Abstract

Background

We previously reported results of a prospective trial evaluating the significance of circulating tumor cells (CTCs) in patients with metastatic colorectal cancer (mCRC). This secondary analysis assessed the relationship of the CTC number with carcinoembryonic antigen (CEA) and overall survival.

Patients and methods

Patients with mCRC had CTCs measured at baseline and specific time points after the initiation of new therapy. Patients with a baseline CEA value ≥10 ng/ml and CEA measurements within ±30 days of the CTC collection were included.

Results

We included 217 patients with mCRC who had a CEA value of ≥10 ng/ml. Increased baseline CEA was associated with shorter survival (15.8 versus 20.7 months, P = 0.012). Among all patients with a baseline CEA value of ≥25 ng/ml, patients with low baseline CTCs (<3, n = 99) had longer survival than those with high CTCs (≥3, n = 58; 20.8 versus 11.7 months, P = 0.001). CTCs added prognostic information at the 3–5- and 6–12-week time points regardless of CEA. In a multivariate analysis, CTCs at baseline but not CEA independently predicted survival and both CTCs and CEA independently predicted survival at 6–12 weeks.

Conclusions

This study demonstrates that both CEA and CTCs contribute prognostic information for patients with mCRC.

introduction

Colorectal cancer (CRC) is the second leading cause of cancer death in the United States [1]. Treatment of patients with metastatic CRC (mCRC) typically consists of palliative chemotherapy. Numerous options are available with several agents in different stages of clinical development, making treatment algorithms complex. Thus, there is a need for the development of predictive and prognostic biomarkers to aid in treatment selection.

Carcinoembryonic antigen (CEA) was first identified in 1965 as an antigen that was present in both fetal colon and colon adenocarcinoma, but was absent from healthy adult colon [2, 3]. Several studies later showed that CEA was also present in healthy tissues, although at much lower concentrations [4]. Subsequently, CEA became one of the most widely used tumor markers. In early-stage CRC, it has prognostic significance [5, 6] and is well established to monitor for recurrence [7–9]. It is also widely used to monitor therapy in mCRC, although prognostic data are generally lacking [10].

Circulating tumor cells (CTCs) are rare malignant cells found in the peripheral blood that originate from the primary tumor and/or metastatic sites [11, 12]. CTCs represent a potential surrogate marker that may influence prognosis and decisions regarding treatment. CTCs have been shown to be a prognostic marker in various malignancies, most notably metastatic breast [13], colorectal [14, 15] and prostate cancer [16]. We have previously reported the results of a prospective clinical trial [14] where we found that the number of CTCs at baseline and during treatment was prognostic factors for progression-free survival (PFS) and overall survival (OS) in patients with mCRC beginning a new line of therapy. This report is a secondary analysis to examine the relationship of CTCs with CEA in this patient cohort.

patients and methods

study design

This was a prospective trial conducted at 55 centers in the United States, the Netherlands and the UK to evaluate the relationship between CTC number and response by imaging and the ability of CTC number to predict PFS and OS in patients with mCRC. Inclusion criteria and study design have been described previously [14]. Briefly, patients with mCRC with measurable disease, Eastern Cooperative Oncology Group (ECOG) performance status of 0–2 and hemoglobin of at least 8 g/dl were eligible. Peripheral blood was collected for CTC evaluation before the initiation of therapy (baseline) and subsequently at 1–20 weeks after initiating treatment. Blood samples were drawn into 10-ml evacuated tubes (CellSave, Veridex LLC, Raritan, NJ). All CTC evaluations were carried out without the knowledge of patient clinical status in one of four central laboratories. The CellSearch System (Veridex LLC), an immunomagnetic separation technique, was used for CTC enumeration, the technical details of which, including accuracy, precision, linearity and reproducibility, have been described previously [14]. Computed tomography or magnetic resonance imaging scans of the chest, abdomen and pelvis were to be carried out at baseline and every 6–12 weeks after initiating treatment.

measurement of CEA

The measurement of CEA was not an inclusion criterion and, therefore, was not mandatory for enrollment on the study. CEA values were drawn and analyzed at local laboratories and recorded on the case report forms for all available time points at baseline and 1–20 weeks after the initiation of treatment. These results were available to treating physicians to be utilized according to typical clinical practice. Only CEA values taken within ±30 days of the date of a given CTC measurement were included.

statistical analysis

The objective of this exploratory analysis was to evaluate the relationship of CEA with CTCs and the clinical outcome. To evaluate CEA in cases where it would be a clinically relevant marker, only patients with baseline CEA values ≥10 ng/ml at baseline were included in this analysis. Since there is no pre-defined clinical benefit threshold for CEA decline post-chemotherapy, we chose 25% and 50% reductions in the CEA value as typical thresholds considered clinically relevant. Proportions of patients among the various clinical classifications in the entire patient cohort and the CEA cohort were compared using Fisher's exact test. The median patient ages and years to metastasis were compared using the non-parametric k-sample χ2 test for the equality of the medians.

