Declines in serum CYFRA21-1 and carcinoembryonic antigen as predictors of chemotherapy response

EXPERIMENTAL AND THERAPEUTIC MEDICINE 4: 243-248, 2012

Declines in serum CYFRA21-1 and carcinoembryonic antigen as predictors of chemotherapy response and survival in patients with advanced non-small cell lung cancer

LIANG YANG1, XIN CHEN1, YUE LI2, JUN YANG1 and LI TANG1

Department of Echocardiography, The First Affiliated Hospital of China Medical University, Shenyang 110001; 2Department of Laboratory Medicine, Fengtian Hospital of Shenyang Medical College, Shenyang 110001, P.R. China

Received October 18, 2011; Accepted November 28, 2011DOI: 10.3892/etm.2012.570

Abstract. The aim of this study was to investigate the clinical value of serum cytokeratin 19 fragment (CYFRA21-1) and carcinoembryonic antigen (CEA) in the prediction of chemo-therapy response and prognosis in patients with advanced non-small cell lung cancer (NSCLC). Serum CYFRA21-1 and CEA levels of 98 patients with advanced NSCLC were measured using immunoradiometric kits prior to and after 2 cycles of chemotherapy. After 2 cycles of chemotherapy, 45 patients achieved a radiological objective response (OR), 30 patients achieved stable disease (SD) and 23 patients had progressive disease (PD). Serum CYFRA21-1 and CEA were significantly decreased compared to baseline levels (P<0.001). By ROC curve analysis, a ≥60% reduction in CYFRA21-1 and a ≥25% reduction in CEA were the optimal cut-off levels with best sensitivity and specificity for the diag-nosis of radiologic OR. The median survival of all patients was 10.2 months (range 2.6-26.3). Univariate survival analysis showed that the Eastern Cooperative Oncology Group (ECOG) performance status (PS) score, radiologic OR, a ≥60% reduction in CYFRA21-1 and a ≥25% reduction in CEA were significant prognostic factors for better overall survival. The median overall survival time in patients with a ≥60% reduction in CYFRA21-1 was significantly longer than in those with a <60% reduction (P<0.001). Similarly, the median overall survival time in patients with a ≥25% reduc-tion in CEA was also significantly longer than in those with a <25% reduction (P<0.001). Multivariate analysis showed that ECOG PS score, a ≥60% reduction in CYFRA21-1 and a ≥25% reduction in CEA were independent prognostic factors of survival, while radiologic OR was not. In conclusion, a ≥60% reduction in CYFRA21-1 and a ≥25% reduction in CEA may be reliable surrogate markers for the prediction of chemothrapy response and prognosis, especially for the diagnosis of radiologic OR.Introduction1Lung cancer is the leading cause of cancer-related death in the world (1). In 2010, an estimated 222,520 new cases and 157,300 deaths were anticipated in the US (2). Non-small cell lung cancer (NSCLC) accounts for more than 85% of all cases of lung cancer (3). Approximately 40% of patients with NSCLC present with advanced-stage disease at the time of diagnosis (3). The standard treatment for these patients is systemic chemotherapy, which improves both quality of life and survival (4). Until recently, platinum- or non-platinum-based, two-drug regimens were considered the standard of care for advanced NSCLC patients (4,5). However, the vast majority of patients with advanced NSCLC failed to benefit from combined chemotherapy (6). The WHO and Response Evaluation Criteria in Solid Tumors (RECIST) criterias, based on radiologic detections, were used to assess objective response after combined chemotherapy (7). Usually, the objective response (OR) of unmeasurable lesions, such as for atelectasis, pericardial effusion, pleural effusion, lymphatic vessel inva-sion and pleural-type tumors are difficult to evaluate using radiologic results (8). Moreover, a decrease in tumor volume as determined by radiologic images cannot accurately predict the survival of patients with advanced NSCLC (9). Therefore, more effective and feasible markers are required for the predic-tion of chemotherapy response and prognosis in patients with advanced NSCLC. Serum tumor markers, as a potential and Correspondence to: Dr Xin Chen, Department of Echocardiography, The First Affiliated Hospital of China Medical University, Shenyang more effective method to determine chemotherapy response and predict prognosis, have been studied extensively in the past. 110001, P.R. ChinaIt has been proven that cytokeratin 19 fragment (CYFRA21-1) E-mail: cmu_cx@http://wendang.chazidian.comand carcinoembryonic antigen (CEA) may be useful predictive Key words: cytokeratin 19 fragment, carcinoembryonic antigen, factors of chemotherapy response and prognosis in advanced non-small cell lung cancer, chemotherapy response, prognosisNSCLC patients. The aim of this study was to investigate the clinical value of serum CYFRA21-1 and CEA in the predic-tion of chemotherapy response and prognosis in patients with advanced NSCLC.

