Cell-free DNA as a prognostic marker in stage I non-small-cell lung cancer patients undergoing stereotactic body radiotherapy.

OBJECTIVE: To evaluate whether plasma cell-free DNA (cfDNA) was related to clinical outcome in inoperable stage I non-small cell lung cancer (NSCLC) patients undergoing stereotactic body radiotherapy (SBRT). MATERIALS AND METHODS: Plasma cfDNA was assessed at baseline, before the last day and 45 days after the end of SBRT, in 22 NSCLC patients. Twenty-two healthy controls were also evaluated. RESULTS: Plasma cfDNA was higher in patients than in controls. An association with unfavourable disease-free survival was found for continuous baseline cfDNA increments (HR = 5.9, 95%CI: 1.7-19.8, p = 0.04). CONCLUSION: Plasma cfDNA may be a promising prognostic biomarker in high-risk NSCLC patients.

Plasma biomarkers of clinical response during chemotherapy plus combination antiretroviral therapy (cART) in HIV+ patients with advanced Kaposi sarcoma.

This study aimed to evaluate plasma concentration of selected cancer-associated inflammatory and immune-modulated cytokines in HIV+ patients with advanced Kaposi sarcoma (KS), and to explore candidate biomarkers capable of predicting clinical outcome in response to chemotherapy (CT) plus combination antiretroviral therapy (cART).Thirty-seven plasma cytokines/chemokines were assessed by Luminex technology in 27 consecutive HIV+ KS patients, followed-up during CT and cART of maintenance (m-cART). Associations between plasma concentration of biomarkers and patient clinical response to m-cART were evaluated by means of Hazard Ratios (HRs) and corresponding 95% Confidence Intervals (CIs).Plasma baseline concentration of Granulocyte colony-stimulating factor (G-CSF), Hepatocyte growth factor (HGF) and endoglin were found to be associated with m-cART clinical response (HR:1.56, 95%CI:1.09-2.22, p = 0.01; HR:0.32, 95% CI:0.10-0.99, p = 0.05; HR:0.72, 95% CI:0.54-0.96, p = 0.03, respectively). The multivariate analysis confirmed the associations of baseline plasma G-CSF and HGF concentration with m-cART clinical complete remission response (HR:1.78, 95% CI:1.15-2.74, p = 0.009; HR:0.19, 95% CI:0.04-0.95, p = 0.04). Our exploratory study suggested that plasma G-CSF, HGF and endoglin may be novel predictors of clinical response during m-cART in HIV+ KS patients. Nonetheless, these findings should be further validated in an independent population study.

γ-Herpesvirus load as surrogate marker of early death in HIV-1 lymphoma patients submitted to high dose chemotherapy and autologous peripheral blood stem cell transplantation.

Autologous stem cell transplantation (ASCT) is a feasible procedure for human immunodeficiency virus-1 (HIV-1) lymphoma patients, whose underlying disease and intrinsic HIV-1- and ASCT-associated immunodeficiency might increase the risk for γ-herpesvirus load persistence and/or reactivation. We evaluated this hypothesis by investigating the levels of Epstein-Barr virus (EBV)- and Kaposi sarcoma-associated herpesvirus (KSHV)-DNA levels in the peripheral blood of 22 HIV-1-associated lymphoma patients during ASCT, highlighting their relationship with γ-herpesvirus lymphoma status, immunological parameters, and clinical events. EBV-DNA was detected in the pre-treatment plasma and peripheral blood mononuclear cells (PBMCs) of 12 (median 12,135 copies/mL) and 18 patients (median 417 copies/10(6) PBMCs), respectively; the values in the two compartments were correlated (r = 0.77, p = 0.0001). Only EBV-positive lymphomas showed detectable levels of plasma EBV-DNA. After debulking chemotherapy, plasma EBV-DNA was associated with lymphoma chemosensitivity (p = 0.03) and a significant higher mortality risk by multivariate Cox analysis adjusted for EBV-lymphoma status (HR, 10.46, 95% CI, 1.11-98.32, p = 0.04). After infusion, EBV-DNA was detectable in five EBV-positive lymphoma patients who died within six months. KSHV-DNA load was positive in only one patient, who died from primary effusion lymphoma. Fluctuations in levels of KSHV-DNA reflected the patient's therapy and evolution of his underlying lymphoma. Other γ-herpesvirus-associated malignancies, such as multicentric Castleman disease and Kaposi sarcoma, or end-organ complications after salvage treatment were not found. Overall, these findings suggest a prognostic and predictive value of EBV-DNA and KSHV-DNA, the monitoring of which could be a simple, complementary tool for the management of γ-herpesvirus-positive lymphomas in HIV-1 patients submitted to ASCT.

