Anti-CD38 monoclonal antibody impairs CD34+ mobilization and affects clonogenic potential in multiple myeloma patients.

BACKGROUND: Induction with daratumumab-based regimens followed by autologous stem cell transplantation is the current standard for newly diagnosed multiple myeloma (NDMM) patients eligible for intensive chemotherapy. However, concerns emerged regarding potential negative effects following daratumumab-based treatment on CD34+ mobilization. We here compared CD34+ mobilization and clonogenic potential between daratumumab and non-daratumumab based therapy without upfront plerixafor administration among patients affected by NDMM. MATERIALS AND METHODS: Clinical, mobilization and clonogenic data from 41 consecutively enrolled NDMM patients were analyzed. Patients underwent collection of autologous CD34+ by apheresis at the ASST Grande Ospedale Metropolitano Niguarda, Milan, Italy, from January 2021 to March 2023. Clonogenicity analysis was performed on BFU-E and CFU-GM. RESULTS: Seventy-five percent of daratumumab-treated patients underwent >1 apheresis, compared to 24% of non-daratumumab patients (p=0.0017). Daratumumab-treated patients had significantly lower CD34+ count (mean 38 vs 79/µL, respectively; p=0.0011), with a median CD34+ harvest of 3.98×106/kg (range 1.68-9.18) vs 6.87×106/kg (range 1.63-16.85) in non-daratumumab-treated (p=0.0006). In multivariate analysis the likelihood of undergoing >1 apheresis was significantly higher in older patients (OR 1.2, 95% CI 1-1.4, Z=2.10, p=0.03) and daratumumab-treated patients (OR 15, 95% CI 2.8-129, p=0.004). Moreover, daratumumab-based induction therapy demonstrated an independent negative association with BFU-E colony formation (p=0.0148), even when accounting for patient age and CD34+ levels. DISCUSSION: Our findings underscore the impact of daratumumab-based treatment on CD34+ mobilization in a real-life, upfront plerixafor-free population of NDMM patients. Higher probability of requiring multiple apheresis occurred among daratumumab-treated patients. Interestingly, the observation that daratumumab might negatively impact BFU-E colony formation, independent of CD34+ cell count, offers novel biological perspectives. Appropriate strategies should be adopted by the Apheresis teams to mitigate these potential negative effects.

Clinical Effects of Driver Somatic Mutations on the Outcomes of Patients With Myelodysplastic Syndromes Treated With Allogeneic Hematopoietic Stem-Cell Transplantation.

PURPOSE: The genetic basis of myelodysplastic syndromes (MDS) is heterogeneous, and various combinations of somatic mutations are associated with different clinical phenotypes and outcomes. Whether the genetic basis of MDS influences the outcome of allogeneic hematopoietic stem-cell transplantation (HSCT) is unclear. PATIENTS AND METHODS: We studied 401 patients with MDS or acute myeloid leukemia (AML) evolving from MDS (MDS/AML). We used massively parallel sequencing to examine tumor samples collected before HSCT for somatic mutations in 34 recurrently mutated genes in myeloid neoplasms. We then analyzed the impact of mutations on the outcome of HSCT. RESULTS: Overall, 87% of patients carried one or more oncogenic mutations. Somatic mutations of ASXL1, RUNX1, and TP53 were independent predictors of relapse and overall survival after HSCT in both patients with MDS and patients with MDS/AML (P values ranging from .003 to .035). In patients with MDS/AML, gene ontology (ie, secondary-type AML carrying mutations in genes of RNA splicing machinery, TP53-mutated AML, or de novo AML) was an independent predictor of posttransplantation outcome (P = .013). The impact of ASXL1, RUNX1, and TP53 mutations on posttransplantation survival was independent of the revised International Prognostic Scoring System (IPSS-R). Combining somatic mutations and IPSS-R risk improved the ability to stratify patients by capturing more prognostic information at an individual level. Accounting for various combinations of IPSS-R risk and somatic mutations, the 5-year probability of survival after HSCT ranged from 0% to 73%. CONCLUSION: Somatic mutation in ASXL1, RUNX1, or TP53 is independently associated with unfavorable outcomes and shorter survival after allogeneic HSCT for patients with MDS and MDS/AML. Accounting for these genetic lesions may improve the prognostication precision in clinical practice and in designing clinical trials.

Down-regulation of microRNAs 222/221 in acute myelogenous leukemia with deranged core-binding factor subunits.

Core-binding factor leukemia (CBFL) is a subgroup of acute myeloid leukemia (AML) characterized by genetic mutations involving the subunits of the core-binding factor (CBF). The leukemogenesis model for CBFL posits that one, or more, gene mutations inducing increased cell proliferation and/or inhibition of apoptosis cooperate with CBF mutations for leukemia development. One of the most common mutations associated with CBF mutations involves the KIT receptor. A high expression of KIT is a hallmark of a high proportion of CBFL. Previous studies indicate that microRNA (MIR) 222/221 targets the 3' untranslated region of the KIT messenger RNA and our observation that AML1 can bind the MIR-222/221 promoter, we hypothesized that MIR-222/221 represents the link between CBF and KIT. Here, we show that MIR-222/221 expression is upregulated after myeloid differentiation of normal bone marrow AC133(+) stem progenitor cells. CBFL blasts with either t(8;21) or inv(16) CBF rearrangements with high expression levels of KIT (CD117) display a significantly lower level of MIR-222/221 expression than non-CBFL blasts. Consistently, we found that the t(8;21) AML1-MTG8 fusion protein binds the MIR-222/221 promoter and induces transcriptional repression of a MIR-222/221-LUC reporter. Because of the highly conserved sequence homology, we demonstrated concomitant MIR-222/221 down-regulation and KIT up-regulation in the 32D/WT1 mouse cell model carrying the AML1-MTG16 fusion protein. This study provides the first hint that CBFL-associated fusion proteins may lead to up-regulation of the KIT receptor by down-regulating MIR-222/221, thus explaining the concomitant occurrence of CBF genetic rearrangements and overexpression of wild type or mutant KIT in AML.

