IGH::NSD2 Fusion Gene Transcript as Measurable Residual Disease Marker in Multiple Myeloma.

Multiple myeloma (MM) is the second most common hematological malignancy. Approximately 15% of MM patients are affected by the t(4;14) translocation resulting in the IGH::NSD2 fusion transcript. Breakage occurs in three major breakpoint regions within the NSD2 gene (MB4-1, MB4-2, and MB4-3), where MB4-1 leads to the production of full-length protein, while truncated proteins are expressed in the other two cases. Measurable residual disease (MRD) has been conclusively established as a crucial prognostic factor in MM. The IGH::NSD2 fusion transcript can serve as a sensitive MRD marker. Using bone marrow (BM) and peripheral blood (PB) samples from 111 patients, we developed a highly sensitive quantitative real-time PCR (qPCR) and digital PCR (dPCR) system capable of detecting fusion mRNAs with a sensitivity of up to 1:100,000. PB samples exhibited sensitivity three orders of magnitude lower compared to BM samples. Patients with an MB4-2 breakpoint demonstrated significantly reduced overall survival (p = 0.003). Our novel method offers a simple and sensitive means for detecting MRD in a substantial proportion of MM patients. Monitoring may be carried out even from PB samples. The literature lacks consensus regarding survival outcomes among patients with different NSD2 breakpoints. Our data align with previous findings indicating that patients with the MB4-2 breakpoint type tend to exhibit unfavorable overall survival.

Low-burden TP53 mutations represent frequent genetic events in CLL with an increased risk for treatment initiation.

TP53 aberrations predict chemoresistance and represent a contraindication for the use of standard chemoimmunotherapy in chronic lymphocytic leukaemia (CLL). Recent next-generation sequencing (NGS)-based studies have identified frequent low-burden TP53 mutations with variant allele frequencies below 10%, but the clinical impact of these low-burden TP53 mutations is still a matter of debate. In this study, we aimed to scrutinise the subclonal architecture and clinical impact of TP53 mutations using a sensitive, NGS-based mutation analysis in a 'real-world' cohort of 901 patients with CLL. In total, 225 TP53 mutations were identified in 17.5% (158/901) of the patients; 48% of these alterations represented high-burden mutations, while 52% were low-burden TP53 mutations. Low-burden mutations as sole alterations were identified in 39% (62/158) of all mutated cases with 82% (51/62) of these being represented by a single low-burden TP53 mutation. Patients harbouring low-burden TP53 mutations had significantly lower time to first treatment compared to patients with wild-type TP53. Our study has expanded the knowledge on the frequency, clonal architecture, and clinical impact of low-burden TP53 mutations. By demonstrating that patients with sole low-burden TP53 variants represent more than one-third of patients with TP53 mutations and have an increased risk for treatment initiation, our findings strengthen the need to redefine the threshold of TP53 variant reporting to below 10% in the routine diagnostic setting.

Liquid biopsy-based monitoring of residual disease in multiple myeloma by analysis of the rearranged immunoglobulin genes-A feasibility study.

The need for sensitive monitoring of minimal/measurable residual disease (MRD) in multiple myeloma emerged as novel therapies led to deeper responses. Moreover, the potential benefits of blood-based analyses, the so-called liquid biopsy is prompting more and more studies to assess its feasibility. Considering these recent demands, we aimed to optimize a highly sensitive molecular system based on the rearranged immunoglobulin (Ig) genes to monitor MRD from peripheral blood. We analyzed a small group of myeloma patients with the high-risk t(4;14) translocation, using next-generation sequencing of Ig genes and droplet digital PCR of patient-specific Ig heavy chain (IgH) sequences. Moreover, well established monitoring methods such as multiparametric flow cytometry and RT-qPCR of the fusion transcript IgH::MMSET (IgH and multiple myeloma SET domain-containing protein) were utilized to evaluate the feasibility of these novel molecular tools. Serum measurements of M-protein and free light chains together with the clinical assessment by the treating physician served as routine clinical data. We found significant correlation between our molecular data and clinical parameters, using Spearman correlations. While the comparisons of the Ig-based methods and the other monitoring methods (flow cytometry, qPCR) were not statistically evaluable, we found common trends in their target detection. Regarding longitudinal disease monitoring, the applied methods yielded complementary information thus increasing the reliability of MRD evaluation. We also detected indications of early relapse before clinical signs, although this implication needs further verification in a larger patient cohort.