Autograft HIV-DNA load predicts HIV-1 peripheral reservoir after stem cell transplantation for AIDS-related lymphoma patients.

Autologous stem cell transplantation (ASCT) is a widely used procedure for AIDS-related lymphomas, and it represents an opportunity to evaluate strategies curing HIV-1 infection. The association of autograft HIV-DNA load with peripheral blood HIV-1 reservoir before ASCT and its contribution in predicting HIV-1 reservoir size and stability during combination antiretroviral therapy (cART) after transplantation are unknown. Aiming to obtain information suggesting new functional cure strategies by ASCT, we retrospectively evaluated HIV-DNA load in autograft and in peripheral blood before and after transplantation in 13 cART-treated HIV-1 relapse/refractoring lymphoma patients. Among them seven discontinued cART after autograft infusion. HIV-DNA was evaluated by a sensitive quantitative real-time polymerase chain reaction (PCR). After debulking chemotherapy/mobilization, the autograft HIV-1 reservoir was higher than and not associated with the peripheral HIV-1 reservoir at baseline [median 215 HIV-DNA copies/10(6) autograft mononuclear cells, range 13-706 vs. 82 HIV-DNA copies/10(6) peripheral blood mononuclear cells (PBMCs), range 13-479, p = 0.03]. After high dose chemotherapy and autograft infusion, HIV-DNA levels reached a plateau between month 6 and 12 of follow-up. No association was found between peripheral HIV-DNA levels at baseline and after infusion in both cART interrupting and not interrupting patients. Only in the last subgroup, a stable significant linear association between autograft and peripheral blood HIV-1 reservoir emerged from month 1 (R(2) = 0.84, p = 0.01) to month 12 follow-up (R(2) = 0.99, p = 0.0005). In summary, autograft HIV-1 reservoir size could be influenced by the mobilization phase and predicts posttransplant peripheral HIV-1 reservoir size in patients on continuous cART. These findings could promote new research on strategies reducing the HIV-1 reservoir by using the ASCT procedure.

Multiplex analysis of blood cytokines as a prognostic tool in HIV related non-Hodgkin lymphoma patients: a potential role of interleukin-7.

BACKGROUND: Recent studies suggest a powerful prognostic value for blood cytokine levels in different diseases. Non-Hodgkin lymphoma (NHL) still represents one of the main causes of death in the HIV setting, with a wide variation in outcome and survival among patients. We measured blood concentrations of 11 cytokines from HIV-NHL patients at diagnosis and correlated these with the patient outcome to evaluate the prognostic value. METHODS: Luminex technology was used to simultaneously measure serum levels of interleukin IL-2/5/6/7/8/10/13/15, INF-γ, TNF-α and VEGF. Eighty-one consecutive HIV-NHL patients, at diagnosis, were studied. Hazard Ratios (HRs) and corresponding 95% confidence intervals (CIs) of disease-free survival (DFS) and overall survival (OS) were computed according to cytokine levels. HRs were also calculated for continuous variation of IL-7. RESULTS: In the multivariate analysis, statistically significant associations to both DFS and OS were found for IL-7 serum levels ≥ 3.2 pg/mL (HR=5.55, 95%CI:2.38-12.95; HR=3.53, 95%CI:1.60-7.77, respectively), IL-8 ≥ 18 pg/mL (HR=2.69, 95%CI:1.15-6.30; HR=2.35, 95%CI:1.01-5.51, respectively) and IL-10 ≥ 13 pg/mL (HR=2.82, 95%CI: 1.19-6.71; HR=2.98, 95%CI:1.21-7.30, respectively). When the multivariate analyses were mutually adjusted for INF-γ, IL-7, IL-8, IL-10 and IL-15, serum IL-7 ≥ 3.2 pg/mL emerged as factor independently associated to increased risk of DFS (HR=3.63, 95%CI:1.47-8.93) and OS (HR=3.97, 95%CI:1.49-10.57). CONCLUSIONS: IL-7, measured at NHL diagnosis, was the only cytokine strongly and independently associated to both DFS and OS. The multiplex analysis of different blood cytokines' concentration might be useful in defining additive predictive markers in HIV-NHL management and ascertainment of their outcome.

Assessment of immunovirological features in HIV related non-Hodgkin lymphoma patients and their impact on outcome.

