Mutations in CCAAT/enhancer binding protein α (CEBPA) occur in 5–10% of cases of acute myeloid leukemia. CEBPA-double-mutated cases usually bear biallelic N- and C-terminal mutations and are associated with a favorable clinical outcome. Identification of CEBPA mutants is challenging because of the variety of mutations, intrinsic characteristics of the gene and technical issues. Several screening methods (fragment-length analysis, gene expression array) have been proposed especially for large-scale clinical use; although efficient, they are limited by specific concerns. We investigated the phenotypic profile of blast and maturing bone marrow cell compartments at diagnosis in 251 cases of acute myeloid leukemia. In this cohort, 16 (6.4%) patients had two CEBPA mutations, whereas ten (4.0%) had a single mutation. First, we highlighted that the CEBPA-double-mutated subset displays recurrent phenotypic abnormalities in all cell compartments. By mutational analysis after cell sorting, we demonstrated that this common phenotypic signature depends on CEBPA-double-mutated multi-lineage involvement. From a multidimensional study of phenotypic data, we developed a classifier including ten core and widely available parameters. The selected markers on blasts (CD34, CD117, CD7, CD15, CD65), neutrophil (SSC, CD64), monocytic (CD14, CD64) and erythroid (CD117) compartments were able to cluster CEBPA-double-mutated cases. In a validation set of 259 AML cases from three independent centers, our classifier showed excellent performance with 100% specificity and 100% sensitivity. We have, therefore, established a reliable screening method, based upon multidimensional analysis of widely available phenotypic parameters. This method provides early results and is suitable for large-scale detection of CEBPA-double-mutated status, allowing gene sequencing to be focused in selected cases.

Relapsed/refractory (R/R) acute myeloid leukemia (AML) is a largely unmet medical need, owing to the lack of standardized, effective treatment approaches, resulting in an overall dismal outcome. The only curative option for R/R AML patients is allogeneic hematopoietic stem cell transplantation (HSCT) which is only applicable in a fraction of patients due to the scarce efficacy and high toxicity of salvage regimens. Recently, a number of targeted agents with relatively favorable toxicity profiles have been explored in clinical trials for R/R AML patients. The Bcl-2 inhibitor venetoclax, in combination with hypomethylating agents or low dose cytarabine, has produced impressive results for newly diagnosed AML, while its role in R/R disease is not well defined yet. We retrospectively analyzed the clinical outcomes of 47 R/R AML patients treated with venetoclax-based regimens between March 2018 and December 2020 at our institution. Overall, we report a composite complete response rate of 55% with an overall acceptable toxicity profile. Outcomes were particularly favorable for NPM1 mutated patients, unlike for FLT3-ITD positive patients irrespective of NPM1 status. For patients treated with intention to transplant, the procedure could be finally performed in 54%. These findings suggest a role for venetoclax-based regimens in R/R AML patients and support the design of prospective studies.

The aim of this study was to measure and identify the reactive carbonyl species (RCSs) released in the blood of humans subjected to hepatic resection. Pre-anesthesia malondialdehyde (MDA) plasma content (0.36 ± 0.11 nmol/mg protein) remained almost unchanged immediately after anaesthesia, before clamping and at the 10th min after ischemia, while markedly increased (to 0.59 ± 0.07 nmol/mg; p < 0.01, Tukey's post test) at the 10th min of reperfusion. A similar trend was observed for the protein carbonyls (PCs), whose pre-anesthesia levels (0.17 ± 0.13 nmol/mg) did not significantly change during ischemia, while increased more than fourfold at the 10th min of reperfusion (0.75 ± 0.17 nmol/mg; p < 0.01, Tukey's post test). RCSs were then identified as covalent adducts to the albumin Cys34, which we previously found as the most reactive protein nucleophilic site in plasma. By using a mass spectrometry (MS) approach based on precursor ion scanning, we found that acrolein (ACR) is the main RCS adducted to albumin Cys34. In basal conditions, the adducted albumin was 0.6 ± 0.4% of the native form but it increased by almost fourfold at the 10th min of reperfusion (2.3 ± 0.7%; p < 0.01, t-test analysis). Since RCSs are damaging molecules, we propose that RCSs, and ACR in particular, are new targets for novel molecular treatments aimed at reducing the ischemia/reperfusion damage by the use of RCS sequestering agents. ARTICLE HISTORY

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