Sub Topic | Secondary Topic: Biotherapeutics and Biotechnology - Biomarkers | Analytical Qualification and Validation
Authors: Srinidi Mohan, University of New England (Main Author, Presenting Author); David Barlow, University of New England; Kelly Frazier, University of New England
Presenting Author: Srinidi Mohan
Purpose: Estrogen-negative (ER–) patients has been recognized as the more aggressive subtype, more difficult to treat, greater ethnic disparity concerns, worse prognosis, and twice the risk of mortality compared to estrogen-positive (ER+) tumors. We have identified a novel maker (viz., Nw-hydroxy L-Arginine, NOHA; U.S. Utility Patent Application., 15/272,841; and International Cooperative Treaty Patent Application., PCT/US2016/053055) as a less-invasive blood based indicator for ER– breast cancer prognosis and disease progression measurement.
Methods: Deidentified ER– breast cancer patient plasma samples of African American (AA), Caucasian (CA), Jewish, Asian and Hispanic origin were tested for NOHA by LC-MS (in a multiple reaction-monitoring mode, lower limit set at 2.5-10 nM), and correlated with genetic risk-profile tumor reoccurrence patient report (viz., mammaPrint®). NOHA stability was assessed at various temperatures (-80˚C, -20˚C, 4˚C, 25˚C, 37˚C and 42˚C), and durations (1-14 days). Metabolic relevance of NOHA accumulation with inducible nitric oxide synthase (NOS2) expression, and functional activity; as well as its correlation with overall arginase activity in ER– versus ER+ tumor 3D-spheroids were assessed using commercial kits. Statistical difference was set at p<0.01.
Results: We identified a ≥49% and ≥65% reduction in NOHA only in ER– tumor groups of CA and AA origin respectively than those of healthy groups (Fig.1). Level of NOHA reduction in ER– Jewish groups was ≥ 24 % more than those of ER– Asian or ER– Hispanic origin. The level of NOHA reduction between ER– Asian, ER–Hispanic, and ER– CA groups were comparable to one another (Fig. 1). In addition to ethnic selectivity, the degree of NOHA reduction was closely associated with the genetic risk level (i.e., high versus low risk), as well as the disease state (i.e., grade 1 versus grade 2) among ER– AA versus ER– CA groups (Fig.2). NOHA stability was maintained for 7 days in plasma, at ≤ 4 ˚C; and for 14 days in dry plasma spots at ≤ 42˚C. The NOHA reduction correlated with at least 1-fold increase in both NOS2 expression and functional activity (measured as increase in total nitrite), with at least 30% increase in arginase activity level by week 1; that showed significant progressive increase over 10-week period of 3D spheroid incubation.
Conclusion: The present study provides the first evidence for NOHA as a sensitive and stable indicator for ER– breast tumor prognosis in ethnically disparate population that is based on disease progression. Further in-depth studies are needed to validate NOHA as a new indicator that can monitor therapy outcome in responsive subgroups and prevent unnecessary exposure of unresponsive patients to ineffective therapy.
See attached abstract pdf for images.