Parasporins 1 and 2: Their structure and activity.
Akiba Toshihiko,Okumura Shiro
Journal of invertebrate pathology
Parasporins are novel protein toxins preferentially cytotoxic against human cancer cells. They are obtained from parasporal inclusions of Bacillus thuringiensis and, accordingly, are considered congeners of the insecticidal Cry toxins. Two types of parasporins have been identified: the three-domain Cry toxin type and the β-pore-forming-toxin (β-PFT) type. Crystal structures of representative members of the two types, PS1Aa1 and PS2Aa1, have been determined and compared with those of well-studied toxins. PS1Aa1 has a typical architecture characteristic of the three-domain insecticidal Cry toxins, though it is cleaved into two polypeptides. It has an extra N-terminal segment found only in the inactive form of the Cry toxins and, hence, it is presumed to act through another mechanism as an activator in the apoptotic signaling pathway rather than a pore-forming toxin. PS2Aa1 shows a remarkable structural similarity to the aerolysin-type β-PFTs, which is much greater than expected from its limited sequence identity to those toxins. This strongly suggests that a pore-forming mechanism similar to that of β-PFTs is involved in the action of this type of parasporin. The structural comparison of PS2Aa1 to other aerolysin-type β-PFTs indicates conserved oligomerization and pore-forming structures in domains 2 and 3, and highly diverse putative receptor binding region structures in domain 1, likely accounting for enhanced cancer cell cytotoxicity as compared to normal control cells. The structural implications for the mechanism of action and cellular specificity of both Cry and β-PFT type parasporins will be enhanced by further experimental validation.
10.1016/j.jip.2016.10.005
Apoptin Regulates Apoptosis and Autophagy by Modulating Reactive Oxygen Species (ROS) Levels in Human Liver Cancer Cells.
Li Yiquan,Zhu Yilong,Fang Jinbo,Li Wenjie,Li Shanzhi,Liu Xing,Liu Zirui,Song Gaojie,Shang Chao,Cong Jianan,Bai Bing,Sun Lili,Jin Ningyi,Li Xiao
Frontiers in oncology
Apoptin is a protein that specifically induces apoptosis in tumor cells. The anti-tumorigenic functions of Apoptin, including autophagy activation and its interaction with apoptosis, have not been precisely elucidated. Here we investigate the main pathways of apoptin-mediated killing of human liver cancer cells, as well as its putative role in autophagy and apoptosis. The anti-proliferative effect of apoptin in liver cancer cells was analyzed by crystal violet staining and MTS detection, and also using a tumor-based model. The main pathway related to apoptin-induced growth inhibition was evaluated by flow cytometry and fluorescence staining. The relationship between apoptosis and autophagy on apoptin-treating cells was analyzed using apoptosis and autophagy inhibitors, mitochondrial staining, Annexin V-FITC/PI flow detection, LC3 staining, and western blotting. The effect of ROS toward the apoptosis and autophagy of apoptin-treating cells was also evaluated by ROS detection, Annexin V-FITC/PI flow detection, LC3 staining, and western blotting. Inhibition of apoptosis in apoptin-treating liver cancer cells significantly reduced the autophagy levels . The overall inhibition increased from 12 h and the effect was most obvious at 48 h. Inhibition of autophagy could increase apoptin-induced apoptosis of cells in a time-dependent manner, reaching its peak at 24 h. Apoptin significantly alters ROS levels in liver cancer cells, and this effect is directly related to apoptosis and autophagy. ROS appears to be the key factor linking apoptin-induced autophagy and apoptosis through the mitochondria in liver cancer cells. Therefore, evaluating the interaction between apoptin-induced apoptosis and autophagy is a promising step for the development of alternate tumor therapies.
10.3389/fonc.2020.01026
Crystal structure of dihydropyrimidine dehydrogenase, a major determinant of the pharmacokinetics of the anti-cancer drug 5-fluorouracil.
Dobritzsch D,Schneider G,Schnackerz K D,Lindqvist Y
The EMBO journal
Dihydropyrimidine dehydrogenase catalyzes the first step in pyrimidine degradation: the NADPH-dependent reduction of uracil and thymine to the corresponding 5,6-dihydropyrimidines. Its controlled inhibition has become an adjunct target for cancer therapy, since the enzyme is also responsible for the rapid breakdown of the chemotherapeutic drug 5-fluorouracil. The crystal structure of the homodimeric pig liver enzyme (2x 111 kDa) determined at 1.9 A resolution reveals a highly modular subunit organization, consisting of five domains with different folds. Dihydropyrimidine dehydrogenase contains two FAD, two FMN and eight [4Fe-4S] clusters, arranged in two electron transfer chains that pass the dimer interface twice. Two of the Fe-S clusters show a hitherto unobserved coordination involving a glutamine residue. The ternary complex of an inactive mutant of the enzyme with bound NADPH and 5-fluorouracil reveals the architecture of the substrate-binding sites and residues responsible for recognition and binding of the drug.
