Treatment and management of primary antibody deficiency: German interdisciplinary evidence-based consensus guideline.

This evidence-based clinical guideline provides consensus-recommendations for the treatment and care of patients with primary antibody deficiencies (PADs). The guideline group comprised 20 clinical and scientific expert associations of the German, Swiss, and Austrian healthcare system and representatives of patients. Recommendations were based on results of a systematic literature search, data extraction, and evaluation of methodology and study quality in combination with the clinical expertise of the respective representatives. Consensus-based recommendations were determined via nominal group technique. PADs are the largest clinically relevant group of primary immunodeficiencies. Most patients with PADs present with increased susceptibility to infections, however immune dysregulation, autoimmunity, and cancer affect a significant number of patients and may precede infections. This guideline therefore covers interdisciplinary clinical and therapeutic aspects of infectious (e.g., antibiotic prophylaxis, management of bronchiectasis) and non-infectious manifestations (e.g., management of granulomatous disease, immune cytopenia). PADs are grouped into disease entities with definitive, probable, possible, or unlikely benefit of IgG-replacement therapy. Summary and consensus-recommendations are provided for treatment indication, dosing, routes of administration, and adverse events of IgG-replacement therapy. Special aspects of concomitant impaired T-cell function are highlighted as well as clinical data on selected monogenetic inborn errors of immunity formerly classified into PADs (APDS, CTLA-4-, and LRBA-deficiency).

Pooling, room temperature, and extended storage time increase the release of adult-specific biologic response modifiers in platelet concentrates: a hidden transfusion risk for neonates?

BACKGROUND: Adult donor platelets (PLTs) are frequently transfused to prevent or stop bleeding in neonates with thrombocytopenia. There is evidence for PLT transfusion-related morbidity and mortality, leading to the hypothesis on immunomodulatory effects of transfusing adult PLTs into neonates. Candidate factors are biologic response modifiers (BRMs) that are expressed at higher rates in adult than in neonatal PLTs. This study investigated whether storage conditions or preparation methods impact on the release of those differentially expressed BRMs. STUDY DESIGN AND METHODS: Pooled PLT concentrates (PCs) and apheresis PCs (APCs) were stored under agitation for up to 7 days at room temperature (RT) or at 2 to 8°C. The BRMs CCL5/RANTES, TGFß1, TSP1, and DKK1 were measured in PCs' supernatant, lysate, and corresponding plasma. PLT function was assessed by light transmission aggregometry. RESULTS: Concerning the preparation method, higher concentrations of DKK1 were found in pooled PCs compared to APCs. In supernatants, the concentrations of CCL5, TGFß1, TSP1, and DKK1 significantly increased, both over standard (≤4 days) and over extended storage times (7 days). Each of the four BRMs showed an up to twofold increase in concentration after storage at RT compared to cold storage (CS). There was no difference in the aggregation capacity. CONCLUSION: This analysis shows that the release of adult-specific BRMs during storage is lowest in short- and CS APCs. Our study points to strategies for reducing the exposure of sick neonates to BRMs that can be specifically associated to PLT transfusion-related morbidity.

Transfusion of red blood cells does not impact progression-free and overall survival after surgery for ovarian cancer.

BACKGROUND: Allogeneic red blood cells (RBCs) have the potential to impact the immunosurveillance of the recipient and may therefore increase the risk of recurrence after cancer surgery. In this article the relationship between perioperative RBC transfusion and the risk of recurrence after ovarian cancer surgery is examined. STUDY DESIGN AND METHODS: This is a retrospective cohort analysis of a prospective database of patients who underwent surgery due to primary ovarian cancer between 2006 and 2014 and who had no residual disease after surgery. Patients who did and did not receive perioperative RBC transfusion were compared. The primary endpoint was progression-free survival (PFS). Propensity score matching (PSM) and Cox proportional hazards regression (CPH) was used to control for between-group differences of prognostic determinants. RESULTS: A total of 529 patients with a median follow-up of 51.4 months (95% CI, 46.1-56.5) were eligible for analysis. Of those, 408 patients (77.1%) received allogeneic, leukoreduced RBCs with a median of 4 units (IQR, 2-6) per patient. There was a strong selection bias of prognostic determinants between patients with and without transfusion. In unadjusted analysis, transfusion of RBCs was associated with an increased risk of cancer recurrence (hazard ratio [HR] of PFS 2.71 [95% CI, 1.94-3.77], p < 0.001). After bias reduction, transfusion of RBCs was no longer associated with an increased risk of cancer recurrence, neither in PSM-adjusted (HR 1.03 [95% CI, 0.59-1.80], p = 0.91), nor in multivariable CPH-adjusted analysis (HR 1.26 [95% CI, 0.85-1.86], p = 0.23). CONCLUSION: Perioperative transfusion of RBCs did not increase the risk of recurrence after ovarian cancer surgery.

