IL-7R-mediated signaling in T-cell acute lymphoblastic leukemia: an update

© 2018 The Authors. Published by Elsevier Ltd. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/BY-NC-ND/4.0/)Interleukin 7 (IL-7) and its receptor (IL-7R, a heterodimer of IL-7Rα and γc) are essential for normal lymphoid development. In their absence, severe combined immunodeficiency occurs. By contrast, excessive IL-7/IL-7R-mediated signaling can drive lymphoid leukemia development, disease acceleration and resistance to chemotherapy. IL-7 and IL-7R activate three main pathways: STAT5, PI3K/Akt/mTOR and MEK/Erk, ultimately leading to the promotion of leukemia cell viability, cell cycle progression and growth. However, the contribution of each of these pathways towards particular functional outcomes is still not completely known and appears to differ between normal and malignant states. For example, IL-7 upregulates Bcl-2 in a PI3K/Akt/mTOR-dependent and STAT5-independent manner in T-ALL cells. This is a 'symmetric image' of what apparently happens in normal lymphoid cells, where PI3K/Akt/mTOR does not impact on Bcl-2 and regulates proliferation rather than survival. In this review, we provide an updated summary of the knowledge on IL-7/IL-7R-mediated signaling in the context of cancer, focusing mainly on T-cell acute lymphoblastic leukemia, where this axis has been more extensively studied.Publication costs were supported by LISBOA-01-0145-FEDER-007391, project cofunded by FEDER, through POR Lisboa2020 Programa Operacional Regional de Lisboa, PORTUGAL 2020, and Fundação para a Ciência e a Tecnologia (FCT, Portugal). The research work in JTB's lab related to the present review was supported by the grants FAPESP/20015/2014 and PTDC/MEC-HEM/31588/2017, from FCT; and by the consolidator grant ERC CoG-648455 from the European Research Council, under the European Union's Horizon 2020 research and innovation programme. JTB is an FCT investigator (consolidator). MLO is a LisbonBioMed PhD student and received a fellowship from FCT. PA received a PhD fellowship from the EU Marie Sklodowska-Curie ITN Protein Conjugates.info:eu-repo/semantics/publishedVersio

Q192R polymorphism of the paraoxonase-1 gene as a risk factor for obesity in Portuguese women

Introduction - Obesity became a major public health problem as a result of its increasing prevalence worldwide. Paraoxonase-1 (PON1) is an esterase able to protect membranes and lipoproteins from oxidative modifications. At the PON1 gene, several polymorphisms in the promoter and coding regions have been identified. The aims of this study were i) to assess PON1 L55M and Q192R polymorphisms as a risk factor for obesity in women; ii) to compare PON1 activity according to the expression of each allele in L55M and Q192R polymorphisms; iii) to compare PON1 activity between obese and normal-weight women. Materials and methods - We studied 75 healthy (35.9±8.2 years) and 81 obese women (34.3±8.2 years). Inclusion criteria for obese subjects were body mass index ≥30 kg/m2 and absence of inflammatory/neoplasic conditions or kidney/hepatic dysfunction. The two PON1 polymorphisms were assessed by real-time PCR with TaqMan probes. PON1 enzymatic activity was assessed by spectrophotometric methods, using paraoxon as a substrate. Results - No significant differences were found for PON1 activity between normal and obese women. Nevertheless, PON1 activity was greater (P<0.01) for the RR genotype (in Q192R polymorphism) and for the LL genotype (in L55M polymorphism). The frequency of allele R of Q192R polymorphism was significantly higher in obese women (P<0.05) and was associated with an increased risk of obesity (odds ratio=2.0 – 95% confidence interval (1.04; 3.87)). Conclusion - 55M and Q192R polymorphisms influence PON1 activity. The allele R of the Q192R polymorphism is associated with an increased risk for development of obesity among Portuguese Caucasian premenopausal women

Factors related with adiponectinemia in obese and normal-weight women and with its variation in weight loss programs

Objective: To assess different factors influencing adiponectinemia in obese and normal-weight women; to identify factors associated with the variation (Δ) in adiponectinemia in obese women following a 6-month weight loss program, according to surgical/non-surgical interventions. Methods: We studied 100 normal-weight women and 112 obese premenopausal women; none of them was on any medical treatment. Women were characterized for anthropometrics, daily macronutrient intake, smoking status, contraceptives use, adiponectin as well as IL-6 and TNF-α serum concentrations. Results: Adiponectinemia was lower in obese women (p < 0.001), revealing an inverse association with waist-to-hip ratio (p < 0.001; r = –0.335). Normal-weight women presented lower adiponectinemia among smokers (p = 0.041); body fat, waist-to-hip ratio, TNF-α levels, carbohydrate intake, and smoking all influence adiponectinemia (r 2 = 0.436). After weight loss interventions, a significant modification in macronutrient intake occurs followed by anthropometrics decrease (chiefly after bariatric procedures) and adiponectinemia increase (similar after surgical and non-surgical interventions). After bariatric intervention, Δ adiponectinemia was inversely correlated to Δ waist circumference and Δ carbohydrate intake (r 2 = 0.706). Conclusion: Anthropometrics, diet, smoking, and TNF-α levels all influence adiponectinemia in normal-weight women, although explaining less than 50% of it. In obese women, anthropometrics modestly explain adiponectinemia. Opposite to non-surgical interventions, after bariatric surgery adiponectinemia increase is largely explained by diet composition and anthropometric changes