As CEA assessments were not mandated by the protocol and their values could influence clinical decision making regarding imaging and PFS, only OS was utilized as a clinical end point. OS was calculated from the time of the blood draw being evaluated until death or the last follow-up. Patients were censored at last follow-up if death had not occurred. Separate Kaplan–Meier survival plots were generated on the basis of (i) CEA levels at baseline and follow-up blood collections, (ii) CTC number at baseline and follow-up and (iii) the correlation of CEA levels and CTC number at baseline and follow-up. Survival curves were compared using log-rank testing. The Cox proportional hazards regression was used to determine univariate and multivariate hazards ratios for OS.

results

patient characteristics

Between February 2004 and November 2006, a total of 481 patients were enrolled. Patient characteristics for the entire cohort have been described previously [14]. Two hundred and seventeen patients had baseline CEA values ≥10 ng/ml. In this group of patients, CEA measurements taken within ±30 days of the CTC measurement were available at baseline in 209 patients, at 3–5 weeks in 115 patients and at 6–8 weeks in 134 patients. Table 1 lists the characteristics for the 217 patients comprising this analysis in comparison with the characteristics of the entire group of assessable patients (n = 430). The two groups had similar demographics (age, gender, race and performance status), tumor and treatment characteristics. A majority of the patients (71%) in this cohort were beginning first line chemotherapy.

Table 1.

Characteristics of patient cohort with baseline carcinoembryonic antigen ≥10 compared with all patients

 All patients (N = 430) CEA ≥ 10 (N = 217) P-valuea 
Age at baseline in years (median; range) 63.0 ± 12.6 (64; 22–92) 63.0 ± 12.3 (63; 22–90) 0.454b 
Gender 
 Female 192 (45%) 95 (44%) 0.867c 
 Male 238 (55%) 122 (56%) 
Race 
 White 311 (72%) 141 (65%) 0.336c 
 Black 44 (10%) 29 (13%) 
 Other 12 (3%) 6 ( 3%) 
 Unknown 63 (15%) 41 (19%) 
Baseline ECOG 
 0 196 (46%) 102 (47%) 0.974c 
 1 187 (43%) 93 (43%) 
 2 31 (7%) 16 (7%) 
 Unknown 16 (4%) 6 (3%) 
Site of disease at primary Dx 
 Colon 292 (68%) 152 (70%) 0.769c 
 Rectal 71 (17%) 31 (14%) 
 Rectosigmoid 66 (15%) 34 (16%) 
 Unknown 1 (0%) 0 (0%) 
Stage at primary Dx 
 1 12 (3%) 7 (3%) 0.638c 
 2 45 (11%) 16 (8%) 
 3 118 (27%) 59 (27%) 
 4 232 (54%) 122 (56%) 
 Unknown 23 (5%) 13 (6%) 
Line of therapy 
 First 308 (72%) 154 (71%) 0.825c 
 Second 96 (22%) 52 (24%) 
 Third 26 (6%) 11 (5%) 
 Unknown 0 (0%) 0 (0%) 
Liver mets 
 No 118 (27%) 45 (21%) 0.069c 
 Yes 312 (73%) 172 (79%) 
 Unknown 0 (0%) 0 (0%) 
Years to metastasis (median; range) 0.9 ± 1.4 (0.1; 0.0–10.8) 0.7 ± 1.3 (0.0; 0.0–7.2) 0.330b 
Follow-up time [average ± SD, median (range)] 
 Alive in months 27.9 ± 7.8, 29.5 (0.9–39.1) 26.9 ± 8.4, 29.0 (0.9–39.1) — 
 Dead 14.2 ± 9.0, 12.7 (0.2–36.0) 14.2 ± 8.4, 13.0 (0.2–35.5) — 
 All patients (N = 430) CEA ≥ 10 (N = 217) P-valuea 
Age at baseline in years (median; range) 63.0 ± 12.6 (64; 22–92) 63.0 ± 12.3 (63; 22–90) 0.454b 
Gender 
 Female 192 (45%) 95 (44%) 0.867c 
 Male 238 (55%) 122 (56%) 
Race 
 White 311 (72%) 141 (65%) 0.336c 
 Black 44 (10%) 29 (13%) 
 Other 12 (3%) 6 ( 3%) 
 Unknown 63 (15%) 41 (19%) 
Baseline ECOG 
 0 196 (46%) 102 (47%) 0.974c 
 1 187 (43%) 93 (43%) 
 2 31 (7%) 16 (7%) 
 Unknown 16 (4%) 6 (3%) 
Site of disease at primary Dx 
 Colon 292 (68%) 152 (70%) 0.769c 
 Rectal 71 (17%) 31 (14%) 
 Rectosigmoid 66 (15%) 34 (16%) 
 Unknown 1 (0%) 0 (0%) 
Stage at primary Dx 
 1 12 (3%) 7 (3%) 0.638c 
 2 45 (11%) 16 (8%) 
 3 118 (27%) 59 (27%) 
 4 232 (54%) 122 (56%) 
 Unknown 23 (5%) 13 (6%) 
Line of therapy 
 First 308 (72%) 154 (71%) 0.825c 
 Second 96 (22%) 52 (24%) 
 Third 26 (6%) 11 (5%) 
 Unknown 0 (0%) 0 (0%) 
Liver mets 
 No 118 (27%) 45 (21%) 0.069c 
 Yes 312 (73%) 172 (79%) 
 Unknown 0 (0%) 0 (0%) 
Years to metastasis (median; range) 0.9 ± 1.4 (0.1; 0.0–10.8) 0.7 ± 1.3 (0.0; 0.0–7.2) 0.330b 
Follow-up time [average ± SD, median (range)] 
 Alive in months 27.9 ± 7.8, 29.5 (0.9–39.1) 26.9 ± 8.4, 29.0 (0.9–39.1) — 
 Dead 14.2 ± 9.0, 12.7 (0.2–36.0) 14.2 ± 8.4, 13.0 (0.2–35.5) — 