Non-malignant, non-infectious lymphoproliferation: challenges in the diagnosis and treatment of autoimmune lymphoproliferative syndrome / Nem malignus, nem infectiosus lymphoproliferatio: kihívások az autoimmun lymphoproliferativ szindróma diagnosztikájában és kezelésében.

Összefoglaló. Az autoimmun lymphoproliferativ szindróma egy ritka, immundeficientiával járó genetikai betegség. Hátterében az extrinszik apoptotikus útvonal génjeinek örökletes vagy szerzett mutációi és a következményesen kialakuló, aktivált lymphocyták negatív szelekciójának a defektusa áll. Az autoimmun lymphoproliferativ szindróma klinikai megjelenésére jellemzo a jóindulatú lymphocytaburjánzás következtében kialakuló lymphadenopathia és lépmegnagyobbodás. Gyakran társul olyan autoimmun kórképekkel, mint az autoimmun haemolyticus anaemia vagy az autoimmun thrombocytopenia. A betegségben jellemzo laboratóriumi eltérések a következok: az αß+ CD4-/CD8- kettos negatív T-sejtek szaporulata, a szolúbilis Fas-ligand, az interleukin-10 és interleukin-18, valamint a B12-vitamin szérumszintjének emelkedése. A kórkép diagnózisához hozzátartozik az in vitro Fas-mediált apoptózis funkciójának vizsgálata, valamint a genetikai vizsgálat. Differenciáldiagnosztikai szempontból fontos elkülöníteni a lymphomáktól, valamint az autoimmun lymphoproliferativ szindrómaszeru betegségektol. A kezelés alapja a társuló autoimmun kórképek tüneteinek csökkentése immunszuppresszív terápiával. Orv Hetil. 2022; 163(4): 123-131. Summary. The autoimmune lymphoproliferative syndrome is a rare genetic disorder causing immunodeficiency. In the background of the disease, germline or somatic mutations of genes participating in the extrinsic apoptotic pathway and the consequential defect in the negative selection of activated lymphocytes were discovered. The clinical appearance of autoimmune lymphoproliferative syndrome consists of non-malignant lymphoproliferation, lymphadenopathy and splenomegaly, it is frequently accompanied by autoimmune disorders such as autoimmune haemolytic anaemia or autoimmune thrombocytopenia. The main diagnostic laboratory findings of this disease are the following: an elevation in αß+, CD4-/CD8- double-negative T cell count, elevated serum levels of soluble Fas-ligand, interleukin-10, interleukin-18 and vitamin B12. Other useful laboratory tests are the in vitro Fas-mediated apoptotic functional assay and the genetic screening for gene mutations. Differential diagnosis should exclude malignant lymphoproliferation in lymphomas and non-malignant autoimmune lymphoprolipherative syndrome-like diseases. The main aim of the treatment is the amelioration of the accompanying autoimmune disease with immunosuppressive therapy. Orv Hetil. 2022; 163(4): 123-131.

Nucleophosmin1 and isocitrate dehydrogenase 1 and 2 as measurable residual disease markers in acute myeloid leukemia.

