Journals

Books

Sign in
Help
Brought to you by:
Biblioteca Central Universitar "Carol I"
Download PDFOpens in a new window. Article
suggestions will be shown in a dialog on return to
ScienceDirect.

Other export options

More options...

Advanced search

Article outline

Show full outline

Abstract

Keywords

1. Introduction

2. Pharmacological properties of betulin derivatives

3. Conclusions and future perspectives

Acknowledgements

References

Figures and tables

1.
2.

3.
Table 1

4.

5.

6.

7.

8.

9.
10.

11.
Table 2

12.
13.
14.
15.

ADVERTISEMENT

European Journal of Pharmaceutical Sciences

Volume 29, Issue 1, September 2006, Pages 113

Review

Pharmacological properties of the ubiquitous natural product betulin

Sami Alakurttib,
Taru Mkela,
Salme Koskimiesa,
Jari Yli-Kauhaluomab, ,
Show more

doi:10.1016/j.ejps.2006.04.006
Get rights and content

Abstract
Betulin (lup-20(29)-ene-3,28-diol) is an abundant naturally occurring triterpene and it is found predominantly
in bushes and trees forming the principal extractive (up to 30% of dry weight) of the bark of birch trees.
Presently, there is no significant use for this easily isolable compound, which makes it a potentially important
raw material for polymers and a precursor of biologically active compounds. Betulin can be easily converted to
betulinic acid, which possesses a wide spectrum of biological and pharmacological activities. Betulinic acid has
antimalarial and anti-inflammatory activities. Betulinic acid and its derivatives have especially shown anti-HIV
activity and cytotoxicity against a variety of tumor cell lines comparable to some clinically used drugs. A new
mechanism of action has been confirmed for some of the most promising anti-HIV derivatives, which makes
them potentially useful additives to the current anti-HIV therapy. Betulinic acid is specifically cytotoxic to
several tumor cell lines by inducing apoptosis in cells. Moreover, it is non-toxic up to 500 mg/kg body weight in
mice. The literature concerning derivatization of betulin for structureactivity relationship (SAR) studies and its
pharmacological properties is reviewed.

Keywords

Betulin;
Betulinic acid;
Birch bark;
Betulin derivatives;
Natural products;
Betula sp.;
Medicinal chemistry

1. Introduction
Betulin 1, lup-20(29)-ene-3,28-diol, also known as betulinol, betuline and betulinic alcohol (Fig. 1), is a
pentacyclic triterpene alcohol with a lupane skeleton. Common structural features of the lupane skeleton are its

five-membered ring E and isopropylidene group (Fig. 1). Although betulin 1 can also be isolated from other
sources in small amounts, the extractive isolation of betulin 1 on an industrial scale from birch bark waste could
be a large and feasible source of raw material. White birches are widespread in the northern latitudes of the
world, and currently there is no economically significant use for this easily isolable compound. Betulin 1can be
isolated (up to 30% dry weight) from the birch bark by extraction with high boiling hydrocarbon solvents or with
water azeotropes of alcohols (Eckerman and Ekman, 1985). The healing properties of birch bark and birch bark
extracts have been known for a long time in folk medicine. Birch bark oil (Betulae pix) has been used for skin
diseases, such as eczema and psoriasis ( Hnsel et al., 1992). Betulin 1 can be used as such or after chemical
modification as a starting compound for other useful materials and compounds, which possess various interesting
pharmacological properties. Some reviews concerning betulin 1 and particularly its biologically more active
derivatives, such as betulinic acid 2 have been published ( Cichewicz and Kouzi, 2003, Baglin et al.,
2003a, Eiznhamer and Xu, 2004 and Aiken and Chen, 2005). Herein, the literature concerning derivatization of
betulin for structureactivity relationship (SAR) studies and its pharmacological properties is reviewed.

Fig. 1.
Structures of lupane skeleton, betulin 1 and betulinic acid 2.
Figure options

2. Pharmacological properties of betulin derivatives

Betulin 1 has three positions in its structure, namely secondary hydroxy group at position C-3, primary hydroxy
group at position C-28 and alkene moiety at position C-20, where chemical modifications can be easily
performed to yield derivatives for structureactivity relationship (SAR) studies. Betulin 1 can be converted in
two steps in 75% overall yield to pharmacologically more active betulinic acid 2 (Fig. 2; Kim et al., 1997).

Fig. 2.
Preparation of betulinic acid 2 from betulin 1.
Figure options

It is clear from the chemical structure of betulin 1 that most of the betulin derivatives presented in this review are
lipophilic and thus poorly soluble in water. This may have significant role in interpretation of the results of the
bioactivity assays that have been carried out mainly in cell cultures. Observed differences in bioactivity between
different modified betulin derivatives may be explained at least partly by different distribution constants between
these analogs.
An essential component of preclinical development of a drug is the elucidation of its mammalian metabolism.
Few studies have been carried out in vitro to predict and prepare potential mammalian metabolites of betulin 1,
betulinic 2 and betulonic acid 3 by utilizing various microorganisms (Chatterjee et al., 2000, Kouzi et al.,
2000 and Akihisa et al., 2002). Betulinic acid 2 is non-toxic at doses up to 500 mg/kg body weight in mice
(Pisha et al., 1995) and its pharmacokinetics and tissue distribution in mice has also been studied ( Udeani et al.,
1999).

2.1. Antitumor activities

The antitumor activity of betulinic acid 2 and its derivatives could be important therapeutically, because these
compounds exhibit a high activity against several tumors and cancers (Table 1), while normal cells are
unaffected by betulinic acid 2, indicating selectivity (Zuco et al., 2002). Studies at the positions C-3 and C-28
demonstrate that simple chemical modifications of the parent structure of betulin 1 can produce potentially
important derivatives, which can act as antitumor drugs. Usually the most cytotoxic compounds show broad
cytotoxic activity against several different cancer cell lines. However, some of these derivatives show increased
general toxicity against normal cells too. It must be also realized that a cytotoxic compound in one cell line may
well be clearly less active or even inactive in other cell line, even within the same cancer type.
Table 1.
Betulinic acid 2 or its derivatives exhibit antitumor activity against the following types of cancer
Cancer type
References
Melanoma

Pisha et al. (1995), Fulda et al. (1997), Kim et al. (1998), Jeong et al. (1999), Selzer et al. (2000),Kim
et al. (2001), Hata et al., 2002 and Hata et al., 2003, Zuco et al. (2002), Symon et al. (2003),Sarek et
al. (2003), Tan et al. (2003), You et al. (2003), Sawada et al. (2004), Urban et al. (2004),Liu et al.
(2004), Zanon et al. (2004), Fulda et al. (2004), Fulda and Debatin (2005), Kasperczyk et al. (2005)

Lung carcinoma

Fulda et al. (1997), Zuco et al. (2002), Sarek et al. (2003), Urban et al. (2004), You et al.
(2003),Mukherjee et al. (2004b)

Colon cancer

Fulda et al. (1997), Kim et al. (2001), Baglin et al. (2003b), Sarek et al. (2003), Mukherjee et al.
(2004b), Urban et al. (2004)

Prostate carcinoma

Kim et al. (2001), Sarek et al. (2003), Mukherjee et al. (2004b), Urban et al. (2004)

Leukemia

Fulda et al. (1997), Noda et al. (1997), Deng and Snyder (2002), Hata et al. (2003), Ehrhardt et al.
(2004), Urban et al. (2004)

Ovarian carcinoma

Deng and Snyder (2002), Zuco et al. (2002), Sarek et al. (2003), Mukherjee et al. (2004b)

Endothelial carcinoma

Mukherjee et al., 2004a and Mukherjee et al., 2004b

Oral epidermoid
carcinoma

Kim et al. (1998), Jeong et al. (1999)

Glioblastoma

Fulda et al. (1999), Sarek et al. (2003), Jeremias et al. (2004), Fulda et al. (2004), Fulda and Debatin
(2005), Kasperczyk et al. (2005)

Breast cancer

Fulda et al. (1997), Sarek et al. (2003), Urban et al. (2004)

Neuroblastoma

Fulda et al. (1997), Schmidt et al. (1997), Fulda et al. (1998a), Hata et al. (2003), Fulda et al.
(2004),Fulda and Debatin (2005), Kasperczyk et al. (2005)

Medulloblastoma

Fulda et al., 1997, Fulda et al., 1999 and Fulda et al., 2004, Fulda and Debatin (2005)

Cervix carcinoma

Zuco et al. (2002)

Hepatocellular
carcinoma

Sarek et al. (2003)

Osteosarcoma

Sarek et al. (2003)

Rhabdomyosarcoma

Sarek et al. (2003)

