[Prevention of progression of allergic bronchial asthma as an occupational disease]. / Prävention eines Progresses des allergischen Asthma bronchiale als Berufskrankheit.

Since January 1st, 2021, termination of a harmful activity as a requirement for its recognition as an occupational disease (OD) has been abolished in Germany. This also includes the OD No. 4301 in accordance with Appendix 1 of the Occupational Diseases Ordinance (ODO) (obstructive respiratory diseases caused by allergenic substances). There has already been a significant increase in the number of diseases recognized under this OD. In this article, we present two patients with allergic (in the second case report mixed-form) bronchial asthma, in whom a medically easily preventable disease progression occurred as a result of continued occupational exposure to allergens as a farmer. According to § 3 (1) ODO, if there is a "risk that an occupational disease will develop, recur or worsen", the statutory accident insurance institutions are obliged to "counteract this danger with all suitable means". These individual preventive measures are compulsory by law and medically sensible, even under circumstances where only the risk of the genesis of an OD is given. Nonetheless, they are likely to be indicated and offered more frequently in the future due to an increased recognition of ODs. Unfavorable clinical courses such as those described in the two case reports presented here should therefore occur less frequently in the future.

Sources of Aerosol Dispersion During Singing and Potential Safety Procedures for Singers.

INTRODUCTION: With respect to the Covid-19 pandemic, singing is assumed to be associated with a high potential person-to-person transmission. However, it remains unclear how the impulse dispersion varies with different types of articulation, intensity levels of diction, or body position. Furthermore, it has not been understood in detail how to prevent aerosol dispersion during singing. MATERIAL AND METHODS: Single professional singers from the Bavarian Radio Chorus were asked to sing in different head positions, with different articulation patterns and different masks after inhaling the basic liquid of an e-cigarette. The vapor cloud was segmented and tracked over time. RESULTS: Consonants and exaggeration of diction enhanced the distance reached by the impulse dispersion. Furthermore, the greatest dispersion was reached for a neutral head position. All protection masks stopped the initial jet of the aerosols but the FFP2 masks were the most effective. CONCLUSION: Some protection equipment has been identified to be promising in reducing aerosol dispersion. However, systematic effects have to be evaluated in greater collectives.

Impulse dispersion of aerosols during playing the recorder and evaluation of safety measures.

INTRODUCTION: Group musical activities using wind instruments have been restricted during the CoVID19 pandemic due to suspected higher risk of virus transmission. It was presumed that the aerosols exhaled through the tubes while playing would be ejected over larger distances and spread into the room due to jet stream effects. In particular, the soprano recorder is widely used as an instrument in school classes, for beginners of all age groups in their musical education, in the context of leisure activities and in professional concert performances. Understanding the aerosol impulse dispersion characteristics of playing the soprano recorder could assist with the establishment of concepts for safe music-making. METHODS: Five adult professionally trained soprano recorder players (4 female, 1 male) played four bars of the main theme of L. van Beethoven's "Ode to Joy" in low and in high octaves, as well as with 3 different potential protection devices in the high octave. For comparison they spoke the corresponding text by F. Schiller. Before each task, they inhaled .5 L of vapor from an e-cigarette filled with base liquid. The vapor cloud escaping during speaking or playing was recorded by cameras and its spread was measured as a function of time in the three spatial dimensions. The potential safety devices were rated for practicability with a questionnaire, and their influence on the sound was compared, generating a long-term average spectrum from the audio data. RESULTS: When playing in the high octave, at the end of the task the clouds showed a median distance of 1.06 m to the front and .57 m diameter laterally (maxima: x: 1.35 m and y: .97 m). It was found that the clouds' expansion values in playing the recorder with and without safety measures are mostly lower when compared to the ordinary, raised speaking voice of the same subjects. The safety devices which covered the instrument did not show clear advantages and were rated as unpractical by the subjects. The most effective reduction of the cloud was reached when playing into a suction funnel. CONCLUSION: The aerosol dispersion characteristics of soprano recorders seem comparable to clarinets. The tested safety devices which covered holes of the instrument did not show clear benefits.

