A set of fifteen triterpenoid pyrazines and pyridines was prepared from parent triterpenoid 3-oxoderivatives (betulonic acid, dihydrobetulonic acid, oleanonic acid, moronic acid, ursonic acid, heterobetulonic acid, and allobetulone). Cytotoxicity of all compounds was tested in eight cancer and two non-cancer cell lines. Evaluation of the structure-activity relationships revealed that the triterpenoid core determined whether the final molecule is active or not, while the heterocycle is able to increase the activity and modulate the specificity. Five compounds (1b, 1c, 2b, 2c, and 8) were found to be preferentially and highly cytotoxic (IC50 ≈ 1 μM) against leukemic cancer cell lines (CCRF-CEM, K562, CEM-DNR, or K562-TAX). Surprisingly, compounds 1c, 2b, and 2c are 10-fold more active in multidrug-resistant leukemia cells (CEM-DNR and K562-TAX) than in their non-resistant analogs (CCRF-CEM and K562). Pharmacological parameters were measured for the most promising candidates and two types of prodrugs were synthesized: 1) Sugar-containing conjugates, most of which had improved cell penetration and retained high cytotoxicity in the CCRF-CEM cell line, unfortunately, they lost the selectivity against resistant cells. 2) Medoxomil derivatives, among which compounds 26-28 gained activities of IC50 0.026-0.043 μM against K562 cells. Compounds 1b, 8, 21, 22, 23, and 24 were selected for the evaluation of the mechanism of action based on their highest cytotoxicity against CCRF-CEM cell line. Several experiments showed that the majority of them cause apoptosis via the mitochondrial pathway. Compounds 1b, 8, and 21 inhibit growth and disintegrate spheroid cultures of HCT116 and HeLa cells, which would be important for the treatment of solid tumors. In summary, compounds 1b, 1c, 2b, 2c, 24, and 26-28 are highly and selectively cytotoxic against cancer cell lines and were selected for future in vivo tests and further development of anticancer drugs.
- MeSH
- antitumorózní látky fytogenní * farmakologie MeSH
- antitumorózní látky * farmakologie MeSH
- chemorezistence MeSH
- HeLa buňky MeSH
- lidé MeSH
- membránový potenciál mitochondrií MeSH
- nádorové buněčné linie MeSH
- prekurzory léčiv * farmakologie MeSH
- pyraziny farmakologie MeSH
- pyridiny farmakologie MeSH
- triterpeny * farmakologie MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
A set of new substituted dienes were synthesized from betulinic acid by its oxidation to 30-oxobetulinic acid followed by the Wittig reaction. Cytotoxicity of all compounds was tested in vitro in eight cancer cell lines and two noncancer fibroblasts. Almost all dienes were more cytotoxic than betulinic acid. Compounds 4.22, 4.30, 4.33, 4.39 had IC50 below 5 μmol/L; 4.22 and 4.39 were selected for studies of the mechanism of action. Cell cycle analysis revealed an increase in the number of apoptotic cells at 5 × IC50 concentration, where activation of irreversible changes leading to cell death can be expected. Both 4.22 and 4.39 led to the accumulation of cells in the G0/G1 phase with partial inhibition of DNA/RNA synthesis at 1 × IC50 and almost complete inhibition at 5 × IC50. Interestingly, compound 4.39 at 5 × IC50 caused the accumulation of cells in the S phase. Higher concentrations of tested drugs probably inhibit more off-targets than lower concentrations. Mechanisms disrupting cellular metabolism can induce the accumulation of cells in the S phase. Both compounds 4.22 and 4.39 trigger selective apoptosis in cancer cells via intrinsic pathway, which we have demonstrated by changes in the expression of the crucial apoptosis-related protein. Pharmacological parameters of derivative 4.22 were superior to 4.39, therefore 4.22 was the finally selected candidate for the development of anticancer drug.
