Prognostic Value of Liver Cancer CTCs Isolated by a Novel Microfluidic Platform

February 7, 2022 updated by: Zhihang Zhou, The Second Affiliated Hospital of Chongqing Medical University

The Prognostic Value of Circulating Tumor Cells Isolated by a Novel Microfluidic Platform in Liver Cancer Patients

This study aims to isolate CTCs in peripheral venous blood of liver cancer patients by inertial focusing principle-based microfluidic device, determine the relationship between the number of CTCs and patient prognosis and treatment response, detect mutation, copy number variation and mutation load in CTC cells and corresponding tissues using single-cell whole genome sequencing technology, and use bioinformatics analysis of CTC heterogeneity and its relationship with clinical outcome. In addition, the culture of CTCs in vitro was explored by organoid culture or sphere culture in order to obtain CTCs cell lines to reveal the metastatic mechanisms of HCC. The partner of this project is Cellomics International Limited, which could provide Cellomics CTC-100 cell sorter and related consumables for this project. Peripheral venous blood from about 300 patients with initial liver cancer will be collected, and CTCs cells will be sorted in 8ml of each patient and typed according to protein expression. Clinical data, treatment effect and survival time of patients will be collected, and finally the relationship between the number of CTCs and subgroup with treatment response and patient prognosis will be analyzed. Uncovering the genomic characteristics of CTCs of HCC provides a new basis for the precise treatment of HCC. The new diagnostic markers for Hcancer were found by miRNA expression spectrum chip and metabolomic testing.In vitro culture methods and cellular characteristics of HCC circulating tumor cells were preliminarily explored.

Study Overview

Status

Recruiting

Conditions

Detailed Description

In China, the incidence of hepatocellular carcinoma (HCC) ranks the fourth, and the mortality rate is the second. Less than 20% of the patients were able to receive curative resection therapy. Most other patients with HCC have a very poor prognosis, with only chemical embolization, ablation treatment or systemic treatment, and the treatment effect is limited. Effective markers are required for the early diagnosis, treatment, prediction of prognosis, and assessment of treatment response in HCC. At present, alpha-fetoprotein (AFP) is the most common marker of HCC, which can be used for its diagnosis and patient prognosis. However, AFP has less sensitivity in the diagnosis of HCC. Although some other markers, such as AFP heteromer, abnormal prothrombin, also have a role in the diagnosis of HCC, they have not been widely used in clinical practice. Liver biopsy helps to directly evaluate the molecular biology of the tumor, but it has the risk of bleeding and causing tumor dissemination, which is currently rarely used in the diagnosis of HCC. Furthermore, liver puncture biopsy due to tumor heterogeneity may not adequately reflect the overall tumor characteristic.

Liquid biopsy can be used in various aspects of tumor diagnosis, detection and treatment, including circulating tumor cells, circulating tumor DNA, circulating miRNA, metabolites, etc. Circulating tumor cells (circulating tumor cells, CTCs) from the tumor group into the circulating system of tumor cells, is the key factor of tumor metastasis. Most CTCs can die from anoikisis, immune attack, or shear stress. To adapt to the environment, CTCs undergo a range of changes, such as epithelial-mesenchymal transition (EMT). Tumor cells undergo EMT and lose epithelial cell markers and acquire stromal cell features. Thus, CTCs are a group of cells derived from different regions of tumor tissue that are heterogeneous and play a critical role in tumor progression. CTCs are not only minimally invasive, but also can reflect the heterogeneity of tumor tissue. The CTCs convey the genomic information from the primary tumor tissue, which can indirectly reflect the information about the gene mutation, the copy number variation, and the tumor mutation load in the tumor tissue through the whole-genome sequencing of the CTCs, helping the clinicians to develop the treatment plan. For example, preliminary studies have shown that copy number variation in CTCs has a high accordance with primary HCC tissues.

