Uptake Of Antibody-drug Conjugates Research Articles

Abstract 5760: Intra-tissue quantitative analysis using LC/MS will determine the mechanisms of interstitial pneumonia induced by antibody-drug conjugates

Abstract Background: Antibody-drug conjugates (ADCs) are a new class of therapeutics that combine a monoclonal antibody with a cytotoxic agent (known as their payloads) via a synthetic linker. While ADCs are one of game changer and are the fastest growing classes with novel clinical benefits recently, interstitial pneumonia (ILD) is a clinical problem as a fatal adverse event. However, the mechanism of ADC-induced ILD is under investigation. Purpose: We clarify ADC-ILD mechanism by analyzing released payload distribution in the lung using a novel Trophoblast cell surface antigen 2 (TROP2)-targeting ADC. Material and Methods: TROP2-eribulin is a novel TROP2-targeting ADC composed of the TROP2 antibody and cytotoxic payload eribulin conjugated by a cleavable linker. Four human cancer cell lines (A549, HeLa, NCI-H2110 and SKOV3) and one human- leukemia monocytic cell line (THP-1) were used in this study. Cell surface TROP2 antigen expression was evaluated by flow cytometry. We investigated the concentration and distribution of the TROP2-eribulin and the payload released from the TROP2-eribulin in tumor and lung tissue, using TROP2 low and high expression human cancer xenograft model that were treated with various doses of TROP2-eribulin. The concentration of erbulin was analyzed by liquid chromatograph mass spectrometry, and tissue distribution of ADCs was assessed by immunofluorescence (IF) assay. Results: In vitro analysis showed that the concentration of intracellularly released eribulin was significantly higher in NCI-H2110 with high TROP2 expression compared to A549 with low TROP2 expression (P < 0.05). In vivo analysis, the concentration of released eribulin in tumor tissue was approximately 10-fold higher in the NIC-H2110 xenograft model than the A549 xenograft model, while analysis of lung tissue showed that TROP2-eribulin was distributed in lung tissue in a dose-dependent manner of TROP2-eribulin regardless of TROP2 expression, with significantly more eribulin released in the high-dose group of TROP2-eribulin than in the other dose groups (P < 0.05). IF assay showed that TROP2-eribuilin localized to alveolar macrophages. In the analysis using THP-1 cell, which mimic macrophage function, the concentration of eribulin released from TROP2-eribuilin was significantly reduced by the use of an Fc receptor inhibitor (P < 0.05). Conclusions: Nonspecific uptake of ADC by alveolar macrophages via Fcγ receptors releases cytotoxic payload into lung tissue, regardless of tumor antigen expression levels. This understanding can help elucidate the pathogenesis of ADC-induced ILD. Citation Format: Shigehiro Koganemaru, Hirobumi Fuchigami, Chihiro Morizono, Hiroko Shinohara, Yasutoshi Kuboki, Keiji Furuuchi, Toshimitsu Uenaka, Toshihiko Doi, Masahiro Yasunaga. Intra-tissue quantitative analysis using LC/MS will determine the mechanisms of interstitial pneumonia induced by antibody-drug conjugates [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 5760.

Abstract 2660: HMBD-501 - a novel Fc engineered, exatecan-based next generation HER3 targeting antibody drug conjugate shows robust efficacy and tolerability in pre-clinical solid tumor models

