Ex vivo expansion of tumor-infiltrating lymphocytes from nasopharyngeal carcinoma patients for adoptive immunotherapy

Establishing Epstein-Barr virus (EBV)-specific cytolytic T lymphocytes (EBV-CTLs) from peripheral blood mononuclear cells (PBMCs) for adoptive immunotherapy has been reported in EBV-associated malignancies including Hodgkin's lymphoma and nasopharyngeal carcinoma (NPC). In the current study,we performed ex vivo expansion of tumor-infiltrating lymphocytes (TILs) obtained from NPC biopsy specimens with a rapid expansion protocol using anti-CD3 monoclonal antibody (OKT3), recombinant human interleukin (IL)-2, and irradiated PBMCs from healthy donors to initiate the growth of TILs. Young TIL cultures comprised of more than 90% of CD3+T cells, a variable percentage of CD3+CD8+and CD3+CD4+T cells, and less than 10% of CD3-CD16+natural killer cells, a similar phenotype of EBV-CTL cultures from PBMCs. Interestingly, TIL cultures secreted high levels of the Th1 cytokines, interferon gamma (IFNγ) and tumor necrosis factor-alpha (TNF-α), and low levels of the Th2 cytokines, IL-4 and IL-10. Moreover, young TILs could recognize autologous EBV-transformed B lymphoblast cell lines, but not autologous EBV-negative blast cells or allogeneic EBV-negative tumor cells. Taken together, these data suggest that ex vivo expansion of TILs from NPC biopsy tissue is an appealing alternative method to establish T cell-based immunotherapy for NPC.

Randomized study of sinusoidal chronomodulated versus flat intermittent induction chemotherapy with cisplatin and 5-fluorouracil followed by traditional radiotherapy for locoregionally advanced nasopharyngeal carcinoma

Neoadjuvant chemotherapy plus radiotherapy is the most common treatment regimen for advanced nasopharyngeal carcinoma(NPC).Whether chronomodulated infusion of chemotherapy can reduce its toxicity is unclear.This study aimed to evaluate the toxic and therapeutic effects of sinusoidal chronomodulated infusion versus flat intermittent infusion of cisplatin(DDP)and 5-fluorouracil(5-FU)followed by radiotherapy in patients with locoregionally advanced NPC.Patients with biopsy-diagnosed untreated stages III and IV NPC(according to the 2002 UICC staging system)were randomized to undergo2 cycles of sinusoidal chronomodulated infusion(Arm A)or flat intermittent constant rate infusion(Arm B)of DDP and 5-FU followed by radical radiotherapy.Using a"MELODIE"multi-channel programmed pump,the patients were given 12-hour continuous infusions of DDP(20 mg/m2)and 5-FU(750 mg/m2)for 5days,repeated every 3 weeks for 2 cycles.DDP was administered from 10:00 am to 10:00 pm,and 5-FU was administered from 10:00 pm to 10:00 am each day.Chronomodulated infusion was performed in Arm A,with the peak deliveries of 5-FU at 4:00 am and DDP at 4:00 pm.The patients in Arm B underwent a constant rate of infusion.Radiotherapy was initiated in the fifth week,and both arms were treated with the same radiotherapy techniques and dose fractions.Between June 2004 and June 2006,125 patients were registered,and 124 were eligible for analysis of response and toxicity.The major toxicity observed during neoadjuvant chemotherapy was neutropenia.The incidence of acute toxicity was similar in both arms.During radiotherapy,the incidence of stomatitis was significantly lower in Arm A than in Arm B(38.1%vs.59.0%,P=0.020).No significant differences were observed for other toxicities.The 1-,3-,and 5-year overall survival rates were 88.9%,82.4%,and 74.8%for Arm A and 91.8%,90.2%,and 82.1%for Arm B.The 1-,3-,and 5-year progression-free survival rates were 91.7%,88.1%,and 85.2%for Arm A and 100%,94.5%,and 86.9%for Arm B.The 1-,3-,and 5-year distant metastasis-free survival rates were 82.5%,79.1%,and 79.1%for Arm A and 90.2%,85.2%,and 81.7%for Arm B.Chronochemotherapy significantly reduced stomatitis but was not superior to standard chemotherapy in terms of hematologic toxicities and therapeutic response.

Detection of nasopharyngeal carcinoma using surface-enhanced laser desorption and ionization mass spectrometry profiles of the serum proteome

Background and Objective: Early diagnosis of nasopharyngeal carcinoma (NPC) is difficult due to the insufficient specificity of the conventional examination method. This study was to investigate potential and consistent biomarkers for NPC, particularly for early detection of NPC. Methods: A proteomic pattern was identified in a training set (134 NPC patients and 73 control individuals) using the surface-enhanced laser desorption and ionization-mass spectrometry (SELDI-MS), and used to screen the test set (44 NPC patients and 25 control individuals) to determine the screening accuracy. To confirm the accuracy, it was used to test another group of 52 NPC patients and 32 healthy individuals at 6 months later. Results: Eight proteomic biomarkers with top-scored peak mass/charge ratios (m/z) of 8605 Da, 5320 Da, 5355 Da, 5380 Da, 5336 Da, 2791 Da, 7154 Da, and 9366 Da were selected as the potential biomarkers of NPC with a sensitivity of 90.9% (40/44) and a specificity of 92.0% (23/25). The performance was better than the current diagnostic method by using the Epstein-Barr virus (EBV) capsid antigen IgA antibodies (VCA/IgA). Similar sensitivity (88.5%) and specificity (90.6%) were achieved in another group of 84 samples. Conclusion: SELDI-MS profiling might be a potential tool to identify patients with NPC, particularly at early clinical stages.

