Defining Aerosol Dose in the Pharmaceutical and Environmental Space: Practical Impacts on Risk Asses

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Defining Aerosol Dose in the Pharmaceutical and
Environmental Space Practical Impacts on Risk
Assessment and Treatment Paradigms in Animals and
Man

• Matthew D. Reed, PhD, DABTDirector of
  Preclinical Drug Development
• Lovelace Respiratory Research Institute
• Albuquerque, NM

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Presentation Overview

• Laying the groundwork. Inhalation particle
  exposure.
• Animal to human continuum (a lungs a lung right?)
  in environmental risk assessment and drug
  development.
• Tidal breathing and the Tidal Equation for
  aerosols
• Environmental Exposures (risk assessment
  general public or occupational and toxicology)
• Drug Development
• Efficacy (Pharmaceutical Nebulizers )
• Regulatory Safety Assessment of Pharmaceuticals
• Forced Maneuvers and Pharmaceutical Inhalers

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Presentation Overview Continued

• Inhalation Constants (what we always measure)
• Aerosol Concentration
• Particle Size and Distribution (impacts
  deposition fraction
• Inhalation Variables
• Minute Volume
• Literature
• Empirical
• Determining Deposition Fraction (in the tidal
  equation and forced maneuvers)
• Literature
• modeling
• Empirical (gamma scintigraphy)

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Inhalation Aerosol Exposure

• Aerosol
• Dispersion of a liquid in a gas or a solid in a
  gas.
• In general, a fine mist or spray which contains
  minute particles

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Inhalation Aerosol Exposure

• Occupational
• Coal Dust
• Silica
• Environmental
• Air Pollution (man made)
• Source Emission Derived
• Volcanic Eruption

In general tidal breathing situations.
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Medicinal Delivery of Aerosols

• Activation Energy
• Breath (DPI)
• Mechanical/ compressed air (DPI)
• Pressure release (MDI)
• Compressed air (nebulizer)
• Vibration (nebulizer)

Applied to..
Aerosol for Inhalation

• Formulation
• Solution
• Suspension
• dry particle/s

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Lungs a lung right??????
Well not really. We have to remember that
the human to animal continuum is not clear cut
when we design our experiments and interpret data.
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Inhalation Exposure Systems Environmental ,
occupational, chemical, NTP, etc.
Whole-body exposure systems
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DIESEL EMISSIONS
2000 Cummins 5.9L ISB 6 cyl. Turbo D-2
Certification Fuel (370 ppm S, 29
aromatics) Shell Rotella-T 15W-40 crankcase
oil Standard exhaust system with muffler Repeated
heavy-duty certification cycle Cold start
excluded Diluted with carbon- and HEPA-filtered
air Exposed at 1000, 300, 100, 30, 0 µg PM/m3
(dilutions 110 to 1300)
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HARDWOOD SMOKE

• Uncertified heating stove (Pineridge, 2 ft2)
• Room scaled to size and air conditioned to absorb
  heat
• Oak from Missouri at 20 moisture
• Flue/chimney 15 ft (4.6 m) with constant draft
  conditions
• 3-phase burn cycle (kindling, high, low)
• Exposed at 1000, 300, 100, 30 µg PM/m3
• (dilutions 1300 to 19000)

PM Concentration in High Level Chamber During Two
Days
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GASOLINE EMISSIONS
1996 4.3 L General Motors V-6 engines
In-use Chevrolet S-10 pickup trucks
Mid-range mileage (40-70k miles) Normal
condition and emissions (California) Unified
Driving Cycle 3-phase cycle mapped from
chassis dynamometer and modified for continuous
use on engine stand Used 2 engines for 2
cold starts/day Gasoline blended to 2002 U.S.
national average regular unleaded No added
oxygenates Reid vapor pressure 10.3 psia 275
ppm sulfur, 30 aromatics Pennzoil 10W-30
multi-grade oil AC Delco Duraguard oil
filters Dilutions 121 ( PM), 201, and 1101

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SIMULATED DOWNWIND COAL EMISSIONS
Specifications for the atmosphere
Low-sulfur sub-bituminous coal - electric furnace
Sulfate to ash PM ratio ? 1001 Carbon
content of ash ? 5-10 SO2SO4 molar
ratio ? 11 N species (NOy)
22 NO, 61 NO2, 17 HNO3 Total SNOy molar
ratio ? 21 Disregard O3 other secondary
reaction products Added Sulfate, SO2, and NOx
Exposed at 1000 ( PM), 300, and 100 PM/m3
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Inhalation Exposure Pharmaceutical
Rodent nose-only and dog face-mask/ sling
exposure systems.
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Rodent nose-only exposure system flow past type
-Nebulizer -Dry Powder -MDI
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Large animal exposure system head dome
-Nebulizer -Dry Powder -MDI
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How do we determine dose under normal
conditions? Environmental , Occupational,
Medicinal

• Tidal Breathing Normal breathing conditions
  where..