Clinical-grade peptide-based inhibition of CK2 blocks viability and proliferation of T-ALL cells and counteracts IL-7 stimulation and stromal support

© 2020 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/).Despite remarkable advances in the treatment of T-cell acute lymphoblastic leukemia (T-ALL), relapsed cases are still a major challenge. Moreover, even successful cases often face long-term treatment-associated toxicities. Targeted therapeutics may overcome these limitations. We have previously demonstrated that casein kinase 2 (CK2)-mediated phosphatase and tensin homologue (PTEN) posttranslational inactivation, and consequent phosphatidylinositol 3-kinase (PI3K)/Akt signaling hyperactivation, leads to increased T-ALL cell survival and proliferation. We also revealed the existence of a crosstalk between CK2 activity and the signaling mediated by interleukin 7 (IL-7), a critical leukemia-supportive cytokine. Here, we evaluated the impact of CIGB-300, a the clinical-grade peptide-based CK2 inhibitor CIGB-300 on T-ALL biology. We demonstrate that CIGB-300 decreases the viability and proliferation of T-ALL cell lines and diagnostic patient samples. Moreover, CIGB-300 overcomes IL-7-mediated T-ALL cell growth and viability, while preventing the positive effects of OP9-delta-like 1 (DL1) stromal support on leukemia cells. Signaling and pull-down experiments indicate that the CK2 substrate nucleophosmin 1 (B23/NPM1) and CK2 itself are the molecular targets for CIGB-300 in T-ALL cells. However, B23/NPM1 silencing only partially recapitulates the anti-leukemia effects of the peptide, suggesting that CIGB-300-mediated direct binding to CK2, and consequent CK2 inactivation, is the mechanism by which CIGB-300 downregulates PTEN S380 phosphorylation and inhibits PI3K/Akt signaling pathway. In the context of IL-7 stimulation, CIGB-300 blocks janus kinase / signal transducer and activator of transcription (JAK/STAT) signaling pathway in T-ALL cells. Altogether, our results strengthen the case for anti-CK2 therapeutic intervention in T-ALL, demonstrating that CIGB-300 (given its ability to circumvent the effects of pro-leukemic microenvironmental cues) may be a valid tool for clinical intervention in this aggressive malignancy.This work was supported by the consolidator grant ERC CoG-648455 from the European Research Council, under the European Union’s Horizon 2020 research and innovation program, and FAPESP/20015/2014 and PTDC/MEC-HEM/31588/2017 grants from Fundação para a Ciência e a Tecnologia (FCT), to JTB.info:eu-repo/semantics/publishedVersio

