Biomedical subjects

Angelo A Cardoso

Publications and source records attributed to Angelo A Cardoso.

15 recordsLinked to original sources

PubMed2006

Leukemia-stimulated bone marrow endothelium promotes leukemia cell survival.

Extensive endothelial cell proliferation and marked neovascularization are the most pronounced microenvironmental changes consistently observed in the bone marrow (BM) of patients with acute lymphoblastic leukemia (ALL). It is not known whether ALL cells induce this phenotype and whether they receive critical signals from the tumor-associated BM endothelium. Here, we show that leukemia cells actively stimulate BM endothelium, promote de novo angiogenesis, and induce neovascularization in the leukemic BM. Soluble factors, present in the leukemic BM microenvironment, promote the proliferation, migration, and morphogenesis of BM endothelial cells, which are critical processes in tumor angiogenesis. We also show in vitro that leukemia cells display directional motion towards assembled BM endothelium and following adherence exhibit cell polarization, pseudopodia, and ultrastructural features that suggest the existence of leukemia-endothelium cross-talk. Finally, we show that BM endothelium promotes leukemia cell survival through a mechanism mediated through the anti-apoptotic molecule bcl-2. These studies indicate that ALL cells actively recruit BM endothelium and mediate the leukemia-associated neovascularization observed in ALL. Therefore, disruption of interactions between leukemia cells and BM endothelium may constitute a valid therapeutic strategy.

Animals↗

PubMed2006

Molecular and functional evidence for activity of murine IL-7 on human lymphocytes.

Although interleukin-7 (IL-7) is essential for human and murine lymphopoiesis and homeostasis, clear disparities between these species regarding the role of IL-7 during B-cell development suggest that other, subtler differences may exist. One basic unsolved issue of IL-7 biology concerns cross-species activity, because in contrast to the human ortholog, the ability of murine (m)IL-7 to stimulate human cells remains unresolved. Establishing whether two-way cross-species reactivity occurs is fundamental for evaluating the role of IL-7 in chimeric human-mouse models, which are the most versatile tools for studying human lymphoid development and disease in vivo. Here, we show that mIL-7 triggers the same signaling pathways as human (h)IL-7 in human T cells, promoting similar changes in viability, proliferation, size, and immunophenotype, even at low concentrations. This ability is not confined to T cells, because mIL-7 mediates cell growth and protects human B-cell precursors from dexamethasone-induced apoptosis. Importantly, endogenous mIL-7 produced in the mouse thymic microenvironment stimulates human T cells, because their expansion in chimeric fetal thymic organ cultures is inhibited by a mIL-7-specific neutralizing antibody. Our results demonstrate that mIL-7 affects human lymphocytes and indicate that mouse models of human lymphoid development and disease must integrate the biological effects of endogenous IL-7 on human cells.

Animals↗

PubMed2006

Evaluation of the anti-proliferative effect the extracts of Allamanda blanchetti and A. schottii on the growth of leukemic and endothelial cells.

PURPOSE: To investigate the anti-proliferative effect of A. blanchetti and A. schottii extracts. METHODS: The anti-proliferative effect of A. blanchetti and A. schottii ethanolic extracts on K562 leukemic cells as well as on BMEC and HUVEC were evaluated. Phytochemical analysis to identify the possible active components was carried out. RESULTS: The root extract of A. schottii was the most active of them. At 80 microg/mL, the root extracts showed a cytostatic effect on K562, whereas at 400 microg/mL, there was a strong cytotoxic effect. Similar cytostatic and cytotoxic effects were seen in the endothelial cells, but at lower doses. The effect of A. schottii root extract on endothelial cells was seen at concentrations ten times lower (8 microg/mL) than the effect of the A. blanchetti root extract (80 microg/mL). Phytochemical investigation of different fractions and parts of the plant led to the isolation of several known compounds, some of which are described for the first time in the genus Allamanda, and with previous evidence of anticancer and antitumoral properties. CONCLUSIONS: Our results suggest that both plants studied exhibit cytostatic and cytotoxic activity, but the most active compounds are located in the roots.

Antineoplastic Agents↗

PubMed2005

Gene expression profiling identifies BAX-delta as a novel tumor antigen in acute lymphoblastic leukemia.