Abstract HER3 is a member of the epidermal growth factor receptor (ERBB) family of tyrosine kinase receptors overexpressed in a broad range of tumors and is associated with disease progression and poor survival. For example, more than 50% of melanoma, cervical, lung, gastric, colorectal, and ovarian cancers show overexpression of HER3, while in breast, pancreatic and prostate cancers the overexpression ranges between 25-40%. In addition, recent studies show that HER3 expression is significantly induced in tumors during metastasis (in colorectal) and during acquired resistance to tyrosine kinase inhibitors (TKI) in lung cancers. This widespread overexpression of HER3 makes it an ideal target for antibody drug conjugates (ADCs). ADCs are a therapeutic modality that harness an antibody’s target specificity to selectively deliver cytotoxic payloads to tumors. Patritumab-Deruxtecan (U3-1402), a HER3-targeting ADC, showed clinical response in a subset of EGFR mutant and TKI resistant lung cancers in a Phase 1 study, albeit with toxicities including severe interstitial lung disease (ILD) and high grade cytopenias. Mechanistic studies show that these ADC-related toxicities are likely driven by Fc- or micropinocytosis-mediated uptake of ADCs by normal cells. We have developed HMBD-501, a novel anti-HER3 ADC with (i) a high-affinity Fab, which can bind to HER3 in the presence or absence of its ligand NRG1, (ii) an engineered Fc-domain to avoid FcgR binding and potential toxicities, and (iii) site-specific conjugation to a next generation cleavable linker and exatecan-based payload. In our preclinical studies, HMBD-501 demonstrates superior inhibition of tumor growth compared to other exatecan-based anti-HER3 ADCs, in a variety of in vitro and in vivo cancer models with a range of HER3 expression. Additionally, the optimized Fc engineering of HMBD-501 shows enhanced tolerability in vivo indicating a potential reduction in Fc mediated toxicities. Finally, the next generation site-specific conjugation-linker-payload technology of HMBD-501 allows optimal plasma stability and pharmacokinetic profile. In conclusion, HMBD-501 represents a next generation HER3 targeting ADC that has a potentially best-in-class efficacy and tolerability profile. Our findings strongly support the development of HMBD-501 as a promising candidate in the treatment of HER3 expressing solid tumors. Citation Format: Suhail Ahmed Kabeer Rasheed, Dipti Thakkar, Shalini Paliwal, Wen Jie Chin, Akshaya Bansal, Ben Ayers, Che Theng ng, Brendon Hanson, Piers Ingram, Jerome Boyd-Kirkup. HMBD-501 - a novel Fc engineered, exatecan-based next generation HER3 targeting antibody drug conjugate shows robust efficacy and tolerability in pre-clinical solid tumor models [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 2660.

Bis‐ethynylphosphonamidates as an Modular Conjugation Platform to Generate Multi‐Functional Protein‐ and Antibody‐Drug‐Conjugates

AbstractBis‐ethynylphosphonamidates allow for a simple chemoselective addition of two thiol‐containing modules in a row. We describe four such bis‐electrophiles that carry different functional O‐substituents with tunable hydrophilicity and enable further subsequent conjugations, thus facilitating a simple protocol for constructing protein‐protein conjugates. An increased spacer length between the two ethynylphosphonamidates simplifies the formation of a conjugate from two bulky proteins. We apply these reagents to obtain homogeneous Antibody‐Drug‐Conjugates (ADCs) from DM1 and trastuzumab with excellent cytotoxicity and selectivity for the targeted cell line. Moreover, a bis‐ethynylphosphonamidate, carrying an additional alkyne for a chemoselective triple conjugation, has been subjected to fluorescent labeling of an ADC specifically at the drug site give an Antibody‐Drug‐Fluorophore‐Conjugate (ADFC), allowing for the observation of intracellular trafficking after ADC uptake into the targeted cell.

Novel antibody-drug conjugate with UV-controlled cleavage mechanism for cytotoxin release

Antibody-drug conjugates (ADCs) are being developed worldwide with the potential to revolutionize current cancer treatment strategies. However, off-target toxicity caused by the instability of linkers remains one of the main issues to be resolved. Developing a novel photocontrol-ADC with good stability and photocontrolled release seemed to be an attractive and practical solution. In this study, we designed, for the first time, a novel ultraviolet (UV) light-controlled ADC by carefully integrating the UV-cleavable o-nitro-benzyl structure into the linker. Our preliminary work indicated that the ADC exhibited good stability and photocontrollability while maintaining a targeting effect similar to that of the naked antibody. Upon irradiation with UV light, the ADC rapidly released free cytotoxins and exerted significant cytotoxicity toward drug-resistant tumor cells. Compared to those of the unirradiated cells, the EC50 values of ADCs increased by up to 50-fold. Furthermore, our research confirmed that the degradation products of unirradiated ADC, Cys-1a, were relatively less toxic, thus potentially reducing the off-target toxicity caused by nonspecific uptake of ADCs. The novel design strategy of UV light-controlled ADCs may provide new perspectives for future research on ADCs and promote the development of photocontrol systems.

