Transcripts with Retained H/ACA Box SnoRNA Sequences As Biomarkers for Estrogen Dependence in Lum-B Breast Carcinomas

Background and Rationale of the Study The primary approach to treating early-stage estrogen receptor-positive breast cancer involves the use of antitumor drugs (e.g., Tamoxifen) that selectively inhibit estrogen receptors, thereby reducing the risk of recurrence and cancer-related mortality. However, individual responses to hormonal therapy can be highly variable, particularly in luminal B breast cancer, necessitating adjuvant chemotherapy for some patients despite its associated toxic effects. The standard approach to determining the most appropriate therapy for these patients relies on clinical-pathological tumor factors, which, however, have suboptimal prognostic and predictive utility for determining chemotherapy benefits.

To overcome this limitation, several multigene tests have been developed to better stratify prognosis and treatment response. Currently, the standard test for estrogen receptor-positive breast cancer is the Oncotype DX Breast Recurrence Score. This test is based on analyzing the expression of a selected list of genes using RNA extracted from paraffin-embedded patient tumor tissue. It provides a Recurrence Score that characterizes the tumor and guides the recommended therapy. Since 2021, it has been part of the standard of care for luminal B breast cancer patients in Italy. However, its optimal use in intermediate-risk patients remains undefined.

In a recently published study, we identified a specific group of transcripts associated with estrogen response in mammary gland tumors. These transcripts are specific mRNA isoforms that exhibit a unique feature: introns containing the H/ACA box snoRNA (small-nucleolar RNA) sequence. For this reason, we named them snoRT (snoRNA Retain Transcripts). The expression of these transcripts is not only associated with hormone receptor status (both estrogen and progesterone receptors) but is also significantly linked to the survival of hormone receptor-positive patients. Furthermore, we showed that the snoRTs identified in the study interact with other protein-coding transcripts involved in mediating the hormone receptor response. The expression of these transcripts is associated with the survival of breast cancer patients, particularly those with luminal B tumors, making them strong candidates for identifying a potential "molecular signature" associated with luminal B breast cancer outcomes.

Importance of the Study and its Clinical Relevance The recent discoveries mentioned in the background reveal an unexplored role of snoRTs and their interacting protein-coding mRNAs in influencing the dependency of specific tumors on nuclear hormone receptors. Our findings suggest that deregulation of these functions may allow luminal B tumors to escape hormonal dependency, becoming more aggressive and potentially unresponsive to anti-estrogen therapy.

Since estrogen-dependent luminal B tumors sometimes develop characteristics enabling them to overcome hormonal dependency, making them resistant to anti-estrogen therapy, studying this new "molecular signature" and these novel cytoplasmic transcripts appears promising. We propose comparing them with the Oncotype DX test to evaluate whether studying these RNAs can add useful insights for identifying patients unlikely to respond well to anti-estrogen treatments (e.g., those classified as intermediate-risk by the Oncotype DX score). If validated, these new biomarkers could lead to the improvement and refinement of current clinical molecular tests.

The proposed study would represent a significant innovation in the field of biomarkers, as the role of this new class of RNAs (snoRTs) has never been thoroughly investigated and is entirely overlooked by traditional RNA-seq analyses based on genes (due to molecular characteristics related to their localization, transcription, and the maturation of snoRNA genes). The role of snoRTs in regulating gene expression and human diseases remains an unexplored field, and their diagnostic and predictive significance has not been previously considered.

WHAT IS ALREADY KNOWN ABOUT THE TOPIC The primary approach to treating estrogen receptor-positive breast cancer involves using estrogen receptor inhibitors. However, individual responses to hormonal therapy can be highly variable, particularly in luminal B breast cancer, necessitating adjuvant chemotherapy for some patients. To determine the most appropriate therapy for these patients, the preferred test for estrogen receptor-positive breast cancer is the Oncotype DX Breast Recurrence Score. This test provides a Recurrence Score that characterizes the tumor and guides treatment recommendations. Although validated, its optimal use in intermediate-risk patients remains undefined.

WHAT THE STUDY COULD ADD This project aims to define innovative approaches for more precise profiling of luminal B patients who may have developed mechanisms of hormonal independence that often escape detection by conventional tests. Thus, the results of this study could contribute to identifying patients who do not respond to estrogen receptor-targeted therapies. This would enable patients to benefit from more appropriate, and therefore more aggressive, treatment, similar to that offered to high-recurrence-risk patients identified by the Oncotype DX test.