transforming-growth-factor-beta and Hodgkin-Disease

Advances in understanding of the immune microenvironment have highlighted the role of immunosuppressive T cell, myeloid, dendritic and monocytic sub-populations in inhibition of the anti-tumor immune response. The role of B cells in modulating the immune response to solid tumors as well as lymphoid malignancies is less well understood. Murine models of autoimmune disease have defined B regulatory cell (Breg) subsets with immune suppressive activity, including B cell subsets that express IL-10, and transforming growth factor-β, which can facilitate T regulatory cell recruitment and expansion. Multiple murine tumor models point to the existence of similar immune suppressive B cell sub-populations that can migrate into tumor deposits and acquire an immune suppressive phenotype, which then leads to attenuation of the local anti-tumor immune response. Other murine models of viral or chemically induced skin carcinogenesis have identified a pivotal role for B cells in promoting inflammation and carcinogenesis. While many human solid tumors demonstrate significant B cell infiltration and/or tertiary lymphoid structure formation, the functional properties of tumor-infiltrating B cells and their effects on immunity are poorly understood. Recent successes in early Phase I/II trials using anti-checkpoint inhibitor antibodies such as nivolumab or pidilizumab directed against PD-1 in the setting of Hodgkin's and non-Hodgkin's lymphomas validate the therapeutic utility of reversing B cell-mediated immune suppression. Further studies to define Breg subsets, and mechanisms of suppression, may provide new avenues for modulation of the immune response and meaningful therapeutic intervention in both lymphoid and solid tumors.

Topics: Animals; Autoimmune Diseases; B-Lymphocytes, Regulatory; Cell Lineage; Clinical Trials as Topic; Disease Models, Animal; Gene Expression Regulation, Neoplastic; Hodgkin Disease; Humans; Immune Tolerance; Interleukin-10; Lymphoma, Non-Hodgkin; Mice; Signal Transduction; T-Lymphocytes, Regulatory; Transforming Growth Factor beta; Tumor Microenvironment

Recent studies provide evidence that Reed-Sternberg (R-S) cells produce factors that may explain the characteristic inflammatory infiltrate in the affected tissues of Hodgkin lymphoma. The various chemokines and cytokines that are produced lead to a preferential influx of Th2-type T cells and suppress Th1-type immune responses. Overall, the immunophenotype of the lymphocytes surrounding the R-S cells is consistent with anergic and/or Th2-type T cells. Therefore, these cells do not support a cytotoxic anti-tumor response. Since the R-S cells are neoplastic B cells, the cytokines produced by these T cells may in fact help their growth and/or survival. The production and induction of various other cytokines may also explain the influx of eosinophils (IL-5, eotaxin) and plasma cells (IL-6). Differences in chemokine and cytokine production may be responsible for the differences between the histological subtypes.

Topics: Cell Communication; Chemokines; Cytokines; Hodgkin Disease; Humans; Immunophenotyping; Interleukin-10; Receptors, Chemokine; Reed-Sternberg Cells; T-Lymphocytes; Transforming Growth Factor beta

Lymphomatoid papulosis and cutaneous CD30+ lymphoma are closely related conditions in which large atypical lymphocytes that have similar immunophenotypic features occur. In lymphomatoid papulosis, the lesions are papules and nodules that spontaneously involute. There are two polar histologic patterns, type A and B, in which the large atypical cells resemble those of Hodgkin's disease and mycosis fungoides, respectively, but in many cases, features of both types are present, either separately or in the same lesions. Variants of lymphomatoid papulosis include cases with a perifollicular distribution and those with lymphocytic vasculitis or dermal mucin deposits. Clinical lesions that tend to be stable, a monomorphous cellular composition, and in the case of immunocompromised patients, the presence of Epstein-Barr viral genome characterize cutaneous CD30+ lymphoma. A loss of response to transforming growth factor-beta, which normally dampens cellular proliferation, may differentiate CD30+ lymphoma from lymphomatoid papulosis.

Topics: Cell Division; Genome, Viral; Hair Follicle; Herpesvirus 4, Human; Hodgkin Disease; Humans; Immunocompromised Host; Immunophenotyping; Lymphocytes; Lymphoma, Large-Cell, Anaplastic; Lymphomatoid Papulosis; Mucins; Mycosis Fungoides; Skin Neoplasms; Transforming Growth Factor beta; Vasculitis, Leukocytoclastic, Cutaneous

The underlying mechanism of fibrosis in classical Hodgkin lymphoma (CHL) remains uncertain. This study aimed to investigate the association of fibrosis in the lymph nodes of patients with CHL through histological examination of the expression of cytokines associated with fibrosis and mast cell proliferation. Additionally, we sought to determine the degree of mast cell infiltration in a nodular sclerosis subtype of CHL (NSCHL) compared with that in non-NSCHL. We analyzed lymph nodes from 22 patients with CHL, of which eight were of the NSCHL and 14 of the non-NSCHL subtype, using immunohistochemical staining of forkhead box P3 (FOXP3), transforming growth factor (TGF)-β, interleukin (IL)-3, IL-13, and stem cell factor (SCF). Mast cells were positive for TGF-β and IL-13, and FOXP3-positive cells were negative for TGF-β. Only the expression of IL-13 in Hodgkin and Reed-Sternberg (HRS) cells was significantly more frequently observed in NSCHL than that in non-NSCHL (P = 0.0028) and was associated with a higher rate of fibrosis (P = 0.0097). The number of mast cells was significantly higher in NSCHL than that in non-NSCHL (P = 0.0001). A significantly positive correlation was observed between the rate of fibrosis and the number of mast cells (correlation coefficient, 0.8524; 95% CI, 0.6725-0.9372) (P <0.0001). The number of mast cells was significantly higher in the group with IL-13-positive HRS cells than that in the group with IL-13-negative HRS cells (P = 0.0157). Based on these findings, we hypothesize that IL-13 production by HRS cells may lead to fibrosis, and furthermore, promote mast cell proliferation and infiltration. This in turn might further produce the fibrotic cytokines IL-13 and TGF-β, resulting in fibrosis typical of NSCHL.