aAll unknown percentages excluded from comparison of all patients and CEA ≥ 10 cohort to determine P-values.

bNon-parametric k-sample test on the equality of medians (continuity-corrected χ2P-value).

cFisher's exact test P-value.

ECOG, Eastern Cooperative Oncology Group; CEA, carcinoembryonic antigen.

CEA and overall survival

At baseline, 157 (75%) had a CEA value of ≥25 ng/ml. Although there was a shorter OS for this subset when compared with patients with a CEA value of <25 ng/ml (Figure 1A and Table 2), the difference was not statistically significant (P = 0.34). The median OS was substantially shorter for patients with baseline CEA levels of ≥50 ng/ml (n = 120) compared with those with baseline CEA levels of <50 ng/ml (Figure 1B and Table 2). Patients with persistently elevated CEA levels of ≥25 ng/ml at 3–5 and 6–12 weeks post-chemotherapy had substantially shorter OS compared with those whose CEA level declined to <25 ng/ml (Supplementary Figure S1, available at Annals of Oncology online and Figure 2, Table 2). Similarly, worse OS was noted for patients with a CEA level of ≥50 ng/ml at 3–5 and 6–12 weeks post-chemotherapy (Supplementary Figure S1, available at Annals of Oncology online and Figure 2, Table 2). As absolute CEA levels may not be the most clinically useful end point, we also analyzed OS based on 25% or 50% reductions of CEA at 3–5 and 6–12 weeks (Supplementary Figure S1, available at Annals of Oncology online and Figure 2C and D, respectively). Table 2 summarizes the results. As expected, OS was substantially longer for patients who had reductions in CEA after initiating treatment.

Table 2.

Relationship of carcinoembryonic antigen values and overall survival at baseline and 3–5-week and 6–12-week time points

Time point n Patients with CEA levels of ≥10 at baseline
 
CEA ≥ 25 ng/ml [n (%)] CEA < 25 ng/ml [median OS in months (95% CI)] CEA ≥ 25 ng/ml [median OS in months (95% CI)] Log-rank (P-value) 
 
 Baseline 209 157 (75%) 18.5 (14.2–23.9) 16.9 (13.2–19.6) 0.3408 
 3–5 weeks 115 68 (59%) 24.9 (20.5–NR) 12.5 (9.4–16.2) 0.0000 
 6–12 weeks 134 69 (52%) 22.0 (18.1–24.8) 13.8 (9.1–16.4) 0.0041 
  CEA ≥ 50 ng/ml [n (%)] CEA < 50 ng/ml [median OS in months (95% CI)] CEA ≥ 50 ng/ml [median OS in months (95% CI)]  
 Baseline 209 120 (57%) 20.7 (15.3–24.5) 15.8 (12.2–18.4) 0.0116 
 3–5 Weeks 115 52 (45%) 23.9 (16.8–29.3) 12.5 (8.8–16.2) 0.0001 
 6–12 Weeks 134 53 (40%) 21.5 (16.8–23.2) 10.7 (6.9–15.6) 0.0004 
  <25% reduction in CEA [n (%)] ≥25% reduction in CEA [median OS in months (95% CI)] <25% reduction in CEA [median OS in months (95% CI)]  
 3–5 Weeks 115 49 (43%) 23.0 (14.5–29.0) 12.6 (9.4–17.1) 0.0062 
 6–12 Weeks 134 35 (26%) 19.4 (14.7–22.6) 10.4 (6.9–17.1) 0.0035 
  <50% reduction in CEA [n (%)] ≥50% reduction in CEA [median OS in months (95% CI)] <50% reduction in CEA [median OS in months (95% CI)]  
 3–5 Weeks 115 73 (63%) 23.0 (13.6–29.3) 15.0 (11.1–18.8) 0.1845 
 6–12 Weeks 134 51 (38%) 21.9 (16.4–24.8) 10.7 (7.3–15.6) 0.0002 
Time point n Patients with CEA levels of ≥10 at baseline
 
CEA ≥ 25 ng/ml [n (%)] CEA < 25 ng/ml [median OS in months (95% CI)] CEA ≥ 25 ng/ml [median OS in months (95% CI)] Log-rank (P-value) 
 
