Effective Elimination of Acute Myeloid Leukemic Cells by Recombinant Bispecific Antibody Derivatives Directed Against CD33 and CD16

Abstract

Single-chain Fv triplebodies (sctb), consisting of a single polypeptide chain with 3 single-chain antibody variable fragments connected in tandem, were generated as antileukemic agents. A CD19-specific sctb of this format has previously been shown to be superior to a bispecific single-chain Fv antibody fragment (bsscFv) for the elimination of leukemic B-lineage cells, but corresponding targeted agents for the treatment of acute myeloid leukemia are still lacking. For this purpose, both a bsscFv and a sctb specific for CD33 and the trigger molecule CD16 (FcgammaRIII) were produced. The sctb displayed 3.5-fold greater avidity for CD33 than the bsscFv 33xds16, whereas both had close to equal affinity for CD16. In antibody-dependent cellular cytotoxicity (ADCC) reactions with human mononuclear cells as effectors, both the bsscFv 33xds16 and the sctb induced lysis of tumor cells with half maximum effective concentrations (EC50) in the low picomolar range. It is interesting to note that the sctb promoted equal lysis of human leukemia-derived cell lines at 10 to 200-fold lower concentrations than the bsscFv. Both molecules mediated ADCC of primary patient cells. In conclusion, both the bsscFv 33xds16 and the sctb 33xds16x33 eliminated acute myeloid leukemia cells in ADCC reactions, but the novel sctb format showed significantly greater specific activity.

Simultaneous engagement of the activatory receptors NKG2D and CD3 for retargeting of effector cells to CD33-positive malignant cells

“…1 The recent discovery of LNK mutations in JAK2V617F-negative MPN suggested an alternative mechanism of JAK-STAT (Janus kinase/signal transducers and activators of transcription) activation and clonal erythrocytosis. 2,3 However, subsequent studies showed that JAK2 and LNK mutations are not necessarily mutually exclusive and the pathogenetic contribution of LNK mutations, in the presence of JAK2V617F, remains unclear. 4 We recently detected two distinct LNK mutations in each of two patients with primary myelofibrosis.…”

“…3,4 For retargeting of T cells bispecific antibodies commonly consist of an anti-CD3 domain and an antibody domain that is directed to a tumor antigen on the cell surface. 4 Among other strategies, 3 for retargeting of NK cells an immuno-ligand for the activating receptor NKG2D was described.…”

“…3,4 For retargeting of T cells bispecific antibodies commonly consist of an anti-CD3 domain and an antibody domain that is directed to a tumor antigen on the cell surface. 4 Among other strategies, 3 for retargeting of NK cells an immuno-ligand for the activating receptor NKG2D was described. 5 This immuno-ligand consists of the soluble extracellular domain of the receptor ULB-P2 and a single-chain fragment variable (scFv) directed to the target antigen.…”

NK cells and cancer: you can teach innate cells new tricks

“…Different tumour types have been targeted by linking these anti-CD16 Fv domains to Fv domains against CD19 (REF. 177) and HLA class II for B cell malignancies 178 , CD30 for Hodgkin's lymphoma 176,179 , epidermal growth factor receptor (EGFR), which is overexpressed in several epithelial cancer types 175 , HER2 (REFS 180,181) for breast cancer, CD33 for AML 182,183 and EPCAM 184 for carcinomas. The in vitro and in vivo efficacy of these optimized proteins in eliciting specific antitumour activity is encouraging and should be further developed in clinical settings.…”

A dual-targeting triplebody mediates preferential redirected lysis of antigen double-positive over single-positive leukemic cells

“…These fusion proteins consist of three different scFv modules connected in tandem by flexible linkers into a single polypeptide chain. [39][40][41][42][43] The distal scFv binding modules can either bind two copies of the same target antigen, or one copy each of two different targets. In the latter case, the triplebody is said to be "dual-targeting."…”