Metabolomic Profiling of Breast Cancer Patients Undergoing Neoadjuvant Chemotherapy for Predicting Disease-Free and Overall Survival.

Breast cancer (BC) remains a significant global health concern, with neoadjuvant chemotherapy (NACT) offering preoperative benefits like tumor downstaging and treatment response assessment. However, identifying factors influencing post-NACT treatment response and survival outcomes is challenging. Metabolomic approaches offer promising insights into understanding these outcomes. This study analyzed the serum of 80 BC patients before and after NACT, followed for up to five years, correlating with disease-free survival (DFS) and overall survival (OS). Using untargeted nuclear magnetic resonance (NMR) spectroscopy and a novel statistical model that avoids collinearity issues, we identified metabolic changes associated with survival outcomes. Four metabolites (histidine, lactate, serine, and taurine) were significantly associated with DFS. We developed a metabolite-related survival score (MRSS) from these metabolites, stratifying patients into low- and high-risk relapse groups, independent of classical prognostic factors. High-risk patients had a hazard ratio (HR) for DFS of 3.42 (95% CI 1.51-7.74; p = 0.003) after adjustment for disease stage and age. A similar trend was observed for OS (HR of 3.34, 95% CI 1.64-6.80; p < 0.001). Multivariate Cox proportional hazards analysis confirmed the independent prognostic value of the MRSS. Our findings suggest the potential of metabolomic data, alongside traditional markers, in guiding personalized treatment decisions and risk stratification in BC patients undergoing NACT. This study provides a methodological framework for leveraging metabolomics in survival analyses.

Metabolomics by NMR Combined with Machine Learning to Predict Neoadjuvant Chemotherapy Response for Breast Cancer.

Neoadjuvant chemotherapy (NACT) is offered to patients with operable or inoperable breast cancer (BC) to downstage the disease. Clinical responses to NACT may vary depending on a few known clinical and biological features, but the diversity of responses to NACT is not fully understood. In this study, 80 women had their metabolite profiles of pre-treatment sera analyzed for potential NACT response biomarker candidates in combination with immunohistochemical parameters using Nuclear Magnetic Resonance (NMR). Sixty-four percent of the patients were resistant to chemotherapy. NMR, hormonal receptors (HR), human epidermal growth factor receptor 2 (HER2), and the nuclear protein Ki67 were combined through machine learning (ML) to predict the response to NACT. Metabolites such as leucine, formate, valine, and proline, along with hormone receptor status, were discriminants of response to NACT. The glyoxylate and dicarboxylate metabolism was found to be involved in the resistance to NACT. We obtained an accuracy in excess of 80% for the prediction of response to NACT combining metabolomic and tumor profile data. Our results suggest that NMR data can substantially enhance the prediction of response to NACT when used in combination with already known response prediction factors.

Electrospun Nanofibrous Architectures of Thrombin-Loaded Poly(ethylene oxide) for Faster in Vivo Wound Clotting.

Wound healing materials to prevent blood loss are crucial during emergency medical treatment because uncontrolled bleeding can lead to patient death. Herein, bioabsorbable fibrous architectures of thrombin-loaded poly(ethylene oxide)-PEO/thrombin-are conceptualized and accomplished via electrospinning for faster wound clotting. Membranes with average fiber diameters ranging from 188 to 264 nm are achieved, where the active thrombin is entrapped within the nanofibers. The results of in vitro and in vivo wound healing activity tests revealed that when the nanofibers with thrombin-loaded capacity are in contact with the wound, the presence of water in the skin or blood catalyzes the degradation of the membranes, thus releasing thrombin. Thrombin then accelerates the wound clotting process. In contrast to other hemostatic materials, PEO/thrombin nanofibers do not require mechanical removal after application, and the viscoelastic nature of such biomaterials enables their conformation to a variety of wound topographies. Remarkably, PEO/thrombin membranes are promising functional materials and their use is a powerful strategy for hemostatic treatment, ranging from simple first aid and sealing to a wound to small surgical procedures.

Using a Fragment-Based Approach to Identify Alternative Chemical Scaffolds Targeting Dihydrofolate Reductase from Mycobacterium tuberculosis.

Dihydrofolate reductase (DHFR), a key enzyme involved in folate metabolism, is a widely explored target in the treatment of cancer, immune diseases, bacteria, and protozoa infections. Although several antifolates have proved successful in the treatment of infectious diseases, they have been underexplored to combat tuberculosis, despite the essentiality of M. tuberculosis DHFR (MtDHFR). Herein, we describe an integrated fragment-based drug discovery approach to target MtDHFR that has identified hits with scaffolds not yet explored in any previous drug design campaign for this enzyme. The application of a SAR by catalog strategy of an in house library for one of the identified fragments has led to a series of molecules that bind to MtDHFR with low micromolar affinities. Crystal structures of MtDHFR in complex with compounds of this series demonstrated a novel binding mode that considerably differs from other DHFR antifolates, thus opening perspectives for the development of relevant MtDHFR inhibitors.

Unsaturated fatty acids as high-affinity ligands of the C-terminal Per-ARNT-Sim domain from the Hypoxia-inducible factor 3α.

Hypoxia-inducible transcription factors (HIF) form heterodimeric complexes that mediate cell responses to hypoxia. The oxygen-dependent stability and activity of the HIF-α subunits is traditionally associated to post-translational modifications such as hydroxylation, acetylation, ubiquitination, and phosphorylation. Here we report novel evidence showing that unsaturated fatty acids are naturally occurring, non-covalent structural ligands of HIF-3α, thus providing the initial framework for exploring its exceptional role as a lipid sensor under hypoxia.

