NT157, an IGF1R-IRS1/2 inhibitor, exhibits antineoplastic effects in pre-clinical models of chronic myeloid leukemia.

Chronic myeloid leukemia (CML) is successfully treated with BCR-ABL1 tyrosine kinase inhibitors, but a significant percentage of patients develop resistance. Insulin receptor substrate 1 (IRS1) has been shown to constitutively associate with BCR-ABL1, and IRS1-specific silencing leads to antineoplastic effects in CML cell lines. Here, we characterized the efficacy of NT157, a pharmacological inhibitor of IGF1R-IRS1/2, in CML cells and observed significantly reduced cell viability and proliferation, accompanied by induction of apoptosis. In human K562 cells and in murine Ba/F3 cells, engineered to express either wild-type BCR-ABL1 or the imatinib-resistant BCR-ABL1T315I mutant, NT157 inhibited BCR-ABL1, IGF1R, IRS1/2, PI3K/AKT/mTOR, and STAT3/5 signaling, increased CDKN1A, FOS and JUN tumor suppressor gene expression, and reduced MYC and BCL2 oncogenes. NT157 significantly reduced colony formation of human primary CML cells with minimal effect on normal hematopoietic cells. Exposure of primary CML cells harboring BCR-ABL1T315I to NT157 resulted in increased apoptosis, reduced cell proliferation and decreased phospho-CRKL levels. In conclusion, NT157 has antineoplastic effects on BCR-ABL1 leukemogenesis, independent of T315I mutational status.

The U2AF homology motif kinase 1 (UHMK1) is upregulated upon hematopoietic cell differentiation.

UHMK1 (KIS) is a nuclear serine/threonine kinase that possesses a U2AF homology motif and phosphorylates and regulates the activity of the splicing factors SF1 and SF3b155. Mutations in these components of the spliceosome machinery have been recently implicated in leukemogenesis. The fact that UHMK1 regulates these factors suggests that UHMK1 might be involved in RNA processing and perhaps leukemogenesis. Here we analyzed UHMK1 expression in normal hematopoietic and leukemic cells as well as its function in leukemia cell line. In the normal hematopoietic compartment, markedly higher levels of transcripts were observed in differentiated lymphocytes (CD4+, CD8+ and CD19+) compared to the progenitor enriched subpopulation (CD34+) or leukemia cell lines. UHMK1 expression was upregulated in megakaryocytic-, monocytic- and granulocytic-induced differentiation of established leukemia cell lines and in erythrocytic-induced differentiation of CD34+ cells. No aberrant expression was observed in patient samples of myelodysplastic syndrome (MDS), acute myeloid (AML) or lymphoblastic (ALL) leukemia. Nonetheless, in MDS patients, increased levels of UHMK1 expression positively impacted event free and overall survival. Lentivirus mediated UHMK1 knockdown did not affect proliferation, cell cycle progression, apoptosis or migration of U937 leukemia cells, although UHMK1 silencing strikingly increased clonogenicity of these cells. Thus, our results suggest that UHMK1 plays a role in hematopoietic cell differentiation and suppression of autonomous clonal growth of leukemia cells.

ANKHD1 silencing inhibits Stathmin 1 activity, cell proliferation and migration of leukemia cells.

ANKHD1 is highly expressed in human acute leukemia cells and potentially regulates multiple cellular functions through its ankyrin-repeat domains. In order to identify interaction partners of the ANKHD1 protein and its role in leukemia cells, we performed a yeast two-hybrid system screen and identified SIVA, a cellular protein known to be involved in proapoptotic signaling pathways. The interaction between ANKHD1 and SIVA was confirmed by co-imunoprecipitation assays. Using human leukemia cell models and lentivirus-mediated shRNA approaches, we showed that ANKHD1 and SIVA proteins have opposing effects. While it is known that SIVA silencing promotes Stathmin 1 activation, increased cell migration and xenograft tumor growth, we showed that ANKHD1 silencing leads to Stathmin 1 inactivation, reduced cell migration and xenograft tumor growth, likely through the inhibition of SIVA/Stathmin 1 association. In addition, we observed that ANKHD1 knockdown decreases cell proliferation, without modulating apoptosis of leukemia cells, while SIVA has a proapoptotic function in U937 cells, but does not modulate proliferation in vitro. Results indicate that ANKHD1 binds to SIVA and has an important role in inducing leukemia cell proliferation and migration via the Stathmin 1 pathway. ANKHD1 may be an oncogene and participate in the leukemia cell phenotype.

