Persistent upregulation of U6: SNORD44 small RNA ratio in the serum of breast cancer patients

Hitesh N. Appaiah, Chirayu P. Goswami, Lida A. Mina, Sunil Badve, George W. Sledge, Yunlong Liu, Harikrishna Nakshatri

Research output: Contribution to journalArticle

55 Citations (Scopus)

Abstract

Introduction: Serum microRNAs have the potential to be valuable biomarkers of cancer. This investigation addresses two issues that impact their utility: a) appropriate normalization controls and b) whether their altered levels persist in patients who are clinically free of the disease.Methods: Sera from 40 age-matched healthy women and 39 breast cancer patients without clinical disease at the time of serum collection were analyzed for microRNAs let-7f, miR-16, miR-21 and miR-155 using quantitative real-time PCR. U6 and 5S, which are transcribed by RNA polymerase III (RNAP-III) and the small nucleolar RNU44 (SNORD44), were also analyzed for normalization. Significant results from the initial study were verified using a second set of sera from 15 healthy patients, 15 breast cancer patients without clinical disease and 15 with metastatic disease, and a third set of 12 healthy and 18 patients with metastatic disease. U6 was further verified in the extended second cohort of 75 healthy and 68 breast cancer patients without clinical disease.Results: U6:SNORD44 ratio was consistently higher in breast cancer patients with or without active disease (fold change range 1.5-6.6, p value range 0.0003 to 0.05). This increase in U6:SNORD44 ratio was observed in the sera of both estrogen receptor-positive (ER+) and ER-negative breast cancer patients. MiR-16 and 5S, which are often used as normalization controls for microRNAs, showed remarkable experimental variability and thus are not ideal for normalization.Conclusions: Elevated serum U6 levels in breast cancer patients irrespective of disease activity at the time of serum collection suggest a new paradigm in cancer; persistent systemic changes during cancer progression, which result in elevated activity of RNAP-III and/or the stability/release pathways of U6 in non-cancer tissues. Additionally, these results highlight the need for developing standards for normalization between samples in microRNA-related studies for healthy versus cancer and for inter-laboratory reproducibility. Our studies rule out the utility of miR-16, U6 and 5S RNAs for this purpose.

Original languageEnglish
Article numberR86
JournalBreast Cancer Research
Volume13
Issue number5
DOIs
StatePublished - Sep 13 2011

Fingerprint

Up-Regulation
RNA
Breast Neoplasms
Serum
MicroRNAs
RNA Polymerase III
Neoplasms
Tumor Biomarkers
Estrogen Receptors
Real-Time Polymerase Chain Reaction

ASJC Scopus subject areas

  • Cancer Research
  • Oncology
  • Medicine(all)

Cite this

Persistent upregulation of U6 : SNORD44 small RNA ratio in the serum of breast cancer patients. / Appaiah, Hitesh N.; Goswami, Chirayu P.; Mina, Lida A.; Badve, Sunil; Sledge, George W.; Liu, Yunlong; Nakshatri, Harikrishna.

In: Breast Cancer Research, Vol. 13, No. 5, R86, 13.09.2011.

Research output: Contribution to journalArticle

@article{3178154abe464291afb8edd34d7ed6a0,
title = "Persistent upregulation of U6: SNORD44 small RNA ratio in the serum of breast cancer patients",
abstract = "Introduction: Serum microRNAs have the potential to be valuable biomarkers of cancer. This investigation addresses two issues that impact their utility: a) appropriate normalization controls and b) whether their altered levels persist in patients who are clinically free of the disease.Methods: Sera from 40 age-matched healthy women and 39 breast cancer patients without clinical disease at the time of serum collection were analyzed for microRNAs let-7f, miR-16, miR-21 and miR-155 using quantitative real-time PCR. U6 and 5S, which are transcribed by RNA polymerase III (RNAP-III) and the small nucleolar RNU44 (SNORD44), were also analyzed for normalization. Significant results from the initial study were verified using a second set of sera from 15 healthy patients, 15 breast cancer patients without clinical disease and 15 with metastatic disease, and a third set of 12 healthy and 18 patients with metastatic disease. U6 was further verified in the extended second cohort of 75 healthy and 68 breast cancer patients without clinical disease.Results: U6:SNORD44 ratio was consistently higher in breast cancer patients with or without active disease (fold change range 1.5-6.6, p value range 0.0003 to 0.05). This increase in U6:SNORD44 ratio was observed in the sera of both estrogen receptor-positive (ER+) and ER-negative breast cancer patients. MiR-16 and 5S, which are often used as normalization controls for microRNAs, showed remarkable experimental variability and thus are not ideal for normalization.Conclusions: Elevated serum U6 levels in breast cancer patients irrespective of disease activity at the time of serum collection suggest a new paradigm in cancer; persistent systemic changes during cancer progression, which result in elevated activity of RNAP-III and/or the stability/release pathways of U6 in non-cancer tissues. Additionally, these results highlight the need for developing standards for normalization between samples in microRNA-related studies for healthy versus cancer and for inter-laboratory reproducibility. Our studies rule out the utility of miR-16, U6 and 5S RNAs for this purpose.",
author = "Appaiah, {Hitesh N.} and Goswami, {Chirayu P.} and Mina, {Lida A.} and Sunil Badve and Sledge, {George W.} and Yunlong Liu and Harikrishna Nakshatri",
year = "2011",
month = "9",
day = "13",
doi = "10.1186/bcr2943",
language = "English",
volume = "13",
journal = "Breast Cancer Research",
issn = "1465-5411",
publisher = "BioMed Central",
number = "5",

}

TY - JOUR

T1 - Persistent upregulation of U6

T2 - SNORD44 small RNA ratio in the serum of breast cancer patients

AU - Appaiah, Hitesh N.

