Exploring the apoptotic effects of sericin on HCT116 cells through comprehensive nanostring transcriptomics and proteomics analysis.

Sericin, a silk protein from Bombyx mori (silkworms), has many applications, including cosmetics, anti-inflammation, and anti-cancer. Sericin complexes with nanoparticles have shown promise for breast cancer cell lines. Apoptosis, a programmed cell death mechanism, stops cancer cell growth. This study found that Sericin urea extract significantly affected HCT116 cell viability (IC50 = 42.00 ± 0.002 µg/mL) and caused apoptosis in over 80% of treated cells. S-FTIR analysis showed significant changes in Sericin-treated cells' macromolecule composition, particularly in the lipid and nucleic acid areas, indicating major cellular modifications. A transcriptomics study found upregulation of the apoptotic signaling genes FASLG, TNFSF10, CASP3, CASP7, CASP8, and CASP10. Early apoptotic proteins also showed that BAD, AKT, CASP9, p53, and CASP8 were significantly upregulated. A proteomics study illuminated Sericin-treated cells' altered protein patterns. Our results show that Sericin activated the extrinsic apoptosis pathway via the caspase cascade (CASP8/10 and CASP3/7) and the death receptor pathway, involving TNFSF10 or FASLG, in HCT116 cells. Upregulation of p53 increases CASP8, which activates CASP3 and causes HCT116 cell death. This multi-omics study illuminates the molecular mechanisms of Sericin-induced apoptosis, sheds light on its potential cancer treatment applications, and helps us understand the complex relationship between silk-derived proteins and cellular processes.

Dual role of azo compounds in inhibiting Plasmodium falciparum adenosine deaminase and hemozoin biocrystallization.

Protein-ligand (GOLD) docking of the NCI compounds into the ligand-binding site of Plasmodium falciparum adenosine deaminase (PfADA) identified three most active azo compounds containing 4-[(4-hydroxy-2-oxo-1H-quinolin-3-yl) moiety. These compounds showed IC50 of 3.7-15.4 µM against PfADA, as well as inhibited the growth of P. falciparum strains 3D7 (chloroquine (CQ)-sensitive) and K1 (CQ-resistant) with IC50 of 1.8-3.1 and 1.7-3.6 µM, respectively. The identified compounds have structures similar to the backbone structure (4-N-(7-chloroquinolin-4-yl)) in CQ, and NSC45545 could mimic CQ by inhibiting the bioformation of hemozoin in parasitic food vacuole. The amount of in situ hemozoin in the ring-stage parasite was determined using a combination of synchrotron transmission Fourier transform infrared microspectroscopy and Principal Component Analysis. Stretching of the C-O bond of hemozoin propionate group measured at 1220-1210 cm-1 in untreated intraerythrocytic P. falciparum strains 3D7 and K1 was disappeared following treatment with 1.85 and 1.74 µM NSC45545, similar to those treated with 0.02 and 0.13 µM CQ, respectively. These findings indicate a novel dual function of 4-[(4-hydroxy-2-oxo-1H-quinolin-3-yl) azo compounds in inhibiting both PfADA and in situ hemozoin biocrystallization. These lead compounds hold promise for further development of new antimalarial therapeutics that could delay the onset of parasitic drug resistance.

Synchrotron Fourier Transform Infrared Microscopy Spectra in Cellular Effects of Janus Kinase Inhibitors on Myelofibrosis Cancer Cells.

Janus kinase (JAK) deregulation of the JAK/signal transducers and activators of transcription pathway leads to myelofibrosis that can be treated by JAK inhibitors including Ruxolitinib and Tofacitinib. Even though both inhibitors are effective against myelofibrosis, each of them has a different mode of action in the cells. Ruxolitinib is an inhibitor for selective JAK1/2, and Tofacitinib is an inhibitor for JAK3. This study evaluated the chemical fingerprints of TF-1 cells after JAK inhibitor treatments by the synchrotron Fourier transform infrared microspectroscopy (S-FTIR) spectrum. Tofacitinib and Ruxolitinib treatments in TF-1 cells were applied with a chemical fingerprint approach in S-FTIR spectroscopy and in vitro cytotoxicity in a cell-based assay. Principal component analysis or PCA was utilized to classify three cell treatments with three biochemical alteration absorbances of lipid vibration by the C-H stretching, protein amide I that appeared from the C=O stretching, and a P=O phosphodiester bond from nucleic acids. The results showed that the inhibition effect of Ruxolitinib on the TF-1 cell lines was two-fold higher than Tofacitinib. PCA distinguishes untreated and drug-treated cells by detecting cellular biochemical alteration. The loading plots identify that proteins and nucleic acids were the different main components in disparate cell treatments. Tofacitinib was distinct from the others in lipid and nucleic acid. The second derivative spectra of the three molecular components had decreased lipid production and accumulation, changes in secondary structures in proteins, and a high level of RNA overexpression in cell treatment. The JAK inhibitors caused different spectroscopic biomarkers of the modifications of secondary protein conformation, stimulated cell lipid accumulation, and phosphorylation from untreated cells. The alteration of cellular biochemical components suggests that FTIR is a potential tool to analyze specific patterns of drug cellular responses at the molecular level.

