scholarly journals Follicular Lymphoma Cells Induce Changes in T-Cell Gene Expression and Function: Potential Impact on Survival and Risk of Transformation

2013 ◽  
Vol 31 (21) ◽  
pp. 2654-2661 ◽  
Author(s):  
Shahryar Kiaii ◽  
Andrew J. Clear ◽  
Alan G. Ramsay ◽  
Derek Davies ◽  
Ajanthah Sangaralingam ◽  
...  
Keyword(s):  
Gene Expression ◽  
Overall Survival ◽  
T Cells ◽  
T Cell ◽  
Follicular Lymphoma ◽  
Peripheral Blood ◽  
Molecular Mechanisms ◽  
Prognostic Significance ◽  
Time Lapse ◽  
Immune Microenvironment

Purpose Previous studies have demonstrated the prognostic importance of the immune microenvironment in follicular lymphoma (FL). To investigate the molecular mechanisms during which tumor-infiltrating T cells (TILs) are altered in the FL microenvironment, we studied highly purified CD4 and CD8 TILs from lymph node biopsies at diagnosis in treatment-naive patients with FL compared with reactive tonsils and the peripheral blood of healthy donors. Patients and Methods Gene expression profiling of highly purified CD4 and CD8 TILs was performed on the Affymetrix platform. Diagnostic tissue microarrays from an independent patient set (n = 172) were used to verify protein expression and analyze any impact of TIL-expressed genes on outcome. Time-lapse imaging was used to assess T-cell motility. Results The most upregulated genes in both CD4 and CD8 TILs were PMCH, ETV1, and TNFRSF9. PMCH is not expressed in peripheral blood T cells, but expression is highly induced on culture with FL. Both CD4 and CD8 TILs from patients with FL have significantly impaired motility compared with those of healthy TILs from reactive tonsils and this can be induced on healthy T cells by FL cells. During multivariate analysis, a model incorporating the number and location of T cells expressing PMCH, NAMPT, and ETV1 showed prognostic significance for overall survival and for time to transformation. Conclusion We showed altered gene expression in TILs in FL and demonstrated that altering the immune microenvironment in FL affects overall survival and time to transformation in this disease.

Differential Gene Expression Profile Identifies Defects and Abnormalities In Infiltrating T Cells In Patients with Follicular Lymphoma at Diagnosis

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 141-141
Author(s):  
Shahryar Kiaii ◽  
Andrew J. Clear ◽  
Alan G. Ramsay ◽  
Ajanthah Sangaralingam ◽  
John G. Gribben
Keyword(s):  
Gene Expression ◽  
T Cells ◽  
T Cell ◽  
Follicular Lymphoma ◽  
Immune Cells ◽  
Peripheral Blood ◽  
Expression Profiling ◽  
Molecular Mechanisms ◽  
Clinical Course ◽  
Cd8 Cells