Montreal cognitive assessment in Brazilian adults with sickle cell disease: The burdens of poor sociocultural background.

Sickle cell disease (SCD) patients are at higher risk of developing silent cerebral infarcts and overt stroke, which may reflect cognitive impairment, functional limitations, and worse quality of life. The cognitive function of Brazilian adult SCD patients (n = 124; 19-70 years; 56 men; 79 SS, 28 SC, 10 S/ß0, 7 S/ß+) was screened through Montreal Cognitive Assessment (MoCA) and correlated the results with possible predictive factors for test performance, including sociocultural, clinical, laboratory data and brain imaging. The Median MoCA score was 23 (8-30); 70% had a 25-or-less score, suggesting some level of cognitive impairment. There were no significant associations between MoCA results and any clinical or laboratory data in SS and SC patients; however, a significant correlation (P = 0.03) with stroke was found in HbS/ß-thalassemic patients. Correlations were further detected according to sociodemographic conditions, such as age (r = -0.316; P < 0.001), age at first job (r = 0.221; P = 0.018), personal (r = 0.23; P = 0.012) and per capita familiar incomes (r = 0.303; P = 0.001), personal (r = 0.61; P = 0), maternal (r = 0.536; P = 0), and paternal educational status (r = 0.441; P = 0). We further sought independent predictors of performance using multivariable regressions and increased education was an independent predictor of better scores in MoCA (0.8099, 95% confidence interval [CI]: 0.509-1.111). Brain imaging analysis showed significant and progressive atrophy in important cerebral areas related to memory, learning, and executive function. These data point to the high prevalence and impact of cognitive decline in adult SCD patients, mirrored in brain atrophic areas. It is also possible to observe the influence of sociodemographic conditions on patients' cognitive performances and the need for creating focused therapeutic plans that address these deficiencies. Moreover, the absence of a significant correlation of MoCA values with stroke in the SS and SC groups may be related to the worst sociocultural and economic conditions of the Brazilian African descent population, in which the impact of low educational stimulation on cognitive function can outweigh even the anatomical damage caused by the disease.

Hematologic Malignancies Patients Face High Symptom Burden and Are Lately Referred to Palliative Consultation: Analysis of a Single Center Experience.

Although hematologic neoplasms have been on the vanguard of cancer therapies that led to notable advances in therapeutic efficacy, many patients face significant symptom burden, which make them eligible for early palliative care (PC) integration. However, previous reports demonstrated that hematological malignancies receive more aggressive care at the end-of-life and are less likely to receive care from specialist palliative services compared to solid tumors. Our aim was to characterize symptom burden, performance status and clinical characteristics of a cohort of hematologic malignancies patients referred to PC outpatient consultation, according to their diagnosis. Fifty-nine hematological malignancies patients referred to PC consultation between January 2018 and September 2021 were included. Clinical and laboratory data were evaluated retrospectively by medical charts analysis. Patients exhibited high ESAS and reduced PPS scores at the time of PC referral. Acute leukemia and multiple myeloma patients had the highest symptom burden scores; in spite of this, median time from the first PC consultation until death was only 3 and 4 months, respectively. In conclusion, we identified that hematologic neoplasms patients are highly symptomatic and are frequently referred to PC in end stages of their disease.

Early integration of palliative care in hematology: an urgency for patients, a challenge for physicians.

