The role of cell transport systems in medicating the toxic effects of drugs and chemicals. Mechanisms of paraquat poisoning.

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The role of cell transport systems in medicating
the toxic effects of drugs and chemicals.
Mechanisms of paraquat poisoning.

• Lewis L Smith
• Syngenta

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Toxicology

• Identification of Hazard
• Characterisation of Hazard
• Relevance to MAN
• RISK Assessment
• RISK Management

Descriptive
Research
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Mechanisms of Toxicity
Relevant to Man?
Similar to Man?
Qualitatively or quantitatively similar to Man?
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Inhibition of putrescine (1µM) accumulation into
rat lung slices by selected amino acids and their
decarboxylated derivatives
Slices of rat lung were incubated in KRP (pH 7.4)
containing 1 µM 14C)-putrescine in the presence
of various concentrations of the compounds
indicated. I50 values were determined by the
method of Ross and Krieger, each animal acting as
its own control. Results are expressed as the
mean SEM, at least 3 animals being employed per
compound studied.
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The inhibitory potencies of various triamines
against putrescine (1µM) accumulation by rat lung
slices
Slices of rat lung were incubated in KRP medium
containing putrescine (1µM) in the presence of
the triamine indicated. I50 values expressed as
the mean SEM (ngt3) derived by the method of
Ross and Krieger.
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The inhibitory potencies of methylglyoxal-bis
(guanylhydrazone) against the accumulation of
putrescine (1 (µM) by rat lung slices
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Eadie-Hofstee plot for the derivation of Km and
Vmax of the accumulation process for MGBG into
rat lung slices
.

100- 90- 80- 70- 60- 50- 40- 30- 20-
10- 0-
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0 10
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V (nmoles/g.wet wt.lung/hr) S (medium substrate
conc. (µM)
Rat lung slices were incubated at 37o in the
presence of various concentrations of 14C-MGBG.
Accumulation of 14C-MGBG(V) was determined
after 30,15 and 5 min for substrate medium
concentrations lt1µM and after 60,30 and 15 min
for substrate medium concentrations gt1µM.
Triplicates were determined at each time point
and all slices taken from the same rat. Rates of
accumulation were determined by linear regression
analysis. The results shown are from a single
experiment typical of 3 and by linear regression
analysis the estimated kinetic parameters from
this experiment were Km4.6µM. Vmax83.5nmols/g
wet wt. Lung/hr (correlation coefficient r
-0.93).
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Compounds
Structure
I50 (µM)
139.9 (50.1)
Ethidium
17.93.8
Pentamidine
Primaquine
51.8 (23.9)
Chloroquine
20.1 (11.3)
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Hanes Woolf Plot of Cystamine Uptake
.
180
VMAX 1/Slope KM Y-Intercept x VMAX
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120
s/v
.
KM 503uM
90
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60
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KM 12uM
30
0
0
100
200
300
400
500
(S) (uM)
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Possible Routes of Cystamine Metabolism
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Cystamine
Cystaldimine
GSH
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3
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Thiocysteamine
Cysteamine
GSSCys
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Hypotaurine
Taurine
-
-
NAD
NADH
1 Postulated thiol-disulphide exchange 2
Amineoxygen oxidoreductase (deaminating) 3
Spontaneous 4 Uncharacterised 5 Cysteamine
dioxygenase 6 Hypotaurine dehydrogenase
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State of 14C-Label after 30 Incubation
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Aims of Current Studies

• Holistic identification of molecular pathways
  involved in PQ-induced lung fibrosis using
  Toxicogenomics
• Develop testable hypotheses for therapeutic
  intervention

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Overview of molecular functions and
pathways associated with PQ-responsive genes
543 Paraquat-responsive genes
Gene Ontology terms (biological functions)