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UW Medicine Researchers Identify Blood Cell Genetic Mutations That Can Disrupt Liquid Biopsy Results

The discovery is yet another factor that must be considered when developing a liquid biopsy test clinical laboratories can use to detect cancer

How often do disruptive elements present in Liquid biopsies result in misdiagnoses and unhelpful drug therapies for cancer? Researchers at the University of Washington School of Medicine (UW Medicine) in Seattle wanted to know. And the results of their study provide another useful insight for pathologists about the elements that circulate in human blood which must be understood so that liquid biopsy tests can be developed that are not affected by that factor.

Based on their case series study of 69 men with advanced prostate cancer, the UW Medicine researchers determined that 10% of men have a clonal hematopoiesis of indeterminate potential (CHIP) that can “interfere” with liquid biopsies and cause incorrect reports and unneeded prostate cancer treatment, according to their paper published in the journal JAMA Oncology.

The process of clonal hematopoiesis occurs when hematopoietic stem cells generate blood cells that mimic blood mutations in the same way as hematopoiesis, Labroots explained in “Potential Problems with Liquid Biopsies.” Hence, the word “clonal” in the description. 

The UW Medicine researchers advised testing for “variants in the cell-free DNA (cfDNA)” shed in blood plasma to enable appropriate treatment for people with already diagnosed prostate cancer, noted to a UW Medicine news release.

According to pathologist Colin Pritchard, MD, PhD, Associate Professor of Laboratory Medicine and Pathology at the UW Medicine, who led the research team, “clonal hematopoiesis can interfere with liquid biopsies. For example, mutations in the genes BRCA1, BRCA2, and ATM have been closely linked to cancer development.

“Unfortunately, these same genes are also commonly mutated as a result of clonal hematopoiesis,” he told Labroots. Pritchard is also Head of the Genetics Division of Laboratory Medicine at UW Medicine, Director of Clinical Diagnostics for the Brotman Baty Institute for Precision Medicine, and Co-Director of the Genetics and Solid Tumors Laboratory at the University of Washington Medical Center.

“The good news is that, by looking at the blood cellular compartment, you can tell with pretty good certainty whether something is cancer, or something is hematopoiesis,” he said in the news release.

What Does CHIP Interference Mean to a Clinical Laboratory Blood Test?

In their published study, the UW Medicine researchers stressed the “urgent need to understand cfDNA testing performance and sources of test interferences” in light of recent US Food and Drug Administration (FDA) clearance of two PARP inhibitors (PARPi) for prostate cancer:

“We found that a strikingly high proportion of DNA repair gene variants in the plasma of patients with advanced prostate cancer are attributable to CHIP,” the researchers wrote. “The CHIP variants were strongly correlated with increased age, and even higher than expected by age group.

“The high rate of CHIP may also be influenced by prior exposure to chemotherapy,” they added. “We are concerned that CHIP interference is causing false-positive cfDNA biomarker assessments that may result in patient harm from inappropriate treatment, and delays in delivering alternative effective treatment options.

“Without performing a whole-blood control, seven of 69 patients (10%) would have been misdiagnosed and incorrectly deemed eligible for PARP-inhibitor therapy based on CHIP interference in plasma. In fact, one patient in this series had a BRCA2 CHIP clone that had been previously reported by a commercial laboratory testing company with the recommendation to use a PARPi. To mitigate these risks, cfDNA results should be compared to results from whole-blood control or tumor tissue,” the researchers concluded.

To find the clinically relevant CHIP interference in prostate cancer cfDNA testing, researchers used the UW-OncoPlex assay (developed and clinically available at UW Medicine). The assay is a multiplexed next-generation sequencing panel aimed at detecting mutations in tumor tissues in more than 350 genes, according to the UW Medicine Laboratory and Pathology website. 

“To improve cfDNA assay performance, we developed an approach that simultaneously analyzes plasma and paired whole-blood control samples. Using this paired testing approach, we sought to determine to what degree CHIP interferes with the results of prostate cancer cfDNA testing,” the researchers wrote in JAMA Oncology

Men May Receive Unhelpful Prostate Cancer Drug Therapies

The research team studied test results from 69 men with advanced prostate cancer. They analyzed patients’ plasma cfDNA and whole-blood control samples.

Tumor sequencing enabled detection of germline (cells relating to preceding cells) variants from CHIP clones.

The UW Medicine study suggested CHIP variants “accounted for almost half of the somatic (non-germline) DNA repair mutations” detected by liquid biopsy, according to the news release.

Colin Pritchard, MD, PhD
“About half the time when the plasma is thought to contain a mutation that would guide therapy with these drugs, it actually contains CHIP variants, not prostate cancer DNA variants. That means that in about half of those tested, a patient could be told that he should be administered a drug that is not indicated to treat to his cancer,” said Colin Pritchard, MD, PhD, pathologist and Associate Professor of Laboratory Medicine and Pathology at UW Medicine in the new release. (Photo copyright: University of Washington School of Medicine.)