Duration of storage influences the hemoglobin rising effect of red blood cells in patients undergoing major abdominal surgery.

BACKGROUND: After transfusion of senescent red blood cells (RBCs) a considerable fraction is rapidly cleared from the recipients' circulation. Thus, transfusion of senescent RBCs may be less effective in terms of increasing hemoglobin concentration (cHb) after transfusion. STUDY DESIGN AND METHODS: Data were retrospectively obtained in patients who underwent major abdominal surgery between 2006 and 2012. Patients were eligible if they received RBCs during surgery and had at least two arterial blood gas analyses performed. The primary endpoint was the increase of recipients' cHb related to the transfusion of 1 unit of RBCs with respect to different storage periods. Four storage periods were defined according to the distribution of RBC storage of the study population. General estimating equation was used for calculation of the primary endpoint and to adjust for confounding variables. RESULTS: A total of 598 arterial blood gas samples from 120 patients, receiving 429 RBC units, were analyzed. Mean (±SD) RBC storage was 21 (±9) days. RBC storage duration and the increase in recipients' cHb were inversely and gradually related; that is, the older the RBCs, the lower the increase in the recipients' cHb after transfusion (storage < 12 days, ΔcHb per unit RBCs +0.82 [95% confidence interval, 0.42-1.21] g/dL, p < 0.01; storage 12-20 days, +0.66 [0.46-0.86] g/dL, p < 0.01; storage 21-29 days, +0.56 [0.33-0.79] g/dL, p < 0.01; storage ≥30 days, +0.39 [0.07 to 0.71] g/dL, p = 0.02). CONCLUSION: Transfusion of senescent RBCs increased cHb less effectively than transfusion of fresher RBCs.

Immobilization and high platelet count are associated with thromboembolic complications in heparin-induced thrombocytopenia.

PURPOSE: Immune-mediated heparin-induced thrombocytopenia (HIT type II, HIT) is a potentially serious adverse drug reaction characterized by an increased risk of venous and arterial thrombosis. This study aimed to identify risk factors associated with the development of these complications. METHODS: Our study cohort included patients with HIT assembled in our pharmacovigilance center by reports from 51 collaborating hospitals in Berlin, Germany. To identify risk factors for thromboembolic complications, patients with thromboembolic events (cases) were compared to those without thromboembolic events (controls) in a case-control design. We applied univariable and multivariable logistic regression analysis to estimate odds ratios (OR) and corresponding 95% confidence intervals (CI) for potential risk factors of thromboembolic complications. RESULTS: Our cohort comprised 209 HIT patients. Of those, 53 developed thromboembolic complications. Most HIT patients received heparin for medical indications (42.1%) or in the context of cardiovascular surgery (40.2%). Of the 78 thromboembolic complications, 49 (63%) and 29 (37%) were observed in the venous and arterial vascular bed, respectively. The main locations were deep vein thrombosis (39.7%), pulmonary embolism (16.7%), and limb artery thrombosis (16.7%). In multivariable analysis, immobilization prior to HIT (OR 4.6, 95% CI 1.2-18.0; P = .026) and higher platelet counts before initiation of heparin therapy (OR 1.004, 95% CI 1.000-1.008; P = .046) were independently associated with the occurrence of thromboembolic events. CONCLUSIONS: Immobilization and a high platelet count (with a low effect size) are additional risk factors of thromboembolic complications in the course of HIT.

Safety and efficacy of eltrombopag for treatment of chronic immune thrombocytopenia: results of the long-term, open-label EXTEND study.

