|dc.description.abstract||The kinamycin antitumor antibiotics, discovered in Japan in the early 1970s as secondary metabolites of the soil bacterium Streptomyces murayamaensis, were at first believed to be derivatives of the N-cyanobenzo[b]carbazole ring system. In 1994, studies in this laboratory revealed
that the initial structural assignment was incorrect. The true structures of the kinamycins are based on
a novel diazobenzo[b]fluorene ring system, a fused 6-6-5-6 carbon skeleton bearing an unusually
stable diazo moiety along with a para-quinone and other functionalities. Additionally, this group
revised the structure of isoprekinamycin (IPK), a metabolite from Streptomyces murayamaensis
previously considered to be a fully aromatized diazobenzo[b]fluorene. IPK was shown to be an
isomeric diazobenzo[a]fluorene possessing a fused 6-5-6-6 carbon skeleton and incorporating an
ortho-quinonediazide moiety. These observations stimulated much research elsewhere in regard to the
synthesis and biological activity of these structurally novel natural products. Among the notable
discoveries in other groups was the isolation and characterization of the lomaiviticins, metabolites of
the marine bacterium Micromonospora lomaivitiensis that are dimeric diazobenzo[b]fluorene
analogues, which are even more potent than the kinamycins as anticancer and antibacterial agents.
The present project was designed to develop new synthetic methods to improve access to the
diazobenzo[b]fluorenes, with a focus on (1R,2R,3R,4S)-11-diazo-1,2,3,4,9-pentahydroxy-2-methyl-
3,4-dihydro-1H-benzo[b]fluorene-5,10-(2H,11H)-dione, also called kinamycin F. The present project
was also designed to carry out experimental and theoretical studies to gain insights into the structures
and chemical properties of kinamycins, to better understand their biological properties and to identify
how such properties might be optimized through specific structural alterations.
A synthetic study was carried out on 2-methyl-1,4-naphthoquinone as a model for a possible
biomimetic generation of the highly oxygenated D-ring of the kinamycins as found in kinamycin F.
Epoxidation of the model quinone, followed by stereoselective reduction of both keto-carbonyl
groups and ring opening of the epoxide with acetate as the nucleophile in a novel process involving
tetramethylammonium triacetoxyborohydride provided (1R*,2R*,3R*,4S*)-2-methyl-1,2,3,4-
tetrahydronaphthalene-1,2,3,4-tetraol in good yield. Comparison of the proton NMR characteristics of
the model tetrol with those of the D-ring of kinamycins led to the conclusion that kinamycin F, unlike
other kinamycins with some of their D-ring oxygen(s) bearing acyl groups, prefers a D-ring
conformation in which the hydroxyl group that is nearest the diazo group is in a pseudo-equatorial
orientation such that the C-O bond is approximately parallel with the diazo group. Ab initio molecular
orbital calculations at the RHF 6-31G level led to the conclusion, supported by experimental
measurements of diazo IR stretching frequencies, that the diazo group of kinamycin F has an
enhanced diazonium ion character in this favoured conformation. This observation is of potential
significance since the electrophilicity of the diazo group may play a role in the mode-of-action of the
kinamycins, and since there is evidence to suggest that the other kinamycins may undergo conversion
into kinamycin F in vivo before exerting their biological effect.
A strategy for applying the results of the model study to the total synthesis of kinamycin F is
disclosed. In addition, the construction of 6-hydroxy-8-methoxy-3-methyl-7,12-dioxo-7,12-
dihydrotetraphen-4-yl methanesulfonate from readily available starting materials is described and
suggestions as to how this compound might serve as a key intermediate in the biomimetic synthesis of
kinamycin F are provided. A critical analysis of this synthetic strategy to the kinamycins in contrast
with several other approaches that have been reported by other groups during the course of this thesis
research is presented. Additionally it is pointed out that this synthetic method could provide
trione as a key intermediate, which might well represent a novel analogue of the kinamycins with
potentially intrinsic anticancer and antibacterial activity of its own, since this compound possesses a
6-6-6-6 carbon skeleton containing an ortho-quinonediazide that could serve as an unique hybrid
between the 6-6-5-6 diazobenzo[b]fluorene and the 6-5-6-6 diazobenzo[a]fluorene systems.
A semi-synthetic method for generating kinamycin F from other natural kinamycins by applying
a modified Zemplen deacylation condition is reported. Electrospray mass spectrometry was employed
to identify products from interaction of kinamycin F with glutathione on a very small scale.
Kinamycin F was found to form a covalent adduct with this thiol that is ubiquitous in mammalian
cells. A discussion of the potential biological significance of this process as well as possible
interactions with other biologically important thiols in specific potential target proteins is provided.
A systematic ab initio molecular orbital analysis at the RHF 6-31 G level of the influence of
substituents in the aromatic D-ring of isoprekinamycin was also carried out. The results have led to
the suggestion of specific structural alterations that might be employed to fine tune the
electrophilicity of the diazo group, which might affect the biological activity of such compounds.
Despite the very high potency of the lomaiviticins as anticancer and antibacterial agents, progress
towards badly needed practical drugs in these areas has been frustrated by a lack of access to adequate
quantities of these complex secondary metabolites either through in vitro fermentation or total
synthesis at the moment. In the hope that a prediction of the three dimensional properties of the
lomaiviticins might inspire the design and synthesis of simpler analogues with comparable biological
activities, a systematic ab initio molecular orbital study at the RHF 6-31G level was undertaken. In
the end, predictions of the most likely conformations of lomaiviticins A and B were achieved and are
provided as potential starting points for medicinal chemists to design simpler but equally potent and
much more accessible analogues.||en