Translational Studies of the High Molecular Weight Polycyclic Aromatic Hydrocarbon, Dibenzo [def ...

133 Pages·2015·1.6 MB·English

AN ABSTRACT OF THE DISSERTATION OF

Erin Madeen for the degree of Doctor of Philosophy in Toxicology presented on
September 1, 2015.

Title: Translational Studies of the High Molecular Weight Polycyclic Aromatic
Hydrocarbon, Dibenzo[def,p] Chrysene; Carcinogenesis and Metabolism

Abstract approved:

______________________________________________________
David E. Williams

The objective of this work is to add to the body of translational data between high
dose animal model research and the environmentally relevant human metabolism of
the persistent pollutant dibenzo[def,p]chrysene (DBC). We furthered the knowledge
of gene/exposure interactions by determining the carcinogenesis risk based on
Cyp1b1 genotype following in utero DBC exposure in mice, finding no difference in
lung carcinogesis by genotype. Cyp1b1 wild-type mice suffered mortality due to
aggressive T-Cell Accute Lymphoblastic Leukemia (T-ALL), while Cyp1b1 null and
CYP1B1 transgenic mice lack susceptibility to T-ALL. We also determined a link for
Cyp1b1 status and fertility status, with a particular vulnerability in Cyp1b1 wild-type
male mice fetally exposed to DBC. Female mice were found to have depleted
primordial and primary follicle reserves in both Cyp1b1 wild-type and Cyp1b1 null
mice. Further research will determine if the extent of depletion is similar across
genotypes.
In addition, sensitive accelerator mass spectrometry (AMS) was utilized to detect the
metabolism of DBC (and BaP) by human volunteers following environmentally
relevant exposures. The human PK of DBC validated rodent based PB/PK models
generated to represent human metabolism for risk assessment. DBC appeared quickly
in plasma, with the majority eliminated quickly over 24 hours and a slower
elimination period to 72 hours. The PD of DBC provided evidence that DBC is
rapidly biotransformed to metabolites circulating in plasma. The products of three or
more bio-transformations (tetrols and beyond) were not present in the plasma,
indicating that they are quickly converted to conjugated species and eliminated from
the plasma compartment. Urine contained primarily DBC metabolites that had
undergone two transformations or more (diols, tetrols, and a putative dione). The
majority of these species exist as conjugated metabolites in urine.

This body of work furthers the field of PAH translation by addressing disease and
disease prevention endpoints from PAH exposure in rodent laboratory research
models. Additionally, we are addressing human risk by removing the translation
element and directly measuring PAH metabolism in humans from the oral route of
exposure by taking a FDA IND “phase 0” approach to pharmacokinetics and
pharmacodynamics.
©Copyright by Erin Madeen
September 1, 2015
All Rights Reserved
Translational Studies of the High Molecular Weight Polycyclic Aromatic
Hydrocarbon, Dibenzo[ def,p] Chrysene; Carcinogenesis and Metabolism

by
Erin Madeen

A DISSERTATION

submitted to

Oregon State University

in partial fulfillment of
the requirements for the
degree of

Doctor of Philosophy

Presented September 1, 2015
Commencement June 2016
Doctor of Philosophy dissertation of Erin Madeen presented on September 1, 2015

APPROVED:

Major Professor, representing Toxicology

Head of the Department of Environmental and Molecular Toxicology

Dean of the Graduate School

I understand that my dissertation will become part of the permanent collection of
Oregon State University libraries. My signature below authorizes release of my
dissertation to any reader upon request.

Erin Madeen, Author
ACKNOWLEDGEMENTS

It has been a privilege for which I am very grateful to have been trainee of the
Superfund Research Program (SRP). The support, resources, training, and
opportunities that the SRP provides to students have been invaluable. The awareness
of global environmental pollution and research to address that pollution has been
inspiring and has helped me keep focus during the more challenging periods of my
graduate career.

The unconditional support of Dr. David Williams, the most laid back PI a student
could ask for, has provided me the freedom to explore and pursue projects that
interest me as I have found my niche in the broad field of toxicology. All AMS
analyses was completed on site at Lawrence Livermore National Laboratory with
continuous support and guidance from Teressa Castro and Drs. Ken Turteltaub and
Graham Bench. LNNL analysis was under the mentorship of Dr. Ted Ognibene, who
deserves sainthood for his patience and grace. Expert pharmacokinetic/
pharmacodynamic training and assistance has been provided by Drs. Rick Corley and
Mark Christensen. Histomorphological and reproductive toxicology training were
provided by Drs. Christiane Löhr and Ulrike Luderer, both of whom have spent
countless hours bouncing ideas, in person training, and providing mentorship and
counsel.
My graduate committee, including Drs. William Baird, Christiane Löhr, Emily Ho,
Ramesh Sagili, and David Williams, provided support and mentorship as well as
much needed reality checks on pertinent matters such as time line and the need for
focus. I’m grateful for the advice and recommendations that they have provided.

The open door/open lab layout of the Linus Pauling Institute has provided an
excellent training and working environment. Dr. Balz Frei was quick to provide help
and references. Lisbeth Siddens has provided detailed technical training and
friendship. Tammie McQuistan and Drs. Sharon Krueger and Pat Iverson made the
challenge of FDA IND compliance possible and successful. It was a pleasure to work
with Dr. Tod Harper as my go-to “How do I do this?” reference on all things. Our lab
super students Hannah You, David Sampson, Alex Van Scoyk, and Jessica Hummel
thought that they were getting training and mentorship, but in reality, they probably
provided me with more training and experience than they received in return.

My professional training would not have been possible without the grounding and
perspective provided by the freaks and misfits that I’ve called friends. It has been said
that “When the going gets weird, the weird turn professional” [1]. It has been
comforting to be surrounded by so many passionate professionals who have
maintained a childlike enthusiasm and a wonder for the natural world as it exists
outside of Microsoft Office Suite ® ©.
CONTRIBUTION OF AUTHORS

Chapter 2: Christiane Löhr provided detailed histomorphological analysis. Hannah
You provided mouse husbandry, generated cDNA data, and helped with necropsy and
tissue collections. Lisbeth E. Siddens helped with mouse gavage, tissue collections,
study design, and Cyp1b1 expression analysis. Sharon K. Krueger provided study
design, experimental animal generation, and ensuring compliance. Roderick H.
Dashwood and Emily Ho provided study design support. Frank J. Gonzalez generated
the Cyp1b1 null and CYP1B1 humanized mouse lines. William M. Baird provided
biochemical advice on DBC metabolism. Lisa Bramer provided detailed statistical
analysis of data from study endpoints. Katrina M. Waters provided assistance with
interpretation of statistical data. David E. Williams assisted with manuscript writing,
study design and technical advice.

Chapter 3: Richard A. Corley and Susan Crowell provided training and advice on
pharmacokinetic analysis. Kenneth Turteltaub, Ted Ognibene, and Mike Malfatti
provided AMS analysis. Tammie J. McQuistan provided sample preparation support.
Mary Garrard provided clinical coordination and clinical nursing support. Dan
Sudakin provided medical supervision as a licensed medical doctor. David E.
Williams provided study design, technical advice, and manuscript preparation
assistance.
Chapter 4: Ted Ognibene provided AMS analysis and data analysis. Rick Corley
provided pharmacokinetic assistance. Mark Christensen provided pharmacodynamics
training and analysis. Tammie McQuistan provided sample preparation assistance.
Ken Turteltaub and David E. Williams provided study design and project support.