An evolved ribosome-inactivating protein targets and kills human melanoma cells in vitro and in vivo.
BACKGROUND
Few treatment options exist for patients with metastatic melanoma, resulting in poor prognosis. One standard treatment, dacarbazine (DTIC), shows low response rates ranging from 15 to 25 percent with an 8-month median survival time. The development of targeted therapeutics with novel mechanisms of action may improve patient outcome. Ribosome-inactivating proteins (RIPs) such as Shiga-like Toxin 1 (SLT-1) represent powerful scaffolds for developing selective anticancer agents. Here we report the discovery and properties of a single chain ribosome-inactivating protein (scRIP) derived from the cytotoxic A subunit of SLT-1 (SLT-1A), harboring the 7-amino acid peptide insertion IYSNKLM (termed SLT-1A IYSNKLM) allowing the toxin variant to selectively target and kill human melanoma cells.


RESULTS
SLT-1A IYSNKLM was able to kill 7 of 8 human melanoma cell lines. This scRIP binds to 518-A2 human melanoma cells with a dissociation constant of 18 nM, resulting in the blockage of protein synthesis and apoptosis in such cells. Biodistribution and imaging studies of radiolabeled SLT-1A IYSNKLM administered intravenously into SCID mice bearing a human melanoma xenograft indicate that SLT-1AI YSNKLM readily accumulates at the tumor site as opposed to non-target tissues. Furthermore, the co-administration of SLT-1A IYSNKLM with DTIC resulted in tumor regression and greatly increased survival in this mouse xenograft model in comparison to DTIC or SLT-1A IYSNKLM treatment alone (115 day median survival versus 46 and 47 days respectively; P values < 0.001). SLT-1A IYSNKLM is stable in serum and its intravenous administration resulted in modest immune responses following repeated injections in CD1 mice.


CONCLUSIONS
These results demonstrate that the evolution of a scRIP template can lead to the discovery of novel cancer cell-targeted compounds and in the case of SLT-1A IYSNKLM can specifically kill human melanoma cells in vitro and in vivo.
Cheung
Melissa C
MC
Department of Pharmaceutical Sciences, University of Toronto, 144 College Street, Toronto, ON, M5S 3 M2, Canada.
Revers
Leigh
L
Perampalam
Subodini
S
Wei
Xin
X
Kiarash
Reza
R
Green
David E
DE
Abdul-Wahid
Aws
A
Gariépy
Jean
J
D000595:Amino Acid Sequence; D000818:Animals; D017209:Apoptosis; D055162:Biocatalysis; D045744:Cell Line, Tumor; D006801:Humans; D007074:Immunoglobulin G / Q000276:immunology; D008545:Melanoma / Q000473:pathology*; D051379:Mice; D016513:Mice, SCID; D008958:Models, Molecular; D008969:Molecular Sequence Data; D019151:Peptide Library; D011485:Protein Binding; D021381:Protein Transport; D011956:Receptors, Cell Surface / Q000378:metabolism; D012074:Remission Induction; D054788:Ribosome Inactivating Proteins / Q000378:metabolism*; D022622:Shiga Toxin 1 / Q000737:chemistry / Q000276:immunology / Q000378:metabolism*; D016019:Survival Analysis; D023041:Xenograft Model Antitumor Assays
D007074:Immunoglobulin G; D019151:Peptide Library; D011956:Receptors, Cell Surface; D022622:Shiga Toxin 1; D054788:Ribosome Inactivating Proteins