Other detailed findings of the UW Medicine Study:

  • CHIP variants of 2% or more were detected in cfDNA from 13 of 69 men.
  • Seven men, or 10%, having advanced prostate cancer “had CHIP variants in DNA repair genes used to determine PARPi candidacy.
  • CHIP variants rose with age: 0% in those 40 to 50; 12.5% in men 51 to 60; 6.3% in those 61 to 70; 20.8% in men 71 to 80; and 71% in men 81 to 90.
  • Whole-blood control made it possible to distinguish prostate cancer variants from CHIP interference variants.

“Men with prostate cancer are at high risk of being misdiagnosed as being eligible for PARPi therapy using current cfDNA tests; assays should use a whole-blood control sample to distinguish CHIP variants from prostate cancer,” the researchers wrote in JAMA Oncology.

Liquid Biopsies Are ‘Here to Stay’

Surgical oncologist William Cance, MD, Chief Medical and Scientific Officer, American Cancer Society (ACS) in Atlanta, recognizes the challenge of tumor biology to liquid biopsies. 

“Genetic abnormalities are only one piece of the puzzle. We need to look comprehensively at tumors for the best therapy, from their metabolic changes and protein signatures in the blood to the epigenetic modifications that may occur, as cancers take hold,” he told Oncology Times. “It’s not just shed DNA in the blood.”

The UW Medicine study demonstrates the importance of understanding how all elements in liquid biopsies interact to affect clinical laboratory test results.

“I think liquid biopsies are here to stay,” Cance told Oncology Times. “They’re all part of precision medicine, tailored to the individual.”

Donna Marie Pocius

Related Information:

Association of Clonal Hematopoiesis in DNA Repair Genes with Prostate Cancer Plasma Cell-free DNA Testing Interference

Potential Problems with Liquid Biopsies

Blood Cell Mutations Confound Prostate Cancer Liquid Biopsy

Pursing and Perfecting Use of Liquid Biopsies in Cancer Early Detection

Researchers at Harvard’s Massachusetts General Hospital Develop a Non-Invasive Liquid Biopsy Blood Test to Detect and Monitor Common Brain Tumors in Adults

Using Extracellular Vesicles, Researchers Highlight Viability of Liquid Biopsies for Cancer Biomarker Detection in Clinical Laboratories

New studies in UK and at Stanford University Show Lung Cancer Cells Circulating in Blood; Findings Could Make It Possible for Pathologists to Diagnose Cancer with ‘Liquid Biopsies’

Scientists Identify Growing Number of COVID-19 Variants, But Not All Clinical Laboratories Have the Capability to Test for Variants

Fear that immunity-resistant mutations of SARS-CoV-2 will emerge are real and the scientific community is paying close attention

Detection of an increasing number of new variants of the SARS-CoV-2 coronavirus raises the possibility that a new strain of COVID-19 might emerge that brings new problems to the management of the pandemic. Public health officials and clinical laboratory scientists are on the alert to determine if any new COVID-19 variant is more virulent or more easily transmissible.

Pathologists, along with the rest of the scientific community worldwide, are following reports of increasing coronavirus mutations with growing concern. The Alpha variant (Lineage B.1.1.7) accounted for most of the COVID-19 cases in April of 2021 in the US, though it was first identified in the United Kingdom. That was followed by the Iota variant (Lineage B.1.526) first identified in New York City. A series of other variants were to follow. Scientists were not surprised. It is normal for viruses to mutate, so they logged and tracked the mutations.

Then, the Delta variant (Lineage B.1.617.2) emerged during a severe outbreak in India. At first, it did not seem more threatening than any other variant, but that changed very quickly. Delta was different.

“The speed with which it dominated the pandemic has left scientists nervous about what the virus will do next. The variant battles of 2021 are part of a longer war, one that is far from over,” The Washington Post reported, which added, “Today, [Delta] has nearly wiped out all of its rivals. The coronavirus pandemic in America has become a Delta pandemic. By the end of July, it accounted for 93.4% of new infections, according to the Centers for Disease Control and Prevention.”

Why is Delta the Worst COVID-19 Variant So Far?

The Delta variant has two advantages that scientists know about:

  • Stickier spike protein than the spike on the original SARS-CoV-2 coronavirus, as well as on the other, earlier variants. This means that the Delta variant stands a better chance of remaining in a person’s nose or throat long enough to reproduce.
  • Faster replication. When a virus mutation has more opportunity to reproduce, it quickly becomes the main viral strain. This is the case with the Delta variant. Experts say that the viral load in patients with Delta is around 1,000 times higher than in patients with the original virus.
Colorized scanning electron micrograph of an apoptotic cell that is infected with the SARS-COV-2 virus

The image above is a “Colorized scanning electron micrograph of an apoptotic cell (tan) heavily infected with SARS-COV-2 virus particles (orange), isolated from a patient sample,” Newsweek reported. (Photo copyright: National Institute of Allergy and Infectious Diseases/Newsweek.)

