drug discovery & development solutions · 2020-04-09 · drug discovery & development...
TRANSCRIPT
Drug discovery & development solutions
FOR DRUG METABOLISM
DRUG METABOLISM
Fast and efficient metabolite identification is critical in today’s
drug discovery pipeline. The goal is to achieve rapid structural
identification and complete characterization of major and
low-level metabolites in a single analysis. AB SCIEX triple
quadrupole-linear ion trap mass spectrometers and accurate
mass approaches make that goal a reality.
– Qual & Quant for the answers you need: Get high-sensitivity
quantification and new qualitative capabilities on a single
platform—the AB SCIEX TripleTOF™ 5600 System and
MetabolitePilot™ Software.
– Fast and efficient low-level metabolite identification:
Acquire predictive MRM (pMRM) and multiple precursor ion
and/or neutral loss survey scans in a single analysis, including
polarity switching, with the AB SCIEX QTRAP® 5500 and
LightSight® Software.
– From early discovery to late stage development: Get the
information you need to take products from the lab to the
clinic with integrated workflows.
The AB SCIEX TripleTOF™ 5600 System and
MetabolitePilot™ Software deliver high
sensitivity quantification and new qualitative
capabilities on a single platform.
The AB SCIEX QTRAP® 5500 and LightSight®
Software use predictive MRM (pMRM) and
multiple precursor ion and/or neutral loss survey
scans in a single analysis, including polarity
switching.
AB SCIEX TripleTOF™ 5600 System
Accurate mass at the speed and sensitivity of a triple quadrupole:
• Fast MS and MS/MS acquisition speeds compatible with
fast chromatography
• Resolution over 30,000
• External mass accuracy ~1ppm
• 4 orders of Linear Dynamic Range
MetabolitePilot™ Software
Accurate mass data processing and interrogation:
• Generate cleaner, more relevant data with Multiple Mass
Defect Filtering.
• Store and retrieve critical information in Compound Library
& Results Database.
• Quickly process multiple sample sets in batches.
• Increase confidence in your results with intelligent scoring
and easy-to-visualize color-coding.
• Predict formulae with a high level of chemical intelligence.
Simple, clear metabolism workflows
AB SCIEX QTRAP® 5500 System
A single platform for drug metabolism and bioanalytical
quantification workflows:
• Targeted and non-targeted workflows
• Increased sensitivity
• Increased speed
• Full quantitative capabilities
LightSight® Software
Exploit the full functionality of QTRAP® technology and processing
strategies for multiple MetID workflows
• pMRM – high sensitivity targeted approach for really low
level detection and confirmation
• PI/NL (+/-ve polarity switching) – structure based filtering
approach ideal for reactive metabolite screening
• Multiple Ion Monitoring (MIM) & Q3 single MS strategies
for a non-targeted approach
AB SCIEX TripleTOF™ 5600 System MetabolitePilot™ Software AB SCIEX QTRAP® 5500 System Lightsight® Software
Drug discovery & development pipelineMetabolite identification is central to many of the activities in the discovery and development
pipeline. From rapid structural identification during the discovery process to provide an early
perspective on the metabolically labile sites, or soft spots of a drug candidate, to a more complete
characterization of the metabolic clearance process to maximize safety and efficacy, there’s an
ever-increasing demand on throughput.
The ability to find, identify and confirm metabolites as fast as possible is critical; not only
for the major metabolites, but the very low-level metabolites as well. Being able to do this in a
single analysis is a highly sought after goal. With improved strategies for metabolite ID from hybrid
triple quadrupole-linear ion trap mass spectrometers and an ever increasing use of accurate mass
approaches, these different but complementary mass spectrometric techniques have brought
the goal to reality.
DRUG DISCOVERY PRE-CLINICAL CLINICAL TRIALS FDA REVIEW
Average time in stage
Phase I20 To 100 volunteers
Phase III1,000 To 5,000 volunteers
Phase II100 to 500 volunteers
5 YEARS 1.5 YEARS 6 YEARS 2 YEARS 2 YEARS
LARGE-SCALE MANUFACTURING
PHASE IV
IND
SUB
MIT
TE
D
ND
A SU
BM
ITT
ED
FDAAPPROVED
DRUG
10,000COMPOUNDS 5 COMPOUNDS250 COMPOUNDS
Application snapshots within the drug discovery & development pipeline
1. EARLY DISCOVERY
Metabolic Stability
GSH Detection
3. DEVELOPMENT
Definitive Identification
Integrated Qual/Quant workflows
2. LATE STAGE DISCOVERY
In-vivo Metabolite ID
Low-level Metabolite ID
Early discovery:metabolic stability screening
C The MS/MS spectrum of Imipramine obtained automatically in IDA mode. The high
mass accuracy and resolution greatly simplify structure elucidation.
B A wide range of Imipramine phase I metabolites detected at t=5 minutes by
TOF MS scan.
A Using a generic IDA method and fast chromatography, metabolic stability profiles were
obtained in rat liver microsomes at a substrate concentration of 1 μM at t=0.
Metabolic stability assessment
Take soft-spot analysis to new levels of speed, efficiency and depth
of data with the TripleTOF™ 5600 and MetabolitePilot™ Software.
Apply completely generic methodology to acquire high quality
quantitative and qualitative data simultaneously for soft-spot
analysis. The AB SCIEX TripleTOF™ 5600 System makes it possible,
with TOF MS and MS/MS scanning at rates suitable for UPLC.
• Generate quantitative data on the parent compound from the
TOF MS scan.
• Automatically acquire MS/MS data on the major metabolites
using high-speed IDA logic.