Impulse dispersion of aerosols during playing wind instruments.

Musical activities, especially singing and playing wind instruments, have been singled out as potentially high-risk activities for the transmission of SARS CoV-2, due to a higher rate of aerosol production and emission. Playing wind instruments can produce condensation, droplets of saliva, and aerosol particles, which hover and spread in the environmental air's convectional flows and which can be potentially infectious. The aim of this study is to investigate the primary impulse dispersion of aerosols that takes place during the playing of different wind instruments as compared to breathing and to speaking. Nine professional musicians (3 trumpeters, 3 flautists and 3 clarinetists) from the Bavarian Symphony Orchestra performed the main theme from the 4th movement of Ludwig van Beethoven's 9th symphony in different pitches and loudness. The inhaled air volume was marked with small aerosol particles produced using a commercial e-cigarette. The expelled aerosol cloud was recorded by cameras from different perspectives. Afterwards, the dimensions and dynamics of the aerosol cloud were measured by segmenting the video footage at every time point. Overall, the flutes produced the largest dispersion at the end of the task, reaching maximum forward distances of 1.88 m. An expulsion of aerosol was observed in different directions: upwards and downwards at the mouthpiece, at the end of the instrument, and along the flute at the key plane. In comparison, the maximum impulse dispersions generated by the trumpets and clarinets were lower in frontal and lateral direction (1.2 m and 1.0 m towards the front, respectively). Also, the expulsion to the sides was lower.

Effects of surgical masks on aerosol dispersion in professional singing.

BACKGROUND: In the CoVID-19 pandemic, singing came into focus as a high-risk activity for the infection with airborne viruses and was therefore forbidden by many governmental administrations. OBJECTIVE: The aim of this study is to investigate the effectiveness of surgical masks regarding the spatial and temporal dispersion of aerosol and droplets during professional singing. METHODS: Ten professional singers performed a passage of the Ludwig van Beethoven's "Ode of Joy" in two experimental setups-each with and without surgical masks. First, they sang with previously inhaled vapor of e-cigarettes. The emitted cloud was recorded by three cameras to measure its dispersion dynamics. Secondly, the naturally expelled larger droplets were illuminated by a laser light sheet and recorded by a high-speed camera. RESULTS: The exhaled vapor aerosols were decelerated and deflected by the mask and stayed in the singer's near-field around and above their heads. In contrast, without mask, the aerosols spread widely reaching distances up to 1.3 m. The larger droplets were reduced by up to 86% with a surgical mask worn. SIGNIFICANCE: The study shows that surgical masks display an effective tool to reduce the range of aerosol dispersion during singing. In combination with an appropriate aeration strategy for aerosol removal, choir singers could be positioned in a more compact assembly without contaminating neighboring singers all singers.

Aerosol Dispersion During Different Phonatory Tasks in Amateur Singers.

INTRODUCTION: Due to increased aerosol generation during singing, choir rehearsals were widely prohibited in the course of the CoVID-19 pandemic. Most studies on aerosol generation and dispersion focus on professional singers. However, it has not been clarified if these data are also representative for amateur singers. METHODS: Nine non-professional singers (four male, five female) were asked to perform five tasks; speaking (T+), singing a text softly (MT-) and loudly (MT+), singing on the vowel [ə] (M+) and singing with a N95 mask (MT+N95). Before performing the tasks, the singers were asked to inhale 0.5 L vapor produced by an e-cigarette consisting of the basic liquid. The spread of the exhaled vapor was recorded in all three dimensions by high-definition cameras and the impulse dispersion was detected as a function of time. RESULTS: Regarding the median dispersion to the front, all tasks showed comparable distances from 0.69 m to 0.82 m at the end of the tasks. However, the maximum aerosol dispersion showed a larger variety among different subjects or tasks, respectively. Especially in the M+ task a maximum distance of 1.96 m to the front was reached by a single subject. Although singing with a N95 mask resulted in a slightly increased median dispersion to the front, the maximum dispersion was decreased from 1.47 m (MT+) to 1.04 m (MT+N95). CONCLUSION: The maximum dispersion distance to the front of 1.96 m at the end of the M+ task and 1.47 m at the end of the MT+ task showed higher values in comparison to professional singers. Differences in phonation, articulation and mouth opening could lead to greater impulse dispersion. Singing in loud phonation with a N95 mask reduced the maximum impulse dispersion to the front to 1.04 m. Taking all results into consideration, a slightly larger safety distance should be necessary for non-professional singers.

