Introduction/Overview
Natural products, as an important source of drug discovery, play an irreplaceable role in the long history of human fight against diseases. One of the core tasks of modern medicinal chemistry and pharmacology research is to isolate, identify, and elucidate the biological functions of active ingredients from traditional herbs. Among numerous natural products with biological activity, diterpenes derived from Euphorbiaceae plants have attracted much attention due to their structural diversity and significant pharmacological activities, especially anti-tumor activity. Lathyrol, as a typical Lathyrane type diterpene, is derived from the traditional Chinese medicine "Lathyrol"（Euphorbia lathyris L. One of the main active ingredients isolated from (). Its unique 5/11/3 tricyclic diterpene skeleton not only endows it with a complex chemical structure, but also indicates its potential for interaction with various biological targets.
In recent years, with the deepening of research on diterpenoid alcohols from Qianjin, their anti-tumor activity, especially their effect on liver cancer, has become a research hotspot. Liver cancer is a malignant tumor with high incidence rate and mortality worldwide. Its pathogenesis is complex, involving abnormal activation of multiple signaling pathways. Although existing treatment methods such as surgery, chemotherapy, and targeted therapy have shown some efficacy, issues such as drug resistance and toxic side effects remain severe. Therefore, the search for efficient and low toxicity new candidate drugs for liver cancer has important clinical significance. Due to its inhibitory effects on the proliferation, migration, and invasion of liver cancer cells, as well as its ability to regulate apoptosis related pathways, quercetin has shown great potential as a lead compound for anti liver cancer treatment. This article will provide a systematic review of the research progress of diterpenoid alcohols from the aspects of chemical structure, plant origin, pharmacological activity, mechanism of action, medicinal properties, and clinical application prospects, aiming to provide comprehensive references for subsequent basic research and drug development.
Chemical structure and physicochemical properties
The chemical structure of Lathyrol belongs to the tetradecane type diterpenes, and its core skeleton is a unique tricyclic system composed of a five membered ring (A ring), an eleven membered ring (B ring), and a three membered ring (C ring) fused together. This rare 5/11/3 ring structure is a characteristic feature of the sesquiterpenes in Euphorbiaceae plants. Specifically, its structural parent nucleus is 5,11,3-tricyclic diterpenes, usually with hydroxyl or ester substituents attached at positions C-3, C-5, C-15, etc. The molecular formula of diterpenoid alcohol in Qianjin is C ₂₀ H ∝₀ O ₄, with a molecular weight of 334.4560 g/mol. Its structure contains multiple chiral centers, endowing the molecule with a specific stereoconfiguration, which is crucial for its precise recognition with biological targets.
From the perspective of physical and chemical properties, diterpenoid alcohols from Qianjin exhibit a certain degree of lipophilicity. Its lipid water partition coefficient (LogP) is 2.0790, indicating that the compound has moderate partition ability in both lipid and water phases, which is beneficial for its penetration into biofilms. The topological polar surface area (TPSA) is 77.7600 Å ², which is typically associated with the oral absorption and blood-brain barrier penetration ability of the compound. According to the Lipinski Five Rules, compounds with TPSA less than 140 Å ² typically have good oral bioavailability, and the TPSA value of quercetin meets this standard. Its water solubility is 0.2993 mg/mL, which belongs to the category of slight solubility. This may limit the development of its formulation to some extent, but it can be improved through prodrug design or novel delivery systems such as liposomes and nanoparticles. It is worth noting that computer simulations predict that quercetin has high blood-brain barrier (BBB) penetration ability, suggesting its potential activity in central nervous system diseases such as brain tumors, but caution should also be exercised about potential central nervous system toxicity. In addition, the predicted results showed that it does not have hERG (human Ether - à - go Related Gene) inhibitory activity (hERG inhibition: No), which reduces its risk of causing cardiac QT interval prolongation and arrhythmia, and is a favorable safety signal. The Ames test result was 0.0, indicating that the compound did not exhibit significant mutagenicity in the bacterial recovery mutation test, suggesting that its genetic toxicity is relatively low.
Plant sources and extraction methods
The main source of diterpenoid alcohols in Euphorbiaceae is the plant Euphorbiaceae, Euphorbiaceae（Euphorbia lathyris L.）， Also known as Xusuizi, Daowanhua, etc. Qianjinzi is an annual or biennial herbaceous plant native to Europe and the Mediterranean region. It has been widely introduced and cultivated in China, Japan, India, and other places. In traditional Chinese medicine theory, its seed (Qianjinzi) is believed to have the effects of promoting diuresis and reducing swelling, breaking blood and resolving symptoms. It is commonly used to treat conditions such as edema, phlegm retention, stagnation, bloating, blood stasis, and meridian closure. Modern pharmacological research has shown that the various pharmacological activities of Qianjingzi, including anti-tumor, antiviral, anti-inflammatory, etc., are mainly attributed to the diterpenoid compounds it contains, among which Qianjingzi diterpenoid alcohols and their ester derivatives are important active components.