Abstract Abstract 141 Patients with follicular lymphoma (FL) have an extremely variable clinical course. Although clinical parameters can be used to define prognostic subgroups, there is a need to identify robust biomarkers that not only aid in prognosis, but also help define the underlying pathophysiology of the disease. Previous gene expression profiling studies demonstrated that among the most important prognostic markers were the molecular features of nonmalignant tumor-infiltrating immune cells present in the tumor at diagnosis (Sandeep et al. NEJM 2004). To investigate the molecular mechanisms whereby T cells are altered in the FL microenvironment we studied highly purified, sorted infiltrating CD4 and CD8 T cells from previously cryopreserved single cell suspensions of lymph node (LN) biopsies at the time of diagnosis in patients with FL (n=12) and compared them to those isolated from reactive tonsils (n=7) as well as from peripheral blood (PB) (n=10) of age matched healthy individuals. These tumor infiltrating T cells (TILs) have impaired proliferative and cytotoxic responses and impaired capacity to form immunologic synapses with antigen presenting cells. Co-culture of FL cells in contact with healthy allogeneic T cells induces similar T cell functional defects, demonstrating that it is the FL cells that drive the altered T cell function. To investigate the molecular mechanisms for this, we performed global gene expression profiling using Affymetrix U133Plus2 chips of highly purified (>95% purity) CD4 and CD8 cells. For both CD4 and CD8 cells, unsupervised analyses distinguished healthy and FL T cell subsets. Using a fold change cut off > 2 and false discovery rate of 5%, 280 genes were differentially expressed for CD4 and 717 genes for CD8, with109 genes overlapped for both subsets. The gene array data was validated for RNA using Real-Time Quantitative-PCR and for protein by flow cytometry and immunohistochemistry. Pathway analysis indicated disruption in multiple pathways including cytokine signaling, T cell differentiation, cell proliferation and, actin-based motility/cytoskeleton formation. In both CD4 and CD8, among the most downregulated genes in FL TILs were ACTN1 and IL17A, and the most upregulated genes were PMCH and ETV1. Using Tissue Microarray (TMA) we demonstrate that the intensity of expression in TILs in FL was significantly higher for PMCH (p<0.0001) and ETV1 (p<0.0001) than that of reactive tissue. PMCH is not expressed in PB T cells, but its expression is highly induced when healthy peripheral blood T cells are cultured, either in cell contact or in transwell culture, with FL cells. Surprisingly, co-culture of healthy T cells with normal B cells also induced its expression. Taken together, these data indicate that TILs in patients with FL are abnormal in terms of their function and gene expression profile, in keeping with our hypothesis that FL induces changes in immune cells in the tumor microenvironment. We are currently further characterizing the functional consequences of the identified changes in the gene expression in tumor-infiltrating T cells of patients with FL in the biology and progression of disease. As non-malignant immune cells have a major role in the clinical course of patients with FL, understanding the nature and impact of the abnormalities induced in infiltrating T cell's in these patients is vital before any immunotherapeutic strategies can be implemented to attempt to alter the immune microenvironment in FL. Disclosures: Gribben: Roche: Consultancy; Celgene: Consultancy; GSK: Honoraria; Napp: Honoraria.

Impact of Tumor Infiltrating T Cells in Patients with Follicular Lymphoma At Diagnosis

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 769-769
Author(s):  
Shahryar Kiaii ◽  
Andrew J. Clear ◽  
Ajanthah Sangaralingam ◽  
John G. Gribben
Keyword(s):  
Gene Expression ◽  
T Cells ◽  
T Cell ◽  
Median Survival ◽  
Immune Cells ◽  
Molecular Mechanisms ◽  
Prognostic Significance ◽  
Lower Percentage ◽  
Molecular Features ◽  
Treatment Naïve

