Liquid biopsy: circulating tumor cells (CTCs), circulating tumor DNA (ctDNA) in lung and colon cancer
Ana Aza, PhD
Liquid biopsy is a non invasive, effective and fast technique which allows early tumor detection and control cancer progression, tumoral heterogeneity and treatment response. UltraSEEK Biochemistry has a detection limit of 0. 1% mutation frequency.
We are living in the era of personalized medicine that focus on the search of the right treatment for the right person by using molecular tests.
- Why do liquid biopsy testing?
- CTCs: circulating tumor cells
- Circulating tumor DNA (ctDNA)
- UltraSEEK Biochemistry by Agena has a detection limit of 0. 1%
- Liquid biopsy panels for lung and colon cancer
Why do liquid biopsy testing?
The traditional biopsy is subject to sampling bias. Sampling bias may be due to difficulties obtaining sample or the heterogeneity of the sample itself depending on the slide. Besides, the tumor molecular profile evolves fast in response to treatment and environment pressure. According to this, human blood contains a lot of information ‘easy’ to reach: circulating tumor cells (CTCs), cell-free tumor DNA (ctDNA) and RNA (ctRNA), proteins and different vesicles including exosomes.
The ability to detect these elements and to monitor disease evolution improving research and patient care attracted oncologists. In general terms, this is what we know as liquid biopsy: obtain information about disease without invasive methods. The term liquid biopsy encompasses not only blood but also other body fluids including urine, saliva, pleural effusions and cerebrospinal fluids (CSF).
Liquid biopsy main characteristics are:
Real time analysis: detect mutations in oncogenes and/or tumor suppressor genes, monitor disease progression, and identify secondary mutations which confer acquired resistance.
Viability: whenever tissue biopsy is not possible or repeated tissue biopsies are impractical.
Advance therapeutic research: test individuals more frequently and evaluate tumor and plasma samples in parallel.
CTCs: circulating tumor cells
Circulating tumor cells (CTCs) are cells which primarily belonged to the tumor but have been shed into bloodstream. It’s not easy to catch CTCs, detection methods go from negative selection, enrichment or positive selection by iron particles. Despite the abundance of CTCs is very low (around 1 cell per 1 x 106 blood cells), it has been demonstrated that the number of CTCs correlates with treatment outcome and overall survival1, so their molecular characterization and size also helps to prognosis.
Circulating tumor DNA (ctDNA)
Back in 1948 (it’s been a while!) Mandel and Metais detected nucleic acids in the bloodstream of different disease patients. Most of ctDNA is produced by apoptosis, a regular process in the body. However, necrosis happens frequently in tumor cells leading to different DNA size fragments. Whereas apoptosis leads to oligonucleosomal fragments (150-166bp) necrosis leads to longer and smaller fragments (300bp) than healthy cell free DNA . It is also described that ctDNA is present in almost all cancers, but its presence varies depending the type. ctDNA is more frequent in colorectal cancer and less in prostate.
Samples from Serum or plasma? Plasma better, for sure! It has been shown that serum has a higher cell free DNA from immune cells lysed during sample processing, thus, increasing background levels.
- Monitoring patient during treatment
- Identify genomic alterations within tumor
- Therapeutic resistance detection
- Detect early tumor evolution
- Characterize tumor heterogeneity and metastatic mutations
UltraSEEK by Agena has a detection limit of 0. 1%
The amount of ctDNA is usually limited, especially in early stages of cancer. In order to monitor the disease we need powerful techniques to detect few ctDNA %.
The current technologies can be divided in 2: Next-generation sequencing (NGS) and PCR approaches. Both are very useful but achieve different goals: NGS is limited by the length depth but it can detect new mutations, while PCR is better to detect known mutations. Generally speaking, PCR is more sensitive than NGS.
When it comes to detection limit , only allele specific PCR, BEAMing, digital PCR and MASS-ARRAY Ultraseek reach the limit of 1% to 0,1 %.
By using a combination of end-point PCR and mass spectrometry technology, UltraSEEK can detect in a single multiplex PCR reaction > 0.1% mutation frequency. This very remarkable detection burden means Agena is the only technique able to multiplex so many mutations with such sensitivity and so little time: from DNA to result in less than 8h. This can result in providing results in less than a week, and time is critical in cancer treatment.
Liquid biopsy panels for lung and colon cancer
As lung biopsies are hard to obtain, risky for patients and not always informative, liquid biopsy opens the door to provide relevant information for physicians just in the right turnaround time.
Many clinical trials compared cytology/biopsy vs. liquid biopsy in mutation analysis concluding that liquid biopsy can be used to determine molecular profiles and to monitor mutation profile in order to target therapies.
Agena has lauched UltraSEEK panels for lung and colon cancer with mutations of known significance included in the last EUROPEAN SOCIETY FOR MEDICAL ONCOLOGY (ESMO) guidelines:
– Lung cancer with 67 mutations in BRAF, EGRF, KRAS, ERBB2 and PIK3CA.
– Colon cancer panel with 107 mutations in BRAF, EGFR, KRAS, NRAS and PIK3CA.
As the interest in ctDNA is constantly increasing, the study of DNA methylation in determined genes as well as early detection screening in some cancers will boost this field to a new era in medicine.
Clinical studies have already been done for both panels, you can request information at firstname.lastname@example.org
If you have some questions about how this technology works, or need any help to set up your assay, our experienced Tech Support Team (all are PhD) will be happy to help you by mail (email@example.com), phone (+34.934464700) or we can visit you. Let’s talk!
More information about MassArray technology here:
Krebs, M. G. et al. Molecular analysis of circulating tumour cells — biology and biomarkers. Nature Reviews Clinical Oncology 11, 129–144 (2014).
Mandel and Metais. 1948
Siravegna et al. Integrating liquid biopsies into the management of cancer. Nature Reviews Clinical Oncology (2017) doi:10.1038/nrclinonc.2017.14. Published online 02 March 2017
Agena bioscience www.agenabioscience.com