Tailored Linker Chemistries for the Efficient and Selective Activation of ADCs with KSPi Payloads.

Several antibody-drug conjugates (ADCs) have failed to achieve a sufficiently large therapeutic window in patients due to toxicity induced by unspecific payload release in the circulation or ADC uptake into healthy organs. Herein, we describe the successful engineering of ADCs consisting of novel linkers, which are efficiently and selectively cleaved by the tumor-associated protease legumain. ADCs generated via this approach demonstrate high potency and a preferential activation in tumors compared to healthy tissue, thus providing an additional level of safety. A remarkable tolerance of legumain for different linker peptides, including those with just a single asparagine residue, together with a modifier of the physicochemical metabolite profile, proves the broad applicability of this approach for a tailored design of ADCs.

HER2 antibody-drug conjugate controls growth of breast cancer brain metastases in hematogenous xenograft models, with heterogeneous blood-tumor barrier penetration unlinked to a passive marker.

Brain metastases of HER2+ breast cancer persist as a clinical challenge. Many therapeutics directed at human epidermal growth factor receptor 2 (HER2) are antibodies or antibody-drug conjugates (ADCs), and their permeability through the blood-tumor barrier (BTB) is poorly understood. We investigated the efficacy of a biparatopic anti-HER2 antibody-tubulysin conjugate (bHER2-ATC) in preclinical models of brain metastases. The compound was evaluated in 2 hematogenous HER2+ brain metastasis mouse models, SUM190-BR and JIMT-1-BR. Endpoints included metastasis count, compound brain penetration, cancer cell proliferation, and apoptosis. Biparatopic HER2-ATC 3 mg/kg prevented metastasis outgrowth in the JIMT-1-BR model. At 1 mg/kg bHER2-ATC, a 70% and 92% reduction in large and micrometastases was observed. For the SUM190-BR model, an 85% and 53% reduction, respectively, in large and micrometastases was observed at 3 mg/kg, without statistical significance. Proliferation was reduced in both models at the highest dose. At the endpoint, bHER2-ATC uptake covered a median of 4-6% and 7-17% of metastasis area in the JIMT-1-BR and SUM190-BR models, respectively. Maximal compound uptake in the models was 19% and 86% in JIMT-1-BR and SUM190-BR, respectively. Multiple lesions in both models demonstrated ADC uptake in the absence or low diffusion of Texas Red Dextran, a marker of paracellular permeability. Using in vitro BTB assays, the ADC was endocytosed into brain endothelial cells, identifying a potentially new mechanism of antibody permeability. Biparatopic HER2-ATC significantly prevented JIMT-1-BR brain metastasis outgrowth and showed activity in the SUM190-BR model. The bHER2-ATC penetration into metastases that are impermeable to fluorescent dye suggested an endocytic mechanism of brain penetration.

A Systems Pharmacology Model for Drug Delivery to Solid Tumors by Antibody-Drug Conjugates: Implications for Bystander Effects.

