Finding a balance in reduced toxicity hematopoietic stem cell transplantation for thalassemia: role of infused CD3+ cell count and immunosuppression.

We performed a retrospective analysis on 124 patients with transfusion-dependent thalassemia who were registered in the German pediatric registry for stem cell transplantation. All patients underwent first allogeneic hematopoietic stem cell transplantation (HSCT) between 2011 and 2020 and belonged mainly to Pesaro risk class 1-2. Four-year overall (OS) and thalassemia-free survival (TFS) were 94.5% ± 2.9% and 88.0% ± 3.4% after treosulfan-fludarabine-thiotepa- and 96.9% ± 3.1% (P = 0.763) and 96.9% ± 3.1% (P = 0.155) after busulfan-fludarabine-based conditioning. Mixed chimerism below 75% occurred predominantly in treosulfan-based regimens (27.5% versus 6.2%). OS and TFS did not differ significantly between matched sibling, other matched family and matched unrelated donor (UD) HSCTs (OS: 100.0%, 100.0%, 96.3% ± 3.6%; TFS: 96.5% ± 2.4%, 90.0% ± 9.5%, 88.9% ± 6.0%). However, mismatched UD-HSCTs performed less favorable (OS: 84.7% ± 7.3% (P = 0.029); TFS: 79.9% ± 7.4% (P = 0.082)). We generated a scoring system reflecting the risk to develop mixed chimerism in our cohort. The main risk-reducing factors were a high CD3+ cell count (≥6 × 107/kg) in the graft, busulfan-conditioning, pre-conditioning therapy and low-targeted ciclosporin A trough levels. Acute GvHD grade III-IV in treosulfan-based concepts predominantly occurred in patients with UD and reduced GvHD prophylaxis but not in the context of high CD3+ cell doses. Taken together, this information might be used to develop more risk-adapted HSCT regimens for thalassemia patients.

Segmental quantification of hepatic lipid content based on volumetric MRI data in patients with suspected iron overload.

PURPOSE: To investigate the segmental distribution of hepatic fat fraction, determined with MRI (MR proton density fat fraction, short MR-PDFF) in patients suspected of having liver iron overload. METHODS: The liver of 44 patients examined with MRI using a 3D multi-echo gradient-echo sequence was segmented semiautomatically and subdivided into nine segments (segment 4 divided in 4a and 4b). Segmental fat content was determined on MR-PDFF maps. Whole-liver steatosis grades were compared to those found in individual segments. Segmental MR-PDFF differences were tested for statistical significance. RESULTS: The most common diseases were thalassemia, various forms of anemia, and hereditary hemochromatosis. No patients suffered from fat metabolism disease. Iron overload was present in 37/44 (84 %) patients. For the whole liver, 22 patients showed a steatosis grade of 0, 21 patients were graded S1, and one patient had a steatosis grade of 2. The grade of steatosis was underestimated in 5 of 21 patients (24 %) in segment 8 and in 8 of 21 patients (38 %) in segment 7. Highly significant segmental MR-PDFF differences were detected with p < 0.00 001, e. g., comparing segment 2 to 5. Segments 1 to 3 had the highest fat content, segments 7 and 8 had the lowest. CONCLUSION: Our results suggest that the storage of fat in the liver is inhomogeneous, so that segment-wise differing fat concentrations were found. Fat distribution in patients with suspected hepatic iron overload was similar to living liver donors. However, it showed significant differences compared with the values published for NAFLD patients, which were less pronounced in the group with high average hepatic MR-PDFF values than in the group with normal lipid content. In patients suspected of having iron overload, segment 8, which is mainly targeted for biopsy, and segment 7 may underestimate steatosis grade. KEY POINTS: · A volumetric analysis of 3D MRI data of patients with suspected hepatic iron overload yielded a markedly elevated MR proton density fat fraction (MR-PDFF) in hepatic segments 1 to 3.. · This hepatic fat distribution, observed for the whole patient cohort, is similar to healthy living liver donors.. · The subgroup of patients with a high average MR-PDFF ≥ 6.5 % shows this effect with lower segmental deviations.. · In patients without fat metabolic disorders, the steatosis grade may be underestimated when taking biopsies in segment 8 or 7..