Patients with chronic immune thrombocytopenia may have bleeding resulting from low platelet counts. Eltrombopag increases and maintains hemostatic platelet counts; however, to date, outcome has been reported only for treatment lasting ≤ 6 months. This interim analysis of the ongoing open-label EXTEND (Eltrombopag eXTENded Dosing) study evaluates the safety and efficacy of eltrombopag in 299 patients treated up to 3 years. Splenectomized and nonsplenectomized patients achieved platelets ≥ 50 000/µL at least once (80% and 88%, respectively). Platelets ≥ 50 000/µL and 2 × baseline were maintained for a median of 73 of 104 and 109 of 156 cumulative study weeks, respectively. Bleeding symptoms (World Health Organization Grades 1-4) decreased from 56% of patients at baseline to 20% at 2 years and 11% at 3 years. One hundred (33%) patients were receiving concomitant treatments at study entry, 69 of whom attempted to reduce them; 65% (45 of 69) had a sustained reduction or permanently stopped ≥ 1 concomitant treatment. Thirty-eight patients (13%) experienced ≥ 1 adverse events leading to study withdrawal, including patients meeting protocol-defined withdrawal criteria (11 [4%] thromboembolic events, 5 [2%] exceeding liver enzyme thresholds). No new or increased incidence of safety issues was identified. Long-term treatment with eltrombopag was generally safe, well tolerated, and effective in maintaining platelet counts in the desired range.

Drug-induced agranulocytosis in the Berlin case-control surveillance study.

PURPOSE: Drug-induced agranulocytosis (DIAG) is a rare but serious adverse drug reaction. The Berlin Case-Control Surveillance Study (FAKOS) aimed to identify pharmaceuticals with an increased risk for this condition. METHODS: Adult patients with acute non-chemotherapy-induced agranulocytosis, developed in hospital or in the outpatient setting, were ascertained by active surveillance in all 51 Berlin hospitals between the years 2000 and 2010. Applying the criteria of the World Health Organization, a standardized drug causality assessment was conducted for each agranulocytosis patient to determine possible drug aetiology. Drug risks were quantified in a case-control design with unconditional logistic regression analysis. RESULTS: Sixty-three out of 88 validated cases of agranulocytosis were identified as being at least probably drug-related. Drug causality assessment resulted in 36 pharmaceuticals with a certain or probable relationship to agranulocytosis. Drugs involved in ≥ 3 cases with a probable or certain causality were metamizole (dipyrone) (N = 10), clozapine (N = 6), sulfasalazine (N = 5), thiamazole (N = 5), and carbamazepine (N = 3). In case-control analysis, six drugs were identified with significant odds ratios for DIAG. The highest odds ratios were observed for clozapine, sulfasalazine, and thiamazole. CONCLUSIONS: Our findings are generally in agreement with those of earlier case-control studies. The spectrum of drugs causing acute agranulocytosis has not changed considerably over recent years, despite many newly marketed drugs. Evidence for induction of agranulocytosis by some new pharmaceuticals is supported.

Genome-wide scan identifies TNIP1, PSORS1C1, and RHOB as novel risk loci for systemic sclerosis.

Systemic sclerosis (SSc) is an orphan, complex, inflammatory disease affecting the immune system and connective tissue. SSc stands out as a severely incapacitating and life-threatening inflammatory rheumatic disease, with a largely unknown pathogenesis. We have designed a two-stage genome-wide association study of SSc using case-control samples from France, Italy, Germany, and Northern Europe. The initial genome-wide scan was conducted in a French post quality-control sample of 564 cases and 1,776 controls, using almost 500 K SNPs. Two SNPs from the MHC region, together with the 6 loci outside MHC having at least one SNP with a P<10(-5) were selected for follow-up analysis. These markers were genotyped in a post-QC replication sample of 1,682 SSc cases and 3,926 controls. The three top SNPs are in strong linkage disequilibrium and located on 6p21, in the HLA-DQB1 gene: rs9275224, P = 9.18×10(-8), OR = 0.69, 95% CI [0.60-0.79]; rs6457617, P = 1.14×10(-7) and rs9275245, P = 1.39×10(-7). Within the MHC region, the next most associated SNP (rs3130573, P = 1.86×10(-5), OR = 1.36 [1.18-1.56]) is located in the PSORS1C1 gene. Outside the MHC region, our GWAS analysis revealed 7 top SNPs (P<10(-5)) that spanned 6 independent genomic regions. Follow-up of the 17 top SNPs in an independent sample of 1,682 SSc and 3,926 controls showed associations at PSORS1C1 (overall P = 5.70×10(-10), OR:1.25), TNIP1 (P = 4.68×10(-9), OR:1.31), and RHOB loci (P = 3.17×10(-6), OR:1.21). Because of its biological relevance, and previous reports of genetic association at this locus with connective tissue disorders, we investigated TNIP1 expression. A markedly reduced expression of the TNIP1 gene and also its protein product were observed both in lesional skin tissue and in cultured dermal fibroblasts from SSc patients. Furthermore, TNIP1 showed in vitro inhibitory effects on inflammatory cytokine-induced collagen production. The genetic signal of association with TNIP1 variants, together with tissular and cellular investigations, suggests that this pathway has a critical role in regulating autoimmunity and SSc pathogenesis.

