Comparative Proteomic Analysis of Drug Trichosanthin Addition to BeWo Cell Line.

Trichosanthin (TCS) is a traditional Chinese herbal medicine used to treat some gynecological diseases. Its effective component has diverse biological functions, including antineoplastic activity. The human trophoblast cell line BeWo was chosen as an experimental model for in vitro testing of a drug screen for anticancer properties of TCS. The MTT method was used in this study to get a primary screen result. The result showed that 100 mM had the best IC50 value. Proteomics analysis was then performed for further investigation of the drug effect of TCS on the BeWo cell line. In this differential proteomic expression analysis, the total proteins extracted from the BeWo cell line and their protein expression level after the drug treatment were compared by 2DE. Then, 24 unique three-fold differentially expressed proteins (DEPs) were successfully identified by MALDI-TOF/TOF MS. Label-free proteomics was run as a complemental method for the same experimental procedure. There are two proteins that were identified in both the 2DE and label-free methods. Among those identified proteins, bioinformatics analysis showed the importance of pathway and signal transduction and gives us the potential possibility for the disease treatment hypothesis.

A Sixty-Year Research and Development of Trichosanthin, a Ribosome-Inactivating Protein.

Tian Hua Fen, a herbal powder extract that contains trichosanthin (TCS), was used as an abortifacient in traditional Chinese medicine. In 1972, TCS was purified to alleviate the side effects. Because of its clinical applications, TCS became one of the most active research areas in the 1960s to the 1980s in China. These include obtaining the sequence information in the 1980s and the crystal structure in 1995. The replication block of TCS on human immunodeficiency virus in lymphocytes and macrophages was found in 1989 and started a new chapter of its development. Clinical studies were subsequently conducted. TCS was also found to have the potential for gastric and colorectal cancer treatment. Studies on its mechanism showed TCS acts as an rRNA N-glycosylase (EC 3.2.2.22) by hydrolyzing and depurinating A-4324 in α-sarcin/ricin loop on 28S rRNA of rat ribosome. Its interaction with acidic ribosomal stalk proteins was revealed in 2007, and its trafficking in mammalian cells was elucidated in the 2000s. The adverse drug reactions, such as inducing immune responses, short plasma half-life, and non-specificity, somehow became the obstacles to its usage. Immunotoxins, sequence modification, or coupling with polyethylene glycerol and dextran were developed to improve the pharmacological properties. TCS has nicely shown the scientific basis of traditional Chinese medicine and how its research and development have expanded the knowledge and applications of ribosome-inactivating proteins.

The use of miR122 and its target sequence in adeno-associated virus-mediated trichosanthin gene therapy.

OBJECTIVE: Plant-derived cytotoxic transgene expression, such as trichosanthin (tcs), regulated by recombinant adeno-associated virus (rAAV) vector is a promising cancer gene therapy. However, the cytotoxic transgene can hamper the vector production in the rAAV producer cell line, human embryonic kidney (HEK293) cells. Here, we explored microRNA-122 (miR122) and its target sequence to limit the expression of the cytotoxic gene in the rAAV producer cells. METHODS: A miR122 target (122T) sequence was incorporated into the 3' untranslated region of the tcs cDNA sequence. The firefly luciferase (fluc) transgene was used as an appropriate control. Cell line HEK293-mir122 was generated by the lentiviral vector-mediated genome integration of the mir122 gene in parental HEK293 cells. The effects of miR122 overexpression on cell growth, transgene expression, and rAAV production were determined. RESULTS: The presence of 122T sequence significantly reduced transgene expression in the miR122-enriched Huh7 cell line (in vitro), fresh human hepatocytes (ex vivo), and mouse liver (in vivo). Also, the normal liver physiology was unaffected by delivery of 122T sequence by rAAV vectors. Compared with the parental cells, the miR122-overexpressing HEK293-mir122 cell line showed similar cell growth rate and expression of transgene without 122T, as well as the ability to produce liver-targeting rAAV vectors. Fascinatingly, the yield of rAAV vectors carrying the tcs-122T gene was increased by 77.7-fold in HEK293-mir122 cells. Moreover, the tcs-122T-containing rAAV vectors significantly reduced the proliferation of hepatocellular carcinoma cells without affecting the normal liver cells. CONCLUSION: HEK293-mir122 cells along with the 122T sequence provide a potential tool to attenuate the cytotoxic transgene expression, such as tcs, during rAAV vector production.

