Abstract OT1-03-03: Investigating circulating lncRNAs in breast cancer patients on neoadjuvant systemic therapy
Authors: Krastev Boris , Timcheva Constanta , Valev Spartak , Zhbantov Georgi , Petrova Mila , Karanikolova Teodora , Gencheva Radostina , Ivanova Anika , Stoykov Ivaylo , Stamenov Georgi 
Published: March 2020

Background. In cells long non-coding RNAs (lncRNAs) control gene expression through various mechanisms, acting on different biological levels: chromatin remodeling, regulation of transcription, posttranscriptional modification. LncRNAs could also be transported via systemic circulation, encapsulated in extracellular vesicles called “exosomes” or in vesicle-free form, bounded to proteins or lipoproteins. Knowing the intracellular function of lncRNAs, it is reasonably speculated that in cancer patients their existence in bloodstream is more than just an extracellular release. It is thought to be a kind of an epigenetic mechanism, rendered by tumor cells, so the last could influence gene expression programs of normal cells in distance, creating tumor-friendly environment in other body compartments - a sine qua non for metastatic spread. Few studies have investigated in vivo circulating lncRNAs in breast cancer and even less have done it in the neoadjuvant setting.

Goals and objectives. We decided to conduct a small exploratory study in patients with locally advanced breast cancer, focusing on changes in plasma lncRNAs during neoadjuvant therapy. The two main goals of the study are: 1. Assessing the effect of systemic anticancer treatment on epigenetic regulators such as circulating lncRNAs; 2. Testing the technical feasibility of using lncRNAs as plasma biomarkers. Our objectives include lncRNA profiling in circulation before onset as well as in the course of systemic therapy, followed by correlation analysis between distinct lncRNA signatures and clinicopathological features. Differential lncRNA expression will be assessed between patients and normal controls as well as among patient subgroups according to time of sampling, age, intrinsic breast cancer phenotype and response to therapy. LncRNAs of interest will be further examined in larger cohorts of patients and healthy controls.

Design/materials/methods. This is a prospective, interventional, case-control clinical trial in two parts - exploration and validation. Patients eligibility criteria: age ≥ 18 years; females; histologically confirmed locally advanced breast cancer; multidisciplinary tumor board decision for neoadjuvant chemotherapy or chemotherapy plus targeted therapy; radiologically excluded distant dissemination; no concomitant malignancy; signed informed consent form (ICF). Healthy controls eligibility criteria: age ≥ 18 years; females; no oncologic history; signed ICF. For part I ten patients and two controls have already been recruited. Blood samples were taken from each patient twice - before onset of neoadjuvant therapy and before 4th cycle (in one patient the second sample was taken before 6th cycle), from control subjects blood was taken once. Plasma was immediately separated and frozen until RNA extraction. Next-generation RNA sequencing was performed with subsequent bioinformatics analysis on lncRNAs. Clinical and pathology information was obtained from each patient’s hospital file. In part II plasma will be collected in the same manner as in part I. Preselected lncRNAs from part I will be measured by real-time polymerase chain reaction.

Statistics. This is an exploratory study with no empirical data and no formal planning for sample size. The correlation between lncRNAs and clinicopathological characteristics and the significance of the differences among groups will be assessed by parametric tests (Pearson correlation, Student’s t-test) and/or multivariate such (ANCOVA).

Present and target accrual. Part I accrual has completed and bioinformatics data are expected. Part II accrual is ongoing with 15 patients already recruited. Target accrual for part II is 50 patients and 20 controls.

Citation Format: Boris Krastev, Constanta Timcheva, Spartak Valev, Georgi Zhbantov, Mila Petrova, Teodora Karanikolova, Radostina Gencheva, Anika Ivanova, Ivaylo Stoykov, Georgi Stamenov. Investigating circulating lncRNAs in breast cancer patients on neoadjuvant systemic therapy [abstract]. In: Proceedings of the 2019 San Antonio Breast Cancer Symposium; 2019 Dec 10-14; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2020;80(4 Suppl):Abstract nr OT1-03-03.