The extraction of diterpenoid alcohols from Qianjin gold is usually carried out using solvent extraction method. Due to its lipophilicity, commonly used extraction solvents include methanol, ethanol, ethyl acetate, chloroform, etc. The classic extraction process is as follows: first, the dried mille gold seeds are crushed and degreased with petroleum ether or n-hexane to remove a large amount of oil and wax. After degreasing, the drug residue should be soaked or refluxed multiple times with polar solvents (such as 95% ethanol or methanol) for extraction. After the extraction solution is concentrated under reduced pressure, the total extract is obtained. Subsequently, the total extract was preliminarily separated using liquid-liquid extraction method. For example, the extract was dispersed in water and extracted sequentially with petroleum ether, ethyl acetate, and n-butanol. Chitose diterpenoid alcohols are usually enriched in the ethyl acetate or chloroform extraction sites.
Further separation and purification require the use of various chromatographic techniques. Silica gel column chromatography is the most commonly used method, usually using gradient elution (such as petroleum ether ethyl acetate or chloroform methanol system) to separate the extraction site. For diterpenoids with similar structures, it may be necessary to combine more refined separation methods such as reverse phase silica gel column chromatography (such as ODS), gel column chromatography (such as Sephadex LH-20), and preparative high performance liquid chromatography (Pre HPLC) to obtain high-purity diterpenoid alcohol monomer of Semen Euphorbiae. In recent years, with the promotion of the concept of green chemistry, some new extraction techniques such as supercritical fluid extraction (SFE), ultrasound assisted extraction (UAE), and microwave-assisted extraction (MAE) have also been attempted for the extraction of diterpenoid components in Qianjin gold. These methods have the advantages of high extraction efficiency, short time, and low solvent consumption, showing good application prospects.
Pharmacological activity research
The pharmacological activity research of quercetin mainly focuses on its anti-tumor effect, especially its inhibitory effect on liver cancer. In addition, its anti-inflammatory and antiviral activities are gradually being revealed.
1. Antitumor activity
- Inhibition effect on liver cancer cells Numerous in vitro experiments have confirmed that quercetin can significantly inhibit the proliferation of various human liver cancer cell lines, such as HepG2, Huh7, SMMC-7721, etc. Its effect is dose-dependent and time-dependent. Through cell viability assays such as MTT or CCK-8, researchers have found that the half maximal inhibitory concentration (IC ₅₀) of quercetin is typically in the micromolar range. In addition to inhibiting proliferation, quercetin can also effectively induce apoptosis in liver cancer cells. Annexin V-FITC/PI dual staining flow cytometry analysis showed that the proportion of early and late apoptotic cells significantly increased after drug treatment. At the same time, it can also arrest the cell cycle in the G2/M or S phase, thereby inhibiting cell division.
- Inhibit migration and invasion Tumor metastasis is the main cause of poor prognosis in liver cancer patients. Scratch experiments and Transwell chamber experiments have shown that quercetin can significantly inhibit the migration and invasion ability of liver cancer cells. This effect is closely related to its regulation of matrix metalloproteinases (MMPs) expression.
- Effects on other tumor cells In addition to liver cancer, the diterpenoid alcohol of Semen Euphorbiae also shows different degrees of cytotoxicity to a variety of tumor cells, such as breast cancer, lung cancer, colon cancer, melanoma, etc. This indicates that its anti-tumor activity has a certain broad-spectrum nature.
2. Anti inflammatory activity
The inflammatory microenvironment plays a crucial role in the occurrence and development of tumors. Research has shown that quercetin can inhibit the production of nitric oxide (NO) and prostaglandin E2 (PGE2) in macrophages (such as RAW264.7 cells) induced by lipopolysaccharide (LPS), and reduce the expression of pro-inflammatory cytokines such as tumor necrosis factor - α (TNF - α), interleukin-6 (IL-6), and interleukin-1 β (IL-1 β). Its anti-inflammatory mechanism may be related to the inhibition of the activation of the nuclear factor kappa B (NF - κ B) signaling pathway.