Clinical Use of Toxic Proteins and Peptides from Tian Hua Fen and Scorpion Venom.

Traditional Chinese Medicine (TCM) has been practiced in China for thousands of years. As a complementary and alternative treatment, herbal medicines that are frequently used in the TCM are the most accepted in the Western world. However, animal materials, which are equally important in the TCM practice, are not well-known in other countries. On the other hand, the Chinese doctors had documented the toxic profiles of hundreds of animals and plants thousand years ago. Furthermore, they saw the potential benefits of these materials and used their toxic properties to treat a wide variety of diseases, such as heavy pain and cancer. Since the 50s of the last century, efforts of the Chinese government and societies to modernize TCM have achieved tremendous scientific results in both laboratory and clinic. A number of toxic proteins have been isolated and their functions identified. Although most of the literature was written in Chinese, this review provide a summary, in English, regarding our knowledge of the clinical use of the toxic proteins isolated from a plant, Tian Hua Fen, and an animal, scorpion, both of which are famous toxic prescriptions in TCM.

Trichosanthin enhances sensitivity of non-small cell lung cancer (NSCLC) TRAIL-resistance cells.

Tumour necrosis factor-related apoptosis-inducing ligand (TRAIL) has a specific antitumour activity against many malignant tumours. However, more than half of lung cancer cells are resistant to TRAIL-relevant drugs. Trichosanthin (TCS) is a traditional Chinese medicine with strong inhibitive effects on various malignancies. Nevertheless, its function on TRAIL resistance has not been revealed in non-small cell lung cancer (NSCLC). To examine the molecular mechanisms of TCS-induced TRAIL sensitivity, we administrated TCS to TRAIL-resistance NSCLC cells, and found that the combination treatment of TCS and TRAIL inhibited cancer cell proliferation and invasion, and induced cell apoptosis and S-phase arrest. This combined therapeutic method regulated the expression levels of extrinsic apoptosis-associated proteins Caspase 3/8 and PARP; intrinsic apoptosis-associated proteins BCL-2 and BAX; invasion-associated proteins E-cadherin, N-cadherin, Vimentin, ICAM-1, MMP-2 and MMP-9; and cell cycle-associated proteins P27, CCNE1 and CDK2. Up-expression and redistribution of death receptors (DRs) on the cell surface were also observed in combined treatment. In conclusion, our results indicated that TCS rendered NSCLC cells sensitivity to TRAIL via upregulating and redistributing DR4 and DR5, inducing apoptosis, and regulating invasion and cell cycle related proteins. Our results provided a potential therapeutic method to enhance TRAIL-sensitivity.

Gualou Guizhi decoction reverses brain damage with cerebral ischemic stroke, multi-component directed multi-target to screen calcium-overload inhibitors using combination of molecular docking and protein-protein docking.

Stroke is a disease of the leading causes of mortality and disability across the world, but the benefits of drugs curative effects look less compelling, intracellular calcium overload is considered to be a key pathologic factor for ischemic stroke. Gualou Guizhi decoction (GLGZD), a classical Chinese medicine compound prescription, it has been used to human clinical therapy of sequela of cerebral ischemia stroke for 10 years. This work investigated the GLGZD improved prescription against intracellular calcium overload could decreased the concentration of [Ca2+]i in cortex and striatum neurone of MCAO rats. GLGZD contains Trichosanthin and various small molecular that they are the potential active ingredients directed against NR2A, NR2B, FKBP12 and Calnodulin target proteins/enzyme have been screened by computer simulation. "Multicomponent systems" is capable to create pharmacological superposition effects. The Chinese medicine compound prescriptions could be considered as promising sources of candidates for discovery new agents.

Trichosanthin attenuates vascular injury-induced neointimal hyperplasia following balloon catheter injury in rats.