CTCs have been extensively studied in multiple solid tumors. For example, in patients with metastatic prostate cancer, the number of CTCs can accurately predict the prognosis of the patients and reflect the of the treatment. Examination HER2-positive CTCs cells in breast cancer patients helps to specify the designated patient's treatment regimen. However, in patients with HCC surgical resection, the number of CTC was associated with of vascular invasion, AFP level, tumor stage, tumor progression, and patient poor prognosis. Recently, it has been shown that postoperative adjuvant TACE is able to prolong the survival time of patients after HCC resection with preoperative EpCAM-positive CTC, with no significant effect on in CTC-negative patients. However, the number of CTCs was minimal, and the heterogeneity existed. There are many limitations on the detection of CTCs, and how to accurately detect CTCs is still a problem that bothers us. The presently used platform for isolating CTCs is based primarily on tumor cell markers, biophysical properties, or strategies without enrichment. Isolation based on tumor surface markers was sorted using specific antibodies: leukocytes were removed from leukocyte surface markers (such as CD45), that is, negative sorting; CTCs were separated from tumor cell surface markers (EpCAM, ASGPR, CK), i. e., positive sorting. The limitation of this approach is the fact that selection of markers fails to reflect tumor heterogeneity and reduces cellular activity. Separation methods based on the cell volume, elasticity, and conductance of CTCs, such as microfiltration, gradient centrifugation, and inertial focus, avoid the limitations of using surface markers for separation while having little effect on cell activity. Without enrichment methods, using high-speed fluorescence imaging, can distinguish CTCs directly in patients' blood.

The CTC-100 circulating tumor cell detection system used by Shenzhen Jingyi Medical Laboratory has been filed for the record of Shenzhen Class I medical devices (record No.: No.20190047) and has been on the market. The system is capable of efficiently capturing the enriched CTC. It mainly relies on the inertial focusing principle of the microfluidic chip and the elastic physical characteristics to realize the screening. This device makes the fluid laminar in the straight channel, the fluid near the channel wall is lowest, while the maximum fluid velocity in the middle of the channel. This flow velocity distribution produces a shear force gradient, and induces the resulting lift will push the particles to the wall. When it moves close to the wall of the channel, the lift induced by the channel wall again pushes the particles away from the channel wall. Both lift forces in opposite directions are called inertial lift, this force acts on the particles, moves it to the equilibrium position. Thus the particles focus on the stable site, form a focused flow. Cell particles migrate with liquid migration in microfluidic chip channels, due to the different size and elasticity of CTC and white blood cells, finally focusing on different equilibrium positions, thus achieving the screening effect. Screened tumor cells can identify their surface glycochemical characteristics and biological characteristics (EpCAM, PDL1, VEGF, etc.), and finally achieve high sensitivity and specific CTC screening and identification. Our preclinical trials showed that the number of CTC tested by this technique was significantly higher in liver cancer patients than in cirrhosis and healthy patients, and was positively associated with liver vascular invasion and tumor size (102 liver cancer, 43 cirrhosis and 10 healthy patients). The detection rate of this technique was 100% in HCC patients, and 64.7% of the cells detected in the CTC were EpCAM negative.Whole-genome sequencing found that the gene mutations in the CTC were consistent with the corresponding cancer tissue (5 CTC samples and the corresponding surgical resection tissue), suggesting that the CTCs can reflect information such as gene mutations in the cancer tissue.

This study aims to isolate CTCs in peripheral venous blood of liver cancer patients by inertial focusing principle-based microfluidic device, determine the relationship between the number of CTCs and patient prognosis and treatment response, detect mutation, copy number variation and mutation load in CTC cells and corresponding tissues using single-cell whole genome sequencing technology, and use bioinformatics analysis of CTC heterogeneity and its relationship with clinical outcome. In addition, the culture of CTCs in vitro was explored by organoid culture or sphere culture in order to obtain CTCs cell lines to reveal the metastatic mechanisms of HCC. The partner of this project is Cellomics International Limited, which could provide Cellomics CTC-100 cell sorter and related consumables for this project. Peripheral venous blood from about 300 patients with initial liver cancer will be collected, and CTCs cells will be sorted in 8ml of each patient and typed according to protein expression. Clinical data, treatment effect and survival time of patients will be collected, and finally the relationship between the number of CTCs and subgroup with treatment response and patient prognosis will be analyzed. Uncovering the genomic characteristics of CTCs of HCC provides a new basis for the precise treatment of HCC. The new diagnostic markers for Hcancer were found by miRNA expression spectrum chip and metabolomic testing.In vitro culture methods and cellular characteristics of HCC circulating tumor cells were preliminarily explored.