The Effect of Singers' Masks on the Impulse Dispersion of Aerosols During Singing.

BACKGROUND: During the Covid-19 pandemic, singing activities were restricted due to several super-spreading events that have been observed during rehearsals and vocal performances. However, it has not been clarified how the aerosol dispersion, which has been assumed to be the leading transmission factor, could be reduced by masks which are specially designed for singers. MATERIAL AND METHODS: Twelve professional singers (10 of the Bavarian Radio-Chorus and two freelancers, seven females and five males) were asked to sing the melody of the ode of joy of Beethoven's 9th symphony "Freude schöner Götterfunken, Tochter aus Elisium" in D-major without masks and afterwards with five different singers' masks, all distinctive in their material and proportions. Every task was conducted after inhaling the basic liquid from an e-cigarette. The aerosol dispersion was recorded by three high-definition video cameras during and after the task. The cloud was segmented and the dispersion was analyzed for all three spatial dimensions. Further, the subjects were asked to rate the practicability of wearing the tested masks during singing activities using a questionnaire. RESULTS: Concerning the median distances of dispersion, all masks were able to decrease the impulse dispersion of the aerosols to the front. In contrast, the dispersion to the sides and to the top was increased. The evaluation revealed that most of the subjects would reject performing a concert with any of the masks. CONCLUSION: Although, the results exhibit that the tested masks could be able to reduce the radius of aerosol expulsion for virus-laden aerosol particles, there are more improvements necessary to enable the practical implementations for professional singing.

Combined molecular and clinical prognostic index for relapse and survival in cytogenetically normal acute myeloid leukemia.

PURPOSE: Cytogenetically normal (CN) acute myeloid leukemia (AML) is the largest and most heterogeneous cytogenetic AML subgroup. For the practicing clinician, it is difficult to summarize the prognostic information of the growing number of clinical and molecular markers. Our purpose was to develop a widely applicable prognostic model by combining well-established pretreatment patient and disease characteristics. PATIENTS AND METHODS: Two prognostic indices for CN-AML (PINA), one regarding overall survival (OS; PINAOS) and the other regarding relapse-free survival (RFS; PINARFS), were derived from data of 572 patients with CN-AML treated within the AML Cooperative Group 99 study (www.aml-score.org). RESULTS: On the basis of age (median, 60 years; range, 17 to 85 years), performance status, WBC count, and mutation status of NPM1, CEBPA, and FLT3-internal tandem duplication, patients were classified into the following three risk groups according to PINAOS and PINARFS: 29% of all patients and 32% of 381 responding patients had low-risk disease (5-year OS, 74%; 5-year RFS, 55%); 56% of all patients and 39% of responding patients had intermediate-risk disease (5-year OS, 28%; 5-year RFS, 27%), and 15% of all patients and 29% of responding patients had high-risk disease (5-year OS, 3%; 5-year RFS, 5%), respectively. PINAOS and PINARFS stratified outcome within European LeukemiaNet genetic groups. Both indices were confirmed on independent data from Cancer and Leukemia Group B/Alliance trials. CONCLUSION: We have developed and validated, to our knowledge, the first prognostic indices specifically designed for adult patients of all ages with CN-AML that combine well-established molecular and clinical variables and that are easily applicable in routine clinical care. The integration of both clinical and molecular markers could provide a basis for individualized patient care through risk-adapted therapy of CN-AML.

Inactivation of TP53 correlates with disease progression and low miR-34a expression in previously treated chronic lymphocytic leukemia patients.