Abstract Abstract 769 Previous global gene expression profiling (GEP) studies have demonstrated non-malignant tumor-infiltrating immune cells present in the tumor at diagnosis were among the molecular features of the most important prognostic markers (Sandeep et al. NEJM 2004). We investigated the molecular mechanisms whereby tumor infiltrating T cells (TILs) are altered in the FL microenvironment and examined GEP of highly purified, sorted infiltrating CD4 and CD8 T cells from previously cryopreserved single cell suspensions of lymph node (LN) biopsies at the time of diagnosis in treatment naive patients with FL (n=12) and compared them to those isolated from reactive tonsils (n=7) as well as peripheral blood (PB) (n=10) of age matched healthy donors. In both CD4 and CD8, among the most downregulated genes in FL TILs were ACTN1 and IL17A, and the most upregulated genes were PMCH, ETV1, TNFRSF9 and NAMPT. PMCH is not expressed in PB T cells, but its expression is highly induced when healthy PB T cells are cultured, either in cell contact or in transwell culture, with FL cells. Using well characterized tissue microarrays of diagnostic FL samples (n=172) we now show that the T-cell expressed genes of PMCH, ETV1 and NAMPT have prognostic significance for survival and time to transformation in FL. Patients with higher percentage (p=0.039, median survival 10.59yr vs 19.20yr) and number (p=0.016, median survival 8.67yr vs 19.01yr) of PMCH expressing cells in intrafollicular and higher percentage (p=0.021, median survival 8.18yr vs 16.5yr) in interfollicular area had significantly longer disease specific survival (DSS) compared with patient with low number of PMCH expressing cells. Patients with higher percentage (p=0.014, median survival 19.01yr vs 11.75) and number (p=0.032, median survival 19.01yr vs 12.41yr) of ETV1 expressing cells in intrafollicular area showed significantly shorter DSS comparing to those with lower percentage and number of ETV1 expressing cells. Furthermore, the higher total MI of NAMPT expression showed significantly longer DSS (p=0.003, median survival 7.62yr vs 18.28yr) as well. Pathway analysis indicated disruption in multiple cellular pathways including actin-based motility/cytoskeleton formation which is in keeping with our previous studies where we have shown altered T cell expression of genes regulating actin in CLL (Gorgun et al, JCI 2005) and AML (De Lieu et al, Blood 2009). Using Time-Lapse Imaging we demonstrated both CD4 and CD8 TILs from patients with FL (n=7) have significantly impaired motility compared to those of healthy TILs from reactive tonsil (n=4) (p<0.025). We have further demonstrated that short term culture of allogeneic healthy T cells with FL cells from treatment naive patients (n=8) directly induce significantly impaired T cell motility compared to culture of these T cells with healthy allogeneic non malignant B cells (n=6)(p<0.05). Taken together, these data indicate that TILs in patients with FL are abnormal in terms of their gene expression and function. Furthermore, we provide evidence supporting the role that FL cells play in inducing changes in immune cells in the tumor microenvironment. In addition, we have shown that changes in the protein level in tumor infiltrating T-cells have an impact on the survival and time to transformation in patients with FL. We are further characterizing the mechanisms of gene expression alteration in TILs of patients with FL and its functional consequences in the biology and of the disease. Since non-malignant infiltrating immune cells have a crucial role in the outcome of patients with FL, understanding the nature and impact of the abnormalities induced in TILs in these patients is vital before any immunotherapeutic strategies can be implemented to attempt to alter the immune microenvironment in FL. Disclosures: Gribben: Roche: Honoraria; Celgene: Honoraria; GSK: Honoraria; Mundipharma: Honoraria; Gilead: Honoraria; Pharmacyclics: Honoraria.

High numbers of tumor-infiltrating FOXP3-positive regulatory T cells are associated with improved overall survival in follicular lymphoma

Blood ◽  
2006 ◽  
Vol 108 (9) ◽  
pp. 2957-2964 ◽  
Author(s):  
Joaquim Carreras ◽  
Armando Lopez-Guillermo ◽  
Bridget C. Fox ◽  
Lluis Colomo ◽  
Antonio Martinez ◽  
...  
Keyword(s):  
Overall Survival ◽  
T Cells ◽  
Regulatory T Cells ◽  
Follicular Lymphoma ◽  
International Prognostic Index ◽  
Prognostic Significance ◽  
Prognostic Index ◽  
Specific Cell ◽  
First Relapse ◽  
The Impact

Abstract The tumor microenvironment plays an important role in the biologic behavior of follicular lymphoma (FL), but the specific cell subsets involved in this regulation are unknown. To determine the impact of FOXP3-positive regulatory T cells (Tregs) in the progression and outcome of FL patients, we examined samples from 97 patients at diagnosis and 37 at first relapse with an anti-FOXP3 monoclonal antibody. Tregs were quantified using computerized image analysis. The median overall survival (OS) of the series was 9.9 years, and the FL International Prognostic Index (FLIPI) was prognostically significant. The median Treg percentage at diagnosis was 10.5%. Overall, 49 patients had more than 10% Tregs, 30 between 5% to 10%, and 19 less than 5%, with a 5-year OS of 80%, 74%, and 50%, respectively (P = .001). Patients with very low numbers of Tregs (< 5%) presented more frequently with refractory disease (P = .007). The prognostic significance of Treg numbers was independent of the FLIPI. Seven transformed diffuse large B-cell lymphomas (DLBCLs) had lower Treg percentages (mean: 3.3%) than FL grades 1,2 (mean: 12.1%) or 3 (mean: 9%) (P < .02). In conclusion, high Treg numbers predict improved survival of FL patients, while a marked reduction in Tregs is observed on transformation to DLBCL.

