Gudrun Göhring

Early molecular and cytogenetic response is predictive for long-term progression-free and overall survival in chronic myeloid leukemia (CML)

In the face of competing first-line treatment options for CML, early prediction of prognosis on imatinib is desirable to assure favorable survival or otherwise consider the use of a second-generation tyrosine kinase inhibitor (TKI). A total of 1303 newly diagnosed imatinib-treated patients (pts) were investigated to correlate molecular and cytogenetic response at 3 and 6 months with progression-free and overall survival (PFS, OS). The persistence of BCR-ABL transcript levels >10% according to the international scale (BCR-ABLIS) at 3 months separated a high-risk group (28% of pts; 5-year OS: 87%) from a group with >1–10% BCR-ABLIS (41% of pts; 5-year OS: 94%; P=0.012) and from a group with ⩽1% BCR-ABLIS (31% of pts; 5-year OS: 97%; P=0.004). Cytogenetics identified high-risk pts by >35% Philadelphia chromosome-positive metaphases (Ph+, 27% of pts; 5-year OS: 87%) compared with ⩽35% Ph+ (73% of pts; 5-year OS: 95%; P=0.036). At 6 months, >1% BCR-ABLIS (37% of pts; 5-year OS: 89%) was associated with inferior survival compared with ⩽1% (63% of pts; 5-year OS: 97%; P<0.001) and correspondingly >0% Ph+ (34% of pts; 5-year OS: 91%) compared with 0% Ph+ (66% of pts; 5-year OS: 97%; P=0.015). Treatment optimization is recommended for pts missing these landmarks.

Impact of additional cytogenetic aberrations at diagnosis on prognosis of CML: long-term observation of 1151 patients from the randomized CML Study IV

The prognostic relevance of additional cytogenetic findings at diagnosis of chronic myeloid leukemia (CML) is unclear. The impact of additional cytogenetic findings at diagnosis on time to complete cytogenetic (CCR) and major molecular remission (MMR) and progression-free (PFS) and overall survival (OS) was analyzed using data from 1151 Philadelphia chromosome-positive (Ph(+)) CML patients randomized to the German CML Study IV. At diagnosis, 1003 of 1151 patients (87%) had standard t(9;22)(q34;q11) only, 69 patients (6.0%) had variant t(v;22), and 79 (6.9%) additional cytogenetic aberrations (ACAs). Of these, 38 patients (3.3%) lacked the Y chromosome (-Y) and 41 patients (3.6%) had ACAs except -Y; 16 of these (1.4%) were major route (second Philadelphia [Ph] chromosome, trisomy 8, isochromosome 17q, or trisomy 19) and 25 minor route (all other) ACAs. After a median observation time of 5.3 years for patients with t(9;22), t(v;22), -Y, minor- and major-route ACAs, the 5-year PFS was 90%, 81%, 88%, 96%, and 50%, and the 5-year OS was 92%, 87%, 91%, 96%, and 53%, respectively. In patients with major-route ACAs, the times to CCR and MMR were longer and PFS and OS were shorter (P < .001) than in patients with standard t(9;22). We conclude that major-route ACAs at diagnosis are associated with a negative impact on survival and signify progression to the accelerated phase and blast crisis.

Marrow fibrosis predicts early fatal marrow failure in patients with myelodysplastic syndromes

Marrow fibrosis (MF) has rarely been studied in myelodysplastic syndromes (MDS). There are no data on occurrence and significance of MF in the context of the World Health Organization (WHO) classification of disease. In total, 349 bone marrow biopsies from 200 patients with primary MDS were examined for MF and its prognostic relevance. MF correlated with multilineage dysplasia, more severe thrombopenia, higher probability of a clonal karyotype abnormality, and higher percentages of blasts in the peripheral blood (P<0.002). Its frequency varied markedly between different MDS types ranging from 0 (RARS) to 16% (RCMD, RAEB, P<0.007). Two patients with MF showed a Janus kinase-2 mutation (V617F). Patients with MF suffered from marrow failure significantly earlier with shortening of the survival time down to 0.5 (RAEB-1/-2), and 1–2 (RCMD, RA) years in median (P<0.00005). The prognostic relevance of MF was independent of the International Prognostic Scoring System and the classification of disease. Conclusion: The risk of MF Differs markedly between various subtypes of MDS. MF indicates an aggressive course with a significantly faster progression to fatal marrow failure and should therefore be considered in diagnosis, prognosis and treatment of disease.

