# DIHYDRODIOL DEHYDROGENASE AND PAH METABOLISM/ACTIVATION

> **NIH NIH R01** · UNIVERSITY OF PENNSYLVANIA · 1993 · $209,176

## Abstract

Dihydrodiol dehydrogenase (DD;EC 1.3.1.20) can suppress the formation of
anti-diol epoxides of polycyclic aromatic hydrocarbons (PAH), which are
ultimate carcinogens, by oxidizing their precursor transdihydrodiols to
yield PAH o-quinones.  For example, DD will catalyze the oxidation of
trans-7,8-dihydroxy-7,8-dihydrobenzoa[a]pyrene (B[a]P-diol) to yield 7,8-
dihydroxybenzo[a]pyrene which then autooxidizes to benzo[a]pyrene-7,8-
dione (BPQ.  BPQ and other PAH o-quinones produced by this enzyme form
conjugates will cellular nucleophiles, enter le minus redox-cycles and
generate presumptive superoxide anion and o-semiquinone radicals and are
cytotoxic in hepatoma cells.  BPQ can also form BPQ-deoxyguanosine (dG)
adducts with DNA.  Since PAH o-quinones may be cyto- and genotoxic, DD
may initiate a new pathway of PAH metabolism and carcinogen activation.

This pathway of PAH metabolism will be assessed in isolated rat
hepatocytes and hamster embryo fibroblasts by measuring the conversion
of [3H]-b[a]P-diol into BPQ and its conjugates.  These metabolites will
be identified by co-chromatography with authentic synthetic standard on
RP-HPLC.  To verify that DD catalyzes the formation of these metabolites
studies will be replicated in the presence of selective inhibitors for
this enzyme.  To amplify this new pathway of PAH metabolism, mammalian
expression vectors containing the cDNA for DD in the sense (pRcCMV/DD)
and anti-sense (pRcCMV/DD anti-sense) direction, will be stably
transfected into cells which metabolize PAH but display only basal levels
of DD expression (Hep-G2 and MCF7 cells).  As a result stable
transfectants should divert [3H]-B[a]P-diol into 7,8-
dihydroxybenzo[a]pyrene and BPQ.  Metabolites that arise from the
intermediate hydroquinone and quinone can then be identified.  The
metabolic fate of [1,3-3H-BPQ will also be elucidated in hepatoma cells
and isolated rat hepatocytes.

To determine if DD initiates a pathway of PAH activation, the formation
of transient superoxide anion radicals during the enzyme catalyzed
oxidation of PAH trans-dihydrodiols will be inferred by measuring oxygen
uptake and H2O2 formation.  Intermediate o-semiquinone radicals will be
spin-trapped with 5,5-dimethyl-1-pyrroline N-oxide for ESR spectroscopy.
o-Semiquinone radicals produced during the activation of PAH o-quinones
by enzymatic le minus redox-cycling will be detected by spin-
stabilization.  The genotoxicity of BPQ will be measured by incubating
hepatoma cells with [3H]-BPQ and detecting BPQ-dG adducts.  The
genotoxicity of ten unlabeled PAH o-quinones generated by DD will be
examined in cell free systems (calf thymus DNA in the presence and
absence of le minus redox-cycling), and in hepatoma cells.  The formation
of covalent adducts will be measured by [32P]-post-labeling and oxidative
DNA damage will be measured by detecting increases in 8-hydroxy-2'-dG
levels.  The mutagenicity of these PAH o-quinones will be assessed using
Salmonella tester strains (Ta...

## Key facts

- **NIH application ID:** 3178561
- **Project number:** 2R01CA039504-08
- **Recipient organization:** UNIVERSITY OF PENNSYLVANIA
- **SAM.gov UEI:** GM1XX56LEP58
- **Principal Investigator:** Trevor M Penning
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 1993
- **Award amount:** $209,176
- **Award type:** 2
- **Project period:** 1986-03-01 → 1998-03-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/3178561

## Citation

> US National Institutes of Health, RePORTER application 3178561, DIHYDRODIOL DEHYDROGENASE AND PAH METABOLISM/ACTIVATION (2R01CA039504-08). Retrieved via MORVS 2026-10-25 from https://api.morvs.ai/grant/nih/3178561. Licensed CC0.

---

*[NIH grants dataset](/datasets/nih-grants) · CC0 1.0*