10.1093/emboj/20.4.650
A Sixty-Year Research and Development of Trichosanthin, a Ribosome-Inactivating Protein.
Toxins
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 -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.
10.3390/toxins14030178
Structures of YAP protein domains reveal promising targets for development of new cancer drugs.
Sudol Marius,Shields Denis C,Farooq Amjad
Seminars in cell & developmental biology
YAP (Yes-associated protein) is a potent oncogene and a major effector of the mammalian Hippo tumor suppressor pathway. In this review, our emphasis is on the structural basis of how YAP recognizes its various cellular partners. In particular, we discuss the role of LATS kinase and AMOTL1 junction protein, two key cellular partners of YAP that bind to its WW domain, in mediating cytoplasmic localization of YAP and thereby playing a key role in the regulation of its transcriptional activity. Importantly, the crystal structure of an amino-terminal domain of YAP in complex with the carboxy-terminal domain of TEAD transcription factor was only recently solved at atomic resolution, while the structure of WW domain of YAP in complex with a peptide containing the PPxY motif has been available for more than a decade. We discuss how such structural information may be exploited for the rational development of novel anti-cancer therapeutics harboring greater efficacy coupled with low toxicity. We also embark on a brief discussion of how recent in silico studies led to identification of the cardiac glycoside digitoxin as a potential modulator of WW domain-ligand interactions. Conversely, dobutamine was identified in a screen of known drugs as a compound that promotes cytoplasmic localization of YAP, thereby resulting in growth suppressing activity. Finally, we discuss how a recent study on the dynamics of WW domain folding on a biologically critical time scale may provide a tool to generate repertoires of WW domain variants for regulation of the Hippo pathway toward desired, non-oncogenic outputs.
10.1016/j.semcdb.2012.05.002
Agrocybe cylindracea fucoglucogalactan induced lysosome-mediated apoptosis of colorectal cancer cell through H3K27ac-regulated cathepsin D.
Carbohydrate polymers
Inducing lysosomal dysfunction is emerging as a promising means for cancer therapy. Agrocybe cylindracea fucoglucogalactan (ACP) is a bioactive ingredient with anti-tumor activity, while its mechanism remains obscure. Herein, we found that ACP visibly inhibited the proliferation of colorectal cancer cells, and the IC50 value on HCT-116 cells (HT29 cells) was 490 μg/mL (786.4 μg/mL) at 24 h. RNA-seq showed that ACP regulated mitochondria, lysosome and apoptosis-related pathways. Further experiments proved that ACP indeed promoted apoptosis and lysosomal dysfunction of HCT-116 cells. Moreover, ChIP-seq revealed that ACP increased histone-H3-lysine-27 acetylation (H3K27ac) on CTSD (cathepsin D) promoter in HCT-116 cells, thus facilitating the binding of transcription factor EB (TFEB), and resulted in ascension of CTSD expression. Additionally, ACP triggered mitochondrial-mediated apoptosis by decreasing mitochondrial membrane potential and increasing pro-apoptotic protein levels. Notably, Pepstatin A (CTSD inhibitor) availably alleviated ACP-induced apoptosis. Taken together, our results indicated that ACP induced lysosome-mitochondria mediated apoptosis via H3K27ac-regulated CTSD in HCT-116 cells. This study indicates that ACP has anti-cancer potential in the treatment of colorectal cancer.
10.1016/j.carbpol.2023.121208
Complex Crystal Structure Determination of Hsp90-NVP-AUY922 and Anti-NSCLC Activity of NVP-AUY922.