Topics: Adolescent; Adult; Aged; Aged, 80 and over; Female; Fibrosis; Hodgkin Disease; Humans; Interleukin-13; Interleukin-3; Lymph Nodes; Male; Mast Cells; Middle Aged; Reed-Sternberg Cells; Transforming Growth Factor beta; Young Adult

Abstract In a phase 2 trial of panobinostat in 129 patients with relapsed or refractory Hodgkin lymphoma, exploratory analyses of chemokines and cytokines were prospectively performed in 109 patients to determine their association with clinical outcomes. Patients were categorized into two groups (reductions > median and reductions ≤ median) based on percentage change from baseline of log10 transformed measurements. Thymus and activation-regulated chemokine (TARC) was most strongly associated with clinical outcome. Early reduction of TARC was observed in responding patients, with the greatest reduction at cycle 1, day 15 (C1D15). Of 93 patients with C1D15 samples, there were three complete and 25 partial responses. The group with TARC reductions > median at C1D15 had more responders (18 [39%] vs. 10 [21%]), longer progression-free survival (10.6 vs. 4.9 months), shorter time to response and longer overall survival than the group with reductions ≤ median. This study is registered at www.ClinicalTrials.gov , NCT00742027.

Topics: Adolescent; Adult; Aged; Antineoplastic Agents; Biomarkers; Chemokine CCL17; Combined Modality Therapy; Female; Hematopoietic Stem Cell Transplantation; Hodgkin Disease; Humans; Hydroxamic Acids; Indoles; Male; Middle Aged; Neoplasm Recurrence, Local; Neoplasm Staging; Panobinostat; Time Factors; Transforming Growth Factor beta; Transplantation, Autologous; Treatment Outcome; Tumor Burden; Young Adult

Neoplastic processes, tumor growth, and tumor cell proliferation and survival are often due to the altered activation of different signaling pathways. The increased activity of PI3K/AKT/mTOR signaling has been shown to be an important regulator of tumor growth in several solid tumors and in mantle cell lymphomas. The active form of mTOR kinase (mammalian target of rapamycin) is a key signaling molecule, and it exists in two different complexes, mTORC1 and mTORC2. In the present work, mTOR activity was investigated in different lymphoma types, in parallel with clinical data. We also examined in Hodgkin lymphomas (HL) the role of mTOR activity in survival mechanisms such as antiapoptotic protein expression and alterations in the microenvironment. We determined which lymphoma types display characteristic high mTOR activity in our TMA (tissue microarray) study. We observed that mTOR activity is increased in mitotic lymphoid cells compared to interphasic cells. The number of diffuse large B cell lymphoma (DLBCL) and HL cases was extended in a further set of TMA. We observed significantly higher mTOR activity in the non-centrum germinativum derived subtype of DLBCL than in the centrum germinativum derived subtype, which was a prognostic marker; 63% of mTOR active cases showed Rictor overexpression, indicating mTORC2 activity. High mTOR activity was also established in 92% of HL cases, which was linked to mTORC1. This finding was not a prognostic marker, however, it can be useful in targeted therapy. We observed the overexpression of the antiapoptotic protein BCL-xL and NFκB-p50 in the majority of mTOR active HLs. HLs showed high numbers of regulatory T cells in the microenvironment and high expression of galectin-1 in tumor cells and in the extracellular matrix, when compared to reactive lymph nodes. We confirmed that mTOR inhibition had significant antiproliferative and antiapoptotic effects in lymphoma cell lines and in lymphoma xenografts (HL, DLBCL, Burkitt lymphoma). We also showed that rapamycin was able to augment the effect of chemotherapeutic agents and TGF-β. Taken together, mTOR activity may be a potential therapeutic target in different lymphoma types. However, patient and inhibitor selection criteria must be carefully considered. The combination of mTOR inhibitors with other agents will probably offer the highest efficiency for achieving the best clinical response, and may also allow dose reduction in order to decrease late treatment toxicity in th. A neopláziás folyamatok kialakulása, a daganat növekedése, a daganatsejtek proliferációja és túlélése hátterében gyakran különbözõ jelutak magváltozott aktivitása áll. A PI3K/AKT/mTOR jelút fokozott aktivitása szolid daganatokban és köpenysejtes lymphomákban a daganatkialakulás és -növekedés fontos szabályozója. Az aktív mTOR (mammalian target of rapamycin) kináz két komplex (mTORC1, mTORC2) meghatározó eleme. Az mTOR-aktivitás szerepét vizsgáltuk különbözõ humán lymphomákban, összefüggéseket keresve a betegek klinikai adataival. Hodgkin-lymphomákban (HL) tanulmányoztuk, hogy a magas mTOR-aktivitás milyen, a daganat túlélésében fontos folyamatokban vesz részt (antiapoptotikus mechanizmusok és mikrokörnyezeti változások). Meghatároztuk azokat a humán lymphomatípusokat, amelyekre magas mTOR-aktivitás jellemzõ. Kimutattuk, hogy a mitotikus lymphoid sejtek mTOR-aktivitása magasabb, mint a nem osztódó sejteké. Nagyobb esetszámot tartalmazó TMA-blokkokon (tissue microarray) tovább vizsgáltuk a diffúz nagy B-sejtes lymphoma (DLBCL) és a HL eseteket. Szignifikáns összefüggést mutattunk ki DLBCL-s betegek altípusmegoszlása (csíraközpont-eredetû és nem csíraközpont-eredetû DLBCL-ek) és az mTOR-aktivitás között. DLBCL-ban a fokozott mTOR-aktivitás negatív prognosztikus markernek bizonyult. A HL-ek 92%-a magas mTOR-aktivitást mutatott (mTORC1-hez köthetõ), ami prognosztikus faktorként nem, viszont terápiás célpontként felhasználható. A HL-ek mikrokörnyezetének vizsgálata szerint a regulátor T-sejtek mennyisége a mikrokörnyezetben, valamint a galektin-1-expresszió a tumorsejtekben és az extracelluláris mátrixban emelkedett. A magas mTOR-aktivitás és a galektin-1-expresszió között kapcsolatot találtunk in vitro kísérleteinkben, ahol az mTOR gátlása transzlációs szinten csökkentette a galektin-1-expressziót. Az mTOR-gátlás jelentõségét – proliferációgátló és apoptotikus hatását – humán lymphoma xenograftokban (HL, DLBCL, Burkitt-lymphoma) bizonyítottuk. In vitro kombinációs kezelésekben a rapamycin apoptotikus hatást fokozó szerepét igazoltuk. Munkánkban meghatároztuk azokat a lymphomatípusokat, amelyekben az mTOR-gátlás célzott terápiaként alkalmazható lehet. Eredményeink alapján annak meghatározása, hogy melyik komplexhez köthetõ az mTOR-aktivitás, nagyon fontos a megfelelõ mTOR-gátló (klasszikus vagy kettõs gátlók) kiválasztásában. A jövõben a magas mTOR-aktivitást mutató lymphomákban várhatóan kombinációs kezelésben az mTOR-gátlók használata hozzájárulhatna a jobb t