The role of capecitabine as maintenance therapy in de novo metastatic nasopharyngeal carcinoma:A propensity score matching study

Background:Capecitabine was previously used as a second-line or salvage therapy for metastatic nasopharyngeal carcinoma(NPC)and has shown satisfactory curative effect as maintenance therapy in other metastatic cancers.This study aimed to explore the role of capecitabine as maintenance therapy in de novo metastatic NPC patients with different plasma Epstein-Barr virus(EBV)DNA levels before treatment.Methods:We selected de novo metastatic NPC patients treated with locoregional radiotherapy(LRRT)for this retrospective study.The propensity score matching(PSM)was applied to balance potential confounders between patients who underwent capecitabine maintenance therapy and those who did not with a ratio of 1:3.Overall survival(OS)was the primary endpoint.The association between capecitabine maintenance therapy and survival was assessed using the log-rank test and a Cox proportional hazard model.Results:Among all patients eligible for this study,64 received capecitabine maintenance therapy after LRRT.After PSM,192 patients were identified in the nonmaintenance group.In the matched cohort,patients treated with capecitabine achieved a higher 3-year OS rate compared with patients in the non-maintenance group(68.5%vs.61.8%,P=0.037).Multivariate analysis demonstrated that capecitabine maintenance therapy was an independent prognostic factor.In subgroup analysis,3-year OS rate was comparable between the maintenance and non-maintenance groups in patients with high pretreatment EBV DNA levels(˃30,000 copies/mL)(54.8%vs.45.8%,P=0.835),whereas patients with low pretreatment EBV DNA levels(≤30,000 copies/mL)could benefit from capecitabine maintenance therapy in OS(90.0%vs.68.1%,P=0.003).Conclusion:Capecitabine maintenance therapy may be superior to non-maintenance therapy in prolonging OS for de novo metastatic NPC patients with pretreatment EBV DNA≤30,000 copies/mL.groups in patients with high pretreatment EBV DNA levels(˃30,000 copies/mL)(54.8%vs.45.8%,P=0.835),whereas patients with low pretreatment EBV DNA levels(≤30,000 copies/mL)could benefit from capecitabine maintenance therapy in OS(90.0%vs.68.1%,P=0.003).Conclusion:Capecitabine maintenance therapy may be superior to non-maintenance therapy in prolonging OS for de novo metastatic NPC patients with pretreatment EBV DNA≤30,000 copies/mL.

The diagnostic and prognostic values of plasma Epstein-Barr virus DNA for residual cervical lymphadenopathy in nasopharyngeal carcinoma patients:a retrospective study

Background:Currently,the diagnosis and treatment of nasopharyngeal carcinoma(NPC)patients with residual cervical lymphadenopathy following radical radiotherapy with or without chemotherapy are challenging.We investigated the prognosis of NPC patients with residual cervical lymphadenopathy and assessed the diagnostic and prognostic values of Epstein-Barr virus(EBV)DNA in these patients.Methods:This study included 82 NPC patients who were diagnosed with suspected residual cervical lymphadenopathy following completion of antitumor therapy.Their plasma EBV DNA levels were measured using quantitative polymerase chain reaction(qPCR)before the initiation of treatment and before neck dissection.Fine needle aspiration cytology(FNAC)was performed in 21 patients.All patients had undergone neck dissection and postoperative pathological examination to identify the nature of residual cervical lymphadenopathy.The overall survival(OS),progression-free survival(PFS),distant metastasis-free survival(DMFS),and locoregional relapse-free survival(LRRFS)were calculated using the Kaplan-Meier method and compared using the log-rank test.The Cox proportional hazards model was used to calculate hazard ratios(HRs)with 95%confidence intervals(CIs).Multivariable analysis was used to estimate the effect of potential prognostic factors on survival.Results:Following a median follow-up of 52.6 months,compared with patients with negative postoperative pathological findings for residual cervical lymphadenopathy,the patients with positive findings had a significantly lower 3-year PFS rate(49.9%vs.83.3%,P=0.008).Among NPC patients with residual cervical lymphadenopathy,the patients with preoperative plasma EBV DNA>0 copy/mL had a lower 3-year PFS rate than did those with no detectable EBV DNA(43.7%vs.61.1%,P=0.031).In addition,combining FNAC with preoperative EBV DNA detection improved the diagnostic sensitivity.Multivariable analysis demonstrated that residual cervical lymphadenopathy with positive postoperative pathological result was an independent prognostic factor for PFS and that detectable preoperative plasma EBV DNA was an independent prognostic factor for OS.Conclusions: Using FNAC combined with preoperative EBV DNA detection improves the sensitivity in diagnosing NPC with residual cervical lymphadenopathy. Compared with patients with undetectable EBV DNA, patients with detectable preoperative plasma EBV DNA have worse prognosis and may require a more aggressive treatment strategy.