In-Bridge Stereochemistry: A Determinant of Stapled Peptide Conformation and Activity.

Peptide side chain stapling has been proven to be an effective strategy for fine-tuning peptide properties. This innovative approach leads to the creation of stapled peptides characterized by stabilized α-helical conformations, enhanced protein-binding affinity, improved cell permeability, superior enzymatic stability, and numerous other advantages. Extensive research has explored the impact of various stapling bridges on the properties of these peptides, with limited investigation into the influence of bridge chirality, until very recently. In this concise review, we provide a brief overview of the current state of knowledge regarding the stereochemistry within the bridges of stapled peptides, offering insights into the potential applications of chiral bridges in the design and development of stapled peptides.

A meta-analysis of the efficacy and safety of first-line chemotherapeutic agents for osteosarcoma.

BACKGROUND: Osteosarcoma is a pleomorphic cancer that frequently affects children and teenagers. Although several chemotherapy regimens have been utilized for many years, the best therapeutic option for the treatment of osteosarcoma has not yet been determined. OBJECTIVES: This meta-analysis was designed to assess the clinical efficacy of a high-dose methotrexate, doxorubicin and cisplatin (MAP) regimen and compare its survival outcomes with those of other chemotherapy strategies in patients diagnosed with osteosarcoma. MATERIAL AND METHODS: We systematically searched databases, namely Embase, the Cochrane Library and PubMed, up to August 2022, for relevant studies investigating the impact of the MAP chemotherapy protocol on survival among patients with osteosarcoma. The odds ratio (OR) pooled estimates and their 95% confidence intervals (95% CIs) were calculated. RESULTS: Twelve studies including 4102 patients were eligible for analysis in this study. The estimated pooled ORs of the 3-year overall survival (OS) and event-free survival (EFS) were OR = 1.08 (95% CI: 0.72-1.62, p = 0.70) and OR = 1.04 (95% CI: 0.81-1.32, p = 0.78, respectively). The 5-year OS and EFS were OR = 0.87 (95% CI: 0.62-1.23, p = 0.42) and OR = 1.13 (95% CI: 0.76-1.68, p = 0.54), respectively, with no statistical differences. The subgroup analysis of MAP compared to a 2-drug regimen (doxorubicin and cisplatin) revealed a significant difference between the 2 chemotherapy strategy groups in 3-year OS rates (OR = 0.72 (95% CI: 0.56-0.92, p = 0.009)) and 5-year EFS rates (OR = 0.57 (95% CI: 0.43-0.76, p < 0.001)). CONCLUSION: The MAP chemotherapy strategy for osteosarcoma showed superiority over other regimens, especially over the 2-drug regimen (doxorubicin/cisplatin), in terms of better prognosis and safety.

Discovery of Itaconate-Mediated Lysine Acylation.

Itaconate is an important antimicrobial and immunoregulatory metabolite involved in host-pathogen interactions. A key mechanistic action of itaconate is through the covalent modification of cysteine residues via Michael addition, resulting in "itaconation". However, it is unclear whether itaconate has other regulatory mechanisms. In this work, we discovered a novel type of post-translational modification by promiscuous antibody enrichment and data analysis with the open-search strategy and further confirmed it as the lysine "itaconylation". We showed that itaconylation and its precursor metabolite itaconyl-CoA undergo significant upregulation upon lipopolysaccharides (LPS) stimulation in RAW264.7 macrophages. Quantitative proteomics identified itaconylation sites in multiple functional proteins, including glycolytic enzymes and histones, some of which were confirmed by synthetic peptide standards. The discovery of lysine itaconylation opens up new areas for studying how itaconate participates in immunoregulation via protein post-translational modification.

Profiling the metabolome of uterine fluid for early detection of ovarian cancer.

