Abstract P118: Analysis of macrophage function and histone deacetylase inhibition in neuroblastoma
Authors: Brumfield Gabrielle L. , Knoche Shelby M. , Larson Alaina C. , Poelaert Brittany J. , Goetz Benjamin T. , Pandey Poomy , Coulter Donald W. , Solheim Joyce C. 
Published: December 2021

Abstract
Background: Neuroblastoma is the third most common childhood cancer and accounts for 12% of cancer-associated deaths in children under the age of 15. Treatment of neuroblastoma with the histone deacetylase inhibitor (HDACi) vorinostat induces increased infiltration of macrophages with upregulated immune cell-surface receptors. Neuroblastoma cells release VEGF and M-CSF, which may alter intratumoral macrophage populations. VEGF has also been implicated in alteration of amyloid precursor protein family processing. Our lab demonstrated that amyloid precursor protein 2 (APLP2), a member of the amyloid precursor protein family, plays an important role in the migration of tumor cells. APLP2 is known to be expressed by macrophages, but no studies have previously examined macrophage functions that are impacted by APLP2 in the context of neuroblastoma disease and its treatment by HDACi drugs. Because of the high morbidity and mortality associated with neuroblastoma, studies such as this one that are designed to comprehend the interaction of immunity and treatment in neuroblastoma are clinically significant. Methods: We treated neuroblastoma tumor cells (Neuro-2a) in vitro with M344, an HDACi with structural similarity to vorinostat, and assessed viability through MTT assay. In addition, we generated mice with APLP2 knockout in cells expressing the Csf-1 receptor (a protein characteristically expressed by macrophages and dendritic cells). Polarization of macrophages isolated from the macrophage-targeted APLP2-knockout mice was achieved through treatment with IFN-γ and LPS (M1) or IL-4 (M2). Macrophages were then analyzed through western blotting and flow cytometry for APLP2 expression and polarization markers. Results: Following polarization, macrophages collected from the bone marrow of macrophage-targeted APLP2-knockout mice have an altered distribution of M1 and M2 sub-populations, which are macrophage sub-populations that differ in their migratory capabilities, as well as in their abilities to stimulate or suppress anti-tumor immunity. Furthermore, we showed that M1 and M2 subpopulations of bone marrow-derived macrophages from normal mice differ in their expression of APLP2. We also demonstrated that M344 decreased neuroblastoma cell growth. Thus, APLP2 is influential in macrophage biology, and we have created a novel mouse model for defining its specific contributions in mice treated with HDACi drugs that influence macrophage biology. Conclusions: We have made progress in understanding the impact of the HDACi drug M344 on neuroblastoma cells and are ready to analyze its impact on macrophage/dendritic cell populations in a syngeneic neuroblastoma mouse model, as well as to define the role of APLP2 in the function of these cell populations in the context of neuroblastoma. In addition to their potential contribution to the development of new neuroblastoma therapies, the results from this study are expected to expand our comprehension of macrophage function and regulation, and thus will be of broad value in the immunology and oncology fields.
Citation Format: Gabrielle L. Brumfield, Shelby M. Knoche, Alaina C. Larson, Brittany J. Poelaert, Benjamin T. Goetz, Poomy Pandey, Donald W. Coulter, Joyce C. Solheim. Analysis of macrophage function and histone deacetylase inhibition in neuroblastoma [abstract]. In: Proceedings of the AACR-NCI-EORTC Virtual International Conference on Molecular Targets and Cancer Therapeutics; 2021 Oct 7-10. Philadelphia (PA): AACR; Mol Cancer Ther 2021;20(12 Suppl):Abstract nr P118.