AU - Goswami, Chirayu P.

AU - Mina, Lida A.

AU - Badve, Sunil

AU - Sledge, George W.

AU - Liu, Yunlong

AU - Nakshatri, Harikrishna

PY - 2011/9/13

Y1 - 2011/9/13

N2 - Introduction: Serum microRNAs have the potential to be valuable biomarkers of cancer. This investigation addresses two issues that impact their utility: a) appropriate normalization controls and b) whether their altered levels persist in patients who are clinically free of the disease.Methods: Sera from 40 age-matched healthy women and 39 breast cancer patients without clinical disease at the time of serum collection were analyzed for microRNAs let-7f, miR-16, miR-21 and miR-155 using quantitative real-time PCR. U6 and 5S, which are transcribed by RNA polymerase III (RNAP-III) and the small nucleolar RNU44 (SNORD44), were also analyzed for normalization. Significant results from the initial study were verified using a second set of sera from 15 healthy patients, 15 breast cancer patients without clinical disease and 15 with metastatic disease, and a third set of 12 healthy and 18 patients with metastatic disease. U6 was further verified in the extended second cohort of 75 healthy and 68 breast cancer patients without clinical disease.Results: U6:SNORD44 ratio was consistently higher in breast cancer patients with or without active disease (fold change range 1.5-6.6, p value range 0.0003 to 0.05). This increase in U6:SNORD44 ratio was observed in the sera of both estrogen receptor-positive (ER+) and ER-negative breast cancer patients. MiR-16 and 5S, which are often used as normalization controls for microRNAs, showed remarkable experimental variability and thus are not ideal for normalization.Conclusions: Elevated serum U6 levels in breast cancer patients irrespective of disease activity at the time of serum collection suggest a new paradigm in cancer; persistent systemic changes during cancer progression, which result in elevated activity of RNAP-III and/or the stability/release pathways of U6 in non-cancer tissues. Additionally, these results highlight the need for developing standards for normalization between samples in microRNA-related studies for healthy versus cancer and for inter-laboratory reproducibility. Our studies rule out the utility of miR-16, U6 and 5S RNAs for this purpose.

AB - Introduction: Serum microRNAs have the potential to be valuable biomarkers of cancer. This investigation addresses two issues that impact their utility: a) appropriate normalization controls and b) whether their altered levels persist in patients who are clinically free of the disease.Methods: Sera from 40 age-matched healthy women and 39 breast cancer patients without clinical disease at the time of serum collection were analyzed for microRNAs let-7f, miR-16, miR-21 and miR-155 using quantitative real-time PCR. U6 and 5S, which are transcribed by RNA polymerase III (RNAP-III) and the small nucleolar RNU44 (SNORD44), were also analyzed for normalization. Significant results from the initial study were verified using a second set of sera from 15 healthy patients, 15 breast cancer patients without clinical disease and 15 with metastatic disease, and a third set of 12 healthy and 18 patients with metastatic disease. U6 was further verified in the extended second cohort of 75 healthy and 68 breast cancer patients without clinical disease.Results: U6:SNORD44 ratio was consistently higher in breast cancer patients with or without active disease (fold change range 1.5-6.6, p value range 0.0003 to 0.05). This increase in U6:SNORD44 ratio was observed in the sera of both estrogen receptor-positive (ER+) and ER-negative breast cancer patients. MiR-16 and 5S, which are often used as normalization controls for microRNAs, showed remarkable experimental variability and thus are not ideal for normalization.Conclusions: Elevated serum U6 levels in breast cancer patients irrespective of disease activity at the time of serum collection suggest a new paradigm in cancer; persistent systemic changes during cancer progression, which result in elevated activity of RNAP-III and/or the stability/release pathways of U6 in non-cancer tissues. Additionally, these results highlight the need for developing standards for normalization between samples in microRNA-related studies for healthy versus cancer and for inter-laboratory reproducibility. Our studies rule out the utility of miR-16, U6 and 5S RNAs for this purpose.

UR - http://www.scopus.com/inward/record.url?scp=80052668330&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=80052668330&partnerID=8YFLogxK

U2 - 10.1186/bcr2943

DO - 10.1186/bcr2943

M3 - Article

C2 - 21914171

AN - SCOPUS:80052668330

VL - 13

JO - Breast Cancer Research

JF - Breast Cancer Research

SN - 1465-5411

IS - 5

M1 - R86

ER -