Clinico-pathological correlation of serial measurement of circulating tumor cells in 24 metastatic colorectal cancer patients receiving chemotherapy reveals interpatient heterogeneity correlated with CEA levels but independent of KRAS and BRAF mutation

Figures & data

Table 1. Blood draw protocol for patients on FOLFOX/FOLFIRI regimes. Blood can be drawn upto 7 times during the course of chemotherapy.

Chemotherapy	Blood samples
Baseline Day −30 to −1	22 ml(3 tubes of 7.5 ml)
Cycle 1 Day 1	15 ml(2 tubes of 7.5 ml)
Cycle 1 Day 7	15 ml(2 tubes of 7.5 ml)
Cycle 2 Day 1	15 ml(2 tubes of 7.5 ml)
Cycle 3 Day 1	22 ml(3 tubes of 7.5 ml)
Cycle 4 Day 1	15 ml(2 tubes of 7.5 ml)
Cycle 6 Day 15	22 ml(3 tubes of 7.5 ml)

Table 2. Blood draw protocol for patients on XELODA (capecitabine)-based chemotherapy.

Chemotherapy	Blood samples
Baseline Day −30 to −1	22 ml(3 tubes of 7.5 ml)
Cycle 1 Day 1	15 ml(2 tubes of 7.5 ml)
Cycle 1 Day 7	15 ml(2 tubes of 7.5 ml)
Cycle 2 Day 1	15 ml(2 tubes of 7.5 ml)
Cycle 3 Day 1	22 ml(3 tubes of 7.5 ml)
Cycle 4 Day 1	15 ml(2 tubes of 7.5 ml)
Cycle 6 Day 1	22 ml(2 tubes of 7.5 ml)
Cycle 8 Day1	22 ml(3 tubes of 7.5 ml)

Figure 1. A (H&E) and 1B (EpCAM). Primary tumor of a case with baseline CTC of 56.33 ± 7 .06. (H&E) and 1D (EpCAM). Metastatic tumor of a different case with baseline CTC of 0.

Figure 2. The burden of metastatic disease in the liver does not correlate with CTC count as determined by the CellSearch method. Shown are CT scans of the abdomen from 2 patients (A and B) with diffuse colorectal cancer with hepatic metastases and high baseline CTC numbers. CT scans were performed before initiation of chemotherapy (baseline). The scans in panel C provide illustrative examples of the observed inter-patient heterogeneity from 4 patients. The numbers in black depict the ratio of total liver volume: volume of disease free liver, and the numbers in red depict the mean CTC count from replicates +/− standard error of the mean.

Table 3A. Comparison of incidence of liver and lung metastases between groups with low baseline CTC count (A) at 0–2 cells/7.5ml of blood and high baseline counts (B) of 3 or more CTCs in 7.5ml of blood (p = 0.39, Fisher's exact test).

Group	Liver metastases only	Lung metastases only	Both liver and lung metastases	None(Metastases to sites other than lung or liver)
A (n=17)	5	5	3	4
B (n=7)	3	0	3	1

Table 3B. Comparison of CEA levels between patient groups with low and high CTC counts, respectively (p=0 .018, Fisher's exact test).

Group	CEA<2.5	CEA=2.5–5	CEA=5−50	CEA=50−200	CEA>200
A (n=17)	4	6	3	3	1
B (n=7)	0	1	0	1	5

Table 3C. Comparison of tumor mutational status between patient groups with low and high CTC counts, respectively (p = 0.67, Fisher's exact test).

Group	Wild-type KRAS	Mutant KRAS	Wild-type BRAF	Mutant BRAF (V600E)
A (n=17)	14	3	14	3
B (n=7)	6	0	6	0

Table 3D. Comparison of time interval between the diagnosis of the primary tumor and the appearance of metastases between lowland high baseline CTC count groups (p = 0.35, Fisher's exact test).

Group	Zero	<6 months	6 months −2years	>2years
A (n=17)	8	5	2	4
B (n=7)	6	1	0	0

Table 4A. Change in CTC counts at first assessment after start of chemotherapy as compared to the counts at baseline assessment prior to treatment.

Total number of patients	Decrease	Increase	Constant
21	11	3	7

Table 4B. Change in CTC counts at end-of-chemotherapy assessment as compared to the counts at baseline assessment prior to treatment.

Total number of patients	Decrease	Increase	Constant
10	5	1	4

Table 4C. Change in CTC counts at end-of-chemotherapy assessment compared to the counts at baseline assessment prior to treatment in the patients with high CTC counts of 3 or more cells in 7.5 ml of blood at baseline.

Total number of patients	Decrease to 0–2 cells/7.5ml	Decrease to 3 or more cells/7.5ml	Increase
7	4	2	1