Exploring the role of IL-7, CK2, and SphK in T-cell acute lymphoblastic leukemia

A leucemia linfoblástica aguda de linfócitos T (LLA-T) é uma doença hemato oncológica infantil agressiva, de mau prognóstico. Apesar das melhorias significativas no tratamento desta doença, são necessárias novas terapias menos tóxicas. A interleucina-7 (IL-7) e o seu receptor (IL-7R) são essenciais para o normal desenvolvimento e homeostasia dos linfócitos-T, podendo, contudo, contribuir também para a viabilidade e proliferação das células LLA-T e acelerar a progressão da leucemia in vivo. Cerca de 9% dos pacientes LLA-T apresentam mutações de ganho-de-função no IL-7R, conduzindo a sinalização constitutiva e oncogénica. Nesta tese, identificámos STAT5 como elemento essencial para uma eficiente sinalização mediada pela IL-7 em LLA-T, envolvido na inibição de apoptose, aumento do crescimento e proliferação das células leucémicas. Demonstrámos igualmente que a via Jak/STAT5 é um dos principais reguladores da rede trascripcional activada por IL-7, revelando PIM1 como alvo directo e caracterizando o seu papel na transmissão dos sinais pro-sobrevivência e proliferação mediados pela IL-7. Também identificámos a caseína cinase (CK2) e esfingosina cinases (SphK) como moduladores-chave dos sinais induzidos pela IL-7. Demonstrámos que a IL 7 regula positivamente a actividade da CK2 e SphK, sendo que a inibição destas enzimas impede a activação, mediada pela IL-7, das vias de sinalização PI3K/AKT e JAK/STAT, e a consequente promoção de viabilidade e entrada no ciclo celular das células LLA-T. Concluindo, os nossos estudos identificam Jak/STAT5/PIM1 como uma via principal dos sinais mediados pela IL-7, e a CK2 e a SphK como moduladores essenciais da activação das vias pro-sobrevivência e proliferação dependentes da IL-7 em LLA-T. Uma vez que mais de 70% dos pacientes de LLA-T respondem à IL-7 ou possuem mutações de ganho-de-função no IL-7R, as nossas observações abrem novas possibilidades terapêuticas para esta patologia.T-cell acute lymphoblastic leukemia (T-ALL) is an aggressive childhood hematological malignancy. Despite the many improvements made in the treatment of T-ALL, it remains a therapeutic challenge. Novel, less toxic therapies are still needed. Interleukin-7 (IL-7) and its receptor (IL-7R) are essential for normal T-cell development and homeostasis. However, IL-7 can also contribute to the viability and proliferation of T-ALL cells and accelerate leukemia progression in vivo. Moreover, around 9% of T-ALL patients display IL-7R gain-of-function mutations that lead to constitutive and oncogenic signaling. Here, we identified STAT5 as an essential element for efficient IL-7-mediated signaling, involved in preventing apoptosis, and promoting cell growth and proliferation of T-ALL cells. Moreover, we showed that Jak/STAT5 signaling pathway is a major regulator of the transcriptional network downstream of IL-7 stimulation in T-ALL. We demonstrated that PIM1 is a direct STAT5 target, involved in IL-7-mediated pro-survival and proliferative signals in T-ALL. Overall, STAT5 and its transcriptional network are essential for IL-7-mediated survival of T-ALL cells. We also identified casein kinase 2 (CK2) and sphingosine kinases (SphK) as key modulators of IL-7-mediated signaling. IL-7 positively regulates CK2 and SphK activity without significantly affecting their expression. Inhibition of these kinases prevents IL-7-mediated activation of both PI3K/AKT and JAK/STAT pathways, and consequent viability and cell cycle progression of T-ALL cells. In summary, our studies identified Jak/STAT5/PIM1 as a main axis downstream from IL-7 stimulation and CK2 and SphK as essential modulators of IL-7-dependent activation of pro-survival and proliferative pathways in T-ALL. Since more than 70% of T-ALL patients respond to IL-7 or have IL-7R gain-of-function mutations, our observations open new possible therapeutic avenues in this pathology

microRNAs regulate TAL1 expression in T-cell acute lymphoblastic leukemia.

The transcription factor TAL1 is a proto-oncogene whose aberrant expression in committed T-cell precursors is associated with the development of T-cell acute lymphoblastic leukemia (T-ALL). The mechanisms leading to aberrant activation of TAL1 in T-ALL patients who lack chromosomal rearrangements involving the TAL1 locus remain largely unknown. We hypothesized that TAL1 levels decrease during normal T-cell development at least in part due to miRNA-dependent silencing, in which case TAL1 over-expression in some T-ALL cases could be the consequence of deregulated miRNA expression. By performing computational prediction of miRNAs that bind to the human TAL1 mRNA we compiled a list of miRNAs that are candidates to regulate TAL1. Using a luciferase reporter system and mutagenesis assays we confirmed the miRNA-TAL1 mRNA interactions and selected candidate miRNAs: miR-101, miR-520d-5p, miR-140-5p, miR-448 and miR-485-5p. Over-expression of these microRNAs in different T-ALL cell lines consistently resulted in the down-regulation of TAL1 protein. In accordance, inhibition of miR-101 and miR-520d-5p promoted TAL1 protein expression. Importantly, we found that miR-101, miR-140-5p, miR-448 and miR-485-5p were down-regulated in T-ALL patient specimens and T-ALL cell lines. Our results show for the first time the existence of epigenetic regulation of TAL1 by specific miRNAs which may contribute, at least in part, to the ectopic expression of TAL1 in some T-ALL cases.These studies were financed by Liga Portuguesa Contra o Cancro (Terry Fox Award) and by Fundacao para a Ciencia e a Tecnologia (project PTDC/BIM-ONC/1548/2012). NCC received an FCT-SFRH PhD fellowship. JTB is supported by an FCT Consolidation Grant. The Cell Division and Cancer group of the CNIO is funded by the Spanish Ministry of Economy and Competitiveness (MINECO; SAF2012-38215: Consolider-Ingenio 2010 Programme SAF2014-57791-REDC; Red Tematica CellSYS BFU2014-52125-REDT), and the Comunidad de Madrid (OncoCycle Programme S2010/BMD-2470).S

Factors Related with Adiponectinemia in Obese and Normal-Weight Women and with Its Variation in Weight Loss Programs

