Could Mass Spectrometry Imaging be a Drug Quantification Technique?
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1. The study investigates using mass spectrometry imaging (MSI) as a quantitative drug analysis technique by developing a methodology for absolute quantification of compounds in tissue sections. 2. The methodology involves calculating tissue extinction coefficients, generating calibration curves from drug standards on tissue, and using the curves to quantify drug levels in tissue sections from preclinical studies of three drugs. 3. The developed quantification MSI (qMSI) technique was validated by comparing quantified drug levels in tissues to levels obtained using other quantitative methods such as liquid chromatography-mass spectrometry, showing good agreement.
![COULD MASS SPECTROMETRY IMAGING BE A DRUG QUANTIFICATION TECHNIQUE?
G. Hamm1, D. Bonnel1, R. Legouffe1, F. Pamelard1, J.-M. Delbos2, F. Bouzom2, C. Piveteau3, N. Willand3, B. Déprez3, J. Stauber1
1: ImaBiotech, Parc Eurasanté, Loos, France. 2: Technologie Servier, Orléans, France. 3: INSERM U761, Biostructures & Drug Discovery, University Lille Nord de France, France.
Introduction
Unlike traditional imaging techniques such as autoradiography, magnetic resonance imaging or positron emission tomography, mass spectrometry imaging (MSI) permits the label-free
study of several compounds of interest simultaneously on the same tissue section. However, the difficulty of obtaining an absolute quantification of experimental data remains one of MSI’s major
disadvantages. Several methods are described in literature in order to address this issue, but none have universal applications. This quantitative MSI feasibility study investigates robustness and
reproducibility in whole-body imaging while taking pharmacokinetic problems into account. Using the example of a propranolol distribution study on whole-body, we report below the methodology
intended to respond to the main obstacles in quantification through MALDI (Matrix-Assisted Laser Desorption/Ionization) imaging. These difficulties are as follows: first, the high dependence of
the detected signal on the matrix deposition/properties and its extraction capacity; secondly, the MALDI ionization yield of specific target molecules; and lastly, the ion suppression effect on tissue.
Walkthrough
Materials and Methods
1. Evaluation of Tissue Extinction Coefficient (TEC)
Applications
1
2
3
Target molecule
Propranolol
BDM31343
Olanzapine
T
C
Structure
Matrix Standard
Whole-body section
2. Calibration curve determination
Samples
R²=0.9999
y=ax+b
Mouse 20min post injection Mouse 30 min post injection
Average ua
Therapeutic area
Anti-hypertension
Preparation
Mouse kidney 2 hours post
injection
Anti-tuberculosis
Sagittal cryosection (20 µm)
Anti-psychotic
Sagittal cryosection (10 µm)
Matrix
DHB
HCCA
Acquisition mode
MS
FAST-SRM
MS
Ion images
m/z 260.2
m/z 303.3→151.2
m/z 313.1
Concentration
4. Quantification
Organs
Average ua normalized
Concentration (µg/g)
Kidney
2563
3408
30.62
Liver
6385
7151
61.96
Brain
16532
26285
192.32
…
3. Drug distribution study
Average ua measured
…
…
…
MALDI MS image
Previously
Calculated TEC
Raster size
300 µm
200 µm
Instrument: MALDI-TOF Mass Spectrometer AutoFlex Speed (Bruker Daltonik GmbH, Bremen, Germany)
equipped with a Smartbeam IIT M laser with a repetition rate of 1000Hz.
y=ax+b
Example of application: Quantification of Propranolol
3. Drug distribution study
2. Calibration curve determination
1. TEC Calculation
Figure 1 : (a) Optical image of a control mouse whole-body section. (b) Distribution of
propranolol at know concentration (10 pmol/µL) mixed with matrix solution is shown. (c)
TEC values for each targeted organ are presented as histograms for brain, lung and
kidney for propranolol.
Figure 2 : (a) MS image of dilution range of propranolol ([M+H]+
ion; m/z 260.2). (b) Calibration curve obtained for propranolol
dilution range (fmol/mm2), equation, linearity coefficient (R2),
limit of detection (LOD) and quantification (LOQ) are reported.