Trichosanthin (TCS), isolated from the root tuber of Trichosantheskirilowii, a well-known traditional Chinese medicinal plant, belonging to the Cucurbitaceae family, was found to exhibit numerous biological and pharmacological activities including anti-inflammatory. However, the effects of TCS on arterial injury induced neointimal hyperplasia and inflammatory cell infiltration remains poorly understood. The aim of study was to examine the effectiveness of TCS on arterial injury-mediated inflammatory processes and underlying mechanisms. A balloon-injured carotid artery induced injury in vivo in rats was established as a model of vascular injury. After 1 day TCS at 20, 40, or 80 mg/kg/day was administered intraperitoneally, daily for 14 days. Subsequently, the carotid artery was excised and taken for immunohistochemical staining. Data showed that TCS significantly dose-dependently reduced balloon injury-induced neointima formation in the carotid artery model rat, accompanied by markedly decreased positive expression percentage proliferating cell nuclear antigen (PCNA). In the in vitro study vascular smooth muscle cells (VSMC) were cultured, proliferation stimulated with platelet-derived growth factor-BB (PDGF-BB) (20 ng/ml) and TCS at 1, 2, or 4 µM added. Data demonstrated that TCS inhibited proliferation and cell cycle progression of VSMC induced by PDGF-BB. Further, TCS significantly lowered mRNA expression of cyclinD1, cyclinE1, and c-fos, and protein expression levels of Akt1, Akt2, and mitogen-activated protein kinase MAPK (ERK1) signaling pathway mediated by PDGF-BB. These findings indicate that TCS inhibits vascular neointimal hyperplasia induced by vascular injury in rats by suppression of VSMC proliferation and migration, which may involve inhibition of Akt/MAPK/ERK signal pathway.

Enhanced anti-tumor activity of trichosanthin after combination with a human-derived cell-penetrating peptide, and a possible mechanism of activity.

Trichosanthin (TCS), a type I ribosome-inactivating protein (RIP-I) and renowned Chinese traditional medicine, displays a broad spectrum of biological and pharmacological properties. Particularly, its anti-tumor activity has received a great deal of attention. However, the cellular mechanism for TCS uptake varies with different tumor cell lines, leading to discrepancies in its reported ability to penetrate cells. In this study, HBD, a human derived cell-penetrating peptide (CPP), was used to improve the delivery of TCS into several types of tumor cells, including HeLa cells. Recombinant TCS (rTCS) with or without the fused HBD peptide was expressed in Escherichia coli cells and successfully purified by Ni-NTA affinity chromatography. The cellular uptake efficiency of FITC-labelled-rTCS-HBD was observed in HeLa cells and compared with the uptake efficiency of non-HBD conjugated rTCS under the same conditions using laser confocal microscopy. Moreover, the IC50 value of rTCS-HBD in the tested tumor cells was much lower than that of rTCS, indicating that HBD could efficiently deliver the rTCS into tumor cells. When compared with rTCS, rTCS-HBD induced higher rates of apoptosis in HeLa cells as analyzed by flow cytometry. Furthermore, the apoptotic events observed in HeLa cells incubated with HBD-fused rTCS included activation of Caspase-9, decrease in the Bcl-2/Bax ratio, and cleavage of PARP. These results strongly suggest the participation of mitochondria in apoptosis. This report illustrates one possible method for achieving the efficient transport of TCS into cells using a CPP as a vector, and increases the likelihood that TCS can be used in the clinic.

Trichosanthin suppresses the proliferation of glioma cells by inhibiting LGR5 expression and the Wnt/ß-catenin signaling pathway.

