Opposing Pathogenesis of Germline SAMD9/SAMD9L Variants in Adult Myelodysplastic Syndrome (MDS)

Familial myelodysplastic syndromes (MDS), leukemias, and bone marrow failure (BMF) syndromes, typically present in children and young adults. Such cases associate with germ line (GL) mutations in DDX41, RUNX1,ETV6, GATA2 and ANKRD26 and were recently designated by WHO to constitute a separate disease. Gain-of-function (GOF) GL mutations in SAMD9 and SAMD9L associate with familial pediatric myelodysplastic syndrome (MDS) cases that often have early onsets and -7/del(7q). It was not known if this was the case in sporadic adult MDS. With this question in mind, we compiled sequencing results on 799 adults with presumably acquired MDS and BMFs, and further enhanced these results through public WES data on 349 myeloid neoplasms. We also performed cell growth assays of exemplary SAMD9 and SAMD9L GL variants identified in adult MDS.

We used Broad Institute's Genome Aggregation Database (gnomAD) as healthy controls (n = 138,632). After comparing ethnicities between gnomAD and our cohort, we matched healthy controls to patients by ethnicities. Defining rare variants as those present in <0.1% of ethnically-matched healthy controls, we found 26 rare SAMD9/SAMD9L GL variants in 3% (24/799) patients. Most were missense; 3 were truncating (nonsense or frameshift All variants were heterozygous except two patients with two variants. We further validated these data were validated in public WES databases. We identified 2% (7/349) of adult MDS patients in public database with GL SAMD9/SAMD9L variants. SAMD9/SAMD9L GL variants were located in the N-terminal significantly more in adult MDS than reported for pediatric MDS [SAMD9; 62% (8/13) vs. 17% (5/30), p=.044, SAMD9L; 62% (7/13) vs. 0% (0/30), p=.0026]. Compared to reported pediatric MDS patients, adult MDS patients had significantly less deletions of chromosome7/del(7q) [8% (2/24) vs. 69% (22/32), p=.0037] and more 5q deletions [33% (8/24) vs. 4% (1/24), p=.022].

Conservative predictions for specific SAMD9 missense variants were made by using SAMD9 GL variants reported in previous papers or public databases of registered healthy donors, and comparing Area Under the Curve of 8 different prediction algorithms. It defined 11 missense variants as "pathogenic". 57% (4/7) of the variants identified in public WES database were also likely pathogenic by our criteria. We evaluated the growth activity of these 11 unique pathogenic missense variants and 1 frameshift variant identified in a patient with -7. Compared to WT-SAMD9, Thr205Pro-SAMD9, Ile247Thr-SAMD9, and Leu574Pro-SAMD9 showed more proliferative cell growth, whereas, I268T-SAMD9 and D550V-SAMD9L had no significant differences compared to the WT. Compared to WT-SAMD9L, 6 variants (Glu220Gly, Leu1323fs, Cys228Tyr, Trp517Arg, Gly235Ser, and Trp333Cys), fused SAMD9L separately, also had higher proliferation rates than WT-SAMD9L. In total, 9 out of 13 variants (69%) cell growth was not suppressed compared to wild-type cells. Pediatric MDS variants showing less cell growth than the wild type cells suggests that they enhance the normal function of SAMD9/SAMD9L, which is to slow cell cycle progression. In this sense they are gain of function mutations (GOF). In contrast, the majority of variants identified in adult MDS were loss-of-function (LOF) mutations.

Thus, given the different distribution of variants, lack of genetic reversion, and opposing functional results, we conclude that GL SAMD9/SAMD9L variants might have different consequences in familial childhood vs. adult disease. Sporadic adult MDS patients lacked early onset and genetic reversions following acquisition of -7/del(7q) or another mutations in cis characterized in pediatric MDS patients. SAMD9/SAMD9L variants could have two conflicting consequences; i) GOF mutations in pediatric patients that result in genetic reversion or MDS associated with aberrations of chromosome 7 with an early onset, or ii) LOF variants resulting in a late development of MDS due to haplo-insufficiency of SAMD9/SAMD9L. Our findings suggest that GL SAMD9/SAMD9L variants may be linked to the pathogenesis of a subset of adult sporadic MDS patients.