lmdit cw1 nowotwory piersi 2

8/13/2019 LMDiT Cw1 Nowotwory Piersi 2

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Breast tissue diagnosis by

Raman spectroscopyB. Broek-Puska*, J.Surmacki*, J. Jaboska**, R. Kordek**,

H. Abramczyk*

*Laboratory of Laser Molecular Spectroscopy,

Institute of Applied Radiation Chemistry,

Technical University of d, Faculty of Chemistry, Poland

**Department of Oncology, Medical University of d, Poland


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GOAL

Some substances are produced by the organism in response to the

cancers presence. They are called tumor markers.

Tumor markers are molecules occurring in blood or tissue that are

associated with cancer and whose measurement or identification is useful in

patient diagnosis or clinical management. The ideal marker would be a

blood test for cancer in which a positive result would occur only in

patients with malignancy, one that would correlate with stage and response

to treatment and that was easily and reproducibly measured. No tumor

marker now available has met this ideal.

We believe that optical methods including Raman spectroscopy will provide such

a marker. Possibly it will help to find the hallmarks of cancer.


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immediate in vivo diagnosis

reduction the number of biopsies

combination of biochemical and histopathological

diagnosis provides more information because pathology

is intimately related to biochemistry

method has a potential to remove human interpretation

non-invasive,non-ionizing method that probes with

chemical specificity vibrations and fluorescence (not just

structure) extremaly high spatial resolution (optical imaging)

Why ?


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Human speciemens obtained from surgery. Upon

removal during the operation, the ex vivo sample is

devided by a doctor into two parts, one goes to our lab

the second goes to the pathology examination

The ex vivo samples are neither frozen in liquidnitrogen for storage nor fixed in formalin, the fresh

tissue is measured immeditely after delivering from

the hospital

The samples for pathology measurements are

passively thawed at room temperature and kept

moist with PBS fixed in formalin

Cut through the marked locations into 5-m-thicksections, and stained with eosin

normal tissue

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514nm, normal tissue

25mW

100mW

Intensity

(arbitrary

units)

Raman shift in wavennumbers [cm-1

]

0 5 00 1 00 0 1 50 0 2 00 0 2 50 0 3 00 0 3 50 0 4 00 0

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514nm, 100mW, cancer

Inten

sity

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units)

Raman shift in wavennumbers [cm-1]

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maligant tissue

Tissue Preparation

carcinoma mammae fibroadenoma mammae


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Measurements Parameters

The laser excitation is 514 nm, the spot is d=500 m in diameter

Light diffusion in the tissue results in a spot of v1mm

Integration time 0.5 s

Spectral resolution 2 and 8 cm -1

The laser excitation power: 25 mW, 100mW


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Patients Statistics

1100 spectra from 100 patients

fibroadenomas

infiltrating carcinoma

infiltrating ductal carcinoma (IDC)

infiltrating lobular carcinoma (ILC)

IDC+ILC

multifocal carcinoma

carcinoma microinvasive

intracystic papillary carcinoma (noninvasive)

carcinoma mucinosumcarcinoma intraductal microinvasive

benign dysplasia

dysplasia benign dysplasia (cystes, apocrinal metaplasia,adenosis)

ductal-lobular hyperplasia

cystic mastopathy

0 1000 2000 3000 4000 5000 6000 7000 80000

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40000014 KT - normal tissue

25mW

514 541 572 607 647 691 743 802 872 [nm]

relative wavenumber [cm-1]

Intensity

[counts/s]

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40000014 KT - carcinoma lobulare infiltrans mammae

25mW

514 541 572 607 647 691 743 802 872 [nm]


Intensity

[counts/s]


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Comparison between the Raman spectra for normal and

malignant tissue (infiltrating ductal carcinoma (IDC))

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15000

25mW

0006 KT - normal tissue

514 541 572 607 647 691 743 802 872 [nm]


Intensity

[counts/s]

0 1000 2000 3000 4000 5000 6000 7000 80000

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25mW

0006 KT - carcinoma ductale infiltrans G2

514 541 572 607 647 691 743 802 872 [nm]


Intensity

[counts/s]


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Quenching of autofluorescence by low temperature Raman

spectroscopy

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-500

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normal tissue - patient nr 90

intensity

(cts/s)

wavenumber (cm-1)

K90T33 - 250K

K90T34 - 200K

K90T35 - 170K

K90T36 - 150K

K90T37 - 110K

K90T38 - 77K

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-100

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cancer tissue - patient nr 90

intensity

9cts/s)

wavenumber (cm-1)

K90T14 - 250K

K90T15 - 200K

K90T16 - 170K

K90T17 - 150K

K90T18 - 110K

K90T19 - 77K


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Comparison between the Raman spectra for normal and

malignant tissue (carcinoma mucinosum )

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-500

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Y

AxisTitle

X Axis Title

K14T2 (carcinoma mucinosum mammae sinistri)K14T2 (normal tissue)

wavenumber [1/cm]

intensity

[cts/s]


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Comparison between the Raman spectra for normal and benign

tumour tissue (fibroadenoma)

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YA

xisTitle

X Axis Title

K31T5 (normal tissue)

K31T8 (Fibroadenoma mamae)

wavenumber [1/cm]

intensity

[cts/s]


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PRINCIPAL COMPONENT ANALYSIS (PCA)PLS_Toolbox Version 4.0

for use with MATLAB

Barry M. WiseNeal B. GallagherRasmus BroJeremy M. Shaver

Willem WindigR. Scott Koch

Eigenvector Research, Inc.3905 West Eaglerock Drive

Wenatchee, WA 98801

[email protected]


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1

PCA score plot mean center, SNV, 1-st derivative

-1 -0.5 0 0.5 1 1.5 2-1.4

-1.2

-1

-0.8

-0.6

-0.4

-0.2

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Scores on PC 1 (66.25%)

ScoresonPC2(7

.70%)

cc

hc

hhh

h

h

c

c h

c

c

c

h

h

c

h

h

hh

hw

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h

hc h

ccc h

c

c

c hc

c

c

hw

ww

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h

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www

hww w

hh

hcc c h

c

cc

h

Samples/Scores Plot of HY

Decluttered

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No Raman peaks Intensive Raman peaks

Normal tissue

Malignant tissue

High autofluorescence

Low autofluorescence

Benign

tissue


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500 1000 1500 2000 2500 3000 3500-0.2

-0.15

-0.1

-0.05

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0.15

0.2

wavenumber (cm-1)

Load

ingsonPC1(66.2

5%)

Variables/Loadings Plot for HY

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C-H

streching

lipids

C-C and

C=C

stretching

carotenoids

PCA loading plot mean center, SNV, 1-st derivative


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CONCLUSIONS

The results clearly illustrates the ability of Raman

spectroscopy to accurately diagnose breast cancer anddemonstrates how the diagnostic scheme can be adjusted toobtain the desired degree of sensitivity and specificity(88%,72%)

The normal tissue has a characteristic bands: C-C (1110 cm-1)and C=C (1520 cm-1) stretching bands of carotenoids and at

2840-2900 cm-1 for the C-H symmetric and asymmetric bands oflipids (fat) which are not visible in the malignant tissue and inbenign tumor tissue. Moreover, the fluorescence is much higherin the malignant tissue.

We belive that in a very near future a good quality Raman

signal will be obtained with the optical fibers coupled with abiopsy needle and incorporated into Raman spectrometer forbreast tissue measurements in vivo.

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