Topics: Animals; Antineoplastic Agents; Apoptosis; Apoptosis Regulatory Proteins; bcl-X Protein; Burkitt Lymphoma; Cell Line; Drug Synergism; Galectin 1; Gene Expression Regulation, Neoplastic; Hodgkin Disease; Humans; Immunosuppressive Agents; Interphase; Lymphoma; Lymphoma, Large B-Cell, Diffuse; Mechanistic Target of Rapamycin Complex 1; Mechanistic Target of Rapamycin Complex 2; Mitosis; Multiprotein Complexes; NF-kappa B p50 Subunit; Signal Transduction; Sirolimus; T-Lymphocytes, Regulatory; Tissue Array Analysis; TOR Serine-Threonine Kinases; Transforming Growth Factor beta; Up-Regulation; Xenograft Model Antitumor Assays

FOX genes encode transcription factors which regulate basic developmental processes during embryogenesis and in the adult. Several FOX genes show deregulated expression in particular malignancies, representing oncogenes or tumor suppressors. Here, we screened six Hodgkin lymphoma (HL) cell lines for FOX gene activity by comparative microarray profiling, revealing overexpression of FOXC1 and FOXD1, and reduced transcription of FOXN3, FOXO1, and FOXP1. In silico expression analyses of these FOX gene candidates in HL patient samples supported the cell line data. Chromosomal analyses demonstrated an amplification of the FOXC1 locus at 6p25 and a gain of the FOXR2 locus at Xp11, indicting genomic aberrations for their upregulation. Comparative expression profiling and ensuing stimulation experiments revealed implementation of the TGFβ- and WNT-signaling pathways in deregulation of FOXD1 and FOXN3. Functional analysis of FOXP1 implicated miR9 and miR34a as upstream regulators and PAX5, TCF3, and RAG2 as downstream targets. A similar exercise for FOXC1 revealed repression of MSX1 and activation of IPO7, both mediating inhibition of the B-cell specific homeobox gene ZHX2. Taken together, our data show that aberrantly expressed FOX genes and their downstream targets are involved in the pathogenesis of HL via deregulation of B-cell differentiation and may represent useful diagnostic markers and/or therapeutic targets.

Topics: Burkitt Lymphoma; Cell Line, Tumor; Chromosomes, Human; Forkhead Transcription Factors; Gene Expression Profiling; Gene Expression Regulation, Neoplastic; Gene Regulatory Networks; Genetic Loci; Hodgkin Disease; Homeodomain Proteins; Humans; Karyopherins; Lymphoma, B-Cell; Lymphoma, Follicular; MicroRNAs; MSX1 Transcription Factor; Receptors, Cytoplasmic and Nuclear; Transcription Factors; Transforming Growth Factor beta; Wnt Signaling Pathway

Herein we describe that in classic Hodgkin lymphomas (cHL, n = 25) the lymph node (LN) stroma displayed in situ high levels of transcription and expression of the disulfide-isomerase ERp5 and of the disintegrin-metalloproteinase ADAM10, able to shed the ligands for NKG2D (NKG2D-L) from the cell membrane. These enzymes were detected both in LN mesenchymal stromal cells (MSCs) and in Reed-Sternberg (RS) cells; in addition, MIC-A and ULBP3 were present in culture supernatants of LN MSCs or RS cells. NKG2D-L-negative RS cells could not be killed by CD8(+)αβT or γδT cells; tumor cell killing was partially restored by treating RS cells with valproic acid, which enhanced NKG2D-L surface expression. Upon coculture with LN MSCs, CD8(+)αβT and γδT cells strongly reduced their cytolytic activity against NKG2D-L(+) targets; this seems to be the result of TGF-β, present at the tumor site, produced in vitro by LN MSCs and able to down-regulate the expression of NKG2D on T lymphocytes. In addition, CD8(+)αβT and γδT cells from the lymph nodes of cHL patients, cocultured in vitro with LN MSCs, underwent TGF-β-mediated down regulation of NKG2D. Thus, in cHL the tumor microenvironment is prone to inhibit the development of an efficient antitumor response.

