Hemostasis is impaired during CABG and coagulation abnormalities often result in clinically relevant organ dysfunctions, eventually increasing morbidity and mortality rates. Fifteen consecutive patients with coronary artery disease submitted to conventional extracorporeal circulation (cECC) have been compared with 15 matched patients, using mini-ECC (MECC). Postoperative lung function was evaluated according to gas exchange, intubation time and lung injury score. In the MECC group, thrombin-antithrombin complex levels (TaTc), prothrombin fragments (PF1+2) formation and thromboelastography (TEG) clotting times were lower compared to the cECC group (p=0.002 and p<0.001, respectively) whereas postoperative blood loss was higher in the cECC group (p=0.030) and more patients required blood transfusion (p=0.020). In the MECC group, postoperative gas exchange values were better, intubation time shorter and lung injury score lower (p<0.001 for all comparisons). Our study suggests that MECC induces less coagulation disorders, leading to lower postoperative blood loss and better postoperative lung function. This approach may be advantageous in high-risk patients.
Summary FLT3 internal tandem duplication (ITD) mutations are frequently detected at diagnosis in cytogenetically normal acute myeloid leukaemia (CN‐AML) and predict unfavourable outcome. FLT3 ITD is an unstable aberration and may be lost or acquired at relapse. Recent whole genome sequencing studies have suggested that FLT3 ITD + ve AML relapse may evolve from small subclones undetectable at diagnosis by routine polymerase chain reaction (PCR). We developed a patient‐specific real‐time quantitative‐PCR (RQ‐PCR) to implement FLT3 ITD detection in six AML patients whose blasts carried wild‐type FLT3 at diagnosis and who relapsed with FLT3 ITD by routine PCR. Patient‐specific forward primers were designed after cloning and sequencing the FLT3 ITD in each case. The assay allowed retrospective detection of FLT3 ITD in diagnostic samples of 4/6 cases and to establish the kinetics of clonal evolution preceding relapse. After conventional chemotherapy, all patients had early relapse despite having been classified as NPM1 + ve/ FLT3 ITD − ve at presentation, with shorter remissions being observed in four patients re‐classified as FLT3 ITD + ve by the new assay. Notably, FLT3 ITD clone became detectable by conventional PCR in three patients tested during remission after initial treatment. Our data underscore the need of identifying low FLT3 ITD levels, which are probably associated with relapse in otherwise good prognosis CN‐AML.
Background: Extramedullary acute myeloid leukemia (EM-AML) is a rare manifestation of AML, associated with poor prognosis and shorter survival. Unlike solid tumors, the metastatic behaviors of blasts and the molecular mechanisms of EM tissue invasion by myeloid blasts have not been well characterized. Aims: To elucidate the molecular mechanisms underlying leukemia infiltration, we analyzed, by RNA sequencing, the expression profile of EM-AMLs localizations, as compared to that of paired BM blasts, isolated at the time of diagnosis or of relapse. The final goal was to identify predictive factors of EM-migration of AML cells, and possible druggable targets to be used to prevent or treat this prognostically adverse leukemic manifestation. Methods: Twelve patients (median age 60 years, range 53-75) affected by AML who developed an extramedullary AML localization at onset (N=5) or relapse (N=7), diagnosed at the Oncohematology Unit of Policlinico Tor Vergata in Rome, during the years 2019-2022, were included in the study. Paired samples of bone marrow (BM) and sarcoma tissue at the time of diagnosis and/or relapse were available in 7 EM-AML cases. For RNA-seq analysis, libraries were sequenced on the Illumina-platform with paired-end reads 150bp long, with a depth of 27 million cluster/sample. Statistical significance threshold for differentially expressed genes was set with an adjusted p-value < 0.1, and |log2(Fold Change)|≥ 2 to identify differentially expressed genes (DEGs) and pathways. These were then validated by q-RT-PCR. Functional experiments were then performed using siRNA and Metformin treatment, according to standard methods. Results: After comprehensive analysis of RNA-Seq data, the intersection of DAVID (v. 2021) and GSEA software (v. 3.0) output showed deregulation of the ECM-receptor interaction and Focal adhesion pathways as the most significantly enriched in EM sites, as compared to BM. Among identified pathways, 32 upregulated DEGs were common to both GSEA and DAVID analysis (Figure 1). Upregulation of these selected genes was confirmed by qRT-PCR in an independent EM-AML prospective cohort (n=3 pts). No differences in expression of cancer invasion pathway related genes were observed comparing BM-MNC (mononuclear cells) of EM-AML to that from healthy donors, suggesting that EM-AML cells may acquire specific invasive capabilities. To validate the transcriptome-based model of AML blasts to develop EM localizations, OCI-AML3 cells were transfected with siRNAs against TWIST1, a gene upregulated in EM-AML biopsies, which has an impact on cancer invasion as a result of dysregulation of EMT-related pathways. Forty-eight hours after siRNA transfection, we observed a significant reduction in cell proliferation as compared to non-treated cells. To confirm our model, cells were treated with Metformin, a drug able to repress TWIST1 expression and inhibit cancer invasion in several tumor models. After treatment with 20 mM Metformin for 48h, we confirmed reduction of EMT-related gene expression in OCI-AML3 cells. Finally, transwell assay showed a decreased cell migration and invasion after TWIST1 modulation. Summary/Conclusion: Our analysis suggests an involvement of processes closely associated with neoplastic cell invasion in EM-AML. Addition of Metformin to chemotherapy may help to reduce the development of EM-AML localizations.Keywords: Metastasis, Cell migration, Acute myeloid leukemia, RNA-seq