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YANG et al: SERUM CYFRA21-1 AND CEA IN PATIENTS WITH ADVANCED NSCLC

Materials and methods

Patient inclusion criteria. study, the following criteria were established: i) patients had a To be eligible for inclusion in this histological or cytological confirmation of clinical stage IIIB or IV NSCLC; ii) at least one measurable lesion; iii) patients were able to withstand at least 2 cycles of first-line platinum-based combined chemotherapy; iv) Eastern Cooperative Oncology Group (ECOG) performance status (PS) 0-2; v) no main organ dysfunction and hematopoietic function, normal liver and renal function, without any serious complications.

Assessment criteria of objective response. objective response (OR) were based on WHO and RECIST Assessments of the criteria (7), including complete response (CR), partial response (PR), stable disease (SD) or progressive disease (PD). OR was defined as CR plus PR, and no response (NR) was defined as SD plus PD. OR was evaluated and a confirmative chest computed tomography (CT) scan was performed after every 2 cycles of chemotherapy.

Assessment criteria of serum markers. from untreated lung cancer patients were prospectively Two serum samples collected: the first prior to the first cycle of chemotherapy, and the second after the second cycle of chemotherapy. All samples were frozen and stored at -80?C. All assays were performed using CIS Biointernational, France) (10), with investigators blinded commercial kits: CYFRA21-1 and CEA (ELSA; to clinical information. The cut-off value of CYFRA21-1 and CEA was 3.2 and 3.4 ng/ defined as assessable when at least one serum level of either ml, respectively. Patients were CYFRA21-1 or CEA was above the normal cut-off values.Statistical analysis. SPSS version 17.0 software (SPSS Inc., Chicago, IL, USA). Statistical analysis was performed using Comparisons of serum CYFRA21-1 or CEA prior to and after 2 cycles of chemotherapy were analyzed by Wilcoxon's signed rank test. Associations between categorical variables were evaluated using the the optimal cut-off levels of the declines of serum CYFRA21-1 χ2-test. The ROC curve was used to assess and CEA in the diagnosis of radiologic OR. The Youden index was used to identify the optimal cut-off levels.

survival (OS) was calculated from the initiation of chemotherapy All survival data were updated in May 1, 2011. Overall until death. Time to progression (TTP) was calculated from the date of registration to progression or last contact. Survival curves were generated with the Kaplan-Meier method and compared by the log-rank test and generalized Wilcoxon's test. Meanwhile, the multivariate survival analysis was performed to investigate the independent prognostic factors using Cox proportional hazards regression model. All tests were two-sided, and a value of P<0.05 was considered statistically significant.Results

Baseline characteristics of patients. and May 2010, a total of 98 patients with advanced NSCLC Between May 2006 who were admitted to The First Affiliated Hospital of China Medical University, were enrolled in this study. The median age was 58 years (range 27-84), including 65 males and

Table I. Baseline characteristics of the NSCLC patients.Characteristics

No. of patients (%)

Gender Male Female 65 (66.3)Age (years)

