A very important change in the breast cancer treatment landscape has made it necessary to expand access to routine testing for acquired genetic variants in patients: the availability of new drugs to address mutations associated with treatment resistance.
Patients diagnosed with hormone receptor-positive (HR+) breast cancer are typically treated with first-line endocrine therapies designed to reduce estrogen levels in the body. A common outcome of these treatments is the development of mutations in the ESR1 gene, seen in up to 40% of patients.1 These acquired variants are specific to patients undergoing endocrine therapies, particularly aromatase inhibitors; they are very rarely seen in treatment-naïve patients.
Of the many variants that have been documented in the ESR1 gene, a certain subset can confer treatment resistance. Such mutations actually eliminate the tumor’s need for estrogen, turning on the biological pathway that is usually only active in the presence of estrogen and once again giving the cancer the ingredients it uses to grow. The endocrine therapies blocking the activation of estrogen cease being effective, and the cancer can relapse.
There are a few approved therapies targeting ESR1 mutations (fulvestrant, elacestrant) that can be used to render those tumors susceptible to treatment again. And positive data from ongoing clinical trials indicates that more effective ESR1-targeting treatments may be on the way.
In light of these developments, it has become essential for clinical laboratories to perform routine monitoring for ESR1 variants in patients with HR+ metastatic breast cancer who are being treated with endocrine therapies. Recently, guidelines from the American Society of Clinical Oncology were updated; they now include a recommendation for frequent ESR1 monitoring to detect emerging mutations during the course of treatment for patients who have been diagnosed with estrogen receptor-positive metastatic breast cancer.2 Clinical data clearly demonstrates that intervening early — as soon as a new mutation is detected — gives patients better outcomes. In some cases, monthly testing may be appropriate to catch mutations as early as possible.
Clinical trials that are underway have shown promising interim results to support the idea of using ESR1-targeting therapies to overcome treatment resistance in cancers that previously responded well to endocrine therapies. One major trial has been evaluating a currently available therapy, while another has been assessing a candidate therapy not yet approved by the FDA.
The multi-center, randomized, phase 3 PADA-1 trial has been going on for about seven years. It is testing the outcomes of incorporating an ESR1-targeting therapy for patients who develop an ESR1 mutation. The trial includes about 1,000 patients diagnosed with HR+ metastatic breast cancer, all of whom began on a conventional endocrine therapy.3 Patients in one arm of the trial are monitored with bi-monthly ESR1 genetic testing, and the detection of a relevant mutation triggers a shift to fulvestrant, a treatment that has shown good results when used for cancers with ESR1 mutations. Other patients can be switched to fulvestrant if their regular imaging scans reveal growth of the tumor, indicating that the cancer was no longer responding to the endocrine therapy. The PADA-1 trial is scheduled for completion in 2025, but already, reported results have been promising. While patients who were switched to fulvestrant due to imaging-based detection of tumor progression received little benefit, the patients who made the switch earlier — when ESR1 mutations were first detected by genetic testing — had a significant change in outcome.4,5 For the latter group, median progression-free survival was more than twice that of a control group that stayed on aromatase inhibitor therapy.
A candidate ESR1-targeting therapy from AstraZeneca also appears to be performing well in clinical trials. The SERENA series of trials is evaluating the new therapy, camizestrant, in patients with breast cancer.6 In a phase 2 trial, results demonstrated that patients with ESR1 mutations experienced almost three times as much progression-free survival when using the candidate treatment compared to standard of care.7 A phase 3 trial known as SERENA-6 is now underway, but expectations are that camizestrant will be effective against a broad range of ESR1 mutations for patients whose cancers become resistant to endocrine therapy.
How to test for ESR1 mutations
For clinical laboratories interested in expanding their test menu with ESR1 monitoring, there are a number of options. Due to differences in technology availability and other factors, though, not all approaches will be right for all labs. Before considering any individual method, it is worth reviewing the key attributes of any ESR1 test.
Sensitivity: ESR1 mutations will first emerge at very low levels against a significantly larger wild-type. Any testing platform must be sensitive enough to identify a clinically relevant mutation that may be present at vanishingly small volumes.
Accuracy: While a test must be sensitive enough to detect ESR1 mutations, it must also be accurate enough not to lead to false-positive results. Most molecular detection technologies that have the required sensitivity will also have the necessary specificity, but it is important to confirm this in any platform evaluation.
Easy access: Clinical trials have proven that shifting patients to an ESR1-targeting therapy as soon as possible is the best way to improve health outcomes. Consequently, ESR1 monitoring must be performed frequently — maybe even monthly — for HR+ metastatic breast cancer patients taking endocrine therapies. Given the need for such frequent testing, an easily accessible sample such as a liquid biopsy is preferable.