• Use accurate mass MS/MS for structural elucidation.
t=5 min
Metabolic Stability – Rat Liver Microsomes
Detect low-level reactive metabolites in complex samples using
glutathione (GSH) detection with the AB SCIEX QTRAP® 5500
System and LightSight® Software.
The formation of undetected reactive metabolite species can
potentially cause idiosyncratic adverse drug reactions (IADRs),
which can lead to Black Box warnings and withdrawal of drugs
from the market. GSH detection is one of the most sensitive ways
of detecting low-level, transient reactive metabolites in complex
biological samples.
H3C
OO
O
OO
OH2N
NH
NH
HN
OHS
HO
H3C
OO
O
OO
OH2N
NH
NH
HN
OHS
HO
Neutral Loss m/z 129 Precursor Ion m/z 272
Early discovery: reactive metabolite detection
High sensitivity GSH adduct screening: Fast scanning and polarity switching with the QTRAP® 5500 System enable sensitive detection of GSH-trapped reactive
metabolites with a combined precursor ion (negative ion mode, 272 m/z) and neutral loss (positive ion mode, m/z 129) in a single injection. The survey scans are
specific to the GSH moiety. They do not require knowledge of parent MS/MS fragmentation, or prediction of metabolites. This provides for comprehensive detection
of GSH conjugated metabolites – whether predicted or not.
GSH Adduct Screening2 Survey scans with polarity switching in 1 injection
Total cycle time1.3 sec
+ CNL of 129
- Precursor m/z 272
+EPI m/z 632 +EPI m/z 648
at 20,000 amu/sec
Both triggered via ER scan
- Prec + ER + EPI + CNL + ER + EPI
Identify metabolites of lead drug candidates for structural
elucidation using selective detection techniques such as mass
defect filtering with the TripleTOF™ 5600 System. For the first time,
multiple mass defect filters can be applied in real time to select
peaks for MS/MS data acquisition. There is no need to schedule a
second injection to obtain MS/MS information. With the TripleTOF™
5600 you can detect and characterize major metabolites in vivo
with a single injection.
Late stage discovery: In vivo metabolite identification
A Minor oxidative metabolite (2.59 minutes) of Carbamazepine detected in protein
precipitated plasma. Although eluting just before a major metabolite, multiple mass
defect filtering enabled acquisition of MS/MS spectra of both metabolites in the
presence of strong matrix interferences.
B TOF MS scan of peak at 2.59 min in above chromatogram. Large endogenous
interferences are present in the scan. Real time mass defect filtering ensures successful
acquisition of MS/MS data for the ion of interest in the presence of high background.
C The accurate mass MS/MS spectrum of the minor oxidative metabolite obtained in
IDA mode using real time mass defect filtering.
A
B C
In vivo detection in complex matrices
Detect low levels of metabolites at clinically relevant concentrations
in vivo—even in matrices such as bile, plasma, urine and fecal
extracts—using the QTRAP® 5500 System. The predictive MRM
(pMRM) survey method on the QTRAP® 5500 System is the most
sensitive and selective single-injection strategy for identifying
metabolites in challenging matrices.
Low level in vivo metabolite
detection in protein precipitated
plasma: extracted ion
chromatograms of Carbamazepine
metabolites from a 4 mg/Kg IV
administration in rat, detected by
pMRM (176 transitions monitored
simultaneously) with the QTRAP
5500 System.
Late stage discovery: low-level metabolite ID
Metabolite characterization in lead compounds
Acquire accurate mass data for both parent and fragments in lead
compounds with the TripleTOF™ 5600 System and MetabolitePilot™
Software. An advanced collision cell design and collision energy
spread function produce rich MS/MS fragmentation in the low mass
range for structure elucidation. Rich fragmentation, high resolution
and mass accuracy enable definitive metabolite profiling.
Hydroxymidazolam in rat liver microsomes. Excellent accuracy
and resolution at the low mass range in MS/MS enables
unambiguous assignment of elemental composition.
Development: definitive in vivo metabolite ID
Metabolite quantification
Estimate metabolite concentrations, without a reference standard,
using mass extracted ion chromatograms from high-resolution
experiments using the TripleTOF™ 5600 System, MetabolitePilot™
and MultiQuant™ Software.
Quantification of metabolites and cross-species quantitative
comparisons have received greater emphasis due to the Metabolites
in Safety Testing (MIST) Guidelines. Estimating metabolite
concentrations without a reference standard, however, is challenging.
TOF MS extracted ion chromatograms of OH-Midazolam metabolites and the Midazolam
parent. All peaks are automatically integrated and labeled with area counts.Corresponding TOF MS scans for the metabolite and parent peaks in Figure 6A.
High resolution enables relative quantitation in TOF MS mode while reducing or
eliminating matrix interference.
Development: integrated qual/quant workflows
TripleTOF™ 5600 System, MetabolitePilot™ and MultiQuant™
Software meets the challenge by enabling you to compare the
response in the extracted ion chromatogram (XIC) of the metabolite
to the parent drug, with high resolution, high resolution and mass
accuracy at the low mass range, and more selectivity. Thanks to fast
scanning, you can perform these studies under UPLC.
Biotransformation studies
Acquire and process mass spectrometric and UV or
radiochromatographic data in the same experiment with the
QTRAP® 5500 System and LightSight® Software—and without
time-consuming manual procedures.
All together now: Correlation of UV and MS
data of rat liver microsomal metabolites of
Bromocriptine. The Processing Workspace of
LightSight® Software makes the correlation of UV
absorbance, pMRM, and EPI-MS/MS data easy
and intuitive.
For Research Use Only. Not for use in diagnostic procedures.
© 2011 AB SCIEX. The trademarks mentioned herein are the property of AB Sciex Pte. Ltd. or their respective owners. AB SCIEX™ is being used under license.
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