Conference 2021 Live Talk
Development of novel auristatin F-containing immunoconjugates for the treatment of triple-negative breast cancer
Authors and Affiliations
Neelakshi Mungra1, Fleury Augustin Nsole Biteghe2, Stefan Barth1,3
1. Medical Biotechnology & Immunotherapy Research Unit, Institute of Infectious Disease and Molecular Medicine, Faculty of Health Sciences, University of Cape Town, Cape Town, South Africa
2. Department of Radiation Oncology and Biomedical Sciences, Cedars-Sinai Medical, Los Angeles, CA, USA
3. South African Research Chair in Cancer Biotechnology, Department of Integrative Biomedical Sciences, Faculty of Health Sciences, University of Cape Town, Cape Town, South Africa
Targeted therapy for breast cancer represents the most promising state-of-the-art technology in the field of oncology. Such therapy directly addresses the lack of specificity encountered with conventional treatment regimes. This is achievable through the use of receptor-specific monoclonal antibodies eventually conjugated to potent cytotoxic agents. However, despite showing great promise as anti-cancer compounds, these agents, also known as antibody-drug conjugates (ADCs) have several limitations: they feature stoichiometrically undefined chemical linkages, poor tumor penetration, unpredictable pharmacokinetic profiles and their generation results in a heterogeneous mixture of products. We generated a SNAP-tag based fusion protein targeting chondroitin-sulphate proteoglycan 4 (CSPG4) and CD44; biomarkers of aggressive and resistant cancers, for the treatment of triple-negative breast cancer (TNBC). By conjugating this fusion protein with monomethyl auristatin F (AURIF), a potent antimitotic agent, we propose to alleviate the challenges of early ADC development, while ensuing tumor cell death through the disruption of mitosis.
We designed a CSPG4/CD44-targeting SNAP-tag based fusion protein as a fluorescent mimic for binding. Protein expression was carried out using a transient secretory mammalian expression system and purified from the cell culture supernatant by ion metal affinity chromatography. Binding of αCSPG4/CD44(scFv)-SNAP to CSPG4/CD44-positive and -negative TNBC cell lines was validated both quantitatively by flow cytometry and qualitatively by confocal microscopy. The AURIF-containing immunoconjugates were generated by reacting αCSPG4/CD44(scFv)-SNAP with benzylguanine (BG)-modified AURIF molecules in a defined 1:1 stoichiometry. Specific and dose-dependent biological activity of the resulting compounds was then assessed on TNBC cells.
Preliminary proof-of-concept data demonstrate that the CSPG4/CD44-targeting, fluorescently-labeled αCSPG4/CD44(scFv)-SNAP binds effectively to CSPG4/CD44-positive TNBC cells. Moreover, a dose-dependent reduction in cell viability was observed in all CSPG4/CD44-positive cell lines, highlighting the promising anti-cancer activity of the AURIF-containing drug.
The combination of such agents with a robust companion diagnostic tool such as SNAP-tag technology represents the first step towards the effective management of African TNBC.