Long-term haematological response and maintained immunological function after laparoscopic subtotal splenectomy in patients with hereditary spherocytosis.

INTRODUCTION: Subtotal or total splenectomy are recommended in severe and should be considered in intermediate forms of hereditary spherocytosis (HS). Data on laparoscopic subtotal splenectomy (LSTS) in HS patients are sparse. METHODS: Thirty three patients with HS (median age 10.7 years (yrs), range 1.8-15.5) underwent LSTS. Baseline and follow-up investigation included haematological parameters, microscopic analysis of pitted erythrocytes (pitE), and B-cell subpopulations assessed by flow cytometry. Results were compared to those of non-splenectomised HS patients, HS patients after total splenectomy (TS), and healthy individuals. RESULTS: After LSTS, haemoglobin levels were normalised in all patients. During median long-term follow-up of 3.9 yrs (range 1.1-14.9), only four patients presented mild anaemia. Despite re-growing of the remnant spleen none of the patients required a second surgical intervention. As compared to TS, PitE in LSTS patients were significantly lower and indicated normal to only moderately decreased spleen function. Relative but not absolute IgM memory B-cell counts were reduced in both LSTS and TS patients. CONCLUSIONS: LSTS is effective for the treatment of patients with HS. A small remnant spleen is sufficient to provide adequate phagocytic function and to induce a pool of IgM memory B-cells.

Noninvasive liver iron quantification by MRI using refocused gradient-echo (bSSFP): preliminary results.

PURPOSE: To evaluate the feasibility of using a balanced steady-state free precession sequence (bSSFP) to determine liver iron content (LIC). METHOD: Thirty-five consecutive patients with liver iron overload were examined with bSSFP. Signal intensity ratios of liver parenchyma to paraspinal muscles were retrospectively correlated with LIC values obtained by FerriScan, which was used as the reference method. Combinations of bSSFP protocols were also evaluated. The best combination was utilized to calculate LIC from bSSFP data. The sensitivity and specificity for the therapeutically relevant LIC threshold of 80 µmol/g (4.5 mg/g) were determined. RESULTS: LIC values ranged from 24 to 756 µmol/g. The best SIR-to-LIC correlation of a single protocol was obtained with a 3.5-ms repetition time (TR) and 17° excitation flip angle (FA). A combination of protocols with TRs of 3.5, 5, and 6.5 ms, each at 17° FA, yielded a superior correlation. LIC values calculated using this combination resulted in a sensitivity/specificity of 0.91/0.85. CONCLUSION: bSSFP is basically suitable to determine LIC. Its advantages are high SNR efficiency and the ability to acquire the entire liver in a breath hold without acceleration techniques. KEY POINTS: · The bSSFP sequence is suited to quantify liver iron overload.. · bSSFP has a high scanning efficiency and potential for LIC screening.. · Despite susceptibility artifacts, the LIC determined from bSSFP data showed high accuracy.. CITATION FORMAT: · Wunderlich AP, Cario H, Götz M et al. Noninvasive liver iron quantification by MRI using refocused gradient-echo (bSSFP): preliminary results. Fortschr Röntgenstr 2023; 195: 804 - 808.

Comprehensive in silico and functional studies for classification of EPAS1/HIF2A genetic variants identified in patients with erythrocytosis.

Gain-of-function mutations in the EPAS1/HIF2A gene have been identified in patients with hereditary erythrocytosis that can be associated with the development of paraganglioma, pheochromocytoma and somatostatinoma. In the present study, we describe a unique European collection of 41 patients and 28 relatives diagnosed with an erythrocytosis associated with a germline genetic variant in EPAS1. In addition we identified two infants with severe erythrocytosis associated with a mosaic mutation present in less than 2% of the blood, one of whom later developed a paraganglioma. The aim of this study was to determine the causal role of these genetic variants, to establish pathogenicity, and to identify potential candidates eligible for the new hypoxia-inducible factor-2 α (HIF-2α) inhibitor treatment. Pathogenicity was predicted with in silico tools and the impact of 13 HIF-2b variants has been studied by using canonical and real-time reporter luciferase assays. These functional assays consisted of a novel edited vector containing an expanded region of the erythropoietin promoter combined with distal regulatory elements which substantially enhanced the HIF-2α-dependent induction. Altogether, our studies allowed the classification of 11 mutations as pathogenic in 17 patients and 23 relatives. We described four new mutations (D525G, L526F, G527K, A530S) close to the key proline P531, which broadens the spectrum of mutations involved in erythrocytosis. Notably, we identified patients with only erythrocytosis associated with germline mutations A530S and Y532C previously identified at somatic state in tumors, thereby raising the complexity of the genotype/phenotype correlations. Altogether, this study allows accurate clinical follow-up of patients and opens the possibility of benefiting from HIF-2α inhibitor treatment, so far the only targeted treatment in hypoxia-related erythrocytosis disease.