Topics: ADAM Proteins; ADAM10 Protein; Adult; Aged; Amyloid Precursor Protein Secretases; Cells, Cultured; Coculture Techniques; Female; Fluorescent Antibody Technique; Gene Expression Regulation, Neoplastic; Hodgkin Disease; Humans; Lymph Nodes; Male; Membrane Proteins; Mesenchymal Stem Cells; Middle Aged; NK Cell Lectin-Like Receptor Subfamily K; Protein Disulfide-Isomerases; Receptors, Antigen, T-Cell, alpha-beta; Receptors, Antigen, T-Cell, gamma-delta; Reed-Sternberg Cells; Reverse Transcriptase Polymerase Chain Reaction; T-Lymphocytes; Transforming Growth Factor beta; Tumor Microenvironment; Young Adult

This study was aimed to analyze the relationship between single nucleotide polymorphisms of transforming growth factor-β1 G-800A and C-509T, interleukin-4 receptor V75I and susceptibility of CHL in adults.. Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) was applied to analyze the expressed alleles of the selected SNP loca. The relationship between genomic polymorphisms of TGF-β1 and IL-4R and susceptibility of CHL were coupled with clinical data.. TGF-β1G-800A and TGF-β1C-509T had obvious linkage equilibrium (D' = 0.879, r(2) = 0.83, P = 0.020). GT haplotype distribution frequencies in mixed cellularity Hodgkin lymphoma cases and control group were of 53.1% and 34.2%, respectively, with statistically significant (OR = 2.35, P = 0.000); distribution frequencies of mutant gene T/T in disease and control groups were of 38.8% and 15.3%, respectively, also with statistically significant (OR = 3.654, P = 0.000); frequencies of nodular sclerosis CHL patients with IL-4R V75I mutant gene A/A in disease and control groups were of 19.2% and 41.75%, respectively, also with statistically significant (OR = 3.156, P = 0.000).. Single nucleotide polymorphisms of TGF-β1 G-800A, C-509T and IL-4R V75I has a significant correlation with Chinese susceptibility to classical Hodgkin lymphoma.

Topics: Adult; Alleles; Asian People; Female; Genotype; Haplotypes; Hodgkin Disease; Humans; Male; Middle Aged; Polymorphism, Restriction Fragment Length; Polymorphism, Single Nucleotide; Receptors, Interleukin-4; Transforming Growth Factor beta; Young Adult

Mesenchymal stem cells (MSCs) have received much attention because of their capabilities of differentiating into multiple mesenchymal lineages and supporting hematopoiesis. Recently, MSCs have gained further interests after the demonstration of an immunosuppressive role. However, it's still unclear whether the immunosuppressive capability of MSCs will be altered with disease state. In this study, our results showed that MSCs derived from patients with lymphoblastic leukemia (ALL), Hodgkin disease (HD), and non-Hodgkin lymphoma (NHL) capable of suppressing the proliferation of T-lymphocyte stimulated in a mixed-lymphocyte reaction (MLR). The immunosuppressive effect of MSCs derived from ALL, HD and NHL on T-cell proliferation was dose-dependent. The supernatants of MSCs derived from ALL, HD and NHL had effect on T-cell proliferation. By using neutralising monoclonal antibodies, we found that transforming growth factor beta1 (TGFbeta1) and hepatocyte growth factor were major mediators of T-cell suppression by MSCs derived from ALL, HD and NHL. Although MSCs derived from patients with myelodysplastics syndromes (MDS) could inhibit T-cell proliferation stimulated with mitogen or in MLR, the inhibitory effect of MDS-MSCs was impaired. However, adherent cells derived from patients with acute myeloid leukemia (AML) showed abnormal immunomodulatory functions. Adherent cells derived from AML failed to suppress the proliferation of T-cell stimulated in MLR.

Topics: Bone Marrow Cells; Hematologic Neoplasms; Hepatocyte Growth Factor; Hodgkin Disease; Humans; Immunologic Factors; Immunosuppression Therapy; Lymphocyte Culture Test, Mixed; Lymphoma, Non-Hodgkin; Mesenchymal Stem Cells; Precursor Cell Lymphoblastic Leukemia-Lymphoma; T-Lymphocytes; Transforming Growth Factor beta

The Epstein-Barr virus (EBV) contributes to the growth and survival of Hodgkin lymphoma (HL) cells. Here we report that down-regulation of the transforming growth factor-beta (TGF-beta) target gene, protein tyrosine phosphatase receptor kappa (PTPRK), followed EBV infection of HL cells and was also more frequently observed in the Hodgkin and Reed-Sternberg (HRS) cells of EBV-positive compared with EBV-negative primary HL. The viability and proliferation of EBV-positive HL cells was decreased by overexpression of PTPRK, but increased following the knockdown of PTPRK expression in EBV-negative HL cells, demonstrating that PTPRK is a functional tumor suppressor in HL. EBV suppressed the TGF-beta-mediated activation of PTPRK expression, suggesting disruption of TGF-beta signaling upstream of PTPRK. This was confirmed when we showed that the Epstein-Barr nuclear antigen-1 (EBNA1) decreased Smad2 protein levels and that this was responsible for PTPRK down-regulation. EBNA1 decreased the half-life of Smad2 but did not interact with Smad2. By down-regulating Smad2 protein expression, EBNA1 apparently disables TGF-beta signaling, which subsequently decreases transcription of the PTPRK tumor suppressor. We speculate that loss of the phosphatase function of PTPRK may activate as-yet-unidentified growth-promoting protein tyrosine kinases, which in turn contribute to the pathogenesis of EBV-positive HL.

