Early transfusion patterns improve the Molecular International Prognostic Scoring System (IPSS-M) prediction in myelodysplastic syndromes.

BACKGROUND: The Molecular International Prognostic Scoring System (IPSS-M) is the new gold standard for diagnostic outcome prediction in patients with myelodysplastic syndromes (MDS). This study was designed to assess the additive prognostic impact of dynamic transfusion parameters during early follow-up. METHODS: We retrieved complete transfusion data from 677 adult Swedish MDS patients included in the IPSS-M cohort. Time-dependent erythrocyte transfusion dependency (E-TD) was added to IPSS-M features and analyzed regarding overall survival and leukemic transformation (acute myeloid leukemia). A multistate Markov model was applied to assess the prognostic value of early changes in transfusion patterns. RESULTS: Specific clinical and genetic features were predicted for diagnostic and time-dependent transfusion patterns. Importantly, transfusion state both at diagnosis and within the first year strongly predicts outcomes in both lower (LR) and higher-risk (HR) MDSs. In multivariable analysis, 8-month landmark E-TD predicted shorter survival independently of IPSS-M (p < 0.001). A predictive model based on IPSS-M and 8-month landmark E-TD performed significantly better than a model including only IPSS-M. Similar trends were observed in an independent validation cohort (n = 218). Early transfusion patterns impacted both future transfusion requirements and outcomes in a multistate Markov model. CONCLUSION: The transfusion requirement is a robust and available clinical parameter incorporating the effects of first-line management. In MDS, it provides dynamic risk information independently of diagnostic IPSS-M and, in particular, clinical guidance to LR MDS patients eligible for potentially curative therapeutic intervention.

Identification and surveillance of rare relapse-initiating stem cells during complete remission after transplantation.

ABSTRACT: Relapse after complete remission (CR) remains the main cause of mortality after allogeneic stem cell transplantation for hematological malignancies and, therefore, improved biomarkers for early prediction of relapse remains a critical goal toward development and assessment of preemptive relapse treatment. Because the significance of cancer stem cells as a source of relapses remains unclear, we investigated whether mutational screening for persistence of rare cancer stem cells would enhance measurable residual disease (MRD) and early relapse prediction after transplantation. In a retrospective study of patients who relapsed and patients who achieved continuous-CR with myelodysplastic syndromes and related myeloid malignancies, combined flow cytometric cell sorting and mutational screening for persistence of rare relapse-initiating stem cells was performed in the bone marrow at multiple CR time points after transplantation. In 25 CR samples from 15 patients that later relapsed, only 9 samples were MRD-positive in mononuclear cells (MNCs) whereas flowcytometric-sorted hematopoietic stem and progenitor cells (HSPCs) were MRD-positive in all samples, and always with a higher variant allele frequency than in MNCs (mean, 97-fold). MRD-positivity in HSPCs preceded MNCs in multiple sequential samples, in some cases preceding relapse by >2 years. In contrast, in 13 patients in long-term continuous-CR, HSPCs remained MRD-negative. Enhanced MRD sensitivity was also observed in total CD34+ cells, but HSPCs were always more clonally involved (mean, 8-fold). In conclusion, identification of relapse-initiating cancer stem cells and mutational MRD screening for their persistence consistently enhances MRD sensitivity and earlier prediction of relapse after allogeneic stem cell transplantation.

Erythroid Differentiation Enhances RNA Mis-Splicing in SF3B1-Mutant Myelodysplastic Syndromes with Ring Sideroblasts.

