Background Genetic reports of patients with severe AA (SAA) often identify heterozygous variants of unknown significance in recessive genes associated with inherited bone marrow failure syndromes (IBMFS), such as Fanconi anemia (FA) and telomere biology diseases (TBD). More recently, heterozygous SAMD9 and SAMD9L (SAMD9/9L) variants have also been reported in children and young adults with BMF, but whether all variants are gain-of-function (GoF) and pathogenic is unknown. In a large cohort of SAA patients, we investigated the prevalence of heterozygous variants in SAMD9/9L and FA/TBD genes and correlated them to disease characteristics and clinical outcomes.

Methods SAA patients (n= 198; median age [range]: 32 [3-82]) enrolled on a phase II trial for treatment with horse anti-thymocyte globulin, cyclosporine, and eltrombopag (NCT01623167) at the National Institutes of Health from 2012-2022 were included (Table 1). All patients were screened for germline mutations in IBMFS related genes. SAMD9/9L variants were segregated by rarity according to frequency in the gnomAD database as common (< 0.1% in overall population, including variants with maximum population frequency [MaxPopFreq]> 1%), rare (MaxPopFreq< 0.1%), very rare (MaxPopFreq< 0.01%), ultra-rare (MaxPopFreq< 0.005%), and novel (absent in gnomAD).

Results We first investigated whether patients had an undiagnosed SAMD9/9L disorder. Although 40/198 patients (20%) had a heterozygous SAMD9/9L variant, including 8 (20%) with 2 variants, these were mostly classed as common (73%; n= 35), followed by rare, very rare, ultra-rare and novel variants seen in 17%(n= 8), 4%(n= 2), 2%(n= 1), and 4%(n= 2), respectively. Most variants were missense (n= 39; 81%), followed by nonsense (n= 6; 13%) and frameshift mutations (n= 3; 6%). Except for ultra-rare and novel, SAMD9/9L often co-occurred with typical features of immune AA (6p-LOH, PNH clones, HLA and PIGA mutations). None of the 40 patients with germline SAMD9/9L variants had primary somatic genetic reversions (SGR) such as somatic loss of function (LoF) mutations (VAF> 0.5%) in SAMD9/9L or UPD7q. Since GoF SAMD9/9L variants are located on chr7 and they associate with development of monosomy 7, we investigated whether SAMD9/9L variants in our cohort were associated with clonal evolution after treatment. However, only 2/40 patients (5%) with chr7 abnormalities had these variants, both common.