Topics: Cell Division; Cell Line; Cell Survival; Down-Regulation; Epstein-Barr Virus Infections; Epstein-Barr Virus Nuclear Antigens; Female; Gene Expression Regulation, Neoplastic; Gene Expression Regulation, Viral; Genes, Tumor Suppressor; Herpesvirus 4, Human; Hodgkin Disease; Humans; Male; Receptor-Like Protein Tyrosine Phosphatases, Class 2; Smad2 Protein; Transforming Growth Factor beta

A hallmark of various human malignancies is the expression of immunoinhibitory factors within the tumor microenvironment. There is indirect evidence based on in vitro experiments that tumor-infiltrating T cells in human malignancies are suppressed by such factors. Still, direct evidence of the influence of individual inhibitory factors on immune cells in human cancer in vivo is lacking. To address this question, we used Hodgkin lymphoma (HL) as a model because histopathological characteristics of HL are thought to be due mostly to the effects of a wide variety of cytokines, including TGFbeta or membrane-bound receptors such as PD-1 that are suspected to contribute to immune evasion of tumor cells. Using a genome-wide transcriptional approach, we established specific RNA fingerprints of TGFbeta and PD-1 signaling in human T cells in vitro. Applying these specific fingerprints, we directly demonstrate that CD4+ T cells in HL--but not in follicular lymphoma (FL)--are under the inhibitory influence of both TGFbeta and PD-1 in vivo. This approach can be easily generalized to provide direct evidence of the impact of any given soluble or cell-bound factor on any cell type within diseased tissue.

Topics: Antigens, CD; Apoptosis Regulatory Proteins; CD4-Positive T-Lymphocytes; Cluster Analysis; Female; Gene Expression Profiling; Gene Expression Regulation, Neoplastic; Hodgkin Disease; Humans; In Vitro Techniques; Male; Nucleotide Mapping; Oligonucleotide Array Sequence Analysis; Programmed Cell Death 1 Receptor; RNA; Transforming Growth Factor beta

Adoptive immunotherapy with Epstein-Barr virus (EBV)-specific cytotoxic T cells (CTL) is effective for the prophylaxis and treatment of EBV-induced lymphoma in hematopoietic stem cell recipients. However, in EBV-positive Hodgkin's disease (HD) the efficacy of adoptively transferred EBV-specific CTL may be limited by tumor-derived immunosuppressive factors, such as T-cell growth factor (TGF) beta, interleukin (IL)13 and the chemokine TARC. Local delivery of IL12 to tumor sites by tumor-specific CTL could provide direct antitumor effects and overcome the CTL-inhibitory effects of the Th2 tumor environment while avoiding the systemic toxicity of recombinant IL12. EBV-specific CTL transduced with a retrovirus vector expressing the p40 and p35 subunits of IL12 as a single molecule (Flexi-IL12), produced IL12 following antigenic stimulation. This resulted in an elevated production of Th1 cytokines, including interferon gamma and tumor necrosis factor alpha, and a reduction in the Th2 cytokines IL4 and IL5. Flexi-IL12-transduced CTL resisted the antiproliferative and anticytotoxic effects of exogenous TGFbeta, likely by antagonizing the TGFbeta-induced downregulation of the Th1 transcriptional factor T-bet. In addition, Flexi-IL12-transduced CTL demonstrated a proliferative advantage in the presence of inhibitory supernatants from HD-derived cell lines. Tumor-specific, Flexi-IL12-transduced EBV-specific CTL should have a functional advantage over unmodified CTL, particularly in the presence of the adverse Th2 cytokine environment produced by Hodgkin tumor cells.

Topics: Antigens, Neoplasm; Cell Line; Cytokines; Gene Expression; Genetic Vectors; Herpesvirus 4, Human; Hodgkin Disease; Humans; Immunotherapy, Adoptive; Interleukin-12; Retroviridae; T-Lymphocytes, Cytotoxic; Transforming Growth Factor beta; Vaccines, Synthetic

We assessed mRNA for chosen pro- and anti-inflammatory cytokines in T-lymphocytes of peripheral blood in paediatric patients with leukemias and lymphomas. Levels of four different cytokine mRNAs (IFN-gamma, IL-10, IL-4, TGF-beta) were determined by the real-time PCR technique. In the whole examined group, at the time of diagnosis, we noted lower amounts of mRNA for TGF-beta1, comparing to respective values in the control patients. In the ALL group, we observed the following: 1) at the time of diagnosis: lower amounts of mRNA for IL-4 and for TGF-beta1, comparing to respective values in the control group; 2) lower amounts of mRNA for IL-10 after remission induction, comparing to the time of diagnosis. In our opinion, "immunedysregulation" in lymphoproliferative diseases in children is not caused by IFN-gamma deficiency. The deficit of anti-inflammatory cytokines, i.e., IL-4, TGF-beta, with higher amounts of IL-10, suggests their role in cancer development.