The identification of new tumor-associated antigens (TAA) is critical for the development of effective immunotherapeutic strategies, particularly in diseases like B-cell acute lymphoblastic leukemia (B-ALL), where few target epitopes are known. To accelerate the identification of novel TAA in B-ALL, we used a combination of expression profiling and reverse immunology. We compared gene expression profiles of primary B-ALL cells with their normal counterparts, B-cell precursors. Genes differentially expressed by B-ALL cells included many previously identified as TAA in other malignancies. Within this set of overexpressed genes, we focused on those that may be functionally important to the cancer cell. The apoptosis-related molecule, BAX, was highly correlated with the ALL class distinction. Therefore, we evaluated BAX and its isoforms as potential TAA. Peptides from the isoform BAX-delta bound with high affinity to HLA-A*0201 and HLA-DR1. CD8+ CTLs specific for BAX-delta epitopes or their heteroclitic peptides could be expanded from normal donors. BAX-delta-specific T cells lysed peptide-pulsed targets and BAX-delta-expressing leukemia cells in a MHC-restricted fashion. Moreover, primary B-ALL cells were recognized by BAX-delta-specific CTL, indicating that this antigen is naturally processed and presented by tumor cells. This study suggests that (a) BAX-delta may serve as a widely expressed TAA in B-ALL and (b) gene expression profiling can be a generalizable tool to identify immunologic targets for cancer immunotherapy.

Adolescent↗

PubMed2005

Proangiogenic stimulation of bone marrow endothelium engages mTOR and is inhibited by simultaneous blockade of mTOR and NF-kappaB.

Most bone marrow (BM) malignancies develop in association with an angiogenic phenotype and increased numbers of endothelial cells. The molecular mechanisms involved in the modulation and recruitment of BM endothelium are largely unknown and may provide novel therapeutic targets for neoplastic diseases. We observed that angiogenic stimulation of BM endothelial cells activates mTOR and engages its downstream pathways 4E-BP1 and S6K1, which are inhibited by the mTOR-specific blockers rapamycin and CCI-779. Both mTOR blockers significantly inhibit growth factor- and leukemia-induced proliferation of BM endothelium by inducing G0/G1 cell-cycle arrest. This effect is associated with down-regulation of cyclin D1 and cdk2 phosphorylation, and up-regulation of the cdk inhibitors p27(kip1) and p21(cip1). Under conditions that reproduce the biomechanical fluidic environment of the BM, CCI-779 is equally effective in inhibiting BM endothelial-cell proliferation. Finally, simultaneous blockade of mTOR and NF-kappaB pathways synergize to significantly inhibit or abrogate the proliferative responses of BM endothelial cells to mitogenic stimuli. This study identifies mTOR as an important pathway for the proangiogenic stimulation of BM endothelium. Modulation of this pathway may serve as a valid therapeutic intervention in BM malignancies evolving in association with an angiogenic phenotype.

Adult↗

PubMed2005

Notch1 modulates timing of G1-S progression by inducing SKP2 transcription and p27 Kip1 degradation.

Cyclin-dependent kinase inhibitors (CKIs) and Notch receptor activation have been shown to influence adult stem cells and progenitors by altering stem cell self-renewal and proliferation. Yet, no interaction between these molecular pathways has been defined. Here we show that ligand-independent and ligand-dependent activation of Notch1 induces transcription of the S phase kinase-associated protein 2 (SKP2), the F-box subunit of the ubiquitin-ligase complex SCF(SKP2) that targets proteins for degradation. Up-regulation of SKP2 by Notch signaling enhances proteasome-mediated degradation of the CKIs, p27 Kip1 and p21 Cip1, and causes premature entry into S phase. Silencing of SKP2 by RNA interference in G1 stabilizes p27 Kip1 and p21 Cip1 and abolishes Notch effect on G1-S progression. Thus, SKP2 serves to link Notch1 activation with the cell cycle machinery. This novel pathway involving Notch/SKP2/CKIs connects a cell surface receptor with proximate mediators of cell cycle activity, and suggests a mechanism by which a known physiologic mediator of cell fate determination interfaces with cell cycle control.

3T3 Cells↗

PubMed2005

Failure to define window of time for autologous tumor vaccination in patients with newly diagnosed or relapsed acute lymphoblastic leukemia.

