Thomas Jefferson University

Information on Use of Human Subjects:

This project does not involve the use of multiple protocols/subprojects.

Identifier or number: ER63055

Institutional Review Board (IRB) Review:
Type of Review: Expedited
Approving Institution: Thomas Jefferson University
Most recent approval: 07/12/06
IRB approval number: 00.1138
Explanation of IRB approval:
Project is complete.

Number of human subjects who participated in this project/protocol/subproject in the last reporting period: 0
Reporting period for number of human subjects: Fiscal Year 2007

Type(s) of Human Subjects Involvement:

Collection of personally identifiable bodily materials (blood or blood products, urine, cells, tissue, teeth, organs, excreta, etc):

• Using cells cultured in a laboratory.
• Using bodily materials collected specifically for this project.

(a. Objectives, b. Methodology, c. Ionizing Radiation, Radioactive Substances, or Chemical Substances to which human subjects are exposed, d. Involvement of Human Subjects [d.1. procedures used, d.2. risks if any])

a. Objectives:
In 2005, approximately 40,810 women in the U.S. will die from breast cancer. Mammography and physical examination miss up to 50 percent of early breast cancers. Moreover, if an abnormality is found, an invasive diagnostic procedure must still be performed to determine if the breast contains atypia or cancer, even though 66 to 85 percent of abnormalities are benign. However, scintigraphic imaging of gene expression in vivo by non-invasive means could precisely direct physicians to appropriate intervention at the onset of disease and could contribute extensively to the management of patients. Until now, no method has been available to image specific overexpressed oncogene mRNAs in vivo by scintigraphic imaging. We have now observed that Tc-99m-PNA-peptides specific for the insulin-like growth factor 1 (IGF1) receptor and CD1 mRNA can delineate human ER+ breast cancer xenografts in nude mice, while peptide mismatches or PNA mismatches do not. We hypothesize that gamma-emitting Tc-99m-PNA-peptides and positron-emitting Cu-64-PNA-peptides will be taken up by human ER+ and ER- breast cancer cells, hybridize to complementary mRNA targets, and permit molecular analysis of oncogene activation in human breast cancer xenografts in nude mice by noninvasive imaging. This method will enable noninvasive detection of precancer, as well as invasive or recurrent breast cancer. Oncogenes cyclin D1 (CCND1), IGF1 receptor (IGF1R), myelocytomatosis (MYC), human epidermal growth factor 2 receptor (HER2), insulin receptor substrate 1 (IRS1), and tumor suppressor p53 will be probed. In principle, noninvasive molecular imaging of gene expression will be useful for diagnosis of other solid tumors and other proliferative abnormalities.

b. Methodology:
1. We will prepare gamma-emitting Tc-99m-PNA-peptides and positron-emitting Cu-64-PNA-peptides with antisense sequences known to hybridize specifically to CCND1, IGF1R, MYC, HER2, IRS1, and p53 mRNAs, and specific for the IGF1 receptor protein on ER+ breast cancer cells, or Her2 protein on ER- breast cancer cells.

2. We will test the specificity of uptake and mRNA hybridization of our radionuclide-PNA-peptides in ER+ and ER- breast cancer cell xenografts in nude mice by scintigraphic imaging or positron emission tomography of CCND1, IGF1R, MYC, HER2, IRS1, and p53 mRNAs in the tumors, compared with gel electrophoretic analysis of excised tumor cell RNA.

3. We will test the specificity of uptake, intracellular localization, and mRNA hybridization of fluorophore-PNA-peptides specific for CCND1, IGF1R, MYC, HER2, IRS1, and p53 mRNAs in freshly excised ER+ and ER- cancerous human breast tissue.

c. Ionizing Radiation:
Only excised cells will be exposed to Tc-99m or Cu-64 labeled probes.

d. Involvement of Human Subjects:
1. Patients with resectable mammary tumors will be asked for permission to analyze the tissues removed during surgery and a blood sample. As soon as possible after removal from the chest wall, the tumor will be aspirated on a back table in the operating room, in Dr. Sauter's laboratory, or in the pathology suite for specimen collection. Tumor levels of cyclin D1, ERBB2, c-MYC, and p53 mRNAs, measured by (1) gamma imaging of Tc-99m-PNA-peptides specific for cyclin D1, HER2, MYC, IRS1, and p53 mRNAs; and (2) reverse transcriptase/polymerase chain reaction. As a baseline control, white blood cell levels of those mRNAs will be measured by reverse transcriptase/polymerase chain reaction. To attain the goal of 10 matched pairs of organotypic cultures of breast cancer cells and normal cells probed with Tc-99m antisense PNA to quantitate oncogene mRNAs, up to 30 patient samples will be collected and processed.

2. Risk to the subject due to blood draws and laboratory analysis of excised tumor sample is negligible, which is less than minimal. Despite the potential benefits for future patients, no benefit may be claimed for the subjects of this experiment. No members of vulnerable populations will be enrolled.

3. Collection of patient samples did not begin until the protocol was approved by the Cancer Clinical Research Review Committee and the Institutional Review Board. Written informed consent was obtained from all patients before their surgical samples were collected for analysis. Records and data will be kept confidential to protect patient privacy.