Monitoring measurable residual disease (MRD) in acute myeloid leukemia (AML) plays an important role in predicting relapse and outcome. The applicability of the leukemia-initiating nucleophosmin1 (NPM1) gene mutations in MRD detection is well-established, while that of isocitrate dehydrogenase1/2 (IDH1/2) mutations are matter of debate. The aim of this study was to investigate the stability of NPM1 and IDH1/2 mutations at diagnosis and relapse retrospectively in 916 adult AML patients. The prognostic value of MRD was evaluated by droplet digital PCR on the DNA level in a selected subgroup of patients in remission. NPM1 re-emerged at relapse in 91% (72/79), while IDH1/2 in 87% (20/23) of mutation-positive cases at diagnosis. NPM1 mutation did not develop at relapse, on the contrary novel IDH1/2 mutations occurred in 3% (3/93) of previously mutation-negative cases. NPM1 MRD-positivity after induction (n = 116) proved to be an independent, adverse risk factor (MRDpos 24-month OS: 39.3±6.2% versus MRDneg: 58.5±7.5%, p = 0.029; HR: 2.16; 95%CI: 1.25-3.74, p = 0.006). In the favorable subgroup of mutated NPM1 without fms-like tyrosine kinase 3 internal tandem duplication (FLT3-ITD) or with low allelic ratio, NPM1 MRD provides a valuable prognostic biomarker (NPM1 MRDpos versus MRDneg 24-month OS: 42.9±6.7% versus 66.7±8.6%; p = 0.01). IDH1/2 MRD-positivity after induction (n = 62) was also associated with poor survival (MRDpos 24-month OS: 41.3±9.2% versus MRDneg: 62.5±9.0%, p = 0.003; HR 2.81 95%CI 1.09-7.23, p = 0.032). While NPM1 variant allele frequency decreased below 2.5% in remission in all patients, IDH1/2 mutations (typically IDH2 R140Q) persisted in 24% of cases. Our results support that NPM1 MRD even at DNA level is a reliable prognostic factor, while IDH1/2 mutations may represent pre-leukemic, founder or subclonal drivers.

Optimising the mutation screening strategy in Marfan syndrome and identifying genotypes with more severe aortic involvement.

BACKGROUND: Marfan syndrome (MFS) is a systemic connective tissue disorder with life-threatening manifestations affecting the ascending aorta. MFS is caused by dominant negative (DN) and haploinsufficient (HI) mutations of the FBN1 gene. Our aim was to identify mutations of MFS patients with high detection rate and to investigate the use of a gene panel for patients with Marfanoid habitus. We also aimed to examine correlations between genotype and cardiovascular manifestations to predict "malignant" mutations. METHODS: 136 individuals were enrolled. In the first phase, next-generation sequencing (NGS) and Sanger sequencing were performed for 57 patients to screen the FBN1 gene, followed by multiplex ligation-dependent probe amplification (MLPA) in negative cases. For repeated negative results, NGS gene panel involving 9 genes was used. In the second phase, 79 patients were tested primarily with the same gene panel, negative samples were tested by MLPA. RESULTS: 84 pathogenic mutations were detected, out of which 78 affected FBN1, 6 non-FBN1 mutations (2 TGFB2, 1 TGFBR2, 2 TGFBR1, 1 SMAD3) are associated with Loeys-Dietz syndrome (LDS). LDS patients had lower systemic score and they were younger, but their aortic involvement did not differ. MLPA detected 4 multi-exon deletions of FBN1 gene, which could not be identified by our first-step screening method. Aortic involvement (aortic dissection and/or dilation) did not differ significantly among HI and DN mutations (p = 0.061). Combined group of HI and DN mutations eliminating a disulphide-bonding cysteine (DN Cys) had significantly higher aortic involvement rate than DN mutations not eliminating a disulphide-bonding cysteine (DN non-Cys) (p < 0.001). Patients with DN Cys required significantly more aortic surgeries than HI and DN non-Cys mutations (p = 0.042 and p = 0.015, respectively). CONCLUSIONS: Due to the relevant number of mutations affecting genes other than FBN1, preferred approach for testing individuals with Marfanoid habitus is using a gene panel rather than single-gene analysis, followed by MLPA for negative samples. DN Cys and HI mutations should be considered as risk factors for aortic involvement. Genetic testing for patients with Marfanoid features and a systemic score under 7 is recommended, as LDS patients may have lower scores, but they may have severe cardiovascular manifestations.

Investigation of TGFB1 -1347C>T variant as a biomarker after allogeneic hematopoietic stem cell transplantation.

