Post-treatment Plasma Epstein-Barr Virus DNA Research Articles

Improved risk stratification of nasopharyngeal cancer by targeted sequencing of Epstein–Barr virus DNA in post-treatment plasma

Quantitative measurement of plasma Epstein-Barr virus (EBV) DNA by real-time PCR at the end of primary treatment is a robust prognostic marker for nasopharyngeal carcinoma (NPC) patients. However, up to 40% of patients who would later develop disease recurrence had undetectable post-treatment plasma EBV DNA. Targeted sequencing for the entire EBV genome potentially allows a more comprehensive and unbiased detection of plasma EBV DNA and enables the use of other parameters such as fragment size as biomarkers. Hence, we explored if plasma EBV DNA sequencing might allow more accurate prognostication of NPC patients. Plasma samples collected from 769 patients with stage IIB-IVB NPC at 6-8 weeks after radiotherapy were analysed using targeted sequencing for EBV DNA. The sensitivities of the PCR-based analysis, at a cut-off of any detectable levels of plasma EBV DNA, for prediction of local and distant recurrences were 42.3% and 85.3%, respectively. The sequencing-based analysis (involving quantitation and size profiling) achieved better performance for both local and distant recurrences than PCR. Using a cut-off of the proportion of plasma EBV DNA deduced by sequencing at 0.01%, the sensitivities of the sequencing-based analysis for local and distant recurrences were 88.5% and 97.1%, with the resultant negative predictive values of 99.1% and 99.4%, respectively. Among patients with undetectable EBV DNA on quantitative PCR, sequencing could further define a subgroup that enjoyed superior survival outcomes based on the proportion of plasma EBV DNA, with a 5-year progression-free survival (PFS) approaching 90%. On multivariate analysis, sequencing-based quantitative level of plasma EBV DNA was the independent prognostic factor with the highest hazard ratio for prediction of overall survival and PFS. NPC prognostication using post-treatment plasma EBV DNA could be enhanced through sequencing.

Prognostic Value of Plasma Epstein-Barr Virus DNA Levels Pre- and Post-Neoadjuvant Chemotherapy in Patients With Nasopharyngeal Carcinoma.

BackgroundIn this study, we evaluated the prognostic value of the plasma levels of Epstein-Barr virus (EBV) DNA in patients with nasopharyngeal carcinoma (NPC) at different treatment stages.MethodsWe retrospectively analyzed the Data of 206 patients with NPC. Pre-neoadjuvant chemotherapy (pre-NACT), post-NACT, post-radiotherapy, and post-treatment plasma EBV DNA levels were used to establish prognostic nomograms. The concordance index (C-index) and calibration curves were used to compare the prognostic accuracy of the nomograms. The results were confirmed in a validation cohort consisting of patients who were tested for EBV DNA levels at all four stages of treatment. The Kaplan-Meier method was used to calculate the progression-free survival (PFS) and overall survival (OS). Survival differences were calculated using the log-rank test.ResultsEBV DNA-positive patients had worse 3-year PFS and 5-year OS than EBV DNA-negative patients; this was true for pre-NACT (PFS: 82.7% vs. 57.3%, P < 0.001; OS: 90.9% vs. 68.7%, P = 0.08) and post-NACT (PFS: 85.0% vs. 50.6%, P < 0.001; OS: 91.7% vs. 65.7%; P = 0.001) EBV DNA levels but not for post-radiotherapy (PFS: 72.2% vs. 60.9%, P = 0.192; OS: 73.1% vs. 77.2%, P = 0.472) or post-treatment (PFS: 77.3% vs. 59.2%, P = 0.063; OS: 77.5% vs. 79.7%, P = 0.644) levels. Nomograms combining pre-NACT and post-NACT EBV DNA levels had a superior prognostic ability than those of post-radiotherapy and post-treatment EBV DNA levels.ConclusionPre-NACT EBV DNA levels combined with post-NACT EBV DNA levels can more reliably predict survival outcomes in patients with NPC.

Integrating pre- and post-treatment Plasma Epstein-Barr Virus DNA levels for better prognostic prediction of Nasopharyngeal Carcinoma.

