Abstract P062: The CL2A-SN38 linker-payload system conjugated to trastuzumab results in improved cellular cytotoxicity over time relative to T-DM1
Notice bibliographique
Résumé
Abstract Background The approved antibody-drug conjugate (ADC) sacituzumab-govitecan (SG) that exploits the topoisomerase I inhibitor SN38, represents a substantial advance in the ADC field. SN38 is the first payload with low nanomolar cytotoxicity. This deviates from current design principles, which utilize ultracytotoxic payloads. The CL2A linker has a short non-cleavable seven polyethylene glycol segment that enables a higher number of hydrophobic SN38 molecules to be attached to reduced cysteines. CL2A is also designed to permit the release of SN38 in an acidic environment via a pH-sensitive benzyl carbonate bond to SN38’s lactone ring. There is no cathepsin B cleavage site. Key to the performance of SG is the SN38 release half-life of 1-2 days. This also deviates from very stable linker systems that require full ADC internalization and lysosomal processing for payload release. As a result of these novel ADC properties, SG is given in higher and repeated doses, which results in slow release at the tumor site and enhanced uptake by target cancer cells and reduced toxicity on healthy cells due to low potency SN38 alone. Our goal is to further evaluate the CL2A-SN38 system when conjugated to trastuzumab. Material and methods Trastuzumab was conjugated to CL2A-SN38 to reduced cysteines (T-SN38). Monitoring the changes in DAR over time was performed to determine the stability of T-SN38 in PBS at pH values of 7.4, 5.72, and 4.58, to mimic circulating blood, early endosomes, and lysosomes, respectively. Cytotoxicity assays for 72 h on HER2-positive ovarian cancer SKOV3.ip and HER2-negative CHO cells were performed and compared to the approved anti-HER2 ADC trastuzumab-emtansine (T-DM1). Cytotoxicity assays were also performed between T-SN38 and T-DM1 at their respective IC50 values and the percent of cell survival determined daily from 24-96 h. Results T-SN38 was generated with a DAR of 6.97. Surprisingly, the half-life of SN38 release in acidic pH was extremely slow (t1/2=~120 h) and at the final evaluated time point (120 h) 69% of SN38 remained conjugated to trastuzumab. As anticipated, as a function of concentration T-DM1 was significantly more potent than T-SN38 (IC50=0.0013 vs 0.0832 mg/mL). However, when cytotoxicity was evaluated as a function of time T-SN38 switched to the more potent ADC at the late time points. T-SN38 killed SKOV3.ip cells 36% more effectively than T-DM1 at 96 h. Conclusions Our data from SN38 release experiments in acidic buffer suggest that pH-induced cleavage of the CL2A linker is not a contributable release mechanism of SN38 when bound to trastuzumab and is counterintuitive for what is currently described for SG. However, traceless release of hydrophobic SN38 capable of traversing cell membranes is only possible by hydrolysis of the benzyl carbonate bond. Our cytotoxicity studies over time suggest that pH-dependent release does occur. Further investigations are ongoing to more precisely determine the contribution of the carbonate bond for SN38 release and cytotoxic potency. Citation Format: Hardeep Singh, Victor Jeffrey Leyton. The CL2A-SN38 linker-payload system conjugated to trastuzumab results in improved cellular cytotoxicity over time relative to T-DM1 [abstract]. In: Proceedings of the AACR-NCI-EORTC Virtual International Conference on Molecular Targets and Cancer Therapeutics; 2021 Oct 7-10. Philadelphia (PA): AACR; Mol Cancer Ther 2021;20(12 Suppl):Abstract nr P062.
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Comment cette classification a été obtenuedéplier
Prédiction machine sur la base complète
Imitation des enseignantsNi prévalence calibrée, ni vérité terrain. Validation humaine à venir. Le volet Gemma est une étiquette directe du modèle pour chaque travail de la base, lue sur la notice réduite au titre. Le volet Codex est un classifieur appris des 10 348 étiquettes directes de Codex et calibré sur les taux pondérés de l'échantillon; les champs sans appui suffisant ne portent aucun appel Codex. Le mode candidate est l'union des deux volets; le consensus est leur intersection. Ces sorties portent le statut machine_predicted_unvalidated et ne sont pas des étiquettes humaines.
Scores du classifieur distillé par catégorie (deux têtes)
| Catégorie | Codex | Gemma |
|---|---|---|
| Métarecherche | 0,000 | 0,000 |
| Méta-épidémiologie (sens strict) | 0,000 | 0,000 |
| Méta-épidémiologie (sens large) | 0,000 | 0,000 |
| Bibliométrie | 0,000 | 0,000 |
| Études des sciences et des technologies | 0,000 | 0,000 |
| Communication savante | 0,000 | 0,000 |
| Science ouverte | 0,000 | 0,000 |
| Intégrité de la recherche | 0,000 | 0,000 |
| Charge utile insuffisante (le modèle a refusé de juger) | 0,003 | 0,001 |
Scores machine (provisoires)
Les deux têtes enseignantes du modèle étudiant, lues sur ce travail. Un score ordonne la base pour la relecture; il n'affirme jamais une catégorie, et le statut de validation accompagne chaque rangée tel quel.
Scores de référence d'un modèle non mature (critères de maturité non atteints, 7 itérations). Un score ordonne; il n'affirme jamais une catégorie.
score_only:v0-immature-baseline · tel quel depuis la passe de notation : score_only signifie que le nombre peut ordonner les travaux, et qu'aucune étiquette de catégorie n'en découleClassification
machine, non validéePrédiction automatique; un appel candidat d’une seule source (Gemma direct ou Codex distillé), pas un consensus.
Le détail, modèle par modèle et score par score, se trouve en fin de page sous « Comment cette classification a été obtenue ».