Antibody-drug conjugates (ADCs) are cancer drugs composed of a humanized antibody linked to a cytotoxic payload, allowing preferential release of payload in cancer cells expressing the antibody-targeted antigen. Here, a systems pharmacology model is used to simulate ADC transport from blood to tumor tissue and ADC uptake by tumor cells. The model includes effects of spatial gradients in drug concentration in a three-dimensional network of tumor blood vessels with realistic geometry and accounts for diffusion of ADC in the tumor extracellular space, binding to antigen, internalization, intracellular processing, and payload efflux from cells. Cells that process an internalized ADC-antigen complex may release payload that can be taken up by other "bystander" cells. Such bystander effects are included in the model. The model is used to simulate conditions in previous experiments, showing good agreement with experimental results. Simulations are used to analyze the relationship of bystander effects to payload properties and single-dose administrations. The model indicates that exposure of payload to cells distant from vessels is sensitive to the free payload diffusivity in the extracellular space. When antigen expression is heterogeneous, the model indicates that the amount of payload accumulating in non-antigen-expressing cells increases linearly with dose but depends only weakly on the percentage of antigen-expressing cells. The model provides an integrated mechanistic framework for understanding the effects of spatial gradients on drug distribution using ADCs and for designing ADCs to achieve more effective payload distribution in solid tumors, thereby increasing the therapeutic index of the ADC.

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.

Highlights of This Issue

Deregulated cell cycle progression and gene regulation in cancer cells exposes vulnerabilities that can be exploited in developing new therapies. For example, CDK7 is an important protein kinase that controls cell cycle progression through the phosphorylation of cyclin-CDKs and transcription initiation by phosphorylating RNA polymerase II. Here Patel, Periyasamy and colleagues describe a new CDK7 inhibitor, ICEC0942 (CT7001), which effectively inhibits tumour cell growth, both as a single agent and in combination with hormonal therapies in estrogen receptor-positive breast cancer cells. Good pharmacological characteristics, including oral bioavailability, highlight potential for the clinical development of ICEC0942, with Phase I trials now underway (clinicaltrials.gov: NCT03363893).Glioblastoma is the most common and lethal form of primary brain tumor with a dismal median survival of 14.5 months. Cancer genomics studies have identified a wealth of new genetic targets, many of which amenable to silencing by oligonucleotide drugs, such as small interfering RNAs (siRNAs). Here, Osborn, Coles, and colleagues have demonstrated that administration of metabolically stabilized, cholesterol-conjugated, self-delivering siRNA (hsiRNA) supports widespread tumor distribution and highly efficient gene silencing in established, orthotopic gliomas. Direct delivery of hsiRNAs allows straightforward in vivo target validation, a critical step to the development of effective therapeutics for these devastating tumors.Chemotherapy-induced autophagy is a proposed mechanism of chemoresistance and potential therapeutic target in osteosarcoma. Here, Livingston and colleagues report the occurrence and prognostic significance of autophagy and heat shock protein 27 (HSP27) in osteosarcoma patients. Autophagy, as measured by punctate LC3B expression (LC3B+), was increased in post-treatment specimens and highest in osteosarcoma metastasis. Following preoperative chemotherapy, expression of HSP27 was associated with poor survival whereas LC3B+ expression conferred a favorable prognosis. These results identify novel prognostic biomarkers in osteosarcoma and support further investigation into targeting HSP27 or modulating autophagy in osteosarcoma treatment.In this article, Durbin and colleagues demonstrate how a mechanistic model can determine key inefficiencies in the processing of an antibody-drug conjugates (ADC) at the cellular level. Using techniques such as imaging flow cytometry and mass spectrometry, the cellular ADC processing steps (i.e., binding, internalization, lysosomal processing, and intracellular drug accumulation) were analyzed and integrated into a novel model. Sensitivity analysis then highlighted parameters that, when modified, would ultimately yield the highest increase in intracellular free drug levels. These outcomes are important for understanding the steps driving ADC uptake and which steps could be improved upon in future rounds of drug development.

Toward a Ferrous Iron-Cleavable Linker for Antibody-Drug Conjugates.