Probing the Anti-Cancer Potency of Sulfated Galactans on Cholangiocarcinoma Cells Using Synchrotron FTIR Microspectroscopy, Molecular Docking, and In Vitro Studies.

Sulfated galactans (SG) isolated from red alga Gracilaria fisheri have been reported to inhibit the growth of cholangiocarcinoma (CCA) cells, which was similar to the epidermal growth factor receptor (EGFR)-targeted drug, cetuximab. Herein, we studied the anti-cancer potency of SG compared to cetuximab. Biological studies demonstrated SG and cetuximab had similar inhibition mechanisms in CCA cells by down-regulating EGFR/ERK pathway, and the combined treatment induced a greater inhibition effect. The molecular docking study revealed that SG binds to the dimerization domain of EGFR, and this was confirmed by dimerization assay, which showed that SG inhibited ligand-induced EGFR dimer formation. Synchrotron FTIR microspectroscopy was employed to examine alterations in cellular macromolecules after drug treatment. The SR-FTIR-MS elicited similar spectral signatures of SG and cetuximab, pointing towards the bands of RNA/DNA, lipids, and amide I vibrations, which were inconsistent with the changes of signaling proteins in CCA cells after drug treatment. Thus, this study demonstrates the underlined anti-cancer mechanism of SG by interfering with EGFR dimerization. In addition, we reveal that FTIR signature spectra offer a useful tool for screening anti-cancer drugs' effect.

FTIR spectra signatures reveal different cellular effects of EGFR inhibitors on nonsmall cell lung cancer cells.

ATP-analogue inhibitors, Gefitinib (Iressa) and Erlotinib (Tarceva) had been approved for advanced and metastatic nonsmall cell lung cancer (NSCLC) cells against tyrosine kinase domain of epidermal growth factor receptor (EGFR). Many techniques have been developed to better understand the drug mechanism which is multistep, time-consuming and expensive. Herein, we performed Fourier-transform infrared (FTIR) microscopy for evaluating the biochemical change on NSCLC (A549) cells after treatment. At levels that produced equivalent effects, Gefitinib dramatically induced cell apoptosis via impaired mitochondrial transmembrane potential. Whereas, Erlotinib had a slight effect on A549. Principal component analysis was performed to distinguish the effect of EGFR inhibitors on A549. FTIR spectra regions were divided into three regions: lipids (3000-2800 cm-1 ), proteins (1700-1500 cm-1 ) and carbohydrates and nuclei acids (1200-1000 cm-1 ). Biochemical changes can be evaluated by these spectral regions. This work may be a novel concept for utilizing FTIR spectroscopy for high-throughput discriminative effects of a drug or compound and its derivatives on cells.

Crystal structure of Plasmodium falciparum adenosine deaminase reveals a novel binding pocket for inosine.

Plasmodium falciparum (Pf), a malarial pathogen, can only synthesize purine nucleotides employing a salvage pathway because it lacks de novo biosynthesis. Adenosine deaminase (ADA), one of the three purine salvage enzymes, catalyzes the irreversible hydrolytic deamination of adenosine to inosine, which is further converted to GMP and AMP for DNA/RNA production. In addition to adenosine conversion, Plasmodium ADA also catalyzes the conversion of 5'-methylthioadenosine, derived from polyamine biosynthesis, into 5'-methylthioinosine whereas the human enzyme is not capable of this function. Here we report the crystal structure of a surface engineered PfADA at a resolution of 2.48 Å, together with results on kinetic studies of PfADA wild-type and active site variants. The structure reveals a novel inosine binding pocket linked to a distinctive PfADA substructure (residues 172-179) derived from a non-conserved gating helix loop (172-188) in Plasmodium spp. and other ADA enzymes. Variants of PfADA and human (h) ADA active site amino acids were generated in order to study their role in catalysis, including PfADA- Phe136, -Thr174, -Asp176, and -Leu179, and hADA-Met155, equivalent to PfADA-Asp176. PfADA-Leu179His showed no effect on kinetic parameters. However, kinetic results of PfADA-Asp176Met/Ala mutants and hADA-Met155Asp/Ala showed that the mutation reduced adenosine and 5'-methylthioadenosine substrate affinity in PfADA and kcat in hADA, thereby reducing catalytic efficiency of the enzyme. Phe136Leu mutant showed increased Km (>10-fold) for both substrates whereas Thr174Ile/Ala only affected 5'-methylthioadenosine binding affinity. Together, the structure with the novel inosine binding pocket and the kinetic data provide insights for rational design of inhibitors against PfADA.