lapchak csmc stroke bioassay facility
TRANSCRIPT
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BIOASSAY FACILITY FOR STROKE
THERAPY DEVELOPMENT AND DE-
RISKING FOR IND/NDA
Paul A. Lapchak, Ph.D.
Director of Translational Research
Professor- Department of Neurology & Neurosurgery
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Barbara and Marvin Davis
Research Building Advanced Health Sciences Pavilion
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PLEASE CONTACT US IF YOU REQUIRE
ASSISTANCE TO DEVELOP A STROKE
THERAPY OR DE-RISK YOUR CURRENT
CANDIDATE
EMAIL: [email protected]
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PLEASE CONTACT US IF YOU WANT TO
DONATE FUNDS TO SUPPORT OUR ONGOING
RESEARCH TO HELP TREAT DEVASTATED
STROKE VICTIMS
EMAIL: [email protected]
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New publications:
1) A cost-effective rabbit embolic stroke bioassay: insight into the
development of acute ischemic stroke therapy.
Lapchak PA.Transl Stroke Res. 2015 Apr;6(2):99-103.
2) A blinded, randomized study of l-arginine in small clot embolized
rabbits. Lapchak PA, Daley JT, Boitano PD.
Exp Neurol. 2015 Feb 20;266C:143-146.
3) Fast neuroprotection (fast-NPRX) for acute ischemic stroke
victims: the time for treatment is now.
Lapchak PA. Transl Stroke Res. 2013 Dec;4(6):704-9.
4) RIGOR guidelines: escalating STAIR and STEPS for effective
translational research. Lapchak PA, Zhang JH, Noble-Haeusslein LJ.
Transl Stroke Res. 2013 Jun;4(3):279-85.
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CEDARS-SINAI MEDICAL CENTER STROKE SCREENING FACILITY
SCREENING IN 2 SPECIES USING MULTIPLE MODELS &
MULTIPLE ENPOINTS
1) RABBIT SMALL CLOT EMBOLIC STROKE
MODEL
2) RAT MCAO MODEL
MRI SCANS if required
PK Analysis conducted in 2 species
Toxicity Analysis (Blood chemistry) conducted in 2 species
Safety (hemorrhage-morbidity-mortality) conducted in 2
species
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ALL CSMC STUDIES ARE CONDUCTED
ACCORDING TO CURRENT RIGOR GUIDELINES:
Lapchak PA. Recommendations and practices to
optimize stroke therapy: Developing effective
translational research programs. Stroke.
2013;44:841-843
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YOUNG MALE AND/OR FEMALE SUBJECTS
MATURE MALE AND/OR FEMALE SUBJECTS
AGED MALE AND/OR FEMALE SUBJECTS
- BEHAVIORAL ENDPOINTS USING QUANTAL
ANALYSIS (HETEROGENEOUS POPULATION
ANALYSIS)
- HISTOCHEMICAL ENDPOINTS FOR INFARCT
VOLUME AND HEMORRHAGE INCIDENCE (SAFETY
OF YOUR DRUG/DEVICE)
RABBIT SMALL CLOT EMBOLIC STROKE DRUG DEVELOPMENT MODEL
REFERENCE: Lapchak PA. Translational stroke research using a rabbit embolic stroke model:
A correlative analysis hypothesis for novel therapy development. Transl Stroke Res. 2010;1:96-107
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RAT MCAO MODEL W/QUANTAL ANALYSIS
YOUNG MALE/FEMALE
AGED MALE/FEMALE
RAT MCAO DRUG SCREENING MODEL
REFERENCE: Lyden, P.D., et al., Synergistic combinatorial stroke therapy: A quantal
bioassay of a GABA agonist and a glutamate antagonist. Exp Neurol, 2000. 163(2): p. 477-89.
ENDPOINTS: BEHAVIOR USING QUANTAL
ANALYSIS AND INFARCT VOLUME USING TTC
STAINING
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Pathophysiological Measures Using the Rabbit Embolic Stroke Model
Cerebral Blood Flow
Cortical Surface
Infarcts or Ischemic areas
TTC- 2,3,5-triphenyltetrazolium chloride stain
Brain Energy Metabolism
ATP Levels (Mitochondrial Function)
Behavioral Analysis
Quantal Curves
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MODELING AN EMBOLIC STROKE
• Common carotid artery ligated, catheter inserted retrograde
into common carotid artery- external carotid ligated
• Result- internal carotid artery access to brain
• Blood clots (100-250 µm) are injected through the catheter
into cerebral vasculature in unanesthetized rabbits
• Embolization causes numerous infarcts & ischemic cores
Result: Altered pathophysiology, cerebral blood flow and
behavioral deficits
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TTC Analysis: Distribution of Infarcts
Ischemia (Infarct cores) are heterogeneous
— Standard TTC staining- TTC taken up by mitochondria and converted
to RED color- White areas are ischemic tissue.