Dose (C x T x M x F) ? W C drug
concentration (mg/L) T duration of exposure
(min/day) M minute volume (L/min) F
pulmonary deposition factor (dependent on
particle size) W body weight (kg)
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Defining Exposure to Humans and Animals is
similar

• Determining Aerosol Concentrations
• Filter collections (real time as indicator)
• Impinger sampling
• Integrated to volume of air passing through (flow
  and collection time) filter or impinger
• Human environmental or occupational settings
• Isolated field sampling
• Used in epidemiology to investigate PM and health
• Personal monitors
• Similar use with isolated populations

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Analyses of components..
Many times this is one of only two variables
actually measured in inhalation studies.

• Equipment
• HPLC with radio, UV, fluorescence, diode array,
  etc.
• MS
• LC/MS/MS
• Unique equipment for atmospheric analyses
• ELISA development and validation

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How do we determine dose under normal
conditions? Environmental , Occupational,
Medicinal, and Safety

• Tidal Breathing Normal breathing conditions
  where..

Dose (C x T x M x F) ? W C drug
concentration (mg/L) T duration of exposure
(min/day) M minute volume (L/min) F
pulmonary deposition factor (dependent on
particle size) W body weight (kg)
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Minute Volume

• Literature
• Most contemporary Calculated based on
  relationship to body mass. RMV 0.499BW0.809,
  where BW is average exposure day body weight in
  kilograms (Bide, RW, Armour SJ, and Yee E,
  Allometric respiration/body mass data for animals
  to be used for estimates of inhalation toxicity
  to young adult humans, Journal of Applied
  Toxicology, Vol. 20, 273-290, 2000). 
• Preclinical Association of Inhalation
  Toxicologists (AIT) working party recommendation
  for standard delivered dose calculation and
  expression in non-clinical aerosol inhalation
  toxicology studies with pharmaceuticals.
  Alexander et al., Inhal Toxicol. 2008
  Oct20(13)1179-89.

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Empirical Minute Volume
Poiseuilles law.  Constants Llength of
pipe rradius ?viscosity of the air,
etc.   The pneumotach works by giving you a
delta P, and you can either solve for Q (flow),
or just accept that the linear relationship will
allow you to construct a 4- or 5-point curve with
known flows.
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Empirical Minute Volume
Empirical Assessment of RVM.   The
pneumotach works by discerning delta P. You can
either solve for Q (flow), or just accept that
the linear relationship will allow you to
construct a 4- or 5-point curve with known flows.
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How do we determine dose under normal
conditions? Environmental , Occupational,
Medicinal

• Tidal Breathing Normal breathing conditions
  where..

Dose (C x T x M x F) ? W C drug
concentration (mg/L) T duration of exposure
(min/day) M minute volume (L/min) F
pulmonary deposition factor (dependent on
particle size) W body weight (kg)
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Estimation of Deposited Dose
Particle size drives dose to ALL areas of the
respiratory tract.
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Aerosol Deposition Based on Particle Size

• Particle-size characterization
• Andersen cascade impactor
• Marple/Miller cascade impactor
• Lovelace multi-jet cascade impactor
• Aerodynamic Particle Spectrometer (TSI Inc.)
• NGI (Next Generation Impactor)

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Pulmonary Deposition Fraction

• Literature
• Most contemporary comparison Comparative
  deposition of inhaled aerosols in experimental
  animals and humans a review. RB Schlesinger. J
  Toxicol Environ Health. 198515(2)197-214.
  Focuses on nasal AND mouth breathing
• Mouth breathing generally generates greater
  pulmonary deposition
• Modeling
• Most contemporary human and rodent (rat) .The
  MPPD modeling software (v1.1 CIIT/RIVM Anjilvel
  and Asgharian, 1995 ).

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MPPD Estimates Comparing Rodents and Humans
(tidal breathing).
Percentage deposition of 1-µm particle in the
lung of a rat breathing under normal conditions.
Head nasal cavity, throat, and larynx TB
tracheal-bronchial region P pulmonary (deep
lung region).
Percentage deposition of 1-µm particle in the
lung of a human breathing under normal
conditions. Head nasal cavity, throat, and
larynx TB tracheal-bronchial region P
pulmonary (deep lung region).
http//www.thehamner.org/
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Pulmonary Deposition Fraction Regulatory FDA GLP

• Accepted dogma for deposition in regulatory
  safety studies at particle sizes of 1-5 microns.
• Large animals (dogs and NHPs) 25 pulmonary (deep
  lung) deposition fraction
• Rodents 10 pulmonary (deep lung) deposition
• Humans assumed to 100 deposition
• These values are used for setting safety factors
  for safe starting doses in human clinical trials.

