Novel PSCA targeting scFv-fusion proteins for diagnosis and immunotherapy of prostate cancer

Autor: Mehmet Kemal Tur, Stefan Barth, Alessa Pardo, Claudia Kessler, Rainer Fischer, Stefan Gattenlöhner, Katharina Kolberg
Rok vydání: 2017
Předmět:
Male
0301 basic medicine
Cancer Research
Virulence Factors
Recombinant Fusion Proteins
media_common.quotation_subject
medicine.medical_treatment
Bacterial Toxins
Exotoxins
chemical and pharmacologic phenomena
Immunologic Tests
GPI-Linked Proteins
law.invention
Flow cytometry
03 medical and health sciences
0302 clinical medicine
Antigens
Neoplasm
law
medicine
Humans
Pseudomonas exotoxin
Molecular Targeted Therapy
Internalization
media_common
ADP Ribose Transferases
medicine.diagnostic_test
Chemistry
Immunotoxins
Prostatic Neoplasms
General Medicine
Immunotherapy
respiratory system
Flow Cytometry
Fusion protein
Molecular biology
Neoplasm Proteins
Prostate Stem Cell Antigen
HEK293 Cells
030104 developmental biology
Oncology
030220 oncology & carcinogenesis
Recombinant DNA
Single-Chain Antibodies
Alkyltransferase
Zdroj: Journal of Cancer Research and Clinical Oncology. 143:2025-2038
ISSN: 1432-1335
0171-5216
DOI: 10.1007/s00432-017-2472-9
Popis: Despite great progress in the diagnosis and treatment of localized prostate cancer (PCa), there remains a need for new diagnostic markers that can accurately distinguish indolent and aggressive variants. One promising approach is the antibody-based targeting of prostate stem cell antigen (PSCA), which is frequently overexpressed in PCa. Here, we show the construction of a molecular imaging probe comprising a humanized scFv fragment recognizing PSCA genetically fused to an engineered version of the human DNA repair enzyme O6-alkylguanine-DNA alkyltransferase (AGT), the SNAP-tag, enabling specific covalent coupling to various fluorophores for diagnosis of PCa. Furthermore, the recombinant immunotoxin (IT) PSCA(scFv)-ETA′ comprising the PSCA(scFv) and a truncated version of Pseudomonas exotoxin A (PE, ETA′) was generated. We analyzed the specific binding and internalization behavior of the molecular imaging probe PSCA(scFv)-SNAP in vitro by flow cytometry and live cell imaging, compared to the corresponding IT PSCA(scFv)-ETA′. The cytotoxic activity of PSCA(scFv)-ETA′ was tested using cell viability assays. Specific binding was confirmed on formalin-fixed paraffin-embedded tissue specimen of early and advanced PCa. Alexa Fluor® 647 labeling of PSCA(scFv)-SNAP confirmed selective binding to PSCA, leading to rapid internalization into the target cells. The recombinant IT PSCA(scFv)-ETA′ showed selective binding leading to internalization and efficient elimination of target cells. Our data demonstrate, for the first time, the specific binding, internalization, and cytotoxicity of a scFv-based fusion protein targeting PSCA. Immunohistochemical staining confirmed the specific ex vivo binding to primary PCa material.
Databáze: OpenAIRE
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