Volumetric Evaluation of 3D Multi-Gradient-Echo MRI Data to Assess Whole Liver Iron Distribution by Segmental R2* Analysis: First Experience.

PURPOSE: MR transverse relaxation rate R2* has been shown to be useful for monitoring liver iron overload. A sequence enabling acquisition of the whole liver in a single breath hold is now available, thus allowing volumetric hepatic R2* distribution studies. We evaluated the feasibility of computer-assisted whole liver segmentation of 3 D multi-gradient-echo MRI data, and compared whole liver R2* determination to analyzing only a single slice. Also, segmental R2* differences were studied. MATERIALS AND METHODS: The liver of 44 patients, investigated by multi-gradient echo MRI at 1.5 T, was segmented and divided into nine segments. Segmental R2* values were examined for all patients together and with respect to two criteria: average R2* values, and reason for iron overload. Correlation of single-slice and volumetric data was tested with Spearman's rank test, segmental and group differences were evaluated by analysis of variance. RESULTS: Whole-liver R2* values correlated excellent to single slice data (p < 0.001). The lowest R2* occurred in segment 1 (S1), differences of S1 with regard to other segments were significant in five cases and highly significant in two cases. Patients with high average R2* showed significant differences between S1 and segments 2, 6, and 7. Disease-related differences with respect to S1 were significant in segments 3 to 5 and 7. CONCLUSION: Our results suggest inhomogeneous hepatic iron distribution. Low R2* in S1 may be explained by its special vascularization. KEY POINTS: · Hepatic R2* distribution is not as homogeneous as previously thought.. · Liver segments might have a functional relevance.. · Segmental and total liver R2* values coincide best in segment 8.. CITATION FORMAT: · Wunderlich AP, Cario H, Kannengießer S et al. Volumetric Evaluation of 3D Multi-Gradient-Echo MRI Data to Assess Whole Liver Iron Distribution by Segmental R2* Analysis: First Experience. Fortschr Röntgenstr 2023; 195: 224 - 233.

Iron deficiency anemia and thrombosis risk in children-revisiting an old hypothesis.

Iron deficiency anemia has a high prevalence in children and has repeatedly been implicated as a risk factor for arterial and venous thrombosis. As an effective therapy for iron deficiency anemia is available, understanding the association between this form of anemia and the potentially severe thrombosis phenotype is of major clinical interest. Recent findings shed light on pathophysiology of hypercoagulability resulting from iron-restricted erythropoiesis. Specifically, an animal model of induced iron deficiency allowed identifying multiple mechanisms, by which iron deficiency anemia results in increased thrombus formation and thrombus progression both in arterial and venous thrombosis. These findings complement and support conclusions derived from clinical data. The purpose of this mini review is to summarize current evidence on the association of iron deficiency anemia and thrombosis. We want to increase the awareness of iron deficiency as a risk factor for thrombosis in the pediatric population. We discuss how novel pathophysiological concepts can be translated into the clinical settings and suggest clinical studies on prevention and treatment strategies in high-risk patient groups.

Transition in Sickle Cell Disease (SCD): A German Consensus Recommendation.

Sickle cell disease (SCD) is considered a rare disease in Germany. Due to the increasing prevalence, the acute and chronic morbidities associated with the disease and the sharp increase in the mortality rate of young adults, a need-based transition structure for patients with SCD in Germany is explicitly required. This is the first multicenter German consensus statement addressing the importance of implementing a standardized transition guideline that allows adolescents and young adults to safely transition from pediatric to adult care. Early identification of medical needs and intervention remains important in the context of chronic diseases. Effective measures can improve health care in general, as they lead to a reduction in disease and the consequential economic burden. It is noteworthy that improving structural barriers remains a key challenge even in highly developed countries such as Germany. Inclusion of these transition services for patients with SCD into the regular care of chronically ill adolescents and young adults should be ensured, as well as the coverage of costs associated with a structured transition process.