Allogeneic hematopoietic stem cell transplantation (allo-HSCT) is a potentially curative therapeutic option for malignant hematopoietic diseases. Cytokines including transforming growth factor ß1 (TGFß1) play a pivotal role in immune reconstruction, and the development of graft versus host disease (GvHD) or infections. The aim of this study was to investigate the role of TGFB1 gene -1347C>T variant in the outcome of HSCT in a cohort of 409 adult recipient-donor pairs. TGFB1 variant was analysed from genomic DNA with LightCycler hybridisation probe method. In case of myeloablative conditioning, donor TGFB1 genotype correlated with overall survival (60-month OS for CC: 62.1 ± 4.8%; CT: 46.8 ± 4.8%; TT: 35.6 ± 9.3%; p = 0.032), which was independent of age, donor type and GvHD prophylaxis in multivariate analysis (HR:2.35, 95%CI:1.35-4.10, p = 0.003). The cumulative incidence of acute GvHD grade III-IV [CC:10%; CT:17%; TT:24%], and non-relapse mortality was higher in TT-carriers (24-month NRM: CC:24%; CT:26%; TT:46%, p = 0.035). We did not find any association between recipient TGFB1 -1347C>T polymorphism and HSCT outcome. Our results suggest that donor TGFB1 -1347C>T may exert an adverse influence on the outcome of myeloablative conditioning transplantation.

[Importance of next generation sequencing in precision oncology approach of acute myeloid leukemia]. / Az új generációs szekvenálás jelentõsége az akut mieloid leukémia precíziós onkológiai megközelítésében.

In contrast to solid tumours, the genetic background of acute myeloid leukemia (AML) is characterized by a relatively low number of alterations per sample (average 3-5 mutations similarly to paediatric malignancies). Although the mutational background is rather heterogeneous, the detection of genetic alterations has diagnostic, prognostic and therapeutic relevance. We investigated cytogenetic and most commonly occurring molecular genetic alterations, and their co-occurrence in 830 AML patients diagnosed and treated in our institute between 2001 and 2019. Results from the recently introduced next generation sequencing for seven AML patients are also presented. Both methods (previously performed standard PCR-based tests and NGS) achieved the same results for commonly occurring mutations, but NGS technique was capable to identify further, rarely occurring mutations which bear diagnostic and prognostic importance according to the recent European LeukemiaNet recommendations. The introduction of NGS techniques to routine laboratory diagnostic applications is a required step following international expertise.

Current Trends in Applications of Circulatory Microchimerism Detection in Transplantation.

Primarily due to recent advances of detection techniques, microchimerism (the proportion of minor variant population is below 1%) has recently gained increasing attention in the field of transplantation. Availability of polymorphic markers, such as deletion insertion or single nucleotide polymorphisms along with a vast array of high sensitivity detection techniques, allow the accurate detection of small quantities of donor- or recipient-related materials. This diagnostic information can improve monitoring of allograft injuries in solid organ transplantations (SOT) as well as facilitate early detection of relapse in allogeneic hematopoietic stem cell transplantation (allo-HSCT). In the present review, genetic marker and detection platform options applicable for microchimerism detection are discussed. Furthermore, current results of relevant clinical studies in the context of microchimerism and SOT or allo-HSCT respectively are also summarized.

Donor KIR2DS1 reduces the risk of transplant related mortality in HLA-C2 positive young recipients with hematological malignancies treated by myeloablative conditioning.