Figure 3 : (a) Scanned optical image of 20 µm thick sagittal
whole-body section of a mouse, 20 min post injection of
propranolol. (b) Distribution of propranolol ([M+H]+ ion; m/z
260) in corresponding tissues sections.
y
Conclusion
4. Quantification
Propranolol
qMSI
Tissue
Conc. (µg/g tissue)
5.6
Kidney
17.7
Lung
10.8
Brain
% RSD
15.9%
13.2%
18.9%
QWBA[1]
Method Comparison
% RSD
Conc. (µg/g tissue)
2.1%
5.5
7.8%
19.2
5.0%
10.3
BDM31343
qMSI
Tissue
Conc. (µg/g tissue)
Lung
39.1
% RSD
12.5%
LC-MS2[2]
Conc. (µg/g tissue)
34.2
Method comparison
% RSD
12.4%
Olanzapine
qMSI
Tissue
Conc. (µg/g tissue)
Kidney
41.6
% RSD
9.3%
LC-MS2[3]
Conc. (µg/g tissue)
41.1
Method comparison
% RSD
1.1%
Table 1 : Quantification data obtained by qMSI methodology of propranolol,
BDM31343 and olanzapine compared with other techniques (liquid
chromatography or quantitative whole-body autoradiography)
www.imabiotech.com
qMSI vs others quantification techniques: Advantages and disadvantages
Tissue quantitative
techniques
Autoradiography
Preparation
Labelling
time
High
Yes
Speed
Slow
Simultaneous
Data
Distribution Metabolite
Treatement time
detection
Yes
No
Low
Tissue extraction
LC-MS²
Low
No
Fast
No
Yes
High
Horizontal sectionning
Low
No
Fast
No
Yes
High
Spectroscopic methods
High
Yes
Fast
Yes
No
Low
qMSI
Low
No
Fast
Yes
Yes
High
1. Kertesz et al, Analytical Chemistry 2008 80 (13), 5168-5177
2. Cornett et al, Analytical Chemistry 2008, 80 (13), 5648-5653
3. Data from INSERM
Contact : stauber.jonathan@imabiotech.com
qMSI Methodology:
Fast preparation
Simultaneous organ analysis
(particularly adapted to wholebody studies)
Huge set of data and long
treatment
qMSI calculation software in
development
MALDI Imaging is a drug
quantification technique:
LOD/LOQ range (ng-µg/g tissue)
Patent FR1152334
US Patent pending
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Could Mass Spectrometry Imaging be a Drug Quantification Technique?
- 1. COULD MASS SPECTROMETRY IMAGING BE A DRUG QUANTIFICATION TECHNIQUE? G. Hamm1, D. Bonnel1, R. Legouffe1, F. Pamelard1, J.-M. Delbos2, F. Bouzom2, C. Piveteau3, N. Willand3, B. Déprez3, J. Stauber1 1: ImaBiotech, Parc Eurasanté, Loos, France. 2: Technologie Servier, Orléans, France. 3: INSERM U761, Biostructures & Drug Discovery, University Lille Nord de France, France. Introduction Unlike traditional imaging techniques such as autoradiography, magnetic resonance imaging or positron emission tomography, mass spectrometry imaging (MSI) permits the label-free study of several compounds of interest simultaneously on the same tissue section. However, the difficulty of obtaining an absolute quantification of experimental data remains one of MSI’s major disadvantages. Several methods are described in literature in order to address this issue, but none have universal applications. This quantitative MSI feasibility study investigates robustness and reproducibility in whole-body imaging while taking pharmacokinetic problems into account. Using the example of a propranolol distribution study on whole-body, we report below the methodology intended to respond to the main obstacles in quantification through MALDI (Matrix-Assisted Laser Desorption/Ionization) imaging. These difficulties are as follows: first, the high dependence of the detected signal on the matrix deposition/properties and its extraction capacity; secondly, the MALDI ionization yield of specific target molecules; and lastly, the ion suppression effect on tissue. Walkthrough Materials and Methods 1. Evaluation