In chronic lymphocytic leukemia (CLL) patients, disruptions of the TP53 tumor suppressor pathway by 17p13 deletion (del17p), somatic TP53 mutations, or downregulation of microRNA-34a have been associated with a poor prognosis. So far, the impact of the various TP53 defects has not been evaluated in a large cohort of previously treated and relapsed CLL patients. Here, we present the results of TP53 gene sequencing and fluorescence in situ hybridization for del17p in a phase 3 clinical trial (REACH [Rituximab in the Study of Relapsed Chronic Lymphocytic Leukemia]). Of the 457 patients, 52 had TP53 mutations and 37 had del17p. In 24 (46%) of the TP53 mutated patients, no del17p was found and in 9 of the del17p patients, no TP53 mutation was identified. Based on a predicted proportion of TP53 disruption, a complete disruption of TP53 function, either by a combination of point mutations and/or del17p, was associated with a high risk for disease progression. Progression-free survival of patients with a heterozygous TP53 mutation was not significantly different from patients with a completely intact TP53 locus. In addition, only a complete loss of TP53 function correlated with low microRNA-34a expression levels. This trial was registered at www.clinicaltrials.gov as #NCT00090051.

GATA2 zinc finger 1 mutations associated with biallelic CEBPA mutations define a unique genetic entity of acute myeloid leukemia.

Cytogenetically normal acute myeloid leukemia (CN-AML) with biallelic CEBPA gene mutations (biCEPBA) represents a distinct disease entity with a favorable clinical outcome. So far, it is not known whether other genetic alterations cooperate with biCEBPA mutations during leukemogenesis. To identify additional mutations, we performed whole exome sequencing of 5 biCEBPA patients and detected somatic GATA2 zinc finger 1 (ZF1) mutations in 2 of 5 cases. Both GATA2 and CEBPA are transcription factors crucial for hematopoietic development. Inherited or acquired mutations in both genes have been associated with leukemogenesis. Further mutational screening detected novel GATA2 ZF1 mutations in 13 of 33 biCEBPA-positive CN-AML patients (13/33, 39.4%). No GATA2 mutations were found in 38 CN-AML patients with a monoallelic CEBPA mutation and in 89 CN-AML patients with wild-type CEBPA status. The presence of additional GATA2 mutations (n=10) did not significantly influence the clinical outcome of 26 biCEBPA-positive patients. In reporter gene assays, all tested GATA2 ZF1 mutants showed reduced capacity to enhance CEBPA-mediated activation of transcription, suggesting that the GATA2 ZF1 mutations may collaborate with biCEPBA mutations to deregulate target genes during malignant transformation. We thus provide evidence for a genetically distinct subgroup of CN-AML. The German AML cooperative group trials 1999 and 2008 are registered with the identifiers NCT00266136 and NCT01382147 at www.clinicaltrials.gov.

The FLT3ITD mRNA level has a high prognostic impact in NPM1 mutated, but not in NPM1 unmutated, AML with a normal karyotype.

The impact of a FLT3-internal tandem duplication (FLT3ITD) on prognosis of patients with acute myeloid leukemia (AML) is dependent on the ratio of mutated to wild-type allele. In 648 normal karyotype (NK) AML patients, we found a significant independent effect of the quantitative FLT3ITD mRNA level--measured as (FLT3ITD/wtFLT3)/(FLT3ITD/wtFLT3+1)--on outcome. Moreover, this effect was clearly seen in 329 patients with a mutated NPM1 gene (NPM1+), but not in 319 patients without a NPM1 mutation (wtNPM1). In a multivariate Cox regression model, the quantitative FLT3ITD mRNA level showed an independent prognostic impact on overall survival (OS) and relapse-free survival (RFS) only in the NPM1+ subgroup (OS: hazard ratio, 5.9; [95% confidence interval [CI]: 3.1-11.2]; RFS: hazard ratio, 7.5 [95% CI: 3.4-16.5]). The FLT3ITD mRNA level contributes to relapse risk stratification and might help to guide postremission therapy in NPM1-mutated AML.