3. Antiviral activity
There are research reports that quercetin and its derivatives have inhibitory effects on certain viruses. For example, they may exhibit anti HIV, anti influenza virus and other activities by interfering with the virus replication cycle or inhibiting the synthesis of viral proteins. However, there is relatively little research in this area, and its antiviral spectrum and specific mechanisms need further clarification.
Mechanism of action and molecular targets
The molecular mechanism of the anti liver cancer activity of triterpenoids in Qianjin gold is the result of the synergistic effect of multiple targets and pathways. Based on existing research, its mechanism of action mainly involves the following aspects and is closely related to the given target:
1. Regulating the apoptotic signaling pathway
- Targeting the BCL2 family BCL2 (B-cell lymphoma 2) is a key anti apoptotic protein that is highly expressed in various tumor cells. Qianjingzi diterpenoid alcohol can downregulate the expression of BCL2 in liver cancer cells, while upregulating the expression of pro apoptotic protein BAX, leading to an increase in the BAX/BCL2 ratio, thereby promoting mitochondrial release of cytochrome c, activating the Caspase cascade reaction, and ultimately inducing cell apoptosis.
- Inhibition of STAT3 signaling pathway：STAT3（Signal transducer and activator of transcription 3） It is an important transcription factor whose sustained activation is closely related to the proliferation, survival, angiogenesis, and immune escape of tumor cells. Chitose diterpenoid alcohol can inhibit the phosphorylation of STAT3 (Tyr705 site), thereby blocking its nuclear translocation and transcriptional activity. The inactivation of STAT3 further downregulates its downstream target genes, including anti apoptotic proteins (such as BCL2, Survivin), cell cycle regulatory proteins (such as Cyclin D1), and angiogenic factors (such as VEGF), thereby synergistically inhibiting tumor growth.
2. Inhibit tumor cell proliferation and induce cycle arrest
- Inhibition of PI3K/AKT/mTOR pathway：PIK3CA（Phosphatidylinositol-4,5-bisphosphate 3-kinase catalytic subunit alpha） The catalytic subunit encoding PI3K is commonly mutated or overactivated in liver cancer. Qianjingzi diterpenoid alcohol can inhibit the activity of PI3K, thereby reducing the phosphorylation level of downstream AKT. Inhibition of the AKT signaling pathway weakens its activation of mTOR, thereby inhibiting protein synthesis and cell proliferation. Meanwhile, the inactivation of AKT can also release the inhibition of FOXO transcription factors and promote the expression of pro apoptotic genes.
- Regulating the MAPK/ERK pathway MAPK1 (Mitogen activated protein kinase 1, ERK2) is a key component of the RAS-RAF-MEK-ERK signaling pathway, regulating cell proliferation and differentiation. Qianjin diterpenoid alcohol may have a bidirectional regulatory effect on the activity of MAPK1, manifested in some studies by inhibiting its phosphorylation, thereby blocking the transmission of proliferation signals.
3. Inhibit tumor metastasis and invasion
- Inhibition of MMP9 expression Matrix metalloproteinase-9 (MMP9) is an enzyme that can degrade the extracellular matrix (ECM), and its high expression is positively correlated with the invasion and metastasis ability of tumors. Qianjingzi diterpenoid alcohol can reduce the invasion and migration ability of liver cancer cells by inhibiting the activity of transcription factors such as NF - κ B or AP-1, downregulating the mRNA and protein levels of MMP9.
- Targeting HIF1A and RELA HIF1A (Hypoxia inducible factor 1-alpha) is a key transcription factor for cells to adapt to low oxygen environments, often activated in solid tumors, promoting angiogenesis and glycolysis. RELA (v-rel retina viral homolog A, also known as p65) is a subunit of the NF - κ B complex. Qianjingzi diterpenoid alcohol may inhibit tumor angiogenesis and metastasis by suppressing the stability of HIF1A and/or nuclear translocation of RELA, downregulating downstream target genes such as VEGF and MMP9.
4. Inducing DNA damage and telomerase inhibition
- Inhibit TOP1 activity TOP1 is an enzyme essential for DNA replication and transcription. Qianjingzi diterpenoid alcohol may act as a TOP1 inhibitor, stabilizing the TOP1-DNA cleavage complex, hindering DNA reconnection, causing DNA damage, and ultimately inducing cell apoptosis.
- Inhibition of TERT expression TERT (Telomerase Reverse Transcriptase) is the catalytic subunit of telomerase, and its reactivation is a key step for cells to acquire infinite proliferation ability (immortalization). Research has shown that quercetin can downregulate the expression of TERT in liver cancer cells, inhibit telomerase activity, shorten telomere length, and induce cell aging and apoptosis.
