Insect Control: Biological and Synthetic Agents - Index of

142 4: Insect Growth- and Development-Disrupting Insecticides
Figure 14 Chemical structures of new ecdysone agonists reported in literature: (1) 3,5-di-t-butyl-4-hydroxy-N-isobutyl-benzamide
(DTBHIB) from Sumitomo; (2)8-O-acetylhrapagide from Merck Research Laboratories, Westpoint, PA, USA and (3) tetrahydroquinoline
from FMC Corporation, Princeton, NJ, USA. In (3) R ¼ halide.
(DTBHIB; Figure 14) and Elbrecht et al. (1996) at
Merck Research Laboratories, Westpoint, PA, USA,
reported the isolation of an iridoid glycoside, 8-Oacetylharpagide
(Figure 14), from Ajuga reptans.
Both these compounds were reported to induce
20E-like morphological changes in Drosophila Kc
cells, as well as competitively displace tritiated
ponasterone A from Drosophila ecdysteroid receptors
with potencies similar to that of RH-5849, the
unsubstituted bisacylhydrazine. However, the insecticidal
activity of these compounds was not
described. Attempts to replicate the results of
Mikitani (1996) using DTBHIB and analogs failed
to demonstrate that these compounds were competitive
inhibitors of tritiated ponasterone A binding to
DmEcR/DmUSP produced by in vitro transcription
and translation (Dhadialla, unpublished observations).
On the other hand, Dinan et al. (2001)
demonstrated that the results obtained by Elbrecht
et al. (1996) were due to co-purification of ecdysteroids
in their 8-O-acetylharpagide preparation.
When used as a highly purified preparation, Dinan
et al. (2001) found that 8-O-acetylharpagide was
not active as an agonist or an antagonist in
D. melanogaster BII cell bioassay, and neither did
it compete with tritiated ponasterone A for binding
to the lepidopteran ecdysteroid receptor complex
from C. fumiferana.
Finally, scientists at FMC discovered a new
tetrahydroquinoline (THQ) class of compounds
(Figure 14) that competitively displaced tritiated
ponasterone A from both dipteran (D. melanogaster)
and lepidopteran (H. virescens) EcR and USP
heterodimers (unpublished data). Interestingly, the
most active analogs of this class of compounds
bound the DmEcR/DmUSP with much higher affinity
than the HvEcR/HvUSP. This is the reverse of
what was observed with bisacylhydrazines. When
tested for ligand binding to EcR/USP proteins from
L. migratoria, B. argentifoli, and T. molitor, to
which bisacylhydrazines show no measurable
affinity, members of the THQ were found to bind
with measurable affinity (mM range; Dhadialla and
colleagues, unpublished results). Further work on
this chemistry was continued at Rohm and Haas
Company, Spring House, PA, USA, and its subsidiary,
RheoGene LLC, Malvern, PA, USA, which
resulted in the synthesis of a number of analogs
that were active in transactivating reporter genes
fused to different insect EcRs and heterodimeric
partners (L. migratoria RXR, and human RXR) in
mouse NIH3T3 cells (Michelotti et al., 2003).
The discovery of THQs with ligand binding activities
to various EcRs (Michelotti et al., 2003), some
of which do not interact with bisacylhydrazines,
and the interpretation of X-ray crystal structure
results of liganded HvEcR/HvUSP (Billas et al.,
2003) provides good evidence for the potential to
discover new chemistries with ecdysone agonist
activities. It should also be possible to design chemistries
that specifically interact with EcR/USPs
from a particular insect order.
4.2.9. Noninsecticide Applications of Nonsteroidal
Ecdysone Agonists; Gene
Switches in Animal and Plant Systems
A number of researchers started to explore the utilization
of the ecdysone receptor as an inducible gene
switch due to the knowledge that neither mammals
nor plants have ecdysone receptors, and the discovery
of bisacylhydrazine as true ecdysone agonists
with reduced risk ecotoxicology and mammalian
profiles. Gene switches are inducible gene regulation
systems that can be used to control the expression
of transgenes in cells, plants, or animals. There
is a recognized need for tightly regulated eukaryotic
molecular gene switch applications, such as for
gene therapy, and in understanding the role played