InsideScientific is an online educational environment designed for life science researchers. Our goal is to aid in the sharing and

distribution of scientific information regarding innovative technologies, protocols, research tools and laboratory services.

Today’s Presenters:

R. Dustan Sarazan, DVM, PhD

Vice President & CSO, Data Sciences International

Steve Fox, BS

Associate Principal Scientist, Merck & Co., Inc.

Anna Honko, PhD

Staff Scientist, NIH/NIAID Integrated

Research Facility

years – 1984 to 2014

Integrated Physiologic Monitoring for Life Science Research

DSI is a pioneering research company focused on systems physiology and pharmacology. We are celebrating 30 years of advancing the field of physiologic monitoring this year. Over the past 30 years, we’ve delivered proven solutions that

have been used to monitor more than 500,000 research subjects and study findings have been published in over 1000 peer-reviewed journal articles. DSI’s 200 scientists, engineers, and personnel are based out of St. Paul, MN, but we also

have offices in Europe and Asia where sales and technical support staff are located to help our customers abroad.

Benefits of Telemetry

1. Freely moving, conscious animals

2. Eliminates exit site infections

3. Chronically instrumented animals can be used sequentially as their own controls, or in several studies, thus improving animal welfare

4. Decreases costs

5. Enhances safety

Additional Considerations

• Number of animals

• Physiologic endpoints of interest

• Study duration

• Interventions

• Data collection schedule

• Data analysis

Utilizing Continuous EEG in Drug Discovery: From Mouse to Man

Steve Fox Associate Principal Scientist

Pharmacology - EEG Merck Research Laboratories

Copyright S. Fox, Data Sciences International, and InsideScientific. All Rights Reserved.

Challenges of Drug Discovery for CNS Indications

a Nature Reviews Drug Discovery 11, 191-200 (March 2012)

Increasing POS of Drug Candidates:

1. Expose at the target site of action over a desired period of time

2. Binding to the pharmacological target as expected for its mode of action

3. Expression of pharmacological activity commensurate with the demonstrated target exposure and target binding

For a development candidate to have potential to elicit the desired effect over the necessary period of time, three fundamental elements need to be demonstrated:

CNS Biomarkers

Body

Brain

Functional Brain Area

Cortex

Mid - Brain Nuclei

ms Second Minute Hour Day Week Months Years

PET

CSF PK

MRI

Genetic Traits

Leve

l of A

nato

mic

Acc

urac

y

Cognition/Mood Space

EEG

EEG Spans a Large Temporal Space

Measure Pharmaco-Dynamic? Pharmaco-Kinetic? Comment:

PET

No

Yes

- Expensive, patient location locked to scanner

MRI

Yes

No

- Expensive, patient location locked to scanner

Genetics No No - Possibly response predictive

EEG

Yes

Yes

- Patient is location and time independent - Spans large temporal and anatomic space - Inexpensive, portable, scaffold for other clinical measures

Symphony in the Brain • Tuba section • Trombone section • French horn section • Trumpet section

EEG • Delta activity (0.5 - 4 Hz) • Theta activity (4 - 8 Hz) • Alpha activity (8 -13 Hz) • Beta activity (14 - 26 Hz)

Beta

Active Cognition

Alpha

Relaxed Wake

Delta

Deep Sleep

Theta

REM Sleep

In Humans: Recording Brain Activity is Done Readily Using External Measurements

Sanna et al. 2004, Int J Sports Med; 25: 457-460

Circadian Physiology

Moore-Ede et al., 1983 NEJM

Sleep is Assessed Using Broadly Useful Measures (EEG, EMG, EOG)

Including: • ECG • cognition • mood • respiration • activity • body temperature • vocalization

& more…

Polysomnography

Sleep is Assessed Using Broadly Useful Measures (EEG, EMG, EOG)

Including: • ECG • cognition • mood • respiration • activity • body temperature • vocalization

& more… NEJM 1974, 290(9) 487

Sleep is Translational by Nature…

Pharmacological Confidence in EEG

1. MRL Sleep / EEG Lab Translation • Rely on natural translation of sleep & EEG

• Employ multiple species (mice, rats, dogs, monkeys & man)