He Chun-Xia,Lv You,Guo Meng,Zhou Huan,Qin Wei,Zhao Dong,Li Hui-Jin,Xing Lu,Zhou Xin,Li Peng-Quan,Yu Feng,He Jian-Hua,Cao Hui-Ling
Frontiers in oncology
New targeted chemotherapy agents greatly improved five-year survival in NSCLC patients, but which were susceptible to drug resistance. NVP-AUY922, terminated in phase II clinical trials, exhibited promising anti-NSCLC (non-small-cell lung cancer) activity targeting to Hsp90 (heat shock protein), which demonstrated advantages in overcoming drug resistance as a broad-spectrum anti-cancer target. It was expected to develop novel anti-NSCLC drugs to overcome drug resistance by the structural optimization of NVP-AUY922. However, the absence of high-resolution complex crystal structure of Hsp90-NVP-AUY922 blocked the way. Herein, 1.59 Å-resolution complex crystal structure of Hsp90-NVP-AUY922 (PDB ID 6LTI) was successfully determined by X-ray diffraction. Meanwhile, there was a strong binding capability between NVP-AUY922 and its target Hsp90 verified by TSA (ΔTm, -15.56 ± 1.78°C) and ITC ( , 5.10 ± 2.10 nM). Results by the complex crystal structure, TSA and ITC verified that NVP-AUY922 well accommodated in the ATP-binding pocket of Hsp90 to disable the molecular chaperone activity of Hsp90. Therefore, NVP-AUY922 exhibited approving inhibitory activity on NSCLC cell line H1299 (IC, 2.85 ± 0.06 μM) by inhibiting cell proliferation, inducing cell cycle arrest and promoting cell apoptosis. At the basis of the complex crystal structure and molecular interaction analysis, thirty-two new NVP-AUY922 derivatives were further designed, and among which twenty-eight new ones display enhanced binding force with Hsp90 by molecular docking evaluation. The results would promote anti-NSCLC new drug development to overcome drug resistance based on the lead compound NVP-AUY922.
10.3389/fonc.2022.847556
Cyclin-dependent kinase-2 as a target for cancer therapy: progress in the development of CDK2 inhibitors as anti-cancer agents.
Chohan Tahir Ali,Qian Haiyan,Pan Youlu,Chen Jian-Zhong
Current medicinal chemistry
Cyclin-dependent kinase-2 (CDK2) is a member of protein kinase family. It plays an important role in regulating various events of eukaryotic cell division cycle. Accumulated evidences indicated that over expression of CDK2 should cause the abnormal regulation of cell-cycle, which would be directly associated with hyperproliferation in cancer cells. Therefore, CDK2 was regarded as a potentially therapeutic target for cancer therapy. Knowledge of crystallography and availability of X-ray crystal structure of CDK2 have enabled us to understand the mode of CDK2 inhibition, which facilitated the development of numerous CDK2 inhibitors. Some of the CDK2 inhibitors were investigated clinically for their potential as anti-cancer agents. In this review, we present the structure, functions and activation of CDK2 by cyclin binding with special focus on recent advances in the development of different classes of CDK2 inhibitors. We also summarize different strategies to achieve subtype specificity either by targeting a binding pocket other than ATP, i.e. allosteric ligand binding site or by natural protein inhibitors capable to disrupt CDK2-cyclin complexes. It is possible to develop pharmacologically relevant cytotoxic agents by specifically inhibiting CDK2 activity with lesser toxicity than traditional chemotherapeutic agents.
Heparanase: A Challenging Cancer Drug Target.
Coombe Deirdre R,Gandhi Neha S
Frontiers in oncology
Heparanase has been viewed as a promising anti-cancer drug target for almost two decades, but no anti-heparanase therapy has yet reached the clinic. This endoglycosidase is highly expressed in a variety of malignancies, and its high expression is associated with greater tumor size, more metastases, and a poor prognosis. It was first described as an enzyme cleaving heparan sulfate chains of proteoglycans located in extracellular matrices and on cell surfaces, but this is not its only function. It is a multi-functional protein with activities that are enzymatic and non-enzymatic and which take place both outside of the cell and intracellularly. Knowledge of the crystal structure of heparanase has assisted the interpretation of earlier structure-function studies as well as in the design of potential anti-heparanase agents. This review re-examines the various functions of heparanase in light of the structural data. The functions of the heparanase variant, T5, and structure and functions of heparanase-2 are also examined as these heparanase related, but non-enzymatic, proteins are likely to influence the efficacy of anti-heparanase drugs. The anti-heparanase drugs currently under development predominately focus on inhibiting the enzymatic activity of heparanase, which, in the absence of inhibitors with high clinical efficacy, prompts a discussion of whether this is the best approach. The diversity of outcomes attributed to heparanase and the difficulties of unequivocally determining which of these are due to its enzymatic activity is also discussed and leads us to the conclusion that heparanase is a valid, but challenging drug target for cancer.
10.3389/fonc.2019.01316