Myelodysplastic syndromes with ring sideroblasts (MDS-RS) commonly develop from hematopoietic stem cells (HSC) bearing mutations in the splicing factor SF3B1 (SF3B1mt). Direct studies into MDS-RS pathobiology have been limited by a lack of model systems that fully recapitulate erythroid biology and RS development and the inability to isolate viable human RS. Here, we combined successful direct RS isolation from patient samples, high-throughput multiomics analysis of cells encompassing the SF3B1mt stem-erythroid continuum, and functional assays to investigate the impact of SF3B1mt on erythropoiesis and RS accumulation. The isolated RS differentiated, egressed into the blood, escaped traditional nonsense-mediated decay (NMD) mechanisms, and leveraged stress-survival pathways that hinder wild-type hematopoiesis through pathogenic GDF15 overexpression. Importantly, RS constituted a contaminant of magnetically enriched CD34+ cells, skewing bulk transcriptomic data. Mis-splicing in SF3B1mt cells was intensified by erythroid differentiation through accelerated RNA splicing and decreased NMD activity, and SF3B1mt led to truncations in several MDS-implicated genes. Finally, RNA mis-splicing induced an uncoupling of RNA and protein expression, leading to critical abnormalities in proapoptotic p53 pathway genes. Overall, this characterization of erythropoiesis in SF3B1mt RS provides a resource for studying MDS-RS and uncovers insights into the unexpectedly active biology of the "dead-end" RS. SIGNIFICANCE: Ring sideroblast isolation combined with state-of-the-art multiomics identifies survival mechanisms underlying SF3B1-mutant erythropoiesis and establishes an active role for erythroid differentiation and ring sideroblasts themselves in SF3B1-mutant myelodysplastic syndrome pathogenesis.

Integrated Genomic and Transcriptomic Analysis Improves Disease Classification and Risk Stratification of MDS with Ring Sideroblasts.

PURPOSE: Ring sideroblasts (RS) define the low-risk myelodysplastic neoplasm (MDS) subgroup with RS but may also reflect erythroid dysplasia in higher risk myeloid neoplasm. The benign behavior of MDS with RS (MDSRS+) is limited to SF3B1-mutated cases without additional high-risk genetic events, but one third of MDSRS+ carry no SF3B1 mutation, suggesting that different molecular mechanisms may underlie RS formation. We integrated genomic and transcriptomic analyses to evaluate whether transcriptome profiles may improve current risk stratification. EXPERIMENTAL DESIGN: We studied a prospective cohort of MDSRS+ patients irrespective of World Health Organization (WHO) class with regard to somatic mutations, copy-number alterations, and bone marrow CD34+ cell transcriptomes to assess whether transcriptome profiles add to prognostication and provide input on disease classification. RESULTS: SF3B1, SRSF2, or TP53 multihit mutations were found in 89% of MDSRS+ cases, and each mutation category was associated with distinct clinical outcome, gene expression, and alternative splicing profiles. Unsupervised clustering analysis identified three clusters with distinct hemopoietic stem and progenitor (HSPC) composition, which only partially overlapped with mutation groups. IPSS-M and the transcriptome-defined proportion of megakaryocyte/erythroid progenitors (MEP) independently predicted survival in multivariable analysis. CONCLUSIONS: These results provide essential input on the molecular basis of SF3B1-unmutated MDSRS+ and propose HSPC quantification as a prognostic marker in myeloid neoplasms with RS.

Pseudouridine-modified tRNA fragments repress aberrant protein synthesis and predict leukaemic progression in myelodysplastic syndrome.

Transfer RNA-derived fragments (tRFs) are emerging small noncoding RNAs that, although commonly altered in cancer, have poorly defined roles in tumorigenesis1. Here we show that pseudouridylation (Ψ) of a stem cell-enriched tRF subtype2, mini tRFs containing a 5' terminal oligoguanine (mTOG), selectively inhibits aberrant protein synthesis programmes, thereby promoting engraftment and differentiation of haematopoietic stem and progenitor cells (HSPCs) in patients with myelodysplastic syndrome (MDS). Building on evidence that mTOG-Ψ targets polyadenylate-binding protein cytoplasmic 1 (PABPC1), we employed isotope exchange proteomics to reveal critical interactions between mTOG and functional RNA-recognition motif (RRM) domains of PABPC1. Mechanistically, this hinders the recruitment of translational co-activator PABPC1-interacting protein 1 (PAIP1)3 and strongly represses the translation of transcripts sharing pyrimidine-enriched sequences (PES) at the 5' untranslated region (UTR), including 5' terminal oligopyrimidine tracts (TOP) that encode protein machinery components and are frequently altered in cancer4. Significantly, mTOG dysregulation leads to aberrantly increased translation of 5' PES messenger RNA (mRNA) in malignant MDS-HSPCs and is clinically associated with leukaemic transformation and reduced patient survival. These findings define a critical role for tRFs and Ψ in difficult-to-treat subsets of MDS characterized by high risk of progression to acute myeloid leukaemia (AML).