Background: Pre- and post-treatment plasma Epstein‐Barr virus (EBV) DNA are important biomarkers for the prognosis of nasopharyngeal carcinoma (NPC). This study was performed to determine the prognostic potential of integrating EBV DNA levels in plasma measured pre-treatment (pre-EBV) and 3 months post-treatment (3 m-EBV).Materials and methods: A total of 543 incident non-metastatic NPC patients treated with intensity-modulated radiotherapy, with or without chemotherapy, were reviewed. Patients were divided into four subgroups based on pre-EBV and 3 m-EBV status. The data for pre-EBV and 3 m-EBV samples were integrated, and the predictability of the survival of patients with NPC was analyzed.Results: There were significant differences in the 5-year progression-free survival, distant metastasis-free survival, locoregional relapse-free survival, and overall survival among the four patient subgroups (P<0.001). Patients who tested negative for both pre-EBV and 3 m-EBV had the best prognosis, followed by patients who tested positive for pre-EBV and negative for 3 m-EBV, and those who tested negative for pre-EBV and positive for 3 m-EBV; however, patients who tested positive for both pre-EBV and 3 m-EBV had the poorest chances of survival. Multivariate analyses demonstrated that integration of pre-EBV and 3 m-EBV data was an independent predictor of NPC progression in patients. Receiver operating characteristic curve analysis further confirmed that the combination of pre-EBV and 3 m-EBV had a greater prognostic value than pre-EBV or 3 m-EBV alone.Conclusions: Integrating pre-EBV and 3 m-EBV data could provide more accurate risk stratification and better prognostic prediction in NPC.

Superiority of Epstein-Barr Virus (EBV) DNA in the Plasma over Whole Blood in Prognostication of Extranodal NK/T Cell Lymphoma, Nasal Type (ENKTL)