BACKGROUND: Recognition of HLA-C2 group alleles on recipient cells by activating killer immunoglobulin like receptors, KIR2DS1 on donor natural killer cells may lead to increased graft-versus-leukemia effect or immunomodulation in patients treated by allogeneic hematopoietic stem cell transplantation (allo-HSCT) influencing disease free and overall survival (OS). OBJECTIVE: In the present study, 314 consecutive, allo-HSCT recipient and donor pairs were included with retrospective donor KIR-genotyping and clinical parameters analyzes. RESULTS: After a median follow-up of 23.6 months, recipients with HLA-C2 group allele (rC2) showed improved (p = 0.046) OS if transplanted with KIR2DS1 positive donors (d2DS1) compared to those without one or both of this genetic attribute. Within the myeloablative conditioning (MAC) subgroup (n = 227), rC2 homozygous+d2DS1 patients (n = 14) showed a 5 years OS of 93% followed by rC2 heterozygous+d2DS1 patients (n = 48, 65%) compared to rC2 and/or d2DS1 negatives (47%, p = 0.018). Multivariate analyses indicated rC2+d2DS1 positivity as an independent predictor of OS (HR:0.47, 0.26-0.86, p = 0.014) besides donor type, presence of CMV-reactivation or chemoresistant disease. Among MAC-treated patients, the combined rC2+d2DS1 presence was associated with a markedly decreased cumulative incidence of transplant related mortality (p = 0.0045). CONCLUSION: The combination of rC2+d2DS1 may be a favorable genetic constellation in allo-HSCT with MAC potentially reducing transplant related mortality.

Hungarian Marfan family with large FBN1 deletion calls attention to copy number variation detection in the current NGS era.

Copy number variations (CNVs) comprise about 10% of reported disease-causing mutations in Mendelian disorders. Nevertheless, pathogenic CNVs may have been under-detected due to the lack or insufficient use of appropriate detection methods. In this report, on the example of the diagnostic odyssey of a patient with Marfan syndrome (MFS) harboring a hitherto unreported 32-kb FBN1 deletion, we highlight the need for and the feasibility of testing for CNVs (>1 kb) in Mendelian disorders in the current next-generation sequencing (NGS) era.

Functional polymorphisms of innate immunity receptors are not risk factors for the non-SBP type bacterial infections in cirrhosis.

BACKGROUND & AIMS: Pattern recognition receptors (PRRs) have a key role in the innate host defense. Functional polymorphisms of various PRRs have been established to contribute to an increased susceptibility to spontaneous bacterial peritonitis (SBP). Their role in the development of cirrhosis-associated bacterial infections (BI), beyond SBP or progressive disease course related to pathological bacterial translocation (BT) remains unknown. METHODS: Three hundred and forty-nine patients with cirrhosis were genotyped for common NOD2 (R702W, G908R and L1007PfsinsC), TLR2 (-16934T>A), and TLR4 (D299G) variants. Incidence of BIs, decompensating events and liver-related death were assessed in a 5-year follow-up observational study. Pathological BT was assessed based on the presence of antimicrobial antibodies or lipopolysaccharide-binding protein (LBP) level. RESULTS: In patients with ascites (n = 88) only NOD2 gene variants were associated with an increased cumulative probability of SBP (76.9% ± 19.9%) compared to wild-type (30.9% ± 6.9%, PLogRank  = .047). Individual or combined PRR genetic profiles were associated with the risk of non-SBP type BI. Advanced disease stage (HR [95% CI]: 2.11 [1.38-3.25]) and prior history of a BI episode (HR: 2.42 [1.58-3.72]) were the major clinical risk factors of a subsequent BI. The risk of a non-SBP type BI in patients with advanced disease and a prior BI was even higher (HR: 4.74 [2.68-8.39]). The frequency of antimicrobial antibodies and LBP levels did not differ between various PRR genotypes. Correspondingly, PRR genetic profile was not able to predict the long-term disease course. CONCLUSIONS: In cirrhosis, functional polymorphisms of PRRs did not improve the identification of patients with high risk of BI beyond SBP or progressive diseases course.

Sex-specific survival difference in association with HLA-DRB1∗04 following allogeneic haematopoietic stem cell transplantation for lymphoid malignancies.