5. Regulating other signal nodes
- Targeting IKBKB IKBKB (inhibitor of nuclear factor kappa-B kinase subunit beta, IKK β) is a key kinase that activates the NF - κ B signaling pathway. Qianjingzi diterpenoid alcohol may inhibit the activity of IKBKB, prevent the phosphorylation and degradation of I κ B α, and thus retain NF - κ B (RELA/p50) complexes in the cytoplasm, inhibiting their nuclear translocation and transcription of pro-inflammatory and pro survival genes.
In summary, the diterpenoid alcohol of Qianzi gold forms a complex regulatory network by simultaneously acting on multiple key targets such as BCL2, STAT3, TOP1, HIF1A, RELA, MAPK1, IKBKB, TERT, PIK3CA, and MMP9, exerting its anti liver cancer activity at multiple levels including inducing apoptosis, inhibiting proliferation, blocking the cell cycle, anti metastasis, and anti angiogenesis. This multi-target mode of action is its advantage, but it also increases the complexity of mechanism research.
Evaluation of drug properties and pharmacokinetics
To develop natural products into clinical drugs, a systematic evaluation of their drug like and pharmacokinetic (ADME) properties is necessary. Based on the provided parameters and existing literature, a preliminary analysis was conducted on the pharmacological properties of diterpenoid alcohols from Qianjin.
1. Physical and chemical properties and the "Lipinski Five Rules"
The molecular weight of diterpenoid alcohol in Qianzi gold is 334.46 (<500), with a LogP of 2.079 (<5). The number of hydrogen bond donors (usually 3-4 hydroxyl groups in the structure,<5) and hydrogen bond acceptors (4 oxygen atoms,<10) are present. These parameters fully comply with the Lipinski Five Rules, indicating its good oral drug potential. The TPSA is 77.76 Å ², which is also within the ideal range (<140 Å ²), indicating its good intestinal absorption capacity.
2. Pharmacokinetic prediction
- absorb As mentioned earlier, its good lipid solubility and moderate polarity are beneficial for oral absorption. However, low water solubility (0.2993 mg/mL) may be a limiting factor, which could lead to low bioavailability. Improvements are expected through the production of salts, prodrugs, or the use of nano formulations.
- distribution Prediction shows that quercetin has high blood-brain barrier penetration ability. This is advantageous for treating brain tumors or central nervous system diseases, but it may also pose a risk of central neurotoxicity, which requires further experimental verification. The binding rate between it and plasma proteins is still unclear and needs further research.
- Metabolism As a diterpenoid compound, quercetin is likely to be metabolized by the cytochrome P450 enzyme system (CYP450) in the liver, mainly involving oxidation, reduction, and hydrolysis reactions. Its metabolites may have different activities or toxicity. At present, there is very limited information about its specific metabolic pathways and enzymes.
- excretion It is expected that diterpenoid alcohols and their metabolites will be mainly excreted through bile and/or urine.
3. Security assessment
- HERG inhibition: Negative. This is a very favorable safety indicator that reduces the risk of cardiac toxicity.
- Ames test Negative (0.0). Preliminary indications suggest that it is non mutagenic and has a low risk of genetic toxicity.
- Other toxicities It should be noted that as a traditional Chinese medicine, Qianjinzi itself has certain toxicity (such as diarrhea and irritation). As its active ingredient, diterpenoid alcohols from Qianjin may also have certain cytotoxicity, especially at high concentrations. Its toxicity to normal cells (selectivity index) is the key to evaluating its safety. At present, research is mostly focused on tumor cells, and toxicity studies on normal liver cells (such as LO2 cells) are not yet sufficient. In addition, its potential immunotoxicity, reproductive toxicity, etc. also need to be comprehensively evaluated.
4. Existing problems and challenges
Despite exhibiting some advantages in medicinal properties, the triterpenoids of Qianjin gold still face many challenges
- Poor water solubility Restricted its formulation development and bioavailability.
- Metabolic stability The diterpene skeleton may be easily metabolized and inactivated, resulting in a short half-life in the body.
- Complex mechanism of action Although multi-target effects have obvious advantages, they also increase the risk of off target effects and toxic side effects.
- Low natural content Extracting and isolating from plants is costly and difficult to meet the demands of large-scale production. The development of chemical synthesis or semi synthesis routes is the key to solving the source problem.
