making translational research happen
TRANSCRIPT
Making Translational Research Happen
Wadsworth Center
New York State Department of Health
Division of Environmental Health Sciences
Kenneth M. Aldous, Ph.D.
Funding Approaches
•Collaboration with Commercial Partner (NIH SBIR)
• Instrumentation development
• Prototype building collaboration with partner R&D staff
• Optimization of digital electronics and software
• Incentive to produce rugged and commercial product
• Develop instrument for RO1 funded study.
•R21 Research Funding for assay development.
• Work performed in-house with PHL staff
• Using existing instrumentation and PHL cores
• Develop pilot data for subsequent funding
Small Business Innovative Research (SBIR)
•Phase I. The objective of Phase I is to establish the technical merit, feasibility and potential for commercialization of the proposed R/R&D efforts and to determine the quality of performance of the small business awardee organization prior to providing further Federal support in Phase II. Support normally may not exceed the following:
•$150,000 total costs for 6 months
•Phase II. The objective of Phase II is to continue the R/R&D efforts initiated in Phase I. Only Phase I awardees are eligible for a Phase II award. Support normally may not exceed:
•$1,000,000 total costs for 2 years
R21 Application• Application Characteristics
• You may request a project period of up to two years
• The combined budget for direct costs for the two year project period may not exceed $275,000. No more than $200,000 may be requested in any single year.
• The R21 can not be renewed
• One resubmission (A1) is allowed
• Introduction required for a resubmission is limited to one page.
• No preliminary data are required but may be included if available.
• The Research Strategy may not exceed 6 pages.
• Scope
• Exploratory, novel studies that break new ground or extend previous discoveries toward new directions or applications.
• High risk high reward studies that may lead to a breakthrough in a particular area, or result in novel techniques, agents, methodologies, models or applications that will impact biomedical, behavioral, or clinical research.
• Projects should be distinct from those supported through the traditional R01 mechanism.
Funding Development
•Background
• Novel X-Ray Optical Systems
• XRF, Monochromatic XRF, Doubly Curved Crystal Optics
•Lead in children‟s toys and consumer products
• CPSC, commercial handheld XRF, XOS HD-Prime
• Archived toy samples and reference materials
•Assessing personal exposure to toxic metals
• NIH funding for new technologies for assessing environmental
exposures
• XOS analyzer for toxic elements in environmental samples
• Public health response to „unusual‟ sources of metal exposure
X-Ray Optical Systems
• Local industry collaborator on this
research
• Leader in development and manufacture
of x-ray optics
• Producer of XRF instrumentation
• Sindie for sulfur in petroleum fuels
• HD-Prime for lead in toys
• Interest in developing new
technologies for environmental
applications
• 3 prototype instruments based on DCC
optics for specific public health problems
• Funded by 2 NIH SBIR grants with
Wadsworth
East Greenbush, NY
XOS Body Fluids AnalyzerX-Beam®
X-Ray Fluorescence Spectrometry
www.nitonuk.co.uk/ images/xrfinfo.jpg
Monochromatic XRF (M-XRF)
17.5 keV
7 keV
31 keV
50 W
Mo K
Advantages
• Reduced background
provides significantly
improved P/B
• No loss in intensity
due to scatter in filter,
so low power source
can be used
• Small spot sizes can
be produced
Monochromatic XRF (M-XRF)
sample
holder
detector
(SDD)
XYZ stage
DCC optic
x-ray tube
45
Energy resolution:
~140 eV at 5.9 keV
X-Beam®:
Integrates source and
DCC into unit with
forced air thermal
management to
ensure proper
alignment
Lead in children‟s toys and
consumer products
Mattel Recalls 19 Million Toys Sent From China
August 2007
Nearly 1 Million Fisher-Price Toys Recalled Because of Lead Paint
XOS HD-Prime
Dimensions: 36x26x32 inches Weight: 240 lbs
cameras
sample chamber
Calibration modes:
Plastic, metal, wood,
glass, rubber, leather,
textiles, bulk paint
For painted products:
1) Measure painted region
2) Measure substrate
≤7 minutes
1 mm spot
size
Specific objectives- lead in toys
•Assess and compare the performance of new XRF
instrumentation designed for the determination of lead in
toys
• At regulatory limits, which were recently reduced
• For bulk substrate materials and paint films
• Use archived toy samples, and CRM/RM as available
•Characterize the advantages and limitations of the XRF
approach compared to conventional methods
•Evaluate the use of area concentration (µg/cm2) versus
mass fraction (mg/kg) for assessing toy safety
Lead in toys
•Consumer Product Safety Improvement Act (CPSIA) of 2008• New regulations for products intended for children <12 years old
2 µg/cm2≠
CPSIA
Thermo Scientific Niton XL3t
•Purchased in 2009 to
evaluate side-by-side
with M-XRF
•Calibration modes
•Metals (%)
•Soil (mg/kg)
•Filters (µg/filter)
•Plastics (mg/kg)
•Painted products (µg/cm2)
•Reports >30 elementsDimensions: 9x9x4 inches Weight: <3 lbs
2 minutes
Reference methods for Pb in paint
Remove paint from substrate
Filter and bring to volume
Analyze with Flame Atomic Absorption Spectrometry
>2 hours
Microwave
assisted heating
Conclusions
•Determination of lead content in toys is a very difficult
analytical problem• Primary advantage of XRF is ability to test small areas and the substrate,
which are difficult with FAAS
•Both XRF instruments can be used for quantitative
assessment of lead in bulk substrates <100 mg/kg limit
•The XL3t is of limited use for determination of lead in
paint, given false negatives for non-PVC substrates
•Though the HD-Prime is approved for certifying
compliance and reported no false negatives in mg/kg
units, quantitation at 90 mg/kg is not reliable
•The CPSC should reduce the 2 µg/cm2 limit
Assessing personal exposure to metals
NIH R01 Application to investigate the application of
XRF in epidemiologic exposure studies.