Ovarian cancer (OC) causes high mortality in women because of ineffective biomarkers for early diagnosis. Here, we perform metabolomics analysis on an initial training set of uterine fluid from 96 gynecological patients. A seven-metabolite-marker panel consisting of vanillylmandelic acid, norepinephrine, phenylalanine, beta-alanine, tyrosine, 12-S-hydroxy-5,8,10-heptadecatrienoic acid, and crithmumdiol is established for detecting early-stage OC. The panel is further validated in an independent sample set from 123 patients, discriminating early OC from controls with an area under the curve (AUC) of 0.957 (95% confidence interval [CI], 0.894-1). Interestingly, we find elevated norepinephrine and decreased vanillylmandelic acid in most OC cells, resulting from excess 4-hydroxyestradiol that antagonizes the catabolism of norepinephrine by catechol-O-methyltransferase. Moreover, exposure to 4-hydroxyestradiol induces cellular DNA damage and genomic instability that could lead to tumorigenesis. Thus, this study not only reveals metabolic features in uterine fluid of gynecological patients but also establishes a noninvasive approach for the early diagnosis of OC.

Metformin sensitises osteosarcoma to chemotherapy via the IGF-1R/miR-610/FEN1 pathway.

Metformin can enhance cancer cell chemosensitivity to anticancer drugs. IGF-1R is involved in cancer chemoresistance. The current study aimed to elucidate the role of metformin in osteosarcoma (OS) cell chemosensitivity modulation and identify its underlying mechanism in IGF-1R/miR-610/FEN1 signalling. IGF-1R, miR-610, and FEN1 were aberrantly expressed in OS and participated in apoptosis modulation; this effect was abated by metformin treatment. Luciferase reporter assays confirmed that FEN1 is a direct target of miR-610. Moreover, metformin treatment decreased IGF-1R and FEN1 but elevated miR-610 expression. Metformin sensitised OS cells to cytotoxic agents, while FEN1 overexpression partly compromised metformin's sensitising effects. Furthermore, metformin was observed to enhance adriamycin's effects in a murine xenograft model. Metformin enhanced OS cell sensitivity to cytotoxic agents via the IGF-1R/miR-610/FEN1 signalling axis, highlighting its potential as an adjuvant during chemotherapy.

Fine Tuning the Properties of Stapled Peptides by Stereogenic α-Amino Acid Bridges.

Peptide stapling represents a versatile strategy to generate peptide derivatives with stable helical structures. While a wide range of skeletons have been investigated for cyclizing the side chains of peptides, the stereochemical outcomes from the linkers remain to be better understood. In this study, we incorporated α-amino acids (α-AAs) as bridges to construct side chain-stapled analogs of an interleukin-17A-binding peptide (HAP) and evaluated the impacts of the staples on the peptide's properties. While all AA-derived peptidyl staples drastically increase the enzymatic stability of HAP, our results indicate that compared to the D-amino acid bridges, the L-AA-based staples may generate more significant impacts in increasing the helicity and enhancing the interleukin-17A(IL-17A)-binding affinity of the modified peptide. Using Rosetta modelling and molecular dynamics (MD) simulations, we demonstrate that the chirality (L/D) possessed within the AAs substantially influences the conformation of stapled HAP peptides, providing either stabilizing or destabilizing effects. Based on the computational model, a modification of the stapled HAP leads to the discovery of a peptide with further enhanced helicity, enzymatic stability and IL-17A-inhibiting ability. This systematic study reveals that chiral AAs can serve as modulatory linkers for optimizing the structures and properties of stapled peptides.

PDTAC: Targeted Photodegradation of GPX4 Triggers Ferroptosis and Potent Antitumor Immunity.

Targeted degradation of proteins, especially those regarded as undruggable or difficult to drug, attracts wide attention to develop novel therapeutic strategies. Glutathione peroxidase 4 (GPX4), the key enzyme regulating ferroptosis, is currently a target with just covalent inhibitors. Here, we developed a targeted photolysis approach and achieved efficient degradation of GPX4. The photodegradation-targeting chimeras (PDTACs) were synthesized by conjugating a clinically approved photosensitizer (verteporfin) to noninhibitory GPX4-targeting peptides. These chimeras selectively degraded the target protein in both cell lysates and living cells upon red-light irradiation. The targeted photolysis of GPX4 resulted in dominant ferroptotic cell death in malignant cancer cells. Moreover, the dying cells resulting from the PDTACs exhibited potent immunogenicity in vitro and efficiently elicited antitumor immunity in vivo. Our approach therefore provides a novel method to induce GPX4 dysfunction based on noncovalent binding and specifically trigger immunogenic ferroptosis, which may boost the application of ferroptosis in cancer immunotherapy.