Objective: To assess different factors influencing adiponectinemia in obese and normal-weight women; to identify factors associated with the variation (&#x0394;) in adiponectinemia in obese women following a 6-month weight loss program, according to surgical/non-surgical interventions. Methods: We studied 100 normal-weight women and 112 obese premenopausal women; none of them was on any medical treatment. Women were characterized for anthropometrics, daily macronutrient intake, smoking status, contraceptives use, adiponectin as well as IL-6 and TNF-α serum concentrations. Results: Adiponectinemia was lower in obese women (p 2 = 0.436). After weight loss interventions, a significant modification in macronutrient intake occurs followed by anthropometrics decrease (chiefly after bariatric procedures) and adiponectinemia increase (similar after surgical and non-surgical interventions). After bariatric intervention, &#x0394; adiponectinemia was inversely correlated to &#x0394; waist circumference and &#x0394; carbohydrate intake (r2 = 0.706). Conclusion: Anthropometrics, diet, smoking, and TNF-α levels all influence adiponectinemia in normal-weight women, although explaining less than 50% of it. In obese women, anthropometrics modestly explain adiponectinemia. Opposite to non-surgical interventions, after bariatric surgery adiponectinemia increase is largely explained by diet composition and anthropometric changes

Adult B-cell acute lymphoblastic leukemia cells display decreased PTEN activity and constitutive hyperactivation of PI3K/Akt pathway despite high PTEN protein levels

© 2014 Ferrata Storti Foundation. This is an open-access paperAdult B-cell acute lymphoblastic leukemia remains a major therapeutic challenge, requiring a better characterization of the molecular determinants underlying disease progression and resistance to treatment. Here, using a phospho-flow cytometry approach we show that adult diagnostic B-cell acute lymphoblastic leukemia specimens display PI3K/Akt pathway hyperactivation, irrespective of their BCR-ABL status and despite paradoxically high basal expression of PTEN, the major negative regulator of the pathway. Protein kinase CK2 is known to phosphorylate PTEN thereby driving PTEN protein stabilization and concomitant PTEN functional inactivation. In agreement, we found that adult B-cell acute lymphoblastic leukemia samples show significantly higher CK2 kinase activity and lower PTEN lipid phosphatase activity than healthy controls. Moreover, the clinical-grade CK2 inhibitor CX-4945 (Silmitasertib) reversed PTEN levels in leukemia cells to those observed in healthy controls, and promoted leukemia cell death without significantly affecting normal bone marrow cells. Our studies indicate that CK2-mediated PTEN posttranslational inactivation, associated with PI3K/Akt pathway hyperactivation, are a common event in adult B-cell acute lymphoblastic leukemia and suggest that CK2 inhibition may constitute a valid, novel therapeutic tool in this malignancy.This study was supported by grant PTDC/IC/83023/2007, from Fundação para a Ciência e a Tecnologia (FCT). A.M.G. and V.P. received BI fellowships from FCT. L.R.M. and A.M. received a post-doctoral and a PhD fellowship, respectively, both from FCT.info:eu-repo/semantics/publishedVersio

STAT5 is essential for IL-7-mediated viability, growth, and proliferation of T-cell acute lymphoblastic leukemia cells

T-cell acute lymphoblastic leukemia (T-ALL) constitutes an aggressive subset of ALL, the most frequent childhood malignancy. Whereas interleukin-7 (IL-7) is essential for normal T-cell development, it can also accelerate T-ALL development in vivo and leukemia cell survival and proliferation by activating phosphatidylinositol 3-kinase/protein kinase B/mechanistic target of rapamycin signaling. Here, we investigated whether STAT5 could also mediate IL-7 T-ALL-promoting effects. We show that IL-7 induces STAT pathway activation in T-ALL cells and that STAT5 inactivation prevents IL-7-mediated T-ALL cell viability, growth, and proliferation. At the molecular level, STAT5 is required for IL-7-induced downregulation of p27kip1 and upregulation of the transferrin receptor, CD71. Surprisingly, STAT5 inhibition does not significantly affect IL-7-mediated Bcl-2 upregulation, suggesting that, contrary to normal T-cells, STAT5 promotes leukemia cell survival through a Bcl-2-independent mechanism. STAT5 chromatin immunoprecipitation sequencing and RNA sequencing reveal a diverse IL-7-driven STAT5-dependent transcriptional program in T-ALL cells, which includes BCL6 inactivation by alternative transcription and upregulation of the oncogenic serine/threonine kinase PIM1 Pharmacological inhibition of PIM1 abrogates IL-7-mediated proliferation on T-ALL cells, indicating that strategies involving the use of PIM kinase small-molecule inhibitors may have therapeutic potential against a majority of leukemias that rely on IL-7 receptor (IL-7R) signaling. Overall, our results demonstrate that STAT5, in part by upregulating PIM1 activity, plays a major role in mediating the leukemia-promoting effects of IL-7/IL-7R