Environmental sampling
• Important for some public health studies as it helps identify possible sources of exposure• Toxic elements are of most interest
•Use of environmental sampling is relatively limited• Lack of sensitive field-portable instrumentation
• Cost associated with laboratory analysis
•Questionnaires used as a substitute but don‟t give a reliable picture of exposure pathways to multiple elements • Epidemiologists want a large N at low cost
Environmental exposure
NIH Genes, Environment, and
Health Initiative (GEI)
Focus on interaction of environmental exposures
and genetic factors to better understand how these
contribute to disease; seeking new environmental
monitoring technologies
Specific objectives- environmental samples
•Participate in the development of an XRF
instrument intended for field measurement of
samples from the personal environment
•Evaluate performance in a laboratory setting, with
a focus on Pb, As, Hg, Cd
•Sample preparation requirements
•Application to „unusual‟ samples
•Comparison to existing field-portable XRF technology
Phases of PEA
α-prototype β-prototypes
Personal Environmental Analyzer (PEA)
Feasibility study
Qualitative assessment with SRM
• NIST SRM 1547 Peach Leaves (powder)
• Detected all elements with certified mass fractions >1 mg/kg, except those outside measureable range (e.g., B, Na, Mg)
0
20
40
60
80
100
120
140
160
180
1 3 5 7 9 11 13
Energy (keV)
Sq
rt (
co
un
ts)
K Ca
Ca
Ba V
Mn
Fe
FeCu
Zn
Zn Pb/As
Rb
Sr
Al P Cl
Br
S
(Ar)
32 W, 300 s
Phases of PEA
α-prototype β-prototypes
Personal Environmental Analyzer (PEA)
Feasibility study Laboratory validation study
Preparation for field evaluation
Instrumental detection limits
ISO/ICPAC 3SD, 10 measurements of aqueous solutions
Samples of public health interest
•Origin
• NYC DOHMH (n=35)
• NJ DHSS (n=3)
•Types
• Herbal medicine products (HMP)
• Ethnic spices
• Personal products
•Many implicated in lead
poisoning cases
• All but one product imported
• Banned for sale in NYC
Prescribed by a NYS-licensed acupuncturist for joint
pain; contains nearly 5% Pb by weight, 46-year old
male with BLL of 91 µg/dL
Results for selected samples
Lead0
100
200
300ICP-OES
FAAS
PEA
XL3t
ma
ss fr
actio
n (m
g/k
g)
Mercury0
10
20
30
40
ma
ss fra
ctio
n (m
g/k
g)
Error bars (XRF) ±SD, n=3, (ICP-OES & FAAS) ±10%, n=1
Chili powder
Excellent agreement
between FAAS and ICP-OES
(2% difference)
Screening in bag,
PEA 130±9
30% bias
Skin lightening cream
Both identify
low-level Hg!
Screening of routine samples
•HMP submitted as part of lead-poisoning investigation
•PEA• 0.3% Pb by weight
• 6% Hg by weight
• ICP-OES• 0.2% Pb by weight
• 15% Hg by weight
• Identified possible additional health risk (<5 min)• Physician notified
2 3 4 5 6 7 8 9 10 11 12 13 14 150
500
1000
1500
2000 Fe
HgHg
Pb
Energy (keV)
Inte
nsity
(cp
s)
Conclusions
•Accuracy and precision for PEA were reasonable (≤20%)
for most CRM
• Metal calibration mode needs development
•Though quantitation was poor, the PEA (and XL3t) were
able to correctly identify the presence of toxic elements in
real world samples
• Consider use of a major, minor, and trace classification for results
in field study
•Advantages of PEA
• Liquid calibration mode
• Detects toxic elements at lower mass fractions that XL3t
Summary
•Rapid (≤10 minutes) elemental characterization of:
• Quantitative
• Metal and plastic substrates (Pb)
• Semi-quantitative
• Paint films (Pb)
• HMPs, personal products (Pb, Hg, As, Cd)
• Qualitative
• Routine/emergency unknowns (Pb, Hg)
•Provide information not otherwise available, which may
itself be sufficient or can aid in additional analyses
Collaborations
Wadsworth Center
Dr. Patrick Parsons
Kathryn McIntosh
X-Ray Optical Systems
Dr. Walter Gibson
David Gibson
Dr. Zewu Chen
Dr. Danhong Li
Satbir Nayar
Matthew Cusack
Alexei Vershinin
Retrospective analysis of Hydrogen Cyanide Exposure
via Mass Spectral Detection of a Unique C-terminal
Cys-SCN Human Serum Albumin Adduct
Michael J Fasco Ph.D.