Background Extranodal natural killer T cell lymphoma (ENKTL) is a rare subtype of non-Hodgkin lymphoma, of which pathogenesis is related to Epstein-Barr virus (EBV) infection. The presence of EBV-DNA in blood is a well-known prognosticator, which is also included in a prognostic model, PINK-E (Kim SJ et al, Lancet Oncol 2016). Furthermore, its presence after treatment is known to be a predictor of the treatment failure (Kim SJ et al, Lancet Haematol 2015). However, no consensus have been made on which blood sample (i.e, whole blood vs. plasma) is best to be used for the test. While EBV in plasma is known to have a higher specificity and sensitivity for EBV-related diseases as compared with EBV in peripheral blood mononuclear cells (Kanakry JA et al., Blood 2016), PINK-E model incorporated EBV status regardless of types of samples. We aimed to compare the prognostic performance of EBV DNA titers from whole blood (WB) and plasma. Method The EBV DNA viral load was measured in both WB and plasma samples by real-time quantitative PCR at the time of diagnosis, during and after completion of planned treatment. DNA was extracted using Artus® EBV RG PCR kit (Qiagen Inc.). The limit of detection level was 2.66 log copies/mL according to the manufacturer. Prognostic accuracy was compared by using time-dependent receiver operating characteristic (ROC) curves and corresponding area under the curve (AUC) values. Results A total of 60 patients with newly diagnosed ENKTL between September 2014 and September 2018 were included in this analysis. Twenty nine patients (48.3%) were in stage I, 6 (10.0%) in stage II, and 25 (41.7%) in stage IV. Forty six (76.7 %) patients had nasal involvement. Among them, 33 patients underwent concurrent chemoradiotherapy with weekly cisplatin and 53 received L-asparaginase containing chemotherapy (VIDL, n=46, 86.8%; SMILE, n=7, 13.2%) as definitive treatment or after chemoradiotherapy. Fifty two patients were evaluable for response: 33 (63.5%) complete response, 9 (17.3%) partial response, and 10 (19.2%) progressive disease. With a median follow-up duration of 34.1 months (range, 1.2-57.5 months), 2-year progression-free survival (PFS) and overall survival (OS) rates were 55.0% and 63.0%, respectively. EBV-DNA was detected in 37 (61.7%) in pretreatment WB and 23 patients (38.3%) in plasma samples respectively (Table 1). There was moderate agreement (ƙ=0.49) between the tests. Discordant results were noted in 16 (26.7%) patients, 15 of whom were WB (+) and plasma (-). Presence of EBV DNA in plasma was significantly correlated with unfavorable clinical characteristics than that in WB (Table 1). WB EBV positivity (2-year PFS rates, 42% vs 74%; HR 2.61, 95% CI, 1.04-6.52, p=0.041) and plasma EBV positivity (2-year PFS rates, 23% vs 75%; HR 4.87, 95% CI, 2.15-11.03, p&lt;0.0001) were both associated with poor PFS. Plasma EBV positivity at diagnosis was a highly significant prognosticator of OS (2-year OS rates of 27% vs. 85%; HR 8.38, 95% CI, 3.03-23.19, p&lt;0.0001). EBV status in the WB also tended to be associated with survival but without statistical significance (2-year OS rates, 47% vs 79%, HR 3.34, 95% CI, 1.12-9.96, P=0.071). Furthermore, WB EBV positivity in those with plasma EBV negativity was not prognostic for OS (2-year OS rates, 86% vs 84%, p=0.944) (Figure 1). On the other hand, plasma EBV status in those with WB EBV positivity was a significant predictor of OS (2yr OS rates, 84% vs. 29%, p&lt;0.001). Multivariable analysis showed plasma EBV positivity was significantly association with poor PFS (HR 4.22, 95% CI, 1.79-9.97, p=0.001) and OS (HR 8.39, 95% CI, 3.03-23.19, p=0.002) (Table 2). The prognostic accuracy assessed by the time dependent ROC curve analysis for 2-year OS rates also showed higher AUCs with plasma than WB (0.79, 95% CI, 0.63-0.92 for plasma; 0.66, 95% CI, 0.37-0.81 for WB). After completion of treatment, 22 patients were tested for EBV titer. Among those with plasma EBV positivity, WB EBV positivity was not associated with OS (2-year OS rates, 83% vs 62%, p=0.488). By contrast, plasma EBV positivity among those with WB EBV positivity was a significant predictor of OS (2yr OS rates, 62% vs. 17%, p=0.008) (Figure 2). In addition, post treatment plasma EBV DNA titer was a significant prognostic factors for PFS and OS by multivariable analysis (Table 3). Conclusion EBV DNA test in plasma is more accurate prognostic marker for ENKTL than that in WB as it can prognosticate regardless of EBV status in the WB. Disclosures No relevant conflicts of interest to declare.

Prognostic models for prediction of overall survival after first-line platinum-based chemotherapy for primary metastatic nasopharyngeal carcinoma.

e17501 Background: Platinum-based chemotherapy remains the main treatment strategy for patients with primary metastatic nasopharyngeal carcinoma (mNPC). However, survival outcomes are heterogeneous. The present study sought to develop clinical predictive models for overall survival (OS) in mNPC. Methods: Institutional database was queried for patients with mNPC who received first-line platinum-based chemotherapy between 2008 and 2015. Eligible patients (n = 460) were split into a training (2008-2013, n = 296) and an internal validation cohort (2014-2015, n = 164) according to the period of diagnosis. Potential prognostic factors for OS were evaluated using univariate and multivariate analyses. The performance of prognostic models was assessed by discrimination and calibration. The clinical usefulness of prognostic models was evaluated by decision curve analysis. Results: We established two nomogram-based prognostic models to predict OS probability, the first (baseline model) based on baseline factors, the second (composite model) incorporating posttreatment plasma Epstein-Barr virus DNA (EBV DNA) and objective response (OR) to first-line chemotherapy. The incorporation of posttreatment plasma EBV DNA and OR to the model improved the predictive accuracy. The C-index of composite model for predicting OS was 0.767 (P &lt; .001), which was statistically higher than the C-index values for baseline model (0.686). The results were confirmed in the internal validation cohort. Conclusions: We developed and validated two prognostic models that can be used before and after completion of first-line chemotherapy, respectively. The proposed models can be used for prognosis prediction and better selection of patient into clinical trials.

