Recombinant anti-CD25 immunotoxin RFT5(scFv)-ETA' demonstrates successful elimination of disseminated human hodgkin lymphoma in SCID mice

Abstract

Since clinical phase-I/II trials in patients with resistant Hodgkin's lymphoma treated with the chemically linked anti-CD25 ricin-A-chain immunotoxin RFT5-SMPT-dgA indicate promising results for patients with minimal residual disease, we constructed a new immunotoxin by fusing the RFT5 single-chain variable fragment to a deletion mutant of Pseudomanas exotoxin A (ETA'). The recombinant protein was directed into the periplasmic space of E. coli by means of the pET-derived expression vector pBM1.1 and our newly developed expression/purification method. Biologically active RFT5(scFv)-ETA' was isolated by freezing/thawing and purified by immobilized metal-ion affinity and molecular-size-chromatography. RFT5(scFv)-ETA' was subsequently used for the treatment of disseminated human Hodgkin's lymphoma in a SCID- mouse model. The mean survival time (MST) of L540rec-challenged SCID mice was 38.1 days. A single i.v. injection of 40 μg recombinant immunotoxin (rIT) I day after tumor inoculation resulted in 100% tumor-free mice, extending the MST to more than 220 days (p < 0.0001). The blood-distribution time T(1/2)α was 39.65 min, the serum elimination time T(1/2)α, 756.6 min. All animals were assessed for soluble interleukin-2 receptor α, which is directly correlated to tumor burden. Soluble CD25 was not detectable in mice treated with the rIT. Our findings, concerning potent anti-tumor effects of a recombinant anti-CD25 immunotoxin against disseminated Hodgkin's lymphoma in SCUD mice reported here demonstrate that RFT5(scFv)-ETA' might be suitable for further evaluation against Hodgkin's lymphoma in humans. (C) 2000 Wiley- Liss, Inc.