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Betulinic acid

miller rellim at MAILHOST.TCS.TULANE.EDU
Fri Jan 15 10:26:54 EST 1999

Here are a few references. Betulinic acid is an interesting triterpene
derivative, and it appears that there is some controversy regarding its
mechanism(s) of inducing cytotoxicity.

Best of results,


Bioorg Med Chem Lett 1998 Jul 7;8(13):1707-12
Synthesis of betulinic acid derivatives with activity against human
Kim DS, Pezzuto JM, Pisha E
Department of Medicinal Chemistry and Pharmacognosy (m/c 833), College of
Pharmacy, University of Illinois at Chicago 60612-7231, USA.

[Medline record in process]

Betulinic acid has been modified at C-3, C-20, and C-28 positions and the
toxicity of the derivatives has been evaluated against cultured human
melanoma (MEL-2) and human epidermoid carcinoma of the mouth (KB) cell
lines. This preliminary investigation demonstrates that simple modifications
of the parent structure of betulinic acid can produce potentially important
derivatives, which may be developed as antitumor drugs.
PMID: 9873420, UI: 99090106

Cancer Res 1998 Dec 15;58(24):5876-7
Correspondence re: S. Fulda et al., Betulinic acid triggers CD95 (Apo1/Fas)-
and p53-independent apoptosis via activation of caspases in neuroectodermal
tumors. Cancer Res., 57: 4956-4964, 1997.
Rieber M, Rieber MS
[Medline record in process]
PMID: 9865749, UI: 99081347

J Biol Chem 1998 Dec 18;273(51):33942-8
Activation of mitochondria and release of mitochondrial apoptogenic factors
by betulinic acid.
Fulda S, Scaffidi C, Susin SA, Krammer PH, Kroemer G, Peter ME, Debatin KM
University Children's Hospital, Prittwitzstrasse 43, D-89075 Ulm, Germany.
[Medline record in process]

Different classes of anticancer drugs may trigger apoptosis by acting on
different subcellular targets and by activating distinct signaling pathways.
Here, we report that betulinic acid (BetA) is a prototype cytotoxic agent
that triggers apoptosis by a direct effect on mitochondria. In isolated
mitochondria, BetA directly induces loss of transmembrane potential
independent of a benzyloxycarbonyl-Val-Ala-Asp-fluoromethyl
ketone-inhibitable caspase. This is inhibited by bongkrekic acid, an agent
that stabilizes the permeability transition pore complex. Mitochondria
undergoing BetA-induced permeability transition mediate cleavage of
caspase-8 (FLICE/MACH/Mch5) and caspase-3 (CPP32/Yama) in a cell-free
system. Soluble factors such as cytochrome c or apoptosis-inducing factor
released from BetA-treated mitochondria are sufficient for cleavage of
caspases and nuclear fragmentation. Addition of cytochrome c to cytosolic
extracts results in cleavage of caspase-3, but not of caspase-8. However,
supernatants of mitochondria, which have undergone permeability transition,
and partially purified apoptosis-inducing factor activate both caspase-8 and
caspase-3 in cytosolic extracts and suffice to activate recombinant
caspase-8. These findings show that induction of mitochondrial permeability
transition alone is sufficient to trigger the full apoptosis program and
that some cytotoxic drugs such as BetA may induce apoptosis via a direct
effect on mitochondria.
PMID: 9852046, UI: 99069377

J Nat Prod 1998 Nov;61(11):1343-7
Preparation and cytotoxic effect of ceanothic acid derivatives.
Lee SS, Chen WC, Huang CF, Su Y
School of Pharmacy, College of Medicine, National Taiwan University, and
Institute of Pharmacology, National Yang-Ming University, Taipei, Taiwan,
Republic of China.
[Medline record in process]

Six ceanothane and 1-norceanothane derivatives (1, 2, 8-11) were prepared
from ceanothic acid dibenzyl ester. These ring-A homologues of betulinic
acid exhibited cytotoxic effects. Among these, 1-decarboxy-3-oxo-ceanothic
acid (2) was found to be the most cytotoxic against OVCAR-3 and HeLa cancer
cell lines, with an IC50 of 2.8 and 6.6 &mgr;g/mL, respectively, and an IC50
of 11.3 &mgr;g/mL against normal cell line FS-5.
PMID: 9834149, UI: 99055021