OBJECTIVES: We and others have shown that B cell precursor acute lymphoblastic leukemia cells (ALL) stimulated with CD40 ligand become efficient antigen-presenting cells (APC) capable of expanding autologous, tumor-specific T cells from patients. Translation of these preclinical findings to a novel treatment strategy required four separate issues to be determined: (1) if a CD40-ALL vaccine could be generated for clinical use; (2) whether clinical translation could be achieved; (3) whether the vaccination was safe; and (4) whether a window of time could be identified that would optimize the efficacy of vaccination. PATIENTS AND METHODS: Nine patients with relapsed/refractory ALL were enrolled in a phase I trial of vaccination with autologous CD40-ALL. Immunologic reconstitution was measured in a separate cohort of 23 patients with newly diagnosed ALL. RESULTS: We successfully prepared autologous vaccines for all nine patients in the phase I trial. CD40-ALL were potent APC, capable of stimulating allogeneic and peptide-specific T cells in vitro. Two patients were vaccinated without adverse events. Five patients died or progressed before vaccination, suggesting that rapid disease progression limits vaccination in patients with relapse disease, thus limiting clinical translation. We therefore sought to identify a window of time for vaccination during which this approach might be feasible. To achieve this end, we evaluated immunological reconstitution in newly diagnosed patients with ALL patients. Despite recovery of myelopoiesis, most patients had profound defects in T, B, and natural killer (NK) cell numbers that failed to recover at any point during therapy. CONCLUSION: Autologous tumor vaccination at a time of ALL relapse is not feasible. Alternative strategies for immunotherapy of ALL may require ex vivo generation of antigen specific T cells and adoptive therapy.

Adolescent↗

PubMed2005

Interleukin-7 in T-cell acute lymphoblastic leukemia: an extrinsic factor supporting leukemogenesis?

The malignant transformation and expansion of tumor cells involve both cell-autonomous mechanisms and microenvironment signals that regulate viability, nutrient utilization, metabolic activity and cell growth. In T-cell acute lymphoblastic leukemia (T-ALL), the co-culture of leukemic cells with stroma or the addition of particular cytokines prevents ex vivo spontaneous apoptosis. Interleukin-7 (IL-7), a cytokine produced by thymic and bone marrow stroma, increases the viability and proliferation of T-ALL cells. IL-7 induces the activation of Jak/STAT, MEK/Erk and PI3K/Akt signaling pathways in T-ALL cells. PI3K/Akt is the dominant pathway that mediates the effects of IL-7 on T-ALL. PI3K signaling is required for the induction of Bcl-2, the down-regulation of p27(kip1) and cell cycle progression. PI3K signaling is also required for the expression of the glucose transporter Glut1, uptake of glucose, activation of the metabolic machinery, increase in cell size, and maintenance of mitochondrial integrity. These observations suggest that substrates of molecular pathways activated by microenvironmental factors represent attractive molecular targets for the regulation of the viability and proliferation of T-ALL cells and provide the means for the development of novel treatment strategies.

Animals↗

PubMed2004

Activation of PI3K is indispensable for interleukin 7-mediated viability, proliferation, glucose use, and growth of T cell acute lymphoblastic leukemia cells.

Interleukin (IL)-7 is essential for normal T cell development. Previously, we have shown that IL-7 increases viability and proliferation of T cell acute lymphoblastic leukemia (T-ALL) cells by up-regulating Bcl-2 and down-regulating the cyclin-dependent kinase inhibitor p27kip1. Here, we examined the signaling pathways via which IL-7 mediates these effects. We investigated mitogen-activated protein kinase (MEK)-extracellular signal-regulated kinase (Erk) and phosphatidylinositol-3-kinase (PI3K)-Akt (protein kinase B) pathways, which have active roles in T cell expansion and have been implicated in tumorigenesis. IL-7 induced activation of the MEK-Erk pathway in T-ALL cells; however, inhibition of the MEK-Erk pathway by the use of the cell-permeable inhibitor PD98059, did not affect IL-7-mediated viability or cell cycle progression of leukemic cells. IL-7 induced PI3K-dependent phosphorylation of Akt and its downstream targets GSK-3, FOXO1, and FOXO3a. PI3K activation was mandatory for IL-7-mediated Bcl-2 up-regulation, p27kip1 down-regulation, Rb hyperphosphorylation, and consequent viability and cell cycle progression of T-ALL cells. PI3K signaling was also required for cell size increase, up-regulation of CD71, expression of the glucose transporter Glut1, uptake of glucose, and maintenance of mitochondrial integrity. Our results implicate PI3K as a major effector of IL-7-induced viability, metabolic activation, growth and proliferation of T-ALL cells, and suggest that PI3K and its downstream effectors may represent molecular targets for therapeutic intervention in T-ALL.