BACKGROUND: Despite the era of highly active antiretroviral therapy, non-Hodgkin lymphoma (NHL) remains one of the main causes of death in HIV-infected patients, with a wide variation on the outcome. OBJECTIVES: We investigated immunological status and EBV, HHV8, HIV viral load in a group of HIV-infected patients at diagnosis of NHL to evaluate their prognostic significance. STUDY DESIGN: Eighty-one consecutive HIV+ NHL patients were studied. CD4 and CD8 cell counts, HHV8 DNA, EBV DNA, HIV RNA and HIV DNA were assessed at diagnosis and at 3 months after chemotherapy initiation. Hazard ratios (HRs) and corresponding 95% confidence intervals (CIs) of disease free survival (DFS) and overall survival (OS) were computed according to CD4 and CD8 cell counts, EBV DNA, HIV RNA and HIV DNA. HRs were, thereafter, computed also for continuous variation of CD4, CD8 cell counts and EBV DNA. RESULTS: In the multivariate analysis, CD4<160 and CD8<590 cell/µl and EBV DNA≥300 c/ml were independently associated to DFS (HR=2.98; 95%CI: 1.26-7.03; HR=2.65, 95%CI: 1.13-6.19; HR=4.01; 95%CI: 1.81-8.91) and OS (HR=3.32; 95%CI: 1.41-7.83; HR=4.62, 95%CI: 1.91-11.19; HR=3.11, 95%CI: 1.42-6.80). HRs for DFS and OS decreased continuously with increasing CD4 and CD8 cell counts, while they increased continuously with increasing EBV DNA levels. CONCLUSIONS: The association with survival of low CD4 and CD8 cell counts and detectable EBV viremia, measured at lymphoma's diagnosis, identified three independent prognostic biomarkers that might help in the management of NHL HIV+ patients, offering complementary information in the ascertainment of their outcome.

Predictive value of HIV type 1 DNA levels on overall survival in HIV-related lymphoma Patients treated with high-dose chemotherapy (HDC) plus autologous stem cell transplantation (ASCT).

The kinetics and predictive value of HIV-1 DNA (HIV DNA) levels in relapsed or refractory HIV lymphoma patients, treated with high-dose chemotherapy (HDC) followed by autologous stem cell transplantation (ASCT), were investigated. HIV DNA was measured by real-time PCR in the peripheral blood mononuclear cells (PBMCs) of 22 patients observed for a median follow-up of 31.0 months. At baseline, HIV DNA was found to be correlated with HIV-1 RNA (HIV RNA) (r = 0.56), but not with CD4(+) counts (r = -0.10). HIV RNA load was under control for the entire follow-up, while HIV DNA levels were almost always detectable (baseline levels vs. 1 year from ASCT levels, p > 0.05). Baseline HIV DNA levels were significantly different between alive and deceased patients (p = 0.03), and the overall survival (OS) analysis showed that for patients with higher HIV DNA levels at baseline there was a higher and nearly significant risk of death if compared to patients with lower levels (HR, 8.33, 95% CI, 0.99-70.06, p = 0.05). Our study demonstrated that high HIV DNA levels at baseline could predict overall survival after ASCT in one of the largest cohorts of HIV lymphoma patients treated with salvage therapy.

The Epstein Barr virus DNA levels as a tumor marker in EBV-associated cancers.

The Epstein Barr virus (EBV) is causally associated to several tumors of epithelial and lymphoid origin. The cancerogenic role in other than B cells has not been proven. This virus has been considered as a target in the effective diagnosis of EBV-associated tumors. For this purpose, molecular biology methods to measure EBV DNA load in the circulation of patients suffering from EBV-related cancers have been recently developed. In this review, we discuss the role of EBV DNA determination, the technical limitations of molecular assays measuring viral load and their impact on the clinical management of patients with EBV-associated tumors arising in the immunocompetent host. Several studies have recently clarified the biological and clinical characteristics of herpesvirus-associated tumors. However, some additional issues must be clarified before introducing viral load determinations into clinical practice. Firstly, since the various EBV-related tumors have different etiopathological and clinical characteristics, the most appropriate biological samples and analytical cut off values must be clearly defined in each group of patients. Secondly, a standardization of the assay, including the definition of the gene segment to be amplified, the use of an international reference for the standard curve and disease-related cut-off values, is strongly required. Thirdly, the interpretation of laboratory data may benefit from an improved design of the studies and obtaining an aggregate of patients from different institutions, pooling these together, in order to have a sample size that is adequate to reinforce the statistical power of the studies.