INTRODUCTION: Early integration between palliative care and other medical specialties in the care of patients with serious illnesses is consolidating itself as good medical practice, based on scientific and ethical evidence. Despite this, palliative care is still not part of the routine care of patients with hematological diseases, even in specialized centers. OBJECTIVE AND METHOD: In this article, we review the benefits and the main barriers described in the literature for early integration of hematology and palliative care. We also point out the challenges encountered in clinical practice, such as end-of-life prognosis assessment in patients with hematological diseases and management of the most common symptoms in hematology. Finally, we review models of integration between palliative care and oncology centers in outpatient and inpatient settings. RESULTS AND CONCLUSION: Patients with hematological diseases can greatly benefit from early integration with palliative care, with improvement in symptom control, quality of life, reduction of emotional distress and the development of advanced care directives. It is necessary to make hematologists aware of the benefits of palliative care, provide adequate training for multidisciplinary teams and encourage specific studies of palliative care in patients with hematological diseases.

Low SARS-CoV-2 seroprevalence in a cohort of Brazilian sickle cell disease patients: Possible effects of emphasis on social isolation for a population initially considered to be at very high risk.

Despite being initially considered at higher risk for severe COVID-19, sickle cell disease (SCD) patients have mostly presented clinical severity similar to the general population. As their vulnerability to become infected remains uncertain, we assessed the seroreactivity for SARS-CoV-2 to estimate the prevalence of infection and possible phenotypic and socioeconomic determinants for their contagion. Serologic evaluation was performed on 135 patients with an overall prevalence of 11%; positivity was associated with older age and use of public transportation. We speculate that social distancing instructions recommended by our clinic may have contributed to lower levels of infection, but potential protection factors need further investigation.

Reduced expression of FLIP SHORT in bone marrow of low risk myelodysplastic syndrome.

Apoptosis is dysregulated in patients with myelodysplastic syndrome (MDS) and acute myelogenous leukaemia (AML). FLIP (FLICE (FAS-associated death-domain-like IL-1 beta-converting enzyme)-inhibitory protein) has been described as an anti-apoptotic protein. Here, we characterize the expression level of FLIP(LONG) and FLIP(SHORT) mRNA in bone marrow aspirates from 61 patients diagnosed with MDS or AML. FLIP(SHORT) mRNA expression was significantly lower in low risk MDS, compared to high risk MDS, according to FAB classification (RA/RARS versus RAEB/RAEBt, P=0.0127) and IPSS (low risk/intermediate-1 versus intermediate-2/high risk, P=0.0345). Furthermore, FLIP(SHORT) mRNA expression was significantly lower in low risk MDS, compared to MDS-AML/AML de novo (P=0.0006), according to FAB classification. FLIP(LONG) expression did not differ between these groups. Increased levels of FLIP(SHORT) in RAEB and AML may be related to apoptosis resistance in these diseases and to MDS progression.

De novo AML exhibits greater microenvironment dysregulation compared to AML with myelodysplasia-related changes.

The interaction between the bone marrow microenvironment and malignant hematopoietic cells can result in the protection of leukemia cells from chemotherapy in both myelodysplastic syndromes (MDS) and acute myeloid leukemia (AML). We, herein, characterized the changes in cytokine expression and the function of mesenchymal stromal cells (MSC) in patients with MDS, AML with myelodysplasia-related changes (MRC), a well-recognized clinical subtype of secondary AML, and de novo AML. We observed a significant inhibitory effect of MDS-MSC on T lymphocyte proliferation and no significant differences in any of the cytokines tested. AML-MSC inhibited T-cell proliferation only at a very low MSC/T cell ratio. When compared to the control, AML-MRCderived MSC presented a significant increase in IL6 expression, whereas de novo AML MSC presented a significant increase in the expression levels of VEGFA, CXCL12, RPGE2, IDO, IL1ß, IL6 and IL32, followed by a decrease in IL10 expression. Furthermore, data indicate that IL-32 regulates stromal cell proliferation, has a chemotactic potential and participates in stromal cell crosstalk with leukemia cells, which could result in chemoresistance. Our results suggest that the differences between AML-MRC and de novo AML also extend into the leukemic stem cell niche and that IL-32 can participate in the regulation of the bone marrow cytokine milieu.

