Organic compounds -- part of the class 532-570 series – Organic compounds – Heterocyclic carbon compounds containing a hetero ring...
Reexamination Certificate
1998-08-21
2001-05-01
Kight, John (Department: 1612)
Organic compounds -- part of the class 532-570 series
Organic compounds
Heterocyclic carbon compounds containing a hetero ring...
Reexamination Certificate
active
06225481
ABSTRACT:
FIELD OF THE INVENTION
The present invention relates to the robustaflavone, intermediates for preparing robustaflavone, and robustaflavone analogues as well as compositions containing the same. The present invention also relates to synthesis of robustaflavone, intermediates and analogues thereof.
BACKGROUND OF THE INVENTION
Robustaflavone, a naturally occurring biflavanoid, is a potent non-nucleoside inhibitor of hepatitis B virus (HBV) replication
1
. HBV is one of the most serious health problems in the world today, and is listed as the ninth leading cause of death by the World Health Organization
2
. Approximately 300 million persons are chronically infected with HBV worldwide, with over one million of those in the United States. The Centers for Disease Control estimates that over 300,000 new cases of acute HBV infection occurs in the United States each year, resulting in 4,000 deaths due to cirrhosis and 1,000 due to hepatocellular carcinoma
3
. The highest incidence of HBV infection occurs in the Far East and sub-Saharan Africa, where approximately 20% of the population are chronically infected
4
. Infection can be prevented through the use of several extremely effective recombinant vaccines
5
. Despite the availability of these vaccines, HBV infection remains the most significant viral pathogen infecting man, particularly in under-developed countries.
Preliminary in vitro evaluations had shown that robustaflavone possesses anti-hepatitis B activity comparable to several nucleoside analogues currently in clinical trialsl, as well as acts synergistically with these agents
22
. Robustaflavone was first
6
isolated in 1973, as its hexa-O-methyl ether, from leaf extracts of
Agathis robusta
, and later
7
in larger quantities from the seed-kernels of
Rhus succedanea
. While robustaflavone represents an important lead compound in the search for potential anti-hepatitis B agents, no total synthesis of robustaflavone has been reported to date. A recent synthesis of a related biflavanoid, amentoflavone (I8,II3′-biapigenin), which consists of two apigenin units connected via a biaryl linkage between respective 8- and 3′-positions, was achieved using Suzuki coupling
8
of an apigenin 8-boronic acid derivative with an appropriate 3′-iodoapigenin analogue
9
. However, there is no disclosure or suggestion that this approach can be extended to the total synthesis of robustaflavone.
SUMMARY OF THE INVENTION
The present invention relates to a method for preparing robustaflavone, a biologically active biflavanoid composed of two apigenin (5,7,4′-trihydroxyflavone) units connected via a biaryl linkage between their 6- and 3′-positions. The present invention also relates to robustaflavone intermediates and analogues useful as potential antiviral agents and/or for preparing other biflavanoids and method for preparing the
same. The method of the present invention includes a thallium-assisted regioselective iodination of apigenin 7,4′-dimethyl ether in the 6-position, allowing efficient access to 6-iodoapigenin trimethyl ether (8). See Scheme 2. This step allows for an efficient route to 6-halogenated flavones that has not been described previously. Another step in the method of the invention entails conversion of 3′-iodoapigenin trimethyl ether (5a) to its corresponding 3′-pinacol boronate derivative 9b, via the palladium-catalyzed cross-coupling of 5a with bis(pinacolato)diboron. See Scheme 3. The corresponding 3′-stannane 9a failed to couple with iodide 8 under Stille conditions, and attempts to convert either of iodides 5a or 8 to their corresponding boronic acids using standard methods (halogen-lithium exchange/trialkylborate quench) failed. See Scheme 3. Identification of reaction conditions which allowed Suzuki coupling between boronate 9b with iodide 8 furnished the critical 6-3′ biaryl linkage, to afford the desired robustaflavone skeleton in the form of its hexamethyl ether (10). See Scheme 4. Finally, demethylation under non-aqueous conditions using BBr
3
provided access to synthetic robustaflavone, which was identical in all respects to the natural product. See Scheme 4. The method of the present invention will allow access to needed quantities of robustaflavone for further biological studies, as well as provide an efficient method for the synthesis of intermediates and structural analogues.