 Baseline 209 157 (75%) 18.5 (14.2–23.9) 16.9 (13.2–19.6) 0.3408 
 3–5 weeks 115 68 (59%) 24.9 (20.5–NR) 12.5 (9.4–16.2) 0.0000 
 6–12 weeks 134 69 (52%) 22.0 (18.1–24.8) 13.8 (9.1–16.4) 0.0041 
  CEA ≥ 50 ng/ml [n (%)] CEA < 50 ng/ml [median OS in months (95% CI)] CEA ≥ 50 ng/ml [median OS in months (95% CI)]  
 Baseline 209 120 (57%) 20.7 (15.3–24.5) 15.8 (12.2–18.4) 0.0116 
 3–5 Weeks 115 52 (45%) 23.9 (16.8–29.3) 12.5 (8.8–16.2) 0.0001 
 6–12 Weeks 134 53 (40%) 21.5 (16.8–23.2) 10.7 (6.9–15.6) 0.0004 
  <25% reduction in CEA [n (%)] ≥25% reduction in CEA [median OS in months (95% CI)] <25% reduction in CEA [median OS in months (95% CI)]  
 3–5 Weeks 115 49 (43%) 23.0 (14.5–29.0) 12.6 (9.4–17.1) 0.0062 
 6–12 Weeks 134 35 (26%) 19.4 (14.7–22.6) 10.4 (6.9–17.1) 0.0035 
  <50% reduction in CEA [n (%)] ≥50% reduction in CEA [median OS in months (95% CI)] <50% reduction in CEA [median OS in months (95% CI)]  
 3–5 Weeks 115 73 (63%) 23.0 (13.6–29.3) 15.0 (11.1–18.8) 0.1845 
 6–12 Weeks 134 51 (38%) 21.9 (16.4–24.8) 10.7 (7.3–15.6) 0.0002 

CEA, carcinoembryonic antigen.

Figure 1

Relationship of ‘baseline carcinoembryonic antigen’ with overall survival using cutoffs of 25 ng/ml (A) and 50 ng/ml (B).

Figure 1

Relationship of ‘baseline carcinoembryonic antigen’ with overall survival using cutoffs of 25 ng/ml (A) and 50 ng/ml (B).

Figure 2

Relationship of carcinoembryonic antigen values at the 6–12-week blood draw with overall survival using cutoffs of 25 ng/ml (A), 50 ng/ml (B), 25% reduction (C) and 50% reduction (D).

Figure 2

Relationship of carcinoembryonic antigen values at the 6–12-week blood draw with overall survival using cutoffs of 25 ng/ml (A), 50 ng/ml (B), 25% reduction (C) and 50% reduction (D).

CTC number and the absolute CEA value

We also analyzed outcomes based on the CTC number (<3 and ≥3 CTCs/7.5 ml) and CEA values at baseline, 3–5 weeks and at 6–12 weeks. As expected, patients with <3 CTCs/7.5 ml and with a CEA level of <25 ng/ml at baseline (n = 48) had a longer median OS compared with those with ≥3 CTCs/7.5 ml and with a CEA level of ≥25 ng/ml (n = 58; 19.9 versus 11.7 months, P = 0.0065, Supplementary Figure S2, available at Annals of Oncology online and Figure 3A). Of note, among all patients with a baseline CEA value of ≥25 ng/ml, there was a distinct separation of survival curves among patients with low (<3 CTCs, n = 99) and high (≥3 CTCs, n = 58) baseline CTC numbers with superior median survival of 20.8 versus 11.7 months, P < 0.001. Among patients with a baseline CEA level of <25 ng/ml, too few patients had ≥3 CTCs for subgroup analysis.

Figure 3

Relationship between circulating tumor cell (CTC) number and carcinoembryonic antigen (CEA) values at baseline and overall survival using the CTC threshold of 3/7.5 ml and the CEA threshold of either 25 ng/ml (A) or 50 ng/ml (B)

Figure 3

Relationship between circulating tumor cell (CTC) number and carcinoembryonic antigen (CEA) values at baseline and overall survival using the CTC threshold of 3/7.5 ml and the CEA threshold of either 25 ng/ml (A) or 50 ng/ml (B)

A similar relationship was noted when a CEA cutoff of 50 ng/ml at baseline was used (Supplementary Figure S2, available at Annals of Oncology online and Figure 3B). Patients with <3 CTCs/7.5 ml and with a CEA level of <50 ng/ml (n = 77) had longer median OS compared with those with ≥3 CTCs/7.5mL and with a CEA level of ≥50 ng/ml (n = 50), 22.5 versus 12.1 months, P = 0.0002. As seen with the 25 ng/ml cutoff, among all patients with a CEA level of ≥50 ng/ml, patients with a low CTC number (<3 CTCs, n = 70) had a trend toward longer survival than those with a high CTC number (≥3 CTCs, n = 50), 17.5 versus 12.1 months, P = 0.06. Among all patients with a CEA level of <50 ng/ml, the elevated CTC number was associated with inferior survival (Figure 3B).