Topics: CD4-Positive T-Lymphocytes; CD8-Positive T-Lymphocytes; Cytokines; Hodgkin Disease; Humans; Interleukin-10; Lymphoma; Lymphoma, Non-Hodgkin; Polymerase Chain Reaction; Precursor Cell Lymphoblastic Leukemia-Lymphoma; Remission Induction; RNA, Messenger; T-Lymphocytes; Transforming Growth Factor beta

To characterize Reed-Sternberg (R-S) cells by proteomic analysis in order to gain insight into the molecular pathways that control their growth and thereby to discern potential molecular interventions in Hodgkin's disease.. Ten cases of the nodular sclerosing (NS) subtype and 4 cases of the lymphocyte-predominant (LP) subtype were studied. Immunohistochemical procedures were performed to detect the following antigens: CD20, CD30, c-kit, platelet-derived growth factor receptor (PDGFR)-alpha, cathepsin D, angiotensin-converting enzyme (ACE), angiotensin II type 1 (AT1) receptor, phosphorylated c-Jun N-terminal kinase (p-JNK), c-Jun, Ki-67, the latency-associated peptide (LAP) of transforming growth factor-beta 1 (TGF-beta1), and the TGF-beta receptor (TGF-betaRII). Immunoreactivities were scored from 0 to 3+ positivity using bright-field microscopy.. The tyrosine kinase signal transducer, PDGFR-alpha, the AT1 receptor transactivator, the p-JNK downstream effector, Ki-67, and proapoptoticTGF-1 (LAP) were detected in R-S cells of the NS and LP subtypes; companion dendritic cells expressed cathepsin D and ACE. Intranuclear c-Jun was present in the NS subtype and stronger immunoreactivity for TGF-betaRII was evident in the LP subtype.. These data corroborate observations in the literature, characterizing R-S cells as possessing molecular pathways that incorporate PDGFR-alpha signaling and angiotensin transactivation with a potential for growth inhibition through activation of TGF-beta1 and upregulation of its receptor. Specific therapies to target R-S cells in Hodgkin's disease might include ST1571, an AT1 receptor inhibitor, and retinoids.

Topics: Activin Receptors, Type I; Angiotensins; Cell Division; Hodgkin Disease; Humans; Immunohistochemistry; Peptidyl-Dipeptidase A; Protein Serine-Threonine Kinases; Proteome; Receptor, Angiotensin, Type 1; Receptor, Platelet-Derived Growth Factor alpha; Receptor, Transforming Growth Factor-beta Type I; Receptors, Angiotensin; Receptors, Transforming Growth Factor beta; Reed-Sternberg Cells; Signal Transduction; Transforming Growth Factor beta; Transforming Growth Factor beta1

EBV proteins present in the malignant Hodgkin Reed-Sternberg (HR-S) cells of about 40% of patients with Hodgkin's Disease (HD) provide targets for immunotherapy with virus-specific cytotoxic T lymphocytes (CTL). However, Hodgkin tumors use multiple strategies to avoid CTL, including down-regulation of immunodominant EBV antigens, and secretion of cytokines and chemokines such as TGF-beta, that inhibit the activation of CTL and professional antigen-presenting cells (APC). To be effective against this tumor, CTL must resist some or all of these strategies. Thirteen patients with multiply-relapsed HD received EBV-specific CTL, generated ex vivo using the autologous EBV-transformed B cells (LCL) as stimulator cells. After CTL infusion, EBV-specific immunity increased, virus load decreased, CTL homed to sites of malignancy and persisted for up to ten months. Clinically, CTL produced resolution of B symptoms and mixed tumor responses including one complete remission of residual disease remaining after autologous bone marrow transplant. However, no complete remission of bulky disease was achieved. Although LMP2-specific CTL activity could be detected in some of the infused CTL lines, they were present in low frequency. In pre-clinical studies, LMP1 and LMP2-specific CTL could be produced by stimulating PBMC from patients and normal donors with autologous dendritic cells expressing LMP1 or LMP2 from adenoviral vectors. Further, CTL could be rendered resistant to the devastating effects of TGF-beta by transduction with a retrovirus vector expressing a dominant-negative TGF-beta receptor, while transgenic IL-12 could increase the expression of Th1 and decrease that of Th2 cytokines. Future clinical studies will test the efficacy of CTL with improved antigen-specificity and resistance to Hodgkin immune evasion strategies.

Topics: Antigens, Viral; B-Lymphocytes; Bone Marrow Transplantation; Cell Line, Transformed; Gene Expression; Herpesvirus 4, Human; Hodgkin Disease; Humans; Immunotherapy; Interleukin-12; Recurrence; Stem Cell Transplantation; T-Lymphocytes, Cytotoxic; Transforming Growth Factor beta; Viral Matrix Proteins

Transforming growth factor beta (TGF-beta), a pleiotropic cytokine that regulates cell growth and differentiation, is secreted by many human tumors and markedly inhibits tumor-specific cellular immunity. Tumors can avoid the differentiating and apoptotic effects of TGF-beta by expressing a nonfunctional TGF-beta receptor. We have determined whether this immune evasion strategy can be manipulated to shield tumor-specific cytotoxic T lymphocytes (CTLs) from the inhibitory effects of tumor-derived TGF-beta. As our model we used Epstein-Barr virus (EBV)-specific CTLs that are infused as treatment for EBV-positive Hodgkin disease but that are vulnerable to the TGF-beta produced by this tumor. CTLs were transduced with a retrovirus vector expressing the dominant-negative TGF-beta type II receptor HATGF-betaRII-Deltacyt. HATGF-betaRII-Deltacyt- but not green fluorescence protein (eGFP)-transduced CTLs was resistant to the antiproliferative and anticytotoxic effects of exogenous TGF-beta. Additionally, receptor-transduced cells continued to secrete cytokines in response to antigenic stimulation. TGF-beta receptor ligation results in phosphorylation of Smad2, and this pathway was disrupted in HATGF-betaRII-Deltacyt-transduced CTLs, confirming blockade of the signal transduction pathway. Long-term expression of TGF-betaRII-Deltacyt did not affect CTL function, phenotype, or growth characteristics. Tumor-specific CTLs expressing HATGF-betaRII-Deltacyt should have a selective functional and survival advantage over unmodified CTLs in the presence of TGF-beta-secreting tumors and may be of value in treatment of these diseases.