Studies have indicated that trichosanthin (TCS), a bioactive protein extracted and purified from the tuberous root of Trichosanthes kirilowii (a well­known traditional Chinese medicinal plant), produces antitumor effects on various types of cancer cells. However, the effects of TCS on glioma cells are poorly understood. The objective of this study was to investigate the antitumor effects of TCS on the U87 and U251 cell lines. The in vitro effects of TCS on these two cell lines were determined using a Cell Counting Kit­8 (CCK­8) assay, Annexin V­FITC staining, DAPI staining, Transwell assays, terminal deoxynucleotidyl transferase­mediated dUTP nick end­labeling (TUNEL) assays, 5,5',6,6'­tetrachloro­1,1',3,3'­tetraethyl­imidacarbocyanine iodide (JC­1) staining and western blotting, which was utilized to assess the expression of leucine­rich repeat­containing G protein­coupled receptor 5 (LGR5) and key proteins in the Wnt/ß­catenin signaling pathway. Our data indicated that TCS inhibited the proliferation of glioma cells in a dose­ and time­dependent manner and played a role in inhibiting glioma cell invasion and migration. Additional investigation revealed that the expression levels of LGR5 and of key proteins in the Wnt/ß­catenin signaling pathway were markedly decreased after TCS treatment. The results suggest that TCS may induce apoptosis in glioma cells by targeting LGR5 and repressing the Wnt/ß­catenin signaling pathway. In the future, in vivo experiments should be conducted to examine the potential use of this compound as a novel therapeutic agent for gliomas.

Effect of Radix Trichosanthis and Trichosanthin on Hepatitis B Virus in HepG2.2.15 Cells.

Radix Trichosanthis is a Chinese herbal medicine that has great medical value and pharmacological actions. There is already a long history of using the plant Radix Trichosanthis as treatment for hepatitis B virus in China. This research mainly focused on investigating the therapeutic effect of different extracts from Radix Trichosanthis on hepatitis B virus, on a cellular level (ex vivo). Cell survival rate of HepG2.2.15 cells was detected by MTT assay. HBsAg and HBeAg in HepG 2.2.15 cell supernatant were evaluated by enzyme linked immunosorbent assay (ELISA). Results showed that water extract from Radix Trichosanthis had a stronger inhibitive effect on expression of HBsAg and HBeAg in HepG2.2.15 cells than the alcohol extract from the same plant. Considering that the most active component of Radix Trichosanthis was in its aqueous extract and this might be related to the active component Trichosanthin. Trichosanthin was further used for related experiments to confirm this hypothesis. The results showed that Trichosanthin, in the aqueous extract from Radix Trichosanthis, is likely the main component responsible for the anti-hepatitis B viral effect.

Smac is another pathway in the anti-tumour activity of Trichosanthin and reverses Trichosanthin resistance in CaSki cervical cancer cells.

Trichosanthin (TCS), or Tin Hua Fen, is a renowned traditional Chinese medicine and is still used in Chinese clinics for midterm abortion and the treatment of choriocarcinoma. Many studies have demonstrated that TCS has anti-tumour action as a type I ribosome-inactivating protein. We hypothesized that there is another pathway of the anti-tumour activity of TCS. cDNA array analysis was applied to profile changes in gene expression of human CaSki in response to TCS stimulation. Smac, a mitochondrial protein, was identified as the highly upregulated protein in response to TCS treatment. The mRNA and protein levels of Smac were determined by real-time RT-PCR and Western blotting respectively. We analysed the methylation status of Smac using methylation-specific PCR (MSP) and indicates that TCS promotes Smac demethylation and increases its expression in cervical CaSki cells. Tumour cells develop resistance to TCS during prolonged treatment, as with other classic chemotherapeutic agents. Smac expression was downregulated and Twist was upregulated in TCS-resistant cells. These results indicate that TCS has demethylating activity and that Smac is involved in both TCS response and TCS resistance.

Cytotoxic genes from traditional Chinese medicine inhibit tumor growth both in vitro and in vivo.

