Folnetti A Alvarez1, Hussein Kaddour1,2, Yuan Lyu1,3, Christina Preece4,5, Jules Cohen6,7, Lea Baer6,7, Alison T Stopeck6,7, Patricia Thompson4,7, Chioma M Okeoma8,9. 1. Department of Pharmacology, Stony Brook University Renaissance School of Medicine, Stony Brook, NY, 11794-8651, USA. 2. Regeneron Pharmaceuticals, Inc, Tarrytown, NY, 10591, USA. 3. Medical Research Center, The Third Affiliated Hospital of Zhengzhou University, Zhengzhou, 450052, China. 4. Department of Pathology, Stony Brook University Renaissance School of Medicine, Stony Brook, NY, 11794-8651, USA. 5. Department of Medicine, Cedars Sinai Medical Center, Los Angeles, CA, 90048, USA. 6. Department of Medicine, Division of Hematology and Medical Oncology, Stony Brook University, Stony Brook, NY, 11794-8651, USA. 7. Stony Brook University Cancer Center, Stony Brook, NY, 11794-8651, USA. 8. Department of Pharmacology, Stony Brook University Renaissance School of Medicine, Stony Brook, NY, 11794-8651, USA. chioma.okeoma@stonybrook.edu. 9. Department of Pathology, Microbiology, and Immunology, New York Medical College, Valhalla, NY, 10595-1524, USA. chioma.okeoma@stonybrook.edu.
Abstract
PURPOSE: Circulating blood plasma derived extracellular vesicles (BEVs) containing proteins hold promise for their use as minimally invasive biomarkers for predicting response to cancer therapy. The main goal of this study was to establish the efficiency and utility of the particle purification liquid chromatography (PPLC) BEV isolation method and evaluate the role of BEVs in predicting breast cancer (BC) patient response to neoadjuvant chemotherapy (NAC). METHODS: PPLC isolation was used to separate BEVs from non-EV contaminants and characterize BEVs from 17 BC patients scheduled to receive NAC. Using LC-MS/MS, we compared the proteome of PPLC-isolated BEVs from patients (n = 7) that achieved a pathological complete response (pCR) after NAC (responders [R]) to patients (n = 10) who did not achieve pCR (non-responders [NR]). Luminal MCF7 and basaloid MDA-MB-231 BC cells were treated with isolated BEVs and evaluated for metabolic activity by MTT assay. RESULTS: NR had elevated BEV concentrations and negative zeta potential (ζ-potential) prior to receipt of NAC. Eight proteins were enriched in BEVs of NR. GP1BA (CD42b), PECAM-1 (CD31), CAPN1, HSPB1 (HSP27), and ANXA5 were validated using western blot. MTT assay revealed BEVs from R and NR patients increased metabolic activity of MCF7 and MDA-MB-231 BC cells and the magnitude was highest in MCF7s treated with NR BEVs. CONCLUSION: PPLC-based EV isolation provides a preanalytical separation process for BEVs devoid of most contaminants. Our findings suggest that PPLC-isolated BEVs and the five associated proteins may be established as predictors of chemoresistance, and thus serve to identify NR to spare them the toxic effects of NAC.
PURPOSE: Circulating blood plasma derived extracellular vesicles (BEVs) containing proteins hold promise for their use as minimally invasive biomarkers for predicting response to cancer therapy. The main goal of this study was to establish the efficiency and utility of the particle purification liquid chromatography (PPLC) BEV isolation method and evaluate the role of BEVs in predicting breast cancer (BC) patient response to neoadjuvant chemotherapy (NAC). METHODS: PPLC isolation was used to separate BEVs from non-EV contaminants and characterize BEVs from 17 BC patients scheduled to receive NAC. Using LC-MS/MS, we compared the proteome of PPLC-isolated BEVs from patients (n = 7) that achieved a pathological complete response (pCR) after NAC (responders [R]) to patients (n = 10) who did not achieve pCR (non-responders [NR]). Luminal MCF7 and basaloid MDA-MB-231 BC cells were treated with isolated BEVs and evaluated for metabolic activity by MTT assay. RESULTS: NR had elevated BEV concentrations and negative zeta potential (ζ-potential) prior to receipt of NAC. Eight proteins were enriched in BEVs of NR. GP1BA (CD42b), PECAM-1 (CD31), CAPN1, HSPB1 (HSP27), and ANXA5 were validated using western blot. MTT assay revealed BEVs from R and NR patients increased metabolic activity of MCF7 and MDA-MB-231 BC cells and the magnitude was highest in MCF7s treated with NR BEVs. CONCLUSION: PPLC-based EV isolation provides a preanalytical separation process for BEVs devoid of most contaminants. Our findings suggest that PPLC-isolated BEVs and the five associated proteins may be established as predictors of chemoresistance, and thus serve to identify NR to spare them the toxic effects of NAC.
Authors: Jennifer L Welch; Hussein Kaddour; Patrick M Schlievert; Jack T Stapleton; Chioma M Okeoma Journal: J Virol Date: 2018-10-12 Impact factor: 5.103
Authors: Yuan Lyu; Hussein Kaddour; Steven Kopcho; Tyler D Panzner; Nadia Shouman; Eun-Young Kim; Jeremy Martinson; Heather McKay; Otoniel Martinez-Maza; Joseph B Margolick; Jack T Stapleton; Chioma M Okeoma Journal: Cells Date: 2019-09-03 Impact factor: 7.666
Authors: Hussein Kaddour; Tyler D Panzner; Jennifer L Welch; Nadia Shouman; Mahesh Mohan; Jack T Stapleton; Chioma M Okeoma Journal: Viruses Date: 2020-10-01 Impact factor: 5.048