Challenging conventional karyotyping by next-generation karyotyping in 281 intensively treated patients with AML.

Although copy number alterations (CNAs) and translocations constitute the backbone of the diagnosis and prognostication of acute myeloid leukemia (AML), techniques used for their assessment in routine diagnostics have not been reconsidered for decades. We used a combination of 2 next-generation sequencing-based techniques to challenge the currently recommended conventional cytogenetic analysis (CCA), comparing the approaches in a series of 281 intensively treated patients with AML. Shallow whole-genome sequencing (sWGS) outperformed CCA in detecting European Leukemia Net (ELN)-defining CNAs and showed that CCA overestimated monosomies and suboptimally reported karyotype complexity. Still, the concordance between CCA and sWGS for all ELN CNA-related criteria was 94%. Moreover, using in silico dilution, we showed that 1 million reads per patient would be enough to accurately assess ELN-defining CNAs. Total genomic loss, defined as a total loss ≥200 Mb by sWGS, was found to be a better marker for genetic complexity and poor prognosis compared with the CCA-based definition of complex karyotype. For fusion detection, the concordance between CCA and whole-transcriptome sequencing (WTS) was 99%. WTS had better sensitivity in identifying inv(16) and KMT2A rearrangements while showing limitations in detecting lowly expressed PML-RARA fusions. Ligation-dependent reverse transcription polymerase chain reaction was used for validation and was shown to be a fast and reliable method for fusion detection. We conclude that a next-generation sequencing-based approach can replace conventional CCA for karyotyping, provided that efforts are made to cover lowly expressed fusion transcripts.

A three-dimensional in vitro model of erythropoiesis recapitulates erythroid failure in myelodysplastic syndromes.

Established cell culture systems have failed to accurately recapitulate key features of terminal erythroid maturation, hampering our ability to in vitro model and treat diseases with impaired erythropoiesis such as myelodysplastic syndromes with ring sideroblasts (MDS-RS). We developed an efficient and robust three-dimensional (3D) scaffold culture model supporting terminal erythroid differentiation from both mononuclear (MNC) or CD34+-enriched primary bone marrow cells from healthy donors and MDS-RS patients. While CD34+ cells did not proliferate beyond two weeks in 2D suspension cultures, the 3D scaffolds supported CD34+ and MNC erythroid proliferation over four weeks demonstrating the importance of the 3D environment. CD34+ cells cultured in 3D facilitated the highest expansion and maturation of erythroid cells, including generation of erythroblastic islands and enucleated erythrocytes, while MNCs supported multi-lineage hemopoietic differentiation and cytokine secretion relevant for MDS-RS. Importantly, MDS-RS 3D-cultures supported de novo generation of ring sideroblasts and maintenance of the mutated clone. The 3D cultures effectively model a clonal disease characterized by terminal erythroid failure and can be used to assess therapeutic compounds.

Megakaryocytes harbour the del(5q) abnormality despite complete clinical and cytogenetic remission induced by lenalidomide treatment.

The mechanisms underlying lenalidomide-resistance of del(5q) MDS stem cells remain to be elucidated and may include cell-intrinsic as well as microenvironmental causes. Abnormal hypolobated megakaryocytes constitute one of the hallmarks of del(5q) MDS. We hypothesized that these cells have potential implications for the regulation of haematopoietic stem cells (HSC) similarly to what has recently been described for megakaryocytes in the murine system. Therefore, we conducted a study to determine the response of abnormal hypolobated megakaryocytes to lenalidomide therapy. We studied lenalidomide-treated patients in the MDS-004 trial as well as a cohort seen at our institution. Morphological evaluation at time of complete cytogenetic remission (CCyR) demonstrated the persistence of hypolobated megakaryocytes in all evaluable patients (n = 9). Furthermore, we provide evidence that the abnormal hypolobated morphology is restricted to del(5q) megakaryocytes, both at diagnosis and during CCyR. Using fluorescence in situ hybridisation analysis on flow-sorted stem- and progenitor populations, we observed a similar degree of clonal involvement in megakaryocyte-erythroid-progenitors as in HSC. Taken together, our findings suggest that megakaryocyte morphology might aid in the evaluation of patients where discontinuation of lenalidomide is considered and offers interesting hypotheses for further investigation of lenalidomide resistance.