Vascular Endothelial Growth Factor Mimetic Peptide and Parathyroid Hormone (1-34) Delivered via a Blue-Light-Curable Hydrogel Synergistically Accelerate Bone Regeneration.

Safe and effective biomaterials are in urgent clinical need for tissue regeneration and bone repair. While numerous advances have been made on hydrogels promoting osteogenesis in bone formation, co-stimulation of the angiogenic pathways in this process remains to be exploited. Here, we have developed a gelatin-based blue-light-curable hydrogel system, functionalized with an angiogenic vascular endothelial growth factor (VEGF) mimetic peptide, KLTWQELYQLKYKGI (KLT), and an osteoanabolic peptide, parathyroid hormone (PTH) 1-34. We have discovered that the covalent modification of gelatin scaffold with peptides can modulate the physical properties and biological activities of the produced hydrogels. Furthermore, we have demonstrated that those two peptides orchestrate synergistically and promote bone regeneration in a rat cranial bone defect model with remarkable efficacy. This dual-peptide-functionalized hydrogel system may serve as a promising lead to functional biomaterials in bone repair and tissue engineering.

Long non-coding RNA long intergenic non-coding 00641 mediates cell progression with stimulating cisplatin-resistance in osteosarcoma cells via microRNA-320d/myeloid cell leukemia-1 axis.

As a staple chemotherapy medicine, cisplatin (DDP) is extensively applied in cancer patients, but its drug resistance is limited. Numerous studies have elucidated that long non-coding RNA (lncRNA) performs as a pivotal agent in osteosarcoma (OS). Nevertheless, lncRNA long intergenic non-coding 00641 (LINC00641)'s functions in DDP resistance for OS remain obscure. The purpose of this study was to investigate the effect and mechanism of LINC00641 on drug resistance of OS. The tissues of both clinical cancer patients and the normal control were gathered. Detection of LINC00641, microRNA-320d (miR-320d) and myeloid cell leukemia-1 (MCL1) was conducted. After the selection of OS cell lines, the detection of cell advancement was applied. Series of experiments were conducted to verify the interaction of LINC00641, miR-320d and MCL1. Xenografted tumor model in vivo was utilized to determine the function of LINC00641. The data displayed, LINC00641 was prominently elevated in OS tissues and cells, especially in DDP-resistant tumors and cell lines. Knock-down LINC00641 was able to attenuate progression of DDP-resistant OS cells thus dampening their drug resistance toward DDP. Moreover, knock-downing LINC00641 gene was also able to manifest antagonism toward DDP-resistance in vivo. On the grounds of bioinformatics prediction, a direct binding of LINC00641 with miR-320d existed, whose target was MCL1. Meanwhile, LINC00641 modulated MCL1 via targeting miR-320d. Additionally, repressive LINC00641 blocked MCL1 via emulative interaction with miR-320d, thus expediting DDP-sensitivity of OS cells. All in all, it is found that LINC00641 is available to escalate drug resistance of DDP-resistant OS cells via mediation of miR-320d/MCL1 axis.

Rational Design of Self-Assembled Mitochondria-Targeting Lytic Peptide Conjugates with Enhanced Tumor Selectivity.

Membrane lytic peptides (MLP) are widely explored as cellular delivery vehicles or antitumor/antibacterial agents. However, the poor selectivity between cancer and normal cells slims their prospects as potential anti-tumor drugs. Herein, we have developed a rationally designed self-assembly strategy to enhance tumor selectivity of MLP-based conjugates, incorporating a hydrophobic triphenylphosphonium (TPP) group for mitochondria targeting, and a hydrophilic arginine-glycine-aspartic acid (RGD) sequence targeting integrins. The self-assembly nanoparticles can enhance the stability of the peptides in vitro plasma and be endocytosed selectively into the cancer cells. The histidine-rich lytic peptide component assists the disruption of endosomal/lysosomal membranes and subsequent the mitochondria membrane, which leads to apoptosis. This rational design of MLP-based conjugates provides a practical strategy to increase the application prospects of lytic peptides in cancer treatment.

