Bola Hanna

Depletion of CLL-associated patrolling monocytes and macrophages controls disease development and repairs immune dysfunction in vivo.

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 Ly6Clow patrolling or nonclassical phenotype. In addition, the percentage of MHC-IIhi 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.

Chaetoglobosin A preferentially induces apoptosis in chronic lymphocytic leukemia cells by targeting the cytoskeleton

Chronic lymphocytic leukemia (CLL) is an incurable malignancy of mature B cells. One of the major challenges in treatment of CLL is the achievement of a complete remission to prevent relapse of disease originating from cells within lymphoid tissues and subsequent chemoresistance. In search for novel drugs that target CLL cells in protective microenvironments, we performed a fungal extract screen using cocultures of primary CLL cells with bone marrow-derived stromal cells. A secondary metabolite produced by Penicillium aquamarinium was identified as Chaetoglobosin A (ChA), a member of the cytochalasan family that showed preferential induction of apoptosis in CLL cells, even under culture conditions that mimic lymphoid tissues. In vitro testing of 89 CLL cases revealed effective targeting of CLL cells by ChA, independent of bad prognosis characteristics, like 17p deletion or TP53 mutation. To provide insight into its mechanism of action, we showed that ChA targets filamentous actin in CLL cells and thereby induces cell-cycle arrest and inhibits membrane ruffling and cell migration. Our data further revealed that ChA prevents CLL cell activation and sensitizes them for treatment with PI3K and BTK inhibitors, suggesting this compound as a novel potential drug for CLL.

Chd7 is indispensable for mammalian brain development through activation of a neuronal differentiation programme

Mutations in chromatin modifier genes are frequently associated with neurodevelopmental diseases. We herein demonstrate that the chromodomain helicase DNA-binding protein 7 (Chd7), frequently associated with CHARGE syndrome, is indispensable for normal cerebellar development. Genetic inactivation of Chd7 in cerebellar granule neuron progenitors leads to cerebellar hypoplasia in mice, due to the impairment of granule neuron differentiation, induction of apoptosis and abnormal localization of Purkinje cells, which closely recapitulates known clinical features in the cerebella of CHARGE patients. Combinatory molecular analyses reveal that Chd7 is required for the maintenance of open chromatin and thus activation of genes essential for granule neuron differentiation. We further demonstrate that both Chd7 and Top2b are necessary for the transcription of a set of long neuronal genes in cerebellar granule neurons. Altogether, our comprehensive analyses reveal a mechanism with chromatin remodellers governing brain development via controlling a core transcriptional programme for cell-specific differentiation.

Extracellular vesicles in chronic lymphocytic leukemia.

Abstract Extracellular vesicles (EVs) are membrane-enclosed nanoparticles 30 to 1000 nm in size and represent a novel mechanism of cell communication. By transferring RNA and protein from their cell of origin, they can reprogram target cells and thus are involved in changes within the cellular microenvironment – a key player in CLL pathogenesis. In the current study, we were able to isolate EVs of 20 to 300 nm from blood plasma of CLL patients as well as from supernatant of primary CLL cells in culture. Further, proteomic profiling by Coomassie staining of SDS-PAGE gels and by mass spectrometry revealed an EV-specific protein profile. These findings suggest that EVs represent an important mean of CLL cells to interact with other cells, which might contribute to the establishment of a pro-survival microenvironment for CLL cells.

Beyond bystanders: Myeloid cells in chronic lymphocytic leukemia

Tumor-promoting inflammation and escape from immune-mediated tumor destruction have been recognized as hallmarks of cancer, and myeloid cells are key players in these processes. By exploiting the tremendous plasticity of myeloid cells, tumors induce a variety of tumor-supportive and immunosuppressive cell phenotypes like tumor-associated macrophages (TAMs) and myeloid-derived suppressor cells (MDSCs). The relevance of these cell types in hematopoietic malignancies has only recently gained a stronger attention. Chronic lymphocytic leukemia (CLL) is a malignancy of mature B cells that expand in secondary lymphoid organs and the bone marrow, and accumulate in the blood of patients. A large body of evidence suggests that the interactions between CLL cells and non-malignant cells in the tumor microenvironment play a key role in the pathology of this disease. CLL is associated with an inflammatory milieu and defective immune responses. A severe skewing of myeloid and T cells toward leukemia-supportive and immunosuppressive phenotypes have been observed in patient samples and the Eμ-TCL1 mouse model of CLL. Myeloid cells were thereby shown to enhance survival of CLL cells and contribute to apoptosis-resistance, to suppress anti-tumoral immunity, and to be involved in immune deficiency of leukemia patients. In addition, treatment regimens that are currently used for CLL target not only directly the malignant cells, but have also an impact on non-malignant bystander cells, including myeloid cells. This review summarizes current literature on these aspects and gives a perspective on how our current knowledge might be used to design novel immunotherapeutic approaches that can be combined with CLL-targeting drugs to achieve better therapeutic responses in CLL patients.

Control of chronic lymphocytic leukemia development by clonally-expanded CD8 + T-cells that undergo functional exhaustion in secondary lymphoid tissues

Chronic lymphocytic leukemia (CLL) is associated with substantial alterations in T-cell composition and function. However, the role of T-cells in CLL remains largely controversial. Here, we utilized the Eµ-TCL1 mouse model of CLL as well as blood and lymph node samples of CLL patients to investigate the existence of anti-tumoral immune responses in CLL, and to characterize involved immune cell populations. Thereby, we identified an oligoclonal CD8+ effector T-cell population that expands along with CLL progression and controls disease development. We further show that a higher percentage of CD8+ effector T-cells produces IFNγ, and demonstrate that neutralization of IFNγ results in faster CLL progression in mice. Phenotypical and functional analyses of expanded CD8+ effector T-cells show significant differences in disease-affected tissues in mice, with cells in secondary lymphoid organs harboring hallmarks of activation-induced T-cell exhaustion. Notably, we further describe a respective population of exhausted CD8+ T-cells that specifically accumulate in lymph nodes, but not in peripheral blood of CLL patients. Collectively, these data emphasize the non-redundant role of CD8+ T-cells in suppressing CLL progression and highlight their dysfunction that can be exploited as target of immunotherapy in this malignancy.

Tumor necrosis factor receptor signaling is a driver of chronic lymphocytic leukemia that can be therapeutically targeted by the flavonoid wogonin

Chronic lymphocytic leukemia is a malignancy of mature B cells that strongly depend on microenvironmental factors, and their deprivation has been identified as a promising treatment approach for this incurable disease. Cytokine array screening of 247 chronic lymphocytic leukemia serum samples revealed elevated levels of tumor necrosis factor (TNF) receptor-1 which were associated with poor clinical outcome. We detected a microenvironment-induced expression of TNF receptor-1 in chronic lymphocytic leukemia cells in vitro, and an aberrantly high expression of this receptor in the proliferation centers of patients’ lymph nodes. Stimulation of TNF receptor-1 with TNF-α enhanced nuclear factor κ-light-chain-enhancer of activated B cells (NFκB) activity and viability of chronic lymphocytic leukemia cells, which was inhibited by wogonin. The therapeutic effects of wogonin were analyzed in mice after adoptive transfer of Eμ-T-cell leukemia 1 (TCL1) leukemic cells. Wogonin treatment prevented leukemia development when given early after transplantation. The treatment of full-blown leukemia resulted in the loss of the TNF receptor-1 on chronic lymphocytic leukemia cells and their mobilization to blood. Targeting TNF receptor-1 signaling is therefore proposed for the treatment of chronic lymphocytic leukemia.