Will More Dangerous SARS-CoV-2 Variants Appear?

“The great fear is that nature could spit out some new variant that completely saps the power of vaccines and upends the progress we’ve made against the pandemic. But to virologists and immunologists, such a possibility seems very unlikely,” STAT reported.

That is because, unlike Influenza, which is also a coronavirus, SARS-CoV-2 variants are not able to share genetic materials and recombine into deadlier variants. Thus, scientists are skeptical that a variant could appear and wipe out the progress made with vaccines and treatments.

One of the reasons the Flu vaccine changes every year is Influenza’s ability to recombine into variants that can evade immunity. Therefore, scientists are beginning to suspect that SARS-CoV-2, like the Flu, will likely be around for a while.

“I don’t think eradication is on the table. But I think we could come up with something that’s better than what we have for the flu,” Sharone Green, MD, Associate Professor of Medicine, Division of Infectious Diseases and Immunology and Infection Control Officer at University of Massachusetts Medical School, told Newsweek.

Limiting Infections and Replication

Several factors combined to create the COVID-19 pandemic. But SARS-CoV-2 was a novel coronavirus, meaning it was a new pathogen of a known virus. This meant every person on the planet was a potential host.

The situation now is different. Thanks to natural immunity, vaccines, and treatments that shorten the infection, the SARS-CoV-2 coronavirus has less chance to replicate.

“The pressure is there, but the opportunity is not. The virus has to replicate in order to mutate, but each virus doesn’t get many lottery tickets in a vaccinated person who’s infected,” Jeremy Kamil, PhD, Associate Professor of Microbiology and Immunology at LSU Health in Shreveport, La., told STAT.

Tracking Variants of Interest and Variants of Concern

The World Health Organization (WHO) has been monitoring the viral evolution of SARS-CoV-2 since the beginning of the pandemic. In late 2020, the WHO created categories for tracking variants:

The WHO’s lists of VOIs and VOCs help inform the global response to the COVID-19 pandemic.

According to the CDC’s SARS-CoV-2 Variant Classifications and Definitions:

VOIs are “A variant with specific genetic markers that have been associated with changes to receptor binding, reduced neutralization by antibodies generated against previous infection or vaccination, reduced efficacy of treatments, potential diagnostic impact, or predicted increase in transmissibility or disease severity.”

Current VOIs include:

  • Eta (Lineage B.1.525), detected in multiple countries, designated a VOI in March 2021.
  • Iota (Lineage B.1.526), US, first detected in November 2020, designated a VOI in March 2021.
  • Kappa (lineage B.1.617.1), India, first detected in October 2020, designated a VOI in April 2021.
  • Lambda (lineage C.37), Peru, first detected in December 2020, designated a VOI in June 2021.

VOCs, on the other hand, demonstrate all the characteristics of VOIs and also demonstrate “an increase in transmissibility, more severe disease (e.g., increased hospitalizations or deaths), significant reduction in neutralization by antibodies generated during previous infection or vaccination, reduced effectiveness of treatments or vaccines, or diagnostic detection failures.”

Current VOCs include:

  • Alpha (lineage B.1.1.7), first detected in the UK, September 2020.
  • Beta (lineage B.1.351), first detected in South Africa, May 2020.
  • Gamma (lineage P.1), first detected in Brazil, November 2020.
  • Delta (lineage B.1.617.2), first detected in India, October 2020.

Will Vaccines Stop Working?

With each new variant, there tends to be a flurry of media attention and fearmongering. That a variant could emerge which would render our current vaccines ineffective has the scientific community’s attention.

“There is intense interest in whether mutations in the spike glycoprotein mediate escape from host antibodies and could potentially compromise vaccine effectiveness, since spike is the major viral antigen in the current vaccines,” wrote Adam S. Lauring, MD, PhD, and Emma B. Hodcroft, PhD, in “Genetic Variants of SARS-CoV-2­—What Do They Mean?” published in the Journal of the American Medical Association (JAMA). 

“Because current vaccines provoke an immune response to the entire spike protein, it is hoped that effective protection may still occur despite a few changes at antigenic sites in SARS-CoV-2 variants,” they added.

Future events may justify the optimism that the ongoing effectiveness of vaccines will help with many COVID-19 variants. But pathologists and clinical laboratory leaders may want to be vigilant, because as infection rates increase, so do workloads and demands on critical resources in their medical laboratories.

Dava Stewart

Related Information

‘Goldilocks Virus’: Delta Vanquishes All Variant Rivals as Scientists Race to Understand Its Tricks

Viral Evolution 101: Why the Coronavirus Has Changed as It Has, and What It Means Going Forward

A Doomsday COVID Variant Worse than Delta and Lambda May Be Coming, Scientists Say

Tracking SARS-CoV-2 Variants

Genetic Variants of SARS-CoV-2—What Do They Mean?