OBJECTIVE: Little effort has been made to study the protein-encoding genes isolated from traditional Chinese medicine (TCM) drugs, and the delivery of these genes into malignant cells through recombinant adeno-associated virus (rAAV) vectors has not been attempted. METHODS: We synthesized the cDNAs of five known cytotoxic proteins isolated from TCM drugs and the FLAG epitope-tagged cDNAs were subcloned into a rAAV plasmid vector. The protein expression was confirmed by Western blot assay. Various cancer cell lines were transfected with the above plasmids and cell growth was monitored both in vitro and in vivo. The best cytotoxic gene was further packaged into rAAV vectors, under the control of a liver cancer-specific promoter. The liver tumor growth was then monitored following intratumor administration of the rAAV vectors. RESULTS: The expression plasmids, encoding individual potential cytotoxic genes tagged with FLAG epitope, were successfully generated and sequenced. Among these genes, trichosanthin (TCS) gene yielded the most promising results for the inhibition of cancer cell growth in vitro. The over-expressed TCS functioned as a type I ribosome-inactivating protein, followed by inducing apoptosis that is associated with the Bcl-PARP signaling pathway. Furthermore, intratumor injection of rAAV vectors containing the TCS gene significantly inhibited the growth of human hepatocellular carcinoma tumors in a murine xenograft model. CONCLUSION: Our studies suggest that the use of TCM cytotoxic genes is a useful therapeutic strategy for treating human cancers in general, and liver tumors in particular.

A novel extraction of trichosanthin from Trichosanthes kirilowii roots using three-phase partitioning and its in vitro anticancer activity.

CONTEXT: Three-phase partitioning (TPP), a unique technique which has been explored for protein separation, was used for extraction of trichosanthin (TCS). OBJECTIVE: TPP was used to optimize the TCS extraction and to determine its anticancer activity. MATERIALS AND METHODS: The process consists of the simultaneous addition of t-butanol and ammonium sulfate to the aqueous slurry of Trichosanthes kirilowii Maxim (Cucurbitaceae) root powder. The extraction of TCS was optimized with respect to the concentration of ammonium sulfate loading, the ratio of t-butanol to slurry, extraction time and pH. The anticancer activity was performed using 3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazolium bromide (MTT) assay in vitro. RESULTS: The extraction time with this technique is lower in comparison to conventional solvent extraction. The optimized protocol resulted in maximum recovery of 98.68% (w/w) protein within 1 h. The in vitro cytotoxic activity of the TCS was evaluated against HepG2 and WRL 68 cancer cell line and results showed that TCS possesses quite highly significant anticancer activity having IC50 values of 10.38 and 15.45 µmol/l, respectively, comparable to standard drugs. CONCLUSION: This framework is utilized as a basis for optimization for protein separation using TPP technique which is economical and eco-friendly.

Cytotoxic genes from traditional Chinese medicine inhibit tumor growth both in vitro and in vivo / 中西医结合学报

<p><b>OBJECTIVE</b>Little effort has been made to study the protein-encoding genes isolated from traditional Chinese medicine (TCM) drugs, and the delivery of these genes into malignant cells through recombinant adeno-associated virus (rAAV) vectors has not been attempted.</p><p><b>METHODS</b>We synthesized the cDNAs of five known cytotoxic proteins isolated from TCM drugs and the FLAG epitope-tagged cDNAs were subcloned into a rAAV plasmid vector. The protein expression was confirmed by Western blot assay. Various cancer cell lines were transfected with the above plasmids and cell growth was monitored both in vitro and in vivo. The best cytotoxic gene was further packaged into rAAV vectors, under the control of a liver cancer-specific promoter. The liver tumor growth was then monitored following intratumor administration of the rAAV vectors.</p><p><b>RESULTS</b>The expression plasmids, encoding individual potential cytotoxic genes tagged with FLAG epitope, were successfully generated and sequenced. Among these genes, trichosanthin (TCS) gene yielded the most promising results for the inhibition of cancer cell growth in vitro. The over-expressed TCS functioned as a type I ribosome-inactivating protein, followed by inducing apoptosis that is associated with the Bcl-PARP signaling pathway. Furthermore, intratumor injection of rAAV vectors containing the TCS gene significantly inhibited the growth of human hepatocellular carcinoma tumors in a murine xenograft model.</p><p><b>CONCLUSION</b>Our studies suggest that the use of TCM cytotoxic genes is a useful therapeutic strategy for treating human cancers in general, and liver tumors in particular.</p>

Anti-tumor action of trichosanthin, a type 1 ribosome-inactivating protein, employed in traditional Chinese medicine: a mini review.