SF3B1-initiating mutations in MDS-RSs target lymphomyeloid hematopoietic stem cells.

Mutations in the RNA splicing gene SF3B1 are found in >80% of patients with myelodysplastic syndrome with ring sideroblasts (MDS-RS). We investigated the origin of SF3B1 mutations within the bone marrow hematopoietic stem and progenitor cell compartments in patients with MDS-RS. Screening for recurrently mutated genes in the mononuclear cell fraction revealed mutations in SF3B1 in 39 of 40 cases (97.5%), combined with TET2 and DNMT3A in 11 (28%) and 6 (15%) patients, respectively. All recurrent mutations identified in mononuclear cells could be tracked back to the phenotypically defined hematopoietic stem cell (HSC) compartment in all investigated patients and were also present in downstream myeloid and erythroid progenitor cells. While in agreement with previous studies, little or no evidence for clonal (SF3B1 mutation) involvement could be found in mature B cells, consistent involvement at the pro-B-cell progenitor stage was established, providing definitive evidence for SF3B1 mutations targeting lymphomyeloid HSCs and compatible with mutated SF3B1 negatively affecting lymphoid development. Assessment of stem cell function in vitro as well as in vivo established that only HSCs and not investigated progenitor populations could propagate the SF3B1 mutated clone. Upon transplantation into immune-deficient mice, SF3B1 mutated MDS-RS HSCs differentiated into characteristic ring sideroblasts, the hallmark of MDS-RS. Our findings provide evidence of a multipotent lymphomyeloid HSC origin of SF3B1 mutations in MDS-RS patients and provide a novel in vivo platform for mechanistically and therapeutically exploring SF3B1 mutated MDS-RS.

Comprehensive mapping of the effects of azacitidine on DNA methylation, repressive/permissive histone marks and gene expression in primary cells from patients with MDS and MDS-related disease.

Azacitidine (Aza) is first-line treatment for patients with high-risk myelodysplastic syndromes (MDS), although its precise mechanism of action is unknown. We performed the first study to globally evaluate the epigenetic effects of Aza on MDS bone marrow progenitor cells assessing gene expression (RNA seq), DNA methylation (Illumina 450k) and the histone modifications H3K18ac and H3K9me3 (ChIP seq). Aza induced a general increase in gene expression with 924 significantly upregulated genes but this increase showed no correlation with changes in DNA methylation or H3K18ac, and only a weak association with changes in H3K9me3. Interestingly, we observed activation of transcripts containing 15 endogenous retroviruses (ERVs) confirming previous cell line studies. DNA methylation decreased moderately in 99% of all genes, with a median ß-value reduction of 0.018; the most pronounced effects seen in heterochromatin. Aza-induced hypomethylation correlated significantly with change in H3K9me3. The pattern of H3K18ac and H3K9me3 displayed large differences between patients and healthy controls without any consistent pattern induced by Aza. We conclude that the marked induction of gene expression only partly could be explained by epigenetic changes, and propose that activation of ERVs may contribute to the clinical effects of Aza in MDS.

Progression in patients with low- and intermediate-1-risk del(5q) myelodysplastic syndromes is predicted by a limited subset of mutations.

