June-Won Cheong

Genomic profiling plays a central role in risk stratification and therapeutic decision-making in acute myeloid leukemia (AML), yet the clinical implications of population-specific genomic architectures remain incompletely defined. We conducted a prospective, multicenter study of 603 adults with newly diagnosed AML in Korea, integrating targeted sequencing of 83 recurrently mutated genes with comprehensive clinical annotation across treatment intensities, including allogeneic hematopoietic stem cell transplantation (allo-HSCT). For contextual comparison, genomic profiles were evaluated against the Beat AML cohort. The overall genomic landscape was broadly conserved, supporting shared core disease biology across populations. However, RUNX1::RUNX1T1, CEBPA, GATA2, KIT, and DDX41 mutations were more frequent in the Korean cohort, whereas FLT3 and NPM1 mutations were less common. These differences translated into a distinct distribution of European LeukemiaNet (ELN) 2022 risk categories, with implications for therapeutic stratification. Notably, most DDX41 alterations were germline (3.2%), highlighting the need for systematic germline evaluation with implications for genetic counseling and donor selection. Although unadjusted overall survival appeared longer in the Korean cohort, this difference was not significant after adjustment for key clinical variables. These findings indicate that population-specific genomic distributions reshape the clinical application of risk stratification and support population-aware precision medicine strategies in AML.

Jin Ju Kim, M.D., Yu Ri Kim, M.D., Hongkyung Kim, M.D., Hye Won Kook, M.D., Haerim Chung, M.D., Doh Yu Hwang, M.D., Ji Eun Jang, M.D., June-Won Cheong, M.D., Seung-Tae Lee, M.D., Jong Rak Choi, M.D., Saeam Shin, M.D. , and Jin Seok Kim, M.D.. Ann Lab Med -0001;0:. https://doi.org/10.3343/alm.2026.0122

Background/Objectives: Myeloproliferative neoplasms (MPNs) are clonal hematopoietic stem cell disorders driven by somatic mutations, most commonly those in JAK2, CALR, and MPL. While conventional molecular testing often focuses on these canonical mutations, emerging data suggest that additional mutations and low-variant-allele-frequency (VAF) subclones may contribute to disease progression and prognosis. Methods: We applied ultra-deep targeted sequencing with an error-correction algorithm to analyze bone marrow and peripheral blood samples from 134 patients with essential thrombocythemia (ET), polycythemia vera (PV), primary myelofibrosis (PMF), secondary myelofibrosis (MF), or post-MPN acute leukemia. Targeted sequencing panels were used to detect disease-associated and clonal driver mutations. Matched germline controls were not systematically available. Results: Low-VAF variants ( 5%, suggesting clonal expansion during progression. Paired sequencing at initial MPN diagnosis and leukemic transformation was available for three patients. JAK2 p.V617F VAFs differed according to treatment timing (Kruskal–Wallis test, p = 0.0138), with lower VAFs observed in untreated patients; however, this association may be influenced by treatment-selection confounding. Mutations in ASXL1 and SRSF2 were occasionally observed in ET and secondary MF, although their prognostic significance remains uncertain. Progression analyses were exploratory because of the limited number of events and longitudinally sampled patients, and formal survival or prognostic modeling was not performed. Conclusions: Ultra-deep error-corrected sequencing enables sensitive detection of low-VAF mutations in MPNs, including subclonal events potentially missed by less sensitive assays. These findings demonstrate the potential utility of high-sensitivity panel-based sequencing for molecular characterization of MPN, while larger longitudinal studies are required to establish the prognostic and clinical significance of these findings.

In the phase 3 ECHELON-2 trial, brentuximab vedotin, cyclophosphamide, doxorubicin, and prednisone (BV-CHP) significantly improved progression-free survival (PFS) and overall survival (OS) compared with cyclophosphamide, doxorubicin, vincristine, and prednisone (CHOP) in patients with CD30+ peripheral T-cell lymphoma, benefits that were maintained at 5 years. Interim positron emission tomography (PET) scan can be used to assess prognosis and risk-stratify patients. The prognostic value of interim PET was assessed in this post hoc exploratory analysis from ECHELON-2, evaluating interim 18F-fluorodeoxyglucose PET scans after cycle 4 (PET4) and end-of-treatment–based response, and correlated with PFS per investigator and OS. PET4 response was determined by Deauville score (scores of 1-3 were considered negative [PET4-negative] and 4-5 positive [PET4-positive]) by independent review. Overall, 452 patients were randomized 1:1 to the BV-CHP (n = 226) and CHOP (n = 226) arms. Of these, 32 in the BV-CHP arm and 41 in the CHOP arm were not evaluable for PET4. In both arms, PET4-negative status was associated with improved PFS (BV-CHP: HR, 0.36; 95% CI, 0.19-0.66; CHOP: HR, 0.26; 95% CI, 0.17-0.41) and OS (BV-CHP: HR, 0.38; 95% CI, 0.18-0.78; CHOP: HR, 0.24; 95% CI, 0.14-0.41) compared with PET4-positive status. Among patients with systemic anaplastic large cell lymphoma, PET4-negative patients had improved PFS (BV-CHP: HR, 0.28; 95% CI, 0.14-0.60; CHOP: HR, 0.31; 95% CI, 0.17-0.56) and OS (BV-CHP: HR, 0.38; 95% CI, 0.16-0.94; CHOP: HR, 0.25; 95% CI, 0.12-0.55) compared with PET4-positive patients. In this exploratory analysis, PET4-negative status by Deauville score was associated with improved long-term PFS and OS in both the BV-CHP and CHOP arms. This trial was registered at www.ClinicalTrials.gov as #NCT01777152.