Serum antibody response to lytic and latent Epstein-Barr virus antigens in undifferentiated nasopharyngeal carcinoma patients from an area of nonendemicity.

Epstein-Barr virus (EBV)-associated undifferentiated carcinoma of the nasopharyngeal type (UCNT) is highly prevalent in southeast China, where immunoglobulin A (IgA) antibodies to viral capsid antigen and early antigen (EA) represent important markers, routinely used to assist in diagnosing this malignancy. Our study aimed at determining the EBV serological profiles of 78 UCNT patients from Italy, an area of nonendemicity for this tumor, using different assays specific for both lytic and latent EBV antigens. Serum IgA against both EA and EBNA1 and IgG and IgA to the latent membrane protein 1 (LMP1), to EA, and to the EBV transactivator ZEBRA protein were assessed. These serological responses were then evaluated according to the clinicopathologic parameters at diagnosis. The sensitivities of the IgG assays were 37.7% for LMP1, 73.6% for EA, and 61.0% for ZEBRA. EA/EBNA1 IgA reactivity was 84.4%, and a high association (odds ratio [OR], 2.6; 95% confidence interval [CI], 1.7 to 4.0) with UCNT was observed. When EBV serological reactivities were analyzed according to the tumor, node, and metastasis staging system (TNM), a statistically significant association was found between N stage and IgG antibody rates for EA (OR, 3.6; 95% CI, 1.2 to 10.9) and ZEBRA (OR, 2.6; 95% CI, 1.2 to 5.5) and between M stage and IgG antibody rates for ZEBRA (OR, 7.1; 95% CI, 3.2 to 16.0) and LMP1 (OR, 14.0; 95% CI, 1.8 to 110.9). Our results show that no single serological marker allows the detection of all UCNT cases. EA/EBNA1 IgA represents a reliable marker for diagnosis, with a high predictive value also in areas where UCNT is not endemic, such as Italy. The analysis of serological results according to TNM classification is consistent with a progressive impairment of humoral immune response to EBV as the disease advances and may be used to improve the accuracy of diagnosis.

High serum levels of soluble CD40-L in patients with undifferentiated nasopharyngeal carcinoma: pathogenic and clinical relevance.

BACKGROUND: Engagement of CD40 promotes survival of undifferentiated nasopharyngeal carcinoma (UNPC) cells and similar effects are induced by the EBV oncoprotein LMP-1 that is expressed in a fraction of cases. Considering that CD40 may be activated also by the soluble isoform of CD40L (sCD40L), we investigated the serum levels of sCD40L in a series of 61 UNPC patients from Italy, a non-endemic area for this disease. RESULTS: At diagnosis, serum samples of UNPC patients contained significantly higher levels of sCD40L than age-matched healthy controls (p < 0.001). High levels of sCD40L (i.e., >18 ng/ml) were more frequently found in patients <40 years of age (p = 0.03) and with distant metastases at presentation (p = 0.03). Serum levels of sCD40L were inversely associated with the expression of the EBV oncoprotein LMP-1 (p = 0.03), which mimics a constitutively activated CD40. The amount of sCD40L decreased in a fraction of patients treated with local radiotherapy alone. Moreover, CD40L+ lymphoid cells admixed to neoplastic UNPC cells were detected in cases with high serum levels of sCD40L, suggesting that sCD40L is probably produced within the tumor mass. CONCLUSION: sCD40L may contribute to CD40 activation in UNPC cells, particularly of LMP-1-negative cases, further supporting the crucial role of CD40 signalling in the pathogenesis of UNPC. sCD40L levels may be useful to identify UNPC patients with occult distant metastases at presentation.

Effects on virological and immunological parameters during CD34 mobilization in HIV patients with lymphoma.

The effects of CD34 mobilization with chemotherapy and G-CSF administration were evaluated in 13 HIV-positive patients with relapsed lymphomas and low CD4 counts. After mobilization, CD4+ cells increased significantly while HIV-RNA and integrated HIV-DNA showed no increases. G-CSF led to an increase of CD4+ cells with limited effects on HIV replication.

Clinical value of Epstein-Barr virus DNA levels in peripheral blood samples of Italian patients with undifferentiated carcinoma of nasopharyngeal type.