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Empirical Deposition Imaging

• Gamma Scintigraphy
• Requires Labeled Particle Usually technetium 99
• Generally used for medicinal purposes
• Highly valuable for use as a tool to study
  deposition in normal and diseased animal/ humans
• Requires specialized equipment
• Gamma camera
• Small bore gamma camera (preclinical)

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LRRI Primary Research Foci
Rodent SPECT/CT images following nose-only
inhalation of Tech-99m radiolabeled particles.
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Empirical deposition using labeled polydispersed
aerosols in rodents by gamma SPECT/CT
Rat and Mouse Deposition Fraction as a Function
of Particle Size
Notice the differential between regulatory dogma
and data.
McDonald et al., 2009 American Thoracic Society
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Empirical deposition using labeled polydispersed
aerosols in rodents by gamma scintigraphy.
30 Deep Lung
7 Deep Lung
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Kuehl et al., 2008 Respiratory Drug Delivery,
Cheng et al., 2009
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What about dose in non-tidal situations? Forced
maneuver.

• Pharmaceutical in natures generally involving
  Forced Inspiratory Volume (FIV).
• Human use generally MDI or DPI.
• May involve a breath hold.
• Dose dependent on..
• Concentration (emitted dose)
• Particle size
• Plume velocity (important in MDI)
• Patient training (breath hold)

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LRRI SPECT Gamma Camera
Used for human Phase I and large animals (dog,
primate and rabbit).
Technetium or other radiolabel colloidally
bound to drug
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Gamma Scintigraphy Depicting Deposition Pattern
of 1 and 3.2 µm Particles (mouth breathing with
breath hold)
CFC-BDP
HFA-BDP
Note lack of upper respiratory deposition
Note lack of upper respiratory deposition
and overall deposition.
4 Lung 94 Oral
59 Lung 31 Oral
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Impact of particle size on dose Primate SPECT
Images.
1 micron aerosol
3.2 micron aerosol
Data generated by C. Leach.
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Preclinical Forced Maneuver Bolus
Forced Inspiratory Volume Similar to FEV

• Robust (chemical and heat resistance facilitates
  sterilization)
• Efficiently ejects test article (typically 80,
  as high as 94)
• Efficiently Delivers test article (typically gt60
  of what is ejected)
• Used to deliver small amounts (1-5 mg) of dry
  powder test article via endotracheal tube to the
  deep lung of a test species

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Impact of disease on dose

• Animal to human continuum needs to be considered
  when considering normal animals and diseased
  animals
• 1-3 microns is not always ideal especially when
  discerning efficacy endpoints in diseased models
• Impact of aerosol size can impact infectious
  disease course and outcome.
• Latter may also effect treatment potential with
  inhaled therapies AND vaccines.

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Inflammation of the Rat Lung after LPS Exposure
(1 mg)
(0.5 mg)
Inflammation is more homogenously distributed in
lungs exposed to LPS aerosols compared to LPS
delivered by intratracheal instillation.
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Emphysema
Expansion of Airspaces Distal to the Terminal
Bronchioles Due to Destruction of Alveolar Walls

• Normal
• Regularly sized alveoli
• Continuous alveolar septa
• Normal elastic recoil

• Emphysematous
• Fragmented alveolar septa
• Irregularly enlarged air spaces
• Loss of elasticity

CS-exposed
FA control
(Cigarette smoke CS-induced emphysema in the
centriacinar region of A/J mice)
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Orthotopic Nude Rat Model
Tumor developmentIrradiation, instillation with
20 x 106 Calu-6 (anaplastic carcinoma derived)
suspended in EDTA solution. Tumors were allowed
to grow 9 wks prior to treatment
Example A549 xenografts in nude rats. Top
control Bottom treated with C75 weekly X3
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Inhalation concepts overlap defense,
environmental, and drug development research.
Characterizing exposure is a critical overriding
theme.
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Parting Thoughts

• There is more to inhaled dose than meets the eye.
• Inhalation concepts are extremely important when
  considering infection, progression of disease,
  therapeutic options, treatment paradigms and risk
  assessment.
• I love inhalation..

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Key Contacts

• Matt Reed, Director of Preclinical Drug
  Development
• 505-348-9451 mreed_at_LRRI.org
• Bill Bechtold, Business Development
• 505-348-9456 wbechtol_at_LRRI.org
• Jake McDonald, Director Chemistry and Exposure
  Operations
• 505-348-9455 jmcdonal_at_LRRI.org

Lovelace Respiratory Research Institute 2425
Ridgecrest Dr. SE Albuquerque, NM 87108
www.LRRI.org
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http//ap.google.com/article/ALeqM5jzFNdB08IY4qoF
Tzjv37XOMtSPqAD93KD9DG0
http//www.sandiapeak.com/