The role of HLA system in allogeneic haematopoietic stem cell transplantation (allo-HSCT) outcome is unarguable. In this study we investigated association of HLA-A,-B and-DRB1 alleles with overall survival (OS) in 186 patients undergoing allo-HSCT for lymphoid malignancies. Analyses confirmed significantly better OS for HLA-DRB1∗04 carriers compared with non-carriers (p = 0.01). Survival benefit was confined to male patients (in multivariate analyses p = 0.034, hazard ratio 0.35, 95% confidence interval 0.13-0.92), whereas in females no difference was noted (p = 0.82). Furthermore, donor gender also affected outcome and transplantation from female HLA-DRB1∗04 carrier donors resulted in superior survival compared with female non-carrier donors (p = 0.01). Combined analyses including recipient/donor gender and HLA-DRB1∗04 showed that survival of male patients varied significantly according to donor gender and HLA-DRB1∗04 carriership (p = 0.04) with best survival among HLA-DRB1∗04 carriers transplanted from female donors. Of relevance to our results, HLA-DRB1∗04 has been documented as risk allele group for lymphoid malignancies, and studies described a male-specific risk. We believe that our findings provide further supporting evidence for sex-specific alterations secondary to HLA-DRB1∗04 or related genes. Further studies are warranted to evaluate whether in contrast to general favour of male donors HLA-DRB1∗04 carrier patients with lymphoid malignancies could benefit from transplantation from female donors.

The adverse effect of FOPNL genomic variant is reversed by bortezomib-based treatment protocols in multiple myeloma.

Fibroblast growth factor receptor 1 oncogene partner N-terminal like gene (FOPNL) rs72773978 polymorphism was identified as an adverse prognostic factor in multiple myeloma (MM). We aimed to investigate the associations of rs72773978 with clinical characteristics and treatment outcome in 373 Hungarian MM patients. In our cohort, FOPNL polymorphism showed differential prognostic effect that depended on the treatment applied. Among patients treated with non-proteasome inhibitor (PI)-based therapy, carriership of the minor allele was significantly associated with adverse overall survival (p=.022). In contrast, the adverse effect was overcome by the application of PI-containing treatment (p=.048). Multivariate analyses revealed the independent adverse effect of rs72773978 on survival in the non-PI-treated group (p=.045), but not in PI treatment (OS: p=.093). We confirmed the adverse prognostic effect of rs72773978 associated with non-PI-based treatment regimens. Our results point to the importance of genotypic prognostic information associated with complex clinical background MM.

Recipient and donor JAK2 46/1 haplotypes are associated with acute graft-versus-host disease following allogeneic hematopoietic stem cell transplantation.

Several genetic polymorphisms have been implicated to affect the outcome of allogeneic hematopoietic stem cell transplantation (allo-HSCT). The role of cytokines in acute graft-versus-host disease (aGvHD) is well established and many of the involved cytokines signal through the Janus kinase (JAK) pathways. In this study, we assessed the association of recipient and donor JAK2 46/1 haplotypes and allo-HSCT outcome in a cohort of 124 acute myeloid leukemia patients. Both, recipient and donor 46/1 haplotypes significantly affected aGvHD grades II-IV development (p = 0.006 and p = 0.031, respectively), furthermore the influence of the haplotypes seemed to be additive. In multivariate analyses the recipient haplotype remained independently related (p = 0.012) to aGvHD, while the donor not (p = 0.08). We observed significantly less relapses among haplotype carriers (p = 0.004), but overall survival did not differ (p = 0.732). Our findings suggest that recipient and donor JAK2 46/1 haplotypes might be involved in the regulation of aGvHD.

Lipoprotein Lipase as a Prognostic Marker in Chronic Lymphocytic Leukemia.

The marked clinical heterogeneity of CLL makes early prognosis assessment important. Lipoprotein lipase (LPL) has been shown to confer adverse prognosis in CLL, recent data indicating it might also contribute to CLL cell survival and metabolism. We determined LPL mRNA expression in unselected peripheral blood of 84 CLL patients by RT PCR. Results were correlated with other prognostic markers and outcome. 30/84 (40 %) of cases were LPL positive based on the cutoff established by ROC analysis. In LPL positive patients significantly shorter median survival (136 vs 258 months, p < 0.0001) and time to first treatment intervals (36 vs 144 months, p < 0.002) were documented. LPL values correlated with male gender, higher stages, more treatment requirement, CD38 positivity and unmutated IgVH genes. Among cases with 13q deletion, LPL positivity identified a subcohort with poor outcome (median survival 108 months vs NR, p < 0.0001). In multivariate analysis, cytogenetic aberrations and LPL had significant impact on survival. Our results confirm that LPL is a strong predictor of outcome in CLL, able to improve prognostic accuracy in good risk cytogenetic subgroups. The relationship between its prognostic and functional role in CLL needs to be explored further.