of Tissue Extinction Coefficient (TEC) Applications 1 2 3 Target molecule Propranolol BDM31343 Olanzapine T C Structure Matrix Standard Whole-body section 2. Calibration curve determination Samples R²=0.9999 y=ax+b Mouse 20min post injection Mouse 30 min post injection Average ua Therapeutic area Anti-hypertension Preparation Mouse kidney 2 hours post injection Anti-tuberculosis Sagittal cryosection (20 µm) Anti-psychotic Sagittal cryosection (10 µm) Matrix DHB HCCA Acquisition mode MS FAST-SRM MS Ion images m/z 260.2 m/z 303.3→151.2 m/z 313.1 Concentration 4. Quantification Organs Average ua normalized Concentration (µg/g) Kidney 2563 3408 30.62 Liver 6385 7151 61.96 Brain 16532 26285 192.32 … 3. Drug distribution study Average ua measured … … … MALDI MS image Previously Calculated TEC Raster size 300 µm 200 µm Instrument: MALDI-TOF Mass Spectrometer AutoFlex Speed (Bruker Daltonik GmbH, Bremen, Germany) equipped with a Smartbeam IIT M laser with a repetition rate of 1000Hz. y=ax+b Example of application: Quantification of Propranolol 3. Drug distribution study 2. Calibration curve determination 1. TEC Calculation Figure 1 : (a) Optical image of a control mouse whole-body section. (b) Distribution of propranolol at know concentration (10 pmol/µL) mixed with matrix solution is shown. (c) TEC values for each targeted organ are presented as histograms for brain, lung and kidney for propranolol. Figure 2 : (a) MS image of dilution range of propranolol ([M+H]+ ion; m/z 260.2). (b) Calibration curve obtained for propranolol dilution range (fmol/mm2), equation, linearity coefficient (R2), limit of detection (LOD) and quantification (LOQ) are reported. Figure 3 : (a) Scanned optical image of 20 µm thick sagittal whole-body section of a mouse, 20 min post injection of propranolol. (b) Distribution of propranolol ([M+H]+ ion; m/z 260) in corresponding tissues sections. y Conclusion 4. Quantification Propranolol qMSI Tissue Conc. (µg/g tissue) 5.6 Kidney 17.7 Lung 10.8 Brain % RSD 15.9% 13.2% 18.9% QWBA[1] Method Comparison % RSD Conc. (µg/g tissue) 2.1% 5.5 7.8% 19.2 5.0% 10.3 BDM31343 qMSI Tissue Conc. (µg/g tissue) Lung 39.1 % RSD 12.5% LC-MS2[2] Conc. (µg/g tissue) 34.2 Method comparison % RSD 12.4% Olanzapine qMSI Tissue Conc. (µg/g tissue) Kidney 41.6 % RSD 9.3% LC-MS2[3] Conc. (µg/g tissue) 41.1 Method comparison % RSD 1.1% Table 1 : Quantification data obtained by qMSI methodology of propranolol, BDM31343 and olanzapine compared with other techniques (liquid chromatography or quantitative whole-body autoradiography) www.imabiotech.com qMSI vs others quantification techniques: Advantages and disadvantages Tissue quantitative techniques Autoradiography Preparation Labelling time High Yes Speed Slow Simultaneous Data Distribution Metabolite Treatement time detection Yes No Low Tissue extraction LC-MS² Low No Fast No Yes High Horizontal sectionning Low No Fast No Yes High Spectroscopic methods High Yes Fast Yes No Low qMSI Low No Fast Yes Yes High 1. Kertesz et al, Analytical Chemistry 2008 80 (13), 5168-5177 2. Cornett et al, Analytical Chemistry 2008, 80 (13), 5648-5653 3. Data from INSERM Contact : stauber.jonathan@imabiotech.com qMSI Methodology: Fast preparation Simultaneous organ analysis (particularly adapted to wholebody studies) Huge set of data and long treatment qMSI calculation software in development MALDI Imaging is a drug quantification technique: LOD/LOQ range (ng-µg/g tissue) Patent FR1152334 US Patent pending Imagine your next innovations