Decitabine Or 5-Azacitidine Pre-Treatment Does Not Compromise The Novel Fc-Engineered Antibody Mab 33.1-Mediated ADCC Against Primary AML Blasts - A Rationale For Combination Therapy
Authors: He Shun , Cheney Carolyn , Whitman Susan P. , Yu Jianhua , Vasu Sumithira , Blum William , Heider Karl-Heinz , Marcucci Guido , Muthusamy Natarajan 
Published: October 2019

Introduction

Acute Myeloid leukemia (AML) in patients older than 60 years is a devastating diagnosis with long-term survival rates of 10%. Elderly patients have poor survival both due to chemoresistance and presence of concomitant comorbidities rendering them ineligible for induction chemotherapy. Hence novel treatment options are warranted in this patient population. Promising activity of monoclonal antibodies such as alemtuzumab and rituximab for chronic lymphocytic leukemia (CLL) and rituximab for lymphomas has raised the potential use of antibody therapies in AML. CD33 is expressed on greater than 90% of AML blast cells while absent from all non-hematopoietic tissues. Hence CD33 is a viable target for antibody-based therapeutics in AML. Here, we tested the ex vivo efficacy of the mAb 33.1, a fully human anti-CD33 antibody Fc-engineered for increased binding to Fcγ receptors on AML cell lines and primary AML blasts. The goals of this study are to evaluate 1) the efficacy of mAb33.1 on purified allogeneic and autologous natural killer (NK) cell-mediated antibody-dependent cellular cytotoxicity (ADCC) against primary AML Blasts; 2) to evaluate efficacy of mAb 33.1 in combination with azanucleosides (i.e. decitabine, 5-azacitidine) that are currently used in AML therapy on NK cell-mediated ADCC against primary AML blasts; and 3) to correlate the levels of surface expression of CD33 on AML blasts to the mAb 33.1 mediated ADCC.

Methods

mAb 33.1 mediated NK cell activation was determined by NK degranulation as determined by CD107a induction, and ADCC was determined by standard 4-hour 51Cr-release assay. An AML cell line HL60 and a total of 15 AML blast samples were used as targets in this study. NK cells enriched from normal donor PBMC (for allogeneic assays) or sorted from AML blast samples (for autologous assays) were used as effector cells.

Results

The mAb 33.1 induced potent ADCC activity (>40%) compared to control non-Fc engineered antibody at the concentration of 10 μg/ml in the HL60 cell line. For the AML blasts, mAb 33.1 mediated significantly higher ADCC activity when compared to the control antibody (p<0.05). The relative cytotoxicity mediated by mAb 33.1 varied among different patients, ranging from 4.4% to 65.8%. Subsequent quantification of CD33 showed that there is a positive correlation between ADCC activity and the number of surface CD33 molecules on the AML blasts. Induction of CD107a expression was also observed in both allogeneic and autologous NK cells when the blasts were labeled with mAb 33.1. Pre-treatment of the NK cells and/or target blasts with decitabine or 5-azacitidine for 48hrs, did not alter the mAb 33.1 mediated ADCC activity or CD107 induction.

Conclusion

mAb33.1 mediated potent ADCC activity and NK activation against AML cell lines and primary AML blasts. Both autologous and allogeneic NK cell-mediated ADCC against primary blast cells from AML patients was observed. The level of NK cell-mediated ADCC was positively associated with the levels of the surface CD33 expression on target AML blasts. Pre-treatment of either AML blasts and/or NK effector cells with Decitabine or 5-azacitidine did not compromise mAb 33.1-mediated ADCC. These pre-clinical studies support further clinical development of mAb 33.1 in combination with relevant anti-AML therapies such as decitabine or 5-azacitidine in patients with CD33 expression.

Disclosures:

Heider: boehringer-ingelheim: Employment.