Dendritic cell development in haematological malignancies and neoplasia

Abstract

Dendritic cells (DC) play a major role in the detection of antigens, initiation of immunity and induction and regulation of tolerance. DCs are Bone Marrow (BM) derived and their development may be influenced by haematological malignancy in several ways. Firstly, myelodysplastic, myeloproliferative or leukaemic transformation of bone marrow progenitors may involve DC precursors directly, when they become part of a malignant clone, or indirectly when neoplastic expansion of other lineages compromises the development of DCs. Secondly, neoplasia of the dendritic cell lineage itself may occur in a heterogeneous group of histiocytic disorders including Langerhans cell histiocytosis (LCH) and Erdheim Chester disease (ECD). The first part of this thesis concerns the effect of haematological malignancies on the generation of DCs; in particular the relationship between DC, monocyte, B and NK lymphoid cell (DCML) deficiency, caused by GATA2 gene mutations, sporadic myelodysplastic syndrome (MDS) and acute myeloid leukaemia (AML). Comprehensive flow cytometric methods for profiling of the DCs and monocytes in the peripheral blood (PB) and peripheral tissues, and the CD34+ progenitors of the BM have been developed. Using these methods, it was possible to see that patients with the three diseases differed on several grounds. DCML deficiencies associated with a younger age of presentation, better preserved haemoglobin, neutrophils and platelets and much more severe defects of DCs, monocytes, and lymphoid cells. In patients with MDS, deficiencies are more moderate and mononuclear cell generation in AML patients is surprisingly preserved. Serum levels of Fms-like tyrosine kinase 3 ligand (FLT3L) also differed, with massively increased levels in DCML deficiency, stable levels in MDS and deficiencies in AML. Finally, strategies of grouping of AML patients have proven to be possible by using progenitor cell or DC related phenotypic markers which may correlate to known cytogenetic abnormalities. The second part of this thesis explores the origin of the cells of LCH and ECD, both neoplasia involving DC-like cells. RT-PCR assays were developed to detect the BRAFV600E mutation, which is present in ~60% of LCH and ECD viii cases. Surprisingly, it was seen that LCH and ECD express BRAFV600E in differing peripheral blood fractions. In LCH, 78-94% of all BRAFV600E alleles were found in HLA-DR+Lineage- cells, localised to the monocytes and CD1c+ myeloid DCs (mDCs). However in ECD, 80-82% of mutated alleles were recovered from the HLA-DR-Lineage- quadrant, localised to CD33+ early myeloid cells, and no mutation was found in the monocytes or mDCs. The final part of the thesis examines whether the circulating cells which carry the BRAFV600E mutation in LCH and ECD can actually develop into LCH-like cells. In order to do this, culture systems were developed with a variety of cytokines and conditions. These experiments showed that CD14+ and CD16+ monocytes and CD1c+ mDCs could all induce the Langerhans Cell (LC) markers CD1a and Langerin in response to GM-CSF, TGFβ and BMP7, however only the CD1c+ mDCs could express Langerin at the high levels seen in LCs, along with EpCam and Birbeck granules. Several techniques have been developed to study DCs in haematological malignancies. Throughout this thesis, these techniques have provided valuable data on the development and homeostasis of human DCs.