Antibody-drug conjugates (ADCs) are antigen-targeted therapeutics that employ antibodies to deliver potent, cytotoxic effectors to cells with potentially high specificity. While promising clinical results have been achieved, significant pitfalls remain including internalization of ADCs in nontargeted cells expressing target antigen, which can limit therapeutic windows. Novel ADC linkers that are cleaved selectively in cancer cells but not in normal cells could minimize collateral damage caused by ADC uptake in nontargeted tissues. Here, we describe a prototypical ADC linker based on an Fe(II)-reactive 1,2,4-trioxolane scaffold (TRX) that by itself has demonstrated tumor-selective activity in preclinical cancer models. We prepared TRX-linked ADCs by site-selective conjugation to two sites in trastuzumab and compared their activity in Her2 positive and negative cells to ADC controls based on established linker chemistry. Our results confirm that the TRX moiety efficiently releases its payload following ADC uptake, affording picomolar potencies in antigen-positive cells. We also identified a destabilizing interaction between these initial TRX linkers and nearby antibody residues and suggest an approach to improve upon these prototypical designs.

Abstract 351: Uptake of antibody-drug conjugates by cultured Kupffer cells can predict pharmacokinetics

Abstract Interest in antibody-drug conjugates (ADCs) has increased rapidly in the oncology field over the past several years. Recent advances in the ADC field have been achieved in part by an improved understanding of how conjugation with drug-linkers impacts the biophysical, pharmacokinetic (PK), and biodistribution properties of a monoclonal antibody. Most of these advances have been driven by in vivo preclinical observations, an inevitably low throughput endeavor. A higher throughput in vitro method that would allow some level of predictive power for ADC disposition in vivo would be highly advantageous, requiring less test article and allowing for much faster turnaround. Thus far no such methods for ADCs have been reported, in contrast to the situation for the prediction of the metabolic clearance of small molecule therapeutics, for which cultured hepatocytes are widely used as an in vitro tool. It has been generally assumed that clearance of ADCs is driven by cells of the monocyte phagocytic system, and we recently published immunohistochemistry (IHC) data implicating hepatic sinusoidal endothelium and the resident liver macrophages, Kupffer cells (KCs), in the accelerated clearance of highly-loaded ADCs. We sought to use this observation to develop a convenient assay that might correlate ADC uptake in vitro with in vivo PK. Initial fluorescent microscopy experiments revealed that cultured rat KCs stain intensely when incubated with fluorescently labeled ADCs, but much less so with unconjugated antibody, similar to the observed IHC results. In an effort to further improve the throughput of this method to allow the rapid comparison of many ADCs, we moved to a FACS-based platform to quantify the amount of KC-associated fluorescent conjugate. To quantify the effect of different drug-linkers, we prepared homogeneous ADCs with 8 drugs per antibody and observed that the KC staining intensity varied widely depending upon the characteristics of the drug-linker that was conjugated. KCs incubated with ADCs loaded with vc-PAB-MMAE had a mean fluorescent intensity (MFI) >20 fold higher than unconjugated Antibody. Additionally, ADCs loaded with MMAE that contain a polyethylene glycol (PEG) masking moiety in the linker had a much lower MFI than those without PEG. This FACS method allowed us to compare fluorescent uptake by KCs with ADC clearance determined in vivo. We have observed that ADC uptake by cultured KCs in vitro correlates well with ADC PK, allowing for more rapid assessment of the PK impact of new drug classes, drug-linker designs, and conjugation methodologies. Citation Format: David Meyer, Sara Shum, Mechthild Jonas, Martha Anderson, Joshua Hunter, Nagendra Chemuturi, Nicole Okeley, Robert Lyon. Uptake of antibody-drug conjugates by cultured Kupffer cells can predict pharmacokinetics. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 351.