Monoallelic CEBPA mutations in normal karyotype acute myeloid leukemia: independent favorable prognostic factor within NPM1 mutated patients.

We and others have shown that cytogenetically normal (CN)-AML patients with biallelic CEBPA gene mutations (biCEBPA) represent a molecularly distinct group with a favorable prognosis. Patients carrying a monoallelic CEBPA mutation (moCEBPA), however, show no different outcome compared to patients with wildtype CEBPA, and these mutations are frequently associated with mutated NPM1 or FLT3-ITD. So far, no molecular or clinical hallmark has been identified to prognostically distinguish moCEBPA patients from patients with wildtype CEBPA. Therefore, we used the data of 663 CN-AML patients treated within the AMLCG 1999 trial to explore the prognostic value of moCEBPA in the context of concomitant clinical and molecular markers (mutated NPM1, FLT3-ITD). Multiple Cox regression in 515 patients adjusting for all available potential confounders revealed that the NPM1 mutation modified the prognostic value of moCEBPA with respect to overall survival (OS, p = 0.017) and event-free survival (EFS, p = 0.011). MoCEBPA was beneficial in NPM1 mutated patients: adjusted OS-hazard ratio (HR) 0.09, 95% confidence interval (CI) 0.01-0.63, p = 0.016; EFS-HR (95% CI) 0.16 (0.04-0.65), p = 0.010. In contrast, moCEBPA had no prognostic impact in patients with wildtype NPM1: OS-HR (95% CI) 1.08 (0.59-1.97), p = 0.804; EFS-HR (95% CI) 1.12 (0.64-1.96), p = 0.682. We found no prognostic effect modification for moCEBPA by FLT3-ITD. The presence of a moCEBPA mutation was shown to be associated with prolonged survival in NPM1 mutated CN-AML patients. Confirmation of these results in larger studies will clarify whether an additional moCEBPA mutation influences the risk stratification of patients with an NPM1 mutated/FLT3-ITD positive genotype.

Age-dependent frequencies of NPM1 mutations and FLT3-ITD in patients with normal karyotype AML (NK-AML).

Prognosis of AML in elderly patients is poor due to adverse patient characteristics and comorbidities. In addition, disease-associated parameters reveal differences between older and younger patients with AML. Survival in normal karyotype AML (NK-AML) is influenced by different clinical and molecular markers. The aim of this work was to investigate the frequencies of molecular markers in patients with NK-AML with a focus on NPM1 mutations and FLT3-ITD in different age groups. In the present study, we analyzed the frequencies of mutations of NPM1 and FLT3-ITD in a cohort of 1,321 adult patients and 148 children with AML treated within the AMLCG99, the AML98, and AML04 trials and their distribution in different age groups. Additionally, the frequencies of mutations in CEBPA genes, FLT3-TKD, and MLL-PTD were analyzed in the cohort with NK-AML (n = 729). Our data show that the presence of mutations of NPM1 (from 60% to 40%) and FLT3-ITD (from 50% to 20%) significantly decreased with age in adult AML. Consequently, the proportion of NPM1-/FLT3-ITD- patients increased with age. The decreasing frequency of NPM1 mutations in elderly patients was paralleled by a reduced complete remission (CR) rate in the elderly of 55% compared to 80% in the younger patients. By contrast, the frequencies of other gene mutations, like FLT3-TKD and MLL-PTD, and mutations in CEBPA were not age-dependent. The decreasing frequency of the favorable NPM1 mutations with increasing age may partially explain the worse outcome in the elderly patients. Furthermore, the increasing amount of elderly patients without NPM1 mutations or FLT3-ITD suggests that other molecular and clinical risk factors may influence prognosis in this age group.

Acute myeloid leukemia with biallelic CEBPA gene mutations and normal karyotype represents a distinct genetic entity associated with a favorable clinical outcome.