Table options

2.1.1. Cytotoxicity and mechanisms of action of betulin and betulinic acid

Betulin 1 itself is inactive or rather inactive against several tested cancer cell lines such as melanoma (MEL-2),
epidermoid carcinoma (KB; Kim et al., 1998), leukemia (HL60, U937, K562) and neuroblastoma (GOTO and
NB-1; Hata et al., 2003). Originally betulinic acid 2 was thought to be selectively cytotoxic against melanoma
(MEL-14) by inducing apoptosis of cancer cells and it was found to be inactive against several other cancer cell
lines, such as breast (BC-1), colon (COL-2), epidermoid carcinoma (KB), prostate (LNCaP) and lung (LU-1)
cancer (Pisha et al., 1995). Subsequent studies confirmed its broader cytotoxicity against several different cancer
cell lines. It was found to be cytotoxic against all nine tested neuroblastoma cell lines (e.g. SKNSH, IMR-5,
NBL-S, NBAS5 and LAN-5; Schmidt et al., 1997). Betulinic acid 2 also triggers an apoptotic cascade in human
malignant glioma (e.g. LN-229, U87 MG and T98G) cells (Wick et al., 1999). Growth inhibition caused by
betulinic acid 2 was observed in neoplastic cell lines, such as ovarian carcinoma (A2780, OVCAR-5 and
IGROV-1), lung carcinoma (H460 and POGB) and cervix carcinoma (A431; Zuco et al., 2002). Betulinic
acid 2 effectively induced apoptosis in primary medulloblastoma (e.g. D283 Med, MHH1 and MEB1) and
glioblastoma (e.g. A172, U138MG, U373 and SK14) cells cultured from patients (Fulda et al., 1999). In vitro,
betulinic acid 2 induced specific cell death 75% in primary glioblastoma multiforme cells at substantially
higher rates (43% 9/21 of patients) than established cytotoxic drugs N,N-bis(2-chloroethyl)-N-nitrosourea
(BCNU) and vincristine (less than 5%; Jeremias et al., 2004). When betulinic acid 2 was compared for in vitro
efficiency against leukemia cells (several cell lines of common acute lymphatic leukemia, T-cell acute lymphatic

leukemia, B-cell acute lymphatic leukemia and acute myeloid leukemia) with conventionally used cytotoxic
drugs, betulinic acid 2 was more active than 9 out of 10 standard therapeutics, such as L-asparaginase,
dexamethasone, prednisolone and 6-thioguanine ( Ehrhardt et al., 2004). The pH of solid tumor tissues is
generally lower than that of normal tissues. This could be beneficial, because the cytotoxicity of betulinic
acid 2 is enhanced at low pH (6.8) in leukemia (L1210) culture medium (Noda et al., 1997).
Betulinic acid 2 has a synergistic cytotoxic effect on the growth and metastasis of melanoma cells (B16F10) in
vivo when it was used in combination with the anticancer drug vincristine. These two drugs induced cell cycle
arrest at different points and caused apoptosis of melanoma cells (Sawada et al., 2004). In combination treatment
betulinic acid 2 and TRAIL (tumor necrosis factor (TNF)-related apoptosis-inducing ligand) co-operated to
induce apoptosis in SHEP neuroblastoma cells, but not in normal human fibroblasts (Fulda et al., 2004). In the
combined treatment, betulinic acid 2 together with anticancer drug doxorubicin, VP16, taxol and actinomycin D,
co-operated to induce apoptosis in SHEP neuroblastoma cells. Betulinic acid 2 also sensitized medulloblastoma
(Daoy), glioblastoma (A172) and melanoma (Mel-Juso) cells for anticancer drug doxorubicin-induced apoptosis,
without affecting normal human fibroblasts (Fulda and Debatin, 2005). The effects of betulinic acid 2 together
with irradiation on survival and growth inhibition of several melanoma (e.g. A375, 518A2, MES20 and Neo IItr) cell lines are clearly additive, because their modes of action are probably different (Selzer et al., 2000). On the
other hand, combination of betulinic acid 2 with cisplatin showed no synergistic effects on head and neck cancer
cell lines (SCC25 and SCC9; Eder-Czembirek et al., 2005).
Apoptosis is recognized by distinct morphological changes including cell shrinkage, DNA fragmentation,
nuclear condensation, membrane blebbing and caspase activation. The activity of betulinic acid 2 is independent
of p53 tumor suppressor gene status and independent of death inducing ligand/receptor CD95/CD95L pairs
(Fulda et al., 1997, Wick et al., 1999, Selzer et al., 2000 and Zuco et al., 2002). Betulinic acid 2 induces
cytoplasmic shrinking and surface blebbing in melanoma (MEL-14) cells (Pisha et al., 1995) and DNA or
nuclear fragmentation in melanoma (MEL-14; Pisha et al., 1995), (B16; Liu et al., 2004), several
neuroblastoma (e.g. SHEP), medulloblastoma (e.g. Daoy and MHH1), glioblastoma (e.g. A172, U343 and SK17)
and Ewing's sarcoma (A17/95) cells (Fulda et al., 1997, Fulda et al., 1998b and Fulda et al., 1999). Betulinic
acid 2 also induces caspase activity (Fulda et al., 1997 and Fulda et al., 1998b) particularly caspases 3 and 8
activation (Fulda et al., 1999) associated with poly(ADP-ribose)polymerase cleavage in glioma (e.g. LN-18 and
U87 MG) cell lines (Wick et al., 1999). Zanon et al. (2004) reported that betulinic acid 2promoted enzymatic
activity of caspases 2, 3, 8 and 9 independently of APAF-1 (apoptosis protease activator protein-1) status in
melanoma cells (e.g. SK-Mel-5). However, Tan et al. (2003) reported that activity of betulinic acid 2 was
independent of caspases 3, 8 and 9 in melanoma (UISO-Mel-1) cells. Betulinic acid 2 has also direct effect on
mitochondria by induction of mitochondrial permeability transition (Fulda et al., 1997,Fulda et al.,
1998b and Fulda et al., 1999) and depolarization of mitochondrial membrane potential (Tan et al., 2003 and Liu
et al., 2004). Betulinic acid 2 also induces releasing of cytochrome c (Fulda et al., 1998b and Fulda et al., 1999).
It also induces formation of reactive oxygen species (Fulda et al., 1998b; Wick et al., 1999 and Liu et al., 2004)
and upregulation of KILLER/DR5 mRNA expression (pro-apoptotic death-domain-containing receptor) in
melanoma (7336) and glioblastoma (H80, G139 and U373) cell lines (Meng and El-Deiry, 2001). Betulinic
acid 2 induces increased phosphorylation pattern and thus activation of p38 and stress activated protein (SAP)
kinase/c-Jun NH2-terminal (JNK) kinase [proapoptotic mitogen-activated protein kinase (MAPK)], without
changes in the phosphorylation of extracellular signal-regulated kinases (ERK, anti-apoptotic MAPKs; Tan et al.,
2003). However, Qiu et al. (2005) reported that betulinic acid 2induces ERK activation in melanoma cells.
Betulinic acid 2 suppresses carcinogen induced NF-B transcription factor activation through inhibition of IB
kinase phosphorylation and degradation, p65 phosphorylation and nuclear translocation. It also suppresses the
production of NF-B-regulated gene products cyclooxygenase-2 and metalloprotease-9 in epithelial (HCT 116),
colon carcinoma (Caco 2) and lung adenocarcinoma (H 1299; Takada and Aggarwal, 2003).
However, Kasperczyk et al. (2005) reported that betulinic acid 2 activates NF-B in neuroblastoma (SHEP and
SH-SY5Y), glioblastoma (LN229 and U373) and melanoma (MeWo) cells, resulting in increased IKappa B
kinase (IKK) activity and phosphorylation of IB- at serine 32/36 followed by degradation of IB- in SHEP
neuroblastoma cells. Moreover, Qiu et al. (2005) reported that betulinic acid 2 is contributing to less sensitivity
in human melanoma cells by inducing EGFR (epithelial growth factor receptor) tyrosine phosphorylation,
serinethreonine kinase (AKT) phosphorylation, survivin expression and weak activation of Jun N-terminal
kinase (JNK) and p38.
Topoisomerase inhibitors are anticancer drugs with clinical importance. Betulinic acid 2 was shown to be a
potent inhibitor of mammalian type I DNA topoisomerase with IC 50 = 5 M (IC50 = 50% inhibition
concentration; Chowdhury et al., 2002).