Trichosanthin (TCS) as a midterm abortifacient medicine has been used clinically in traditional Chinese medicine for centuries. Additionally, TCS manifests a host of pharmacological properties, for instance, anti-HIV and anti-tumor activities. TCS has been reported to inhibit cell growth of a diversity of cancers, including cervical cancer, choriocarcinoma, and leukemia/lymphoma, etc. This article purported to review the various anti-tumor activities of TCS and the mechanism of apoptosis it induced in these tumor cells. These research progresses provide an insight into cancer research and treatment as well as disclose new pharmacological properties of the ancient but popular Chinese medicine.

Trichosanthin inhibits breast cancer cell proliferation in both cell lines and nude mice by promotion of apoptosis.

Breast cancer ranks as a common and severe neoplasia in women with increasing incidence as well as high risk of metastasis and relapse. Translational and laboratory-based clinical investigations of new/novel drugs are in progress. Medicinal plants are rich sources of biologically active natural products for drug development. The 27-kDa trichosanthin (TCS) is a ribosome inactivating protein purified from tubers of the Chinese herbal plant Trichosanthes kirilowii Maximowicz (common name Tian Hua Fen). In this study, we extended the potential medicinal applications of TCS from HIV, ferticide, hydatidiform moles, invasive moles, to breast cancer. We found that TCS manifested anti-proliferative and apoptosis-inducing activities in both estrogen-dependent human MCF-7 cells and estrogen-independent MDA-MB-231 cells. Flow cytometric analysis disclosed that TCS induced cell cycle arrest. Further studies revealed that TCS-induced tumor cell apoptosis was attributed to activation of both caspase-8 and caspase-9 regulated pathways. The subsequent events including caspase-3 activation, and increased PARP cleavage. With regard to cell morphology, stereotypical apoptotic features were observed. Moreover, in comparison with control, TCS- treated nude mice bearing MDA-MB-231 xenograft tumors exhibited significantly reduced tumor volume and tumor weight, due to the potent effect of TCS on tumor cell apoptosis as determined by the increase of caspase-3 activation, PARP cleavage, and DNA fragmentation using immunohistochemistry. Considering the clinical efficacy and relative safety of TCS on other human diseases, this work opens up new therapeutic avenues for patients with estrogen-dependent and/or estrogen-independent breast cancers.

Trichosanthin inhibits DNA methyltransferase and restores methylation-silenced gene expression in human cervical cancer cells.

Epigenetic silencing of tumor suppressor genes is a well-established oncogenic process and the reactivation of tumor suppressor genes that have been silenced by promoter methylation is an attractive molecular target for cancer therapy. In this study, we investigated the demethylation activity of trichosanthin (TCS, the main bioactive component isolated from a Chinese medicinal herb) and its possible mechanism of action in cervical cancer cell lines. HeLa human cervical adenocarcinoma and CaSki human cervical squamous carcinoma cells were treated with various concentrations (0, 20, 40 and 80 µg/ml) of TCS for 48 h and the mRNA and protein expression levels of the tumor suppressor genes adenomatous polyposis coli (APC) and tumor suppressor in lung cancer 1 (TSLC1) were detected using reverse transcription (RT)-PCR and western blotting, respectively. We analyzed the methylation status of APC and TSLC1 using methylation-specific PCR (MSP). The expression levels and enzyme activity of DNA methyltransferase 1 (DNMT1) were also examined. The mRNA and protein expression levels of APC and TSLC1 were increased following treatment with various concentrations (0, 20, 40 and 80 µg/ml) of TCS for 48 h. The expression of the APC gene increased 2.55±0.29-, 3.44±0.31- and 4.36±0.14-fold, respectively. The expression of the TSLC1 gene increased 2.28±0.15-, 4.23±0.88- and 6.09±0.23-fold, respectively. MSP detection showed that TCS induced demethylation in HeLa and CaSki cells and that this demethylation activity was accompanied by the decreased expression of DNMT1 and reduced DNMT1 enzyme activity. Our experimental results demonstrate for the first time that TCS is capable of restoring the expression of methylation-silenced tumor suppressor genes and is potentially useful as a demethylation agent for the clinical treatment of human cervical cancer.

