Abstract
ADCs targeting cytotoxic payloads to cancer cells using antibodies that recognize tumor-associated antigens are an expanding therapeutic area. CD123, the IL-3 receptor alpha-subunit, is an attractive cancer target implicated in AML cell survival and proliferation. While CD123 is expressed on AML blasts and is associated with aggressive disease, it is also differentially expressed on AML stem cells relative to normal hematopoietic cells. We report the pre-clinical evaluation of IMGN632, an ADC comprising a CD123-binding antibody and a DNA-alkylating, indolino-benzodiazepine payload (termed an IGN), in disseminated and subcutaneous (SC) AML xenograft models which possess poor prognosis markers or demonstrate drug resistance.
Novel anti-CD123 antibodies were generated in mice by immunization with a human CD123-expressing cell line. Following antibody selection and humanization in the IgG1 format, IMGN632 was produced by conjugating a novel DNA-alkylating payload (DGN549), via a cleavable peptide linker, to the anti-CD123 antibody at engineered cysteine residues, resulting in an ADC with ~2 DGN549 molecules per antibody. The in vitro cytotoxicity of IMGN632 and of a non-targeted control ADC (the DGN549 payload conjugated to an antibody against Kunitz soybean trypsin inhibitor) on human cancer cell lines were evaluated by determining cell viability using the WST-8 reagent after continuous ADC exposure for up to 7 days. The IC50 was determined for each ADC and the specificity ratio (IC50 of control ADC: IC50 of IMGN632) was calculated for each cell line. The antitumor activity of IMGN632 and the control ADC were assessed in vivo in immuno-deficient mice bearing Molm-13, Kasumi-3-Luc or MV4-11 disseminated or EOL-1 sc human AML xenografts. Subcutaneous tumor volumes were measured twice weekly. Mice bearing Kasumi-3-Luc were live-animal imaged approximately twice weekly to quantify bioluminescent tumor burden. The maximum tolerated dose (MTD) of IMGN632 was determined by administering single intravenous (IV) injections of IMGN632 to non-tumor bearing female CD-1 mice. In all studies, assessment was terminated for weight loss > 20% or for clinical signs or, in the EOL-1 model, when SC tumor volume reached approximately 1000 mm3.
IMGN632 was highly potent in vitro towards CD123-expressing human AML cell lines with poor prognostic markers (EOL-1, Molm-13 (FLT3-ITD), MV4-11 (FLT3-ITD) and Kasumi-3 (p53, MDR1+)), with IC50 values of < 3 pMol and high specificity ratios ranging from 100 to 2000. In contrast, IMGN632 had IC50 values of > 8000 pMol and a specificity ratio of 1 in CD123-negative human cell lines (Namalwa, K562). These data confirm the CD123-directed activity of IMGN632 in vitro.
IMGN632 displayed antigen-specific antitumor activity in immuno-deficient mice bearing disseminated xenografts, resulting in prolonged survival, with an increase in life span from 50% to > 262%. In these models, the control ADC was inactive, with no increase in life span. In the Kasumi-3-Luc disseminated model, treatment with highly active doses of 240 or 800 mcg/kg IMGN632 (by antibody) resulted in decreased bioluminescent tumor burden. In the other disseminated models, IMGN632 was highly active at doses of 80 or 240 mcg/kg in MV4-11 and at doses as low as 8 mcg/kg in Molm-13, which, when taken together with the MTD of 8000 mcg/kg, generated a therapeutic index of 1000. In the azacitidine- and cytarabine-resistant EOL-1 SC model, IMGN632 was highly active at a dose of 240 mcg/kg, resulting in 8/8 long-term complete responses, while unconjugated DGN549, anti-CD123 antibody and control ADC were inactive at IMGN632-matched doses.
In conclusion, IMGN632 exhibits potent, CD123-specific in vitro activity against AML cell lines, including those with markers of poor prognosis such as MDR1, p53 and FLT3-ITD. IMGN632 is highly active in vivo against Molm-13, Kasumi-3 and MV4-11 disseminated xenografts, resulting in prolonged survival and reduced tumor burden. In addition, IMGN632 is highly active against EOL-1 xenografts, which appear to be resistant to azacitidine and cytarabine in vivo, but highly sensitive to the ADC, resulting in tumor regression and prolonged tumor-free survival. These findings support advancing IMGN632 into clinical trials.
Disclosures
Adams: ImmunoGen, Inc.: Employment. Wilhelm:ImmunoGen, Inc.: Employment. Harvey:ImmunoGen, Inc.: Employment. Bai:ImmunoGen, Inc.: Employment. Yoder:ImmunoGen, Inc.: Employment. Kovtun:ImmunoGen, Inc.: Employment. Chittenden:ImmunoGen, Inc.: Employment. Pinkas:ImmunoGen, Inc.: Employment.
PDTM-25. STUDY OF ONC201 IN PRE-CLINICAL MODELS OF DIPG