33 (33.7) Median (range) ≤65 58 (27-84) >65 60 (61.2)Histology

38 (38.8) Squamous cell carcinoma Adenocarcinoma

28 (28.6) Adenosquamous carcinomas 61 (62.2)Clinical stage 9 (9.2) III IV

53 (54.1)ECOG PS score45 (45.9) 0-1 2

54 (55.1)CYFRA21-1 baseline (ng/ml)44 (44.9) Median value (range) Normal (≤3.2) 6.4 (1.5-144.7) Abnormal (>3.2) 22 (22.4)CEA baseline (ng/ml)76 (77.6) Median value (range) Normal (≤3.4) 13.9 (1.1-985.1)

Abnormal (>3.4)

16 (16)Chemotherapy response82 (82)

CR PR 1 (1.0) SD 44 (44.9) PD

30 (30.6)Last follow-up status23 (23.5) Alive Dead

19 (19.4)79 (80.6)

CYFRA21-1, antigen; formance status; CR, complete response; PR, partial response; SD, ECOG, cytokeratin Eastern Cooperative 19 fragment; Oncology CEA, carcinoembryonic Group; PS, per-stable disease; PD, progressive disease.

33 females. There were 61 patients with adenocarcinomas, 28 with squamous carcinomas and 9 with adenosquamous carcinomas. According to the TNM staging system for lung cancer by the 6th edition of the International Union Against Cancer (UICC) and the American Joint Committee on Cancer (AJCC) (11), the study included 53 patients with stage IIIB and 45 patients with stage IVPS score of 0-1. All patients received a median of 4 cycles . There were 54 patients with a ECOG of chemotherapy (range 2-6). Baseline characteristics of the patients are shown in Table I.

Association between chemotherapy response and decreases in the serum markers. 45.9% (45/98) achieved OR after 2 cycles of chemotherapy,

Among 98 patients with advanced NSCLC,

EXPERIMENTAL AND THERAPEUTIC MEDICINE 4: 243-248, 2012245

Figure 1. (A) CYFRA21-1 and (B) CEA natural logarithm prior to and after 2 cycles of chemotherapy according to radiologic objective response (OR).

NR, no response.

including 1 patient with CR, 44 patients with PR, 30 patients with SD and 23 patients with PD. The median values of serum CYFRA21-1 prior to and after chemotherapy were 6.4 ng/ml (range 1.5-144.7) and 3.4 ng/ml (range 0.6-97.7), respectively. The median values of serum CEA prior to and after chemo-therapy were 13.9 ng/ml (range 1.1-985.1) and 3.9 ng/ml (range 1.0-375.5), respectively. After 2 cycles of chemotherapy, serum CYFRA21-1 and CEA were significantly decreased compared to baseline levels (P<0.0001).

sent serum CYFRA21-1 and CEA prior to chemotherapy, and As shown in Fig. 1, the values in the horizontal axis repre-those in the vertical axis represent serum CYFRA21-1 and CEA after 2 cycles of chemotherapy. This reflected the relationship between serum CYFRA21-1 or CEA and radiologic OR. If there was no significant difference in serum CYFRA21-1 or CEA prior to and after 2 cycles of chemotherapy, the corresponding points should fall along the straight line. In fact, most of the data points were below the line, especially for those of serum CYFRA21-1, suggesting that chemotherapy induced declines in serum CYFRA21-1 and/or CEA in the majority of patients.Figure 2. ROC curves analysis of serum (A) CYFRA21-1 and (B) CEA in the diagnosis of radiologic OR after 2 cycles of chemotherapy.ROC curves of serum CYFRA21-1 and CEA. ROC curves was carried out to assess the correlation between Analysis of the declines in serum CYFRA21-1 or CEA and radiologic OR after 2 cycles of chemotherapy (Fig. 2). The area under the ROC curve (AUC) was 0.727 (95% CI 0.600-0.857) for CYFRA21-1 and 0.629 (95% CI 0.48-0.771) for CEA. After 2 cycles of chemotherapy, a ≥60% reduction in CYFRA21-1 and a ≥25% reduction in CEA were the optimal cut-off levels with best sensitivity and specificity for the diagnosis of radiologic OR. When there was a ≥60% reduction in serum CYFRA21-1, the sensitivity and specificity values were 77.9 and 78.2%, respectively. When there was a ≥25% reduction in serum CEA, the -sensitivity and specificity were 70.5 and 68.7%, respectively. Therefore, a ≥60% reduction in CYFRA21-1 and a ≥25% reduction in CEA were defined as ‘serum marker response’.Univariate and multivariate survival analysis. study period, 79 of 98 (80.6%) patients with advanced NSCLC During the died. The median survival of all patients was 10.2 months