Co-occurrence of Myeloproliferative Neoplasms and Solid Tumors Is Attributed to a Synergism Between Cytoreductive Therapy and the Common TERT Polymorphism rs2736100.

BACKGROUND: The germline telomerase reverse transcriptase (TERT) rs2736100_C variant was identified as a susceptibility factor for a variety of solid tumors and recently for myeloproliferative neoplasms (MPN). METHODS: LightCycler melting curve analysis was applied to detect risk alleles of TERT rs2736100_C and Janus kinase 2 (JAK2) rs12343867_C tagging 46/1 haplotype in 584 BCR-ABL1-negative MPN, 308 acute, and 86 chronic myeloid leukemia (AML and CML) patients and 400 healthy individuals. RESULTS: TERT rs2736100_C showed an increased allele frequency in BCR-ABL1-negative MPN patients compared with controls (62.7%±2.8% vs. 48.8%±3.5%, P < 0.0001) regardless of molecular background or disease type, but not in CML or AML. Combined TERT and JAK2 hetero- or homozygosity conferred even higher risk for classic MPN. Common complications (thrombosis, myelofibrosis, or leukemia) were not associated with the TERT variant; however, adverse survival was noted in TERT variant carrier polycythemia vera patients. MPN patients with the TERT CC genotype had a higher probability (44.4%) to die from solid tumors compared with TERT AC/AA individuals (5.3%; P = 0.004). TERT rs2736100_C carriers had increased risk of solid tumors independently from cytoreductive treatment [3.08 (1.03-9.26), P = 0.045]. CONCLUSIONS: TERT rs2736100_C polymorphism predisposes to the development of BCR-ABL1-negative MPN with the co-occurrence of solid tumors, especially with the usage of cytoreductive treatment. IMPACT: The high frequency of TERT variant in the classic MPN population highlights the importance of the avoidance of long-term cytoreductive treatment in MPN patients.

Carrier and prenatal diagnostic strategy and newly identified mutations in Hungarian haemophilia A and B families.

Deficiencies of blood coagulation factors VIII and IX (haemophilia A and haemophilia B) represent the most common inherited bleeding disorders with a wide range of causative mutations. Carrier and prenatal diagnostics are preferably performed by direct mutation detection; however, in certain situations, indirect family studies may also be useful. We aimed to utilize a combination of direct and indirect techniques for carrier and prenatal diagnostics in both haemophilias in a single national centre. Two hundred and eleven haemophilia A families were investigated by screening for inversions of introns 1 and 22, and by family studies using polymorphic markers. Twenty-eight haemophilia A and 39 haemophilia B families were investigated by Sanger-sequencing of the coding regions. Among severe haemophilia A families, frequencies of intron 22 and 1 inversions were 82 out of 145 (57%) and two out of 145 (1.4%). Sequencing of the entire coding region of the respective factor gene was performed and 12 (haemophilia A) and 5 (haemophilia B) previously unpublished disease-causing mutations were identified. For genetic markers used for haemophilia A indirect family testing, heterozygosity rates varied between 137 out of 327 [42% intragenic BclI restriction fragment length polymorphism (RFLP], 168 out of 254 (66% intragenic F8Civs13CA) and 202 out of 261 (77% extragenic DXS15CA) with a combined rate of 92% (intragenic markers) and 97% (all three markers). For male fetuses, prenatal diagnostics was provided to 43 haemophilia A families (n = 22 with direct mutation detection and n = 21 by indirect family testing) and to three haemophilia B families. The combination of direct and indirect molecular genetics approaches is a successful and cost-effective approach to provide carrier and prenatal diagnostics and risk assessment for inhibitor formation.