2.1.2. Carbon-3 derivatives and ring A modified compounds

Betulonic acid 3 is more active than betulinic acid 2 against melanoma cell line MEL-2 (Kim et al.,
1998 and Kim et al., 2001), whereas the effect of C-3 ketone carbonyl moiety on melanoma cell lines G361 and
SK-MEL-28 was small (Hata et al., 2003). With respect to betulinic acid 2, betulonic acid 3 is also more active
against endothelial (ECV304; Mukherjee et al., 2004a), human epidermoid carcinoma of the mouth (KB; Kim et
al., 1998), ovary (PA-1; Mukherjee et al., 2004c), colon (HT 29), prostate (PC 3), leukemia (CEM) and breast
(MCF 7; Urban et al., 2004) cancer cell lines. However, betulonic acid 3 is less active against human
lymphoblastic and human B-cell lymphoma leukemia (CEM.CM3 and BRISTOL8), prostate (DU145) and lung
(L132) cancer cell lines (Mukherjee et al., 2004c). With respect to betulinic acid 2, acetylation of the hydroxy
group of betulinic acid 2 at C-3 does not seem to have influence on the cytotoxic effect on melanoma (MELB16-2F2; Hata et al., 2002), (M14-MEL, SK-MEL-2 and UACC-257), colon (HCT-116) and prostate (PC3; Kim
et al., 2001) cell lines. The introduction of a methyl oxime or oxime group to betulinic acid 2 at C-3 resulted in
the loss of cytotoxicity against the cultured human melanoma (MEL-2) cell line. The amino analogue showed a
comparable influence on cytotoxicity (MEL-2) with respect to betulinic acid 2 (Kim et al., 1998). The cytotoxic
activity of epibetulinic acid increased slightly toward the Bro melanoma cells and decreased toward MS
melanoma cells (Symon et al., 2003). Incorporation of the bulky arylpropenoyl scaffold at C-3 made betulinic
acid derivatives non-toxic, when tested against the colon cancer (HT 29) cell line (Baglin et al., 2003b). 3Hydrazono-20,29-dihydrobetulinic acid derivatives 46 (Fig. 3) were more active than betulinic acid 2 against
the endothelial (ECV304) cell line. These derivatives also exhibited high endothelial specificity against the lung
carcinoma (A549) cell line. The 3-O-acyl betulinic acid derivative having an electron withdrawing group in
aromatic ring at C-3 side chain (compound 7, Fig. 4) is a more active anti-angiogenic agent on endothelial
(ECV304) cell line than the compounds with bulky pentyl (compound 8) or heptyl group or electron donating
group in the aromatic ring (compound 9; Mukherjee et al., 2004a).

Fig. 3.
3-Hydrazono betulinic acid derivatives with stronger cytotoxicity against endothelial cell line than betulinic acid 2.
Figure options

Fig. 4.
Derivative 7 with an electron withdrawing group is more active anti-angiogenic agent against endothelial cell line than
compounds 89.
Figure options

In 20,29-dihydrobetulinic acid analogues the effect was reversed. Compounds possessing fluoro substituents
showed higher cytotoxicity and a compound bearing an electron withdrawing trifluoromethyl group exhibited
lower cytotoxicity (Mukherjee et al., 2004b). Ring A modified compounds 10 and 11 having a hydroxy group at
C-2 (Fig. 5) were found to be slightly less cytotoxic than betulinic acid 2 when tested over 60 different cell lines.
However, compound 10 showed significant cytotoxicity against all six leukemia cell lines screened.
Compound 11 had a strong activity against ovarian (SK-OV-3) cancer cell line with GI 50 < 10 nM (GI50 = 50%
growth inhibition; Deng and Snyder, 2002). The 2-bromo-20,29-dihydrobetulonic acid 12 showed improved
cytotoxicity against human lymphoblastic leukemia (MOLT-4 and CEM.CM3) and ovary (PA-1) cell lines
(Mukherjee et al., 2004c). Several ring A modified betulinic acid derivatives with minimal steric hindrance were
found to be more cytotoxic than betulinic acid 2, when tested on melanoma (SK-MEL-2 and B16-F10) and lung
(A-549) cancer cell lines. Especially, the compounds having an enone structure with an electron withdrawing
group such as 2-chloro 13, 2-cyano or 2-formyl at position C-2 displayed strong cytotoxicity (You et al., 2003).
Ring A seco derivatives (1417, 14 being the most active, Fig. 6) of betulinic acid showed strong cytotoxicity
against human lymphoblastic leukemia (CEM), leukemia (K562 and K562 Tax), colon (HT 29), prostate (PC 3
and DU 145), breast (MCF 7) and melanoma (SK-MEL2) cell lines. In addition, secoanhydrides 18 and 19 as
well as triol 20 ( Fig. 7) showed greatly improved cytotoxicity ( Urban et al., 2004).

Fig. 5.
Ring A modified compounds with strong cytotoxicity.
Figure options

Fig. 6.
Ring A seco derivatives with strong cytotoxicity.
Figure options

Fig. 7.
Betulin triol derivative with strong cytotoxicity.
Figure options

2.1.3. Carbon-28 derivatives and ring E modified compounds

Betulin 1 is slightly more cytotoxic than lupeol against melanoma (MEL B16 2F2, G361 and SK-MEL-28),
neuroblastoma (GOTO and NB-1) and leukemia (HL60, U937 and K562) cell lines. When C-28 hydroxy group
of betulin 1 is oxidized to the corresponding betulinic aldehyde, cytotoxicity is increased against the same cancer
cell lines by several folds. Further oxidation of betulinic aldehyde to betulinic acid 2 did not have so much effect
on cytotoxicity. In addition, methyl betulinate has approximately the same cytotoxicity profile as betulinic
aldehyde (Hata et al., 2002 and Hata et al., 2003). Betulinic acid 2 is much more cytotoxic on melanoma (MEL2; Kim et al., 1998) and mouse epidermal (JB6; Gao et al., 2003) cell lines, when compared to betulin 1. The
results indicate that carbonyl, not the carboxyl group, at C-28 is essential for cytotoxic activity. The activity of
the corresponding alkyl esters was clearly lower. 3,28-Dioxime betulin is cytotoxic against human oral
epidermoid carcinoma (KB), whereas 3-oxime betulinic acid is inactive. 3,28-Dimethyloxime betulin is inactive
against melanoma (MEL-2) and human oral epidermoid carcinoma (KB;Kim et al., 1998). Esterification of the
carboxylic moiety with a glycosyl group at C-28 led to inactive compounds against colon (HT 29) cancer cell
line (Baglin et al., 2003b). Methyl esters of amide derivatives of alanine 21 and valine 22 (Fig. 8) and free acid
of alanine 23 and glycine 24 showed cytotoxicity against the melanoma (MEL-2) cell line comparable to that of
betulinic acid 2. Only free acids of alanine 23 and valine 25also showed cytotoxicity against human oral
epidermoid carcinoma (KB; Jeong et al., 1999). In another study, it was demonstrated that compounds 26
28 (Fig. 9) containing lupane E-ring-derived unsaturated ketone or carbonyl moieties possessed broad in vitro
cytotoxic activity against several tumor cell lines by inducing apoptosis in cells. However, cytotoxicity of these
derivatives increased considerably against normal human lymphocytes too (Sarek et al., 2003).

Fig. 8.

Structures of the most cytotoxic betulinic acid amide derivatives.

Figure options

Fig. 9.
Lupane E-ring-derived carbonyl compounds possessing stronger cytotoxicity than betulinic acid 2 against several tumor
cell lines (Sarek et al., 2003).
Figure options

2.1.4. Derivatives via the isopropylidene moiety

Hydrogenation of the isopropylidene side chain at C-20-29 of betulinic acid 2 had a slightly decreasing effect on
cytotoxicity, when tested on melanoma (MEL-2) cell line. Hydrogenation of the side chain of betulonic
acid3 and 3-methyloxime betulinic acid made compounds inactive against human oral epidermoid carcinoma
(KB; Kim et al., 1998). 20,29-Dihydrobetulinic acid and 20,29-dihydromethyl betulinate had enhanced
inhibitory effect on topoisomerase I when compared to betulinic acid 2 (Chowdhury et al., 2002). 20,29Dihydrobetulinic acid had a slightly improved cytotoxicity against leukemia/lymphoma (MOLT-4) and prostate
(DU145) cell lines. 20,29-Dihydrobetulinic acid is also cytotoxic on ovary (PA-1) and lung (A549) cancer cell
lines, whereas betulinic acid 2 is inactive. The isopropylidene double bond between C-20 and C-29 in betulinic
acid 2 plays a crucial role in eliciting high endothelial (ECV304) cell cytotoxicity, the endothelial cell specificity
and inhibition of tube-like structures (Mukherjee et al., 2004b and Mukherjee et al., 2004c). The effect of
chemical modifications, such as insertion of ketone carbonyl, hydroxy, oxime, methyl oxime, hydroxylamino,
methoxylamino and ether at C-20 of betulinic acid 2, was found to be sensitive to the size and the electron
density of the substituents on retaining the cytotoxicity of betulinic acid 2 against human melanoma (M14-MEL,
SK-MEL-2 and UACC-257), colon carcinoma (HCT-116) and human prostate adenocarcinoma (PC3) cell lines.
The majority of these C-20 modified compounds were clearly less active or inactive, when compared to betulinic
acid 2 (Kim et al., 2001).

