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Potdar et al. Circulating tumor cells in liquid biopsies

INTRODUCTION Chemotherapy is a standard mode of treatment for
all cancers. CTC levels are determined before and
Cancer cells are invasive and can metastasize; once after rounds of chemotherapy. In the majority of cases
metastasis has occurred, disease is usually incurable. of non-metastatic breast cancer, reduction in the
Recent technological developments have enabled number of CTCs is observed after the first round of
evaluation of the metastatic potential of cells using chemotherapy; however, CTCs can be resistant to
“liquid biopsies”, [1,2] noninvasive tests for circulating therapy, leading to administration of increasing doses
tumor cells (CTCs), which are present even at early in subsequent rounds of chemotherapy. In one study,
stages of disease. Additionally, molecular profiling patients with > 80 CTCs in 7.5 mL of blood died within
[3]
and enumeration of CTCs can predict their metastatic one year of diagnosis due to metastases. In another
[16]
potential, thereby facilitating improved treatment and study of patients with metastatic breast cancer, when
[3]
prognosis. Such testing has potential to revolutionize CTCs continued to be detected after the first round
personalized cancer medicine by avoiding biopsies, of chemotherapy, there was rapid progression of
since it captures cancer cells that tumors shed into the disease; it was suggested that, in such cases, it
the circulation, rather than sampling the tumor itself. would be preferable to opt for alternative treatment,
Liquid biopsies permit repeated, noninvasive sample rather than continuing with the same chemotherapy.
[17]
collection from cancer patients that can be profiled for A review of CTCs in patients with breast cancer in
gene expression, allowing clinicians to choose specific 2013 considered the clinical application of CTCs in
therapies targeting particular mutations. Ultimately, breast cancer therapy and assessed the use of HER2
liquid biopsies could become a routine cancer treatment in HER2-negative breast cancer patients,
screening method, alongside current methods, such as selected on the basis of CTC detection. [13,14] Rack et al.
[18]
mammograms and colonoscopies. [4,5] Liquid biopsies concluded that there is an independent prognostic
could be useful in the majority of invasive solid tumors, relevance of CTC determination both before and after
including breast, colon, lung, prostate, and pancreatic adjuvant chemotherapy in a study of > 2,000 patients
cancers. with primary breast cancer. Bidard et al. also studied
[13]
the clinical utility of CTCs in metastatic breast cancer,
Isolation and enumeration of CTCs is difficult because mainly focusing on first and second line treatments.
of their low numbers, size, and heterogeneity.
Several methods have been established for isolation Completion of the human genome project and
and enumeration of CTCs from liquid biopsies, the advent of molecular profiling has led to an
including the Ficoll-Hypaque gradient method, understanding of the genetic profiles of cancer cells,
[6]
®
[9]
[19]
flow cytometry, [7,8] the CELLSEARCH system, including CTCs. MMP1, MMP2, MMP9, VEGFA,
isolation by size of epithelial tumor cells (ISET), [10,11] MT-CO2, ICAM1, CD44, and PROM1 are major genes
and microfluidic systems. The CELLSEARCH involved in the processes of invasion and metastasis.
[12]
®
system is primarily based on selection of EpCAM- and Moreover, cadherin-1, N-cadherin, fibronectin, and
cytokeratin-positive CTCs, and has been approved integrin β are adhesion molecules involved in the
by the Food and Drug Administration (FDA) and used metastatic process. During invasion and metastasis,
in several clinical studies; [13,14] however, one study CTCs frequently undergo the process of EMT,
reported that CTCs which have undergone epithelial which involves several genes including TWIST1 and
mesenchymal transition (EMT) exhibit down-regulation TWIST2, in combination with TGFB1, WNT, and
[20]
of epithelial markers, including EpCAM and keratin, NOTCH2. The majority of CTCs isolated from breast
[15]
and such CTCs would be missed using this technology. cancer patients express EMT markers, including ETV5,
[21]
Similarly, in some cases, CTCs from HER2-positive NOTCH1, SNAIL, TGFB1, ZEB1, and ZEB2. Keratin
metastatic breast cancer patients include EpCAM- analysis is also a major molecular test in cancer, while
negative cells. Hence, EpCAM-independent methods CK19 and TP53 mutations are frequently found in
[15]
may enable improved detection of CTCs in certain CTCs from triple-negative breast cancer patients,
[22]
cancer patients. Advances in microfluidic technologies, and are a factor in the progression of the disease to
biomaterials, and molecular profiling have led to rapid the triple-negative stage. Patients with breast cancer
growth and interest in achieving liquid biopsies for expressing CK19, SCGB2A2, and ERBB2 showed
cancer diagnosis and treatment. CTCs are primarily poor survival rates. Riethdrof et al. demonstrated
[23]
[22]
characterized and identified by their morphology that HER2+ CTCs can be identified in HER2- breast
and immunostaining pattern with specific antibodies; cancer patients, leading to consideration of revision of
however, the heterogeneity of CTC’s is a major obstacle ongoing treatment, with trastuzumab-based therapy
to their isolation, identification, and characterization applied to patients with HER2+ CTCs and HER2-
from liquid biopsies. primary tumors, demonstrating that CTCs can assist in
Journal of Cancer Metastasis and Treatment ¦ Volume 3 ¦ January 23, 2017 7