undesired PTMs such as succinimide ring hydrolysis. Mirroring comparison using the purified ADC visibly indicated that Lys248 in the Fc region was conjugated to the drug-linker. MS/MS analysis also provided evidence to support Lys248 conjugation. Finally, extracted ion-chromatogram methodology suggested the site-specificity of AJICAP™ conjugation. Purified ADCs by preparative HIC-HPLC showed clear visual results and more than 93% sequence coverage by a single enzymatic digestion. The analytical strategy described herein demonstrated a robust analytical methodology for revealing the conjugation site of ADCs.

ADCs, as Novel Revolutionary Weapons for Providing a Step Forward in Targeted Therapy of Malignancies.

Antibody drug conjugates (ADCs), as potent pharmaceutical trojan horses for cancer treatment, provide superior efficacy and specific targeting along with low risk of adverse reactions compared to traditional chemotherapeutics. In fact, the development of these agents combines the selective targeting capability of monoclonal antibody (mAb) with high cytotoxicity of chemotherapeutics for controlling the neoplastic mass growth. Different ADCs (more than 60 ADCs) in preclinical and clinical trials were introduced in this novel pharmaceutical field. Various design-based factors must be taken into account for improving the functionality of ADC technology, including selection of appropriate target antigen and high binding affinity of fragment (miniaturized ADCs) or full mAbs (preferentially use of humanized or fully human antibodies compared to murine and chimeric ones), use of bispecific antibodies for dual targeting effect, linker engineering and conjugation method efficacy to obtain more controlled drug to antibody ratio (DAR). Challenging issues affecting therapeutic efficacy and safety of ADCs, including bystander effect, on- and off-target toxicities, multi drug resistance (MDR) are also addressed. 4 FDA-approved ADCs in the market, including ADCETRIS ®, MYLOTARG®, BESPONSA ®, KADCYLA®. The goal of the current review is to evaluate the key parameters affecting ADCs development.

Comparison of Analytical Methods for Antibody–Drug Conjugates Produced by Chemical Site-Specific Conjugation: First-Generation AJICAP

Antibody-drug conjugates (ADCs) have become a major class of oncology biopharmaceuticals. Traditional ADCs have a stochastic distribution of cytotoxic drugs attached at several different sites on the antibody. The heterogeneous nature of stochastic ADCs results in a complex compositional analysis. To improve on traditional ADC technology, we have developed a chemical conjugation platform termed "AJICAP" for the site-specific modification of native antibodies using a class of IgG Fc affinity reagents. Here we report further investigation focusing on several analyses of a first-generation AJICAP-ADC (Angew. Chem., Int. Ed. 2019, 58, 5592-5597). For drug-antibody ratio (DAR) determination, we examined and compared six different analytical methods. To the best of our knowledge, this is the first report of a comparison of analytical techniques to measure the DAR for ADCs produced by a site-specific technology such as AJICAP. Furthermore, a rapid analytical process for confirmation of the site selectivity of AJICAP conjugation was established by SEC-Q-TOF-MS. The analytical strategy reported here can be applied to the DAR determination of site-specific ADCs.

Potential mechanisms of target-independent uptake and toxicity of antibody-drug conjugates.

Antibody-drug conjugates (ADCs) are a promising therapeutic modality for oncology indications. The concept of an ADC platform is to increase the therapeutic index (TI) of chemotherapeutics through more selective delivery of cytotoxic agents to tumor cells while limiting exposure to healthy normal cells. Despite the use of antibodies targeting antigens abundantly and/or exclusively expressed on cancer cells (i.e., target cells), dose limiting toxicities (DLTs) in normal cells/tissues are frequently reported even at suboptimal therapeutic doses. Although advancement of ADC technology has helped to optimize all three key components (i.e., mAb, linker, and payload), DLTs remain a key challenge for ADC development. Mechanisms of ADC toxicity in normal cells/tissues are not clearly understood, but the majority of DLTs are considered to be target-independent. In addition to linker-drug instability contributing to the premature release of cytotoxic drug (payload) in circulation, uptake/trafficking of intact ADCs by both receptor-dependent (FcγRs, FcRn and C-type lectin receptors), and-independent (non-specific endocytosis) mechanisms may contribute to off-target toxicity in normal cells. In this article, we review potential mechanisms of target-independent ADC uptake and toxicity in normal cells, as well as discuss components of ADCs which may influence these mechanisms. This information will provide a deeper understanding of the underlying mechanisms of ADC off-target toxicity and prove helpful toward improving the overall TI of the next generation of ADCs.

The Latest Research and Development into the Antibody-Drug Conjugate, [fam-] Trastuzumab Deruxtecan (DS-8201a), for HER2 Cancer Therapy.

A major limitation of traditional chemotherapy for cancer is dose-limiting toxicity, caused by the exposure of non-tumor cells to cytotoxic agents. Use of molecular targeted drugs, such as specific kinase inhibitors and monoclonal antibodies, is a possible solution to overcome this limitation and has achieved clinical success so far. Use of an antibody-drug conjugate (ADC) is a rational strategy for improving efficacy and reducing systemic adverse events. ADCs use antibodies selectively to deliver a potent cytotoxic agent to tumor cells, thus drastically improving the therapeutic index of chemotherapeutic agents. Lessons learned from clinical failure of early ADCs during the 1980s to 90s have recently led to improvements in ADC technology, and resulted in the approval of four novel ADCs. Nonetheless, further advances in ADC technology are still required to streamline their clinical efficacy and reduce toxicity. [fam-] Trastuzumab deruxtecan (DS-8201a) is a next-generation ADC that satisfies these requirements based on currently available evidence. DS-8201a has several innovative features; a highly potent novel payload with a high drug-to-antibody ratio, good homogeneity, a tumor-selective cleavable linker, stable linker-payload in circulation, and a short systemic half-life cytotoxic agent in vivo; the released cytotoxic payload could exert a bystander effect. With respect to its preclinical profiles, DS-8201a could provide a valuable therapy with a great potential against HER2-expressing cancers in clinical settings. In a phase I trial, DS-8201a showed acceptable safety profiles with potential therapeutic efficacy, with the wide therapeutic index.

Hydrophilic Monomethyl Auristatin E Derivatives as Novel Candidates for the Design of Antibody-Drug Conjugates

Antibody-drug conjugates (ADCs) are promising state-of-the-art biopharmaceutical drugs for selective drug-delivery applications and the treatment of diseases such as cancer. The idea behind the ADC technology is remarkable as it combines the highly selective targeting capacity of monoclonal antibodies with the cancer-killing ability of potent cytotoxic agents. The continuous development of improved ADCs requires systematic studies on the nature and effects of warhead modification. Recently, we focused on the hydrophilic modification of monomethyl auristatin E (MMAE), the most widely used cytotoxic agent in current clinical trial ADCs. Herein, we report on the use of micellar electrokinetic chromatography (MEKC) for studying the hydrophobic character of modified MMAE derivatives. Our data reveal a connection between the hydrophobicity of the modified warheads as free molecules and their cytotoxic activity. In addition, MMAE-trastuzumab ADCs were constructed and evaluated in preliminary cytotoxic assays.