2.2. Anti-HIV activities

Anti-HIV-1 triterpenes are classified into five different classes depending on their action mechanism and their
molecular targets (Table 2; Huang and Chen, 2002). Currently approved anti-HIV drugs are either HIV-1 reverse
transcriptase or protease inhibitors. A new mechanism of action has been confirmed for the most promising antiHIV agents derived from betulin. Statine-derived IC9564 29a (Fig. 10) and its diastereomer
RPR103611 29b (Mayaux et al., 1994, Soler et al., 1996, Holz-Smith et al., 2001, Sun et al., 2002 and Yuan et
al., 2004) act as entry inhibitors and block HIV adsorption or membrane fusion. The dimethylsuccinyl derivative
of betulin DSB 30 (also known as YK-FH312 and PA-457; Fig. 11) (Sun et al., 1998a, Sun et al.,
1998b, Kashiwada et al., 1996, Kashiwada et al., 2000, Kashiwada et al., 2001, Hashimoto et al.,
1997,Kanamoto et al., 2001, Li et al., 2003, Huang et al., 2004 and Zhou et al., 2004) acts as a virus maturation
inhibitor.
Table 2.
Classes of anti-HIV triterpenes
Class
Site of action
1

Entry inhibitors that block HIV adsorption or membrane fusion

Reverse transcriptase inhibitors

Protease inhibitors

Virus maturation inhibitors that do not inhibit HIV-1 protease

Inhibitors with unknown mechanism of action

Table options

Fig. 10.
Structures of potent anti-HIV compounds IC9564 29a, RPR103611 29b.
Figure options

Fig. 11.
3-O-(3,3-dimethylsuccinyl) betulinic acid (DSB) 30.
Figure options

Even more potential compounds against HIV-1 (III B) are diesters 31 and 32 (Fig. 12) with therapeutic index (TI)
values of 21,515 and 42,400 and EC 50 values of 0.66 and 0.87 nM, respectively. Comparable TI and EC 50values
for approved anti-HIV drug AZT are between 33,33341,622 and 1545 nM, respectively (Sun et al.,
1998a; Kashiwada et al., 2001). When 3,3-dimethylsuccinyl side chain at C-3 was combined with aminoalkanoic
side chain at C-28, superior bifunctional anti-HIV compounds LH55 33 (Fig. 13) and LH15 were obtained.
IC50 values of LH55 33 (between 6.5 and 32 nM) are 453 times better than in IC9564 29a or in
DSB30 (depending on the tested HIV-1 strain). Inhibition of the viruses, which are resistant to HIV-1 reverse
transcriptase inhibitors or protease inhibitors and viruses that are resistant to IC9564 29a or DSB 30 alone, was
also obtained. The antifusion activity of LH55 33 blocks HIV-1 before it enters the cell. On the other hand,
viruses which bypass this blockage, have to face the maturation inhibition activity of these compounds (Huang et
al., 2004).

Fig. 12.
Structures of potent anti-HIV compounds 31 and 32.
Figure options

Fig. 13.
Structure of LH-55 33.
Figure options

2.2.1. Carbon-3 derivatives

Betulin 1 and 3,28-diacetylbetulin are inactive as anti-HIV agents. Epibetulin, betulinic acid 2, betulonic
acid 3and 3,28-dioxime are also rather inactive as anti-HIV agents (Hashimoto et al., 1997, Sun et al.,
1998a and Sun et al., 1998b). In the series of C-28 -undecanoic amide derivatives, 3--hydroxy as well as 3keto groups led to less active compounds. 3--Deoxy, 3--methoxy and 3--amino compounds were inactive.
Structural variations in ring A of the triterpene highlight the importance of the 3--hydroxy group. None of the
tested compounds showed anti-HIV activity against HIV-2 (ROD). These studies demonstrate that even minor
modifications at ring A lead to considerable loss in anti-HIV activity (Evers et al., 1996). Later results indicate
the importance of the C-3 acyl group to the anti-HIV-1 (III B) effect. The reduced activity of amide, 3-ester and
oxime analogues suggested that the orientation and linkage of the C-3 acyl side chain plays an important role in
the anti-HIV activity (Sun et al., 1998b). A series of compounds, where C-3 hydroxy group is esterified with e.g.
diglycolic, glutaric or succinic anhydrides, was studied. The highest anti-HIV-1 (III B) activity was observed with

compounds containing isovaleryl domain incorporated with 3,3-dimethylsuccinyl (DSB, Fig. 11)30 or -glutaryl
moieties with terminal carboxylic group (Hashimoto et al., 1997 and Sun et al., 1998a). DSB 30was 4000-fold
more active and had a 2150-fold higher therapeutic index than betulinic acid 2 (Kashiwada et al.,
1996 and Hashimoto et al., 1997). DSB 30 exhibits a mean (six different virus strains) IC 50 value of 10.3 nM,
which is comparable to that of approved anti-HIV drugs AZT and indinavir (4.3 and 8.8 nM, respectively) and
better than the non-nucleoside reverse transcriptase (RT) inhibitor nevirapine (40.0 nM). Importantly,
DSB 30 displayed a similar level of activity against a panel of virus isolates resistant to the three classes of drugs
targeting the viral reverse transcriptase (RT) and protease (PR) enzymes (Li et al., 2003). Results confirm that
DSB 30 affects the step(s) of virion assembly and/or budding of virions (Kanamoto et al., 2001, Li et al.,
2003 and Zhou et al., 2004). In particular, a mechanism is proposed, where DSB 30 acts late in the HIV-1 life
cycle and binds to the CA-p2 junction of Gag polyprotein (single polyprotein which is sufficient for virus
particle assembly) by viral protease during HIV-1 particle assembly and sterically inhibits cleavage of this site.
Incomplete processing of Gag polyprotein results to virions, which lack a functional core and are thus noninfectious (Zhou et al., 2004). DSB-related analogues of ursolic acid and oleanolic acid, which contain a sixmembered E-ring, led to clearly less active compounds (Kashiwada et al., 2000).

2.2.2. Carbon-28 derivatives

Oxidation of betulin 1 to betulinic acid 2 increases anti-HIV activity considerably, but both compounds are still
rather inactive (Sun et al., 1998a and Sun et al., 1998b). Studies with modifications of the alkanoic acid side
chain at C-28 showed that the presence of hydrogen bond-donating amide bond within the side chain was
important for the optimal anti-HIV activity. Variations at the position C-28 carboxyl, such as N-methylation of
the amide moiety, replacement of the amide by an ester or replacement of the carbonyl by a methylene led to the
complete loss of anti-HIV activity. Reversed amide and urea derivatives were also inactive against HIV-1 (Evers
et al., 1996). Also bulky non-carboxylic group at C-28 produced less active anti-HIV compounds (Sun et al.,
1998b). Systematic alterations, for example to determine the optimal chain length have been studied in
condensation products from betulinic acid 2 with two aminoalkanoic acids in C-28 side chain and condensation
product from (betulinylamino)octanoic acid with -, -, and -amino acids. Statine derivative
RPR103611 29b showed the strongest anti-HIV activity. Altering of the chain length of IC9564 29a from
aminoalkanoic side led to a decrease in anti-HIV activity (Soler et al., 1996). However, the statine moiety could
be replaced with L-leucine moiety without affecting anti-HIV activity. Stereoisomers IC9564 29a and
RPR103611 29b were found equal in anti-HIV activity with an EC 50 (concentration which inhibits HIV-1
replication by 50%, 50% effective concentration) value of 0.40 M (Sun et al., 2002). However, mutations in the
HIV-1 envelope glycoprotein gp120 sequence (such as in NL4-3 and M2-NLDH HIV-1 variants) makes HIV-1
resistant to IC9564 29a (Holz-Smith et al., 2001 and Yuan et al., 2004).

2.2.3. Derivatives via the isopropylidene moiety

Various substituents were introduced to position C-30 of the C-28--undecanoic amide, but none of them was
better than the corresponding C-30-unsubstituted compound. Anti-HIV activity of 30-(hydroxyethyl)thio, 30-[2(diethylamino)ethyl]thio, 30-(1-pyrrolidine) and 30-hydroxy derivatives remained high, but they were not better
than the unsubstituted compound. Acidic groups, primary or secondary amine groups or an aromatic group at C30 led to significant loss of anti-HIV activity. An acetyl or carboxyl group at C-19 led to inactive compounds
against HIV-1. Hydrogenation of the isopropylidene group also reduced anti-HIV activity (Evers et al., 1996).
Hydroxylation of betulin 1 at C-30 decreased anti-HIV activity considerably in acutely infected H9 lymphocytes
(Sun et al., 1998b). In the series of 3,28-diesterified compounds, saturated compounds were found to be several
fold less active than unsaturated diesters (Sun et al., 1998a and Sun et al., 1998b). The saturated analogue of
IC9564 29a was equipotent, suggesting that double bond does not play a crucial role in this type of compounds
(Sun et al., 2002). Saturated analogue of DSB 30 is only slightly less active (Hashimoto et al., 1997).

