From High-Throughput Microarray-Based Screening to Clinical Application: The Development of a Second Generation Multigene Test for Breast Cancer Prognosis

Abstract

Several multigene tests have been developed for breast cancer patients to predict the individual risk of recurrence. Most of the first generation tests rely on proliferation-associated genes and are commonly carried out in central reference laboratories. Here, we describe the development of a second generation multigene assay, the EndoPredict test, a prognostic multigene expression test for estrogen receptor (ER) positive, human epidermal growth factor receptor (HER2) negative (ER+/HER2−) breast cancer patients. The EndoPredict gene signature was initially established in a large high-throughput microarray-based screening study. The key steps for biomarker identification are discussed in detail, in comparison to the establishment of other multigene signatures. After biomarker selection, genes and algorithms were transferred to a diagnostic platform (reverse transcription quantitative PCR (RT-qPCR)) to allow for assaying formalin-fixed, paraffin-embedded (FFPE) samples. A comprehensive analytical validation was performed and a prospective proficiency testing study with seven pathological laboratories finally proved that EndoPredict can be reliably used in the decentralized setting. Three independent large clinical validation studies (n = 2,257) demonstrated that EndoPredict offers independent prognostic information beyond current clinicopathological parameters and clinical guidelines. The review article summarizes several important steps that should be considered for the development process of a second generation multigene test and offers a means for transferring a microarray signature from the research laboratory to clinical practice.

Is Axillary Sentinel Lymph Node Biopsy Required in Patients Who Undergo Primary Breast Surgery

“…Even in the era of a more thorough understanding of tumor biology in early breast cancer, nodal status remains an important prognostic factor. When developing a gene expression signature for estrogen receptor(ER)-positive/HER2-negative early breast cancer, we realized that the prognostic accuracy could be substantially improved after including the number of involved lymph nodes in the model [55]. The molecular information of EndoPredict® (EP) (Myriad Genetics, Salt Lake City, UT, USA) was consequently combined with the clinical parameters nodal status and tumor size, resulting in the molecular and clinical risk score, EndoPredict-clin (EPclin) which outperformed all conventional clinicopathologic risk factors in prospective-retrospective validation studies [56].…”

Prognostic Value of EndoPredict in Women with Hormone Receptor–Positive, HER2-Negative Invasive Lobular Breast Cancer

“…However, the development and validation of these assays have been mostly performed in women without considering histologic subtype and little is known about the performance of these assays specifically in ILCs. EndoPredict (EPclin) covers several cellular processes, such as proliferation, apoptosis, DNA repair, cell adhesion, and cell signaling, and combines the expression of proliferative and ER signaling-associated genes with information on nodal status and tumor size (9)(10)(11). This combined algorithm is used as a diagnostic test in the clinical setting.…”

Selecting postoperative adjuvant systemic therapy for early‐stage breast cancer: An updated assessment and systematic review of leading commercially available gene expression assays

“…Details of the development and validation of this assay are summarized elsewhere. 13,30 The primary validation studies were prospective-retrospective, meeting TMUGS standards for level IB evidence. 5,18,[31][32][33] p < 0.0001), as well as for LN+ patients limited to 1-3 positive nodes (HR: 3.7; 95% CI: 1.7-7.7; p = 0.0003).…”

“…Oncotype DX ®The Oncotype DX assay is an RT-PCR-based 21-gene assay that was designed to assess DR risk in ER+ ESBC. A proprietary algorithm produces a continuous Recurrence Score ® (RS) result from 0 to 100, with low (0-17), intermediate(18)(19)(20)(21)(22)(23)(24)(25)(26)(27)(28)(29)(30), and high-risk categories. Alternative risk cut points have been proposed and increasingly utilized 11,12,50.…”