Breast tissue diagnosis by Raman spectroscopy

Transcription

1 Breast tissue diagnosis by Raman spectroscopy B. Brożek-Płuska*, J.Surmacki*, J. Jabłońska**, R. Kordek**, H. Abramczyk* *Laboratory of Laser Molecular Spectroscopy, Institute of Applied Radiation Cemistry, Tecnical University of Łódź, Faculty of Cemistry, Poland **Department of Oncology, Medical University of Łódź, Poland

2 GOAL Some substances are produced by te organism in response to te cancer s presence. Tey are called tumor markers. Tumor markers are molecules occurring in blood or tissue tat are associated wit cancer and wose measurement or identification is useful in patient diagnosis or clinical management. Te ideal marker would be a blood test for cancer in wic a positive result would occur only in patients wit malignancy, one tat would correlate wit stage and response to treatment and tat was easily and reproducibly measured. No tumor marker now available as met tis ideal. We believe tat optical metods including Raman spectroscopy will provide suc a marker. Possibly it will elp to find te allmarks of cancer.

3 Wy? immediate in vivo diagnosis reduction te number of biopsies combination of biocemical and istopatological diagnosis provides more information because patology is intimately related to biocemistry metod as a potential to remove uman interpretation non-invasive,non-ionizing metod tat probes wit cemical specificity vibrations and fluorescence (not just structure) extremaly ig spatial resolution (optical imaging)

4 Intensity (arbitrary units) Intensity (arbitrary units) Tissue Preparation carcinoma mammae fibroadenoma mammae 514nm, 1mW, cancer maligant tissue normal tissue 514nm, normal tissue 1mW 25mW Raman sift in wavennumbers [cm -1 ] Raman sift in wavennumbers [cm -1 ] Human speciemens obtained from surgery. Upon removal during te operation, te ex vivo sample is devided by a doctor into two parts, one goes to our lab te second goes to te patology examination Te ex vivo samples are neiter frozen in liquid nitrogen for storage nor fixed in formalin, te fres tissue is measured immeditely after delivering from te ospital Te samples for patology measurements are passively tawed at room temperature and kept moist wit PBS fixed in formalin Cut troug te marked locations into 5-μm-tick sections, and stained wit eosin

5 Measurements Parameters Te laser excitation is 514 nm, te spot is d=5 μm in diameter Ligt diffusion in te tissue results in a spot of v 1mm Integration time.5 s Spectral resolution 2 and 8 cm -1 Te laser excitation power: 25 mw, 1mW

6 Intensity [counts/s] Intensity [counts/s] Patients Statistics 4 14 KT - normal tissue 11 spectra from 1 patients 3 2 fibroadenomas infiltrating carcinoma infiltrating ductal carcinoma (IDC) infiltrating lobular carcinoma (ILC) IDC+ILC multifocal carcinoma carcinoma microinvasive intracystic papillary carcinoma (noninvasive) carcinoma mucinosum carcinoma intraductal microinvasive benign dysplasia dysplasia benign dysplasia (cystes, apocrinal metaplasia, adenosis) ductal-lobular yperplasia cystic mastopaty 1 25mW relative wavenumber [cm -1 ] [nm] KT - carcinoma lobulare infiltrans mammae 25mW relative wavenumber [cm -1 ] [nm]

7 Intensity [counts/s] Intensity [counts/s] Comparison between te Raman spectra for normal and malignant tissue (infiltrating ductal carcinoma (IDC)) 15 6 KT - normal tissue 15 6 KT - carcinoma ductale infiltrans G mW relative wavenumber [cm -1 ] [nm] mW relative wavenumber [cm -1 ] [nm]

8 intensity (cts/s) intensity 9cts/s) Quencing of autofluorescence by low temperature Raman spectroscopy 25 2 normal tissue - patient nr 9 K9T33-25K K9T34-2K K9T35-17K K9T36-15K K9T37-11K K9T38-77K 6 5 cancer tissue - patient nr 9 K9T14-25K K9T15-2K K9T16-17K K9T17-15K K9T18-11K K9T19-77K wavenumber (cm-1) wavenumber (cm-1)

9 Y Axis Title intensity [cts/s] Comparison between te Raman spectra for normal and malignant tissue (carcinoma mucinosum ) 5 K14T2 (carcinoma mucinosum mammae sinistri) K14T2 (normal tissue) wavenumber X Axis [1/cm] Title

10 Y Axis Title intensity [cts/s] Comparison between te Raman spectra for normal and benign tumour tissue (fibroadenoma) 18 K31T5 (normal tissue) K31T8 (Fibroadenoma mamae) X Axis Title wavenumber [1/cm]

11 PRINCIPAL COMPONENT ANALYSIS (PCA) PLS_Toolbox Version 4. for use wit MATLAB Barry M. Wise Neal B. Gallager Rasmus Bro Jeremy M. Saver Willem Windig R. Scott Koc Eigenvector Researc, Inc. 395 West Eaglerock Drive Wenatcee, WA

12 Scores on PC 2 (7.7%) PCA score plot mean center, SNV, 1-st derivative Low autofluorescence w w w c w c w w c w w w c c c c w w c c c c c c c w w c w c Normal tissue c c Malignant tissue c c Samples/Scores Plot of HY Benign tissue Decluttered No Raman peaks Hig autofluorescence Scores on PC 1 (66.25%) Intensive Raman peaks 12

13 Loadings on PC 1 (66.25%) PCA loading plot mean center, SNV, 1-st derivative.2 Variables/Loadings Plot for HY C-C and C=C stretcing carotenoids C-H strecing lipids wavenumber (cm-1)

14 CONCLUSIONS Te results clearly illustrates te ability of Raman spectroscopy to accurately diagnose breast cancer and demonstrates ow te diagnostic sceme can be adjusted to obtain te desired degree of sensitivity and specificity (88%,72%) Te normal tissue as a caracteristic bands: C-C (111 cm -1 ) and C=C (152 cm -1 ) stretcing bands of carotenoids and at cm -1 for te C-H symmetric and asymmetric bands of lipids (fat) wic are not visible in te malignant tissue and in benign tumor tissue. Moreover, te fluorescence is muc iger in te malignant tissue. We belive tat in a very near future a good quality Raman signal will be obtained wit te optical fibers coupled wit a biopsy needle and incorporated into Raman spectrometer for breast tissue measurements in vivo.