Primary myelofibrosis: current therapeutic options.

Primary myelofibrosis is a Philadelphia-negative myeloproliferative neoplasm characterized by clonal myeloid expansion, followed by progressive fibrous connective tissue deposition in the bone marrow, resulting in bone marrow failure. Clonal evolution can also occur, with an increased risk of transformation to acute myeloid leukemia. In addition, disabling constitutional symptoms secondary to the high circulating levels of proinflammatory cytokines and hepatosplenomegaly frequently impair quality of life. Herein the main current treatment options for primary myelofibrosis patients are discussed, contemplating disease-modifying therapeutics in addition to palliative measures, in an individualized patient-based approach.

Low Ten-eleven-translocation 2 (TET2) transcript level is independent of TET2 mutation in patients with myeloid neoplasms.

BACKGROUND: New sequencing technologies have enabled the identification of mutations in Ten-eleven-translocation 2 (TET2), an enzyme that catalyzes the conversion of 5-methylcytosine into 5-hydroxymethylcytosine (5-hmC) in myeloid neoplasms. We have recently identified reduced TET2 mRNA expression in myelodysplastic syndromes (MDS) and acute myeloid leukemia (AML), which is associated with a poor overall survival in MDS. We herein aimed to investigate TET2 mutations and their impact on TET2 expression in a cohort of patients with myeloid neoplasms, including MDS and AML patients. FINDINGS: TET2 mutations were observed in 8 out of 19 patients (42 %) with myeloid neoplasms. The TET2 expression profile was similar between in wild type and in TET2 mutated patients. CONCLUSION: Our results suggest that TET2 expression is reduced in MDS/AML patients, independently of mutational status.

IRS2 silencing increases apoptosis and potentiates the effects of ruxolitinib in JAK2V617F-positive myeloproliferative neoplasms.

The recurrent V617F mutation in JAK2 (JAK2V617F) has emerged as the primary contributor to the pathogenesis of myeloproliferative neoplasms (MPN). However, the lack of complete response in most patients treated with the JAK1/2 inhibitor, ruxolitinib, indicates the need for identifying pathways that cooperate with JAK2. Activated JAK2 was found to be associated with the insulin receptor substrate 2 (IRS2) in non-hematological cells. We identified JAK2/IRS2 binding in JAK2V617F HEL cells, but not in the JAK2WT U937 cell line. In HEL cells, IRS2 silencing decreased STAT5 phosphorylation, reduced cell viability and increased apoptosis; these effects were enhanced when IRS2 silencing was combined with ruxolitinib. In U937 cells, IRS2 silencing neither reduced cell viability nor induced apoptosis. IRS1/2 pharmacological inhibition in primary MPN samples reduced cell viability in JAK2V617F-positive but not JAK2WT specimens; combination with ruxolitinib had additive effects. IRS2 expression was significantly higher in CD34+ cells from essential thrombocythemia patients compared to healthy donors, and in JAK2V617F MPN patients when compared to JAK2WT. Our data indicate that IRS2 is a binding partner of JAK2V617F in MPN. IRS2 contributes to increased cell viability and reduced apoptosis in JAK2-mutated cells. Combined pharmacological inhibition of IRS2 and JAK2 may have a potential clinical application in MPN.

Molecular matching for Rh and K reduces red blood cell alloimmunisation in patients with myelodysplastic syndrome.