(range 2.6-26.3). As shown in Fig. 3, the median OS time in

246YANG et al: SERUM CYFRA21-1 AND CEA IN PATIENTS WITH ADVANCED NSCLC

Figure 3. Overall survival curve in patients with different serum (A) CYFRA21-1 and (B) CEA responses.

Table II. Univariate survival analysis.

Prognostic factors

Age (years) ≤65 >65 Gender Male Female No. of patients 60 38 65 33 37 61 59 39 28 61 9 55 53 22 76 16 82 36 62 40 58 MST (months) 10.1 11.3 11.2 10.2 95% CI 9.4-10.8 9.9-12.5 10.3-12.1 9.3-10.1 9.1-10.3 9.7-11.3P-value 0.504 0.170 0.761 0.001 0.088Clinical stage III IV ECOG PS score 0 1-2 10.6 10.9 11.7 9.6 11.2 9.7 10.4 11.3 9.8 10.7 11.1 10.7 10.2 11.6 9.3 11.2 8.9 Histology Squamous cell carcinoma Adenocarcinoma Adenosquamous carcinomas Radiologic OR Yes No 9.9-12.3 8.3-10.9 9.5-12.9 9.0-10.4 9.2-11.1 10.4-13.0 9.0-10.6 9.1-10.5 9.4-11.6 9.7-12.7 9.5-10.5 11.5-12.98.8-9.8 0.034 0.401 0.683<0.001<0.001CYFRA21-1 baseline level Normal >3.2 ng/ml CEA baseline level Normal >3.4 ng/ml ≥60% reduction in CYFRA21-1 Yes No ≥25% reduction in CEA Yes No 10.9-13.19.1-9.9

MST, median survival time; CI, confidence interval; ECOG, Eastern Cooperative Oncology Group; PS, performance status; CYFRA21-1, cytokeratin 19 fragment; CEA, carcinoembryonic antigen; OR, objective response.

EXPERIMENTAL AND THERAPEUTIC MEDICINE 4: 243-248, 2012247

Table III. Multivariate survival analysis.

Prognostic factors HR 95% CI P-valueECOG PS 0 <0.0001 1-2 1.000

Radiologic OR 2.904 1.763-4.784

Yes 1.000 0.1910 No ≥60% reduction in 1.563

CYFRA21-1 0.807-2.924 0.0010 No Yes 1.000

≥25% reduction in CEA 0.254 0.110-0.588

No 1.000 0.0380 Yes 0.417 0.182-0.954

HR, hazards ratio; CI, confidence interval, ECOG, Eastern Cooperative Oncology CYFRA21-1, Group; PS,

antigen.cytokeratin performance 19 fragment; status; CEA, OR, objective carcinoembryonic response. patients with a ≥60% reduction in CYFRA21-1 was significantly longer compared to those with a <60% reduction (11.6 -vs. 9.3 months, P<0.001). In addition, the median OS time in patients with a ≥25% reduction in CEA was also significantly longer compared to those with a <25% reduction (11.2 vs. 8.9 months, P<0.001).

radiologic OR, a ≥60% reduction in CYFRA21-1 and a ≥25% Univariate survival analysis showed that ECOG PS score, reduction in CEA were significant prognostic factors for better OS (Table II). However, age, gender, clinical stage, histological type and baseline levels of CYFRA21-1 and CEA were not related to prognosis. Results from the Cox regression analysis are shown in Table III. In this analysis, the independent prognostic roles of ECOG PS score, a ≥60% reduction in CYFRA21-1 and a ≥25% reduction in CEA were confirmed, while radiologic OR was not an independent prognostic factor.Discussion