Abstract 1285: Tumor associated macrophages can process antibody-drug conjugates and contribute to antitumor activity in preclinical xenograft models

Abstract The primary mechanism of antibody-drug conjugates (ADCs) is the targeted delivery of a cytotoxic payload via cancer antigen mediated internalization. However, stromal components in the tumor microenvironment may also play a role in ADC penetration, distribution, and processing. Here, we study the potential roles of Fc-FcγR interaction between tumor-associated macrophages (TAMs) and ADCs in the antitumor activity observed in xenograft models. In the CD30+ L428 Hodgkin lymphoma (HL) model, the anti-CD30 ADC (cAC10-vcMMAE) and a non-binding control ADC (h00-vcMMAE) showed similar antitumor activity over a two-week period post treatment initiation. The antitumor activity was correlated with payload release, as h00-vcMMAE produced intratumoral MMAE concentration comparable to cAC10-vcMMAE in this time frame. Histopathology analysis of L-428 xenografts revealed high abundance of TAMs, leading to the hypothesis that TAMs can internalize and process ADCs for payload release. Using immunohistochemistry and flow cytometry, we confirmed that h00-vcMMAE bound to macrophage cell lines and TAMs. We further examined the presence of TAMs in additional xenograft models and correlated that to the antitumor activity of non-binding h00-vcMMAE. High levels of TAMs were observed in the KM-H2 HL and BR620 breast cancer models that were sensitive to h00-vcMMAE. In contrast, xenograft models with much fewer TAMs (DOHH2, SU-DHL8, and Karpas-299) did not respond to h00-vcMMAE treatment. Interestingly, h00-vcMMAF, releasing a membrane non-permeable payload, had no activity on the L-428 tumor model. These data suggest TAM-processed drug can mediate bystander tumor killing when the released payload is membrane permeable. To evaluate whether Fc-FcγR interaction plays a role in the ADC uptake by TAMs, we mutated the Fc region of h00-vcMMAE to decrease FcγR binding affinity (E233P:L234V:L235, G1V1). h00G1V1-vcMMAE lost its cytotoxicity activity in FcγR+ THP-1 monocytes. Furthermore, h00G1V1-vcMMAE could no longer mediate tumor regression or growth delay in three xenograft models that are sensitive to h00-vcMMAE treatment. These results suggest ADC-FcγR interaction is required for ADC processing by TAMs in these xenografts. These results suggest that TAMs can contribute to ADC processing through FcγR interaction in preclinical tumor models. Moreover, TAM-processed membrane permeable payload can mediate bystander tumor cell killing and contribute to ADC activity. Although this phenomenon may be an additional mechanism of ADCs in vivo, whether TAMs play a role in patients’ response to ADCs requires further correlated studies in clinical trials. Citation Format: Fu Li, Michelle Ulrich, Mechthild Jonas, Germein Linares, Xinqun Zhang, Lori Westendorf, Dennis Benjamin, Che-Leung Law. Tumor associated macrophages can process antibody-drug conjugates and contribute to antitumor activity in preclinical xenograft models. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1285.

Abstract 365: Using radionuclear imaging and mechanistic modeling to assess the therapeutic potential of antibody-drug conjugates (ADCs)