Differential clinical significance of pre-, interim-, and post-treatment plasma Epstein–Barr virus DNA load in NK/T-cell lymphoma treated with P-GEMOX protocol

Circulating EBV-DNA is an accurate biomarker of tumor load in extranodal natural killer (NK)/T cell lymphoma (ENKTL); however, its role in patients treated with P-GEMOX has not been evaluated. In this study, we examined plasma EBV-DNA of 99 patients at different time points by real-time quantitative polymerase chain reaction. Multivariate analysis revealed that ECOG PS score, response rate, and post-treatment EBV-DNA level were independent predictors of progression-free survival (PFS) and overall survival (OS). Positive post-treatment plasma EBV-DNA was associated with poor OS in ENKTL patients. The 3-year OS for patients with positive pre-, interim-, post-treatment EBV-DNA was significantly lower than that for patients with negative EBV-DNA; the values were 70.2% vs. 93.9% (p = .022), 53.8% vs. 99.1% (p < .001), and 40.6% vs. 91.8% (p < .001), respectively. We conclude that monitoring dynamic changes in plasma EBV-DNA in ENKTL patients treated with P-GEMOX could predict important outcomes such as OS.

Levels of plasma Epstein-Barr virus DNA prior and subsequent to treatment predicts the prognosis of nasopharyngeal carcinoma.

The level of Epstein-Barr virus DNA (EBV-DNA) in the plasma prior and subsequent to treatment is a reliable biomarker for the screening, diagnosis, monitoring and prognosis of nasopharyngeal carcinoma (NPC). The present retrospective study aimed to determine whether pre- and post-treatment levels of plasma EBV-DNA were predictive of survival in a large sample of patients with NPC. The level of plasma EBV-DNA in 637 NPC patients prior and subsequent to treatment was determined by quantitative polymerase chain reaction. The value of pre- and post-treatment plasma EBV-DNA in predicting the survival of NPC patients was then analyzed. The results revealed that pre-treatment plasma EBV-DNA loads were significantly higher in patients with NPC than those in healthy controls (P<0.001). The percentage of patients with positive plasma EBV-DNA was markedly higher prior to treatment (70.64%; median, 1150 copies/ml; range, 0-9.75×106 copies/ml) than following treatment (25.99%; median, 0 copies/ml; range, 0-3.83×106 copies/ml) (P<0.001). Patients with a high plasma EBV-DNA load presented with a higher clinical tumor classification, lymph node status, metastatic status and overall cancer stage. The risk of NPC relapse and mortality was higher in patients with pre-treatment plasma EBV-DNA levels of ≥1,500 copies/ml than that in patients with <1,500 copies/ml. Furthermore, the risk of relapse and mortality was higher in patients with positive post-treatment plasma EBV-DNA than in patients with negative post-treatment plasma EBV-DNA. Detectable post-treatment plasma EBV-DNA was the most significant prognostic factor to affect relapse-free survival, whilst metastasis was the prognostic factor with the greatest effect on overall survival. These data indicated that pre- and post-treatment levels of plasma EBV-DNA were able to predict the prognosis of NPC. This finding may provide novel references for research and clinical practice.

The clinical utility of plasma Epstein-Barr virus DNA assays in nasopharyngeal carcinoma: the dawn of a new era?: a systematic review and meta-analysis of 7836 cases.