- MeSH
- alkadieny chemická syntéza chemie farmakologie MeSH
- antitumorózní látky chemická syntéza chemie farmakologie MeSH
- apoptóza účinky léků MeSH
- buněčný cyklus účinky léků MeSH
- kultivované buňky MeSH
- léky antitumorózní - screeningové testy MeSH
- lidé MeSH
- molekulární struktura MeSH
- pentacyklické triterpeny chemie farmakologie MeSH
- proliferace buněk účinky léků MeSH
- psi MeSH
- vztah mezi dávkou a účinkem léčiva MeSH
- vztahy mezi strukturou a aktivitou MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- psi MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
A set of 41 glycosidic conjugates of pentacyclic triterpenes was synthesized in order to improve the solubility of highly cytotoxic parent compounds. Their in vitro cytotoxic activity was evaluated in 25 cancer cell lines and 2 noncancer fibroblasts. Fifteen compounds had high cytotoxicity on the T-lymphoblastic leukemia cell line CCRF-CEM and 6 of them were active in multiple cell lines of various histogenic origin and not toxic in fibroblasts. Compound 11a had IC50 of 0.64 μM in CCRF-CEM cells, 0.60 μM in K-562 cells, and 0.37 μM in PC-3 cells; compound 12a had IC50 of 0.64 μM in CCRF-CEM cells and 0.71 μM in SW620 cells; compound 17b had IC50 of 0.86 μM in HCT116 cells and 0.92 μM in PC-3 cells. Compounds 11b and 12b were slightly less active than the previously mentioned derivatives; however, their solubility was significantly better, and therefore they were selected for the in vivo evaluation of the pharmacokinetic profile in mice. In both compounds, the maximum concentration in plasma was achieved very rapidly-the highest level in plasma was found 1 h after administration (22.2, respectively, 6.4 μM). For compound 12b, the resorption was followed with fast elimination, and 12 h after administration, the compound was not detected in plasma. In contrast, compound 11b was eliminated more slowly; it was still present in plasma after 12 h, but its concentration dropped below the detection limit after 24 h. The elimination half-time determined for compound 11b was 2.4 h and for compound 12b just about 1.4 h. These values are reasonable for further drug development.
- MeSH
- antitumorózní látky chemie farmakologie MeSH
- deoxycukry chemie MeSH
- glykosidy chemie MeSH
- léky antitumorózní - screeningové testy MeSH
- lidé MeSH
- myši MeSH
- nádorové buňky kultivované MeSH
- nádory farmakoterapie metabolismus patologie MeSH
- proliferace buněk * MeSH
- tkáňová distribuce MeSH
- triterpeny chemie farmakokinetika farmakologie MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- myši MeSH
- ženské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
To improve the properties of cytotoxic triterpenoid acids 1-5, a large set of hydrophilic esters was synthesized. We choose betulinic acid (1), dihydrobetulinic acid (2), 21-oxoacid 3 along with highly active des-E lupane acids 4 and 5 as a model set of compounds for esterification of which the properties needed to be improved. As ester moieties were used - methoxyethanol and 2-(2-methoxyethoxy)ethanol and glycolic unit (type a-d), pyrrolidinoethanol, piperidinoethanol and morpholinoethanol (type f-h), and monosaccharide groups (type i-l). As a result, 56 triterpenic esters (49 new compounds) were obtained and their cytotoxicity on four cancer cell lines and normal human fibroblasts was tested. All new compounds were fully soluble at all tested concentrations, which used to be a problem of the parent compounds 1 and 2. 16 compounds had IC50 < 10 μM on at least one cancer cell line, 12 compounds had cytotoxicity of <10 μM against at least three of four tested cancer cell lines. The highest activity was found for compound 3c (1.8 μM on MCF7, 2.8 μM on HeLa, and 1.6 μM on G-361 cells) which also had no toxicity on non-cancerous BJ fibroblasts at the highest tested concentration (50 μM). High selective cytotoxicity was also found in compounds 1k, 2k, 3c, and 3i that are ideal candidates for drug development. Therefore, more studies to identify the mechanism of action were performed in case of 1k, 3c, and 3g such as effects on cell cycle and apoptosis. It was found that compounds 3c and 3g can induce apoptosis via caspase-3 activation and modulation of protein Bcl-2 in G-361 cells. In conclusion, compounds 1k, 3c, and 3g show high and selective cytotoxicity, therefore they are significantly better candidates for anti-cancer drug development than the parent acids 1-5.