Clinical application prospects and prospects
As a natural product with multi-target anti liver cancer activity, quercetin has broad clinical application prospects, but also faces many challenges. Future research should focus on the following aspects:
1. In depth mechanism research and target validation
Although multiple potential targets have been identified, the direct target protein (i.e., its "receptor") of quercetin has not been clearly defined. The core task of elucidating its mechanism of action is to use chemical proteomics techniques such as drug affinity reaction target stability DARTS, thermal transfer analysis CETSA, and photoaffinity labeling to identify and validate the protein targets it directly binds to. At the same time, it is necessary to use techniques such as gene knockout/knock in and RNA interference to verify the necessity of key targets such as BCL2, STAT3, and PIK3CA in the anti liver cancer effect of quercetin at the cellular and animal levels.
2. Research on structural optimization and structure-activity relationship
Systematic structural modification using diterpenoid alcohols as lead compounds is an effective way to enhance their activity, improve pharmacokinetic properties, and reduce toxicity. For example:
- Improve water solubility Introducing hydrophilic groups (such as phosphate groups and amino acid ester groups) onto the mother nucleus, or preparing prodrugs.
- Enhance activity Esterification, etherification, and other modifications were performed on hydroxyl groups at C-3, C-5, C-15, and other sites to explore the effects of different substituents on activity.
- Reduce toxicity Improving selectivity towards tumor cells and reducing toxicity towards normal cells through structural modifications.
3. Development of new drug delivery systems
Given its poor water solubility and potential toxicity, it is crucial to develop new delivery systems using modern pharmaceutical technologies. For example:
- Liposomes/Nanoparticles Encapsulating quercetin in liposomes or polymer nanoparticles can enhance its water solubility, prolong in vivo circulation time, and achieve passive tumor targeting through EPR effect.
- Targeted delivery system Conjugate targeting ligands (such as antibodies or peptides against liver cancer specific antigens) on the surface of nanoparticles to achieve active targeting, further improve therapeutic efficacy, and reduce systemic toxicity.
4. Combination therapy strategy
Given the complexity of liver cancer, a single drug often fails to achieve the desired effect. The multi-target properties of quercetin make it an ideal candidate for combination therapy. You can explore its relationship with:
- chemotherapeutic drugs The combination of drugs such as sorafenib, cisplatin, and doxorubicin enhances therapeutic efficacy through synergistic effects and may reduce the dosage and toxicity of chemotherapy drugs.
- targeted drug Combination therapy with drugs such as sorafenib and lenvatinib, while blocking multiple signaling pathways, overcomes or delays the development of drug resistance.
- Immune checkpoint inhibitors The combination of PD-1/PD-L1 antibodies enhances anti-tumor immune response by regulating the tumor microenvironment.
5. Sustainable development of resources
The natural sources of diterpenoid alcohols from Qianjin are limited. Therefore, it is necessary to develop sustainable production methods:
- biosynthesis Analyze the biosynthetic pathways of diterpenoid compounds in Qianjin gold, clone key enzyme genes, and utilize synthetic biology techniques (such as constructing heterologous synthetic pathways in yeast or Escherichia coli) to achieve efficient and green biomanufacturing.
- chemical synthesis Develop concise and efficient chemical synthesis or semi synthesis routes to provide stable raw material supply for drug development.
Conclusion
As the core active ingredient of traditional Chinese medicine, quercetin, with its unique 5/11/3 tricyclic diterpenoid skeleton and significant anti liver cancer activity, has become a new star in the field of natural product drug development. Existing research has revealed the complex mechanism by which it exerts synergistic anti-tumor effects in inducing apoptosis, inhibiting proliferation, anti metastasis, and anti angiogenesis by regulating multiple key targets such as BCL2, STAT3, PIK3CA, MMP9, TOP1, and HIF1A. Its preliminary pharmacological evaluation shows good drug like properties and low risks of cardiac and genetic toxicity, but poor water solubility, metabolic instability, and potential off target toxicity remain the main obstacles to its clinical translation.
Looking ahead to the future, the research on diterpenoid alcohols in Qianjin is at a critical stage of transition from "discovery" to "development". By combining modern medicinal chemistry, chemical biology, nanopharmaceuticals, and systems biology, and conducting in-depth research on structure-activity relationships, direct target discovery, construction of novel delivery systems, and exploration of combination therapy strategies, it is expected to transform this natural product into a highly effective and low toxicity candidate drug for liver cancer, bringing new therapeutic hope to liver cancer patients. At the same time, breakthroughs in the analysis of its biosynthetic pathway and chemical synthesis methods will provide fundamental guarantees for solving its source problem. The research process of triterpenoids in Qianjin gold is not only a story of the development of a single compound, but also a vivid example of the deep integration of traditional Chinese medicine wisdom and modern pharmaceutical science.