Charles R Hauer, III Ph.D.
Erasmus Schneider Ph.D.
Michael Dailey MD
Kenneth M Aldous Ph.D.
Substances containing cyanide have been used as poisons for centuries. In 1792a Swedish chemist named Scheele identified cyanide as the active ingredient and may have died in a laboratory accident from cyanide poisoning.
NIEHS 1R21ES016858-01 7/16/2009 -2012
Pilot Study Funded by
National Institute of Environmental Health
Sciences (NIEHS)
Study Aims
• Develop a validated, simple, quantitative assay of CN adducted to
Human Serum Albumen (HSA).
• Measure concentrations of unbound CN and CN adducted to HSA in
human blood from a background exposed population. (Aim I)
• Measure levels in a cohort of firefighters who are at constant risk for
higher than normal exposure to CN. (Aim II)
• Determine if cumulative exposure can be measured and provide new
information on potential ways to minimize exposure. (Aim III)
Why Cyanide and not Carbon Monoxide?
•CN is 35x more toxic than CO
•Both are present as combustion products and
in smoke at fires in varying amounts.
•CN exposure is difficult to measure accurately
because of its short half life (1 hour).
•CN HSA adducts half life in blood is ~21 days.
•Do firefighters CN HSA levels accumulate as
a result of repeat responses to fires?
•Toxic composition of smoke varies from fire to fire
•Nature of the burning materials
•Temperature
•Oxygen level
•Ventilation
• Conditions of high temperature and low oxygen enhance degradation of
synthetics quickening chemical release
Historically, carbon monoxide asphyxiation has been considered
the primary cause of deaths of those overcome by smoke.
CO is the focus of gas monitoring
There is mounting evidence that hydrogen cyanide is directly
responsible for many more deaths than previously assumed
Cumulative effect with CO worse than either individually
“ Cyanide toxicity from smoke inhalation in a structural or
enclosed space fire is the most likely cause of cyanide
toxicity that EMS & fire professionals will encounter”
JEMS Communications Summer 2004
Gaseous Composition of Smoke
Narrow Range of Toxicity – almost always fatal at concentrations above 3 g per ml of blood -- <0.5 g per ml is assumed to be nontoxic.
Once in the body cyanide exists almost exclusively as the gas.
Approximately 90% of CN in blood is noncovalently (reversibly)
bound to hemoglobin in the red blood cells (RBCs); the remainder is
in the plasma compartment.
The half-life of CN in blood is less than 1 hr. CN is extensively
metabolized -- primarily to thiocyanate; other metabolites are formed
from reaction of CN ion with the disulfide bond of cystine. By the time
that blood from people potentiallly exposed to CN is drawn several
half-lives have already passed.
CYANIDE
Biomarker Tests for Cyanide Exposure
1. Measure “free” CN in blood (CDC Method using
GC/MS)
2. Measure CN adduct with Human Serum Albumen (HSA)
new method developed in this study using LC/MS/MS.
Liquid Chromatograph/MS/MS
FOX Study Sampling Kit
Kit Contents
2 Part shipping container
1 - Red Top Vacutainer
1 - Purple Top Vacutainer
Butterfly
Adapter
Alcohol Pad
Questionaire form
Logistics•Firefighter Enrollment for AIM II and AIM III
•Reading and Signing Informed Consent Forms
•Sample collection after enrollment (Baseline
questionaire).
•Sample collection at up to two Acute Exposure
Events (AEE). Questionaire for each AEE)
•Sample transportation/pickup. (Contact
Wadsworth)
•Confidentiality – Samples separated from
questionaire forms and mailed.
Potential Applications
Assessment of current procedures and protective devices,
including clothing, to safeguard firefighters and other first
responders against CN poisoning from smoke
Assessment of CN exposure to smoke inhalation from
residential and commercial fires and war zones
Determine potentially dangerous exposure in the workplace
Construction of a database capable of identifying adverse,
long term health consequences
Study Investigators
• Kenneth M. Aldous, PhD
• Ryan Bennett, MS
• Michael Fasco, PhD
• Michael Dailey, MD Medical Officer
• Ed Fitzgerald, PhD Epidemiologist
Questions?