Background Hemophilia A (HA) and hemophilia B (HB) are rare X-linked bleeding disorders. Hemophilia genotype is important to inform reproductive planning, pregnancy, neonatal management, inhibitor risk, disease severity, and understanding of disease mechanisms. MyLifeOurFuture (MLOF) was formed as a collaboration between the American Thrombosis and Hemostasis Network (ATHN), the National Hemophilia Foundation (NHF), Bloodworks (BW), and Biogen. Together with participating hemophilia treatment centers (HTCs), MLOF provided hemophilia genotype analysis for patients in the U.S. and created a Research Repository.
Methods HTCs contracted through ATHN, enrolled patients, obtained samples, and provided clinical results to patients. ATHN offered provider education, secure infrastructure for clinical data collection, and access for research proposals. NHF educated the bleeding disorders community about the initiative and supported recruitment. Biogen provided scientific collaboration and financial support. BW served as the central genotyping laboratory and houses the research sample repository. Genotyping was performed using a custom next generation sequencing screen followed by confirmation with a second method. Clinical results were returned to providers.
Results 107 HTCs enrolled 11,341 patients for testing: 8976 for HA (6180 males, 2796 females), 2358 for HB (1616 males, 742 females), 3 for HA/HB, 4 for hemophilia NOS. Clinically reportable variants were detected in 98.2%% of male HA and 98.1% of male HB patients. 1919 unique variants were found, 1486 in F8 and 433 in F9. Of these, 744 were novel (F8 n=610, F9 n=134). Two variants were detected in 95 patients, including 36 females.
In severe HA (n=3419), the most common variants were F8 inversions (44.7%), missense (16.8%), frameshift (16.1%), stop-gain (11.3%), large structural variants (SV) (5.7%), and splice (3.4%). In severe HB (n=564), the most common variants were missense (46.8%), stop-gain (24.1%), frameshift (9.4%), SV (8.3%), and splice (4.1%). Missense variants were the most common variants found in non-severe HA (81.1%) and HB (88.2%).
Inhibitor information is reported for 6986 MLOF patients (HA n=5583; HB n=1403) in the ATHNdataset. Inhibitors were more common in severe disease than non-severe disease in HA [29.8% (n=950/3193) vs. 9.0% (n=216/2390)] and HB [12.4% (n=63/508) vs. 1.9% (n=17/895)].
In severe HA, inhibitors were reported in ~50% of patients with large deletions (n=77/80), complex intron 22 inversions (n=9/17), or no variant found (n=7/14). Other gene-disrupting genotypes had intermediate inhibitor rates (25-36%): intron 1 inversions, intron 22 type 1 and type 2 inversions, stop-gain, splice, and frameshift. Lower rates (~7-14%) were reported with in-frame insertion-deletions, missense, and large duplications. In severe hemophilia B, inhibitors were most common with large deletions (57%, n=24/42).
Conclusions We here report our analysis of the complete dataset of MLOF, the largest hemophilia genetics project performed to date. Clinically reportable DNA variants were identified in nearly all patients. Our findings support the need for comprehensive gene sequencing and SV detection. Deletions, complex inversions, and "No variant found" were the highest risk genotypes for inhibitors in severe hemophilia. The incidence of discovery of novel variation was high (38%) and continued throughout the study, indicating additional variation remains undiscovered. In summary, MLOF has been a successful nationwide collaboration to genotype two rare bleeding disorders at a scale not previously done. This effort has contributed significantly towards identifying and better understanding DNA variation in the F8 and F9 genes in hemophilia.
Johnsen:Octapharma: Research Funding. Pierce:Voyager Therapeutics: Membership on an entity's Board of Directors or advisory committees; CRISPR Therapeutics: Consultancy; Decibel Therapeutics: Consultancy; Third Rock Ventures: Consultancy; Takeda: Consultancy; Geneception: Consultancy; Generation Bio: Consultancy; Global Blood Therapeutics: Membership on an entity's Board of Directors or advisory committees; BioMarin: Consultancy; Ambys Medicines: Consultancy; World Federation of Hemophilia.: Membership on an entity's Board of Directors or advisory committees. Recht:Genentech: Consultancy, Other: personal fees, Research Funding; CSL Behring: Consultancy, Other: personal fees; uniQure: Consultancy, Other: personal fees, Research Funding; Novo Nordisk: Consultancy, Other: personal fees, Research Funding; Spark: Research Funding; Takeda: Consultancy, Other: personal fees, Research Funding; BioMarin: Research Funding; Pfizer: Consultancy, Other: personal fees. Konkle:BioMarin: Consultancy; Sanofi: Consultancy, Research Funding; Sigilon: Consultancy, Research Funding; Pfizer: Consultancy, Research Funding; Baxalta: Research Funding; Spark: Consultancy, Research Funding; Takeda: Research Funding; Uniquire: Research Funding; CSL Behring: Consultancy; Roche: Consultancy.
Asterisk with author names denotes non-ASH members.