Preliminary reports have suggested that autologous stem-cell transplantation (ASCT) is feasible in elderly patients with acute myeloid leukemia (AML). The objective of this study was to describe the disease characteristics and treatment results from a series of 22 elderly AML patients undergoing ASCT.The median age was 64 yr (range 61-71). Twenty patients were in first complete remission (CR1), two in CR2, and all were in performance status 0-1. The median interval between CR achievement and ACST was 3 months (range 2-5). In 20 cases peripheral blood stem cells were infused, in two bone marrow.All patients had a successful engrafment. One patient (5%) died from transplant-related complications. The median number of days to granulocytes > 500 mm-3 and platelets > 20 000 mm-3 was 11(range 9-15) and 13 (range 9-20), respectively. Non-hematologic toxicity included WHO grade III-IV stomatitis in 32% patients and grade IV nausea and vomiting in one (4.5%). Seven patients had fever of unknown origin, while in 14 a documented infection was diagnosed. Median duration of hospitalization was 31 d (range 16-60).After a median follow-up of 12 months from ASCT, nine patients are alive in continuous CR and 13 died from AML relapse. Median survival from diagnosis and disease-free survival (DFS) was 19 and 14 months, respectively. Our data show that ASCT with a standard conditioning regimen is feasible in AML patients aged more than 60 yr. Toxicity and hemopoietic recovery do not substantially differ from those observed in young adults. DFS and overall survival (OS) duration are encouraging, but a longer follow up is needed on a larger series of patients.
Background: Following the experience of the GIMEMA AML1310 protocol (Venditti A et al, Blood 2019), in which the choice between autologous (AuSCT) or allogeneic hematopoietic stem cell transplant (ASCT) was risk-adapted (for favorable- and adverse-risk categories) or MRD-driven (for patients in the intermediate-risk category), we designed a next generation, multicenter trial named AML1819 (NCT04168502). This trial recruits young patients (≤ 60 years of age) belonging to the ELN2017 favorable-and intermediate-risk categories, with the exception of cases FLT3 positive and relies on the addition of gemtuzumab ozogamicin (GO) to intensive chemotherapy (Castaigne S et al, Lancet 2012 – Lambert J et al, Haematologica 2019). AML1819 trial, likewise AML1310, is inspired to a MRD-oriented post-remission approach. Aims: The primary outcome measures of AML1819 are to determine (1) the percentage of MRD negativity after consolidation in patients treated in induction and consolidation with GO; (2) the efficacy of a post-transplant maintenance with glasdegib vs clinical observation in terms of disease free survival improvement. The present report illustrates the preliminary results of the post-consolidation MRD analysis. Methods: In AML1819, patients are to receive 1 induction and 1 consolidation with the addition of gemtuzumab ozogamicin (GO) and then, based on the level of post-consolidation MRD, they are submitted to an AuSCT (MRD neg) or ASCT (MRD pos). Following the transplant, the patients are randomized between observation or a maintenance with glasdegib, for 12 months. MRD is assessed by Reverse-Trascriptase quantitative-Polimerase Chain Reaction (RT-qPCR) for patients with a molecular marker (NPM1 mutation or CBF rearrangements) or by multiparametric flow cytometry (MFC) in those lacking a traceable molecular signature (Fig. n.1 Results: Of 171 patients enrolled, 145 patients (85%) are evaluable, median age 53 (18-61), 52% males and 48% females; 76 patients (52%) belonged to the ELN2017 favorable-risk (FR) category and 69 (48%) to the intermediate-risk (IR) one. Of 145 patients, 107 (74%) achieved a CR/CRi. In the FR and IR category, 63 of 76 (83%) and 44 of 69 (64%) achieved CR/CRi, respectively. Of 107 patients, 105 (98%) received the consolidation course and 96 (91%) [58 FR (60%) and 38 IR (40%)] are evaluable for post-consolidation MRD assessment. Overall, 74% (71) of 96 were MRD neg and 26% (25) MRD pos. When the analysis was split as per risk category, 81% (47) of 58 patients with FR AML became MRD neg after consolidation and 19% (11) were MRD pos. In the IR category, of 38 patients 63% (24) achieved a MRD neg status whereas 37% (14) remained MRD pos. The post-consolidation frequency of MRD negativity for patients belonging to the IR category of the AML1310 protocol was 46% (42 patients out of 92). With a median follow-up of 17.4 months, 1-year overall survival of FR and IR patients is 83.1% (SD 69.2%, 99.8%) and 72.1% (53.0%, 98.0%), respectively. Summary/Conclusion: In the AML1819 trial, the preliminary analysis of the MRD status after consolidation indicates that a remarkable proportion of patients become negative when GO is added to intensive chemotherapy. With all the limits of such a comparison, we also found that the proportion of patients being MRD negative after consolidation was higher in AML1819 trial than in AML1310 one, in which no GO was added to chemotherapy. Such a finding has practical implications since, in AML1819 trial, a lower fraction of patients is submitted to ASCT. These figures need confirmation in a more advanced phase of trial development, showing that the high frequency of MRD negativity translates into a survival benefit.Fig. 1. GIMEMA AML1819 Trial Design Keywords: Gemtuzumab ozogamicin, Acute myeloid leukemia, Chemotherapy, Measurable residual disease