Accordingly, it is one object of the invention to provide a direct method for preparing robustaflavone, intermnediates and analogues thereof.
It is another object of the invention to provide intermediates useful in preparing robustaflavone analogues and other biflavanoid compounds having linkages through the 6-position or the 3′ position such as hinokiflavane, rhusflavone and succedaneaflavone.
It is a further object of the invention to provide robustaflavone analogues useful as potential anti-viral, e.g., anti-hepatitis B virus, agents.
These and other objects of the invention will become apparent in light of the detailed description below.
DETAILED DESCRIPTION OF THE INVENTION
According to the present invention, the total synthesis of robustaflavone was approached via construction of two apigenin derivatives, one substituted in the 6-position and one substituted in the 3′-position, with groups that could be coupled using transition metal-catalyzed cross coupling methodology. The synthesis of apigenin derivatives substituted in the 3′-position is straightforward (Scheme 1), involving esterification of phloroacetophenone dimethyl ether (1) with 3-substituted p-anisoyl chlorides, such as 3-iodo-p-anisoyl chloride (2a). Rearrangement of the resulting ester (3a) to the &bgr;-diketone 4a was achieved by heating in pyridine at 100° C. in the presence of powdered KOH. Cyclization of the diketone 4a under acidic conditions provided 3′-iodoapigenin trimethyl ether (5a)
10,23
. This route was also utilized to prepare apigenin trimethyl ether (5b), starting with 1 and p-anisoyl chloride.
The preparation of apigenin derivatives substituted in the 6-position presented a more difficult challenge, as direct electrophilic substitution of apigenin ethers occurs preferentially in the 8-position. An extensive search of the chemical literature yielded only two examples of 6-halogenated apigenin derivatives; the first, described in 1939, reported 6-bromoapigenin trimethyl ether as an intermediate in a total synthesis of apigenin
11
. However, it was later determined that the position of the ring bromination had been incorrectly assigned in the original report, and that the actual intermediate was 8-bromoapigenin trimethyl ether
12
. A second route described the iodination of apigenin 7,4′-dimethyl ether with iodine in an iodic acid solution, which provided a mixture of the 6-iodo and 8-iodo derivatives, in a 1:4 ratio
13
. The desired 6-iodinated derivative was reportedly purified by fractional recrystallization, but in very poor yield.
The desired 6-iodinated species was prepared in excellent yield with almost exclusive regioselectivity by exploiting the ortho-directing capabilities of thallium(I) salts in the iodination of phenols
14
(Scheme 2). Selective demethylation of apigenin trimethyl ether (5b) in the 5-position may be accomplished with boron tribromide in an amount generally ranging between about 0.9 equiv and about 3.0 equiv, preferably about 1.1 equiv boron tribromide, to afford apigenin 7,4′-dimethyl ether (6). The reaction is generally carried out a temperature ranging between about −40° C. and about 50° C., preferably about 25° C., for a time period ranging between about 0.5 h and about 24 h, preferably about 5 h, in the presence of a suitable solvent such as methylene chloride, chloroform, benzene and toluene, preferably methylene chloride, until a thick precipitate is formed. The precipitate is then collected and recrystallized in a suitable solvent such as ethanol, methanol, ethyl acetate/hexane, preferably ethanol. If desired, other boron compounds such as boron trichloride may be used in
Flavin Michael T.
Lin Yuh-Meei
Zembower David E.
Zhang Heping
Advanced Life Sciences, Inc.
Covington Raymond
Kight John
McDonnell & Boehnen Hulbert & Berghoff
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