Total serum tryptase: a predictive marker for KIT mutation in acute myeloid leukemia.

Human tryptase is a serine protease expressed in mast-cells. We previously observed that AML blast cells, cultured in vitro from a KIT D816Y patient, give rise to adherent cells with mast-cell like phenotype and tryptase was released in the serum-free medium. To correlate total serum tryptase (ts-try) levels with cytogenetic features and KIT mutational status, we analyzed serum samples from AML patients at diagnosis. In 70 out of 155 patients (45%) we detected elevated ts-try (>15 ng/mL), significantly linked to t(8;21) (P < .001) and inv(16) (P = .007). In patients that achieved complete remission the ts-try decreased to normal values. In 75 patients screened for KIT mutation, we found a clear relationship between elevated ts-try and mutated patients with t(8;21) (P < .001). In conclusion, we propose that checking for ts-try at diagnosis of AML may be a simple tool to select patients to be addressed to KIT mutation screening.

Molecular analysis of PDGFRA and PDGFRB genes by rapid single-strand conformation polymorphism (SSCP) in patients with core-binding factor leukaemias with KIT or FLT3 mutation.

BACKGROUND: Mutations involving KIT and FLT3 genes, encoding tyrosine kinase (TK) membrane receptors, are detected in core-binding factor leukaemia (CBFL) patients. PDFGRA and PDGFRB encode class III TK receptors and are involved both in physiological processes and in the pathogenesis of haematological and solid tumours. The aim of this study was to investigate if PDGFR mutations are involved in CBFL. PATIENTS AND METHODS: In order to detect PDGFR mutations in CBFL, 35 patients without KIT or FLT3 mutations patients were screened by rapid and sensitive single-strand conformation polymorphism (SSCP) analysis. Sequence analysis was performed in polymerase chain reaction (PCR) products showing altered mobility in SSCP analysis in order to determine the nucleotide changes. RESULTS: Three types of single-nucleotide polymorphism (SNP) were detected in the PDGFRA gene (exon 12, exon 13 and exon 18) while no mutation of PDGFRB was detected in the tested CBFLs. CONCLUSION: These data showed that no pathogenic mutations in PDGFRA and PDGFRB were detected in the context of CBFL without KIT and FLT3 mutations. Thus, PDGFR genes do not seem to be involved in CBFL and future studies are needed to establish the genetic causes of the disease in these particular patients.

Prognostic impact of c-KIT mutations in core binding factor leukemias: an Italian retrospective study.

Distinct forms of tyrosine kinase domain (TKD), juxtamembrane domain, exon 8, and internal tandem duplication (ITD) mutations of c-KIT, were observed in about 46% of core binding factor leukemia (CBFL) patients. To evaluate their prognostic significance, 67 adult patients with CBFL were analyzed to ascertain the c-KIT mutation status. In acute myeloid leukemia (AML) with t(8;21), the presence of c-KIT TKD mutation at codon 816 (TKD(816)) was associated with a high white blood cell count at diagnosis (median, 29.60 x 10(9)/L) and a higher incidence (33%) of extramedullary leukemia (EML) during the course of the disease. Data also showed that the TKD(816) mutated patients (n = 12) had a significantly higher incidence of relapse and a lower overall survival (OS) at 24 months, compared with the 17 c-KIT unmutated (c-KIT(-)) patients (90% vs 35.3%, P = .002; 25% vs 76.5%, P = .006, respectively). No difference in relapse incidence (P = .126) and OS (P = .474) was observed between the c-KIT mutated other than TKD(816) (n = 7) and the c-KIT(-) patients. These findings indicate that c-KIT TKD(816) mutation has a negative impact on the outcome of AML with t(8;21).

Epstein-Barr virus (EBV) load and interleukin-10 in EBV-positive and EBV-negative post-transplant lymphoproliferative disorders.

Post-transplant lymphoproliferative disorders (PTLDs) are heterogeneous severe complications occurring in 1-10% of transplanted patients. In most cases, PTLDs are associated with Epstein-Barr virus (EBV) infection but, recently, some clinical studies have reported an increasing number of EBV-negative PTLDs. Several studies have emphasized the critical role of the early identification of patients at risk for PTLD, in prompting the adoption of either pre-emptive strategies or timely treatment. To this purpose, monitoring of EBV DNA load in peripheral blood mononuclear cells is considered to be a useful test. Moreover, recently, the role of interleukin (IL)-10 in EBV-related diseases has been remarked, and high levels of IL-10 have been detected in PTLD patients. In this study, both EBV load and IL-10 were monitored in 38 PTLD patients at diagnosis and during follow-up, as well as in a control group, in order to establish the diagnostic role of the two tests, their relationship with the different PTLD subsets (EBV-positive and EBV-negative) and their behaviour during treatment. Results of our study suggest that the usefulness of IL-10 assay for early diagnosis of PTLD is similar to that of EBV load quantification, and its clinical diagnostic value is lower in EBV-negative than in EBV-positive PTLDs.