Study Type

Observational

Enrollment (Anticipated)

300

Contacts and Locations

This section provides the contact details for those conducting the study, and information on where this study is being conducted.

Study Contact

Study Locations

  • China
      • Chongqing, China
        • Recruiting
        • The Second Affiliated Hospital of Chongqing Medical University
        • Contact:

Participation Criteria

Researchers look for people who fit a certain description, called eligibility criteria. Some examples of these criteria are a person's general health condition or prior treatments.

Eligibility Criteria

Ages Eligible for Study

18 years to 70 years (Adult, Older Adult)

Accepts Healthy Volunteers

Yes

Genders Eligible for Study

All

Sampling Method

Non-Probability Sample

Study Population

Peripheral venous blood samples are expected to collect 300 liver cancer patients and 50 patients with liver cirrhosis, taking 8ml of venous blood from each patient. Among them, about 100 patients underwent surgery (about 5 * 5 * 5mm).After receiving follow-up for 1-2 years, information on patient treatment regimen, treatment response, recurrence, metastasis, survival time and other patients was collected.

Description

Inclusion Criteria:

  • confirmed by imaging and serological examination for benign liver cancer;
  • patients without any previous treatment;
  • informed consent;
  • age: 18 years or older (adult).

Exclusion Criteria:

  • with other malignancies;
  • metastatic liver tumors;
  • with major organic lesions;
  • with acute complications such as acute liver failure, massive gastrointestinal bleeding and so on.

Study Plan

This section provides details of the study plan, including how the study is designed and what the study is measuring.

How is the study designed?

Design Details

Cohorts and Interventions

Group / Cohort
HCC patients
HCC patients with initial diagnosis
Liver cirrhosis patients

What is the study measuring?

Primary Outcome Measures

Outcome Measure
Measure Description
Time Frame
Time to progression or death (months)
Time Frame: 2 years
Time to progression or death (months) after initial diagnosis will be recorded.
2 years

Collaborators and Investigators

This is where you will find people and organizations involved with this study.

Sponsor

The Second Affiliated Hospital of Chongqing Medical University

Collaborators

Cellomics International Limited

Investigators

  • Principal Investigator: ZHOU ZHIHANG, The Second Affiliated Hospital of Chongqing Medical University

Publications and helpful links

The person responsible for entering information about the study voluntarily provides these publications. These may be about anything related to the study.

General Publications

Study record dates

These dates track the progress of study record and summary results submissions to ClinicalTrials.gov. Study records and reported results are reviewed by the National Library of Medicine (NLM) to make sure they meet specific quality control standards before being posted on the public website.

Study Major Dates

Study Start (Actual)

December 1, 2021

Primary Completion (Anticipated)

January 31, 2024

Study Completion (Anticipated)

January 31, 2024

Study Registration Dates

First Submitted

January 24, 2022

First Submitted That Met QC Criteria

February 7, 2022

First Posted (Actual)

February 16, 2022

Study Record Updates

Last Update Posted (Actual)

February 16, 2022

Last Update Submitted That Met QC Criteria

February 7, 2022

Last Verified

February 1, 2022

More Information

Terms related to this study

Keywords

Additional Relevant MeSH Terms

Other Study ID Numbers

  • HCC-CTC

Plan for Individual participant data (IPD)

Plan to Share Individual Participant Data (IPD)?

YES

IPD Plan Description

IPD will be shared after the trial has been finished

IPD Sharing Time Frame

will be shared after the trial has been finished

IPD Sharing Supporting Information Type

  • STUDY_PROTOCOL
  • CSR

Drug and device information, study documents

Studies a U.S. FDA-regulated drug product

No

Studies a U.S. FDA-regulated device product

No

This information was retrieved directly from the website clinicaltrials.gov without any changes. If you have any requests to change, remove or update your study details, please contact register@clinicaltrials.gov. As soon as a change is implemented on clinicaltrials.gov, this will be updated automatically on our website as well.

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