Measuring versus reading - transforming the field of quantitative pathology

In this SelectScience interview, we speak with Dr. David Rimm, MD, PhD, Professor of Pathology at Yale University School of Medicine, to find out more about his groundbreaking assay that utilizes the latest cutting-edge multiplex spatial imaging technology to quantify HER2 biomarkers in breast cancer patients, with significantly greater accuracy than conventional tests. This new test can improve patient stratification and help determine which patients are candidates for treatment with antibody-drug conjugates (ADCs), ultimately enabling more breast cancer patients to access potentially life-saving drugs. Rimm discusses the technology behind his pioneering work, the value it has for clinical diagnosis and treatment decision-making and shares his thoughts about the future of quantitative pathology.

Identifying breast cancer biomarkers

Accurate diagnostics for cancer patients are vital to ensure a match between the correct therapies and the cancer phenotype, in order to effectively treat the condition while minimizing toxicity. This is particularly important when determining whether to treat with a biotherapeutic, such as antibody-drug conjugates (ADCs).

HER2-positive breast cancer is a type of breast cancer in which patients have high levels of human epidermal growth factor receptor 2 (HER2). Currently, to determine HER2-positive patients, pathologists carry out immunohistochemistry (IHC) diagnostic tests to identify HER2 biomarkers and score patients on a scale of 0-1+ (negative), 2+ (equivocal, requiring reflex testing via in situ hybridization assay, or ISH) or 3+ (positive). According to ASCO/CAP guidelines for making treatment decisions, a score of at least 2+ and a positive ISH result is required for patients to receive approval for HER2 targeted treatment, as most drugs are only effective when a patient expresses large amounts of the HER2 protein.

But recently, a new ADC called trastuzumab deruxtecan (marketed as Enhertu) has been approved which can effectively treat patients with low levels of HER2. In light of this new development, a new HER2-low category has been defined which covers patients with a score of 1+ and 2+ with a negative ISH score, meaning only patients with a 0 score would be considered HER2-negative and would not receive treatment. However, conventional IHC tests are not sufficiently sensitive in the low HER2 range and results can be ambiguous. Consequently, there is high variability and inaccurate assignment of IHC scores from pathologists, which means some patients may miss out on beneficial treatment or receive treatment that won’t be effective. This disparity highlights the need for a more sensitive, unbiased assay that is accurate in the low HER2 range so that HER2-low patients are more likely to be identified and approved for treatment with Enhertu.

Improving patient stratification with multiplexed immunofluorescent imaging

To tackle this issue, Dr. David Rimm and his team at Yale University set out to design an assay that was effective at identifying HER2-low patients. The scientists developed a test that combined quantitative immunofluorescence, using the CyteFinder II HT multiplexed fluorescent imaging platform from RareCyte, with a cell line array standardized with mass spectrometry to measure HER2 proteins in attomoles per square millimetre (attomol/mm2).

“Our new quantitative assay is much more sensitive and accurate at detecting low levels of HER2 proteins than conventional tests. Before, it was like weighing mice on a scale built for elephants. Now, we have essentially developed a scale for mice. And because our assay actually measures the number or concentration of HER2 molecules, rather than pathologists making estimations by eye, it also removes the subjectivity seen in traditional diagnostics,” explains Rimm. Using this method, the scientists determined that the HER2-low range expression is between 2 and 20 attomol/mm2. “Interestingly, when we applied this assay in our initial tests we discovered that 67% of breast cancer patients that were scored a 0 by traditional IHC tests actually had HER2 expression high enough to suggest they may be categorized as HER2-low,” Rimm adds.

The CyteFinder imaging system was key to the success of the scientists' work, thanks to its ability to deliver robust, precise and repeatable measurements. Rimm says, “with our older equipment, assays and scans were slow and the software that we used was outdated. The CyteFinder platform uses advanced technology and modern software that make it perfectly suited for lab-developed tests. One of the main reasons I enjoyed working with RareCyte was their willingness to work with us on customizing the system to make the assay high throughput”.

Enhancing treatment of breast cancer patients

Dr. Rimm’s trailblazing work in the field of quantitative pathology and his use of cutting-edge multiplex spatial imaging instruments is advancing the triage of breast cancer patients and helping determine which patients are candidates for treatment with Enhurtu. Using traditional IHC HER2 tests, pathologists may score patients 0, even though they may actually have HER2 molecules present and could benefit from the drug. With Rimm’s new and sufficiently sensitized test for low-range HER2, these patients are more likely to fall under the HER2-low category, meaning a significantly higher number of HER2 breast cancer patients can get access to potentially life-saving treatment. On the other hand, with current tests, some patients are wrongly classified as HER2-low. When in fact, they don’t have enough of the HER2 target present to benefit from Enhurtu. This is not only misleading for the patient but also means they may be taking drugs unnecessarily. It’s important to note that the quantitative assay has not yet been included in studies that determine what levels of HER2 are required, and these studies are currently being planned by the team.

Rimm emphasizes how advantageous it is to measure results rather than estimate or read by eye, in order to effectively match patients with the correct drugs. He believes as digital pathologies and technologies like CyteFinder advance, there will be a gradual move toward this type of precise testing. Rimm states: “As additional ADCs emerge, I think quantifying drug targets rather than just identifying if they are present will become more important. For example, there are currently two different approved ADCs for breast cancer treatment - Enhurtu which targets HER2, and sacituzumab govitecan which targets Trop2. At the moment, there is no test that measures both of these targets. This makes it hard for pathologists to know which biomarker is the most abundant and therefore which drug will be the most effective.”

Rimm believes their quantitative standardized test is just the beginning of a new direction for pathologists. He hopes it could be used as a model for many other tests that determine the biology of tumors in order to match patients with the correct drugs. Currently, the team offers their laboratory developed test (LDT) in their clinical lab at Yale University. In the future, they are looking to work with other companies to make their test more available to clinicians and hope to scale up their throughput to improve the triage and stratification of breast cancer patients.

For more information on the CyteFinder II HT multiplexed fluorescent imaging platform and other spatial biology tools, please contact RareCyte