With increasing demand for ESR1 testing, an in-house test is likely to be faster and more cost-effective than a send-out test. Here’s a quick look at the different technology platforms available for this application.
Droplet digital PCR: This is the technology used in the PADA-1 clinical study, with demonstrated sensitivity for detecting ESR1 mutations. Digital PCR is highly sensitive and can be used with blood samples. However, there is limited availability of digital PCR platforms in clinical laboratories because of their high cost and complex setup. For ESR1 mutations, there is the added drawback of having to run several different reactions to test the full range of clinically relevant variants.
Next-generation sequencing: This widely available technology is also sensitive enough to detect ESR1 mutations amid a background of non-tumor DNA from virtually any sample type. Unfortunately, most NGS platforms are not designed for cost-effective analysis of single genes. Clinical labs with very high demand for ESR1 mutation testing might have sufficient sample volume to run an NGS-based test affordably, but most labs will find it too expensive to deploy a modern sequencer for this purpose.
qPCR: Unlike the prior two techniques, qPCR instrumentation does not typically meet the sensitivity threshold for ESR1 variant detection. But a new method may allow clinical lab teams to use this affordable, widely available technology for ESR1 monitoring. Scientists have successfully paired circulating tumor DNA (ctDNA) with exosomal RNA analysis to produce highly sensitive ESR1 mutation detection on a standard qPCR instrument. The idea is to use a liquid biopsy approach, capturing both the low-abundance ctDNA and the high-abundance exosomes shed by a tumor. Either signal on its own may be undetectable with qPCR, but combining them into a single assay overcomes that issue. In a presentation at last year’s annual meeting of the Association for Molecular Pathology, scientists reported the development of a single workflow identifying allele-specific ESR1 mutations from ctDNA and exosomal RNA, achieving the sensitivity required for clinical monitoring.8
Going forward, clinical lab teams may find that one of the approaches above is a good fit for incorporating ESR1 monitoring. With ESR1-targeting therapies available today, and more on the horizon, routine variant testing will be a useful way to spot the signs of treatment resistance among HR+ breast cancer patients undergoing endocrine therapy.
References
1. Hermida-Prado F, Jeselsohn R. The ESR1 Mutations: From Bedside to Bench to Bedside. Cancer Res. 2021;1;81(3):537-538. doi:10.1158/0008-5472.CAN-20-4037.
2. Burstein HJ, DeMichele A, Somerfield MR, et al. Testing for ESR1 Mutations to Guide Therapy for Hormone Receptor-Positive, Human Epidermal Growth Factor Receptor 2-Negative Metastatic Breast Cancer: ASCO Guideline Rapid Recommendation Update. J Clin Oncol. 2023;20;41(18):3423-3425. doi:10.1200/JCO.23.00638.
3. National Library of Medicine. PAlbociclib and Circulating Tumor DNA for ESR1 Mutation Detection (PADA-1). Available at https://clinicaltrials.gov/study/NCT03079011. Accessed February 28, 2024.
4. Bidard FC, Hardy-Bessard AC, Dalenc F, et al. Switch to fulvestrant and palbociclib versus no switch in advanced breast cancer with rising ESR1 mutation during aromatase inhibitor and palbociclib therapy (PADA-1): a randomised, open-label, multicentre, phase 3 trial. Lancet Oncol. 2022;23(11):1367-1377. doi:10.1016/S1470-2045(22)00555-1.
5. Helwick C. PADA-1 Trial: With Early Identification of ESR1 Mutation, Switch to Fulvestrant in Metastatic Breast Cancer. October. 2022;10.
6. National Library of Medicine. Phase III Study to Assess AZD9833+ CDK4/6 Inhibitor in HR+/HER2-MBC With Detectable ESR1m Before Progression (SERENA-6) (SERENA-6). Available at https://www.clinicaltrials.gov/study/NCT04964934. Accessed February 28, 2024.
7. Camizestrant significantly delayed disease progression in advanced ER-positive breast cancer, adding at least 3.5 months benefit versus Faslodex. Astrazeneca.com. Published December 8, 2022. Accessed March 4, 2024. https://www.astrazeneca.com/media-centre/press-releases/2022/camizestrant-significantly-delayed-disease-progression-in-advanced-er-positive-breast-cancer.html.
8. Thibert JR. Development of a Novel Exosome-Based, Multiplexed RT-qPCR Technology for Rapid and Accurate Detection of Circulating Tumor Acquired Resistance Variants in ESR1 at ≤ 0.1% Frequency. Poster Presented at the Annual Meeting of the Association for Molecular Pathology.; 2023.
