Investigating the role of immunosurveillance by macrophage in supporting therapeutic strategies using CD47 blockade in MDS.

Abstract

e18582 Background: Myelodysplastic syndrome (MDS) is characterised by disturbances in the maturation and differentiation of haematopoietic stem cells. Whilst chemotherapy and demethylating agents are the mainstay treatment, patient response to treatment varies and there are currently no curative other then bone marrow transplant. Recent immunotherapeutic approaches targeting the CD47 failed to demonstrate additional value when associated to azacitidine or chemotherapy in both MDS and AML, suggesting a possible mechanism of immune-escape of the myeloid clone. The aim of the study was to develop an indirect co-culture system which recapitulates the MDS tumour microenvironment, within which to determine how macrophage responses impact upon CD47 expression in MDS cell lines and survival in the presence of different combinations of standard therapies and anti-CD47 blockade through use of flow cytometry. Initial results in SKM-1 MDS cell line, showed CD47 blockade’s efficacy in decreasing CD47 cell surface expression, both by itself and in combination with standard therapies (p<0.001). Methods: To establish an indirect co-culture system, THP-1 derived macrophages were polarised into pro-inflammatory M1 and anti-inflammatory/pro-tumorigenic M2 phenotypes, which play a key role in immunosurveillance. These were then cultured with SKM-1 cells, using an insert to allow cell-cell interaction without the macrophages directly phagocytosing the SKM-1 cells. Results: CD47 blockade by monoclonal anti-CD47 antibody alone decreased CD47 cell surface expression on SKM-1 cells (<1% cells expressing CD47), however this reversed in the presence of pro-inflammatory M1 macrophages (with 59.81% of SKM-1 cells expressing CD47) (p<0.0001). Presence of azacitidine improved the efficiency of CD47 blockade despite the presence of M1 macrophage, showing a reduction in CD47 expression (dropping from 68% in the absence of M1 macrophages to 41% of cells expressing CD47 in co-culture conditions). Additionally, presence of SKM-1 in co-culture with M1 macrophages with CD47 blockade +/- standard therapy, was also associated with an increase in both M1 and M2 markers, suggesting that M1 macrophage are being repolarised to pro-tumorigenic M2 macrophage by the SKM-1 cells. Conclusions: This assay helps understanding the macrophages response to different treatment agents. Further work is now needed to confirm these findings, including use of patient samples, to better understand which patient groups could benefit from therapies and to also enhance immunosurveillance mechanisms in MDS to create novel therapeutic approaches for this condition.