Despite advances with novel targeted agents (e.g., BCL-2 or IDH inhibitors) combined with chemotherapy for acute myeloid leukemia (AML), drug resistance persists. We investigated whether blocking Na⁺/H⁺ exchanger 1 (NHE1) could enhance AML cell sensitivity to the BCL-2 inhibitor venetoclax and sought to determine the molecular mechanisms. Our results demonstrated that co-treatment with venetoclax and the NHE1 inhibitor 5-(N,N-hexamethylene) amiloride (HMA) synergistically induced apoptosis in both venetoclax-sensitive and -resistant leukemic cell lines. Specifically, the combination significantly increased apoptosis in venetoclax-resistant THP-1 cells to 72.28% (17.79% with 100 nM venetoclax and 10.15% with 10 μM HMA alone; p < 0.001). Conversely, another venetoclax-resistant line, U-937, showed no significant apoptotic response to the combination. In THP-1 cells, this synergy was mediated via a caspase-dependent programmed cell death pathway, evidenced by an increased BAX/BCL-2 ratio, mitochondrial cytochrome c release, and subsequent caspase-9 and caspase-3 activation. Furthermore, co-treatment downregulated the anti-apoptotic protein MCL-1 and reduced PI3K and Akt phosphorylation, suggesting that inhibition of these survival pathways also contributed to the synergistic effect. Inhibition of NHE1 may substantially enhance venetoclax sensitivity in certain AML models, particularly in venetoclax-resistant THP-1 cells but not in U-937, highlighting biological diversity and the probable involvement of alternative survival pathways.

Tracking transfused platelets is important to evaluate platelet transfusion efficiency. Traditionally, corrected count increments were used; however, quantitative polymerase chain reaction-based methods have recently been developed. As both these methods have some limitations, we developed a new method based on next-generation sequencing (NGS) of platelet mitochondrial DNA (mtDNA). We identified several single nucleotide variant markers by sequencing the entire mtDNA region of platelets, and used NGS to estimate the proportion of each platelet unit. This method was validated using mixed platelets obtained from different donors at various ratios. We confirmed the applicability of this method in patients who received platelet transfusions using pre- and posttransfusion samples from 12 patients with hematological malignancies. The method showed good linearity (r2 >0.99 in the range of mixing ratios from 1:1 to 1:50) in the platelet-mixing experiment. In addition, platelet tracking in patients who received transfusions was feasible using this method. Furthermore, it was possible to track individual platelets in patients who received a single platelet transfusion and in those who received multiple transfusions, including a patient who received 5 platelet transfusions. Hence, this NGS-based platelet-tracking method can be used for patients with various conditions.

Purpose: Allogeneic hematopoietic stem cell transplantation remains a curative option for acute leukemia. While an adequate CD34⁺ cell dose is essential for engraftment, the optimal upper threshold in haploidentical peripheral blood stem cell transplantation (haplo-PBSCT) remains unclear. Methods: We retrospectively analyzed 81 patients with acute leukemia who underwent haplo-PBSCT with reduced-intensity conditioning between 2010 and 2020. Patients were stratified by CD34⁺ cell dose (< 8 × 10⁶/kg vs. ≥ 8 × 10⁶/kg). Clinical outcomes, including overall survival (OS), non-relapse mortality (NRM), graft failure, and graft-versus-host disease (GVHD) incidence, were compared. Results: A higher CD34⁺ cell dose was associated with inferior OS (P = 0.022) and increased NRM (P = 0.002), despite similar rates of graft failure and acute GVHD. Chronic GVHD was more frequent in the higher dose group, though the difference was not statistically significant. Multivariate Cox analysis confirmed a high CD34⁺ cell dose as an independent predictor of poor OS (HR 2.054, P = 0.031). Conclusion: These findings suggest that excessively high doses may adversely affect survival by increasing transplant-related toxicity. Graft cell dose should be carefully balanced to optimize outcomes in haplo-PBSCT.

Emerging evidence indicates that CD4+ T cells contribute to antitumor immunity beyond their traditional roles as helpers or regulators. However, the specific subset of CD4+ T cells mediating beneficial outcomes in patients with multiple myeloma remains unclear. Here, we performed single-cell RNA sequencing and T-cell receptor sequencing on CD4+ T cells sorted from the bone marrow of patients across the stages of myeloma progression. We identified several distinct states of CD4+ cytotoxic T lymphocytes (CTLs) that were significantly increased and clonally expanded in patients with myeloma. CD4+ CTLs displayed transcriptional and phenotypic characteristics indicative of cytotoxicity, demonstrating their ability to directly kill myeloma cells. This cytotoxicity, however, was abrogated by NKG2D blockade. Notably, the abundance of NKG2D+CD4+ CTLs correlated with improved survival in patients with myeloma. Our findings suggest that harnessing CD4+ CTLs could lead to novel strategies for enhancing immunotherapy outcomes in multiple myeloma.