PURPOSE: CEBPA mutations are found as either biallelic (biCEBPA) or monoallelic (moCEBPA). We set out to explore whether the kind of CEBPA mutation is of prognostic relevance in cytogenetically normal (CN) acute myeloid leukemia (AML). PATIENTS AND METHODS: Four hundred sixty-seven homogeneously treated patients with CN-AML were subdivided into moCEBPA, biCEBPA, and wild-type (wt) CEBPA patients. The subgroups were analyzed for clinical parameters and for additional mutations in the NPM1, FLT3, and MLL genes. Furthermore, we obtained gene expression profiles using oligonucleotide microarrays. RESULTS: Only patients with biCEBPA had an improved median overall survival when compared with patients with wtCEBPA (not reached v 20.4 months, respectively; P = .018), whereas patients with moCEBPA (20.9 months) and wtCEBPA had a similar outcome (P = .506). Multivariable analysis confirmed biCEBPA, but not moCEBPA, mutations as an independent favorable prognostic factor. Interestingly, biCEBPA mutations, compared with wtCEBPA, were never associated with mutated NPM1 (0% v 43%, respectively; P < .001) and rarely associated with FLT3 internal tandem duplication (ITD; 5% v 23%, respectively; P = .059), whereas patients with moCEBPA had a similar frequency of mutated NPM1 and a significantly higher association with FLT3-ITD compared with patients with wtCEBPA (44% v 23%, respectively; P = .037). Furthermore, patients with biCEBPA showed a homogeneous gene expression profile that was characterized by downregulation of HOX genes, whereas patients with moCEBPA showed greater heterogeneity in their gene expression profiles. CONCLUSION: Biallelic disruption of the N and C terminus of CEBPA is required for the favorable clinical outcome of CEBPA-mutated patients and represents a distinct molecular subtype of CN-AML with a different frequency of associated gene mutations. These findings are of great significance for risk-adapted therapeutic strategies in AML.

ERG expression is an independent prognostic factor and allows refined risk stratification in cytogenetically normal acute myeloid leukemia: a comprehensive analysis of ERG, MN1, and BAALC transcript levels using oligonucleotide microarrays.

PURPOSE: Recently, several novel molecular prognostic markers were identified in cytogenetically normal acute myeloid leukemia (CN-AML). In addition to the well-known influence of FLT3, NPM1, and CEBPA mutations, high transcript levels of the ERG, BAALC, and MN1 genes have been associated with inferior outcomes, but the relative importance of these risk markers remains to be defined. PATIENTS AND METHODS: We analyzed ERG, BAALC, and MN1 expression levels in a cohort of 210 patients with CN-AML who received intensive chemotherapy. Expression levels of ERG, BAALC, and MN1 were determined in bone marrow samples by using oligonucleotide microarrays. RESULTS: High transcript levels of ERG, BAALC, and MN1 were predictors for inferior overall survival (OS) and a lower rate of complete remissions (CRs). There were significant positive correlations between the expression levels of all three genes. ERG expression levels predicted OS in elderly patients (ie, age 60 years or older) with CN-AML (P = .006) as well as in younger patients (P = .013). In multivariate analyses, high ERG expression was independently associated with a lower CR rate (P = .013), shorter event-free survival (P = .008), and shorter OS (P = .005). Patients who had low ERG levels and absent FLT3 internal tandem duplication (ITD) had a 5-year OS of 44%, and patients who had high ERG expression and FLT3 ITD had a 5-year OS of only 5%. CONCLUSION: We analyzed a comprehensive set of molecular risk factors in a large, homogeneous CN-AML patient cohort. In this study, high ERG expression levels emerged as a strong negative prognostic factor and provided prognostic information in addition to established molecular markers.

NPM1 but not FLT3-ITD mutations predict early blast cell clearance and CR rate in patients with normal karyotype AML (NK-AML) or high-risk myelodysplastic syndrome (MDS).