scholarly journals OS1.6 Peripheral blood immune profiles at first recurrence of IDH wild type glioblastoma after standard chemoradiotherapy predict overall survival: secondary analyses of the phase II DIRECTOR trial

2019 ◽  
Vol 21 (Supplement_3) ◽  
pp. iii6-iii7
Author(s):  
H Wirsching ◽  
E Terksikh ◽  
S Manuela ◽  
K Carsten ◽  
R Patrick ◽  
...  
Keyword(s):  
Overall Survival ◽  
T Cells ◽  
Prognostic Factors ◽  
T Cell ◽  
Peripheral Blood ◽  
Immune Cell ◽  
Cd4 T Cell ◽  
Cox Models ◽  
First Recurrence ◽  
Cell Fractions

Abstract BACKGROUND Isocitrate dehydrogenase (IDH) wildtype glioblastoma is associated with distinctive peripheral blood immune cell profiles that evolve under first line chemoirradiation with temozolomide. Whether peripheral blood immune cell profiles at recurrence are associated with survival of IDH wildtype glioblastoma has not been studied in detail. PATIENTS AND METHODS Peripheral blood mononuclear cells (PBMC) of 21 healthy donors and of 52 clinically well-annotated patients with IDH wildtype glioblastoma were analyzed by 11-color flow cytometry at 1st recurrence after standard chemoirradiation with temozolomide and at 2nd recurrence after dose-intensified temozolomide re-challenge. Patients were treated within the randomized phase II trial DIRECTOR, which explored the efficacy of dose-intensified temozolomide at first recurrence of glioblastoma. Patients were classified based on unsupervised analyses of PBMC profiles at 1stand 2ndrecurrence. Associations with survival were explored in multivariate Cox models controlling for established prognostic and predictive factors. RESULTS At 1strecurrence, two patient clusters were identified which differed in CD4+ T-cell fractions, but not with respect to CD8+ T-cells, CD4+;CD25+;FoxP3+ regulatory T-cells, B-cells or monocytes. The composition of CD4+, CD8+ or regulatory T-cell fractions was similar in both clusters. All control samples clustered with the CD4high cluster. Patients in both clusters did not differ by established prognostic factors, including age, O6-methylguanine-DNA-methyl-transferase (MGMT) gene promoter methylation, tumor volume, Karfnosky performance score or steroid use. Progression-free survival was similar (CD4high vsCD4low 2.1 vs 2.4 months, p=0.19), whereas post-recurrence overall survival was longer among the CD4highcluster (12.7 vs 8.7 months, p= 0.004). At 2nd recurrence, monocyte fractions increased, whereas memory CD4+ T-cell fractions decreased. Unsupervised segregation of patients by CD4+ subpopulations yielded two clusters characterized by the abundance of memory T-cell fractions and higher memory CD4+ fractions were associated with longer overall survival at 2nd recurrence (p=0.004). The reported prognostic associations were retained in multivariate Cox models controlling for established prognostic factors. CONCLUSION We conclude that temozolomide-associated memory CD4+ T-cell depletion may have deteriorating effects on the survival of glioblastoma patients.

Association of peripheral blood CD4+ T-cell depletion under temozolomide with inferior survival of patients with IDH wildtype glioblastoma.

2020 ◽  
Vol 38 (15_suppl) ◽  
pp. 2548-2548
Author(s):  
Michael Weller ◽  
Ekaterina Terksikh ◽  
Manuela Silginer ◽  
Carsten Krieg ◽  
Patrick Roth ◽  
...  
Keyword(s):  
Overall Survival ◽  
T Cells ◽  
Prognostic Factors ◽  
T Cell ◽  
Phase Ii ◽  
Peripheral Blood ◽  
Phase Ii Trial ◽  
Cd4 T Cell ◽  
Cox Models ◽  
Cell Fractions