We investigated EBV viremia in matched serum and peripheral blood mononuclear cells (PBMCs) from one of the largest Italian cohort of Undifferentiated Carcinoma of Nasopharyngeal Type (UCNT) patients (N=34). By using a LMP-1 real-time PCR assay, we found that EBV DNA detection rate was 74% (median 8417 copies/ml) and 24% (median 164 copies/10(6)cells) on serum and PBMCs, respectively. Significantly higher serum EBV DNA levels were detected in patients with advanced UCNT (nodal stage N2 versus N0-1 and N3 versus N0-1, P=0.03 and 0.018; overall stage IV versus I-II, P=0.03). During the follow-up, there was also a statistically significant difference of EBV DNA viral load between patients with and without clinical relapse (P=0.008). We concluded that serum EBV DNA reflects the biological activity of the UCNT and may be a prognostic factor also in a low-incidence region.

Interleukin-10 and interleukin-18 promoter polymorphisms in an Italian cohort of patients with undifferentiated carcinoma of nasopharyngeal type.

PURPOSE: Cytokines such as IL-10 and IL-18 seem to be involved in the inflammatory response of undifferentiated carcinoma of nasopharyngeal type (UCNT). The aim of this study was to evaluate the correlation between functional single nucleotide polymorphisms (SNPs) in the promoter region of IL-10 and IL-18 genes and the virological and clinical characteristics in a large case series of Caucasian patients suffering from UCNT, a tumor regularly associated with the Epstein Barr Virus (EBV). METHODS: Eighty-nine patients with histologically confirmed UCNT and 130 healthy donors were included in our study. DNA was examined for the polymorphisms of IL-10 gene at positions -1082, -819, -592 by direct sequencing and IL-18 gene at position -607 and -137 by allele-specific PCR. EBV DNA serum viremia was evaluated by QC-PCR. RESULTS: The distributions of the IL-10 and IL-18 genetic variants were not different between UCNT patients and healthy controls. The frequency of IL-10 -1082G allele, which is associated with high IL-10 expression, showed a nearly statistically significant increase in UCNT patients EBV DNA-negative as compared to healthy controls (OR=3.3 95% CI: 1.2-9.8). Subjects with C/C or C/G combined IL-18 genotypes showed an increased risk of being with Stages III-IV (OR=2.1 95% CI: 1.2-6.6). CONCLUSION: This study was performed to improve the definition of the pathogenetic factors implicated in UCNT by addressing the correlation between cytokine polymorphisms and clinical parameters. This is the first study investigating the possible role of the IL-18 and IL-10 polymorphisms in the development and outcome of UCNT. In our genetic analysis there is no evidence for involvement of IL-10 promoter polymorphisms alone in the genetic predisposition to this tumor. On the other hand, IL18 genetic variants may represent a genetic risk factor for tumor aggressiveness.

Characterization of immunologic and virological parameters in HIV-infected patients with primary effusion lymphoma during antiblastic therapy and highly active antiretroviral therapy.

BACKGROUND: Primary effusion lymphoma (PEL) represents a peculiar lymphoma infected with human herpesvirus 8 (HHV-8) and occurs predominantly in human immunodeficiency virus (HIV)-infected patients. The aim of the present study was to evaluate the immunologic and virological parameters, including HHV-8 viremia, of 5 HIV-infected patients with PEL whose disease was diagnosed and treated at our institute. METHODS: Five patients were enrolled in the study. Biological parameters, such as latent and lytic HHV-8 antigen levels, plasma HHV-8 load, Epstein-Barr virus plasma DNA load, HIV-1 load, and CD4 cell count, were assessed before treatment, during therapy, and at follow-up. RESULTS: Four patients were treated with chemotherapy and highly active antiretroviral therapy (HAART), and 1 was treated with HAART alone; 3 of 5 patients reached complete remission. HHV-8 could be detected before the initiation of therapy in plasma from all patients analyzed. HHV-8 levels decreased after therapy in 4 patients. During the whole observation period, plasma HHV-8 load showed a statistically significant inverse correlation with CD4 cell count but no significant correlation with HIV load and response to therapy. CONCLUSIONS: Our analysis demonstrates that HHV-8 can be detected in the plasma at the onset of PEL; its prognostic role needs to be explored. CD4 cell count seems to be the most important indicator of progression of PEL.

Evaluation of three molecular biology-based assays for the detection of GB virus C/hepatitis G virus in clinical specimens.