• Add therapeutic area specific measures

2. Sleep & qEEG as Biomarkers • EEG is time scalable

• Dynamic frequencies and transients

• Topography

3. Combine EEG, Cognition & PK • 3-D drug signature

• PK/PD and PD/PD estimates

4. Automation to Drive Probability of Success • Data collection is easy – data analysis is hard

• Alignment of pre-clinical & clinical end points

EEG patterns are conserved among mice, rats, dogs, NHP and humans

- this approach is readily translatable among species…

Ideal Profile for Insomnia Drug:

- reduce sleep latency (sleep onset) and sleep fragmentation (maintenance), improve next-day performance, non-addictive

EEG Translational Strategy for Drug Discovery

• Similar study designs across species

• Control for environmental conditions

• Standardize and optimize analytic approaches

CNS Drug Discovery Improved POS Higher POS Optimal POS Minimal POS

Patients

Human PhI

Rat

Effic

acy,

PK

Patients

Human PhI

Canine

Rat

Mouse (TG)

NHP

Slee

p / q

EEG

/ Ph

arm

aco-

EEG

Effic

acy,

PK

, Cog

nitio

n, T

emp,

EC

G, A

ctiv

ity

Patients

Human PhI

Rat

Mouse (TG)

Slee

p / q

EEG

/ Ph

arm

aco-

EEG

Effic

acy,

PK

Patients

Human PhI

Rat

Mouse (TG)

Slee

p / q

EEG

/ Ph

arm

aco-

EEG

Effic

acy,

PK

, Cog

nitio

n, T

emp,

EC

G, A

ctiv

ity

1. Minimize stress, surgeries, possibilities for infection

2. Maximize predictive ability, animal health, and model durability

3. Measure continuously over multiple circadian cycles/ drug administrations

4. Support discovery and development programs using automated workflows and algorithms

Pre-clinical EEG methods are similar to human but are optimized to:

Measuring Sleep in Pre-Clinical Species

DSI Implant

F20/F40 EET

Quad-ET

D70-EEE

Standard Analytic Approaches for Pre-Clinical Studies

EEG Recording V/t

∆ φ α β γ

Arousal State Average Across Time

Spectral Distribution Spectral Band Average Across Time

Sleep Scoring

FFT Analysis

Compound L

• EEG + EMG + Activity

• Polysomnography

• Quantitative EEG (qEEG) – Fast Fourier Transform (FFT)

• Group frequencies by bands (Delta, Theta, Alpha,…)

• Time course of drug effect

Rat Sleep as High-Throughput PK/PD Screen

• Sleep/EEG signatures of CNS compounds

• Focuses target selection, lead optimization, and biomarker assessment

• Wireless implants allow for additional measures in home cages

Paroxetine (Paxil©)

Buproprion (Wellbutrin©)

Buspirone (Buspar©)

High-Throughput EEG Screen for Seizure Liability

• WAG-Rij rats genetically susceptible to absence-like seizures

• EEG evaluates brain penetrance and thalamic calcium channel target engagement

• Automated scoring reduces cycle time while maintaining accuracy 0%

20%

40%

60%

80%

100%

0 2 4 6 8 10 12 14 16 18 20 22

Perc

ent C

umul

ativ

e Se

izur

e Ti

me

(%)

Untreated (n=32)

200mpk (8)400mpk (8)

Dosed

50mpk (8)

ACTIVE IN-ACTIVE ACTIVE

Hours After Dosing

0%

20%

40%

60%

80%

100%

0 2 4 6 8 10 12 14 16 18 20 22

Perc

ent C

umul

ativ

e Se

izur

e Ti

me

(%)

Untreated (n=32)

0%

20%

40%

60%

80%

100%

0 2 4 6 8 10 12 14 16 18 20 22

Perc

ent C

umul

ativ

e Se

izur

e Ti

me

(%)

Untreated (n=32)

200mpk (8)200mpk (8)400mpk (8)400mpk (8)