Benefits of a Disease Management Program for Sickle Cell Disease in Germany 2011-2019: The Increased Use of Hydroxyurea Correlates with a Reduced Frequency of Acute Chest Syndrome.

Sickle Cell Disease (SCD) is the most common monogenic disorder globally but qualifies as a rare disease in Germany. In 2012, the German Society for Paediatric Oncology and Haematology (GPOH) mandated a consortium of five university hospitals to develop a disease management program for patients with SCD. Besides other activities, this consortium issued treatment guidelines for SCD that strongly favour the use of hydroxyurea and propagated these guidelines in physician and patient education events. In order to quantify the effect of these recommendations, we made use of claims data that were collected by the research institute (WIdO) of the major German insurance company, the Allgemeine Ortskrankenkasse (AOK), and of publicly accessible data collected by the Federal Statistical Office (Statistisches Bundesamt, Destatis). While the number of patients with SCD in Germany increased from approximately 2200 in 2011 to approximately 3200 in 2019, important components of the recently issued treatment guidelines have been largely implemented. Specifically, the use of hydroxyurea has more than doubled, resulting in a proportion of approximately 44% of all patients with SCD being treated with hydroxyurea in 2019. In strong negative correlation with the use of hydroxyurea, the frequency of acute chest syndromes decreased. Similarly, the proportion of patients who required analgesics and hospitals admissions declined. In sum, these data demonstrate an association between the dissemination of treatment guidelines and changes in clinical practice. The close temporal relationship between the increased use of hydroxyurea and the reduction in the incidence of acute chest syndrome in a representative population-based analysis implies that these changes in clinical practice contributed to an improvement in key measures of disease activity.

Introduction of Universal Newborn Screening for Sickle Cell Disease in Germany-A Brief Narrative Review.

Sickle cell disease (SCD) is a severe non-malignant disorder of hemoglobin and is inherited in an autosomal-recessive manner [...].

Liver Iron Content Determination Using a Volumetric Breath-Hold Gradient-Echo Sequence With In-Line R2 * Calculation.

BACKGROUND: Liver iron overload is a serious condition occurring in patients requiring blood transfusions (eg, in thalassemia and different forms of anemia) or with dysfunctional iron resorption, since there is no physiological mechanism to excrete iron. Above a certain level of iron concentration, chelation therapy is indicated. To monitor therapy success, liver iron content should be assessed regularly. A noninvasive method is important for patient management. Existing MRI methods suffer from long acquisition times and cost. PURPOSE: To study the correlation of liver iron content (LIC) reference values to liver R2 * determined using a 3D breath-hold multigradient echo (GRE) MRI sequence, employing accelerated acquisition by parallel imaging and in-line R2 * calculation. STUDY TYPE: Prospective. POPULATION: In all, 117 patients (22.1 ± 14.1 years, 66 men) suspected of iron overload. SEQUENCE: GRE. FIELD STRENGTH: 1.5T. ASSESSMENT: For comparison, a regulatory-approved method with a considerably longer scan time was used, providing LIC reference values. Participants were divided into a calibration group (65 participants), analyzed independently by two observers, and a validation group (52 participants). STATISTICAL TESTS: Linear correlation parameters were evaluated for R2 * values with LIC reference values, and for LIC determined from R2 * for validation group participants with LIC reference values. Sensitivity/specificity for clinical relevant LIC thresholds were analyzed. Interobserver variability was determined by intraclass correlation coefficient (ICC). RESULTS: Interobserver agreement was excellent, with an ICC of 0.99, P < 0.001. Good correlation (R2 = 0.89) and congruence of LIC values obtained with our method to LIC reference values was found, and almost identical diagnostic accuracy. Sensitivity/specificity were 0.98/0.67 for the diagnostic relevant LIC threshold of 4.5 mg/g and 1.0/0.95 for the threshold of 7 mg/g. DATA CONCLUSION: MRI acquisition times for determination of LIC can be significantly reduced by the use of comprehensive in-line R2 * map generation without compromising diagnostic accuracy. LEVEL OF EVIDENCE: 1 TECHNICAL EFFICACY STAGE: 2.

