NEW YORK – A team led by researchers at mass spectrometry firm Protypia and drugmaker Eli Lilly has demonstrated the ability of targeted mass spec to quantify immune checkpoint inhibitor targets in patient tissue.
Detailed in a study published this week in Scientific Reports, the work showed that mass spectrometry can provide a more accurate assessment of the expression of checkpoint proteins like PD-L1 than conventional clinical approaches like immunohistochemistry.
The findings further confirm the widely held view that existing tests for selecting cancer patients for treatment with checkpoint inhibitors have poor accuracy, selecting many patients who are unlikely to benefit while missing many who could profit from the drugs.
They also suggest that technologies like mass spectrometry could improve the patient selection process, said Daniel Liebler, president, CEO, and founder of Nashville, Tennessee-based Protypia, though he noted that broad clinical adoption of such an approach is likely still a somewhat distant prospect.
The work follows a 2017 study Liebler led using mass spec to analyze PD-L1 and PD-L2 expression in human melanoma samples.
Liebler was formerly a professor of biochemistry at Vanderbilt's School of Medicine and a participant in major cancer proteomic initiatives, including the National Cancer Institute's Clinical Proteomic Tumor Analysis Consortium (CPTAC). He left his position and launched Protypia in late 2016 with the aim of more directly applying mass spectrometry and proteomic technologies to cancer drug development.
"I felt like the real action was in therapeutic development, particularly in immune-oncology," he said. "And I could see that the field was kind of limping along with really bad biomarkers."
Given his mass spec background, the technology seemed to Liebler a natural fit for measuring key immuno-oncology biomarkers like PD-L1.
Currently, IHC is the dominant technology for measuring proteins in pathology, including immunotherapy targets. However, Liebler noted, the technology was developed as a qualitative tool, and while efforts have been undertaken to make its measurements more quantitative, these have met with limited success.
In an IHC assay, antibodies are used to stain the proteins of interest. When quantifying proteins using IHC, a cutoff for the intensity of the stain is established with cells with an intensity above the cutoff deemed positive for the target protein and cells below the cutoff deemed negative. The percentage of positive cells is then used to make diagnostic or treatment decisions.
In the case of checkpoint inhibitors, 50 percent positivity is a common threshold for determining who should be treated, though many patients above that cutoff don't respond to the drugs while some below the cutoff do, Liebler noted.
In their new paper, he and his co-authors compared IHC scores with mass spec-based quantitation of PD-L1 levels in non-small cell lung cancer tissues, finding that while the two measurements were correlated, the IHC scores masked significant heterogeneity in PD-L1 expression.
For instance, tissues that according to 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.
"IHC both overestimates and underestimates the true PD-L1 content," Liebler said. "IHC is great in that it lets you look at the tissue and get a sense of which cells express the target, but it's not a reliable quantitative technique."
"If you want to quantify these immune checkpoint drug targets, you need something better than IHC," he said, noting that previous datasets have demonstrated similar results.
Liebler said he also sees a role for mass spec in measuring cancer drug targets for which there are no good antibodies.
He cited the example of the enzyme IDO1, which is involved in T cell regulation and which pharma firm Incyte sought to target in combination with Merck's PD-L1 inhibitor Keytruda to treat melanoma.
"They had promising preclinical data as well as phase I and phase II data," he said. "And they had a phase II trial that combined the Incyte IDO1 inhibitor, called epacadostat, with the Merck PD-L1 inhibitor Keytruda, and it bombed. Nobody who got the combination did any better than the people who just got the Keytruda alone."
Liebler said that in the original MCP study from his former group at Vanderbilt, they showed that only around a third of the melanoma samples they analyzed had IDO1 expression, as measured by mass spec.
He added that in the Scientific Reports study, he and his coauthors similarly found that only a small percentage of NSCLC patients had IDO1 expression and only about 15 percent of patients expressed significant levels of both PD-L1 and IDO1.
"If you're going to use a combo therapy or select patients for a trial, you should select patients based on the knowledge that they have the target," he said. "With Incyte, the problem was that in the trial that failed, there was no IHC assay for IDO1. There was no antibody. And this has been kind of a problem with IDO1. So in that trial, they took all comers with respect to IDO1, because they had no way to gauge who expressed the target."
"People talk about precision medicine a lot, but it has often been treated as kind of a digital phenomenon," he said. "It's 'yes or no' for a patient. They have this mutation, they get this drug. They have that mutation, they get that drug. But when you are drugging targets that vary in abundance, you need a quantitative platform to assess the targets."
Liebler said that he believed mass spec was well suited to that sort of quantitative measurement, but he acknowledged that from a usability standpoint, and a lab and vendor business perspective, widespread use of clinical mass spec assays for guiding immunotherapy treatment decisions remained a ways off.
He said that his goal with Protypia was to drive this process within the pharmaceutical companies themselves, the idea being that this was more likely to be a successful route than moving directly to trying to commercialize diagnostics.
"Working with pharmaceutical companies, we can show them [that mass spec] can measure what they can't and what we can agree is very important to measure, the drug targets," he said. "Furthermore, we can go to archival paraffin sections from completed trials where you have given the patients the drugs, you have the outcome information, and you've got questions. We can analyze what the abundance of the targets or other key coregulators was… and retrospectively test hypotheses about what might have accounted for surprising or disappointing results in clinical trials."
Liebler called the work with Lilly "an intermediate proof-of-concept" study, and added that the company is involved with Lilly and several other drugmakers on other projects in cancer immunotherapy and autoimmune disease.