Significant differences in OS were also noted at the 3–5-week time point utilizing the subgroups above. As expected, patients with <3 CTCs/7.5 ml and with a CEA level of <25 ng/ml had longer survival compared with those with ≥3 CTCs/7.5 ml and with a CEA level of ≥25 ng/ml (median survival 24.4 versus 4.9 months, respectively, P < 0.0001). Similarly, the median OS was 23.2 compared with 4.9 months (P < 0.0001) for patients with <3 CTCs/7.5 ml and with a CEA level of <50 ng/ml versus those with ≥3 CTCs/7.5 ml and with a CEA level of ≥50 ng/ml. In patients with elevated CEA, the CTC number distinguished prognostic groups. Among patients with a CEA level of ≥ 25 or ≥50 ng/ml, patients with <3 CTCs/7.5 ml blood had a median survival of 15.3 versus only 4.9 months for patients with ≥3 CTCs/7.5 ml (P < 0.001). CTCs had less impact on differentiating prognostic groups within the group of patients with a CEA level of <25 ng/ml, perhaps reflecting the small number of patients in this group (n = 4 with ≥3 CTCs; Supplementary Figure S2, available at Annals of Oncology online).

Evaluating the 6–12-week time point, similar findings were noted. For patients with <3 CTCs/7.5 ml and with a CEA level of <25 ng/ml, the median survival was 22 compared with 5 months for patients with ≥3 CTCs/7.5 ml and with a CEA level of ≥25 ng/ml (P < 0.0001, Figure 4A). Patients with <3 CTCs/7.5 ml and with a CEA level of <50 ng/ml had longer survival compared with those with ≥3 CTCs/7.5 ml and with a CEA level of ≥50 ng/ml (median survival 21.9 versus 5 months, P < 0.0001; Figure 4B). Similar to the 3–5-week time point, the CTC number added prognostic information within CEA groupings. For example, among patients with elevated CEA (either ≥25 or ≥50 ng/ml), those with <3 CTCs/7.5 ml had improved survival compared with those with ≥3 CTCs/7.5 ml (P < 0.001 for both comparisons).

Figure 4

Relationship between circulating tumor cell number and carcinoembryonic antigen (CEA) values and OS at 6–12-week blood draw using the circulating tumor cell threshold of 3/7.5 ml and the CEA threshold of either 25 ng/ml (A), 50 ng/ml (B), 25% reduction (C) or 50% reduction (D).

Figure 4

Relationship between circulating tumor cell number and carcinoembryonic antigen (CEA) values and OS at 6–12-week blood draw using the circulating tumor cell threshold of 3/7.5 ml and the CEA threshold of either 25 ng/ml (A), 50 ng/ml (B), 25% reduction (C) or 50% reduction (D).

CTC number and percent CEA reduction

Sixty-six (57%) patients had a ≥25% reduction and 42 (37%) patients had a ≥50% reduction in the CEA value at 3–5 weeks post-chemotherapy when compared with baseline (Supplementary Figure S2, available at Annals of Oncology online). When divided into subgroups based on the number of CTCs per 7.5 ml at 3–5 weeks, a clear difference in OS was appreciated. As expected, the median OS for patients with <3 CTCs/7.5 ml and a CEA reduction of ≥25% was longer than those with ≥3 CTCs/7.5 ml and a CEA reduction of <25% (23.9 versus 8.7 months, P = 0.0025). Similarly, the median survival was 23.9 months for patients with <3 CTCs/7.5 ml and a CEA reduction of ≥50% versus 8.7 months for patients with ≥3 CTCs/7.5 ml and a CEA reduction of <50% (P = 0.0089). This was also true for the same cutoffs at the 6–12-week mark, as shown in Figure 4C and D. Interestingly, among patients with a similar CEA reduction, a important difference was noted in OS based on the CTC number at the 6–12-week blood draw. The median OS was 19.8 months for patients with a ≥25% reduction and <3 CTCs/7.5 ml compared with 5 months for patients with a ≥25% reduction and ≥3 CTCs/7.5 ml (P = 0.0001). Similarly, the median OS was higher for patients with <3 CTCs/7.5 ml (13.8 months) compared with those with ≥3 CTCs/7.5 ml (2.5 months, P = 0.0004) among all patients with a <25% reduction at the 6–12-week blood draw. At the 3–5-week time point, the CTC number did not appear to add additional prognostic information to percent CEA reduction.

univariate and multivariate Cox regression

Univariate Cox regression analysis to predict the risk of death was carried out using the OS time from the baseline blood draw, the 3–5-week blood draw and the 6–12-week blood draw separately (Supplementary Table S1, available at Annals of Oncology online). At baseline, ≥3 CTCs, a CEA level of ≥50 ng/ml, the patient's age at the time of the blood draw, the current line of therapy and the ECOG performance status at the time of study entry were substantial predictors of OS in a univariate analysis. In a multivariate Cox regression model using substantial parameters from the univariate analysis, all but CEA remained important in the prediction of OS from the time of the baseline blood draw (Table 3).

