Abstract Introduction: Pediatric cancers have been reported to be infiltrated with macrophages. Tumor associated macrophages and myeloid derived suppressive cells (MDSCs) are increasingly recognized to play an important role in cancer immune escape, promote tumor progression by suppression of adaptive immunity, and are associated with poor prognosis in adult cancers. Little is known about the presence and the role of these cells in the pathobiology of pediatric cancers. We sought to analyze pediatric sarcomas for the presence of myeloid cells expressing inducible nitric oxide synthase (iNOS), arginase1 (Arg1), and/or indolamine 2,3 deoxygenase (IDO). Concomitantly, we used a murine model of embryonal rhabdomyosarcoma (ERMS) to further explore MDSCs associated with pediatric sarcomas. Methods: We conducted immunohistochemical (IHC) studies on 20 paraffin embedded tumor samples of rhabdomyosarcoma, Ewing's sarcoma, desmoplastic small round cell tumor and malignant peripheral nerve sheath tumor, assessing for CD68, CD163, iNOS, Arg1 and IDO immunoreactivity. In the murine model, 1×106 RMS M3-9-M murine ERMS cells were injected intramuscularly into the gastrocnemius muscle of C57BL/6 mice. Tumors and spleens were excised on days 7, 12, 16 and 19 post injection and tumor and spleen myeloid cell were analyzed for CD11b, CD11c, Gr-1 and IL4R alpha expression and for gene expression of iNOS, Arg1 and IDO using RT-PCR. Results: Pediatric sarcomas demonstrate significant macrophage infiltration with iNOS, Arg1 and IDO present in most of these samples. Murine ERMS also shows substantial numbers of MDSCs both in the tumor and spleen of tumor bearing mice, with increasing numbers as tumor burden increased. Comparisons between different myeloid cells subsets by relative quantitation of gene expression reveal the presence of an expanded population of cells that bear the hallmarks of MDSCs by expressing iNOS and Arg1. Significantly higher IDO expression was seen in MDSCs compared to other myeloid cells as tumors progressed. Conclusion: Macrophages infiltrating pediatric sarcomas expressed markers associated with MDSCs, which have been shown to contribute to immune evasion and augment tumor growth in several other model systems. Similarly, murine ERMS M3-9-M induced accumulation of MDSCs in spleen and tumor microenvironment, with increasing numbers associated with tumor progression, suggesting that they may play an important role in growth of pediatric sarcomas. Immunomodulation of these cells might be a key step in enhancing the effectiveness of immune-based therapies. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 1332.