2.3. Other antiviral and antibacterial activities

The antiviral activity of betulin 1, betulinic acid 2, betulonic acid 3 and their derivatives have also been studied
against influenza A, herpes simplex type 1 (HSV-1), influenza FPV/Rostock and ECHO-6 enterovirus
(Kanamoto et al., 2001, Flekhter et al., 2002b, Flekhter et al., 2002c, Flekhter et al., 2003, Baltina et al.,
2003 and Pavlova et al., 2003). Betulin 1, betulinic acid 2 and betulonic acid 3 showed antiviral activity against
herpes simplex type 1 (Pavlova et al., 2003). Introduction of ureides or the C-28 amide group to betulonic
acid3 clearly increased their antiviral activity against HSV-1 virus (Baltina et al., 2003 and Flekhter et al., 2003).
Potential anti-HIV compound DSB 30 was found inactive against HSV-1 (Kanamoto et al., 2001). Betulin 1and

betulinic acid 2 were inactive against influenza FPV/Rostock virus and betulonic acid 3 showed a weak antiviral
activity (Pavlova et al., 2003).
3-Oxime of betulonic acid was more active against influenza A virus than betulinic acid 2. When the C-28
carboxyl group was substituted by a primary amide group, the activity was further improved, the EC 50 being
0.7 M (Baltina et al., 2003). Also, 3,28-dioxymebetulin caused some antiviral activity against the type A strain
of influenza virus (Flekhter et al., 2002c). Ureido derivatives of betulonic acid caused clear inhibiting activity,
but the effect was quite weak (Flekhter et al., 2003). The potential anti-HIV compound DSB 30 was inactive
against influenza virus (Kanamoto et al., 2001). Betulin 1, betulinic acid 2 and betulonic acid 3showed antiviral
activity against ECHO-6 virus. Betulonic acid 3 caused the most significant reduction of the ECHO-6 virus titer
(Pavlova et al., 2003). 3,28-Dioximebetulin was also found to possess some antiviral activity against ECHO-6
virus (Flekhter et al., 2002c).
Betulinic acid 2 has been found to be inactive against Staphylococcus aureus, Escherichia coli ( Hess et al.,
1995), Bacillus subtilis and Micrococcus luteus ( Nick et al., 1995). Generally, betulin derivatives seem to have
rather poor antibacterial activity.

2.4. Anti-inflammatory activity

Some triterpenes have been shown to be effective as anti-inflammatory agents. From derivatives of ursane,
oleanane and lupane series, betulin 1 and betulinic acid 2 are the most effective compounds in skin inflammation
and ear edema induced by mezerein, DPT (12-deoxyphorbol-13-tetradecanoate), DPP (12-deoxyphorbol-13phenylacetate) or TPA (12-O-tetradecanoylphorbol-13-acetate) in mice. However, betulin 1and betulinic
acid 2 were inactive against inflammation induced by resiniferatoxin, xylene and arachidonic acid ( Huguet et
al., 2000). Betulinic acid 2 shows significant anti-inflammatory activity against rat paw edema induced by
carrageenan and serotonin, which is comparable to the standard anti-inflammatory agents phenylbutazone and
dexamethasone ( Mukherjee et al., 1997). Erythrodiol and ursolic acid were found to be more active than their
five-membered ring E analogues betulin 1 and betulinic acid 2 against chronic dermal inflammation induced by
multiple-dose TPA ( Manez et al., 1997).
Betulin 1 and betulinic acid 2 were found to inhibit phospholipase A2 (PLA2, which plays a crucial role in
inflammation process) activity at 5 M concentrations by 30% and 40%, respectively (Bernard et al., 2001).
Betulin diesters and allobetulin esters, such as bishemiphthalate and allobetulin-3-O-[2-(4-chlorophenyl)-3methyl]butanoate, were found to be active against foot edema induced by formalin and carrageenan in mice
(Flekhter et al., 2002d; Karachurina et al., 2002). Betulonic acid 3 and 28-oxoallobetulone have been shown to
possess antiulcer (induced by indomethacin or acetylsalicylic acid) activity in rats ( Flekhter et al., 2002b). On
the chronic ulceration models induced by ethanol or cryogenic factor, betulin-3,28-diphthalate led to a reliable
decrease in the total area of ulceration ( Karachurina et al., 2002). An antiulcer effect was also seen with betulin
dinicotinate ( Flekhter et al., 2002a).

2.5. Antimalarial activity

Betulin 1, betulinic acid 2, ursolic acid and oleanolic acid have also been tested for antimalarial activity against
chloroquine sensitive (T9-96 strain) and resistant (K1 strain) Plasmodium falciparum. Betulin 1 was inactive,
whereas the others showed moderate activity, betulinic acid 2 being most active in vitro against both strains of P.
falciparum at IC50 values 19.6 g/mL (K1) and 25.9 g/mL (T9-96) respectively. The 2-propenyl side chain of
betulinic acid 2 was not essential for activity, whereas structural changes at C-17 dramatically altered the activity
( Steele et al., 1999). Betulinic acid 2 is also moderately active in vitro against the chloroquine sensitive strain
(3D7) at an IC50 value of 19 M. Comparable IC50 value for chloroquine is 0.024 M ( Duker-Eshun et al., 2004).
Methyl betulinate possessed slightly better in vitro antiplasmodial activity against P. falciparum strain 3D7 with
an IC50 value of 7.0 M ( Ziegler et al., 2004).

3. Conclusions and future perspectives

The high availability of betulin 1 from the bark of birch trees makes it a potentially important raw material for
polymers and it is an interesting precursor of biologically more active compounds. Betulin 1 can be easily
converted to relatively non-toxic betulinic acid 2, which has shown to possess wide spectrum of biological
activities. Betulinic acid 2 and its derivatives have been shown to possess antimalarial, anti-inflammatory and
antifungal activity. Betulinic acid's 2 anti-HIV and anticancer properties have especially been sources of great
interest. Derivatives of betulinic acid have showed cytotoxicity and anti-HIV activity at micromolar or even at
nanomolar concentrations, which are comparable to some clinically used drugs. New mechanism of action has
been confirmed for some of the most promising anti-HIV derivatives, which makes them potentially useful
additives to the current anti-HIV therapy. Betulinic acid 2 has specific cytotoxicity on several tumor cell lines by

inducing apoptosis in cells. However, thorough preclinical animal experimentation is needed to evaluate the
usefulness of betulinic acid 2 and its derivatives in vivo.

Acknowledgements
We thank National Technology Agency (Tekes) for financial support. This study was also supported by the
Academy of Finland (Grant 108376) and the EU grant to Pro-KinaseResearch LSHB-2004-503467.

References
1.

2.

o
o
o

Aiken and Chen, 2005

C. Aiken, C.H. Chen
Betulinic acid derivatives as HIV-1 antivirals

Trends Mol. Med., 11 (2005), pp. 3136

o
o
o
o
o

3.

o
o

Mini Rev. Med. Chem., 3 (2003), pp. 525539

Baglin et al., 2003b

I. Baglin, A. Poumaroux, M. Nour, K. Tan, A.C. Mitaine-Offer, M.A. Lacaille-Dubois, B.
Chauffert, C. Cave
New ursolic and betulinic derivatives as potential cytotoxic agents
J. Enzyme Inhib. Med. Chem., 18 (2003), pp. 111117

o
o
o
o
o

6.

Baglin et al., 2003a

I. Baglin, A.C. Mitaine-Offer, M. Nour, K. Tan, C. Cave, M.A. Lacaille-Dubois
A review of natural and modified betulinic, ursolic and echinocystic acid derivatives as
potential antitumor and anti-HIV agents

o
o
o

5.

J. Nat. Prod., 65 (2002), pp. 278282

o
o
o
o

4.

Akihisa et al., 2002

T. Akihisa, Y. Takamine, K. Yoshizumi, H. Tokuda, Y. Kimura, M. Ukiya, T. Nakahara, T.
Yokochi, E. Ichiishi, H. Nishino
Microbial transformations of two lupane-type triterpenes and anti-tumor-promoting
effects of the transformation products

Baltina et al., 2003

L.A. Baltina, O.B. Flekhter, L.R. Nigmatullina, E.I. Boreko, N.I. Pavlova, S.N. Nikolaeve,
O.V. Savinova, G.A. Tolstikov
Lupane triterpenes and derivatives with antiviral activity
Bioorg. Med. Chem. Lett., 13 (2003), pp. 35493552

o
o
o
o
o

Bernard et al., 2001

P. Bernard, T. Scior, B. Didier, M. Hibert, J. Berthon
Ethnopharmacology and bioinformatic combination for leads discovery: application to
phospholipase A2 inhibitors
Phytochemistry, 58 (2001), pp. 865874

7.
o
o
o

Chatterjee et al., 2000

P. Chatterjee, S.A. Kouzi, J.M. Pezzuto, M.T. Hamann
Biotransformation of the antimelanoma agent betulinic acid by Bacillus megaterium
ATCC 13368
Appl. Environ. Microbiol., 66 (2000), pp. 38503855

8.

o
o
o
o

Chowdhury et al., 2002

A.R. Chowdhury, S. Mandal, B. Mittra, S. Sharma, S. Mukhopadhyay, H.K. Majumder
Betulinic acid, a potent inhibitor of eukaryotic topoisomerase I: identification of the
inhibitory step, the major functional group responsible and development of more potent derivatives
Med. Sci. Monit., 8 (2002), pp. br254br260

9.

o
o
o
o

Cichewicz and Kouzi, 2003

R.H. Cichewicz, S.A. Kouzi
Chemistry, biological activity, and chemotherapeutic potential of betulinic acid for the
prevention and treatment of cancer and HIV infection
Med. Res. Rev., 24 (2003), pp. 90114

10.

o
o
o
o

Deng and Snyder, 2002

Y. Deng, J.K. Snyder
Preparation of a 24-Nor-1,4-dien-3-one triterpene derivative from betulin: a new route to
24-nortriterpene analogues
J. Org. Chem., 67 (2002), pp. 28642873

11.

