Organosulfur Natural Products
Chemistry:
We have launched a broad program of research focusing on the
organosulfur chemistry of natural products isolated from plants
with insecticidal, agricultural or medicinal value, and whose
organoleptic properties imply the presence of organosulfur
derivatives other than ubiquitous sulfur containing compounds
such as cysteine and methionine. Major concerns include 1)
isolation and characterization of novel potentially useful
g-glutamyl cysteines; 2) C-S lyase crystallization and structure
determination, in addition to studies of their enzymatic mechanisms;
3) an assessment of the variety of cysteine sulfoxide precursors
responsible for organosulfur headspace volatiles observed
in selected plants; 4) chemotaxonomic classification of plant
species; 5) determination of the lachrymatory principle(s)
in plants such as Petiveria alliacea; 6) thiosulfinate chemistry
and optical activity; 7) determination of formation pathways
of observed organosulfur natural products; and 8) synthesis
and characterization of observed novel compounds.
In the course of our work on organosulfur compounds in higher
plants, we have determined 1) that C-S lyase mediated ‘allium’
chemistry is quite ubiquitous, occurring in a number of plant
families that are unrelated to the genus Allium; 2) that Nectaroscordum
species may be appropriately classified as alliums, based
on observed chemistry that is analogous to that observed in
garlic and onions; 3) that the principle in P. alliacea that
has a lachrymatory effect on farm animals is the unique sulfine
thiobenzaldehyde S-oxide; 4) that P. alliacea contains not
only a C-S lyase, but also an L -F synthase that appears to
be analogous to that recently observed in onions, and which
is directly responsible for the formation of the lachrymator;
5) that a number of novel cysteine sulfoxide derivatives different
from those typically observed in garlic and onion varieties
exist in some higher plants, and that these and their C-S
lyase mediated decomposition products have antimicrobial and
anticancer activity.
Work on organosulfur natural products is ongoing, and current
studies include complete structural, kinetic and mechanistic
characterization of the C-S lyases and LF synthases, as well
as isolation and characterization of g-glutamyl dipeptide
derivatives which may have novel biological effects and mechanisms
of action. More recently, we have established a collaboration
with Dr. Gunter Siegel, Professor of Physiology at the Institut
fur Physiologie, Freie Universität Berlin, with whom
we are conducting in vivo and in vitro studies on the cardiovascular
effects of novel organosulfur natural products.
Development
of HIV-1 Nucleocapsid Protein Inhibitors:
Currently, therapeutic management of HIV infection and pathogenesis
is generally based upon administration of combination therapy
with multiple inhibitors of the HIV type 1 (HIV-1) reverse
transcriptase and protease. Although this approach has resulted
in significant suppression of viral loads in a substantial
number of patients, the long-term treatment outlook is negatively
impacted by a variety of issues including a high rate of drug
protocol violations, drug toxicity, and the persistence of
latent reservoirs of virus in long-lived populations of infected
cells. Additionally, the extreme mutability of the virus,
and the fact that fairly significant changes in the structures
of key viral enzymes do not always translate into gross inefficiencies
in their respective functions, renders combination therapy
with a particular drug protocol eventually ineffectual in
most cases.
These considerations imply that an appropriate target against
which to develop anti-HIV, and by extension, other antiretroviral
therapies, is a mutationally intolerant protein that plays
essential and diverse roles in various phases of the viral
replication cycle. A heretofore unexploited target that fits
this general profile is the nucleocapsid protein of HIV-1.
This small, very basic and highly conserved protein participates
in numerous obligate stages of the viral replication cycle,
and mutations in its primary structure have been observed
to result in the production of non-infectious virions. By
studying the structure of the nucleocapsid protein when it
is bound to other viral molecular components, we have devised
a plan to mimic these components with small organic molecules.
These molecules may ultimately serve as lead drug compounds.
To aid in the design of the small molecule nucleocapsid protein
inhibitors, we have been using a number of computational methods
and molecular modeling programs-with each enabling us to add
additional layers of refinement to our lead compound structures.
As a consequence, we have made significant progress in the
design of small molecule inhibitors, and this has enabled
us to advance to the stage where the compounds are being synthesized
in the laboratory. Completion of the syntheses will allow
us to test our ‘first generation’ inhibitors,
whose structures we will further refine as the results of
their ability to inhibit nucleocapsid protein functions are
acquired.
Bioassay Guided
Fractionation, Isolation and Structural Characterization of
Natural Products from Medicinal Plants
In principle, there are two general ways in which plants that
may have biological activity can be chosen for study. Plants
can be randomly selected and subjected to a series of biological
assays to determine their potential biological activity, or
they can chosen based upon a history of use against a particular
ailment in a society or culture. A disadvantage of the first
approach is that selected plants, although demonstrating efficacy
in a particular biological assay, may ultimately be found
to be highly toxic. This issue is, to a certain extent, circumvented
in the second approach, in that the toxicology of the herb
is generally know. Put simply, herbs that kill on ingestion
are likely not used in traditional medicine, and those that
have a long and consistent history of use against a particular
ailment have a higher probability of working (otherwise, there
would be no continued use).
We have launched a program to study natural products derived
from various plants and food crops that have been used to
treat disease in some cultures of the world. Thus, plants
that have a history of use in traditional medicine are subjected
to bioassays to determine the active component, or combination
of active components responsible for the manifested therapeutic
effects. Plants that we are examining in this way include
Croton lechleri which produces a red sap known as known as
‘dragon’s blood’, Uncaria tomentosa (also
known as cat’s claw), beta vulgaris (table beets) and
vigna unguiculata (cowpea).
Dragons’ blood is a viscous tree sap that is used extensively
by the indigenous cultures of the Amazon River basin for its
remarkable haling properties. When applied to the skin for
abrasions, cuts scratches, blisters, bites and stings, Dragon’s
blood forms a long-standing barrier possibly due to its ability
to coprecipitate with proteins or other matrix elements. In
doing so it is claimed to foster accelerated wound healing
and does so with reduced pain, inflammation and scarring.
We have found in our work that Dragon’s blood confers
benefit by suppressing the activation of sensory afferent
nerve mechanisms, which supports its ethnomedical use for
disorders characterized by neurogenic inflammation.
Cat’s claw is a vine that grows in the Peruvian Amazon
and has been used in traditional medicine to alleviate inflammation.
Ethnomedically, the bark and root of cat’s claw are
the parts of the plant that are most frequently used, and
are prepared as an aqueous extraction in hot water. Several
groups have reported a wide range of chemical substituents
in cat’s claw, although few studies have demonstrated
that administration of these isolated components exerts consistent
anti-inflammatory effects. Of the compounds that have been
isolated, the most well known are the oxindole alkaloids.
Based on in vitro experiments, it has been indicated that
oxindole alkaloids promote phagocytosis, leading to the claim
that cat’s claw has immunostimulant properties, and
also the ability to induce a lymphocyte-proliferation-regulating
factor in endothelial cells. However, these actions are difficult
to reconcile with the use of cat’s claw to treat chronic
inflammation. Hence, we have continued our efforts to evaluate
alternatives explanations for the mechanisms of action of
cat’s claw. In our own studies, we have found that cat’s
claw exhibits potent antioxidant and anti-inflammatory activity
that is independent of the oxindole content of the plant sample.
There is anecdotal evidence that components of table beets
and cowpea extracts may have anticancer activity. We have
initiated a bioassay guided fractionation study to determine
if there is scientific support for these reports.
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