Lapchak, Translational stroke research: from target selection to clinical trials;
Lapchak & Zhang, eds. Springer, New York, NY. USA
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CORTICAL ATP CONTENT MEASURED IN ISCHEMIC TISSUE 3 HOURS FOLLOWING EMBOLIZATION
Lapchak PA, De Taboada L. Brain Res. 1306: 100-105 (2010).
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Clinical Rating Scores in Embolized Rabbits
Quantal Analysis Uses a Dichotomous Rating Scale
Quantal Curves are based upon S-shaped sigmoid logistics curve
Quantal assay determines how a wide stroke population responds
to a treatment
Neurological Endpoints rated are Motor Functions
Ataxia (Lack of coordination of voluntary muscles )
Leaning & Circling (Loss of balance)
Paraplegia (Total loss of hind limb function)
Death is included as an endpoint (<5% incidence)
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THROMBOLYTIC PHARMACOLOGY
Doppler analysis of MCA blood flow. The arrow points to the cessation of MCA blood flow when
a blood clot occludes or blocks the MCA. This is reversed by tPA administration.
Lapchak EOID 11(11):1623-32 (2002).
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THROMBOLYTIC EFFECTS IN EMBOLIZED RABBITS:
tPA vs. Vehicle
A positive control used in every drug screening study
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THROMBOLYTIC SIDE EFFECTS
In the process of thrombolysis, matrix metalloproteinase enzymes (MMP’s)
are activated
Intracerebral hemorrhage (ICH)
In stroke patients ICH incidence up to 6.5%
In embolized rabbits ICH incidence varies depending upon the model
Lapchak and Han, Brain Res. 1303, 144-150 (2009).
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CONCLUSION
• The RSCEM was originally used to develop tPA and remains
an effective model to predict drug efficacy prior to initiating
expensive clinical trials.
• Based upon correlative analysis, it is hypothesized that the
RSCEM can be used as an effective translational tool for the
development of stroke treatments.
• There is an apparent therapeutic window correlation of 2.43 -
3 hours between the RSCEM and AIS patients.
• The RSCEM should be used as a “GOLD” standard
translational guide in order to gauge the clinical potential of
new treatments.
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Rodent intraluminal suture model
A standardized rodent intraluminal suture ischemia model with multiple
behavioral endpoints to detect deficits/improvement in motor function and
memory. The model is suitable for both neuroprotection and
neurorestoration studies.
1) Foot Fault (w/video)
2) Adhesive test (w/video)
3) Cylinder test (w/video)
4) Rotarod
5) Rotometer
6) Catwalk
7) Garcia Neuroscore Test
8) Bederson Test
9) Barnes maze (w/video)
10) Morris water maze (w/video)
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Rat quantal bioassay
Increase in ischemia duration correlates with improved behavior when
drug X is administered post-stroke
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Post-mortem Analysis
Post-mortem analysis can include
TTC infarct volume
H&E
Immunocytochemistry
Western blot analysis
RNA analysis by PCR
Customized endpoints to suit your therapy development needs
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Left side of panel- Vehicle treated
Right side of panel- Drug X-treated
Drug X improved behavior and decreased TTC
infarct volume
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• Confirmation of drug activity in a second
model in a second species
• Direct measurement of behavior when
a therapy is administered post stroke
• Direct measurement of infarct volume
• Confirmation external to your own facility
for target and efficacy validation
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PLEASE CONTACT ME IF YOU REQUIRE
ASSISTANCE TO DE-RISK YOUR NEXT
ATTEMPT TO DEVELOP A STROKE
THERAPY
OR
IF YOU WANT TO DONATE FUNDS TO
SUPPORT OUR RESEARCH TO HELP TREAT
DEVASTATED STROKE VICTIMS
DIRECT CONTACT
EMAIL: [email protected]