A total of 41 new triterpenoids were prepared from allobetulone, methyl betulonate, methyl oleanonate, and oleanonic acid to study their influence on cancer cells. Each 3-oxotriterpene was brominated at C2 and substituted with thiocyanate; subsequent cyclization with the appropriate ammonium salts gave N-substituted thiazoles. All compounds were tested for their in vitro cytotoxic activity on eight cancer cell lines and two non-cancer fibroblasts. 2-Bromoallobetulone (2 b) methyl 2-bromobetulonate (3 b), 2-bromooleanonic acid (5 b), and 2-thiocyanooleanonic acid (5 c) were best, with IC50 values less than 10 μm against CCRF-CEM cells (e.g., 3 b: IC50 =2.9 μm) as well as 2'-(diethylamino)olean-12(13)-eno[2,3-d]thiazole-28-oic acid (5 f, IC50 =9.7 μm) and 2'-(N-methylpiperazino)olean-12(13)-eno[2,3-d]thiazole-28-oic acid (5 k, IC50 =11.4 μm). Compound 5 c leads to the accumulation of cells in the G2 phase of the cell cycle and inhibits RNA and DNA synthesis significantly at 1×IC50 . The G2 /M cell-cycle arrest probably corresponds to the inhibition of DNA/RNA synthesis, similar to the mechanism of action of actinomycin D. Compound 5 c is new, active, and nontoxic; it is therefore the most promising compound in this series for future drug development. Methyl 2-bromobetulonate (3 b) and methyl 2-thiocyanometulonate (3 c) were found to inhibit nucleic acid synthesis only at 5×IC50 . We assume that in 3 b and 3 c (unlike in 5 c), DNA/RNA inhibition is a nonspecific event, and an unknown primary cytotoxic target is activated at 1×IC50 or lower concentration.
- MeSH
- antitumorózní látky chemická syntéza chemie toxicita MeSH
- apoptóza účinky léků MeSH
- kontrolní body fáze G2 buněčného cyklu účinky léků MeSH
- kontrolní body M fáze buněčného cyklu účinky léků MeSH
- kyselina olenalová analogy a deriváty chemie MeSH
- léky antitumorózní - screeningové testy MeSH
- lidé MeSH
- nádorové buněčné linie MeSH
- thiazoly chemická syntéza chemie toxicita MeSH
- triterpeny chemie MeSH
- vztahy mezi strukturou a aktivitou MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
Lupane derivatives containing an electronegative substituent in the position 2 of the skeleton are often cytotoxic, however, the most active compounds are not selective enough. To further study the influence of a substituent in the position 2 in lupane and 18α-oleanane derivatives on their biological properties, we prepared a set of 38 triterpenoid compounds, 19 of them new, most of them substituted in the position 2. From betulin, we obtained 2-bromo dihydrobetulonic acid and 2-bromo allobetulon and their substitutions yielded derivatives with various substituents in the position 2 such as amines, amides, thiols, and thioethers. Nitration of allobetulon and dihydrobetulonic acid gave 2-nitro and 2,2-dinitro derivatives. Fifteen derivatives had IC50 < 50 μM on a chemosensitive CCRF-CEM (acute lymphoblastic leukemia) cell line and were tested on another seven cancer cell lines including resistant and two non-cancer lines. 2-Amino allobetulin had IC50 4.6 μM and caused significant block of the tumor cells in S and slightly in G2/M transition and caused strong inhibition of DNA and RNA synthesis at 5 × IC50. 2-Amino allobetulin is the most active derivative of 18α-oleanane skeletal type prepared in our research group to date.
- MeSH
- antitumorózní látky chemie farmakologie MeSH
- cytotoxiny chemie farmakologie MeSH
- inhibiční koncentrace 50 MeSH
- kontrolní body buněčného cyklu účinky léků MeSH
- kyselina olenalová analogy a deriváty chemie farmakologie MeSH
- léky antitumorózní - screeningové testy MeSH
- lidé MeSH
- nádorové buněčné linie MeSH
- nukleové kyseliny antagonisté a inhibitory biosyntéza MeSH
- triterpeny chemie farmakologie MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
7-Oxygenated metabolites of dehydroepiandrosterone (DHEA) are known for their neuroprotective and immunomodulatory properties. These neuroactive steroids are currently predominately analysed by mass spectrometry, for which the use of internal deuterated standards is necessary. The aim of this study was to synthesize the deuterated derivatives of 7α-hydroxy-DHEA and 7-oxo-DHEA and test them in liquid chromatography-tandem mass spectrometry (LC-MS/MS) in order to enhance the performance characteristics of this method. Here we report the synthesis of 3α deuterium-labelled 7α-hydroxy-DHEA and 7-oxo-DHEA. Deuterium was introduced into the 3α position by reduction of the corresponding 3-ketone with a protected 17-carbonyl group using NaBD4. Our new procedure allows the easier synthesis of deuterated steroid labelled compounds. The use of these deuterated steroids enabled us to improve the human plasma LC-MS/MS analysis of 7α-hydroxy-DHEA and 7-oxo-DHEA in terms of sensitivity, precision and recovery.