Prognostic effect of calreticulin mutations in patients with myelofibrosis after allogeneic hematopoietic stem cell transplantation

Prognostic effect of calreticulin mutations in patients with myelofibrosis after allogeneic hematopoietic stem cell transplantation

A novel pedigree with heterozygous germline RUNX1 mutation causing familial MDS-related AML: can these families serve as a multistep model for leukemic transformation?

A novel pedigree with heterozygous germline RUNX1 mutation causing familial MDS-related AML: can these families serve as a multistep model for leukemic transformation?

Epigenetic inactivation of tumour suppressor gene KLF11 in myelodysplastic syndromes.

The identification of aberrantly hypermethylated genes may lead to the development of new diagnostic markers and the identification of novel targets of epigenetic therapy in myelodysplastic syndromes (MDS). We therefore investigated the methylation status of transcription factor genes KLF5, KLF11, and MAFB, shown to be aberrantly methylated in myelogeneous leukaemia cells, in a series of 115 MDS patient as well as in 25 control subjects. Using quantitative high‐resolution pyrosequencing methodology, KLF11, MAFB, and KLF5 were shown for the first time to be hypermethylated in 17 (15%), 8 (7%), and 2 (1.7%) cases, respectively, but not in any of the patients with an isolated 5q‐deletion. Patient samples harbouring KLF11 methylation displayed reduced KLF11 mRNA expression and KLF11 hypermethylation correlated with a high International Prognostic Scoring System score (P < 0.05). In conclusion, epigenetic inactivation and subsequent transcriptional repression of the KLF11 gene is quite frequent in MDS. Patients with an isolated 5q‐deletion seem to harbour a distinct epigenetic profile.

Prognostic implications and molecular associations of NADH dehydrogenase subunit 4 ( ND4 ) mutations in acute myeloid leukemia

To study the prevalence and prognostic importance of mutations in NADH dehydrogenase subunit 4 (ND4), a mitochondrial encoded transmembrane component of the electron transport chain respiratory Complex I, 452 AML patients were examined for ND4 mutations by direct sequencing. The prognostic impact of ND4 mutations was evaluated in the context of other clinical prognostic markers and genetic risk factors. In all, 29 of 452 patients (6.4%) had either somatic (n=12) or germline (n=17) ND4 mutations predicted to affect translation. Somatic mutations were more likely to be heteroplasmic (P<0.001), to occur in predicted transmembrane domains (P<0.001) and were predicted to have damaging effects upon translation (P<0.001). Patients with somatically acquired ND4 mutations had significantly longer relapse-free survival (P=0.017) and overall survival (OS) (P=0.021) than ND4wildtype patients. Multivariate analysis also demonstrated a tendency for increased survival in patients with somatic ND4 mutations (RFS: hazard ratio (HR) 0.25, confidence interval (CI) 0.06–1.01, P=0.052; OS: HR 0.29, CI 0.74–1.20, P=0.089). Somatic ND4mutated patients had a higher prevalence of concomitant DNMT3A mutations (P=0.023) and a higher percentage of the NPM1/FLT3-ITD low-risk genotype (P=0.021). Germline affected cases showed higher BAALC (P=0.036) and MLL5 (P=0.051) expression levels. Further studies are warranted to validate the favorable prognostic influence of acquired ND4 mutations in AML.

Acute myeloid leukemia derived from lympho-myeloid clonal hematopoiesis.