Planta Med 1998 Oct;64(7):655-7
Betulinic acid inhibits aminopeptidase N activity.
Melzig MF, Bormann H

The triterpene betulinic acid inhibits the activity of aminopeptidase N (EC in a dose-dependent manner. An IC50 of 7.3 +/- 1.4 microM was
determined for betulinic acid. This inhibitory activity is higher than that
of bestatin' (IC50 = 16.9 +/- 4.1 microM), a well known inhibitor of this
enzyme. The finding supports the idea that betulinic acid acts as
anti-melanoma agent via inhibition of aminopeptidase N activity.
Publication Types:
    a.. Letter
PMID: 9810272, UI: 99027967

Cancer Res 1998 Oct 1;58(19):4453-60
Molecular ordering of apoptosis induced by anticancer drugs in neuroblastoma
Fulda S, Susin SA, Kroemer G, Debatin KM
University Children's Hospital, Ulm, Germany.

Apoptosis mediated by anticancer drugs may involve activation of
death-inducing ligand/receptor systems such as CD95 (APO-1/Fas), cleavage of
caspases, and perturbance of mitochondrial functions. We investigated the
sequence of these events in SHEP neuroblastoma cells transfected with Bcl-2
or Bcl-X(L) using two different drugs, namely, doxorubicin (Doxo), which
activates the CD95/CD95 ligand (CD95-L) system, and betulinic acid (Bet A),
which does not enhance the expression of CD95 or CD95-L and which, as shown
here, directly targets mitochondria. Apoptosis induced by both drugs was
inhibited by Bcl-2 or Bcl-X(L) overexpression or by bongkrekic acid, an
agent that stabilizes mitochondrial membrane barrier function, suggesting a
critical role for mitochondria. After Doxo treatment, enhanced CD95/CD95-L
expression and caspase-8 activation were not blocked by Bcl-2 or Bcl-X(L)
and were found in cells with a mitochondrial transmembrane potential (delta
psi(m)) that was still normal (delta psi(m)high cells). In marked contrast,
after Bet A treatment, caspase-8 activation occurred in a Bcl-2- or
Bcl-X(L)-inhibitable fashion and was confined to cells that had lost their
delta psi(m) (delta psi(m)low cells). Mitochondria from cells treated with
either Doxo or Bet A induced cleavage of both caspase-8 and caspase-3 in
cytosolic extracts. Thus, caspase-8 activation may occur upstream or
downstream of mitochondria, depending on the apoptosis-initiating stimulus.
In contrast to caspase-8, cleavage of caspase-3 or
poly(ADP-ribose)polymerase was always restricted to delta psi(m)low cells,
downstream of the Bcl-2- or Bcl-X(L)-controlled checkpoint of apoptosis.
Cytochrome c, released from mitochondria undergoing permeability transition,
activated caspase-3 but not caspase-8 in a cell-free system. However, both
caspases were activated by apoptosis-inducing factor, indicating that the
mechanism of caspase-8 activation differed from that of caspase-3
activation. Taken together, our findings demonstrate that perturbance of
mitochondrial function constitutes a central coordinating event in
drug-induced cell death.
PMID: 9766678, UI: 98438068

DNA Cell Biol 1998 May;17(5):399-406
Induction of p53 without increase in p21WAF1 in betulinic acid-mediated cell
death is preferential for human metastatic melanoma.
Rieber M, Strasberg Rieber M
IVIC, Tumor Cell Biology Laboratory, Caracas, Venezuela.
mrieber at pasteur.ivic.ve

Because betulinic acid was recently described as a melanoma-specific inducer
of apoptosis, we investigated whether this agent was comparably effective
against metastatic tumors and those in which metastatic ability and 92-kD
gelatinase activity had been decreased by introduction of a normal
chromosome 6. Human metastatic C8161 melanoma cells showed greater DNA
fragmentation and growth arrest and earlier loss of viability in response to
betulinic acid than their non-metastatic C8161/neo 6.3 counterpart. These
effects involved induction of p53 without activation of p21WAF1 and were
synergized by bromodeoxyuridine in metastatic Mel Juso, with no comparable
responses in non-metastatic Mel Juso/neo 6 cells. Our data suggest that
betulinic acid exerts its inhibitory effect partly by increasing p53 without
a comparable effect on p21WAF1.
PMID: 9628583, UI: 98290547