Dosed

50mpk (8)50mpk (8)50mpk (8)

ACTIVE IN-ACTIVE ACTIVE

Hours After Dosing

5 sec

seizure

Automatically scored seizure

Ethosuximide

Summary

= statistically significant effects 0.05, 0.01, 0.001

0 2 4 6 8 10 12 14 16

OlanzapineVehicle

Active Wake

Light Sleep

Delta Sleep

REM Sleep

Hours

Dur

atio

n (M

in)

Olanzapine• 10 mpk, PO• Vehicle- 0.5% Methylcellulose• Analysis of all nights • Dose Time=3:30• Time 0 = 3:00pm• n=8 rats & nights: 56

Study details

-30369

1215

3

6

9

12

15

5

10

15

20

25

0

2

4

6

8

1. 8 hour decrease in active wake starting immediately post dose

2. Increase in light sleep for 2 hours

3. 10.5 hour increase in delta sleep for starting 3 hours post dose

4. 1 hour increase in REM sleep immediately post dose; 5 hour decrease in REM starting 2 hours post dose

Summary

= statistically significant effects 0.05, 0.01, 0.001

0 2 4 6 8 10 12 14 160 2 4 6 8 10 12 14 16

OlanzapineVehicleOlanzapineVehicle

Active Wake

Light Sleep

Delta Sleep

REM Sleep

Hours

Dur

atio

n (M

in)

Olanzapine• 10 mpk, PO• Vehicle- 0.5% Methylcellulose• Analysis of all nights • Dose Time=3:30• Time 0 = 3:00pm• n=8 rats & nights: 56

Study details

-30369

1215

3

6

9

12

15

5

10

15

20

25

0

2

4

6

8

1. 8 hour decrease in active wake starting immediately post dose

2. Increase in light sleep for 2 hours

3. 10.5 hour increase in delta sleep for starting 3 hours post dose

4. 1 hour increase in REM sleep immediately post dose; 5 hour decrease in REM starting 2 hours post dose

Rat Sleep as Adverse Event Screen

• Sleep, EEG, & cognition are also adverse event detection tools

• Seizures also detected automatically if needed

• Zyprexa causes “excessive sedation” when given PO to rats

Rat Sleep Study: Olanzapine

Ancillary Measures Using Quad-ET in Rats

• Acute and chronic changes in SC body temperature and heart rate

• Indicative of adverse effects, though insomnia drugs may reduce body temperature and heart rate to sleep-phase levels

• Activity data not shown, but is also analyzed

Heart Rate SC Body Temperature

Innovative Research

• Flexibility to design and run interesting studies

• Sleep Disruption

• Lights-on, Sounds

• Develop new translatable analysis

• Combine EEG + Cognition for residual sleep effects / arousability

0

2

4

6

0

5

10

15

0

5

10

15

-0.10

0.10.20.30.4

0

10

20

30

Dur

atio

n (M

in)

REM Sleep

Delta Sleep II

Delta Sleep I

Active Wake

Quiet Wake

REM Sleep

Delta Sleep II

Delta Sleep I

Active Wake

Quiet Wake

0 2 4 6 8 10 12 14 16 18 20 220 2 4 6 8 10 12 14 16 18 20 22

Sleep Disruption

Human “Home Cage” EEG

Recursive Vertical Integration

• PD/PK across species using common PD/PK endpoints

• Improve next-generation drug quality

• Decrease clinical discovery time

Cellular Assays (Specificity, Potency)

Rodent Assays (Safety, EEG)

Monkey Assays (EEG, Cognition)

Identify Specific Signals

Identify Efficacy (Success per Year)

2MM

3000

200

3-5

Human Assays (EEG, Cognition)

+ ID

+ ID

+ ID

30

1 FDA Approved!

Can the flow of medicines be improved? Fundamental pharmacokinetic and pharmacological principles toward improving Phase II survival

Sleep Lab Translational Strategy…

Paul Morgan, Piet H. Van Der Graaf, John Arrowsmith, Doug E. Feltner, Kira S. Drummond, Craig D. Wegner and Steve D.A. Street