In this study, we assessed the potential of plasma Epstein–Barr virus (EBV) DNA assays to predict clinical outcomes in a large sample of nasopharyngeal carcinoma (NPC) patients and proposed a risk stratification model based on standardized EBV DNA load monitoring.We conducted a meta-analysis of 14 prospective and retrospective comparative studies (n = 7 836 patients) to evaluate the correlation between pretreatment plasma EBV DNA (pre-DNA), midtreatment plasma EBV DNA (mid-DNA), posttreatment plasma EBV DNA (post-DNA), the half-life value of plasma EBV DNA clearance rate (t1/2), and clinical outcomes. Our primary endpoint was overall survival (OS). Our secondary endpoints were progression-free survival (PFS), distant-metastasis-free survival (DMFS), and local-regional-failure-free survival (LRFS).High pre-DNA, detectable mid-DNA, detectable post-DNA, and slow EBV DNA clearance rates were all significantly associated with poorer OS, with hazard radios (HRs) equal to 2.81, 3.29, 4.26, and 3.58, respectively. Pre-DNA, mid-DNA, and post-DNA had the same effects on PFS, DMFS, and LRFS.Plasma EBV DNA assays are highly prognostic of long-term survival and distant metastasis in NPC patients. Based on the results of this meta-analysis, we propose a 4-grade systematic risk stratification model. Given the inherent limitations of the included studies, future well-designed randomized clinical trials are required to confirm to the findings of this analysis and to contribute to the development of individualized treatment strategies for NPC patients.

Prognostic significance of circulating CD19+ B lymphocytes in EBV-associated nasopharyngeal carcinoma.

In this retrospective study, the correlation between pre- and post-treatment plasma Epstein-Barr virus (EBV) DNA and circulating immune subsets as well as the prognostic implications was investigated in nasopharyngeal carcinoma (NPC) patients. Patients (n=356) were diagnosed and received comprehensive treatment at the First People's Hospital of Foshan from 2006 to 2010. Pre- and post-treatment plasma EBV DNA load and circulating immune subsets (percentage of CD3+ T cell, CD3+ CD4+ T cells, CD3+ CD8+ T cells, CD19+ B cells and CD56+ NK cells) were analyzed by real-time PCR and flow cytometry. Patient age correlated negatively with CD3+ T cells (r=-0.264, P=0.001) and positively with CD56+ NK cells (r=0.272, P=0.001). Pre-treatment plasma EBV DNA correlated negatively with CD19+ B cells (r=-0.223, P=0.009) and CD4/CD8 ratio (r=-0.177, P=0.047). Patients with low CD19+ B cell had poorer 5-year progression-free survival (PFS) (66.6 vs. 81.8%, P=0.036) and 5-year overall survival (OS) (70.5 vs. 81.5%, P=0.097) than patients with high CD19+ B cells. Low CD19+ B cells was identified as a negative prognostic factor for 5-year PFS (hazard ratio [HR] 0.487; P=0.040), but not for 5-year OS (HR 0.550; P=0.102) in multivariate analysis. Post-treatment plasma EBV DNA was the most important prognostic factor for 5-year PFS (HR 2.983; P=0.006) and 5-year OS (HR 3.927; P<0.001). This study demonstrates the clinical value of circulating CD19+ B cell measurements in NPC patients.

Plasma Epstein-Barr virus DNA level strongly predicts survival in metastatic/recurrent nasopharyngeal carcinoma treated with palliative chemotherapy

Plasma Epstein-Barr virus (EBV) DNA is widely used in screening, monitoring, and prediction of relapse in nonmetastatic nasopharyngeal carcinoma (NPC). However, data regarding utility of plasma EBV DNA in metastatic NPC are rare. The current study was to test the prognostic implication of plasma EBV DNA level in metastatic/recurrent NPC patients treated with palliative chemotherapy. Plasma EBV DNA level was measured at baseline and thereafter at the start of each treatment cycle in 127 histologically proven metastatic/recurrent NPC patients treated with palliative chemotherapy. Correlations of pre-treatment and post-treatment plasma EBV DNA levels to survival and response were analyzed. Patients with a low pre-treatment plasma EBV DNA level (<median) had significantly better survival than those with a high pre-treatment plasma EBV DNA level (≥median). Patients with a post-treatment plasma EBV DNA decline to an undetectable level had better survival and better tumor response compared with those with a sustained detectable post-treatment plasma EBV DNA level. The early decrease of post-treatment plasma EBV DNA to an undetectable level after 1 cycle of chemotherapy was associated with significantly increased survival. Patients with low pre-treatment plasma EBV DNA level and undetectable post-treatment plasma EBV DNA showed a favorable prognosis (5-year overall and progression-free survival of 50.6% and 21.7%, respectively). Plasma EBV DNA is of predictive value for prognosis in metastatic/recurrent NPC patients undergoing palliative chemotherapy. The pre-treatment plasma EBV DNA level as well as the early decrease of plasma EBV DNA after chemotherapy enabled easy and early discrimination between patients who will and those who will not benefit from continued treatment.