2548 Background: Standard first line chemoirradiation with temozolomide is associated with distinctive peripheral blood immune cell profiles in IDH wildtype glioblastoma. Whether such profiles at recurrence are associated with survival has not been studied in detail. Methods: Peripheral blood mononuclear cells of 21 healthy donors and of 91 patients with IDH wildtype glioblastoma were analyzed by flow cytometry at 1st recurrence. Patients received either (i) standard chemoirradiation with temozolomide (TMZ) followed by dose-intensified TMZ at first recurrence within the phase II trial DIRECTOR (N = 52) or (ii) hypofractionated radiotherapy with or without bevacizumab (N = 39) followed by investigators’ choice within the phase II trial ARTE. Patients were classified based on unsupervised analyses of PBMC profiles at 1st recurrence. Associations with survival were explored in multivariate Cox models controlling for established prognostic and predictive factors. Results: At 1st recurrence, two patient clusters were identified in the DIRECTOR cohort which differed in CD4+ T-cell fractions, but not with respect to CD8+ T-cells, CD4+;CD25+;FoxP3+ regulatory T-cells, B-cells or monocytes. The composition of CD4+, CD8+ or regulatory T-cell fractions was similar in both clusters. All control samples clustered with the CD4high cluster. Patients in both clusters did not differ by established prognostic factors, including age, O6-methylguanine-DNA-methyl-transferase ( MGMT) gene promoter methylation, tumor volume, Karfnosky performance score or steroid use. Progression-free survival was similar (CD4high vs CD4low 2.1 vs 2.4 months, p = 0.19), whereas post-recurrence overall survival was longer among the CD4high cluster (12.7 vs 8.7 months, p = 0.004). At 2nd recurrence after dose-intensified TMZ re-challenge, monocyte fractions increased, whereas memory CD4+ T-cell fractions decreased. Higher memory CD4+ fractions were associated with longer overall survival at 2nd recurrence (p = 0.004). The reported associations were retained in multivariate Cox models controlling for established prognostic factors. In the ARTE cohort, CD4+ T cell fractions at 1st recurrence did not differ compared to diagnosis (p = 0.91) and there were no associations with bevacizumab (p = 0.28) or survival (p = 0.74), supporting that the effects observed in the DIRECTOR cohort were driven by TMZ. Conclusions: We conclude that TMZ-associated memory CD4+ T-cell depletion may have deteriorating effects on the survival of glioblastoma patients.

scholarly journals Peripheral blood T cells in acute myeloid leukemia (AML) patients at diagnosis have abnormal phenotype and genotype and form defective immune synapses with AML blasts

Blood ◽  
2009 ◽  
Vol 114 (18) ◽  
pp. 3909-3916 ◽  
Author(s):  
Rifca Le Dieu ◽  
David C. Taussig ◽  
Alan G. Ramsay ◽  
Richard Mitter ◽  
Faridah Miraki-Moud ◽  
...  
Keyword(s):  
Gene Expression ◽  
Immune Response ◽  
T Cells ◽  
Acute Myeloid Leukemia ◽  
T Cell ◽  
Peripheral Blood ◽  
Myeloid Leukemia ◽  
Absolute Number ◽  
Immune Synapses ◽  
Acute Myeloid

Abstract Understanding how the immune system in patients with cancer interacts with malignant cells is critical for the development of successful immunotherapeutic strategies. We studied peripheral blood from newly diagnosed patients with acute myeloid leukemia (AML) to assess the impact of this disease on the patients' T cells. The absolute number of peripheral blood T cells is increased in AML compared with healthy controls. An increase in the absolute number of CD3+56+ cells was also noted. Gene expression profiling on T cells from AML patients compared with healthy donors demonstrated global differences in transcription suggesting aberrant T-cell activation patterns. These gene expression changes differ from those observed in chronic lymphocytic leukemia (CLL), indicating the heterogeneous means by which different tumors evade the host immune response. However, in common with CLL, differentially regulated genes involved in actin cytoskeletal formation were identified, and therefore the ability of T cells from AML patients to form immunologic synapses was assessed. Although AML T cells could form conjugates with autologous blasts, their ability to form immune synapses and recruit phosphotyrosine signaling molecules to the synapse was significantly impaired. These findings identify T-cell dysfunction in AML that may contribute to the failure of a host immune response against leukemic blasts.

scholarly journals Comprehensive Analysis of Cell Population Dynamics and Related Core Genes During Vitiligo Development

