Steven Best

The HBS1L-MYB intergenic interval associated with elevated HbF levels shows characteristics of a distal regulatory region in erythroid cells

HBS1L-MYB intergenic polymorphism (HMIP) on chromosome 6q23 is associated with elevated fetal hemoglobin levels and has pleiotropic effects on several hematologic parameters. To investigate potential regulatory activity in the region, we have measured sensitivity of the sequences to DNase I cleavage that identified 3 tissue-specific DNase I hypersensitive sites in the core intergenic interval. Chromatin immunoprecipitation with microarray (ChIP-chip) analysis showed strong histone acetylation in a defined interval of 65 kb corresponding to the core HBS1L-MYB intergenic region in primary human erythroid cells but not in non-MYB-expressing HeLa cells. ChIP-chip analysis also identified several potential cis-regulatory elements as strong GATA-1 signals that coincided with the DNase I hypersensitive sites present in MYB-expressing erythroid cells. We suggest that HMIP contains regulatory sequences that could be important in hematopoiesis by controlling MYB expression. This study provides the functional link between genetic association of HMIP with control of fetal hemoglobin and other hematologic parameters. We also present a large-scale analysis of histone acetylation as well as RNA polymerase II and GATA-1 interactions on chromosome 6q, and alpha and beta globin gene loci. The data suggest that GATA-1 regulates numerous genes of various functions on chromosome 6q.

Bio‐electrospraying whole human blood: analysing cellular viability at a molecular level

Bio‐electrosprays, pioneered in 2005, have undergone several developmental studies which have seen this technique evolve as a novel direct in vivo tissue engineering and regenerative medicinal strategy. Those studies have been a hallmark for electrosprays; however, in this communication we report our on‐going developmental investigations for exploring bio‐electrosprays as a potential medical device and diagnostic protocol. The studies reported here demonstrate the ability to directly jet whole human blood without affecting the genetic make‐up, which has been interrogated by way of reverse transcription–polymerase chain reaction (RT–PCR) in comparison to controls (p = 0.7337). These studies demonstrate bio‐electrosprays as a possible diagnostic protocol. Copyright

High concordance of genomic and cytogenetic aberrations between peripheral blood and bone marrow in myelodysplastic syndrome (MDS)

Bone marrow (BM) genetic abnormalities in myelodysplastic syndrome (MDS) have provided important biological and prognostic information; however, frequent BM sampling in older patients has been associated with significant morbidity. Utilizing single-nucleotide polymorphism array (SNP-A) and targeted gene sequencing (TGS) of 24 frequently mutated genes in MDS, we assessed the concordance of genetic abnormalities in BM and peripheral blood (PB) samples concurrently from 201 MDS patients. SNP-A karyotype in BM was abnormal in 108 (54%) and normal in 93 (46%) patients, with 95% (190/201) having an identical PB karyotype. The median copy number (CN) for deletions was significantly lower in BM (CN:1.4 (1–1.9)) than in PB (CN:1.5 (1–1.95), P<0.001). Using TGS, 71% (130/183) patients had BM somatic mutations with 95% (124/130) having identical mutations in PB. The mutant allele burden was lower in PB (median 27% (1–96%)) compared with BM (median 29% (1–100%); P=0.14) with no significant difference in the number, types of mutations or World Health Organization subtype. In all patients with discordant SNP (n=11) and mutation (n=6) profiles between BM and PB, shared abnormalities were always present irrespective of treatment status. Overall, 86% of patients had a clonal aberration with 95% having identical SNP-A karyotype and mutations in PB, thus enabling frequent assessment of response to treatment and disease evolution especially in patients with fibrotic or hypocellular marrows.

CSNK1A1 mutations and isolated del(5q) abnormality in myelodysplastic syndrome: a retrospective mutational analysis

BACKGROUND A mechanism for clonal growth advantage in isolated del(5q) disease remains elusive. CSNK1A1 resides on the critically deleted region, and deletion of this gene has been shown in mouse knockout and transplantation studies to produce some characteristics of bone marrow failure, including a proliferative advantage. We aimed to establish the frequency, nature, and clinical association of CSNK1A1 mutations in patients with myelodysplastic syndrome and associated myeloid neoplasms. METHODS Between June 1, 2004, and May 31, 2014, in Kings College (London, UK), we did whole-exome sequencing of five patients with isolated del(5q) followed by targeted screening for CSNK1A1 mutations and 20 myelodysplastic syndrome-associated mutations in 245 additional patients with myeloid neoplasms. All patients met present WHO diagnostic criteria for myelodysplastic syndrome and other related myeloid neoplasms. FINDINGS 39 (16%) of 250 patients with myeloid neoplasms had isolated del(5q), of whom seven (18%) had CSNK1A1 mutations. All these mutations were missense and presented in a highly conserved region that is implicated in ATP catalysis. Serial sampling and response to lenalidomide treatment showed that CSNK1A1 mutations were highly associated with the del(5q) clone. Only one patient with a CSNK1A1 mutation showed complete cytogenetic response to lenalidomide. Four (57%) of the seven patients carrying a CSNK1A1 mutation showed disease progression coupled with an increase in mutant allele burden (all four were on lenalidomide). We detected coexisting myelodysplastic syndrome-related gene mutations in patients with CSNK1A1 mutations, including TP53. INTERPRETATION Similar to the effect of TP53 mutations on progression of del(5q) abnormality, mutant CSNK1A1 also gives rise to a poor prognosis in del(5q) abnormality, for which a coupled increase in P53 activation is suggested. CSNK1A1 mutations in del(5q) disease are important in the context of therapeutic manipulation and need incorporation into future prospective studies. FUNDING Leukaemia and Lymphoma Research.

Heterogeneity of the epsilon gamma delta beta-thalassaemias: characterization of three novel English deletions

We have characterized three novel ɛγδβ‐thalassaemia deletions in three English families. Two of the deletions, 114 and 439 kb, removed the entire β‐globin gene complex, including a variable number of flanking olfactory receptor (HOR) genes. The 98‐kb deletion extended 90‐kb upstream of the ɛ gene to 8 kb upstream of the Gγ‐gene, leaving the γ,δ and β‐genes intact. The 439 kb deletion is the largest deletion reported so far to cause ɛγδβ‐thalassaemia; heterozygotes for this deletion were variably affected by neonatal haemolytic anaemia. Two of the deletions were de novo. Breakpoints of all three deletions occurred within regions of L1 or Alu repeats and contained short regions of direct homology between the flanking sequences, a feature that is likely to have contributed to the illegitimate recombinations.

Mixed T Cell Chimerism After Allogeneic Hematopoietic Stem Cell Transplantation for Severe Aplastic Anemia Using an Alemtuzumab-Containing Regimen Is Shaped by Persistence of Recipient CD8 T Cells.

Highlights • Clinical outcomes are excellent using an alemtuzumab-containing hematopoietic stem cell transplantation regimen for aplastic anemia.• Outcomes are excellent despite prolonged abnormality of the T cell profile.• Recipient-derived CD8 T cells shape persistent mixed chimerism.

GATA2 monoallelic expression underlies reduced penetrance in inherited GATA2-mutated MDS/AML.

Saudi Arabian Ministry of Higher Education through a doctoral scholarship awarded to A.F.A.S. and a Bloodwise Programme grant (14032) awarded to J.F., T.V., and I.D.

Proteomic and genomic integration identifies kinase and differentiation determinants of kinase inhibitor sensitivity in leukemia cells.

Leukemia accepted article preview online, 12 December 2017. doi:10.1038/leu.2017.349.