A high proportion of patients with lower-risk del(5q) myelodysplastic syndromes will respond to treatment with lenalidomide. The median duration of transfusion-independence is 2 years with some long-lasting responses, but almost 40% of patients progress to acute leukemia by 5 years after starting treatment. The mechanisms underlying disease progression other than the well-established finding of small TP53-mutated subclones at diagnosis remain unclear. We studied a longitudinal cohort of 35 low- and intermediate-1-risk del(5q) patients treated with lenalidomide (n=22) or not (n=13) by flow cytometric surveillance of hematopoietic stem and progenitor cell subsets, targeted sequencing of mutational patterns, and changes in the bone marrow microenvironment. All 13 patients with disease progression were identified by a limited number of mutations in TP53, RUNX1, and TET2, respectively, with PTPN11 and SF3B1 occurring in one patient each. TP53 mutations were found in seven of nine patients who developed acute leukemia, and were documented to be present in the earliest sample (n=1) and acquired during lenalidomide treatment (n=6). By contrast, analysis of the microenvironment, and of hematopoietic stem and progenitor cells by flow cytometry was of limited prognostic value. Based on our data, we advocate conducting a prospective study aimed at investigating, in a larger number of cases of del(5q) myelodysplastic syndromes, whether the detection of such mutations before and after lenalidomide treatment can guide clinical decision-making.

Mutations in histone modulators are associated with prolonged survival during azacitidine therapy.

Early therapeutic decision-making is crucial in patients with higher-risk MDS. We evaluated the impact of clinical parameters and mutational profiles in 134 consecutive patients treated with azacitidine using a combined cohort from Karolinska University Hospital (n=89) and from King's College Hospital, London (n=45). While neither clinical parameters nor mutations had a significant impact on response rate, both karyotype and mutational profile were strongly associated with survival from the start of treatment. IPSS high-risk cytogenetics negatively impacted overall survival (median 20 vs 10 months; p<0.001), whereas mutations in histone modulators (ASXL1, EZH2) were associated with prolonged survival (22 vs 12 months, p=0.01). This positive association was present in both cohorts and remained highly significant in the multivariate cox model. Importantly, patients with mutations in histone modulators lacking high-risk cytogenetics showed a survival of 29 months compared to only 10 months in patients with the opposite pattern. While TP53 was negatively associated with survival, neither RUNX1-mutations nor the number of mutations appeared to influence survival in this cohort. We propose a model combining histone modulator mutational screening with cytogenetics in the clinical decision-making process for higher-risk MDS patients eligible for treatment with azacitidine.

Aberrant splicing of genes involved in haemoglobin synthesis and impaired terminal erythroid maturation in SF3B1 mutated refractory anaemia with ring sideroblasts.

Refractory anaemia with ring sideroblasts (RARS) is distinguished by hyperplastic inefficient erythropoiesis, aberrant mitochondrial ferritin accumulation and anaemia. Heterozygous mutations in the spliceosome gene SF3B1 are found in a majority of RARS cases. To explore the link between SF3B1 mutations and anaemia, we studied mutated RARS CD34(+) marrow cells with regard to transcriptome sequencing, splice patterns and mutational allele burden during erythroid differentiation. Transcriptome profiling during early erythroid differentiation revealed a marked up-regulation of genes involved in haemoglobin synthesis and in the oxidative phosphorylation process, and down-regulation of mitochondrial ABC transporters compared to normal bone marrow. Moreover, mis-splicing of genes involved in transcription regulation, particularly haemoglobin synthesis, was confirmed, indicating a compromised haemoglobinization during RARS erythropoiesis. In order to define the phase during which erythroid maturation of SF3B1 mutated cells is most affected, we assessed allele burden during erythroid differentiation in vitro and in vivo and found that SF3B1 mutated erythroblasts showed stable expansion until late erythroblast stage but that terminal maturation to reticulocytes was significantly reduced. In conclusion, SF3B1 mutated RARS progenitors display impaired splicing with potential downstream consequences for genes of key importance for haemoglobin synthesis and terminal erythroid differentiation.

SF3B1 mutation identifies a distinct subset of myelodysplastic syndrome with ring sideroblasts.