Clinical and immunological characterisation of patients with common variable immunodeficiency related immune thrombocytopenia.

Primary Immune thrombocytopenia (ITP) is an autoimmune disease. Secondary ITP occurs in patients with underlying diseases such as common variable immunodeficiency (CVID). CVID is one of the most common symptomatic primary immunodeficiencies in adults, characterised by infectious and non-infectious symptoms. Amongst CVID patients, ITP is the most frequent autoimmune manifestation. In this single-centre study, we performed a clinical and immunological characterisation of 20 patients with CVID-related ITP and 20 ITP patients without CVID to compare severity and remission rates. We found that patients with CVID-related ITP had a higher WHO Bleeding Scale at initial diagnosis yet showed higher remission rates and required less treatment. Patients with ITP needed up to seven therapy options and were often treated with second-line drug therapy, whilst only one CVID-related ITP patient required second-line drug therapy. Therefore, we show that the course of thrombocytopenia in patients with CVID-related ITP is milder. Furthermore, we show that soluble interleukin-2 receptor (sIL-2R, CD25) was higher in CVID-related ITP compared to ITP patients and could accurately classify patient cohorts with an Area Under the Receiver Operating Characteristic of 0.92. Whilst none of the ITP patients had a history of immunodeficiency, we found immunological abnormalities in 12 out of 18 patients. Therefore, we recommend screening ITP patients for CVID and other immunodeficiencies to detect immune abnormalities early, as we found patients with reduced immunoglobulin levels as well as severe lymphocytopenia in our ITP cohort.

Drug-induced immune thrombocytopaenia: results from the Berlin Case-Control Surveillance Study.

PURPOSE: Drug-induced immune thrombocytopaenia is a rare, serious condition that can be triggered by numerous medications. To characterize the spectrum of drugs associated with immune thrombocytopaenia (ITP) in the Berlin Case-Control Surveillance Study (FAKOS). METHODS: Adult hospitalized patients with new onset idiopathic, secondary or drug-induced acute ITP and hospital control patients were ascertained by active surveillance in 50 Berlin hospitals (>180 clinical departments) between 2000 and 2009. Drug exposures were obtained in a personal interview. Chronic cases were excluded in a follow-up after 6 or more months. A standardized causality assessment was conducted for each ITP patient to assess possible drug aetiology. Drug risks were quantified in a case-control design with unconditional logistic regression analysis. RESULTS: Ninety out of 169 validated cases of acute ITP were assessed as being at least possibly drug-related (n= 85 different drugs overall, n = 30 drugs with certain or probable causality). Drugs involved in ≥ 2 cases with a probable or certain relationship were tirofiban (n = 10 cases), abciximab (n = 4), trimethoprim/sulphamethoxazole (n = 4), influenza vaccine (n = 3), and citalopram (n = 2). Pneumococcal and poliomyelitis vaccine were assessed as probably causing ITP in one case each. In the case-control analyses, significantly increased risks were observed for tirofiban, abciximab, trimethoprim/sulphamethoxazole, gentamicin, triamterene/hydrochlorothiazide, drospirenone/ethinylestradiol, and influenza vaccination. CONCLUSIONS: Our study confirms known ITP risks for glycoprotein IIb/IIIa receptor antagonists and sulphonamides and generates signals for several other drugs and vaccines. New onset of ITP should not only direct attention to drugs as possible aetiological agents, but also to vaccines that are known to cause autoimmune phenomena.

Platelet Kinetics in Idiopathic Thrombocytopenic Purpura Patients Treated with Thrombopoietin Receptor Agonists.

AIM: Thrombopoietin receptor agonists (Tpo RA) increase platelet counts in the majority of chronic autoimmune thrombocytopenia (idiopathic thrombocytopenic purpura; ITP) patients. It is unknown whether this treatment may also improve platelet survival (PS) in these patients. METHODS: In order to determine platelet survival (PS), autologous platelets were labeled with (111)In oxine and retransfused in six patients under treatment with Tpo RA (romiplostim n = 3; eltrombopag n = 3). RESULTS: Stable platelet counts of greater than 100 × 10(3)/µl were observed in all 6 patients. Platelet survival was decreased in all cases (mean 2.10 days; range 0.13-3.73 days). No correlation was found between platelet count and PS. Similarly, there was no significant relationship between platelet turnover and platelet count. However, a high platelet turnover, exceeding 25 or three times the norm was observed in 2 patients who presented the lowest PS (0.13 or 0.83 days). Two patients had a moderately shortened PS (1.91 or 2.42 days), and, correspondingly, a moderately increased platelet turnover rate (63,072 or 72,872 platelets/µl/day). CONCLUSION: These results indicate that Tpo RA may not only overcompensate platelet destruction in ITP, but may interfere with other mechanisms, which, in some cases, results in a reduced platelet destruction rate.

