Synthesis and characterization of an antihuman T-lymphocyte saporin
immunotoxin (OKT1-SAP) with in vivo stability into nonhuman primates
S Siena, DA Lappi, M Bregni, A Formosa, S Villa, M Soria, G Bonadonna and AM Gianni
Cristina Gandini Bone Marrow Transplantation Unit, Istituto Nazionale
Tumori, Milan, Italy.
The authors conjugated, by a disulphide bond, the antihuman T- lymphocyte
(CD5) monoclonal antibody (MoAb) OKT1 to the saporin-6 (SAP)
ribosome-inactivating protein of the plant Saponaria officinalis. The
resulting OKT1-SAP immunotoxin bound to CD5-expressing target cells and
under standard culture conditions specifically suppressed mitogen-
induced-T-lymphocyte DNA and protein synthesis in a dose-related manner.
T-lymphocyte killing was achieved by five-minute exposure of the target
cells to OKT1-SAP. The concentration inhibiting 50% (IC50) of T-lymphocyte
DNA synthesis was 0.32 nmol/L. The potency of OKT1-SAP was moderately
enhanced by amantadine (IC50 0.08 nmol/L) but not by ammonium chloride or
chloroquine. Whole blood components did not interfere with the efficacy of
OKT1-SAP, as in vitro treatment of fresh whole blood resulted in effective
elimination of clonable peripheral blood T-lymphocytes assessed by a
limiting dilution assay. Because these characteristics of T-lymphocyte
killing by OKT1-SAP (ie, rapidity of action, potency also without
potentiators) and lack of inhibition by whole blood components may be
relevant for the use of an immunotoxin as a therapeutic agent in humans,
the authors evaluated the stability in vivo and the circulatory clearance
of OKT1-SAP in cynomolgus monkeys. Following a single intravenous (IV)
injection of nontoxic dosages (0.16 to 1.3 mg/kg), an initial rapid decline
(t1/2 alpha = 1.0 to 4.1 hours) was followed by a long-lasting slower
decrease (t1/2 beta = 11.6 to 20.6 hours) of OKT1-SAP plasma concentrations
as detected by double- antibody solid phase enzyme-linked immunosorbent
assay (ELISA) assay. Not only did OKT1-SAP remain intact immunologically
but it also retained its biological activity, as measured by the ability of
plasma samples from monkeys given immunotoxin to inhibit DNA synthesis in
human T-lymphocytes. Taken together the findings presented in this article
indicate the feasibility of using OKT1-SAP as a therapeutic tool and
provide information that will facilitate the rational use of immunotoxins
as a treatment modality in humans.
Volume 72,
Issue 2,
pp. 756-765,
08/01/1988
Copyright © 1988 by The American Society of Hematology