O-Glycosylation Induces Amyloid-ß To Form New Fibril Polymorphs Vulnerable for Degradation.

Brain accumulation of amyloid-ß (Aß) peptides (resulting from a disrupted balance between biosynthesis and clearance) occurs during the progression of Alzheimer's disease (AD). Aß peptides have diverse posttranslational modifications (PTMs) that variously modulate Aß aggregation into fibrils, but understanding the mechanistic roles of PTMs in these processes remains a challenge. Here, we chemically synthesized three homogeneously modified isoforms of Aß (1-42) peptides bearing Tyr10 O-glycosylation, an unusual PTM initially identified from the cerebrospinal fluid samples of AD patients. We discovered that O-glycans significantly affect both the aggregation and degradation of Aß42. By combining cryo-EM and various biochemical assays, we demonstrate that a Galß1-3GalNAc modification redirects Aß42 to form a new fibril polymorphic structure that is less stable and more vulnerable to Aß-degrading enzymes (e.g., insulin-degrading enzyme). Thus, beyond showing how particular O-glycosylation modifications affect Aß42 aggregation at the molecular level, our study provides powerful experimental tools to support further investigations about how PTMs affect Aß42 fibril aggregation and AD-related neurotoxicity.

Probing N-Glycan Functions in Human Interleukin-17A Based on Chemically Synthesized Homogeneous Glycoforms.

N-Glycosylation represents an essential type of posttranslational modification for proteins. However, deciphering the functions of N-glycosylation remains a challenge due to the lack of analytical and biochemical methods to accurately differentiate the protein glycoforms with various intact glycans. Here we report our synthesis and evaluation of homogeneously glycosylated interleukin-17A (IL-17A), based on a synthetic approach combining solid-phase synthesis of (glyco)peptides, chemoenzymatic glycan modification on segments, and chemical ligations. The obtained homogeneous glycoproteins allow for the demonstration of the stabilizing role of N-glycans during the folding step. A comparison of three IL-17A glycoforms in a normal human dermal fibroblast (NHDF) assay reveals dose-dependent interleukin-6-inducing activities in all cases, wherein the glycoform with sialyl undecasaccharides displays much weaker stimulatory effect than that of the GlcNAc- or GlcNAc(ß1→4)GlcNAc-modified proteins. Further surface plasmon resonance (SPR) and hydrogen/deuterium exchange mass spectroscopic experiments confirm that the evaluated complex type N-glycan impedes the binding between IL-17A and its receptor IL-17RA. This structure-activity relationship study on glycoproteins highlights the viability of applying the de novo approach to probe the roles of N-glycans.

Predicted CD4+ T cell infiltration levels could indicate better overall survival in sarcoma patients.

OBJECTIVE: The role of tumor-infiltrating lymphocytes (TILs) has not yet been characterized in sarcomas. The aim of this bioinformatics study was to explore the effect of TILs on sarcoma survival and genome alterations. METHODS: Whole-exome sequencing, transcriptome sequencing, and survival data of sarcoma were obtained from The Cancer Genome Atlas. Immune infiltration scores were calculated using the Tumor Immune Estimation Resource. Potential associations between abundance of infiltrating TILs and survival or genome alterations were examined. RESULTS: Levels of CD4+ T cell infiltration were associated with overall survival of patients with pan-sarcomas, and higher CD4+ T cell infiltration levels were associated with better survival. Somatic copy number alterations, rather than mutations, were found to correlate with CD4+ T cell infiltration levels. CONCLUSIONS: This data mining study indicated that CD4+ T cell infiltration levels predicted from RNA sequencing could predict sarcoma prognosis, and higher levels of CD4+ T cells infiltration indicated a better chance of survival.

Development and Validation of an Immune-Related Gene Pair Signature in Skin Cutaneous Melanoma.