Drug Discover Today – Volume 17, Number 9/10 – May 2012

EEG as a Model for CNS Drug Discovery

1. Sleep & qEEG as biomarkers for CNS penetration and vertical translation to patients

2. EEG is high throughput & highly translational 3. Multi-species for optimal human predictions

• Sleep • qEEG • Cognition • Aging

4. Ambulatory PSG/EEG • FIM • CNS Penetration • Dose selection

5. Database building • CNS target specific EEG signals • PK/PD & PD/PD across species

Patients

Human

Non-Human Primate

Canine

Rat

Mouse

Telemetry for Biodefense Research Applications

Anna Honko, PhD Staff Scientist

NIH/NIAID Integrated Research Facility

Frederick, MD

Copyright A. Honko, Data Sciences International, and InsideScientific. All Rights Reserved.

What will we cover today…

1. Introduce what makes biodefense research unique from other pharm/tox or drug discovery applications in the context of disease models

2. Discuss some of the different study designs where telemetry can be utilized

Biosafety Level 3 (BSL-3)

• Agents associated with serious or lethal human disease for which preventive or therapeutic interventions may be available

• Personnel may have to change into scrubs before entering the laboratory and take a complete shower before exiting

• Other personal protective equipment may be required as well, depending on the tasks to be performed

CDC Biosafety in Microbiological and Biomedical Laboratories (BMBL) 5th Edition

Biosafety Level 4 (BSL-4)

• BSL-4 is required for work with dangerous and exotic agents that pose a high risk of aerosol-transmitted laboratory infections and life-threatening disease that is frequently fatal, for which there are no approved vaccines or treatments

CDC Biosafety in Microbiological and Biomedical Laboratories (BMBL) 5th Edition

BSL-4 viral agents:

Ebola virus Marburg virus

Smallpox (Variola virus)

Lassa Fever Virus Junin Virus

Crimean-Congo hemorrhagic

fever virus

BSL-3 bacterial agents:

Yersinia pestis Bacillus

anthracis Francisella tularensis

Mycobacterium tuberculosis

BSL-3 viral agents:

Venezuelan equine encephalitis virus

Eastern equine encephalitis virus Western equine

encephalitis virus SARS coronavirus Yellow Fever Virus

West Nile Virus Chikungunya

Rift Valley Fever Virus

Due to engineering design requirements of laboratory, telemetry system reliability is critical… 1. Only persons whose presence in the facility or individual laboratory

rooms is required for scientific or support purposes are authorized to enter

2. Equipment must be decontaminated using an effective and validated method before repair, maintenance, or removal from the laboratory

This means… • Only able to install or make major repairs when laboratory is decontaminated, or • Personnel must be able to install or make minor adjustments if necessary • Telemetry device sterilization/decontamination

[Either one-time use or decontaminate and replace/exchange (factor into costs)]

CDC Biosafety in Microbiological and Biomedical Laboratories (BMBL) 5th Edition

Sequence of Entry Rooms

• Since there is frequently a physical separation between the telemetry base station and the animals being monitored, communication is critical…

Biodefense research and the FDA Animal Rule

1. Non-traditional regulatory pathway for new drug approval when human efficacy studies are not ethical or feasible

2. The Animal Rule states that FDA will rely on evidence from animal studies to provide substantial evidence of effectiveness only when the following four criteria are met: 1. There is a reasonably well-understood pathophysiological mechanism of the

toxicity of the substance and its prevention or substantial reduction by the product;

2. The effect is demonstrated in more than one animal species expected to react with a response predictive for humans, unless the effect is demonstrated in a single animal species that represents a sufficiently well-characterized animal model for predicting the response in humans;

3. The animal study endpoint is clearly related to the desired benefit in humans, generally the enhancement of survival or prevention of major morbidity; and

4. The data or information on the kinetics and pharmacodynamics of the product or other relevant data or information, in animals and humans, allows selection of an effective dose in humans. Paraphrased from “Guidance for Industry, Product

Development Under the Animal Rule, May 2014”