To better understand the mechanism of action of antitumor triterpenes, we are developing methods to identify their molecular targets. A promising method is based on combination of quantitative proteomics with SILAC and uses active compounds anchored to magnetic beads via biotin-streptavidin interaction. We developed a simple and fast solid-phase synthetic technique to connect terpenes to biotin through a linker. Betulinic acid was biotinylated from three different conjugation sites for use as a standard validation tool since many molecular targets of this triterpene are already known. Then, a set of four other cytotoxic triterpenoids was biotinylated. Biotinylated terpenes were similarly cytotoxic to their nonbiotinylated parents, which suggests that the target identification should not be influenced by linker or biotin. The developed solid-phase synthetic approach is the first attempt to use solid-phase synthesis to connect active triterpenes to biotin and is applicable as a general procedure for routine conjugation of triterpenes with other molecules of choice.
- MeSH
- antitumorózní látky chemická syntéza chemie farmakologie MeSH
- biotin chemická syntéza chemie farmakologie MeSH
- biotinylace MeSH
- lidé MeSH
- nádorové buněčné linie MeSH
- nádory farmakoterapie MeSH
- techniky syntézy na pevné fázi MeSH
- triterpeny chemická syntéza chemie farmakologie MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
In this article, we describe the preparation and cytotoxic properties of a small focused library of lupane and 18α-oleanane triterpenoids that contain a combination of two structural motifs known to enhance the biological activities. First, we introduced two fluorine atoms to position 2 of the skeleton. Second, we synthesized a set of hemiester prodrugs, which were intended to increase the solubility and activity. Starting from betulin, we obtained two hydroxyketones (derivatives of dihydrobetulinic acid and allobetulin) and their fluorination using DAST provided 2,2-difluoro-3-oxo-compounds as the main products. Then the 3-oxo group in each derivative was reduced by NaBH4 to obtain 3β-hydroxy compounds suitable for modifying by various hemiesters. We prepared 21 compounds, 11 of them new, their cytotoxicity was tested on T lymphoblastic leukemia CCRF-CEM cells first and the most active derivatives were selected for screening on another six tumor and two non-tumor cell lines. All of them showed selectivity against cancer lines with therapeutic index between 2 and 8. All hemiesters had activity in the same range as the free hydroxyl derivatives and they would be suitable prodrugs for future in vivo experiments. Interestingly, all hemiesters of 2,2-difluorodihydrobetulonic acid had higher activity against p53 knock-out p53-/- cancer cell line than against the non-mutated analog. In active derivatives, the cell cycle was analyzed by flow cytometry and several compounds slowed down cell cycle progression through G0/G1 or S-phase.
- MeSH
- antitumorózní látky fytogenní chemická syntéza chemie farmakologie MeSH
- apoptóza účinky léků MeSH
- buněčný cyklus účinky léků MeSH
- fibroblasty účinky léků MeSH
- fluorované uhlovodíky chemická syntéza chemie farmakologie MeSH
- léky antitumorózní - screeningové testy MeSH
- lidé MeSH
- molekulární konformace MeSH
- nádorové buňky kultivované MeSH
- proliferace buněk účinky léků MeSH
- triterpeny chemie MeSH
- vztah mezi dávkou a účinkem léčiva MeSH
- vztahy mezi strukturou a aktivitou MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Triterpenoids are natural compounds with a variety of biological activities and are usually produced by plants as secondary metabolites. In recent decades, scientists focused on the properties of triterpenoids have discovered many activities, such as antitumor, antiviral, antimicrobial, anti-inflammatory and others. Thousands of new triterpenoids with various skeletal modifications have now been synthesized. One of the most important modifications is the formation of a new heterocyclic ring. The simple fact that the vast majority of currently used drugs are heterocyclic compounds has encouraged a lot of researches to synthesize analogous triterpenoid derivatives in order to find new molecules with higher activities and consequently optimize their pharmacological profile. The biological properties of triterpenoid heterocycles are very promising and many of them have been studied, especially as antitumor agents. This review is mainly focused on the synthesis of various types of nitrogen and occasionally sulfur-containing heterocyclic triterpenoids for their potential use as future drugs in medicine and in addition discusses their overall biological activities.
- MeSH
- antiflogistika MeSH
- antiinfekční látky MeSH
- antitumorózní látky MeSH
- antivirové látky MeSH
- dusík chemie MeSH
- heterocyklické sloučeniny * chemická syntéza chemie farmakologie MeSH
- molekulární struktura MeSH
- síra chemie MeSH
- triterpeny * chemická syntéza chemie farmakologie MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- přehledy MeSH
