Monitoring of Circulating Tumor Cells by a Combination of Immunomagnetic Enrichment and RT-PCR in Colorectal Cancer Patients Undergoing Surgery

Abstract

Background. The presence of circulating tumor cells (CTC) has been reported in patients with advanced colorectal cancer. Monitoring CTC (also known as a liquid-biopsy) has recently become the center of interest for low-invasive monitoring of cancer progression and predictive biomarkers testing. Along with high-cost technology and a complex methodology, a straightforward method based on magnetic beads enrichment followed by RT-PCR is set to allow for routine CTC analysis in colorectal cancer patients. Objectives. The main purpose of this study was to evaluate the possibility of CTC detection in routine monitoring of patients starting before and continuing after surgery. Material and Methods. The investigated group consisted of 30 patients mainly in advanced stages of colorectal cancer. In all patients, CTC detection was performed prior to surgery, in a subset of 14 patients additional sampling was done during and after surgery. In all cases, peripheral blood was processed using AdnaTest ColonCancer kit, which relies on enriching CTCs using EpCAM-functionalized magnetic beads and subsequently identifying tumorspecific CEA, EGFR and GA733-2 mRNA transcripts. Results. Out of all the tested samples, CTC were found in one patient suffering from advanced disease with lung and liver metastases. There, however, the positive finding was confirmed in 3 consecutive samples acquired before, during and shortly after palliative R2 resection. Conclusions. The presence of CTC may be used to observe post-operative disease development. Due to the overall low CTC detection, further technology development may be necessary before its universal applicability to manage colorectal cancer patients (Adv Clin Exp Med 2016, 25, 6, 1273-1279).

Circulating tumor cell-related transcripts in blood as prognostic biomarkers of early recurrence after liver resection for colorectal metastases

“…In fact, only one out of three patients was positive for CTCs: this stage 4 CRC patient was positive for EGFR mRNA only in the first of five samples (before surgical treatment), while the remaining ones (during the surgery, 1 and 7 days after surgery, and finally 3 months later), were positive only for CEA mRNA. Similar results were also obtained by Pesta et al 30 These findings completely overlap with those obtained in our study; therefore, we can agree with the conclusions of Vojtechova et al 29 who stated that as there are numerous reports in the literature showing that the presence of CTC in colorectal patients (especially in advanced metastatic disease stages) it can hardly be questioned. 8,29 In fact, the successful CTC recovery naturally relies on: (a) proper timing and methodology applied during the sample acquisition; (b) leukocyte contamination whose genetic material could saturate the RT-PCR reaction, thus hampering detection of EGFR, CEA, and GA733-2 transcripts originating from CTCs.…”

“…Similar results were also obtained by Pesta et al 30 These findings completely overlap with those obtained in our study; therefore, we can agree with the conclusions of Vojtechova et al 29 who stated that as there are numerous reports in the literature showing that the presence of CTC in colorectal patients (especially in advanced metastatic disease stages) it can hardly be questioned. 8,29 In fact, the successful CTC recovery naturally relies on: (a) proper timing and methodology applied during the sample acquisition; (b) leukocyte contamination whose genetic material could saturate the RT-PCR reaction, thus hampering detection of EGFR, CEA, and GA733-2 transcripts originating from CTCs. 29 Interestingly, 52% of patients developed recurrence within 12 months after surgery.…”

“…Therefore, although it would be conceivable that at least one-third of metastatic patients might also show increased levels of circulating RNA, in our setting, both the three blood sources used (peripheral vein, CVC, and portal vein) at the T0 and the following T1 and T2 peripheral blood samples did not show any difference in the levels of EGFR. These results are not so surprising if we consider data recently reported by Vojtechova et al, 29 where they were not able to enrich CTCs from the blood of stage 3 and 4 CRC patients. In fact, only one out of three patients was positive for CTCs: this stage 4 CRC patient was positive for EGFR mRNA only in the first of five samples (before surgical treatment), while the remaining ones (during the surgery, 1 and 7 days after surgery, and finally 3 months later), were positive only for CEA mRNA.…”

Predicting Outcome and Therapy Response in mCRC Patients Using an Indirect Method for CTCs Detection by a Multigene Expression Panel: A Multicentric Prospective Validation Study

“…Also, the quantification of CTCs combining two methodologies, CellSearch and AdnaTest, demonstrated an improved detection of CTCs in a 47 mCRC patient cohort [22]. The requirement for improved sensitivity in CTCs detection has also been identified for the management of patients undergoing surgery [23]. We must also consider that the CTCs cohort in mCRC patients may be composed not only of epithelial tumor cells, but also tumor cells that have undergone transformation due to the epithelial-mesenchymal transition, and tumor stem cells [24].…”

A Direct Comparison between the Lateral Magnetophoretic Microseparator and AdnaTest for Isolating Prostate Circulating Tumor Cells

“…Among the macroscale techniques, the AdnaTest (Qiagen) is a representative method for detecting cancer-associated genes in CTCs isolated from various types of cancer, including breast [ 19 ], colon [ 20 ], ovarian [ 21 ], and prostate [ 22 ] cancer. Its ease of use and high sensitivity means that the AdnaTest is widely used in detecting gene expression profiling in CTCs and identifying potential prognostic and diagnostic biomarkers for cancer.…”