Bioorg Med Chem 1997 Dec;5(12):2133-43
Anti-AIDS agents--XXVII. Synthesis and anti-HIV activity of betulinic acid
and dihydrobetulinic acid derivatives.
Hashimoto F, Kashiwada Y, Cosentino LM, Chen CH, Garrett PE, Lee KH
Faculty of Pharmaceutical Sciences, Kumamoto University, Japan.

Two series of lupane-type triterpenoic acid derivatives were synthesized and
evaluated for their inhibitory activity against HIV-1 replication in acutely
infected H9 cells, based on the fact that betulinic acid (1) and
dihydrobetulinic acid (9) were identified as anti-HIV agents. Among the
derivatives, 3-O-(3',3'-dimethylsuccinyl)-betulinic acid (3) and
3-O-(3',3'-dimethylsuccinyl)-dihydrobetulinic acid (11) both demonstrated
extremely potent inhibitory activity with EC50 values of < 3.5 x 10(-4)
microM, and remarkable in vitro therapeutic index (TI) values of 20,000 and
14,000, respectively. 3-O-(3',3'-dimethylglutaryl)-betulinic acid (4)
and-dihydrobetulinic acid (12), 3-O-diglycolyl-betulinic acid (5)
and -dihydrobetulinic acid (13) and 3-O-glutaryl-betulinic acid (6) were
also potent inhibitors of HIV replication with EC50 values ranging from 0.04
to 2.3 x 10(-3) microM and TI values from 292 to 2344. In addition,
compounds 11 and 12 were also active against HIV replication in a monocyte
cell line and in peripheral blood mononuclear cells. Our in vitro assay
indicated that these compounds are not inhibitors of HIV-1 reverse
transcriptase, whereas they inhibited syncytia formation completely in a
concentration range of 20-40 micrograms/mL. However,
3-O-(2',2'-dimethylsuccinyl)-betulinic acid (2) was also found to be an
inhibitor of HIV-induced membrane fusion with an IC100 value of 20
micrograms/mL, though it displayed significantly lower anti-HIV activity
than foregoing compounds with an EC50 value of 2.7 microM and TI of 6.7.
Further study is underway to determine the mechanisms of action of these
PMID: 9459011, UI: 98120632

Cancer Res 1997 Nov 1;57(21):4956-64
Betulinic acid triggers CD95 (APO-1/Fas)- and p53-independent apoptosis via
activation of caspases in neuroectodermal tumors.
Fulda S, Friesen C, Los M, Scaffidi C, Mier W, Benedict M, Nunez G, Krammer
PH, Peter ME, Debatin KM
Division of Hematology/Oncology, University Children's Hospital, German
Cancer Research Center, Heidelberg.

Betulinic acid (BA), a melanoma-specific cytotoxic agent, induced apoptosis
in neuroectodermal tumors, such as neuroblastoma, medulloblastoma, and
Ewing's sarcoma, representing the most common solid tumors of childhood. BA
triggered an apoptosis pathway different from the one previously identified
for standard chemotherapeutic drugs. BA-induced apoptosis was independent of
CD95-ligand/receptor interaction and accumulation of wild-type p53 protein,
but it critically depended on activation of caspases (interleukin
1beta-converting enzyme/Ced-3-like proteases). FLICE/MACH (caspase-8),
considered to be an upstream protease in the caspase cascade, and the
downstream caspase CPP32/YAMA/Apopain (caspase-3) were activated, resulting
in cleavage of the prototype substrate of caspases PARP. The broad-spectrum
peptide inhibitor benzyloxycarbonyl-Val-Ala-Asp-fluoromethylketone, which
blocked cleavage of FLICE and PARP, also completely abrogated BA-triggered
apoptosis. Cleavage of caspases was preceded by disturbance of mitochondrial
membrane potential and by generation of reactive oxygen species.
Overexpression of Bcl-2 and Bcl-XL conferred resistance to BA at the level
of mitochondrial dysfunction, protease activation, and nuclear
fragmentation. This suggested that mitochondrial alterations were involved
in BA-induced activation of caspases. Furthermore, Bax and Bcl-xs, two
death-promoting proteins of the Bcl-2 family, were up-regulated following BA
treatment. Most importantly, neuroblastoma cells resistant to CD95- and
doxorubicin-mediated apoptosis were sensitive to treatment with BA,
suggesting that BA may bypass some forms of drug resistance. Because BA
exhibited significant antitumor activity on patients' derived neuroblastoma
cells ex vivo, BA may be a promising new agent for the treatment of
neuroectodermal tumors in vivo.
PMID: 9354463, UI: 98014586

