Ex-Vivo Evaluation of Stx-0712, a CCR2 Cytotoxicity Targeting Chimera (CCR2-CyTAC) for the Treatment of Acute Myeloid Leukemia

Introduction:

Acute Myeloid Leukemia (AML) is a serious and lethal hematologic malignancy characterized by the accumulation of pathogenic cells including abnormal immature myeloid blasts and differentiated malignant monocytes. There currently exists no effective therapies available to specifically target the malignant monocytes in AML patients. Because AML malignant monocytes express high and homogeneous levels of the homing chemokine receptor CCR2, we have developed STX-0712, a CCR2 Cytotoxicity Targeting Chimera (CCR2-CyTAC), which potently binds CCR2 on cells, and when combined with the CyTAC platform antibody, eliminates target cells via enhanced effector mediated killing. We present here the characterization of CCR2 expression in AML patient samples, ex vivo depletion of AML malignant monocytes via treatment with STX-0712, and combination efficacy data with Venetoclax (Ven) and Azacitidine (Aza), supporting AML as a promising indication for STX-0712.

Methods:

For CCR2 characterization we used bone marrow biopsy specimens and peripheral blood mononuclear cells (PBMCs) obtained from AML patients and controls from healthy donors. CCR2 expression was evaluated via flow cytometry. To evaluate the activity of STX-0712 in AML, OncoPrecision's Patient Micro-Avatar (PMAs) technology was employed. This platform promotes the ex-vivo survival of Patient Derived Cells (PDCs) by mimicking the cancer microenvironment through the co-culture with heterologous stromal cells. As such, PMAs allow the simultaneous tracking of pathological and healthy cells using cluster differentiation markers and multiparametric flow cytometry. Heterologous NK cells are also added to the AML PMA system at a 1:1 target to effector ratio.

Results:

First, we evaluated CCR2 expression in peripheral blood and bone marrow of samples from 15 AML patients by flow cytometry. The percentage of CCR2 positive tumor cells in AML samples ranged from 2.4% to 97.1% across different AML subtypes with primitive forms of AML (M0, M1, M2) expressing low levels of CCR2 and monocytic AML (M4, M5) expressing high levels. Analysis of CCR2 expression in subpopulations of AML cells revealed a high and homogeneous expression of CCR2 on CD11b+ malignant monocytes and generally low expression on CD34+ blasts, with the exception M4 subtypes of AML, which in some cases exhibited CD34+ CCR2+ co-expression.

Next, PMAs from the 15 AML samples were treated with STX-0712 and the depletion of AML cells, mediated by Fc gamma receptor 3a positive (FcγRIIIa) natural killer cells, was assessed. The results showed a robust activity of STX-0712 in the CD11b+ malignant population in 12 of the 15 (80%) patients evaluated with an average potency of 3nM and a range of 13-74% target cell killing. STX-0712 had minimal effect on CCR2- populations such as CD34+ blasts or lymphocytes.

Finally, Aza+Ven alone or STX-0712 combined with Aza+Ven were evaluated in the same 15 AML samples and the depletion of tumor cells was assessed. In 9/15 (60%) of AML patient samples, the combination outperformed the single agents alone. STX-0712 outperforms Aza+Ven in targeting differentiated monocytic cells, while Ven-Aza outperformed STX-0712 in targeting undifferentiated blasts. Therefore, the improved therapeutic outcome of the combination of STX-0712 and Aza+Ven could be attributed to the differential targeting of subpopulations of pathological cells rather than to additive/synergic effects against individual cells

Conclusion: These results demonstrate high and homogenous CCR2 expression on malignant populations, ex vivo human target engagement and pharmacodynamic activity of STX-0712 and support single agent efficacy of STX-0712 in myelomonocytic/monocytic AML and the potential value of combining STX-0712 with Aza+Ven in earlier lines of AML treatment.