3. Adequate and well-controlled animal efficacy studies are required for approval under the Animal Rule = well-documented and GLP if possible

Purpose of natural history studies for rare diseases

From a presentation by Anne R. Pariser, M.D. , former Associate Director for Rare Diseases, CDER, FDA:

• “The natural course of a disease from the time immediately prior to its inception, progressing through its presymptomatic phase and different clinical stages to the point where it has ended and the patient is either cured, chronically disabled or dead without external intervention”1

There is limited natural history data in humans for high containment diseases for a number of reasons:

• Sporadic nature of the outbreaks and the remote locations of the cases, availability of resources at sites

• Variability in dose, route of exposure and time to presentation for individual cases

1Posada de la Paz M; Groft SC. 2010. Rare diseases epidemiology. Vol. 686

Benefits of natural disease progression studies in non-human primates using telemetry

1. Studies are observational and longitudinal, allowing for

the characterization of the pathophysiology of the disease in a more controlled system

• Does the disease in nonhuman primates model the disease in humans?

• Are we able to identify potential triggers for therapeutic intervention, biomarkers or correlates of disease?

Benefits of natural disease progression studies in non-human primates using telemetry

2. Some disease manifestations that are common in human cases, such as shortness of breath or fever, are difficult to objectively measure in nonhuman primates cageside • Telemetry is considered a “refined technique” for collecting this data

3. Additionally, if mortality is used as a primary endpoint, considerations must be made for the effects of euthanasia criteria • Real-time monitoring using telemetry has provided an additional means

of objective euthanasia criteria, reducing observational study bias

• Increasing sample integrity since terminal sample collection can occur

Study designs that benefit from telemetry data acquisition

1. Longitudinal natural history studies and basic pathophysiology modeling studies

2. Comparison of nonhuman primate species

3. Agent dose-seeking studies

4. Identification / application of therapeutic interventions, establishment of euthanasia criteria

5. Efficacy trials of potential vaccine or therapeutic

Longitudinal natural history studies and basic pathophysiology modeling studies

Characterization of disease progression differences between fatal and non-fatal viral infection

Non-survivor Survivor

1

Longitudinal natural history studies and basic pathophysiology modeling studies 1

Ability to match physiological parameters by telemetry with commonly collected clinical parameters

Plasma RNA levels Clinical serum chemistry

Febrile response of macaques following EEE virus exposure

Longitudinal natural history studies and basic pathophysiology modeling studies 1

Comparisons of viral disease in different NHP models

Comparison of Non-human Primate Species 2

Comparison of febrile responses between dose groups of aerosol exposed macaques (WEE)

Low Dose High Dose

Agent Dose-Seeking Studies 3

Trigger to initiate therapeutic intervention • Results of natural disease modeling studies can be used to determine triggers for treatment

• In combination with RT-PCR assessment, fever as calculated by ≥1.5°C above pre-challenge baseline using remote telemetry monitoring was used to determine starting point for mAb treatment for Ebola

Identification / application of therapeutic interventions, establishment of euthanasia criteria 4

Comparison of vaccine efficacy: Sham-immunized controls challenged against EEEV

Efficacy trials of potential vaccine or therapeutic 5

Comparison of vaccine efficacy: SIN-SAEEEV vaccinated (nonprotective – 17% survival)

Efficacy trials of potential vaccine or therapeutic 5

Comparison of vaccine efficacy: SIN/NAEEEV vaccinated (significant protection – 82% survival)

Efficacy trials of potential vaccine or therapeutic 5

Summary

Real-time monitoring of physiological parameters using less-invasive implanted telemetry devices will provide critical data for:

1. Development of animal models for biodefense or emerging diseases

2. Evaluation of potential vaccines and therapeutics

3. Establishment of objective criteria for therapeutic interventions or endpoints

Thank You! For additional information on telemetry applications and various DSI solutions for continuous monitoring of physiology in pre-clinical research studies please visit:

www.datasci.com

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InsideScientific is an online educational environment designed for life science researchers. Our goal is to aid in the sharing and

distribution of scientific information regarding innovative technologies, protocols, research tools and laboratory services.