12.

o
o
o
o

Duker-Eshun et al., 2004

G. Duker-Eshun, J.W. Jaroszewski, W.A. Asomaning, F. Oppong-Boachie, S.B. Christensen
Antiplasmodial constituents of Cajanus cajan

Phytother. Res., 18 (2004), pp. 128130

o
Eckerman and Ekman, 1985
C. Eckerman, R. Ekman
Comparison of solvents for extraction and crystallization of betulinol from birch bark

o
o
o
waste

Paperi ja Puu, 67 (1985), pp. 100106

13.

o
Eder-Czembirek et al., 2005
C. Eder-Czembirek, C. Czembirek, B.M. Erovic, E. Selzer, D. Turhani, L. Vormittag, D.

o
o
Thurnher
o
o

14.

Combination of betulinic acid with cisplatindifferent cytotoxic effects in two head and
neck cancer cell lines
Oncol. Rep., 14 (2005), pp. 667671

o
o

Ehrhardt et al., 2004

15.

16.

o
o

H. Ehrhardt, S. Fulda, M. Fuhrer, K. Debatin, I. Jeremias

Betulinic acid-induced apoptosis in leukemia cells

Leukemia, 18 (2004), pp. 14061412

o
o
o
o

Eiznhamer and Xu, 2004

D.A. Eiznhamer, Z. Xu
Betulinic acid: a promising anticancer candidate

IDrugs: Investig. Drugs J., 7 (2004), pp. 359373

o
o
o
o

Evers et al., 1996

M. Evers, C. Poujade, F. Soler, Y. Ribeill, C. James, Y. Lelievre, J.C. Gueguen, D. Reisdorf, I.
Morize, R. Pauwels, E. De Clercq, Y. Henin, A. Bousseau, J.F. Mayaux, J.B. Le Pecq, N. Dereu
Betulinic acid derivatives: a new class of Human Immunodeficiency Virus type 1 specific
inhibitors with a new mode of action
J. Med. Chem., 39 (1996), pp. 10561068

17.

o
o
o
o

Flekhter et al., 2002a

O.B. Flekhter, L.T. Karachurina, L.R. Nigmatullina, T.A. Sapozhnikova, L.A. Baltina, F.S.
Zarudii, F.Z. Galin, L.V. Spirikhin, G.A. Tolstikov, O.A. Plyasunova, A.G. Pokrovskii
Synthesis and pharmacological activity of betulin dinicotinate
Russ. J. Bioorg. Chem., 28 (2002), pp. 494500

18.

o
o
o
o

Flekhter et al., 2002b

O.B. Flekhter, L.R. Nigmatullina, L.A. Baltina, L.T. Karachurina, F.Z. Galin, F.S. Zarudii,
G.A. Tolstikov, E.I. Boreko, N.I. Pavlova, S.N. Nikolaeva, O.V. Savinova
Synthesis of betulinic acid from betulin extract and study of the antiviral and antiulcer
activity of some related terpenoids
Pharm. Chem. J., 36 (2002), pp. 484487

19.

o
o
o
o

Flekhter et al., 2002c

O.B. Flekhter, O.Y. Ashavina, E.I. Boreko, L.T. Karachurina, N.I. Pavlova, N.N. Kabalnova,
O.V. Savinova, F.Z. Galin, S.N. Nikolaeva, F.S. Zarudii, L.A. Baltina, G.A. Tolstikov
Synthesis of 3-O-acetylbetulinic and betulonic aldehydes according to Svern and the
pharmacological activity of related oximes
Pharm. Chem. J., 36 (2002), pp. 303306

20.

o
Flekhter et al., 2002d
O.B. Flekhter, N.I. Medvedeva, L.T. Karachurina, L.A. Baltina, F.S. Zarudii, F.Z. Galin, G.A.

o
o
Tolstikov

1.

Synthesis and antiinflammatory activity of new acylated betulin derivatives

Pharm. Chem. J., 36 (2002), pp. 488491

o
o
o

Flekhter et al., 2003

O.B. Flekhter, E.I. Boreko, L.R. Nigmatullina, E.V. Tretyakova, N.I. Pavlova, L.A. Baltina,
S.N. Nikolaeva, O.V. Savinova, F.Z. Galin, G.A. Tolstikov

Synthesis and antiviral activity of ureides and carbamates of betulinic acid and its

o
derivatives

Russ. J. Bioorg. Chem., 29 (2003), pp. 594600

2.

3.

o
o
o
o

Fulda and Debatin, 2005

S. Fulda, K. Debatin
Sensitization for anticancer drug-induced apoptosis by betulinic acid

Neoplasia, 7 (2005), pp. 162170

o
o
o
o

Fulda et al., 1997

S. Fulda, C. Friesen, M. Los, C. Scaffidi, W. Mier, M. Benedict, G. Nunez, P.H. Krammer,
M.E. Peter, K. ebatin
Betulinic acid triggers CD95 (APO-1/Fas)- and p53-independent apoptosis via activation
of caspases in neuroectodermal tumors
Cancer Res., 57 (1997), pp. 49564964

4.

5.

6.

7.

o
o
o
o

Fulda et al., 2004

S. Fulda, I. Jeremias, K. Debatin
Cooperation of betulinic acid and TRAIL to induce apoptosis in tumor cells

Oncogene, 23 (2004), pp. 76117620

o
o
o
o

Fulda et al., 1999

S. Fulda, I. Jeremias, H.H. Steiner, T. Pietsch, K. Debatin
Betulinic acid: a new cytotoxic agent against malignant brain-tumor cells

Int. J. Cancer, 82 (1999), pp. 435441

o
o
o
o

Fulda et al., 1998a

S. Fulda, S.A. Susin, G. Kroemer, K. Debatin
Molecular ordering of apoptosis induced by anticancer drugs in neuroblastoma cells

Cancer Res., 58 (1998), pp. 44534460

o
Fulda et al., 1998b
S. Fulda, C. Scaffidi, S.A. Susin, P.H. Krammer, G. Kroemer, M.E. Peter, K. Debatin
Activation of mitochondria and release of mitochondrial apoptogenic factors by betulinic

o
o
o
acid
o

8.

J. Biol. Chem., 273 (1998), pp. 3394233948

o
o
o
o
o

Gao et al., 2003

H. Gao, L. Wu, M. Kuroyanagi, K. Harada, N. Kawahara, T. Nakane, K. Umehara, A.
Hirasawa, Y. Nakamura
Antitumor-promoting constituents from Chaenomeles sinensis Koehne and their
activities in JB6 mouse epidermal cells
Chem. Pharm. Bull., 51 (2003), pp. 13181321

9.

10.

o
o
o
o

Hnsel et al., 1992

R. Hnsel, K. Keller, H. Rimpler, G. Schneider
Drogen A-D: Betula

Springer Verlag, Berlin (1992)

o
o
o
o
o

11.

13.

Hata et al., 2003

K. Hata, K. Hori, H. Ogasawara, S. Takahashi
Anti-leukemia activities of lup-28-al-20(29)-en-3-one, a lupane triterpene

Toxicol. Lett., 143 (2003), pp. 17

Hess et al., 1995

S.C. Hess, R.L. Brum, N.K. Honda, A.B. Cruz, E. Moretto, R.B. Cruz, I. Messana, F. Ferrari,
V. Cechinel Filho, R.A. Yunes
Antibacterial activity and phytochemical analysis of Vochysia divergens (Vochysiaceae)
J. Ethnopharmacol., 47 (1995), pp. 97100

o
o
o
o
o

16.

J. Nat. Prod., 65 (2002), pp. 645648

o
o
o

15.

Hata et al., 2002

K. Hata, K. Hori, S. Takahashi
Differentiation- and apoptosis-inducing activities by pentacyclic triterpenes on a mouse
melanoma cell line

o
o

14.

Bioorg. Med. Chem., 5 (1997), pp. 21332143

o
o
o
o

12.