Topics: Adjuvants, Immunologic; DNA-Binding Proteins; Genetic Therapy; Herpesvirus 4, Human; Hodgkin Disease; Humans; Immunotherapy; Mutation; Protein Serine-Threonine Kinases; Receptor, Transforming Growth Factor-beta Type II; Receptors, Transforming Growth Factor beta; Smad2 Protein; T-Lymphocytes, Cytotoxic; Trans-Activators; Transduction, Genetic; Transforming Growth Factor beta

To determine whether changes in the plasma Transforming Growth Factor beta1 (TGF beta1) concentration during radiotherapy could identify patients at risk for developing symptomatic radiation pneumonitis.. Thirty-six patients who received radiation therapy with curative intent for lung cancer (n = 31), Hodgkin's disease (n = 4), or thymoma (n = 1) were evaluated prospectively. All patients had serial plasma TGF beta1 measurements obtained before, during, and after treatment. Plasma TGF beta1 was quantified using an enzyme-linked immunosorbent assay. Pneumonitis was defined clinically. Plasma TGF beta1 levels were considered to have normalized if the following occurred: the last on-treatment TGF beta1 level was both <7.5 ng/ml and lower than the pretreatment level.. Thirteen of these 36 patients developed pneumonitis. Significant changes in plasma TGF beta1 levels during treatment were seen only in the subset of patients whose TGF beta1 levels were >7.5 ng/ml at baseline (n = 22). Failure of plasma TGF beta1 to normalize by the end of treatment, as defined above, much more accurately identified patients at risk for symptomatic pneumonitis if their baseline TGF beta1 was >7.5 ng/ml than if it was <7.5 ng/ml.. Changes in plasma TGF beta1 levels during radiotherapy appears to be a useful means by which to identify patients at risk for the development of symptomatic radiation pneumonitis, particularly in the subset of patients whose pretreatment TGF beta1 levels are >7.5 ng/ml.

Topics: Biomarkers; Hodgkin Disease; Humans; Lung Neoplasms; Prospective Studies; Radiation Pneumonitis; Sensitivity and Specificity; Thymoma; Transforming Growth Factor beta

To determine the cellular origin of the most potent cytokine present in Hodgkin's disease, transforming growth factor (TGF) beta, the polycellular population of Hodgkin's tissue was studied using in situ hybridisation.. A biotin labelled oligo-complementary DNA (cDNA) was constructed according to the previously determined sequence for TGF beta 1 cDNA. Forty three frozen and paraffin wax embedded tissue samples replaced by Hodgkin's disease or non-Hodgkin's lymphoma, three Reed-Sternberg cell lines, one Ki1 positive lymphoma cell line, and an epithelial cell line were studied for expression of TGF beta 1 messenger RNA (mRNA) as well as secretion of the TGF beta 1 protein and expression of the CD30 epitope.. The results obtained with the 24 frozen tissue samples confirmed that the TGF beta antigen is found predominantly in the nodular sclerosing Hodgkin's disease (NSHD) subtype. Nineteen paraffin wax embedded tissue samples were used to measure the simultaneous expression of CD30 and TGF beta 1 mRNA. The latter was found in eight of eight NSHD samples, two of six mixed cellularity samples, and two of five non-Hodgkin's lymphoma samples. No evidence of fibroblast expression of TGF beta 1 mRNA was noted.. Activated lymphocytes in NSHD express TGF beta 1 mRNA, but binucleate Reed-Sternberg cells and mononuclear Hodgkin's cells are the primary sources of activated TGF beta in Hodgkin's disease.

Topics: Base Sequence; Hodgkin Disease; Humans; In Situ Hybridization; Molecular Sequence Data; Oligonucleotide Probes; Reed-Sternberg Cells; RNA, Messenger; Transforming Growth Factor beta

The depressed cellular immunity observed in patients with Hodgkin's disease (HD) has been attributed to production of transforming growth factor (TGF)-beta or TGF-beta-like substances by Hodgkin's Reed-Sternberg (H-RS) cells. The TGF-beta produced by L-428 cells (an H-RS cell line) is a 130-kd molecular weight glycoprotein that apparently differs from the TGF-beta (molecular weight, 25 kd) produced by most lymphoid and hematopoietic cells. Among several distinct types of TGF-beta that have been purified, only TGF-beta 1 and TGF-beta 2 have thus far been identified in hematopoietic cells. By using monoclonal antibodies (1D11 and 3C7) and oligonucleotide probes specific for TGF-beta 1 and TGF-beta 2, were confirmed that a cultured H-RS cell line, KM-H2, can produce both TGF-beta types, whereas another line, HDLM-1, produces only TGF-beta 1. Despite the abundance of mRNA in both of these cells, only small amounts of TGF-beta activity were detected, probably because of rapid degradation of TGF-beta 1 mRNA by specific nuclease. No degraded TGF-beta 2 RNA products were observed in KM-H2 cells. The TGF-beta produced by both types of H-RS cells had a molecular weight of approximately 25 kd. In tissues expression of TGF-beta was observed in a small portion (30%) of H-RS cells in 16 of 20 cases examined. A large number of small to medium-sized lymphoid cells (T lymphocytes) in tissues involved by HD also were positive for TGF-beta. These results indicate that there is functional heterogeneity among H-RS cells, and that H-RS cells are not the only source of TGF-beta in tissues involved by HD. Hodgkin's Reed-Sternberg cells are known to secrete several other cytokines, including interleukin (IL)-1, IL-6, and tumor necrosis factor-alpha. These cytokines could be responsible for the increased number of T lymphocytes in tissues involved by HD. Furthermore, T lymphocytes can respond to IL-1 and IL-6 secreted by H-RS cells by increasing their production of TGF-beta. Abundant expression of TGF-beta by T lymphocytes was not observed in lymphoid tissues other than those involved by HD.