Influence of Intravenous Immunoglobulin Treatment on Thrombopoiesis.

AIM: The mechanisms by which intravenous immunoglobulins (IVIg) result in an increase in platelet counts in most patients with autoimmune thrombocytopenia (ITP) have not yet been fully explained. One of these mechanisms may be related to stimulation of thrombopoiesis. METHODS: A total of 13 adult patients who received IVIg were studied: 11 patients with primary ITP, 1 patient with ITP related to common variable immunodeficiency (CVID), and 1 patient with uncharacterized thrombocytopenia. IVIg (0.5-1.5 g/kg body weight) was administered on consecutive days (days 1-3). Endogenous thrombopoietin (eTPO) was measured prior to and at least 1 day following treatment. In addition, IL-6 was measured in 5 of the treated patients. RESULTS: In 10 of 13 patients, IVIg treatment resulted in an increase in platelet counts. eTPO remained unchanged or elevated in almost all cases where the platelet count remained low (<100 × 10(3)/µ0. In all cases with normal or increased platelet counts (>100 × 10(3)/µ0, the eTPO concentration decreased. Furthermore, IVIg induced IL-6 synthesis in all 5 examined patients. CONCLUSION: Our data indicate that the induction of eTPO synthesis by IL-6 may be a potential mechanism in which IVIg may stimulate thrombopoiesis. Further studies are required to characterize this mechanism.

Infection-related transient loss of response and other early side effects in chronic ITP during treatment with thrombopoietin mimetics.

BACKGROUND: The new thrombopoietin mimetics (TM) romiplostim (Nplate®) and eltrombopag (Revolade®) have been demonstrated to increase platelet counts in the majority of patients with chronic autoimmune thrombocytopenia (ITP). Platelet counts in chronic ITP may fluctuate significantly due to infections, but it is unknown if this holds true in patients being treated with TM. PATIENTS AND METHODS: 13 adult patients with refractory chronic ITP were treated with the new TM. 10 patients were only treated with eltrombopag, 2 were initially treated with eltrombopag and subsequently switched to romiplostim, and 1 patient was only treated with romiplostim. RESULTS: Ten patients responded to eltrombopag, and 4 of these patients experienced a transient loss of response following infection. Two patients developed bleeding symptoms, with 1 of these patients requiring platelet transfusion. All 3 patients treated with romiplostim responded, and 1 of these patients experienced also a transient loss of response following infections. CONCLUSIONS: Patients receiving treatment with TM may experience adverse platelet count fluctuations during infections.

Treatment patterns in adults with immune thrombocytopenia before, during and after use of thrombopoietin receptor agonists: a longitudinal prescription database study from Germany.

OBJECTIVE: To assess real-world treatment patterns in patients with immune thrombocytopenia (ITP) who received thrombopoietin receptor agonists (TPO-RAs) in Germany. METHODS: This was a longitudinal, retrospective study using anonymized patient-level data (IQVIA healthcare prescription database, covering 82% of German statutory prescriptions). Eligible patients (aged ≥18 years) had received ≥1 TPO-RA prescription (romiplostim/eltrombopag) from July 2016 to June 2019 (treatment duration ≥30 days). ITP medication use was assessed for 18 months prior to, during and for ≥6 months after TPO-RA treatment. RESULTS: A total of 3553 patients (median age 64 years) were included. Median persistence on TPO-RAs was 12 months (range 1-34). In the periods before, during and after TPO-RA treatment, oral corticosteroids were the most commonly used therapy (64.4%, 43.4% and 36.1% of patients, respectively); median cumulative doses across each period were 2521.9, 2000.0 and 2277.8 mg. The median total duration of corticosteroid use before, during and after TPO-RA therapy was 15, 18 and 32 weeks, respectively. The total median cumulative corticosteroid dose was 6799.7 mg. CONCLUSION: We identified a potential overuse of corticosteroids in patients with ITP in Germany. Earlier use of TPO-RA therapy after a short course of corticosteroids could avoid side effects associated with long-term use.