Trichosanthin down-regulates Notch signaling and inhibits proliferation of the nasopharyngeal carcinoma cell line CNE2 in vitro.

The prognosis of nasopharyngeal carcinoma (NPC) is still poor. Trichosanthin (TCS) has abortifacient, anti-virus, immunoregulation and various anti-tumor pharmacological activities, but there are no reports about its effect on NPC and the exact mechanisms that TCS inhibits tumor are not well known. In this study, the proliferation, apoptosis and soft agar colony formation abilities of CNE2 cells were examined with various assays in vitro followed by treatment with TCS. Furthermore, the activation status of Notch signaling pathway in TCS and control cells also was examined. The results revealed that TCS could inhibit NPC cell line CNE2 in vitro, reduce clone formation ability and induce apoptosis of CNE2 cells. Down-regulation of Notch signaling may be one of the mechanisms that TCS inhibits NPC.

Recent progress in medicinal investigations on trichosanthin and other ribosome inactivating proteins from the plant genus Trichosanthes.

Ribosome inactivating proteins (RIPs) are toxic RNA N-glycosidases that cleave an adenine-ribose glycosidic bond at position adenine(4324) with the conserved ricin/α-sarcin loop in the eukaryotic 28S ribosomal RNA. RIPs have captured the attention of botanists, biochemists, and drug discoverers, due to their diverse potent defensive activities, and inter alia, their antitumor and anti-HIV activities. Out of the 145 families of plants, Trichosanthes ranks among the top 5 genera with a good potential of use for discovery of anticancer drugs. Trichosanthin (TCS) is a famous type I RIP purified from T. kirilowii that has been known for around 30 years. Based on the results of voluminous in vitro and in vivo investigations, TCS is considered a good candidate for the treatment of HIV/AIDS and neoplasms. Here we integrate recent progress of the research on the different medicinal activities of TCS. In addition to TCS, other promising RIPs from the same species (such as TAP29 and trichoanguin), and from the same genus Trichosanthes are included. This review presents a brief panorama of the studies on Trichosanthes RIPs. Regarding the debilitating nature of AIDS and different tumors, further understanding of these multifunctional proteins is worthwhile since it may help to open a novel therapeutic window for these stubborn diseases.

Trichosanthin enhances anti-tumor immune response in a murine Lewis lung cancer model by boosting the interaction between TSLC1 and CRTAM.

Trichosanthin (TCS), extracted from the Chinese medicinal herb Trichosanthes kirilowi, has shown promise for the inhibition of tumor growth. However, its immunomodulatory effect on tumor-host interaction remains unknown. In this study, we focused on the effect of TCS on murine anti-tumor immune response in the 3LL Lewis lung carcinoma tumor model and explored the possible molecular pathways involved. In addition to inhibiting cell proliferation and inducing apoptosis in the 3LL tumor, TCS retarded tumor growth and prolonged mouse survival more significantly in C57BL/6 immunocompetent mice than in nude mice. This reflected the fact that the host immune system was involved in tumor eradication. Using FACS analysis, we found that TCS increased the percentage of effector T cells, particularly Interferon-gamma (IFN-γ) producing CD4(+) and CD8(+) T cells from tumor-bearing mice. TCS also promoted the vigorous proliferation of antigen-specific effector T cells, markedly increased Th1 cytokine secretion and elicited more memory T cells in tumor-bearing mice, consequently enhancing the anti-tumor response and inducing immune protection. Furthermore, we found that TCS upregulated the expression of tumor suppressor in lung cancer 1 (TSLC1) in 3LL tumor cells and the expression of its ligand, class I-restricted T cell-associated molecule (CRTAM), in effector T cells. Blocking TSLC1 expression with small interfering RNA (siRNA) significantly eliminated the effects of TCS on the proliferation and cytokine secretion of effector T cells, suggesting that TCS enhances anti-tumor immune response at least partially by boosting the interaction between TSLC1 and CRTAM. Collectively, our data demonstrate that TCS not only affects tumor cells directly, but also enhances anti-tumor immunity via the interaction between TSLC1 and CRTAM. These findings may lead to the development of a novel approach for tumor regression.