Background Germline DDX41 mutations are the most common genetic predisposition to myeloid neoplasms, representing a distinct subtype associated with favorable outcomes, although evidence highlighting the prognostic relevance of second somatic hits, particularly p.Arg525His, is emerging. Treatment strategies and their outcomes in this setting remain poorly described.

Methods We retrospectively analyzed patients with ≥1 DDX41 variant detected by sequencing at a tertiary center between Jan 1998–Jul 2025. Germline (GL) origin was confirmed by skin biopsy or inferred by VAF (>/<40%), public GL databases, and known hotspots mutations. Clinical, hematological, and molecular data were collected, together with treatment strategies including transplant outcomes.

Results A total of 240 patients with DDX41 variants were detected: 82 were unreported VUS and 8 benign, and were not included in full analysis. Of the remaining 150, all were pathogenic/likely pathogenic except for 9 reported ‘hot’ VUS, with evidence points at least 4-5 following interpretation according to ACMG guidelines. DDX41 variants with probable GL aetiology were found in 123 cases (82%), 87 of which were truncating (stop gain, frameshift indels, splice variants). The most common alterations were p.Met1? (29/87) and p.Asp140GlyfsTer2 (18/87). Germline confirmation was done in 57 (43%). A somatic DDX41 second hit, median VAF 10% (2-49), was present in 85/123 (69%), most commonly p.Arg525His (64%).

Median age was 70 years (26–91) and69% of patients were male;median follow-up was 25.7 months (IQR 12.5–54.5).Diagnoses included AML(66), MDS(63), CCUS(14), and others. MDS risk categories were mainly PSS-R (36%) intermediate, and IPSS-M low (49%). AML was mostly intermediate-risk by ELN 2022 (62%). Baseline marrow aspirate/trephine was hypocellular for age in 37.5%, with median BM blasts 15%(0-96). The latter were higher in carriers with a biallelic acquired DDX41 variant(20% vs 2%, p=0.003). Karyotype was normal in 74.5%. Co-mutations were present in 60% (median 1, 0–5),most frequentlyASXL1 (20), DNMT3A (17)CUX1 (16), enriched in AML;10 (7%) harbored a TP53 mutation.

Overall, 45 patients (35%) died, with a median OS of 61 months. Survival analysis showed that GL missense variants without a biallelic acquired DDX41 variant or with non-p.Arg525His biallelic acquired variant had the best OS (NR and 98.5 months, respectively),while the worst prognosis was observed in patients with truncating GL plus p.Arg525His (mOS 47 months).The same pattern was seen for AML-free survival.

Regarding treatment, 24% of patients had no indication for treatment, 5 received only best supportive care (BSC) and 10 had ESAs. Active treatments included azacitidine (21), venetoclax and azacitidine (VENAZA, 17), high-dose chemotherapy (26), and others (4). Watch & wait was more common in patients without a somatic second-hit (44% vs 19%, p=0.03), whereas high-dose chemo was more frequent with second hit (25% vs 7%, p=0.04). First line overall response rate was 74% for the whole cohort, 66% in MDS (54% complete), 78% in AML, all complete. Treatment-specific response rates were: EPO 20%, azacitidine 62%, VENAZA 50% (18% stopped for toxicity), high-dose chemotherapy 84%. Relapse occurred in 30%; salvage regimens included azacitidine (18%), VEN-based (45%), high-dose chemo (18%), other (13%) with 40% ORR, or BSC (5%).

64 patients (43%) received HMA (alone or in combination) either frontline or at relapse: their ORR was 80%, complete in 70%. Best response occurred at median 2 cycles (1–9); median total cycles was 7 (1–40) and treatment duration 20 months (4–56), mOS for this cohort was 57.6 months. Reasons for discontinuation were relapse (25), transplant (11), toxicity (5), or durable CR (1); 12 patients remain on therapy. HMA was more often used in patients with a biallelic acquired variant (26% vs 7%, p=0.04) and associated with higher response (82% vs 60%, p=0.05). Finally, 24 patients (17%) underwent transplant; 9 relapsed after median 12.4 months (0-46).

Conclusion In this cohort, truncating GL mutations accompanied by somatic p.Arg525His confirmed to be associated with inferior outcomes and advanced disease at presentation. Treatment strategies largely mirrored standard MDS/AML approaches, but demonstrated limited efficacy for EPO and high responses to HMAs and intensive chemotherapy. HMA therapy induced durable responses even in the higher risk group of patients with biallelic acquired DDX41 variants.

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2025
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