Chronic lymphocytic leukemia (CLL) is characterized by apoptosis resistance and a dysfunctional immune system. Previous reports suggested a potential role of myeloid cells in mediating these defects. However, the composition and function of CLL-associated myeloid cells have not been thoroughly investigated in vivo. Using the Eμ-TCL1 mouse model, we observed severe skewing of myeloid cell populations with CLL development. Monocytes and M2-like macrophages infiltrated the peritoneal cavity of leukemic mice. Monocytes also accumulated in the spleen in a CCR2-dependent manner, and were severely skewed toward Ly6C(low) patrolling or nonclassical phenotype. In addition, the percentage of MHC-II(hi) dendritic cells and macrophages significantly dropped in the spleen. Gene expression profiling of CLL-associated monocytes revealed aberrantly high PD-L1 expression and secretion of multiple inflammatory and immunosuppressive cytokines like interleukin-10, tumor necrosis factor-α and CXCL9. In vivo myeloid cell depletion using liposomal Clodronate resulted in a significant control of CLL development accompanied by a pronounced repair of innate immune cell phenotypes and a partial resolution of systemic inflammation. In addition, CLL-associated skewing of T cells toward antigen-experienced phenotypes was repaired. The presented data suggest that targeting nonmalignant myeloid cells might serve as a novel immunotherapeutical strategy for CLL.

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Abstract:The purpose of this study was to identify and characterize fungal natural products (NPs) with in vitro bioactivity towards leukemia cells. We based our screening on a combined analytical and bio-guided approach of LC-DAD-HRMS dereplication, explorative solid-phase extraction (E-SPE), and a co-culture platform of CLL and stromal cells. A total of 289 fungal extracts were screened and we tracked the activity to single compounds in seven of the most active extracts. The novel ophiobolin U was isolated together with the known ophiobolins C, H, K as well as 6-epiophiobolins G, K and N from three fungal strains in the Aspergillus section Usti. Ophiobolins A, B, C and K displayed bioactivity towards leukemia cells with induction of apoptosis at nanomolar concentrations. The remaining ophiobolins were mainly inactive or only slightly active at micromolar OPEN ACCESSMolecules 2013, 18 14630 concentrations. Dereplication of those ophiobolin derivatives possessing different activity in combination with structural analysis allowed a correlation of the chemical structure and conformation with the extent of bioactivity, identifying the hydroxy group at C3 and an aldehyde at C21, as well as the A/B-cis ring structure, as indispensible for the strong activity of the ophiobolins. The known compounds penicillic acid, viridicatumtoxin, calbistrin A, brefeldin A, emestrin A, and neosolaniol monoacetate were identified from the extracts and also found generally cytotoxic.