Topics: Base Sequence; Blotting, Northern; Cell Line; Gene Expression; Hodgkin Disease; Humans; Immunohistochemistry; Interleukin-1; Interleukin-6; Molecular Sequence Data; Reed-Sternberg Cells; RNA; T-Lymphocytes; Transforming Growth Factor beta; Tumor Necrosis Factor-alpha

Transforming growth factor-beta 1 (TGF-beta 1) is a multifunctional cytokine which promotes fibroblast growth and collagen synthesis, but suppresses growth and differentiation of immune lymphocytes and killer cells. Immunohistochemical detection of TGF-beta 1 in Hodgkin's disease (HD) has been shown to correlate with the histologic feature of nodular sclerosis, which is associated with a favorable prognosis (American Journal of Pathology 1990, 136:1209). In that study, TGF-beta 1 was localized mainly at the margins of broad collagen bands (presumably sites of new collagen synthesis) and in areas containing numerous Hodgkin/Reed-Sternberg cells (H/RS). In these areas, TGF-beta 1 protein was found on the membrane and occasionally within the cytoplasm of H/RS cells. To determine whether TGF-beta 1 is synthesized by H/RS cells or secondarily bound to their membrane and sometimes internalized, we performed in situ hybridization (ISH) using 1.5 Kb 35S-labeled anti-sense and sense RNA probes to TGF-beta 1. Paraffin-embedded tissues of 10 cases from all histologic types of HD were examined. Somewhat unexpectedly, the major site of TGF-beta 1 mRNA was in eosinophils; TGF-beta 1 mRNA was not detected in H/RS cells. TGF-beta 1 mRNA was found in eosinophils in all cases of nodular sclerosis but not in other types of HD, despite the presence of numerous eosinophils in mixed cellularity cases. The presence of TGF-beta 1 mRNA coincided with immunohistochemical detection of TGF-beta 1 protein using antibody CC (1-30). These results confirm the role of TGF-beta 1 in the histogenesis of nodular sclerosing HD and indicate that eosinophils are the major source of TGF-beta 1 in this type of HD.

Topics: Adult; Collagen; Eosinophils; Hodgkin Disease; Humans; Lymph Nodes; RNA, Messenger; RNA, Neoplasm; Sclerosis; Transforming Growth Factor beta

Activated lymphocytes and malignant lymphoma cells derived from them (Ki-1 positive lymphoma cells) share similar mechanisms of proliferation. To further examine the inhibitory role of endogenous transforming growth factor beta (TGF beta) in Ki-1 positive lymphoma cells, the authors studied anti-TGF beta antibodies and measured their effect on proliferation. A monoclonal antibody (T1A5) prepared against a unique antigenic epitope of high molecular weight Hodgkin's TGF beta and a polyclonal rabbit antibody prepared against highly purified 25,000 D porcine platelet TGF beta 1 were used. Both antibodies are shown here to inhibit the biological activity of Hodgkin's TGF beta and to crossreact with their respective antigens in immunoblotting. DNA synthesis by Ki-1 lymphoma cells was increased 138-fold by anti-TGF beta 1 antibody and 262-fold by anti-Hodgkin's TGF beta. Exogenous TGF beta 1 suppression was completely reversed by anti-TGF beta 1 antibody and IL-2-induced proliferation was markedly potentiated (41 fold). L-428 Reed-Sternberg cells secrete physiologically active TGF beta but have fewer than 500 TGF beta receptor sites per cell; no significant proliferative response was measured for either anti-TGF beta 1 or anti-Hodgkin's TGF beta. These results show the suppressive effect of exogenous TGF beta 1 on indolent Ki-1 lymphoma cells and suggest that the endogenous secretion of high molecular weight physiologically active TGF beta is important in maintaining the indolent nature of this low-grade Ki-1 positive lymphoma.

Topics: Antibodies; Antigens, CD; Antigens, Neoplasm; Cell Division; Hodgkin Disease; Humans; Interleukin-2; Ki-1 Antigen; Lymphoma; Neutralization Tests; Transforming Growth Factor beta; Tumor Cells, Cultured

To measure the in vivo secretion of high molecular weight (HMW) transforming growth factor (TGF)beta by Reed-Sternberg cells from patients with nodular sclerosing Hodgkin's disease, we studied the urine samples from untreated patients. The urinary proteins did not promote the proliferation of NIH-3T3 cells in monolayer culture and contained similar amounts of total TGF activity when compared with normal controls. Urinary proteins from 24 different control and test urines were analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and immunoblotting. Either of two primary antibodies were used for immunoblot detection: (a) affinity column purified polyclonal anti-TGF beta 1 prepared against platelet TGF beta 1 or (b) monoclonal anti-HMW-TGF beta prepared against HMW-TGF beta secreted by cloned L-428 Reed-Sternberg cells. All patients with active nodular sclerosing Hodgkin's disease had a detectable HMW-TGF beta (approximately 300,000) which cross-reacted with both anti-TGF beta 1 and anti-HMW-TGF beta. Purification demonstrated HMW-TGF beta which was active at physiological pH. Twelve control urine samples from healthy adults and 5 follow-up samples from the Hodgkin's patients after successful treatment contained no detectable urinary HMW-TGF beta. The in vivo production of HMW-TGF beta in untreated nodular sclerosing Hodgkin's disease supports the conclusion that this growth factor is secreted in large amounts by Reed-Sternberg cells or cells stimulated by Reed-Sternberg cells.

Topics: 3T3 Cells; Adolescent; Adult; Animals; Female; Hodgkin Disease; Humans; Immunoblotting; Male; Mice; Molecular Weight; Sclerosis; Transforming Growth Factor beta