2021 ◽  
Vol 12 ◽  
Author(s):  
Jingzhan Zhang ◽  
Shirong Yu ◽  
Wen Hu ◽  
Man Wang ◽  
Dilinuer Abudoureyimu ◽  
...  
Keyword(s):  
Gene Expression ◽  
Immune Response ◽  
T Cells ◽  
T Cell ◽  
Peripheral Blood ◽  
Immune Cell ◽  
Cd8 T Cell ◽  
Cd4 T Cell ◽  
Healthy Controls ◽  
Cell Abundance

Vitiligo is a common immune-related depigmentation condition, and its pathogenesis remains unclear. This study used a combination of bioinformatics methods and expression analysis techniques to explore the relationship between immune cell infiltration and gene expression in vitiligo. Previously reported gene expression microarray data from the skin (GSE53146 and GSE75819) and peripheral blood (GSE80009 and GSE90880) of vitiligo patients and healthy controls was used in the analysis. R software was used to filter the differentially expressed genes (DEGs) in each dataset, and the KOBAS 2.0 server was used to perform functional enrichment analysis. Compared with healthy controls, the upregulated genes in skin lesions and peripheral blood leukocytes of vitiligo patents were highly enriched in immune response pathways and inflammatory response signaling pathways. Immunedeconv software and the EPIC method were used to analyze the expression levels of marker genes to obtain the immune cell population in the samples. In the lesional skin of vitiligo patients, the proportions of macrophages, B cells and NK cells were increased compared with healthy controls. In the peripheral blood of vitiligo patients, CD8+ T cells and macrophages were significantly increased. A coexpression analysis of the cell populations and DEGs showed that differentially expressed immune and inflammation response genes had a strong positive correlation with macrophages. The TLR4 receptor pathway, interferon gamma-mediated signaling pathway and lipopolysaccharide-related pathway were positively correlated with CD4+ T cells. Regarding immune response-related genes, the overexpression of IFITM2, TNFSF10, GZMA, ADAMDEC1, NCF2, ADAR, SIGLEC16, and WIPF2 were related to macrophage abundance, while the overexpression of ICOS, GPR183, RGS1, ILF2 and CD28 were related to CD4+ T cell abundance. GZMA and CXCL10 expression were associated with CD8+ T cell abundance. Regarding inflammatory response-related genes, the overexpression of CEBPB, ADAM8, CXCR3, and TNIP3 promoted macrophage infiltration. Only ADORA1 expression was associated with CD4+ T cell infiltration. ADAM8 and CXCL10 expression were associated with CD8+ T cell abundance. The overexpression of CCL18, CXCL10, FOS, NLRC4, LY96, HCK, MYD88, and KLRG1, which are related to inflammation and immune responses, were associated with macrophage abundance. We also found that immune cells infiltration in vitiligo was associated with antigen presentation-related genes expression. The genes and pathways identified in this study may point to new directions for vitiligo treatment.

scholarly journals Gene expression of T lymphocytes of gravid cows during preimplantation

2013 ◽  
Vol 82 (2) ◽  
pp. 131-134 ◽  
Author(s):  
Yousuke Maeda ◽  
Kana Yamamoto ◽  
Hiromichi Ohtsuka ◽  
Takaaki Ando ◽  
Michiko Tomioka ◽  
...  
Keyword(s):  
Gene Expression ◽  
T Cells ◽  
T Cell ◽  
Peripheral Blood ◽  
Mononuclear Cells ◽  
Γδ T Cells ◽  
Interferon Γ ◽  
T Cell Subsets ◽  
Interleukin 4 ◽  
Blocking Factor

An interaction between the conceptus and the immune system of animals is important during implantation. The aim of this study was to clarify the gene expression of T cell subsets in gravid cows during the preimplantation period. Peripheral blood from 14 Holstein dairy cows was taken 14 days after artificial insemination. Based on the gravidity, cows were divided into gravid (n = 8) and nongravid (n = 6) groups. Mononuclear cells from peripheral blood were stimulated with phytohaemagglutinin and then CD4+, CD8+, and WC1+ γδ T cell subsets were isolated using magnetic cell sorting. The expression of interferon γ, interleukin 4, and progesterone induced blocking factor were determined using real-time PCR. The expression of interleukin 4 and progesterone induced blocking factor was significantly higher in WC1+ γδ T cells from gravid cows. In addition, interleukin 4 expression in WC1+ γδ T cells from gravid cows was significantly higher than that in CD4+ and CD8+ T cells. This study describes for the first time the important role of WC1+ γδ T cells during the preimplantation period in cows.