Table 3.

Multivariate Cox regression analysis for overall survival risk at baseline blood draw

Parameter Categories
 
OS risk from baseline draw
 
Positive (1) Negative (0) HR (95% CI) P-value Number of patients 
Baseline circulating tumor cell number ≥3 <3 1.5 (1.0–2.1) 0.053 203 
Baseline carcinoembryonic antigen value (ng/ml) ≥50 <50 1.3 (0.9–1.9) 0.122 
Age at baseline blood draw (years) ≥65 years <65 years 1.4 (1.0–2.0) 0.039 
Line of therapy Second or third First 1.7 (1.2–2.4) 0.005 
ECOG status at study entry 2 versus 1 versus 0 1.4 (1.1–1.8) 0.012 
Parameter Categories
 
OS risk from baseline draw
 
Positive (1) Negative (0) HR (95% CI) P-value Number of patients 
Baseline circulating tumor cell number ≥3 <3 1.5 (1.0–2.1) 0.053 203 
Baseline carcinoembryonic antigen value (ng/ml) ≥50 <50 1.3 (0.9–1.9) 0.122 
Age at baseline blood draw (years) ≥65 years <65 years 1.4 (1.0–2.0) 0.039 
Line of therapy Second or third First 1.7 (1.2–2.4) 0.005 
ECOG status at study entry 2 versus 1 versus 0 1.4 (1.1–1.8) 0.012 

ECOG, Eastern Cooperative Oncology Group; OS, overall survival; HR, hazard ratio.

At the time of the 3–5-week blood draw, in a univariate analysis, only CEA (≥25 and ≥50 ng/ml), a ≥25% reduction in CEA and the current line of therapy were important in the prediction of OS (Supplementary Table S1, available at Annals of Oncology online). Both CEA and the line of therapy remained important in multivariate Cox regression models.

At the time of the 6–12-week blood draw, ≥3 CTCs, CEA (≥25 and ≥50 ng/ml), reductions in CEA (≥25% and ≥50% reductions) and the current line of therapy were univariately important in the prediction of OS (Supplementary Table S1, available at Annals of Oncology online). Multivariate Cox regression analysis was modeled including CTCs, one measure of CEA and the current line of therapy. CTCs were the most substantial predictor of OS and only CEA at thresholds of >50 ng/ml or a ≥50% reduction in CEA remained multivariately important (Supplementary Table S2A–D, available at Annals of Oncology online).

correlation of CTCs and CEA

At baseline, there appeared to be no real correlation between the CTC and CEA, and the Pearson's correlation coefficient was 0.0370 (n = 209, P = 0.5950) indicating no significant correlation. Similarly, there was no significant correlation between the CTC and CEA at the other time points (weeks 1–2, weeks 3–5 and weeks 6–12).

discussion

This secondary analysis from our previously reported multicenter study demonstrates that CTCs add prognostic information to CEA at baseline and shortly after a new therapy begins for mCRC. We analyzed absolute levels and percent decrease in CEA at different time points with consistent findings regardless of the analysis method. To our knowledge, this trial is the first to report the relationship of absolute CEA values and CEA percentage reductions to CTCs and the outcome in patients with mCRC.

Despite widespread availability and common use of CEA, its role in patients with mCRC is still not well-defined. Preoperative CEA levels have been shown to be a reliable prognostic marker, with worse outcome seen with high preoperative CEA [17, 18]. An inverse relationship of preoperative CEA with prognosis has also been noted in patients with mCRC undergoing liver meta-statectomy in two independent studies [19, 20]. After the resection of primary CRC, serial elevations of CEA are often used as a trigger to evaluate for possible recurrence [21]. For patients with metastatic disease undergoing chemotherapy, CEA has not been shown to predict antitumor response. Despite lack of clear consensus on CEA monitoring in mCRC patients, CEA levels are widely used for monitoring response to chemotherapy [22]. There are no clear criteria or treatment algorithms to modify management based on the CEA value alone in this subset of patients. In our study of mCRC patients undergoing chemotherapy, higher baseline CEA was associated with inferior survival. In a univariate analysis, CTCs at a threshold of ≥3/7.5 ml, CEA at a threshold of ≥50 ng/ml and the current line of therapy were important in the prediction of OS at both baseline and the 6–12-week time points. The CTC was the only substantial variable in a multivariate analysis at baseline. At the 6–12-week time point, CTCs were the most substantial predictor of OS and only CEA at a threshold of ≥50 ng/ml or a ≥50% reduction in CEA remained multivariately important.

The treatment of mCRC has become substantially more nuanced over the last decade, with five classes of systemic agents available and various strategies for treatment. Patients may begin with one, two or three chemotherapy drugs with or without an antibody [23, 24]. They may drop part of their therapy or take complete treatment breaks [25, 26]. As patients are living longer, the need for non-radiologic assessment to evaluate the intensity and sequence of systemic therapy is increasing.