CYFRA21-1 is an acidic protein of 40 kDa that is part of the cytoskeleton of epithelial cells (12). CYFRA21-1 is a specific and reproducible negative-prognostic marker for NSCLC (13). Many studies have confirmed that CYFRA21-1 is both a sensitive and specific tumor marker for NSCLC and especially for squamous cell carcinoma (14). It appears more sensitive and more specific than other tumor markers, such as CEA and NSE, and slightly better than squamous cell carcinoma-antigen (SCC) in squamous cell carcinoma (15). CEA is a glycoprotein expressed during early fetal life, and is the product of the CEACAM5-gen (16). CEA is an oncofetal protein attached to epithelial cell apical membrane via its C-terminal glycosylphosphatidylinositol anchor, a member of the immunoglobulin superfamily of cell adhesion molecules (17). Usually, CEA is overexpressed in a variety of neoplasms, such as colorectal, breast, bladder, gastric, pancreatic and lung carcinomas (16). CEA is a good monitoring marker for conventional chemotherapy. High serum CEA levels have been associated with disease progression and relapse in patients with advanced NSCLC (18). Several reports have been published concerning the prognostic value of serum tumor markers in patients with advanced NSCLC, for example CYFRA21-1, CEA, NSE and SCC (19). However, no reports concerning the relationship of declines in serum CYFRA21-1 or CEA with chemotheapy response and prognosis in patients with advanced NSCLC have been previously published. To our knowledge, this study is the first to propose that a ≥60% reduction in CYFRA21-1 and a ≥25% reduction in CEA after 2 cycles of chemotherapy can be regarded as possible surrogate markers of chemotherapy response and prognosis in patients with advanced NSCLC.CYFRA21-1 and CEA prior to and after 2 cycles of chemo-In the present study, we found that decreases in serum therapy also found that a ≥60% reduction in CYFRA21-1 and a ≥25% were correlated with chemotherapy response. We reduction in CEA after 2 cycles of chemotherapy were inde-pendent prognostic factors for patients with advanced NSCLC in multivariate survival analysis, while radiologic OR was not an independent prognostic factor. Nisman there was no correlation between radiologic OR and survival, et al (20) found that while declines in serum CYFRA21-1 after 2 cycles of chemo-therapy were closely related to survival. Ardizzoni studied 107 patients with advanced NSCLC and also observed et al (21) that declines in serum CYFRA21-1 and CEA were closely related to chemotherapy response and survival, whereas radiologic OR had no correlation with survival. Similar to previous studies, our study also demonstrated that the declines in serum CYFRA21-1 and CEA were closely related to chemotherapy response and survival, especially related to radiologic OR. After 2 cycles of chemotherapy, a ≥60% reduction in CYFRA21-1 and a ≥25% reduction in CEA were the -optimal cut-off levels, with best sensitivity and specificity for the diagnosis of radiologic OR. Univariate survival analysis showed that ECOG PS score, radiologic OR, a ≥60% reduction in CYFRA21-1 and a ≥25% reduction in CEA were significant prognostic factors. After 2 cycles of chemotherapy, the median OS time in patients with a ≥60% reduction in CYFRA21-1 was significantly longer compared to those with a <60% reduction. Similarly, the median OS time in patients with a ≥25% -reduction in CEA was also significantly longer compared to those with a <25% reduction. Multivariate analysis further confirmed the clinical value of declines in serum CYFRA21-1 and CEA in the prediction of chemotherapy response and prognosis in patients with advanced NSCLC. In multivariate analysis, the independent prognostic roles of ECOG PS score, ≥60% reduction in CYFRA21-1 and ≥25% reduction in CEA were confirmed, while radiologic OR was not an independent prognostic factor.tion in CYFRA21-1 and a ≥25% reduction in CEA may be In conclusion, our study demonstrated that a ≥60% reduc-reliable surrogate markers for the prediction of chemotherapy response and prognosis, particularly for the diagnosis of radio-logic OR. Due to a limitation in the sample of patients, this conclusion should be further confirmed by large case-control studies with an adequate methodological quality and properly controlled for possible confounds.