Tumor-Infiltrating T Cells Are Not Predictive of Clinical Outcome in Follicular Lymphoma.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 824-824 ◽  
Author(s):  
Wei Yun Z. Ai ◽  
Debra Czerwinski ◽  
Sandra J. Horning ◽  
John Allen ◽  
Robert Tibshirani ◽  
...  
Keyword(s):  
Gene Expression ◽  
Flow Cytometry ◽  
Overall Survival ◽  
T Cells ◽  
Clinical Outcome ◽  
Follicular Lymphoma ◽  
Clinical Features ◽  
Training Set ◽  
Hla Dr ◽  
Validation Set

Abstract Background: Follicular lymphoma (FL) has variable clinical outcomes. It has been suspected that tumor-infiltrating immune cells affect the biology and outcome of this disease. Using gene expression profiling and immunoperoxide tissue staining techniques, T cells and macrophages have been related to the survival outcome in some studies, but not others. In the current study, we used flow cytometry to analyze T cells and their subsets in follicular lymphoma biopsy specimens and determined whether these cell populations correlated with clinical features and outcomes. Methods: Two hundred and eighty-nine follicular lymphoma patients (pt) presented from 1997 to 2003 underwent an excisional lymph node biopsy prior to any treatment. The median age of pt at diagnosis was 45.7 yrs, median follow-up was 8.6 yrs for living pts, and median survival was 15.7 yrs. All but 8 patients had stage III/IV disease, 5 had stage I/II, and 3 were unknown. The histological grades were: 162 (56%) grade 1, 112 (39%) grade 2, 13 (4.5%) grade 3 and 2 (0.5%) unknown. Among the 289 patients, 41(17%) had low FLIPI score, 150 (63%) intermediate, 48 (20%) high and 50 unknown. All biopsies were analyzed for CD20, CD3, CD4, CD8 and HLA-DR expression by single-parameter flow cytometry. The 289 pts were divided into a training set of 147 and a validation set of 142, stratified by age and era of diagnosis. We used these two factors to stratify the pts because age at diagnosis is the most important prognostic factor for survival, and, in our data set, the era of diagnosis had an impact on survival and on the time from diagnosis to first treatment. For our analysis, we began with the training set and used the percentages of each immune cell population as a continuous variable in a univariate analysis in relation to clinical features and outcomes. We chose 8 phenotypic variables: CD20, CD3, CD4, CD8, HLA-DR, CD4/CD3 ratio, CD8/CD3 ratio, and activated T cells [defined as (HLA-DR-CD20)/CD3]. Five parameters were used as clinical endpoints: overall survival, FLIPI score at diagnosis, the time from diagnosis to first treatment (defined as the time from the first treatment to second treatment), response to CVP as the first treatment and the duration of the benefit from the first treatment (defined as the time interval between initiation of first treatment and initiation of second treatment). Results: The number of pt evaluable for each of the outcome parameters was as follows: 289 for time to first treatment and for overall survival, 239 for FLIPI scores, 164 for response to CVP and 129 for duration of the benefit from the first treatment., Of the 8 variables tested in the training set, only CD4/CD3 ratio and CD8/CD3 ratio were marginally significant for the survival endpoint, with p 0.034 and 0.088, respectively. None of the variables was significant for any of the other endpoints. A multivariate analysis yielded CD4/CD3 as the only significant predictor for survival. When CD4/CD3 was tested in the validation set, it yielded a p value of 0.48. Conclusion: We find no evidence that the percentage of tumor-infiltrating T cells or their subsets is predictive of clinical outcome in follicular lymphoma. Any gene expression signature involving T cells that does relate to clinical outcome could therefore be a property of the activity of the cells rather than a simple reflection of their numbers.

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