Antigens, CD↗

PubMed2004

pH-triggered microparticles for peptide vaccination.

Improving vaccine delivery to human APCs is a way to increase the CTL response to vaccines. We report the use of a novel pH-triggered microparticle that exploits the ability of APCs to cross-present MHC I-restricted Ags that have been engulfed in the low pH environment of the phagosome. A model MHC class I-restricted peptide Ag from the influenza A matrix protein was encapsulated in spray-dried microparticles composed of dipalmitoylphosphatidylcholine and the pH-sensitive polymethacrylate Eudragit E100. Release of the peptide from the particle was triggered by a drop in pH to the acidity normally found in the phagosome. The particles were efficiently phagocytosed by human monocytes and dendritic cells with minimal cellular toxicity and no functional impairment. Encapsulation of the peptide in the microparticles resulted in efficient presentation of the peptide to CD8(+) T cells by human dendritic cells in vitro, and was superior to unencapsulated peptide or peptide encapsulated in an analogous pH-insensitive particle. Vaccination of human HLA-A*0201 transgenic mice with peptide encapsulated in pH-triggering microparticles resulted in priming of CTL responses. These microparticles can be modified to coencapsulate a range of adjuvants along with the Ag of interest. Encapsulation of MHC I epitopes in pH-triggered microparticles increases Ag presentation and may improve CD8(+) T cell priming to peptide vaccines against viruses and cancer.

1,2-Dipalmitoylphosphatidylcholine↗

PubMed2004

Phylogenetic footprinting reveals extensive conservation of Sonic Hedgehog (SHH) regulatory elements.

Sonic Hedgehog (SHH) plays a fundamental role in numerous developmental processes including morphogenesis of limbs, nervous system, and teeth. Using a Bayesian alignment algorithm for phylogenetic footprinting we analyzed approximately 28 kb of noncoding DNA in the SHH locus of human and mouse. This showed that the length of conserved noncoding sequences (4196 nt) shared by these species was approximately 3 times larger than the SHH coding sequence (1386 nt). Most segments were located in introns (53%) or within 2-kb regions upstream (16%) or downstream (20%) of the first and last SHH codon. Even though regions more than 2 kb upstream or downstream of the first and last SHH codon represented 57% (16 kb) of the sequence compared, they accounted for only 11% (494 nt) of the total length of conserved noncoding segments. One region of 650 nt downstream of SHH was identified as a putative scaffold/matrix attachment region (SMAR). Human-mouse analysis was complemented by sequencing in apes, monkeys, rodents, and bats, thus further confirming the evolutionary conservation of some segments. Gel-shift assays indicated that conserved segments are targeted by nuclear proteins and showed differences between two cell types that expressed different levels of SHH, namely human endothelial cells and breast cancer cells. The relevance of these findings with respect to regulation of SHH expression during normal and pathologic development is discussed.

Animals↗

PubMed2004

Common gamma chain-signaling cytokines promote proliferation of T-cell acute lymphoblastic leukemia.

BACKGROUND AND OBJECTIVES: The identification of signals critical for the pathophysiology of T-cell acute lymphoblastic leukemia (T-ALL) should contribute to the development of novel, more effective therapeutic strategies. Common gamma-chain signaling cytokines (gammac-cytokines) - interleukins 2, 4, 7, 9 and 15 - differentially regulate T-cell development, survival, proliferation and differentiation. Although studies exist on some individual cytokines, no comprehensive analysis of the effects of the Zc-cytokine family on malignant T cells has been reported. Here, we examined the effect of Zc-cytokines on T-ALL proliferation. DESIGN AND METHODS: Primary leukemic cells were collected at diagnosis from the blood or bone marrow of children with T-ALL. The cells were immunophenotyped and classified according to maturation stage. Proliferative responses to gammac-cytokines were assessed by 3H-thymidine incorporation. RESULTS: All gammac-cytokines promoted proliferation of primary T-ALL cells. Interleukin (IL)-7 was the cytokine that most frequently induced leukemic cell proliferation and promoted the most robust responses. IL-4 preferentially stimulated proliferation of samples with a more mature immunophenotype, whereas CD1a-positive cortical T-ALL cells were less responsive to IL-9. Finally, combinations of two Zc-cytokines showed synergistic or additive proliferative effects. INTERPRETATION AND CONCLUSIONS: This study indicates that all the gammac-cytokines tested can stimulate proliferation of leukemic T cells and suggests that synergistic effects may occur in vivo. We present the first demonstration that IL-9 and IL-15 can provide a proliferative signal to T-ALL cells. Importantly, our results support the hypothesis that IL-7 may function as a critical regulator of T-ALL and that its activity may be potentiated by other Zc-cytokines.