Refractory anemia with ring sideroblasts (RARS) is a myelodysplastic syndrome (MDS) characterized by isolated erythroid dysplasia and 15% or more bone marrow ring sideroblasts. Ring sideroblasts are found also in other MDS subtypes, such as refractory cytopenia with multilineage dysplasia and ring sideroblasts (RCMD-RS). A high prevalence of somatic mutations of SF3B1 was reported in these conditions. To identify mutation patterns that affect disease phenotype and clinical outcome, we performed a comprehensive mutation analysis in 293 patients with myeloid neoplasm and 1% or more ring sideroblasts. SF3B1 mutations were detected in 129 of 159 cases (81%) of RARS or RCMD-RS. Among other patients with ring sideroblasts, lower prevalence of SF3B1 mutations and higher prevalence of mutations in other splicing factor genes were observed (P < .001). In multivariable analyses, patients with SF3B1 mutations showed significantly better overall survival (hazard ratio [HR], .37; P = .003) and lower cumulative incidence of disease progression (HR = 0.31; P = .018) compared with SF3B1-unmutated cases. The independent prognostic value of SF3B1 mutation was retained in MDS without excess blasts, as well as in sideroblastic categories (RARS and RCMD-RS). Among SF3B1-mutated patients, coexisting mutations in DNA methylation genes were associated with multilineage dysplasia (P = .015) but had no effect on clinical outcome. TP53 mutations were frequently detected in patients without SF3B1 mutation, and were associated with poor outcome. Thus, SF3B1 mutation identifies a distinct MDS subtype that is unlikely to develop detrimental subclonal mutations and is characterized by indolent clinical course and favorable outcome.

FISH Detection of X and Y Chromosomes in Combination with Immunofluorescence to Study Contribution of Transplanted Cells to Skeletal Muscle Fibers.

During the past decades, several studies in animals have displayed the ability of cells from the bone marrow (BM) to participate in regeneration of various tissues including skeletal muscle tissue. Studies in mice have demonstrated that regular physical activity is sufficient to induce contribution of BM derived cells to the skeletal muscle tissue, suggesting that this is part of the physiological remodeling of skeletal muscle. To analyze whether BM-derived cells participate in skeletal muscle remodeling in human, we developed a protocol of immunofluorescence in combination with fluorescence in situ hybridization (FISH) that enables the detection of male donor bone marrow cell contribution to female skeletal muscle tissue.

Bone marrow derived cells in adult skeletal muscle tissue in humans.

BACKGROUND: During the past decade, several animal studies have demonstrated that in addition to local cells, cells from the bone marrow (BM) possess the ability to contribute to regeneration of injured skeletal muscle tissue. In addition, in mice, regular physical activity has been displayed to be a sufficient stimulus for BM-derived cell contribution to the muscle, indicating that this is part of the ongoing physiological remodeling of skeletal muscle. However, whether BM-derived cells participate in human skeletal muscle remodeling is not known. To this end, we analyzed the incorporation of BM-derived cells in healthy human skeletal muscle in women transplanted with male BM. METHODS: Skeletal muscle biopsies were obtained from the m. vastus lateralis of women transplanted with male donor hematopoietic stem cells 6 to 12 years earlier. Healthy women served as controls. Immunohistochemical staining for skeletal muscle fibers, satellite cells (SCs) or endothelial cells (ECs) combined with fluorescent in situ hybridization (FISH) of X and Y chromosomes was used to identify cells of BM origin within the biopsies. Three dimensional confocal imaging was performed to demonstrate colocalization of Y chromosome and DAPI within muscle fibers. To further investigate whether BM-derived cells incorporate into the SC niche, myoblasts were extracted from the biopsies from the transplanted women, cultured, and analyzed using XY FISH and immunocytochemistry. RESULTS: Three dimensional confocal imaging indisputably demonstrated colocalization of Y chromosome and DAPI within muscle fibers. Some Y chromosomes were found within centrally located nuclei. No Y chromosomes were detected in CD56+ SCs in the tissue sections nor in the myoblasts cultured from the extracted SCs. Y chromosome+ ECs were found in all sections from the transplanted subjects. No Y chromosomes were found in the skeletal muscle biopsies obtained from healthy control women. CONCLUSIONS: We demonstrate that BM-derived cells contribute to skeletal muscle fibers and ECs. Our results support that BM contribution to skeletal muscle occurs via direct fusion to muscle fibers, and that the contributing cells derive from the hematopoietic lineage. Thus, the present findings encourage further studies of the importance of this process for the physiological adaptation occurring throughout life.

The transporter ABCB7 is a mediator of the phenotype of acquired refractory anemia with ring sideroblasts.

