Studying Circulating Tumor Cells at the Protein Level

Abstract and Introduction

Abstract

Metastasis is the main cause of cancer death. As the tumor progresses, cells from the primary tumor site are shed into the bloodstream as circulating tumor cells (CTCs). Eventually, these cells colonize other organs and form distant metastases. It is therefore imperative that we gain a better understanding of the biological characteristics of CTCs for development of novel treatment modalities to minimize metastasis-associated cancer deaths. In recent years, rapid developments in technologies for the study of CTCs have taken place. We now have a variety of tools for the isolation and examination of CTCs which were not available before. This review introduces some commonly used protein markers in CTC investigations and summarizes a few advanced technologies which have been successfully applied for studying CTC biology at the protein level.

Introduction

Cancer is a leading cause of death worldwide where mortalities are mainly caused by occurrence of distant metastases through hematogenous spread.^[1,2] There is an urgent need for development of novel biomarkers to allow earlier diagnosis, better disease monitoring, improved treatment efficacy and personalization of therapeutic regimens. Moreover, a more detailed understanding of the intermediate steps between primary tumor formation and development of metastasis is critical to the development of treatment strategies for reduction of cancer-associated deaths.

During metastasis, tumor cells are shed off from the primary tumor site and enter the bloodstream where they travel to distant organs and form secondary tumors.^[3–5] In recent years, these circulating tumor cells (CTC) have become the topic of intense research which aimed to investigate their value in prognosis, disease monitoring and treatment response in cancer patients.^[6–11] Numerous reports have now shown that CTCs provide real-time data about the patients' tumor characteristics such as their expression profile and mutation status as well as other clinically important information such as whether a patient is suitable for a certain treatment (e.g., trastuzumab).^[8,12,13] Thus, it has been suggested that CTCs may serve a 'liquid biopsy' for examination of tumor characteristics in place of tissue samples.^[14] In this sense, blood is an ideal source for both patients and clinicians as a blood draw is routinely performed, simple, minimally invasive and cost effective.

A difficulty which arises in the detection of CTC from blood is their rarity, with cell numbers as low as having only 1 CTC per ml of blood, even in patients with metastatic carcinoma.^[15,16] In this respect, immense technological achievements in the recent years have accelerated research into the clinical utility of CTCs. In this review, we discuss some of the methods commonly used for isolation and detection of CTCs with focus on methods utilizing protein markers. We first list some of the most frequently used protein markers for such application (Table 1) and then introduce the readers to some technologies which have been successfully used for CTC investigations.

Table 1. Protein markers commonly used for circulating tumor cells investigations.
Target protein	Examples of cell/tissue types which express this protein	Ref.
EpCAM (also called HEA or BerEP4)	Epithelial cells such as breast, lung and colon	[17–19]
Cytokeratins (e.g., CK8, CK18, CK19)	Epithelial cells such as breast, lung and colon	[27–29]
Mucin 1	Colon, lung, breast, ovarian, pancreatic, prostate, kidney, stomach, head and neck	[37–40]
HER2	Breast	[5,46–55]
CD45	Hematopoietic cells	[56–61]
CD133	Stem cells from hematopoietic cells, brain, prostate liver, pancreas, lung colon and ovary	[62–73]
Chemokine receptors (e.g., CXCR4, CCR6, CCR7, CCR9)	Stomach, lung, kidney, colon, breast	[86–91]
Adhesion molecules (e.g., VCAM-1, ICAM-1)	Endothelial cells, skin, pancreas, breast	[93–99]