Hashimoto et al., 1997

F. Hashimoto, Y. Kashiwada, L.M. Cosentino, C. Chen, P.E. Garrett, K. Lee
Anti-AIDS agents: XXVII. Synthesis and anti-HIV activity of betulinic acid and
dihydrobetulinic acid derivatives

Holz-Smith et al., 2001

S.L. Holz-Smith, I. Sun, L. Jin, T.J. Matthews, K. Lee, C.H. Chen
Role of human immunodeficiency virus (HIV) type 1 envelope in the anti-HIV activity of
the betulinic acid derivative IC9564
Antimicrob. Agents Chemother., 45 (2001), pp. 6066

o
o
o
o

Huang and Chen, 2002

L. Huang, C.H. Chen
Molecular targets of anti-HIV-1 triterpenes

Curr. Drug Target Infect. Disord., 2 (2002), pp. 3336

o
o
o

Huang et al., 2004

L. Huang, X. Yuan, C. Aiken, C.H. Chen

Bifunctional anti-human immunodeficiency virus type 1 small molecules with two novel
mechanisms of action
Antimicrob. Agents Chemother., 48 (2004), pp. 663665

17.

o
o
o
o

Huguet et al., 2000

A.I. Huguet, M. del Carmen Recio, S. Manez, R.M. Giner, J.L. Rios
Effect of triterpenoids on the inflammation induced by protein kinase C activators,
neuronally acting irritants and other agents
Eur. J. Pharmacol., 410 (2000), pp. 6981

18.

o
Jeong et al., 1999
H. Jeong, H. Chai, S. Park, D.S.H.L. Kim
Preparation of amino acid conjugates of betulinic acid with activity against human

o
o
o
melanoma

Bioorg. Med. Chem. Lett., 9 (1999), pp. 12011204

19.

o
o
o
o

Jeremias et al., 2004

I. Jeremias, H.H. Steiner, A. Benner, K.M. Debatin, C. Herold-Mende
Cell death induction by betulinic acid, ceramide and TRAIL in primary glioblastoma
multiforme cells
Acta Neurochir., 146 (2004), pp. 721729

20.

o
Kanamoto et al., 2001
T. Kanamoto, Y. Kashiwada, K. Kanbara, K. Gotoh, M. Yoshimori, T. Goto, K. Sano, H.

o
o
Nakashima
o
o

1.

2.

3.

Anti-human immunodeficiency virus activity of YK-FH312 (a betulinic acid derivative), a

novel compound blocking viral maturation
Antimicrob. Agents Chemther., 45 (2001), pp. 12251230

o
o
o
o

Karachurina et al., 2002

L.T. Karachurina, T.A. Sapozhnikova, F.S. Zarudii, O.B. Flekhter, F.Z. Galin
Antiinflammatory and antiulcer properties of betulin bis-hemiphthalate

Pharm. Chem. J., 36 (2002), pp. 432433

o
o
o
o

Kashiwada et al., 2001

Y. Kashiwada, J. Chiyo, Y. Ikeshiro, T. Nagao, H. Okabe, L.M. Cosentino, K. Fowke, K.H. Lee
3,28-Di-O-(dimethylsuccinyl)-betulin isomers as anti-HIV agents

Bioorg. Med. Chem. Lett., 11 (2001), pp. 183185

o
o
o
o

Kashiwada et al., 1996

Y. Kashiwada, F. Hashimoto, L.M. Cosentino, C. Chen, P.E. Garrett, K. Lee
Betulinic acid and dihydrobetulinic acid derivatives as potent anti-HIV agents

J. Med. Chem., 39 (1996), pp. 10161017

4.

5.

o
o
o
o

Kashiwada et al., 2000

Y. Kashiwada, T. Nagao, A. Hashimoto, Y. Ikeshiro, H. Okabe, L.M. Cosentino, K. Lee
Anti-AIDS agents 38. Anti-HIV activity of 3-O-acylursolic acid derivatives

J. Nat. Prod., 63 (2000), pp. 16191622

o
Kasperczyk et al., 2005
H. Kasperczyk, K. La Ferla-Bruehl, M.A. Westhoff, L. Behrend, R.M. Zwacka, K. Debatin, S.

o
o
Fulda
o
o

6.

7.

8.

9.

10.

Oncogene, 24 (2005), pp. 69456956

o
o
o
o

Kim et al., 1997

D.S.H.L. Kim, Z. Chen, N. Van Tuyen, J.M. Pezzuto, S. Qiu, Z. Lu
A concise semi-synthetic approach to betulinic acid from betulin

Synth. Commun., 27 (1997), pp. 16071612

o
o
o
o

Kim et al., 1998

D.S.H.L. Kim, J.M. Pezzuto, E. Pisha
Synthesis of betulinic acid derivatives with activity against human melanoma

Bioorg. Med. Chem. Lett., 8 (1998), pp. 17071712

o
o
o
o

Kim et al., 2001

J.Y. Kim, H.M. Koo, D.S.H.L. Kim
Development of C-20 modified betulinic acid derivatives as antitumor agents

Bioorg. Med. Chem. Lett., 11 (2001), pp. 24052408

o
o
o
o

Kouzi et al., 2000

S.A. Kouzi, P. Chatterjee, J.M. Pezzuto, M.T. Hamann
Microbial transformations of the antimelanoma agent betulinic acid

J. Nat. Prod., 63 (2000), pp. 16531657

o
o
o
o
o

11.

Betulinic acid as new activator of NF-B: molecular mechanisms and implications for
cancer therapy

Li et al., 2003
F. Li, R. Goila-gaur, K. Salzwedel, N.R. Kilgore, M. Reddick, C. Matallana, A. Castillo, D.
Zoumplis, D.E. Martin, J.M. Orenstein, G.P. Allaway, E.O. Freed, C.T. Wild
PA-457: a potent HIV inhibitor that disrupts core condensation by targeting a late step in
Gag processing
Proc. Natl. Acad. Sci. U.S.A., 100 (2003), pp. 1355513560

o
o
o
o

Liu et al., 2004

W. Liu, J.C.K. Ho, F.W.K. Cheung, B.P.L. Liu, W. Ye, C. Che
Apoptotic activity of betulinic acid derivatives on murine melanoma B16 cell line

Eur. J. Pharmacol., 498 (2004), pp. 7178

12.

13.

o
o
o
o

Manez et al., 1997

S. Manez, M.C. Recio, R.M. Giner, J.L. Rios
Effect of selected triterpenoids on chronic dermal inflammation

Eur. J. Pharmacol., 334 (1997), pp. 103105

o
o
o
o

Mayaux et al., 1994

J.F. Mayaux, A. Bousseau, R. Pauwels, T. Huet, Y. Henin, N. Dereu, M. Evers, F. Soler, C.
Poujade, E. De Clercq, J.B. Le Pecq
Triterpene derivatives that block entry of human immunodeficiency virus type 1 into
cells
Proc. Natl. Acad. Sci., 91 (1994), pp. 35643568

14.

o
o
o
o

Meng and El-Deiry, 2001

R.D. Meng, W.S. El-Deiry
p53-Independent upregulation of KILLER/DR5 TRAIL receptor expression by
glucocorticoids and interferon-g
Exp. Cell Res., 262 (2001), pp. 154169

15.

16.

o
o
o
o

Mukherjee et al., 1997

P.K. Mukherjee, K. Saha, J. Das, M. Pal, B.P. Saha
Studies on the anti-inflammatory activity of rhizomes of Nelumbo nucifera

Planta Med., 63 (1997), pp. 367369

o
Mukherjee et al., 2004a
R. Mukherjee, M. Jaggi, P. Rajendran, M.J.A. Siddiqui, S.K. Srivastava, A. Vardhan, A.C.

o
o
Burman

17.

Betulinic acid and its derivatives as anti-angiogenic agents

Bioorg. Med. Chem. Lett., 14 (2004), pp. 21812184

o
o
o
o

Mukherjee et al., 2004b

R. Mukherjee, M. Jaggi, P. Rajendran, S.K. Srivastava, J.A. Siddiqui Mohammad, A. Vardhan,
A.C. Burman
Synthesis of 3-O-acyl/3-benzylidene/3-hydrazone/3-hydrazine/17-carboxyacryloyl ester
derivatives of betulinic acid as anti-angiogenic agents
Bioorg. Med. Chem. Lett., 14 (2004), pp. 31693172

18.

o
Mukherjee et al., 2004c
R. Mukherjee, M. Jaggi, M.J.A. Siddiqui, S.K. Srivastava, P. Rajendran, A. Vardhan, A.C.

o
o
Burman
o
o

Synthesis and cytotoxic activity of 3-O-acyl/3-hydrazine/2-bromo/20,29-dibromo betulinic

acid derivatives
Bioorg. Med. Chem. Lett., 14 (2004), pp. 40874091

19.

o
o
o
o
o

20.

o
o

Fitoterapia, 74 (2003), pp. 489492

Pisha et al., 1995

E. Pisha, H. Chai, I.S. Lee, T.E. Chagwedera, N.R. Farnsworth, G.A. Cordell, C.W.W. Beecher,
H.H.S. Fong, A.D. Kinghorn, D.M. Brown, M.C. Wani, M.E. Wall, T.J. Hieken, T.K. Das Gupta, J.M.
Pezzuto
Discovery of betulinic acid as a selective inhibitor of human melanoma that functions by
induction of apoptosis
Nat. Med., 1 (1995), pp. 10461051

Qiu et al., 2005

L. Qiu, Q. Wang, Q. Jiang, E. Schefeller, S. Derby, H. Wanebo, B. Yan, Y. Wan
Transient activation of EGFR/AKT cell survival pathway and expression of survivin
contribute to reduced sensitivity of human melanoma cells to betulinic acid
Int. J. Oncol., 27 (2005), pp. 823830

o
o
o
o
o

5.

Pavlova et al., 2003

N.I. Pavlova, O.V. Savinova, S.N. Nikolaeva, E.I. Boreko, O.B. Flekhter
Antiviral activity of betulin, betulinic and betulonic acids against some enveloped and
non-enveloped viruses

o
o
o
o

4.