Planta Med 1997 Jun;63(3):255-7
Betulinic acid: isolation from Triphyophyllum peltatum and Ancistrocladus
heyneanus, antimalarial activity, and crystal structure of the benzyl ester.
Bringmann G, Saeb W, Assi LA, Francois G, Sankara Narayanan AS, Peters K,
Peters EM
Institut fur Organische Chemie der Universitat, Wurzburg, Germany.

The known lupane-type triterpene betulinic acid (3) was isolated for the
first time from Triphyophyllum peltatum and Ancistrocladus heyneanus. It was
found to exhibit moderate to good in vitro antimalarial activity against
asexual erythrocytic stages of the human malaria parasite Plasmodium
falciparum. A first X-ray structure analysis succeeded after conversion into
its benzyl ester 4.
PMID: 9225608, UI: 97369120

Nat Med 1995 Oct;1(10):1046-51
Discovery of betulinic acid as a selective inhibitor of human melanoma that
functions by induction of apoptosis.
Pisha E, Chai H, Lee IS, Chagwedera TE, Farnsworth NR, Cordell GA, Beecher
CW, Fong HH, Kinghorn AD, Brown DM, et al
Department of Medicinal Chemistry and Pharmacognosy, College of Pharmacy,
University of Illinois at Chicago 60612, USA.

As a result of bioassay-guided fractionation, betulinic acid, a pentacyclic
triterpene, was identified as a melanoma-specific cytotoxic agent. In
follow-up studies conducted with athymic mice carrying human melanomas,
tumour growth was completely inhibited without toxicity. As judged by a
variety of cellular responses, antitumour activity was mediated by the
induction of apoptosis. Betulinic acid is inexpensive and available in
abundant supply from common natural sources, notably the bark of white birch
trees. The compound is currently undergoing preclinical development for the
treatment or prevention of malignant melanoma.
PMID: 7489361, UI: 96071635

Oncology 1991;48(1):72-6
Sterol and triterpene derivatives from plants inhibit the effects of a tumor
promoter, and sitosterol and betulinic acid inhibit tumor formation in mouse
skin two-stage carcinogenesis.
Yasukawa K, Takido M, Matsumoto T, Takeuchi M, Nakagawa S
College of Pharmacy, Nihon University, Chiba, Japan.

A single topical application of 1 microgram of 12-O-tetradecanoylphorbol-
13-acetate (TPA) to the ears of mice was shown to induce edema, and this
TPA-induced inflammation was inhibited by 4-methylsterol and triterpene
derivatives. The ED50 of these compounds against TPA-induced inflammation
was 0.1-3 mumol. Phytosterols had only slight inhibitory effects.
Furthermore, application of 5 micrograms TPA to mouse skin rapidly caused
accumulation of ornithine decarboxylase (ODC). Similarly, sitosterol and
lupane-type triterpene derivatives markedly inhibited this TPA-induced ODC
accumulation. In addition, 5 mumol betulinic acid markedly inhibited the
promoting effect of 2.5 micrograms TPA applied twice weekly on skin tumor
formation in mice initiated with 50 micrograms of
7,12-dimethylbenz[a]anthracene, and 5 mumol of sitosterol caused slight
suppression. Thus, the inhibitory effects of sterol and triterpene
derivatives on TPA-induced inflammation roughly parallelled their inhibitory
activities against tumor promotion.

PMID: 1898988, UI: 91101822

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