Functional diversification of cerato-platanins in Moniliophthora perniciosa as seen by differential expression and protein function specialization.

Cerato-platanins (CP) are small, cysteine-rich fungal-secreted proteins involved in the various stages of the host-fungus interaction process, acting as phytotoxins, elicitors, and allergens. We identified 12 CP genes (MpCP1 to MpCP12) in the genome of Moniliophthora perniciosa, the causal agent of witches' broom disease in cacao, and showed that they present distinct expression profiles throughout fungal development and infection. We determined the X-ray crystal structures of MpCP1, MpCP2, MpCP3, and MpCP5, representative of different branches of a phylogenetic tree and expressed at different stages of the disease. Structure-based biochemistry, in combination with nuclear magnetic resonance and mass spectrometry, allowed us to define specialized capabilities regarding self-assembling and the direct binding to chitin and N-acetylglucosamine (NAG) tetramers, a fungal cell wall building block, and to map a previously unknown binding region in MpCP5. Moreover, fibers of MpCP2 were shown to act as expansin and facilitate basidiospore germination whereas soluble MpCP5 blocked NAG6-induced defense response. The correlation between these roles, the fungus life cycle, and its tug-of-war interaction with cacao plants is discussed.

Structure, dynamics, and RNA interaction analysis of the human SBDS protein.

Shwachman-Bodian-Diamond syndrome is an autosomal recessive genetic syndrome with pleiotropic phenotypes, including pancreatic deficiencies, bone marrow dysfunctions with increased risk of myelodysplasia or leukemia, and skeletal abnormalities. This syndrome has been associated with mutations in the SBDS gene, which encodes a conserved protein showing orthologs in Archaea and eukaryotes. The Shwachman-Bodian-Diamond syndrome pleiotropic phenotypes may be an indication of different cell type requirements for a fully functional SBDS protein. RNA-binding activity has been predicted for archaeal and yeast SBDS orthologs, with the latter also being implicated in ribosome biogenesis. However, full-length SBDS orthologs function in a species-specific manner, indicating that the knowledge obtained from model systems may be of limited use in understanding major unresolved issues regarding SBDS function, namely, the effect of mutations in human SBDS on its biochemical function and the specificity of RNA interaction. We determined the solution structure and backbone dynamics of the human SBDS protein and describe its RNA binding site using NMR spectroscopy. Similarly to the crystal structures of Archaea, the overall structure of human SBDS comprises three well-folded domains. However, significant conformational exchange was observed in NMR dynamics experiments for the flexible linker between the N-terminal domain and the central domain, and these experiments also reflect the relative motions of the domains. RNA titrations monitored by heteronuclear correlation experiments and chemical shift mapping analysis identified a classic RNA binding site at the N-terminal FYSH (fungal, Yhr087wp, Shwachman) domain that concentrates most of the mutations described for the human SBDS.

Lytic activity and structural differences of amphipathic peptides derived from trialysin.

Trialysin is a pore-forming protein found in the saliva of Triatoma infestans (Hemiptera, Reduviidae), the insect vector of Chagas' disease. The protein is active against a broad range of cell types from bacteria to eukaryotic cells. Recognizing that the N-terminus of trialysin harbors the lytic motif [Amino, R., Martins, R. M., Procopio, J., Hirata, I. Y., Juliano, M. A., and Schenkman, S. (2002) J. Biol. Chem. 277, 6207-6213], we designed a set of peptides scanning this region to investigate the structural basis of its biological function. Peptides encompassing residues 1-32 (P6), 1-27 (P7), and 6-32 (P5) efficiently induced lysis of the protozoan parasite Trypanosoma cruzi and Escherichia coli in the 0.4-9.0 microM range, while much higher concentrations were required to cause hemolysis. Other more internal peptides, including peptide P2 (residues 21-47) and others up to residue 52, were less effective. P6 turned out to be the most active of all. P7 has a significantly higher activity than P5 against E. coli, while P5 has a hemolytic activity comparable to that of P6. CD spectroscopy showed that all tested peptides acquire a comparable helical content in solvent mixtures or in detergent micelles. The solution structure of P2 and P5-P7 was determined in a 30% trifluoroethanol/water mixture by nuclear magnetic resonance. All peptides exhibit a structure characterized by a central helical fold, and except for P2, which does not show a continuous hydrophobic surface, they are amphipathic. The structural models show that P5 and P7 extend their structural similarities with the most active peptide, P6, in either the C-terminus or the N-terminus. Amino acid substitutions in the N-terminus of P6 improved hemolysis but did not change the activity against T. cruzi. These results suggest that while amphipathicity is essential for the lytic activity, the selectivity of the active peptides for specific organisms appears to be associated with the structural features of their N- and C-termini.

The NMR-derived solution structure of a new cationic antimicrobial peptide from the skin secretion of the anuran Hyla punctata.

Amphibian skin secretions constitute an important source of molecules for antimicrobial drug research in order to combat the increasing resistance of pathogens to conventional antibiotics. Among the various types of substances secreted by the dermal granular amphibian glands, there is a wide range of peptides and proteins, often displaying potent antimicrobial activities and providing an effective defense system against parasite infection. In the present work, we report the NMR solution structure and the biological activity of a cationic 14-residue amphiphilic alpha-helical polypeptide named Hylaseptin P1 (HSP1), isolated from the skin secretion of the hylid frog Hyla punctata. The peptide antimicrobial activity was verified against Candida albicans, Staphylococcus aureus, Escherichia coli, and Pseudomonas aeruginosa, whereas no significant lytic effect was detected toward red or white blood cells.