Determinants of utilization of cryopreservation of germ cells in adolescent cancer patients in four European countries.

Infertility is a relevant late-effect following cancer treatment; yet, a large proportion of survivors cannot recall having been informed of this risk. In an intervention study, we examined if and how supportive patient information material on fertility/fertility-preserving measures influences utilization of cryopreservation in adolescent cancer patients. The control group, recruited 03/2014-01/2016, received the usual patient education at initial diagnosis. The intervention group, recruited 04/2016-10/2017, received patient education supported by a fertility flyer and brochure. Patients and parents were each asked questions on utilization of cryopreservation in a questionnaire 3 and 6 months after initial diagnosis. Patient core and therapy data were obtained from medical records. Overall, cryopreservation rates showed no significant difference between the control (32.7%, n = 37/113) and intervention group (36.6%, n = 37/101). In the control group, cryopreservation was associated with gender (OR 0.100, CI 0.023-0.427), age (OR 1.559, CI 1.077-2.258) and recalling information on fertility protection (OR 33.663, CI 2.100-539.574); in the intervention group, cryopreservation was related to gender (OR 0.093, CI 0.026-0.330) and the estimated infertility risk (OR 43.665, CI 2.157-883.974).Conclusion: Cryopreservation rates did not overall increase following the intervention; however, the individual risk seemed to be brought into attention more: Those at risk, including younger patients, cryopreserved at higher rates.What is Known:•Infertility is a relevant late-effect following adolescent cancer.•Guidelines recommend to offer fertility protection before cancer treatment.•A relevant proportion of adolescents with cancer are not aware of this risk.•Fertility protection seems under-used in cancer patients at risk for infertility.What is New:•Information material on fertility and protection in adolescents did not increase overall rates of cryopreservation.•Cryopreservation rates were improved according to individual risk for infertility.•Our flyers and brochures on fertility in cancer patients are available in various languages.

Sickle cell disease in Germany: Results from a national registry.

BACKGROUND: Limited data on the prevalence and medical care of sickle cell disease (SCD) in Germany are available. Here, we make use of a patient registry to characterize the burden of disease and the treatment modalities for patients with SCD in Germany. PROCEDURE: A nationwide German registry for patients with SCD documents basic data on diagnosis and patient history retrospectively at the time of registration. A prospective annual documentation provides more details on complications and treatment of SCD. For the current analyses, data of 439 patients were available. RESULTS: Most patients had homozygous SCD (HbSS 75.1%, HbS/ß-thalassemia 13.2%, and HbSC 11.3%). The median age at diagnosis was 1.9 years (interquartile range, 0.6-4.4 years), most patients were diagnosed when characteristic symptoms occurred. Sepsis and stroke had affected 3.2% and 4.2% of patients, respectively. During the first year of observation, 48.3% of patients were admitted to a hospital and 10.1% required intensive care. Prophylactic penicillin was prescribed to 95.6% of patients with homozygous SCD or HbS/ß thalassemia below the age of six and hydroxycarbamide to 90.4% of patients above the age of two years. At least one annual transcranial Doppler ultrasound was documented for 74.8% of patients between 2 and 18 years. CONCLUSION: With an estimated number of at least 2000, the prevalence of SCD in Germany remains low. Prospectively, we expect that the quality of care for children with SCD will be further improved by an earlier diagnosis after the anticipated introduction of a newborn screening program for SCD.

Fertility knowledge and associated empowerment following an educational intervention for adolescent cancer patients.