Different Clinical Significance of Pre- and Post-treatment Plasma Epstein–Barr Virus DNA Load in Nasopharyngeal Carcinoma Treated with Radiotherapy

Aims To correlate the pre-treatment plasma Epstein–Barr virus (EBV) DNA with tumour burden and to explore the prognostic implications of pre- and post-treatment plasma EBV DNA load in nasopharyngeal carcinoma patients treated with radiotherapy. Materials and methods Plasma EBV DNA load was measured using a real-time quantitative polymerase chain reaction assay in 69 patients with nasopharyngeal carcinoma before and after radiation treatment and correlated with tumour volume and treatment outcome. Tumour volume was calculated by multiplying the sum of the areas of gross extent of the primary tumour and regional lymph nodes shown by computed tomography images and/or magnetic resonance imaging. Prognostic models for distant metastasis and overall survival were constructed using a multivariable fractional polynomial algorithm. Results The pre-treatment plasma EBV DNA concentration was significantly associated with tumour volume (Spearman correlation coefficient, 0.61; P < 0.001). The multivariable fractional polynomial algorithm selected post-treatment EBV DNA and administration of chemotherapy as prognostic factors for distant metastasis ( P < 0.001, P = 0.021, respectively), as well as for overall survival ( P < 0.001, P = 0.018, respectively). Conclusions Pre- and post-treatment plasma EBV DNA load have important clinical significance. Pre-treatment plasma EBV DNA concentration reflects tumour burden, whereas clearance of circulating plasma EBV DNA after treatment predicts the risk of distant metastasis and overall survival.

Plasma Epstein-Barr virus DNA and residual disease after radiotherapy for undifferentiated nasopharyngeal carcinoma.

Epstein-Barr virus (EBV) DNA can be detected and quantified in the plasma of patients with EBV-related tumors, such as nasopharyngeal carcinoma (NPC). Although NPC at early stages can be cured by radical radiotherapy, there is a high recurrence rate in patients with advanced NPC. The pretreatment level of circulating EBV DNA is a prognostic factor for NPC, but the prognostic value of post-treatment EBV DNA has not been studied. We designed a prospective study in Hong Kong, China, to investigate the value of plasma EBV DNA as a prognostic factor for NPC. One hundred seventy NPC patients, without metastatic disease at presentation, were treated with a uniform radiotherapy protocol. Circulating EBV DNA was measured by real-time quantitative polymerase chain reaction before treatment and 6-8 weeks after radiotherapy was completed. Risk ratios (RRs) were determined with a Cox regression model, and associations of various factors with progression-free and overall survival and recurrence rates were determined with a stepwise Cox proportional hazards model. All statistical tests were two-sided. Ninety-nine percent of patients achieved complete clinical remission. Levels of post-treatment EBV DNA dominated the effect of levels of pretreatment EBV DNA for progression-free survival. The RR for NPC recurrence was 11.9 (95% confidence interval [CI] = 5.53 to 25.43) for patients with higher post-treatment EBV DNA and 2.5 (95% CI = 1.14 to 5.70) for patients with higher pretreatment EBV DNA. Higher levels of post-treatment EBV DNA were statistically significantly associated with overall survival (P<.001; RR for NPC recurrence = 8.6, 95% CI = 3.69 to 19.97). The positive and negative predictive values for NPC recurrence for a higher level of post-treatment EBV DNA were 87% (95% CI = 58% to 98%) and 83% (95% CI = 76% to 89%), respectively. Levels of post-treatment plasma EBV DNA in patients with NPC appear to strongly predict progression-free and overall survival and to accurately reflect the post-treatment residual tumor load.