Abstract Antibody-drug conjugates (ADCs) comprise a drug class that allows for direct delivery of a cytotoxic agent to the target tumor cell. The successful development of an ADC involves the assessment of patient characteristics such as tumor antigen expression and tumor vascularity. There is a need to develop a mechanistic understanding of the effect these tumor parameters have on ADC biological activity. This mechanistic insight can be derived from a mathematical model integrating quantitative preclinical data, and will support ADC development. MLN0264 is an investigational ADC consisting of a human anti-guanylyl cyclase C (GCC) antibody linked to the microtubule-disrupting agent monomethyl auristatin (MMAE). Here, we use mathematical modeling, in conjunction with in vivo imaging, to decouple the contribution of different tumor parameters to overall ADC biological activity. We constructed a mathematical model of ADC biological activity by integrating experimental results from (1) in vivo antibody imaging studies, (2) in vitro viability assays, and (3) in vivo xenograft biological activity studies. First, blood pharmacokinetics and tumor disposition were quantitatively constrained using in vivo radiolabeled antibody single-photon emission computed tomography (SPECT) data for both blood and tumor tissues. SPECT data from three xenografts with various antigen expression levels were used to link antigen expression level to ADC uptake. Second, the relationship between bound GCC receptor concentration and cell viability was established using viability assays run on an engineered cell line (293-GCC) with high antigen expression and high sensitivity to MMAE. Finally, using this relationship, we built a tumor growth dynamics model to describe in vivo xenograft biological activity, and to estimate the growth inhibition coefficient of 293-GCC. This mechanistic model can be used to gain insights into the factors driving response of a tumor that is intrinsically sensitive to MMAE. Our results indicate the process of ADC vascular permeability is one of the limiting factors of ADC disposition. This outcome is reasonable given that a large ADC molecular weight decreases the permeability. Furthermore, the model simulations suggest some tumors that are intrinsically sensitive to MMAE may not be affected by the ADC if GCC antigen expression levels are very low. Taken together, the mechanistic model developed here forms the basis of a quantitative understanding for several factors influencing MLN0264 patient selection. Citation Format: Shu-Wen Teng, Ozlem Yardibi, Julie Zhang, Donna Cvet, Johnny Yang, Kelly Orcutt, Melissa Gallery, Arijit Chakravarty, Wen Chyi Shyu, Jerome Mettetal, Daniel Bradley, Petter Veiby. Using radionuclear imaging and mechanistic modeling to assess the therapeutic potential of antibody-drug conjugates (ADCs). [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 365. doi:10.1158/1538-7445.AM2014-365

Proposed mechanism of off-target toxicity for antibody-drug conjugates driven by mannose receptor uptake.

Antibody-drug conjugates (ADCs) are developed with the goal of increasing compound therapeutic index by specific and targeted delivery of a toxic payload to the site of action while considerably reducing damage to normal tissues. Yet, off-target hepatic toxicities have been reported for several ADC. Locations of these off-target toxicities coincide with the reported locations of cell surface mannose receptor (MR). The relative proportion of agalactosylated glycans on the Fc domain (G0F vs. G1F and G2F components) in monoclonal antibody (mAb)-based biotherapeutics is closer to some disease state IgG rather than to a normal serum-derived immunoglobulin. The lack of the terminal galactose on a G0F glycan creates an opportunity for the mAb to interact with soluble and cell surface MRs. MR is a known multi-domain lectin that specifically binds and internalizes glycoproteins and immune complexes with relatively high G0F content and has been found on the surface of various cell types, including immune cells of myeloid lineage, endothelial cells, and hepatic and splenic sinusoids. In this review paper it is proposed that the mechanism of the off-target toxicities for ADC biotherapeutics is at least in part driven by the carbohydrates, specifically agalactosylated glycans, such as G0F, their interactions with MR and resulting glycan-derived cellular uptake of ADCs. Several case studies are reviewed presenting corroborating information.

Désir, plaisir et dépendance : rôle de la dopamine

Antibody-drug conjugates (ADCs) are being developed worldwide with the potential to revolutionize current cancer treatment strategies. However, off-target toxicity caused by the instability of linkers remains one of the main issues to be resolved. Developing a novel photocontrol-ADC with good stability and photocontrolled release seemed to be an attractive and practical solution. In this study, we designed, for the first time, a novel ultraviolet (UV) light-controlled ADC by carefully integrating the UV-cleavable o-nitro-benzyl structure into the linker. Our preliminary work indicated that the ADC exhibited good stability and photocontrollability while maintaining a targeting effect similar to that of the naked antibody. Upon irradiation with UV light, the ADC rapidly released free cytotoxins and exerted significant cytotoxicity toward drug-resistant tumor cells. Compared to those of the unirradiated cells, the EC50 values of ADCs increased by up to 50-fold. Furthermore, our research confirmed that the degradation products of unirradiated ADC, Cys-1a, were relatively less toxic, thus potentially reducing the off-target toxicity caused by nonspecific uptake of ADCs. The novel design strategy of UV light-controlled ADCs may provide new perspectives for future research on ADCs and promote the development of photocontrol systems.