interleukin-8 and Rhabdomyosarcoma

Suppression of the host's immune system plays a major role in cancer progression. Tumor signaling of programmed death 1 (PD1) on T cells and expansion of myeloid-derived suppressor cells (MDSCs) are major mechanisms of tumor immune escape. We sought to target these pathways in rhabdomyosarcoma (RMS), the most common soft tissue sarcoma of childhood. Murine RMS showed high surface expression of PD-L1, and anti-PD1 prevented tumor growth if initiated early after tumor inoculation; however, delayed anti-PD1 had limited benefit. RMS induced robust expansion of CXCR2(+)CD11b(+)Ly6G(hi) MDSCs, and CXCR2 deficiency prevented CD11b(+)Ly6G(hi) MDSC trafficking to the tumor. When tumor trafficking of MDSCs was inhibited by CXCR2 deficiency, or after anti-CXCR2 monoclonal antibody therapy, delayed anti-PD1 treatment induced significant antitumor effects. Thus, CXCR2(+)CD11b(+)Ly6G(hi) MDSCs mediate local immunosuppression, which limits the efficacy of checkpoint blockade in murine RMS. Human pediatric sarcomas also produce CXCR2 ligands, including CXCL8. Patients with metastatic pediatric sarcomas display elevated serum CXCR2 ligands, and elevated CXCL8 is associated with diminished survival in this population. We conclude that accumulation of MDSCs in the tumor bed limits the efficacy of checkpoint blockade in cancer. We also identify CXCR2 as a novel target for modulating tumor immune escape and present evidence that CXCR2(+)CD11b(+)Ly6G(hi) MDSCs are an important suppressive myeloid subset in pediatric sarcomas. These findings present a translatable strategy to improve the efficacy of checkpoint blockade by preventing trafficking of MDSCs to the tumor site.

Topics: Adolescent; Adult; Animals; Antibodies, Monoclonal; Antigens, Ly; Antineoplastic Combined Chemotherapy Protocols; B7-H1 Antigen; Case-Control Studies; CD11b Antigen; Cell Line, Tumor; Chemokine CXCL1; Chemotaxis; Child; Child, Preschool; Cytokines; Female; Humans; Interleukin-8; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Neoplastic Stem Cells; Prognosis; Receptors, Interleukin-8B; Rhabdomyosarcoma; Time Factors; Tumor Burden; Tumor Escape; Tumor Microenvironment; Young Adult

Rhabdomyosarcoma (RMS) is the most common soft-tissue sarcoma of adolescence and childhood. Because RMS tumors are highly vascularized, we sought to determine which factors secreted by RMS cells are crucial in stimulating angiogenesis in response to hypoxia. To address this issue, we evaluated expression of several proangiogenic factors [interleukin (IL)-8, vascular endothelial growth factor (VEGF), fibroblast growth factor (FGF)-2, stromal-derived factor (SDF)-1, hepatocyte growth factor (HGF) and leukemia inhibitory factor (LIF)] in 8 human RMS cell lines in both normal steady-state and hypoxic conditions. We found by real-time quantitative polymerase chain reaction (RQ-PCR) and confirmed by enzyme-linked immunosorbent assay (ELISA) that from all the factors evaluated, IL-8, whose expression is very low in normoxia, had been very highly expressed and secreted by RMS cells lines during hypoxic conditions ( approximately 40-170 times). Interestingly, this upregulation was not affected by knocking down hypoxia-inducible factor (HIF)-1alpha, but was inhibited by mitogen-activated protein kinase (MAPK)p42/44 and phosphatidylinositaol 3-kinase (PI3K)/AKT pathway inhibitors. This suggests that IL-8 expression is regulated in an activating protein (AP)-1- and nuclear factor (NF)-kappaB-dependent manner. Furthermore, we found that conditioned media (CM) harvested from RMS cells exposed to hypoxia activated and stimulated chemotactic responses in human umbilical vein endothelial cells (HUVECs) and that IL-8 was responsible for hypoxia-related effects. Finally, by employing shRNA, the expression of IL-8 in human RH-30 cells was downregulated. We noticed that such RMS cells, if injected into skeletal muscles of immunodeficient mice, have a reduced ability for tumor formation. We conclude that IL-8 is a pivotal proangiogenic factor released by human RMS cells in hypoxic conditions and that the targeting of IL-8 may prove to be a novel and efficient strategy for inhibiting RMS growth.

Topics: Animals; Cell Hypoxia; Cell Line; Cell Line, Tumor; Chemotaxis; Culture Media, Conditioned; Endothelial Cells; Enzyme-Linked Immunosorbent Assay; Humans; Hypoxia; Interleukin-8; Mice; Mice, SCID; Mitogen-Activated Protein Kinase 1; Mitogen-Activated Protein Kinase 3; Neovascularization, Pathologic; Phosphatidylinositol 3-Kinases; Proto-Oncogene Proteins c-akt; Reverse Transcriptase Polymerase Chain Reaction; Rhabdomyosarcoma; RNA Interference; Signal Transduction; Up-Regulation; Xenograft Model Antitumor Assays

LIGHT is a member of the tumor necrosis factor (TNF) superfamily, which binds two known receptors, lymphotoxin-beta receptor (LTbetaR) and the herpesvirus entry mediator (HVEM)/TR2. We investigated the effects of LIGHT on the human rhabdmyosarcoma cell line RD. LIGHT delayed cell proliferation and induced morphological changes of the cells. These effects were not shown by other TNF family ligands such as TNFalpha and LTalpha, which induced the transcriptional activity of nuclear factor-kappaB (NF-kappaB) and NF-kappaB-responsible chemokine productions in the same manner as did LIGHT. LTalpha1beta2, another TNF family ligand for LTbetaR, was shown to have similar activities in RD cells as LIGHT. Both LIGHT and LTalpha1beta2 induced the expression of muscle-specific genes such as smooth muscle (SM) alpha-actin, while TNFalpha and LTalpha did not. These findings indicate that LIGHT may be a novel inducer of RD cell differentiation associated with SM alpha-actin expression through the LTbetaR.

Topics: Actins; Cell Division; Chemokine CCL5; Flow Cytometry; Humans; Interleukin-8; Kinetics; Lymphotoxin-alpha; Membrane Proteins; Muscle Proteins; NF-kappa B; Rhabdomyosarcoma; RNA, Neoplasm; Tumor Cells, Cultured; Tumor Necrosis Factor Ligand Superfamily Member 14; Tumor Necrosis Factor-alpha