Autocrine Communication↗

PubMed2003

IL-7-dependent human leukemia T-cell line as a valuable tool for drug discovery in T-ALL.

The specific targeting of critical signaling molecules may provide efficient therapies for T-cell acute lymphoblastic leukemia (T-ALL). However, target identification and drug development are limited by insufficient numbers of primary T-ALL cells and by their high rate of spontaneous apoptosis. We established a human interleukin-7 (IL-7)-dependent T-ALL cell line, TAIL7, that maintains several biologic and signaling properties of its parental leukemia cells. TAIL7 cells are pre-T-ALL cells that proliferate in response to IL-7 and IL-4. IL-7 stimulation of TAIL7 cells prevents spontaneous in vitro apoptosis and induces cell activation and cell cycle progression. The signaling events triggered by IL-7 include down-regulation of p27(kip1) and hyperphosphorylation of retinoblastoma protein (Rb). Stimulation of TAIL7 cells by IL-7 leads to phosphorylation of Janus kinase 3 (JAK3), signal transducer and activator of transcription 5 (STAT5), Akt/PKB (protein kinase B), and extracellular-regulated kinase 1 and 2 (Erk1/2). Importantly, specific blockade of JAK3 by its inhibitor WHI-P131 abrogates the IL-7-mediated proliferation and survival of TAIL7 cells, suggesting that activation of JAK3 is critical for IL-7 responsiveness by these cells. Because TAIL7 cells seem to be a biologic surrogate for primary leukemia T cells, this cell line constitutes a valuable tool for the study of the signaling pathways implicated in T-ALL. Exploitation of this cell line should allow the identification of molecular targets and promote the rational design and validation of antileukemia signaling inhibitors.

Animals↗

PubMed2002

HER-2/neu and hTERT cryptic epitopes as novel targets for broad spectrum tumor immunotherapy.

Tolerance to tumor-nonmutated self proteins represents a major obstacle for successful cancer immunotherapy. Since this tolerance primarily concerns dominant epitopes, we hypothesized that targeting cryptic epitopes that have a low affinity for HLA could be an efficient strategy to breach the tolerance to tumor Ags. Using the P1Y heteroclitic peptide approach, we identified low affinity cryptic HLA-A*0201-restricted epitopes derived from two widely expressed tumor Ags, HER-2/neu and hTERT. The P1Y variants of four HER-2/neu (neu(391), neu(402), neu(466), neu(650))- and two hTERT (hTERT(572) and hTERT(988))-derived low affinity peptides exhibited strong affinity for HLA-A*0201 and stimulated specific CTL from healthy donor PBMCs. These CTL specifically recognized HER-2/neu- and hTERT-expressing tumor cells of various histological origins. In vivo studies showed that HLA-A*0201 transgenic HHD mice vaccinated with the P1Y variant peptides generated CTL that specifically lysed Ag-expressing tumor cells, thus recognizing the cognate endogenous Ags. These results suggest that heteroclitic variants of low affinity, cryptic epitopes of widely expressed tumor Ags may serve as valid tools for tumor immunotherapy.

Animals↗

PubMed2002

CXCL13 (BCA-1) is produced by follicular lymphoma cells: role in the accumulation of malignant B cells.

Follicular lymphomas (FLs) localize in lymphoid tissues and recapitulate the structure of normal secondary follicles. The chemokine/chemokine receptor pair CXCL13/CXCR5 is required for the architectural organization of B cells within lymphoid follicles. In this study, we showed that CXCL13 was secreted by FL cells. FL cells expressed CXCR5 and migrated in response to CXCL13. Furthermore, we observed a synergistic effect between CXCL13 and CXCL12 (SDF-1), a chemokine produced by stromal cells in lymphoid tissues. The production of CXCL13 by FL cells and CXCL12 by stromal cells probably directs and participates in the accumulation of FL cells within specific anatomic sites.

Apoptosis↗