INTRODUCTION/OBJECTIVES: Fertility preservation is a major concern for adolescent cancer patients; yet, educational gaps remain. Our intervention study examined whether specially designed educational materials regarding fertility preservation increase knowledge and empowerment of patients and parents. METHODS: Eleven paediatric-oncological centres in four European countries agreed to enrol all eligible patients and parents in a questionnaire survey at 3 and 6 months after diagnosis. Treating physicians were surveyed on their medical consultation regarding fertility. RESULTS: Educational intervention increased knowledge in both patients (n = 113 and n = 101 in the control and intervention groups, respectively) and parents (n = 111 and n = 99 in the control and intervention groups, respectively), but the difference did not achieve statistical significance (knowledge difference patients: 5.6% (t0)/13.1% (t1); parents: 6.4% (t0)/3.8% (t1)). Parents of older patients (OR = 1.3, 95%CI = 1.1-1.7) and higher educational groups (OR = 6.2, 95%CI = 2.1-18.3) in the intervention group (OR = 1.9, 95%CI = 1.03-3.7) achieved higher knowledge levels. Empowerment was significantly improved in both patients (p = 0.046, d = 0.27) and parents (p = 0.046, d = 0.48) in the intervention group. DISCUSSION/CONCLUSIONS: In our study, the use of specifically prepared flyers and brochures successfully raised the level of fertility preservation knowledge in parents of older patients as well as parents with higher educational levels. Overall, the intervention improved patient and parent empowerment. Subsequent projects will include simpler information and digital material to particularly reach out to younger and less educated individuals.

[What's new in diagnostics and treatment of hemoglobinopathies?] / Was ist neu in der Diagnostik und Therapie der Hämoglobinopathien?

Improvement of care for thalassemia during previous decades focussed on the development of new diagnostic and treatment modalities concerning secondary hemochromatosis resulting from symptomatic treatment with regular transfusion. In addition, hematopoetic stem transplantation as the so far only curative approach had been developed and constantly improved. New approaches for the treatment of thalassemias already entering clinical practice include e. g. additive gene therapy and medical treatment with new drugs like luspatercept. For sickle cell disease, a variety of treatment approaches either to correct the genetic change by gene therapy, to affect the underlying mechanism of sickling e. g. by R-state- stabilising agents, or later the pathophysiology of vaso-occlusive complications e. g. by inhibiting selectins are currently thoroughly investigated and some of them will probably get approval very soon.

Quantitative DWI predicts event-free survival in children with neuroblastic tumours: preliminary findings from a retrospective cohort study.

BACKGROUND: Quantitative diffusion-weighted imaging (DWI) probes into tissue microstructure in solid tumours. In this retrospective ethically approved study, we investigated DWI as a potential non-invasive predictor of tumour dignity and prognosis in paediatric patients with neuroblastic tumours. METHODS: Nineteen consecutive patients with neuroblastoma (NB, n = 15), ganglioneuroblastoma (GNB, n = 1) and ganglioneuroma (GN, n = 3) underwent 3-T magnetic resonance imaging at first diagnosis and after 3-month follow-up, following a protocol including DWI (b = 50 and 800 s/mm2) in addition to standard sequences. All DWI scans were analysed for tumour volume assessment and apparent diffusion coefficient (ADC) calculation. Correlation with tumour pathology and risk factors (bone-marrow metastases, MYCN-amplification and 1p-deletion), therapeutic regime (observation versus chemotherapy) and clinical follow-up was evaluated. RESULTS: At baseline, mean ADC in NB was lower than in GNB/GN (0.76 vs. 1.47 × 10-3 mm2/s, p = 0.003). An ADC cutoff ≤ 1.05 identified malignant disease with 100.0% sensitivity (95% confidence interval [CI] 29.2-100.0%) and 93.8% specificity (95% CI 69.8-99.8%). Initial ADC was < 0.80 in all NB patients with eventual tumour relapse. During follow-up, tumour ADC values increased in the observation group (NB/GN) without relapse (p = 0.043). In eventually relapsing tumours, ADC values at follow-up tended to decrease further despite reduction in tumour volume. CONCLUSIONS: ADC values at first presentation differed significantly between malignant and benign neuroblastic tumours. Low baseline ADC was predictive of tumour progression and relapse in NB patients. With therapy, increasing ADC values appeared to predict relapse-free survival, while a decreasing ADC during therapy was an indicator of poor prognosis.

PanCareLIFE: The scientific basis for a European project to improve long-term care regarding fertility, ototoxicity and health-related quality of life after cancer occurring among children and adolescents.