OBJECTIVE: The present study was performed to evaluate the reliability of three reverse transcription-polymerase chain reaction (RT-PCR) assays, one commercial and two 'homebrew', for GB virus C (GBV-C)/hepatitis G virus (HGV) RNA detection in clinical specimens. We, therefore, investigated the virus prevalence with the method that gave us the best performances. METHODS: The commercial assay amplified sequences from the viral 5'-untranslated region (5'UTR) and non-structural 3 (NS3) region. The non-commercial assays 1 and 2 were based on different primers for the 5'UTR consensus sequence. RESULTS: The percentage of overall concordance by the three methods was 91.7%, raising to 93.0% when only the two non-commercial methods were compared. Assay 1 showed low sensitivity (57.1% vs. the commercial assay, 58.8% vs. assay 2), with 100% specificity. The commercial assay gave 18 of 54 (33.3%) 'false-negative' results, concordantly negative by the other assays. The prevalence of GBV-C/HGV RNA among the HIV+ patients was 27.0 and 32.6% in HIV/HCV co-infected patients. CONCLUSION: These data suggest that assay 2 has higher reliability as compared to the other two methods and may be used for an accurate GBV-C/HGV RNA detection in clinical and epidemiological studies.

Quantitative plasma/serum EBV DNA load by LMP2A determination in an Italian cohort of NPC patients.

BACKGROUND: Nasopharyngeal carcinoma (NPC) is frequently associated with Epstein-Barr virus (EBV), but little is known about the EBV DNA prevalence on peripheral blood in Western Countries, where the tumour is not endemic and its incidence is low. OBJECTIVES: To set up and evaluate an internally controlled qualitative polymerase chain reaction (PCR) followed by quantitative competitive PCR for the detection of EBV DNA in clinical specimens. To investigate whether EBV DNA load in peripheral blood was a consistent feature of Italian NPC patients. MATERIALS AND METHODS: A PCR assay based on latent membrane protein 2A (LMP2A) sequence amplification was chosen. Best assay conditions, sensitivity and reproducibility were determined. Sixty-four sera and 63 plasma from an Italian cohort of 39 NPC patients were analyzed. Samples from 5 patients followed up after radiotherapy were also assayed. Qualitative and quantitative beta-globin amplification was performed in parallel in order to provide an independent control for amplification competence of DNA and to investigate whether EBV DNA levels could be due to intracellular EBV viral genomes from cells lysed during plasma/serum collection. RESULTS: Twenty-five patients had undifferentiated carcinoma (UC) and 14 squamous cell carcinoma (SCC). EBV DNA has been quantified in 58 and 9% of the UC and SCC cases, respectively. No statistically significative differences were observed between the EBV DNA levels (469 vs 750 copies/ml, P=0.16) and prevalence (64 vs 57%, chi2(1)=0.22, P=0.64) in plasma and serum samples. Increased EBV viremia was found in patients with considerable extension of the primary tumour (172 vs 2250 copies/ml, low vs high tumour burden). Three UC subjects, which had detectable pre-treatment EBV DNA levels, became negative after radiotherapy. Clinical examination revealed that all had complete tumour regression. CONCLUSIONS: These PCR procedures allow an accurate and reproducible estimation of plasma/serum EBV DNA load in NPC patients living in non endemic areas, being strictly associated with UC WHO III and with tumour severity.

Interferon-gamma secretion and perforin expression are impaired in CD8+ T lymphocytes from patients with undifferentiated carcinoma of nasopharyngeal type.

An efficent antitumor and antiviral cellular immune response requires optimal interferon-gamma (IFN-gamma) secretion and perforin expression in CD8(+) T cells. The aim of this study was to define whether CD4(+) and CD8(+) T cells from patients with undifferentiated carcinoma of nasopharyngeal type (UCNT), a tumor regularly associated with the Epstein-Barr virus (EBV), have abnormal phenotype profiles, cytokine production, perforin and CD3-zeta expressions. Our data showed that CD4 and CD8 subset distribution was not grossly altered in the peripheral blood of UCNT patients, while tumor biopsies contained an increased proportion of CD8(+) T cells. The analysis of the CD4(+) subset showed a defect in interleukin-2 (IL-2) production and a moderate increase of IL-10 production, a situation consistent with a Th1/Th2 imbalance. We have also demonstrated that CD8(+) lymphocytes from UCNT patients had a marked impairment of IFN-gamma secretion and perforin expression. This impairment was not related to the presence of detectable EBV DNA in the plasma. In UCNT patients, the blockade of the perforin pathway and of IFN-gamma production may constitute important mechanisms for immune escape by the tumor and for impaired control of EBV replication.