Purpose: The source of glioblastoma (GBM)-associated immunosuppression remains multifactorial. We sought to clarify and therapeutically target myeloid cell-derived peripheral immunosuppression in patients with GBM.Experimental Design: Direct ex vivo T-cell function, serum Arginase I (ArgI) levels, and circulating myeloid lineage populations were compared between patients with GBM and normal donors or patients with other intracranial tumors. Immunofunctional assays were conducted using bulk and sorted cell populations to explore the potential transfer of myeloid cell-mediated immunosuppression and to identify a potential mechanism for these effects. ArgI-mediated immunosuppression was therapeutically targeted in vitro through pharmacologic inhibition or arginine supplementation.Results: We identified a significantly expanded population of circulating, degranulated neutrophils associated with elevated levels of serum ArgI and decreased T-cell CD3z expression within peripheral blood from patients with GBM. Sorted CD11b þ cells from patients with GBM were found to markedly suppress normal donor T-cell function in coculture, and media harvested from mitogen-stimulated GBM peripheral blood mononuclear cell (PBMC) or GBM-associated mixed lymphoid reactions showed ArgI levels that were significantly higher than controls. Critically, T-cell suppression in both settings could be completely reversed through pharmacologic ArgI inhibition or with arginine supplementation. Conclusions: These data indicate that peripheral cellular immunosuppression in patients with GBM is associated with neutrophil degranulation and elevated levels of circulating ArgI, and that T-cell function can be restored in these individuals by targeting ArgI. These data identify a novel pathway of GBM-mediated suppression of cellular immunity and offer a potential therapeutic window for improving antitumor immunity in affected patients. Clin Cancer Res; 17(22); 6992-7002. Ó2011 AACR.

The pathogen Brucella melitensis secretes a Toll/interleukin-1 receptor (TIR) domain containing protein that abrogates host innate immune responses. In this study, we have characterized the biochemical interactions of Brucella TIR-like protein TcpB with host innate immune adaptor proteins. Using protein-fragment complementation assays based on Gaussia luciferase and green fluorescent protein, we find that TcpB interacts directly with MyD88 and that this interaction is significantly stronger than the interaction of TcpB with TIRAP, the only other adaptor protein that detectably interacts with TcpB. Surprisingly, the TcpB–MyD88 interaction depends on the death domain (DD) of MyD88, and TcpB does not interact with the isolated TIR domain of MyD88. TcpB disrupts MyD88DD-MyD88DD, MyD88DD-MyD88TIR and MyD88DD-MyD88 interactions but not MyD88–MyD88 or MyD88TIR-MyD88TIR interactions. Structural models consistent with these results suggest how TcpB might inhibit TLR signaling by targeting MyD88 via a DD–TIR domain interface.

Sea anemones were subjected to mild trauma consisting of a 2 min immersion in calcium-depleted seawater. The trauma caused a loss of vibration sensitivity that spontaneously recovered within 50 min of returning the anemones to calcium containing seawater. Apparently, recovery is conferred by proteins contained in fraction gamma, a chromatographic fraction of homogenized mucus collected at the base of anemones allowed to recover from similar trauma. On silver stained SDS-PAGE gels, fraction gamma consists of a single band having an estimated mass of 55 kDa. Fraction gamma is alone sufficient to repair hair bundle mechanoreceptors in anemones. Its biological activity is enhanced in the presence of exogenously supplied ATP, but not GTP nor ADP-ribose. Biotinylated fraction gamma binds to hair bundles. The hypothesis that fraction gamma consists of Hsp60 proteins was tested. Commercial antibodies to Hsp60 label a band at 55 kDa in western blots. Hsp60 antibodies label hair bundles in traumatized anemones but not in untreated controls. Dilute Hsp60 antiserum (but not nonimmune serum) delays the spontaneous recovery of vibration sensitivity in anemones subjected to mild trauma. Thus, fraction gamma likely consists of Hsp60, or a Hsp60-like protein, that functions on the extracellular face of the plasma membrane to restore function to traumatized hair bundles.

Cells respond to extracellular cues through a variety of receptors on the surface. These signals once transduced across the cell membrane, activate protein tyrosine kinases, which through phosphorylation of substrates on key tyrosine residues, are able to control cellular growth, activation and differentiation pathways. Recent data suggest that protein tyrosine kinases are critical in integrating signals from various cellular receptors, including pathogen detection receptors that mediate the host innate immune response. In this article, we have reviewed the roles of tyrosine kinases of the Tec, FAK, Fps, Fer, Syk, Src and TAM-receptor families in toll-like receptor signaling. The shared roles of these tyrosine phosphorylation mediators in host defense, inflammation, autoimmune disease and oncogenesis provides promising avenues for the use of their inhibitors in multiple disorders.

<p>PDF file - 45K</p>

<p>PDF file - 46K</p>

<div>Abstract<p><b>Purpose:</b> The source of glioblastoma (GBM)-associated immunosuppression remains multifactorial. We sought to clarify and therapeutically target myeloid cell–derived peripheral immunosuppression in patients with GBM.</p><p><b>Experimental Design:</b> Direct <i>ex vivo</i> T-cell function, serum Arginase I (ArgI) levels, and circulating myeloid lineage populations were compared between patients with GBM and normal donors or patients with other intracranial tumors. Immunofunctional assays were conducted using bulk and sorted cell populations to explore the potential transfer of myeloid cell–mediated immunosuppression and to identify a potential mechanism for these effects. ArgI-mediated immunosuppression was therapeutically targeted <i>in vitro</i> through pharmacologic inhibition or arginine supplementation.</p><p><b>Results:</b> We identified a significantly expanded population of circulating, degranulated neutrophils associated with elevated levels of serum ArgI and decreased T-cell CD3ζ expression within peripheral blood from patients with GBM. Sorted CD11b⁺ cells from patients with GBM were found to markedly suppress normal donor T-cell function in coculture, and media harvested from mitogen-stimulated GBM peripheral blood mononuclear cell (PBMC) or GBM-associated mixed lymphoid reactions showed ArgI levels that were significantly higher than controls. Critically, T-cell suppression in both settings could be completely reversed through pharmacologic ArgI inhibition or with arginine supplementation.</p><p><b>Conclusions:</b> These data indicate that peripheral cellular immunosuppression in patients with GBM is associated with neutrophil degranulation and elevated levels of circulating ArgI, and that T-cell function can be restored in these individuals by targeting ArgI. These data identify a novel pathway of GBM-mediated suppression of cellular immunity and offer a potential therapeutic window for improving antitumor immunity in affected patients. <i>Clin Cancer Res; 17(22); 6992–7002. ©2011 AACR</i>.</p></div>

<p>PDF file - 46K</p>