Sensitivity of cancer cell lines to the novel non-toxic type 2 ribosome-inactivating protein nigrin b Raquel Mun Äoz, Yolanda Arias, J. Miguel Ferreras, Pilar Jime Ânez, M. Angeles Rojo, Toma Âs Girbe Âs * Departamento de Bioquõ Âmica y Biologõ Âa Molecular, Facultad de Ciencias, Universidad de Valladolid, 47005 Valladolid, Spain Received 17 January 2001; received in revised form 14 February 2001; accepted 1 March 2001 Abstract The cytotoxicity of the type 2 ribosome-inactivating proteins (RIPs) ricin and nigrin b was determined in a variety of cancer cells. Nigrin b, considered to be a novel non-toxic type 2 RIP as compared with ricin, was approximately 10 4 ±10 5 times less toxic than ricin in all cancer cells studied, with the exception of melanoma cells. Cancer cells displayed considerable hetero- geneity in their sensitivity to ricin, melanoma cells being the least sensitive. Rabbit polyclonal anti-nigrin b antibodies did not cross-react with ricin as analyzed by enzyme-linked immunosorbent assays. The low non-speci®c toxicity of nigrin b as compared with that of ricin and the lack of immunological cross-reaction between anti-nigrin b antibodies and ricin supports the use of nigrin b in the construction of cytotoxic conjugates as an alternative to ricin when anti-ricin antibodies are produced during cancer therapy. q 2001 Elsevier Science Ireland Ltd. All rights reserved. Keywords: Nigrin b; Ricin; Ribosome-inactivating protein; Cancer cells; Toxicity; Cancer therapy 1. Introduction Among the novel approaches for cancer therapy, immunotoxins have emerged as promising tools for targeting and killing malignant cells and tumors [1,2]. Immunotoxins contain a monoclonal antibody that targets a toxic agent, usually a protein toxin, to antigens present in target cells. Among the protein toxins used in the construction of immunotoxins are the ribosome-inactivating proteins (RIPs) like the A chain of the type 2 RIP ricin and the single chain type 1 RIPs like saporin, which are a class of enzymes that inhibit protein synthesis by irreversible inactivation of ribosomes [3]. The molecular mechanism of action of RIPs against mammalian ribosomes is the hydrolysis of the N-glycosidic link- age of the adenine A 4234 present in the highly conserved loop responsible for the interaction of elongation factor 2 with the ribosome [4]. RIPs are currently classi®ed in two categories: type 1 RIPs (having a single polypeptide chain with N-glycosi- dase activity), and type 2 RIPs (having two polypep- tide chains, one enzymic polypeptide chain (A chain) and the other one a lectin (B chain) able to recognize plasma membrane glycoproteins). Traditionally, type 2 RIPs have been considered as highly toxic to most mammalian cells in culture and also to animals. The best-known type 2 RIP is the extremely toxic ricin found by Stillmark in 1888 present in the seeds of Ricinus communis [5]. Both intact ricin and the ricin A chain have been Cancer Letters 167 (2001) 163±169 0304-3835/01/$ - see front matter q 2001 Elsevier Science Ireland Ltd. All rights reserved. PII: S0304-3835(01)00477-3 www.elsevier.com/locate/canlet * Corresponding author. Tel.: 134-983-423082; fax: 134-983- 423083. E-mail address: girbes@bio.uva.es (T. Girbe Âs).