Chem. Pharm. Bull., 45 (1997), pp. 16651670

o
o
o

3.

Noda et al., 1997

Y. Noda, T. Kaiya, K. Kohda, Y. Kawazoe
Enhanced cytotoxicity of some triterpenes toward leukemia L1210 cells cultured in low
pH media: possibility of a new mode of cell killing

o
o
o
o

2.

Phytochemistry, 40 (1995), pp. 16911695

o
o
o
o

1.

Nick et al., 1995

A. Nick, A.D. Wright, T. Rali, O. Sticher
Antibacterial triterpenoids from Dillenia papuana and their structureactivity
relationships

Sarek et al., 2003

J. Sarek, J. Klinot, P. Dzubak, E. Klinotova, V. Noskova, V. Krecek, G. Korinkova, J.O.
Thomson, A. Janostakova, S. Wang, S. Parsons, P.M. Fischer, N.Z. Zhelev, M. Hajduch
New lupane derived compounds with pro-apoptotic activity in cancer cells: synthesis and
structureactivity relationships
J. Med. Chem., 46 (2003), pp. 54025415

o
o
o
o

Sawada et al., 2004

N. Sawada, K. Kataoka, K. Kondo, H. Arimochi, H. Fujino, Y. Takahashi, T. Miyoshi, T.
Kuwahara, Y. Monden, Y. Ohnishi
Betulinic acid augments the inhibitory effects of vincristine on growth and lung
metastasis of B16F10 melanoma cells in mice

Br. J. Cancer, 90 (2004), pp. 16721678

6.

7.

o
o
o
o

Schmidt et al., 1997

M.L. Schmidt, K.L. Kuzmanoff, L. Ling-Indeck, J.M. Pezzuto
Betulinic acid induces apoptosis in human neuroblastoma cell lines

Eur. J. Cancer, 33 (1997), pp. 20072010

o
Selzer et al., 2000
E. Selzer, E. Pimentel, V. Wacheck, W. Schlegel, H. Pehamberger, B. Jansen, R. Kodym
Effects of betulinic acid alone and in combination with irradiation in human melanoma

o
o
o
cells
o

8.

o
o
o
o
o

9.

10.

Steele et al., 1999

J.C.P. Steele, D.C. Warhust, G.C. Kirby, M.S.J. Simmonds
In vitro and in vivo evaluation of betulinic acid as an antimalarial

Phytother. Res., 13 (1999), pp. 115119

o
Sun et al., 2002
I. Sun, C. Chen, Y. Kashiwada, J. Wu, H. Wang, K. Lee
Anti-AIDS agents 49. Synthesis, anti-HIV, and anti-fusion activities of IC9564 analogues
based on betulinic acid
J. Med. Chem., 45 (2002), pp. 42714275

o
o
o
o

Sun et al., 1998a

I. Sun, J. Shen, H. Wang, L.M. Cosentino, K. Lee
Anti-AIDS agents. 32. synthesis and anti-HIV activity of betulin derivatives

Bioorg. Med. Chem. Lett., 8 (1998), pp. 12671272

o
o
o
o
o

13.

J. Med. Chem., 39 (1996), pp. 10691083

o
o
o

12.

Soler et al., 1996

F. Soler, C. Poujade, M. Evers, J.C. Carry, Y. Henin, A. Bousseau, T. Huet, R. Pauwels, E. De
Clercq, J.F. Mayaux, J.B. Le Pecq, N. Dereu
Betulinic acid derivatives: a new class of specific inhibitors of human immunodeficiency
virus type 1 entry

o
o
o

11.

J. Invest. Dermatol., 114 (2000), pp. 935940

Sun et al., 1998b

I. Sun, H. Wang, Y. Kashiwada, J. Shen, L.M. Cosentino, C. Chen, L. Yang, K. Lee
Anti-AIDS agents. 34. Synthesis and structureactivity relationships of betulin
derivatives as anti-HIV agents
J. Med. Chem., 41 (1998), pp. 46484657

o
o
o

Symon et al., 2003

A.V. Symon, A.P. Kaplun, N.K. Vlasenkova, G.K. Gerasomiva, L.B. Shon, E.F. Litvin, L.M.
Kozlova, E.L. Surkova, V.I. Shvets
Epimerization of hydroxyl group in lupan series triterpenoids
Russ. J. Bioorg. Chem., 29 (2003), pp. 185189

14.

o
o
o
o

Takada and Aggarwal, 2003

Y. Takada, B.B. Aggarwal
Betulinic acid suppresses carcinogen-induced NF-kB activation through inhibition of
IkBa kinase and p65 phosphorylation: abrogation of cyclooxygenase-2 and matrix metalloprotease-9
J. Immunol., 171 (2003), pp. 32783286

15.

o
o
o
o

Tan et al., 2003

Y. Tan, R. Yu, J.M. Pezzuto
Betulinic acid-induced programmed cell death in human melanoma cells involves
mitogen-activated protein kinase activation
Clin. Cancer Res., 9 (2003), pp. 28662875

16.

o
o
o
o

Udeani et al., 1999

G.O. Udeani, G. Zhao, Y.G. Shin, B.P. Cooke, J. Graham, C.W.W. Beecher, A.D. Kinghorn,
J.M. Pezzuto
Pharmacokinetics and tissue distribution of betulinic acid in CD-1 mice
Biopharm. Drug Dispos., 20 (1999), pp. 379383

17.

18.

o
o
o
o

Urban et al., 2004

M. Urban, J. Sarek, J. Klinot, G. Korinkova, M. Hajduch
Synthesis of A-seco derivatives of betulinic acid with cytotoxic activity

J. Nat. Prod., 67 (2004), pp. 11001105

o
o
o
o

Wick et al., 1999

W. Wick, C. Grimmel, B. Wagenknecht, J. Dichgans, M. Weller
Betulinic acid-induced apoptosis in glioma cells: a sequential requirement for new
protein synthesis, formation of reactive oxygen species, and caspase processing
J. Pharmacol. Exp. Ther., 289 (1999), pp. 13061312

19.

20.

o
o
o
o

You et al., 2003

Y. You, Y. Kim, N. Nam, B. Ahn
Synthesis and cytotoxic activity of A-ring modified betulinic acid derivatives

Bioorg. Med. Chem. Lett., 13 (2003), pp. 31373140

o
Yuan et al., 2004
X. Yuan, L. Huang, P. Ho, C. Labranche, C.H. Chen
Conformation of gp120 determines the sensitivity of HIV-1 DH012 to the entry inhibitor

o
o
o
IC9564

Virology, 324 (2004), pp. 525530

1.

o
o
o

Zanon et al., 2004

M. Zanon, A. Piris, I. Bersani, C. Vegetti, A. Molla, A. Scarito, A. Anichini
Apoptosis protease activator protein-1 expression is dispensable for response of human
melanoma cells to distinct proapoptotic agents
Cancer Res., 64 (2004), pp. 73867394

2.

o
o
o

Zhou et al., 2004

J. Zhou, X. Yuan, D. Dismuke, B.M. Forshey, C. Lundquist, K. Lee, C. Aiken, C.H. Chen
Small-molecule inhibition of human immunodeficiency virus type 1 replication by
specific targeting of the final step of virion maturation
J. Virol., 78 (2004), pp. 922929

3.

o
o
o

Ziegler et al., 2004

H.L. Ziegler, H. Franzyk, M. Sairafianpour, M. Tabatabai, M.D. Tehrani, K. Bagherzadeh, H.
Hgerstrand, D. Staaerk, J.W. Jaroszewski
Erythrocyte membrane modifying agents and the inhibition of Plasmodium falciparum
growth: structureactivity relationships for betulinic acid analogues
Bioorg. Med. Chem., 12 (2004), pp. 119127

4.

o
o
o

Zuco et al., 2002

V. Zuco, R. Supino, S.C. Righetti, L. Cleris, E. Marchesi, C. Gambacorti-Passerini, F. Formelli
Selective cytotoxicity of betulinic acid on tumor cell lines, but not on normal cells

Cancer Lett., 175 (2002), pp. 1725

Corresponding author. Tel.: +358 9 191 59170; fax: +358 9 191 59556.

Copyright 2006 Elsevier B.V. All rights reserved.

About ScienceDirect

Contact and support

Information for advertisers

Terms and conditions

Privacy policy

Copyright 2014 Elsevier B.V. except certain content provided by third parties. ScienceDirect is a registered trademark of
Elsevier B.V.
Cookies are used by this site. To decline or learn more, visit our Cookies page
Switch to Mobile Site

Recommended articles

1.
1.
2.

Betulin isolation from birch bark by vacuum and atmospheric sublimation. A

thermogravimetric study
2003, Thermochimica Acta

more

2.
1.

The betulinic acid production from betulin through biotransformation by fungi

2.

2009, Enzyme and Microbial Technology

more

3.
1.

Apoptotic activity of betulinic acid derivatives on murine melanoma B16 cell line

2.

2004, European Journal of Pharmacology

more

4.

View more articles

Citing articles (193)
Related book content