Our study thus raises a number of important clinical questions for consideration. Should CTC and CEA measurements dictate treatment decision-making in the absence of radiographic documentation? At the current time, the answer to this question is no. However, clinical trials to assess the utility of an early change in therapy based on poor response by the CTC and CEA are warranted. Further, the technology utilized in this trial (CellSearch® Veridex) yields relatively few CTCs in mCRC compared with other epithelial malignancies such as prostate and breast cancer. Emerging technologies and strategies with potentially increased sensitivity compared with the CellSearch system have promise to increase the CTC yield and thus clinical relevance [27–29].

This study has several limitations which should be acknowledged. For one, cutoffs used for CEA analysis were arbitrary and CEA was not mandated by the protocol. Thus, there could be inherent selection bias for those patients who had it drawn as part of the trial. However, patient characteristics among groups of patients who did and did not have CEA drawn are similar which may mitigate this potential impact. In addition, we specifically selected OS rather than PFS in our analysis so that there would be little bias with the relationship of CEA with our clinical end point. Finally, CEA cutoffs were selected as those felt to be most relevant clinically.

A second potential limitation is that our analysis includes patients from this trial who were receiving various lines of therapy. However, the majority of patients received first line therapy. As mentioned in our prior publications, CTCs retained their relationship with the clinical outcome regardless of line of therapy. The relatively small patient numbers with CEA drawn precluded further subset analysis by line or type of chemotherapy. Finally, the exact timing of CEA was not mandated in this protocol. However, as it usually corresponded to timing of CTC blood draws the current report most accurately reflects clinical practice.

In conclusion, this study demonstrates that CEA and CTCs add independent prognostic information for patients undergoing chemotherapy for mCRC. The potential role of both biomarkers in treatment decision making requires further prospective study. Consideration should be given to incorporating CEA and CTC measurements as stratification factors in future prospective randomized trials of patients with mCRC beginning a new line of chemotherapy.

funding

This study was funded by a grant from (Immunicon) Veridex LLC.

disclosure

MCM is an employee of Veridex, LLC and was instrumental in the analysis of the results of this trial.

references

1
Jemal
A
Siegel
R
Xu
J
, et al.  . 
Cancer statistics
CA Cancer J Clin
 , 
2010
, vol. 
60
 (pg. 
277
-
300
)
2
Gold
P
Freedman
SO
Specific carcinoembryonic antigens of the human digestive system
J Exp Med
 , 
1965
, vol. 
122
 (pg. 
467
-
481
)
3
Gold
P
Freedman
SO
Demonstration of tumor-specific antigens in human colonic carcinomata by immunological tolerance and absorption techniques
J Exp Med
 , 
1965
, vol. 
121
 (pg. 
439
-
62
)
4
Boucher
D
Cournoyer
D
Stanners
CP
, et al.  . 
Studies on the control of gene expression of the carcinoembryonic antigen family in human tissue
Cancer Res
 , 
1989
, vol. 
49
 (pg. 
847
-
852
)
5
Wiggers
T
Arends
JW
Volovics
A
Regression analysis of prognostic factors in colorectal cancer after curative resections
Dis Colon Rectum
 , 
1988
, vol. 
31
 (pg. 
33
-
41
)
6
Wolmark
N
Fisher
B
Wieand
HS
, et al.  . 
The prognostic significance of preoperative carcinoembryonic antigen levels in colorectal cancer. Results from NSABP (National Surgical Adjuvant Breast and Bowel Project) clinical trials
Ann Surg
 , 
1984
, vol. 
199
 (pg. 
375
-
382
)
7
Bhattacharjya
S
Aggarwal
R
Davidson
BR
Intensive follow-up after liver resection for colorectal liver metastases: results of combined serial tumour marker estimations and computed tomography of the chest and abdomen—a prospective study
Br J Cancer
 , 
2006
, vol. 
95
 (pg. 
21
-
26
)
8
Zeng
Z
Cohen
AM
Urmacher
C
Usefulness of carcinoembryonic antigen monitoring despite normal preoperative values in node-positive colon cancer patients
Dis Colon Rectum
 , 
1993
, vol. 
36
 (pg. 
1063
-
1068
)
9
Petrioli
R
Licchetta
A
Roviello
G
, et al.  . 
CEA and CA19.9 as early predictors of progression in advanced/metastatic colorectal cancer patients receiving oxaliplatin-based chemotherapy and bevacizumab
Cancer Invest
 , 
2012
, vol. 
30
 (pg. 
65
-
71
)
10
Aldulaymi
B
Bystrom
P
Berglund
A
, et al.  . 
High plasma TIMP-1 and serum CEA levels during combination chemotherapy for metastatic colorectal cancer are significantly associated with poor outcome
Oncology
 , 
2010
, vol. 
79
 (pg. 
144
-
149
)
11
Gupta
GP
Massague
J
Cancer metastasis: building a framework
Cell
 , 
2006
, vol. 
127
 (pg. 
679
-
695
)
12
Allard
WJ
Matera
J
Miller
MC
, et al.  . 
Tumor cells circulate in the peripheral blood of all major carcinomas but not in healthy subjects or patients with nonmalignant diseases
Clin Cancer Res
 , 
2004
, vol. 
10
 (pg. 
6897
-
6904
)
13
Cristofanilli
M
Budd
GT
Ellis
MJ
, et al.  . 
Circulating tumor cells, disease progression, and survival in metastatic breast cancer
N Engl J Med
 , 
2004
, vol. 
351
 (pg. 
781
-
791
)
14
Cohen
SJ
Punt
CJ
Iannotti
N
, et al.  . 
Relationship of circulating tumor cells to tumor response, progression-free survival, and overall survival in patients with metastatic colorectal cancer
J Clin Oncol
 , 
2008
, vol. 
26
 (pg. 
3213
-
3221
)
15
Tol
J
Koopman
M
Miller
MC
, et al.  . 
Circulating tumour cells early predict progression-free and overall survival in advanced colorectal cancer patients treated with chemotherapy and targeted agents
Ann Oncol
 , 
2010
, vol. 
21
 (pg. 
1006
-
1012
)
16
de Bono
JS
Scher
HI
Montgomery
RB
, et al.  . 
Circulating tumor cells predict survival benefit from treatment in metastatic castration-resistant prostate cancer
Clin Cancer Res
 , 
2008
, vol. 
14
 (pg. 
6302
-
6309
)
17
Wanebo
HJ
Rao
B
Pinsky
CM
, et al.  . 
Preoperative carcinoembryonic antigen level as a prognostic indicator in colorectal cancer
N Engl J Med
 , 
1978
, vol. 
299
 (pg. 
448
-
451
)
18
Grem
J
The prognostic importance of tumor markers in adenocarcinomas of the gastrointestinal tract
Curr Opin Oncol
 , 
1997
, vol. 
9
 (pg. 
380
-
387
)
19
Fong
Y
Fortner
J
Sun
RL
, et al.  . 
Clinical score for predicting recurrence after hepatic resection for metastatic colorectal cancer: analysis of 1001 consecutive cases
Ann Surg
 , 
1999
, vol. 
230
 (pg. 
309
-
318
discussion 318–321
20
Nordlinger
B
Guiguet
M
Vaillant
JC
, et al.  . 
Surgical resection of colorectal carcinoma metastases to the liver. A prognostic scoring system to improve case selection, based on 1568 patients. Association Francaise de Chirurgie
Cancer
 , 
1996
, vol. 
77
 (pg. 
1254
-
1262
)
21
National Comprehensive Cancer Network. Guidelines for management of colon cancer
 