INTRODUCTION: Skin cutaneous melanoma (SKCM) is a common skin malignancy worldwide, and its metastasis and mortality rates are high. The molecular characteristics exhibited by tumor-immune interactions have drawn the attention from researchers. Therefore, increased knowledge and new strategies to identify effective immune-related biomarkers may improve the clinical management of SKCM by providing more accurate prognostic information. PATIENTS AND METHODS: In this study, we established a prognostic immune-related gene pair (IRGP) signature for predicting the survival of SKCM patients. The Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO) databases provided gene expression profiles together with clinical information, and the samples were randomly divided into three groups including the training, testing, and validation datasets. The regression model of least absolute shrinkage and selection operator (LASSO) helped to identify a 13-IRGP signature with a significant relation to the survival of SKCM patients. RESULTS: The training, TCGA, and independent sets have an average value of area under the curve of 0.79, 0.76, and 0.82, respectively. In addition, this 13-IRGP signature can noticeably divide SKCM patients into high-risk group and low-risk group with significantly different prognoses. Many biological activities such as gene family were enriched among the genes in our IRGP signature. While analyzing the risk signature and clinical characteristics, there was a large difference in the risk score between T stage and tumor stage grouping. Finally, we constructed a nomogram and forest plots of the risk score and clinical features. CONCLUSION: In summary, we developed a robust 13-IRGP prognostic signature in SKCM, which can identify and provide new insights into immunological biomarkers.

Increased expression of the RNA-binding motif protein 47 predicts poor prognosis in non-small-cell lung cancer.

Lung cancer is the leading cause of cancer-associated mortality worldwide. In China, in particular, lung cancer mortality has markedly increased and is likely to continue to rise. RNA-binding proteins are pivotal to the development and progression of a variety of cancer types, including non-small cell lung cancer (NSCLC). RNA-binding motif protein 47 (RBM47) has been found to act as a tumor suppressor in breast cancer and NSCLC. However, to the best of our knowledge, RBM47 expression in NSCLC tissues has yet to be investigated. Analysis via the online database, Gene Expression Omnibus, revealed that RBM47 was upregulated in NSCLC and associated with pathological type, suggesting that RBM47 may play different roles in lung adenocarcinoma and lung squamous cell carcinoma. In the present study, the expression of RBM47 was examined by immunohistochemistry in 175 pairs of tumor and adjacent non-cancerous tissues resected from patients with NSCLC. The results indicated that the expression of RBM47 was significantly increased in NSCLC samples compared with that in the matched non-cancerous samples. Furthermore, RBM47 expression was higher in Xuanwei compared with that in non-Xuanwei NSCLC, suggesting that RBM47 is a more sensitive biomarker in Xuanwei NSCLC, and that it may serve as a candidate therapeutic target. In addition, RBM47 expression was associated with the pathological type, however not with the age, sex, lymph node metastasis, pT stage or pathological Tumor-Node-Metastasis stage of the patients. The increased expression level of RBM47 may indicate a worse overall survival rate for patients with NSCLC. In addition, multivariate survival analysis showed that the Xuanwei area is associated with poor prognosis for patients with NSCLC. In conclusion, the present study revealed that the upregulation of RBM47 accelerated the malignant progression of NSCLC, indicating that RBM47 may be a potential biomarker for NSCLC progression and a therapeutic target for NSCLC.

CD19 and CD30 CAR T-Cell Immunotherapy for High-Risk Classical Hodgkin's Lymphoma.

BACKGROUND: In clinical applications of CAR T-cell therapy, life-threatening adverse events including cytokine release syndrome and neurotoxicity can lead to treatment failure. Outcomes of patients treated with anti-CD30 CAR T- cell have been disappointing in relapsing/refractory (r/r) classical Hodgkin's Lymphoma (cHL). METHODS: In order to understand the applicable population of multiple CAR T-cell therapy, we examined the expression of CD19, CD20, and CD30 by immunohistochemistry (IHC) in 38 paraffin-embedded specimens of cHL. In the past two years, we found only one patient with cHL who is eligible for combined anti-CD19 and CD30 CAR T-cell treatment. This patient's baseline characteristics were prone to severe adverse events. We treated this patient with low doses and multiple infusions of anti-CD19 and CD30 CAR T-cell. RESULTS: The positive expression of CD19+ + CD30+ in Reed-Sternberg (RS) cells is approximately 5.2% (2/38). The patient we treated with combined anti-CD19 and CD30 CAR T-cell did not experience severe adverse events related to CAR T-cell therapy and received long term progression-free survival (PFS). CONCLUSION: For high risk r/r cHL patients, low doses of CAR T-cell used over different days at different times might be safe and effective. More clinical trials are warranted for CD19 and CD30 CAR T-cell combination therapy.

miR-193b Increases the Chemosensitivity of Osteosarcoma Cells by Promoting FEN1-Mediated Autophagy.