Disease Severity, Fever, Age, and Sex Correlate With SARS-CoV-2 Neutralizing Antibody Responses.

Clinical trials on the use of COVID-19 convalescent plasma remain inconclusive. While data on safety is increasingly available, evidence for efficacy is still sparse. Subgroup analyses hint to a dose-response relationship between convalescent plasma neutralizing antibody levels and mortality. In particular, patients with primary and secondary antibody deficiency might benefit from this approach. However, testing of neutralizing antibodies is limited to specialized biosafety level 3 laboratories and is a time- and labor-intense procedure. In this single center study of 206 COVID-19 convalescent patients, clinical data, results of commercially available ELISA testing of SARS-CoV-2 spike-IgG and -IgA, and levels of neutralizing antibodies, determined by plaque reduction neutralization testing (PRNT), were analyzed. At a medium time point of 58 days after symptom onset, only 12.6% of potential plasma donors showed high levels of neutralizing antibodies (PRNT50 ≥ 1:320). Multivariable proportional odds logistic regression analysis revealed need for hospitalization due to COVID-19 (odds ratio 6.87; p-value 0.0004) and fever (odds ratio 3.00; p-value 0.0001) as leading factors affecting levels of SARS-CoV-2 neutralizing antibody titers in convalescent plasma donors. Using penalized estimation, a predictive proportional odds logistic regression model including the most important variables hospitalization, fever, age, sex, and anosmia or dysgeusia was developed. The predictive discrimination for PRNT50 ≥ 1:320 was reasonably good with AUC: 0.86 (with 95% CI: 0.79-0.92). Combining clinical and ELISA-based pre-screening, assessment of neutralizing antibodies could be spared in 75% of potential donors with a maximal loss of 10% of true positives (PRNT50 ≥ 1:320).

Massive platelet transfusion is a rapidly effective emergency treatment in patients with refractory autoimmune thrombocytopenia.

Patients with refractory autoimmune thrombocytopenia (ITP) may develop life-threatening bleeding that cannot be immediately controlled by drug administration. To date, there have been no studies conducted to evaluate the efficacy of platelet transfusion alone in such cases. Ten patients with refractory ITP and bleeding or a high bleeding risk were consecutively transfused (one unit/30 min) with apheresis platelet concentrates (APC) without the administration of new drugs. The used APCs (average 3-7 units) contained 2.7 x 10(11) (medium) platelets and were leukodepleted (< or = 1 x 10(6) leukocytes/unit). Platelet serology was performed using standard techniques. Platelet transfusion resulted in an increase in the platelet count to 84 - 157 x 10(3)/microl, and the cessation of bleeding in all patients without any serious adverse effects. Although platelet counts gradually decreased within a few days post-transfusion, bleeding was stopped in all cases. These findings indicate that consecutive platelet transfusion using APCs is a rapidly effective emergency treatment in patients with refractory ITP.

Use of Targeted High-Throughput Sequencing for Genetic Classification of Patients with Bleeding Diathesis and Suspected Platelet Disorder.

Inherited platelet disorders (IPD) form a rare and heterogeneous disease entity that is present in about 8% of patients with non-acquired bleeding diathesis. Identification of the defective cellular pathway is an important criterion for stratifying the patient's individual risk profile and for choosing personalized therapeutic options. While costs of high-throughput sequencing technologies have rapidly declined over the last decade, molecular genetic diagnosis of bleeding and platelet disorders is getting more and more suitable within the diagnostic algorithms. In this study, we developed, verified, and evaluated a targeted, panel-based next-generation sequencing approach comprising 59 genes associated with IPD for a cohort of 38 patients with a history of recurrent bleeding episodes and functionally suspected, but so far genetically undefined IPD. DNA samples from five patients with genetically defined IPD with disease-causing variants in WAS , RBM8A , FERMT3 , P2YR12 , and MYH9 served as controls during the validation process. In 40% of 35 patients analyzed, we were able to finally detect 15 variants, eight of which were novel, in 11 genes, ACTN1 , AP3B1 , GFI1B , HPS1 , HPS4 , HPS6 , MPL , MYH9 , TBXA2R , TPM4 , and TUBB1 , and classified them according to current guidelines. Apart from seven variants of uncertain significance in 11% of patients, nine variants were classified as likely pathogenic or pathogenic providing a molecular diagnosis for 26% of patients. This report also emphasizes on potentials and pitfalls of this tool and prospectively proposes its rational implementation within the diagnostic algorithms of IPD.