Mutations in the NPM1 gene represent the most frequent genetic alterations in patients with acute myeloid leukemia (AML) and are associated with a favorable outcome. In 690 normal karyotype (NK) AML patients the complete remission rates (CRs) and the percentage of patients with adequate in vivo blast cell reduction 1 week after the end of the first induction cycle were significantly higher in NPM1(+) (75% and 80%, respectively) than in NPM1(-) (57% and 57%, respectively) patients, but were unaffected by the FLT3-ITD status. Multivariate analyses revealed the presence of a NPM1 mutation as an independent positive prognostic factor for the achievement of an adequate day-16 blast clearance and a CR. In conclusion, NPM1(+) blast cells show a high in vivo sensitivity toward induction chemotherapy irrespective of the FLT3-ITD mutation status. These findings provide insight into the pathophysiology and help to understand the favorable clinical outcome of patients with NPM1(+) AML.

CBL exon 8/9 mutants activate the FLT3 pathway and cluster in core binding factor/11q deletion acute myeloid leukemia/myelodysplastic syndrome subtypes.

PURPOSE: CBL is a negative regulator of activated receptor tyrosine kinases (RTK). In this study, we determined the frequency of CBL mutations in acute leukemias and evaluated the oncogenic potential of mutant CBL. EXPERIMENTAL DESIGN: The cDNA of 300 acute myeloid leukemia (AML)/myelodysplastic syndrome (MDS) and acute lymphoblastic leukemia (ALL) patients and 82 human leukemic cell lines was screened for aberrations in the linker and RING finger domain of CBL. The oncogenic potential of identified mutants was evaluated in hematopoietic cells. RESULTS: We identified 3 of 279 AML/MDS patients expressing CBL exon 8/9 deletion mutants. Three of four cases at diagnosis expressed deleted transcripts missing exon 8 or exon 8/9. In remission samples a weak or no expression of mutant CBL was detected. No aberrations were found in normal hematopoietic tissues. One of 116 sequenced AML/MDS cases carried a R420G missense mutation. All AML/MDS patients with identified CBL mutants belonged to the core binding factor and 11q deletion AML subtypes. Functionally, CBL negatively regulated FMS-like tyrosine kinase 3 (FLT3) activity and interacted with human FLT3 via the autophosphorylation sites Y589 and Y599 and colocalized in vivo. Expression of CBLDeltaexon8 and CBLDeltaexon8+9 in FLT3-WT-Ba/F3 cells induced growth factor-independent proliferation associated with autophosphorylation of FLT3 and activated the downstream targets signal transducer and activator of transcription 5 (STAT5) and protein kinase B (AKT). FLT3 ligand-dependent hyperproliferation of CBL mutant cells could be abrogated by treatment with the FLT3 PTK inhibitor PKC412 (midostaurin). CONCLUSION: CBL exon8/9 mutants occur in genetically defined AML/MDS subtypes and transform hematopoietic cells by constitutively activating the FLT3 pathway. This phenotype resembles the one of mutated RTKs and suggests that CBL mutant AML patients might benefit from treatment with FLT3 PTK inhibitors.

Rapid and sensitive screening for CEBPA mutations in acute myeloid leukaemia.

The presence of CCAAT/enhancer binding protein alpha (CEBPA) gene mutations in patients with cytogenetically normal acute myeloid leukaemia (CN-AML) confers a favourable prognosis. Routine screening of all CN-AML patients for CEBPA mutations is therefore important for individual risk-adapted post-remission therapy and requires a fast and easy screening method. CEBPA mutations are distributed over the entire CEBPA gene and the functional and clinical consequences of the different mutations are still largely unknown. Therefore, we developed a multiplex polymerase chain reaction-based fragment length analysis mutation screening method for the entire CEBPA coding region. We initially evaluated our method by analysing 120 CN-AML samples both by fragment analysis and nucleotide sequencing and reached a sensitivity of 100% and a specificity of 90%. 349 CN-AML samples were subsequently screened for CEBPA mutations by fragment length analysis. Among a total of 469 CN-AML patient samples, 58 CEBPA mutations were detected in 38 CN-AML patients (8.1%). In conclusion, we established a fast and sensitive CEBPA mutation screening method suitable for inclusion in routine AML diagnostics.