BACKGROUND: Matching for Rh and K antigens has been used in an attempt to reduce antibody formation in patients receiving chronic transfusions but an extended phenotype matching including Fy(a) and Jk(a) antigens has also been recommended. The aim of this study was to identify an efficient transfusion protocol of genotype matching for patients with myelodysplastic syndrome (MDS) or chronic myelomonocytic leukaemia. We also examined a possible association of HLA class II alleles with red blood cell (RBC) alloimmunisation. MATERIALS AND METHODS: We evaluated 43 patients with MDS undergoing transfusion therapy with and without antibody formation. We investigated antigen-matched RBC units for ABO, D, C, c, E, e, K, Fy(a), Fy(b), Jk(a), Jk(b), S, s, Do(a), Do(b) and Di(a) on the patients' samples and on the donor units serologically matched for them based on their ABO, Rh and K phenotypes and presence of antibodies. We also determined the frequencies of HLA-DRB1 alleles in the alloimmunised and non-alloimmunised patients. RESULTS: Seventeen of the 43 patients had discrepancies or mismatches for multiple antigens between their genotype-predicted profile and the antigen profile of the units of blood serologically matched for them. We verified that 36.8% of patients had more than one RBC alloantibody and 10.5% of patients had autoantibodies. Although we were able to find a better match for the patients in our extended genotyped/phenotyped units, we verified that matching for Rh and K would be sufficient for most of the patients. We also observed an over-representation of the HLA-DRB1*13 allele in the non-alloimmunised group of patients with MDS. DISCUSSION: In our population molecular matching for C, c, E, e, K was able to reduce RBC alloimmunisation in MDS patients. An association of HLA-DRB1*13 and protection from RBC alloimmunisation should be confirmed.

Stathmin 1 inhibition amplifies ruxolitinib-induced apoptosis in JAK2V617F cells.

The JAK/STAT pathway is constitutively activated in myeloproliferative neoplasms and can be inhibited by ruxolitinib, a selective JAK1/2 inhibitor. The JAK2(V617F) mutation leads to constitutive STAT3 phosphorylation and potentially leads to inhibition of Stathmin 1 activity via STAT3. In support of this hypothesis, we found that, in HEL JAK2(V617F) cells, ruxolitinib treatment decreased STAT3 and Stathmin 1 association, induced Stathmin 1 activation and microtubule instability. Silencing of Stathmin 1 significantly reduced cell proliferation and clonal growth, and increased apoptosis induced by ruxolitinib. Stathmin 1 silencing also prevented ruxolitinib-induced microtubule instability. To phenocopy the effect of Stathmin 1 inhibition, cells were treated with paclitaxel, a microtubule-stabilizing drug, in association or not with ruxolitinib; combined treatment significantly increased apoptosis, when compared to monotherapy. Notably, Stathmin 1 mRNA levels were highly expressed in CD34(+) cells from primary myelofibrosis patients. We then proposed that an undesired effect of ruxolitinib treatment may constitute Stathmin 1 activation and microtubule instability in JAK2(V617F) cells. Induction of microtubule stability, through Stathmin 1 silencing or paclitaxel treatment, combined with ruxolitinib could be an effective strategy for promoting apoptosis in JAK2(V617F) cells.

Myelodysplastic syndrome with synchronous gastric cancer: when the symptoms suggest something else.

Although myelodysplastic syndromes have a clear definition in theory, the morphologic dysplasia associated with ineffective hematopoiesis may be subtle and difficult to recognize and can commonly be mimicked by systemic conditions, such as infections, autoimmune disorders, nutritional deficiencies, toxic factors and non-hematological malignancies. However, myelodysplastic syndromes may truly coexist with other systemic diseases, which can be masked when the patient's symptoms are attributed exclusively to myelodysplastic syndromes without further investigation. To better illustrate this, we herein describe two cases associated with synchronous gastric cancers.

Stathmin 1 is involved in the highly proliferative phenotype of high-risk myelodysplastic syndromes and acute leukemia cells.

Stathmin 1 is an important cytoplasmic microtubule-destabilizing protein that plays critical roles in proliferation and accurate chromosome segregation through regulation of microtubule dynamics. High levels of Stathmin 1 expression have been reported in leukemia and solid tumors. However, Stathmin 1 has not been studied in myelodysplastic syndrome cells. We, herein, report that significantly higher Stathmin 1 levels were observed in proliferating hematopoietic cells, in high-risk MDS and acute leukemia cells. In addition, Stathmin 1 silencing in U937 and Namalwa leukemia cells reduced cell proliferation and clonogenicity. Our data suggest that Stathmin 1 expression may be related to the highly proliferative phenotype of hematopoietic cells and add new insights into the participation of Stathmin 1 in hematological malignancies.