Refractory anemia with ring sideroblasts (RARS) is characterized by mitochondrial ferritin (FTMT) accumulation and markedly suppressed expression of the iron transporter ABCB7. To test the hypothesis that ABCB7 is a key mediator of ineffective erythropoiesis of RARS, we modulated its expression in hematopoietic cells. ABCB7 up and downregulation did not influence growth and survival of K562 cells. In normal bone marrow, ABCB7 downregulation reduced erythroid differentiation, growth and colony formation, and resulted in a gene expression pattern similar to that observed in intermediate RARS erythroblasts, and in the accumulation of FTMT. Importantly, forced ABCB7 expression restored erythroid colony growth and decreased FTMT expression level in RARS CD34+ marrow cells. Mutations in the SF3B1 gene, a core component of the RNA splicing machinery, were recently identified in a high proportion of patients with RARS and 11 of the 13 RARS patients in this study carried this mutation. Interestingly, ABCB7 exon usage differed between normal bone marrow and RARS, as well as within the RARS cohort. In addition, SF3B1 silencing resulted in downregulation of ABCB7 in K562 cells undergoing erythroid differentiation. Our findings support that ABCB7 is implicated in the phenotype of acquired RARS and suggest a relation between SF3B1 mutations and ABCB7 downregulation.

Clinical effect of increasing doses of lenalidomide in high-risk myelodysplastic syndrome and acute myeloid leukemia with chromosome 5 abnormalities.

BACKGROUND: Patients with chromosome 5 abnormalities and high-risk myelodysplastic syndromes or acute myeloid leukemia have a poor outcome. We hypothesized that increasing doses of lenalidomide may benefit this group of patients by inhibiting the tumor clone, as assessed by fluorescence in situ hybridization for del(5q31). DESIGN AND METHODS: Twenty-eight patients at diagnosis or with relapsed disease and not eligible for standard therapy (16 with acute myeloid leukemia, 12 with intermediate-risk 2 or high-risk myelodysplastic syndrome) were enrolled in this prospective phase II multicenter trial and treated with lenalidomide up to 30 mg daily for 16 weeks. Three patients had isolated del(5q), six had del(5q) plus one additional aberration, 14 had del(5q) and a complex karyotype, four had monosomy 5, and one had del(5q) identified by fluorescence in situ hybridization only. RESULTS: Major and minor cytogenetic responses, assessed by fluorescence in situ hybridization, were achieved in 5/26 (19%) and 2/26 (8%) patients, respectively, who received one or more dose of lenalidomide, while two patients achieved only a bone marrow response. Nine of all 26 patients (35%) and nine of the ten who completed the 16 weeks of trial responded to treatment. Using the International Working Group criteria for acute myeloid leukemia and myelodysplastic syndrome the overall response rate in treated patients with acute myeloid leukemia was 20% (3/15), while that for patients with myelodysplastic syndrome was 36% (4/11). Seven patients stopped therapy due to progressive disease and nine because of complications, most of which were disease-related. Response rates were similar in patients with isolated del(5q) and in those with additional aberrations. Interestingly, patients with TP53 mutations responded less well than those without mutations (2/13 versus 5/9, respectively; P=0.047). No responses were observed among 11 cases with deleterious TP53 mutations. CONCLUSIONS: Our data support a role for higher doses of lenalidomide in poor prognosis patients with myelodysplastic syndrome and acute myeloid leukemia with deletion 5q. (Clinicaltrials.gov identifier NCT00761449).

Constitutional inv(3) in myelodysplastic syndromes.

The constitutional pericentric inversion on chromosome 3, inv(3), is rarely found in a normal population. The aim of our study was to investigate its possible link to hematologic malignancy. Chromosomes from bone marrow cells in 890 patients with hematologic disorders were analyzed with the Q-banding technique. Thirty-four patients had inv(3) (3.8%). In 241 patients with myelodysplastic syndromes the frequency was 6.2% as opposed to 2.9% in the remaining 649 patients (p=0.02). The increased frequency of inv(3) in patients with myelodysplastic syndromes indicates that inv(3) could be a risk factor for the development of the disease.