It has been suggested that 1–2% of acute promyelocytic leukemia (APL) patients present variant rearrangements of retinoic acid receptor alpha (RARα) fusion gene, with the promyelocytic leukaemia zinc finger (PLZF)/RAR_α being the most frequent. Resistance to all-trans-retinoic acid (ATRA) and arsenic trioxide (ATO) has been suggested in PLZF/RAR_α and other variant APLs. Herein, we analyze the incidence, characteristics, and outcomes of variant APLs reported to the multinational PETHEMA (Programa para el Tratamiento de Hemopatias Malignas) registry, and we perform a systematic review in order to shed light on strategies to improve management of these extremely rare diseases. Of 2895 patients with genetically confirmed APL in the PETHEMA registry, 11 had variant APL (0.4%) (9 PLZF-RAR_α and 2 NPM1-RAR_α), 9 were men, with median age of 44.6 years (3 months to 76 years), median leucocytes (WBC) 16.8 × 10⁹/L, and frequent coagulopathy. Eight patients were treated with ATRA plus chemotherapy-based regimens, and 3 with chemotherapy-based. As compared to previous reports, complete remission and survival was slightly better in our cohort, with 73% complete remission (CR) and 73% survival despite a high relapse rate (43%). After analyzing our series and performing a comprehensive and critical review of the literature, strong recommendations on appropriate management of variant APL are not possible due to the low number and heterogeneity of patients reported so far.

NPM1-mutated acute myeloid leukemia (AML) is genetically well-defined, but clinical outcomes remain heterogeneous, suggesting that quantitative clonal features may refine current risk stratification. We analyzed 688 intensively treated NPM1-mutated AML integrating variant allele frequency (VAF), mutation order, and clonal architecture inferred by PyClone and ClonEvol. Co-mutations were present in 97% of patients (median = 3 per case), dominated by DNMT3A (49%), FLT3-ITD (46%), TET2 (22%), and IDH2 (20%). Prognostic modelling of NPM1 VAF identified an optimal cut-off of 31.44%, defining NPM1high and NPM1low groups. NPM1low correlated with splicing-related alterations and independently predicted inferior overall (HR = 1.46; p = 0.037) and relapse-free survival (HR = 1.40; p = 0.036). Gene-specific VAF analyses revealed divergent effects across partners, high DNMT3A, FLT3-OTHER, KRAS, and PTPN11 burdens were adverse, whereas high IDH2 VAF was protective. Combined models showed that patients with NPM1high and favorable co-mutation VAFs had the best outcomes, while dual unfavorable burdens conferred the poorest survival. Mutation ordering inferred from VAFs positioned NPM1 after epigenetic and splicing lesions but before signaling and transcription-factor mutations. Non-canonical orders, such as early FLT3-OTHER/TKD or WT1 prior to NPM1, significantly stratified outcomes. Clonal reconstruction revealed predominantly linear evolutionary trajectories (84.3%), with increased mutational burden and clonal diversity associating with inferior survival. Notably, intra-clonal co-localization of NPM1 with IDH1 or TET2 was associated with improved outcomes, whereas co-localization with WT1 predicted dismal prognosis. These results demonstrate that quantitative and structural dimensions of clonality refine the biological and prognostic landscape of NPM1-mutated AML beyond mutational status alone.

RAS mutations are recurrent in acute myeloid leukemia (AML); yet, their genomic and clinical landscape remain poorly characterized. We analyzed centralized molecular profiling data in 8,553 patients with newly diagnosed AML. Of these, 1,775 (21%) harbored RAS mutations: NRAS in 1,103 (13%), KRAS in 445 (5%), and both in 227 (3%). RAS mutations primarily clustered in P-loop hotspots (G12/G13, 70%), with NRAS showing higher Q61 mutation prevalence (24% vs. 9%) and KRAS showing additional K117/A146 hotspots. Multiple RAS mutations occurred in 23% of patients. Convergent evolution of RAS signaling mutations emerged as a key feature of RASmut AML. RTK/RAS pathway mutations were non-uniformly enriched: PTPN11, KIT and FLT3-TKD mutations were positively associated, CBL mutations showed no association, and FLT3-ITD mutations were negatively associated. The prognostic impact of RAS mutations was dependent on the genomic background and treatment. In multivariate analyses in intensively treated patients, NRAS mutations showed opposing associations with overall survival (OS): adverse in patients with AML, myelodysplasia-related (HR=1.32, p=0.036) and favorable in those with NPM1 (HR=0.61, p=0.036) or FLT3-ITD (HR=0.63, p=0.048) mutations, among others. KRAS mutations predicted inferior OS (HR=1.41; p=0.003). In patients treated with non-intensive, venetoclax-based regimens, RAS mutations were associated with markedly inferior OS in patients with NPM1 (HR=2.98, p=0.007) or IDH1/2 mutations (HR=4.06, p<0.001). These data inform risk stratification and development of RAS-targeting strategies in AML.

Mutations in FLT3 are present in approximately 30% of patients with AML. The addition of midostaurin (MIDO) to intensive chemotherapy (IC) became standard of care following the RATIFY trial, but comprehensive real‐world data spanning the full adult age spectrum and including both FLT3 ‐ITD and FLT3 ‐TKD mutations remain limited. We retrospectively analyzed 1658 adults aged 14–85 years with newly diagnosed FLT3 ‐mutated AML from 129 PETHEMA registry centers: 469 received IC + MIDO and 1189 IC alone. Composite complete remission was higher with IC + MIDO than IC (81.4% vs. 71.7%; p < 0.001) and Day 30 mortality was substantially lower (2.1% vs. 7.1%; p < 0.001). Median overall survival was 47.2 versus 19.3 months (HR 0.64; 95% CI, 0.53–0.76; p < 0.001), and the benefit was sustained after multivariable adjustment (HR 0.73; p = 0.017). In 261 propensity score–matched pairs, the effect was attenuated, remaining significant for event‐free survival (HR 0.77; p = 0.029) and showing a nonsignificant trend for OS (HR 0.77; p = 0.06). Allogeneic hematopoietic stem cell transplantation in first remission was more frequent in the IC + MIDO cohort (48.9% vs. 41.3%; p = 0.023). Time‐dependent analyses showed the largest MIDO effect among autologous and non‐transplanted patients (OS HR 0.45; p = 0.138, and HR 0.69; p = 0.014, respectively). The benefit of MIDO was consistent irrespective of FLT3 mutation type, FLT3 ‐ITD allelic burden, cytogenetic risk, and gender, while less improvement occurred among NPM1 wild type and secondary AML patients. This large real‐world cohort confirms the survival benefit of MIDO plus IC across the full adult age spectrum, supporting its standard‐of‐care status in FLT3 ‐mutated AML.