AIMS: Survival after cancer diagnosed during childhood or adolescence continues to improve with new treatments and supportive therapies. Optimal long-term care requires that risks to vulnerable organs are clearly defined and translated into guidelines that are implemented into practice. PanCareLIFE is a pan-European consortium that addresses survivorship issues comprising fertility, hearing impairment and quality of life. This article describes the scientific basis of PanCareLIFE's studies. METHODS: PanCareLIFE involves 17 partner institutions from eight European countries, with additional 11 data providers from five other countries. Study designs and methods include molecular genetic, cohort and case-control studies, a longitudinal study and an intervention study. Ethics and data protection issues have been taken into account from the beginning. RESULTS: PanCareLIFE will investigate the way that treatment impairs female fertility, by evaluating anti-Müllerian hormone levels and the underlying genetic susceptibility to loss of fertility. For our fertility studies, more than 6000 survivors have completed questionnaires, more than 1500 provided serum samples and more than 400 case-control triads have been identified. Fertility preservation guidelines for boys and girls will be developed. More than 2000 survivors have contributed audiograms for the ototoxicity study. Almost 1000 samples were sent for genetic analysis related to ototoxicity and gonadal reserve. The SF-36 questionnaire will measure quality of life in more than 10,000 survivors. CONCLUSIONS: The large number of subjects enrolled in PanCareLIFE and the detailed information accumulated will allow in-depth evaluation of important outcomes. Fertility preservation guidelines will help patients and their families make informed decisions and contribute to their long-term well-being.

Serum ferritin is not a reliable predictor to determine iron overload in thalassemia major patients post-hematopoietic stem cell transplantation.

OBJECTIVE: Iron overload (IO) in transfusion-dependent anemia persists after hematopoietic stem cell transplantation (HSCT) and can cause long-term organ damage. In many studies, the diagnosis of IO before and after HSCT is based on serum ferritin (SF) levels rather than on assessment of liver iron concentration (LIC) by MRI or SQUID. METHOD: In a retrospective multicenter study, we analyzed the concordance for indication of iron depletion therapy and correlation between LIC and SF of 36 thalassemia patients after HSCT. LIC was determined either by MRI-R2 (FerriScan®) or SQUID. RESULTS: The concordance between LIC and SF varies over time after transplant (P = 0.011). The correlation between SF and LIC was strong in the first year (Spearman's rho 0.75; P < 0.001). In agreement, the concordance between SF and LIC concerning indication for treatment was close to 1 with an overall error rate ca. of 10%. In particular in the first year after HSCT, SF underestimates the degree of iron overload. However, in the longitudinal analysis since the second year post-HSCT onward no association was found between LIC and SF (P = 0.217). Furthermore, in the second year after HSCT, the overall error rate was 35%, whereas in the 3rd, 4th, and >4th year, it was 58%, 60%, and 25%, respectively. CONCLUSIONS: Our data suggest serum ferritin is not a reliable predictor to determine iron overload in thalassemia patients after HSCT.

Identification of a new VHL exon and complex splicing alterations in familial erythrocytosis or von Hippel-Lindau disease.

Chuvash polycythemia is an autosomal recessive form of erythrocytosis associated with a homozygous p.Arg200Trp mutation in the von Hippel-Lindau (VHL) gene. Since this discovery, additional VHL mutations have been identified in patients with congenital erythrocytosis, in a homozygous or compound-heterozygous state. VHL is a major tumor suppressor gene, mutations in which were first described in patients presenting with VHL disease, which is characterized by the development of highly vascularized tumors. Here, we identify a new VHL cryptic exon (termed E1') deep in intron 1 that is naturally expressed in many tissues. More importantly, we identify mutations in E1' in 7 families with erythrocytosis (1 homozygous case and 6 compound-heterozygous cases with a mutation in E1' in addition to a mutation in VHL coding sequences) and in 1 large family with typical VHL disease but without any alteration in the other VHL exons. In this study, we show that the mutations induced a dysregulation of VHL splicing with excessive retention of E1' and were associated with a downregulation of VHL protein expression. In addition, we demonstrate a pathogenic role for synonymous mutations in VHL exon 2 that altered splicing through E2-skipping in 5 families with erythrocytosis or VHL disease. In all the studied cases, the mutations differentially affected splicing, correlating with phenotype severity. This study demonstrates that cryptic exon retention and exon skipping are new VHL alterations and reveals a novel complex splicing regulation of the VHL gene. These findings open new avenues for diagnosis and research regarding the VHL-related hypoxia-signaling pathway.