22
Reiter
W
Stieber
P
Reuter
C
, et al.  . 
Multivariate analysis of the prognostic value of CEA and CA 19–9 serum levels in colorectal cancer
Anticancer Res
 , 
2000
, vol. 
20
 (pg. 
5195
-
5198
)
23
Falcone
A
Ricci
S
Brunetti
I
, et al.  . 
Phase III trial of infusional fluorouracil, leucovorin, oxaliplatin, and irinotecan (FOLFOXIRI) compared with infusional fluorouracil, leucovorin, and irinotecan (FOLFIRI) as first-line treatment for metastatic colorectal cancer: the Gruppo Oncologico Nord Ovest
J Clin Oncol
 , 
2007
, vol. 
25
 (pg. 
1670
-
1676
)
24
Tol
J
Koopman
M
Cats
A
, et al.  . 
Chemotherapy, bevacizumab, and cetuximab in metastatic colorectal cancer
N Engl J Med
 , 
2009
, vol. 
360
 (pg. 
563
-
572
)
25
Chibaudel
B
Maindrault-Goebel
F
Lledo
G
, et al.  . 
Can chemotherapy be discontinued in unresectable metastatic colorectal cancer? The GERCOR OPTIMOX2 Study
J Clin Oncol
 , 
2009
, vol. 
27
 (pg. 
5727
-
5733
)
26
Tournigand
C
Cervantes
A
Figer
A
, et al.  . 
OPTIMOX1: a randomized study of FOLFOX4 or FOLFOX7 with oxaliplatin in a stop-and-Go fashion in advanced colorectal cancer—a GERCOR study
J Clin Oncol
 , 
2006
, vol. 
24
 (pg. 
394
-
400
)
27
Maheswaran
S
Sequist
LV
Nagrath
S
, et al.  . 
Detection of mutations in EGFR in circulating lung-cancer cells
N Engl J Med
 , 
2008
, vol. 
359
 (pg. 
366
-
377
)
28
Nagrath
S
Sequist
LV
Maheswaran
S
, et al.  . 
Isolation of rare circulating tumour cells in cancer patients by microchip technology
Nature
 , 
2007
, vol. 
450
 (pg. 
1235
-
1239
)
29
Sequist
LV
Nagrath
S
Toner
M
, et al.  . 
The CTC-chip: an exciting new tool to detect circulating tumor cells in lung cancer patients
J Thorac Oncol
 , 
2009
, vol. 
4
 (pg. 
281
-
283
)