BACKGROUND: Osteosarcoma (OS) is one of the most common malignant bone tumors and specific microRNAs (miRNAs) are closely associated with malignant OS progression. In this study, we examined the role of microRNA-193b-3p (miR-193b) and the involvement of autophagy and apoptosis in the chemosensitivity of OS cells. METHODS: We employed qRT-PCR, Western blot, and immunohistochemistry to examine the expression levels of miR-193b, flap endonuclease 1 (FEN1), and autophagy-related proteins. Apoptosis was determined by flow cytometry using an Annexin V-FITC/PI apoptosis detection kit. Luciferase reporter assays confirmed the relationship between miR-193b and FEN1. RESULTS: miR-193b was downregulated in OS compared to adjacent normal tissues (p < 0.05). miR-193b overexpression in the OS cell lines induced autophagy and apoptosis, as shown by Western blotting and flow cytometry. Knockdown of FEN1, a structure-specific nuclease overexpressed in OS tissues (p < 0.001), induced apoptosis through activation of autophagy. Luciferase reporter assays confirmed that FEN1 is a direct target of miR-193b, FEN1 knockdown reinforced miR-193b induced apoptosis. Moreover, miR-193b expression enhanced epirubicin-induced autophagy and apoptosis. CONCLUSION: Collectively, the results showed that miR-193b/FEN1 may serve as a novel therapeutic target for OS aimed mainly at the induction of autophagy and apoptosis. The miR-193b/FEN1 axis increased the chemosensitivity of OS cells, while activation of autophagy enhanced the anticancer effects of epirubicin.

Knockdown of STIM1 expression inhibits non-small-cell lung cancer cell proliferation in vitro and in nude mouse xenografts.

Stromal interaction molecule 1 (STIM1) is a calcium-sensing protein localized in the membrane of the endoplasmic reticulum. The expression of STIM1 has been shown to be closely associated with cell proliferation. The aim of the present study was to investigate the role of STIM1 in the regulation of cancer progression and its clinical relevance. The data demonstrated that the expression of the STIM1 was significantly higher in non-small-cell lung cancer (NSCLC) tissues than in benign lesions and was associated with advanced NSCLC T stage. Knockdown of STIM1 expression in NSCLC cell lines A549 and SK-MES-1 significantly inhibited cell proliferation and induces A549 and SK-MES-1 cell arrest at the G2/M and S phases of the cell cycle. Western blotting showed that the expression of cyclin-dependent kinase (CDK) 1 and CDK2 were reduced while knockdown of STIM1 expression. Furthermore, knockdown of STIM1 in NSCLC cells significantly reduced the levels of xenograft tumor growth in nude mice. These data indicate that aberrant expression of the STIM1 protein may contribute to NSCLC progression. Future studies should focus on targeting STIM1 as a novel strategy for NSCLC therapy.

α-Selective Lysine Ligation and Application in Chemical Synthesis of Interferon Gamma.

A traceless ß-mercaptan-assisted α-selective ligation of N-terminal lysine-containing peptides has been developed. In this ligation-desulfurization-based protocol, the ε-amine of lysine is free of protection, thus improving the overall synthetic efficiency and avoiding harsh reactions in preparing large peptides and proteins. The applicability of this methodology has been demonstrated in the synthesis of an acid-labile therapeutic protein, interferon gamma, and the anticancer activity of synthetic protein has also been evaluated.

ß-Glucan as an immune activator and a carrier in the construction of a synthetic MUC1 vaccine.

We describe the preparation of a cancer vaccine candidate by conjugating a MUC1 peptide antigen to the ß-glucan polysaccharide, which serves both as a carrier and an immune activator. In contrast to amorphous polysaccharides, peptide-ß-glucan conjugates form uniform nanoparticles that facilitate the delivery of antigens and binding to myeloid cells, thus leading to the activation of both innate and adaptive immunity.