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Fred Hutch Establishes CLIA Lab Focused on Targeted Mass Spec Protein Assays

NEW YORK – Researchers at the Fred Hutchinson Cancer Research Center have established a CLIA laboratory focused on running targeted proteomic assays by multiple reaction monitoring mass spectrometry, making it one of the few labs anywhere set up specifically to use the method for clinical purposes.

Led by Fred Hutch professor Amanda Paulovich, the lab plans to launch an MRM mass spec assay for HER2 as its first CLIA-certified offering with a test for the immuno-oncology marker PD-L1 likely to follow.

While MRM mass spec is commonly used for targeted protein quantitation in the research space, it is still rarely used clinically, which makes the launch of the Fred Hutch lab a notable development for clinical proteomics. Paulovich is a leader in the use of MRM mass spec for targeted protein quantification and has developed a number of MRM assays and assay panels focused on different aspects of cancer biology. Much of this work is done under the auspices of National Cancer Institute's Clinical Proteomic Tumor Analysis Consortium (CPTAC), which aims to move proteomic technologies into the clinic.

Proponents of mass spec have long highlighted the technology's ability to provide more specific, quantitative protein measurements than immunoassays, as well as its potential to multiplex large numbers of proteins in a single assay. However, mass spec is still infrequently used in the clinic for protein assays due in part to the technique's complexity compared to immunoassays along with other concerns such as its typically lower sensitivity.

National reference labs like Quest Diagnostics and Laboratory Corporation of America have offered mass spec-based tests for a few proteins, most commonly for insulin-like growth factor 1 and thyroglobulin, for nearly a decade, but they have been slow to expand their mass spec-based protein assay offerings. A few diagnostics firms, most notably Integrated Diagnostics and Sera Prognostics, have also brought tests to market using mass spec for targeted protein measurements, but it by and large it remains a niche technology as far as clinical use is concerned.

Paulovich acknowledged these challenges, but said she believed it was still relatively early in the process of pushing mass spec-based proteomics into the clinic.

She noted that she set up her Fred Hutch lab focused on translational proteomics 17 years ago, and has since been working with clinical chemists and others to devise more effective approaches to measure proteins for improving cancer treatment.

"Even though it has been a long time… I don't think it's that long in the life cycle of getting new technologies taken up in the clinical space," she said. "I think the clinical space is very slow and conservative in taking on new things, since the stakes are high in making patient care decisions. It really just takes evangelism and commitment and clinical need."

That latter factor, Paulovich noted, informed her group's decision to launch HER2 as its first assay. She said that some recently developed antibody-drug conjugates targeting HER2 in breast cancer patients appear to have clinical activity even in patients who would generally be considered HER2-negative according to conventional immunohistochemistry assays.

"Our assays are able to quantify the HER2 protein in those low expressing patients," she said. "And we're going to be looking to see if we can add value in terms of selecting patients for therapies like that."

The lab is also working on a mass spec assay for PD-L1, an area where Paulovich noted IHC assays have struggled with accuracy and harmonization, despite being widely used.

Last month, for instance, research from mass spectrometry firm Protypia and drugmaker Eli Lilly, demonstrated that IHC testing masks significant heterogeneity in PD-L1 expression.

For instance, tissues that based on mass spec analysis had PD-L1 concentrations of around .2 femtomoles per microgram ranged, according to IHC, from nearly 0 percent positive to nearly 100 percent positive for the protein.

There were also cells deemed 100 percent positive for PD-L1 by IHC across the full ranges of expression levels measured by mass spectrometry, indicating that existing clinical IHC assays both underestimate and overestimate patient PD-L1 content.

Paulovich noted that given the work involved in CLIA-certifying assays, the lab would move deliberately in adding new tests to its menu.

"When we run our CLIA assay, we have a pathologist signing out the results, and a patient treatment decision can be based on those results," she said. "We won’t CLIA-certify an assay unless a clinical decision might be based on it. So, that's really what determines when an assay gets brought up to the full CLIA" certification.

The lab also offers hundreds of MRM mass spec targeted protein assays that, while not themselves CLIA certified, are run within the lab's CLIA environment and can be used for clinical research purposes.

Paulovich said that she and her colleagues plan in the near future to publish a series of papers describing more than 300 MRM assays to proteins involved in immune-modulation or inflammation that they have validated through work within both CPTAC and the Applied Proteogenomics Organizational Learning and Outcomes (APOLLO) network, a partnership between the National Cancer Institute, the US Department of Defense, and the US Department of Veterans Affairs. Both are national efforts to accelerate the understanding of the molecular basis of cancer through the application of large-scale proteome and genome analysis, or proteogenomics.

She said another area where she thought their assays could be useful is in more rapidly screening patients for inclusion in clinical trials for targeted cancer therapies.

"If you look at precision oncology trials like the NCI-MATCH trial, they have tens of antibody-drug conjugates (ADC) they would like to open up arms for and triage patients into those arms based on which of the ADC protein targets are expressed on the surface of their tumor," she said. "If you have 30 ADC targets – and in some tumor types only 5 percent of patients might express it, but those patients will benefit greatly – you want to pick that up."

Testing hundreds to thousands of patients for tens of different protein targets isn't feasible using IHC, but it could be using targeted, multiplexed mass spec, Paulovich said. "There's a bottleneck there in patient screening that we are wondering if we could help relieve."

She noted that this interest in better selecting patients for cancer treatments has been the main driver of her lab's research since its founding.

"The entire past 17 years has been about optimizing and validating this platform in the preclinical space, and now in the most recent phase of our work, we have built a catalogue of assays that are off the shelf and allow us to do a robust quantification of cancer-related proteins," she said. "Now the ultimate goal and next step is to move those into a space where they become clinically useful."