We studied acute myeloid leukemia (AML) patients with lympho-myeloid clonal hematopoiesis (LM-CH), defined by the presence of DNA methyltransferase 3A (DNMT3A) mutations in both the myeloid and lymphoid T-cell compartment. Diagnostic, complete remission (CR) and relapse samples were sequenced for 34 leukemia-related genes in 171 DNMT3A mutated adult AML patients. AML with LM-CH was found in 40 patients (23%) and was associated with clonal hematopoiesis of indeterminate potential years before AML, older age, secondary AML and more frequent MDS-type co-mutations (TET2, RUNX1 and EZH2). In 82% of AML patients with LM-CH, the preleukemic clone was refractory to chemotherapy and was the founding clone for relapse. Both LM-CH and non-LM-CH MRD-positive AML patients who achieved CR had a high risk of relapse after 10 years (75% and 75%, respectively) compared with patients without clonal hematopoiesis in CR with negative MRD (27% relapse rate). Long-term survival of patients with LM-CH was only seen after allogeneic hematopoietic stem cell transplantation (HSCT). We define AML patients with LM-CH as a distinct high-risk group of AML patients that can be identified at diagnosis through mutation analysis in T cells and should be considered for HSCT.

Long-term transfusion independence in del(5q) MDS patients who discontinue lenalidomide

mice (CD45.1þCD45.2þ ) were transplanted into lethally irradiated C57BL/6 (CD45.2) recipients together with competitor cells (CD45.2). Short-term (2 months) and long-term (4 months) competitive repopulating capability was studied by monitoring peripheral blood donor chimerism of recipient mice by flow cytometry. Mice transplanted with Npm1þ / HSC exhibited markedly lower engraftment than the recipients of Npm1þ /þ HSC as measured by total chimerism (threefold; Figure 2a, Po0.03). The percentage total and myeloid chimerism in individual transplanted mice did not differ significantly at 2 and 4 months (data not shown). We also quantified donor chimerism in the bone marrow of the recipient mice 4 months after transplantation (Figure 2b). Similar to the results in the peripheral blood, the competitive repopulating ability of the HSC from Npm1þ / mice was significantly reduced (8.5-fold; Figure 2b, Po0.02) in the bone marrow of the recipient mice. Although the engraftment potential of Npm1þ / HSC was reduced, we did not observe any effects on the composition of donor-derived mature hematopoietic cells in the peripheral blood of recipient mice (Supplementary Figure 5). Thus, the level of NPM1 expression has a direct effect on HSC repopulating ability in vivo, but has no effect on hematopoietic fate commitment. In summary, our Npm1þ / mouse model indicates that NPM1 has a role in maintaining HSC number and in preserving the functional integrity of these cells in the context of competitive transplantation. However, it does not appear to have a role in regulating HSC differentiation. Npm1 haploinsufficiency did not result in significant alterations in mature blood cell numbers or significant dysplasia in our mice. These results are at some variance with a previous report, in which Npm1 heterozygosity was associated with an MDS-like phenotype in older (6–10 months) mice. We cannot completely exclude the possibility that the different techniques used to generate these mutant mice may explain the differences in our results; however, both models are associated with homozygous null embryonic lethality as well as decreased Npm1 mRNA (data not shown) and protein levels in Npm1þ / mice. Our study did use mice in which the targeted Npm1 allele was functionally eliminated by a single-gene trap event in the NPM1 locus between exons 7 and 8 (genetic background 129/SvEvBrd C57BL/6J), whereas in the previous study exons 2–7 were replaced by green fluorescent protein cassette introducing the stop codon after exon 2 (genetic background 129/Sv C57BL/6). Minor genetic variability between the two different substrains used in these studies (129/Sv vs 129/SvEvBrd) might also influence their respective phenotypes. Although we have no definitive explanation for the difference in phenotypes, only our study directly assessed HSC function. We speculate that the increased number of HSC in Npmþ / mice could be an important early step in myeloid pathogenesis, as these HSC could provide a milieu in which additional genetic events might occur, thereby increasing the susceptibility to subsequent development of MDS or myeloid leukemia. Indeed, prior studies have demonstrated that transgenic Em-myc overexpression in the Npm1þ / mutant setting can facilitate AML development. However, these data also indicate that loss of NPM1 function alone is not sufficient to result in increased development of hematological malignancies, which suggests that other cooperating lesions are likely necessary for disease pathogenesis. Further studies focused on understanding the interactions between Npm1 and other genes associated with MDS or AML progression will help to identify these cooperating events.

Telomere shortening, clonal evolution and disease progression in myelodysplastic syndrome patients with 5q deletion treated with lenalidomide

Telomere shortening, clonal evolution and disease progression in myelodysplastic syndrome patients with 5q deletion treated with lenalidomide