José Carlos Páez Franco

Statins are clinically effective drugs for treating dyslipidemia and have been proposed as promising antineoplastic and adjuvant agents in cancer therapy for years due to their impact on dysregulated cell growth processes, including cell signaling, energetics, and membrane synthesis. Despite being potent inhibitors of mevalonate synthesis and its downstream products, their limited clinical success highlights the need to further explore their mechanistic effects. Leveraging the observed sensitivity of muscle cells to atorvastatin in clinical settings and utilizing untargeted metabolomic analysis of atorvastatin-treated RD rhabdomyosarcoma cells, we identified reduced levels of aminoadipic acid, an intermediate in lysine catabolism. We investigated whether metabolic sensitization of RD cells to lysine-related metabolites (lysine, aminoadipic acid, pipecolic acid, glutamic acid, α-ketoglutarate, and lysine-p-nitroanilide) prior to atorvastatin treatment enhances its cytotoxic effects. Metabolic sensitization or reprogramming involves cellular processes wherein cells adapt their metabolism to environmental changes, reflecting alterations in enzymatic activity, transport, and stress response thresholds. These adaptations enable cells to cope with specific environmental pressures but may impair their ability to respond to other stressors or stimuli. To evaluate the impact of metabolic supplementation, we analyzed cellular stress response markers via western blot. The results revealed that lysine-p-nitroanilide increased BiP, the master regulator of the unfolded protein response, and augmented the phosphorylation at threonine 172 of AMPK, an indicator of altered cellular energetics. Further analysis demonstrated that combining lysine-p-nitroanilide with atorvastatin disrupted mitochondrial homeostasis and reduced glycolysis, both desirable outcomes in antineoplastic treatments. Lysine-p-nitroanilide acts as an in vitro inhibitor of α-aminoadipic semialdehyde synthase, enzyme essential for lysine metabolism via the saccharopine pathway. However, we demonstrated that it is catabolically cleaved to p-nitroanilide, with this molecule driving the cytotoxic activity observed in our experiments. Although lysine metabolism was not fully suppressed by lysine-p-nitroanilide, these findings provide valuable insights for developing novel therapies for rhabdomyosarcoma.

Protein tyrosine phosphatase 1B (PTP1B) is a crucial enzyme involved in regulating insulin and leptin signaling pathways, making it a promising target for treating type 2 diabetes. In this study, we synthesized 14 derivatives of indole- and pyrazole-fused glycyrrhetinic acid (GA) and evaluated their effects on PTP1B, using both its long (hPTP1B_1–400) and short (hPTP1B_1–285) forms. We analyzed enzymatic kinetics and selectivity over T-cell protein tyrosine phosphatase (TCPTP). Molecular docking and molecular dynamics simulations were performed to understand the binding mode of the compounds within PTP1B. Untargeted metabolomics, using gas chromatography–mass spectrometry, assessed metabolic changes caused by the most potent PTP1B inhibitors in HepG2 cells. We also evaluated these inhibitors in vivo to determine their effects on insulin sensitivity through the insulin tolerance test (ITT) in streptozotocin (STZ)-induced diabetic mice. Two compounds, 4b (indole-fused) and 5g (pyrazole-fused), showed greater potency against the long form of PTP1B compared to the short form, indicating that both compounds preferentially bind to the disordered C-terminal region of PTP1B. Molecular docking and molecular dynamics studies supported this finding. Furthermore, enzymatic kinetics revealed that compounds 4b and 5g function as uncompetitive inhibitors of PTP1B, with K _i values of 0.32 and 0.72 μM, respectively. Notably, both GA derivatives exhibited more pronounced inhibition of PTP1B compared to the well-established inhibitors ursolic acid and Ertiprotafib, while also demonstrating selectivity over TCPTP. Metabolomic analysis revealed that these compounds increased pantothenic acid and glycine levels, while decreasing glucose and fatty acid levels in HepG2 cells, suggesting enhanced glycolysis and reduced lipogenesis. Both compounds exhibited low cytotoxicity in HFF-1 cells and significantly reduced glucose levels in the ITT in STZ-induced diabetic mice, outperforming the insulin-sensitizing drug Pioglitazone.

Background/Objective: Human milk is an irreplaceable source of nutrition and is essential for the infant’s growth and development right after birth and for early life stage survival. This study aims to characterize and compare the metabolite profiles of colostrum and transitional and mature milk using an untargeted GC-MS approach. Additionally, it explores potential correlations between the identified metabolites and maternal nutritional factors. Methods: This was a longitudinal, prospective, and observational study. We included human milk samples from 113 Mexican women who practiced exclusive breastfeeding. Partial least squares-discriminant analysis (PLS-DA) was performed to assess differences among lactation stages. Metabolites showing significant variation across lactation stages were further analyzed using Friedman tests with post hoc Wilcoxon tests and Bonferroni correction. Correlations with maternal anthropometric measures were evaluated. Results: Twenty-three metabolites were identified, including amino acids and derivatives, sugars, fatty acids, and energetic metabolites. Alanine and creatinine levels decreased during lactation, while aspartate, serine, and valine levels increased. Rhamnose level was higher in colostrum, whereas decanoic, dodecanoic, and tetradecanoic acid levels increased over time, and that of 11,14-eicosadienoic acid decreased. Lactic acid levels declined across stages. Negative correlations were found between several amino acids and maternal anthropometric variables, while glyceric acid, rhamnose and lactic acid correlated positively. Conclusions: Human milk metabolomic profiles display distinct, stage-specific variations shaped by maternal characteristics, reflecting the dynamic physiological and nutritional demands of the developing infant

Background/Objective: Human milk is an irreplaceable source of nutrition, is essential for the infant’s growth and development right after birth and for early life stages survival. This study aims to characterize and compare the metabolite profiles of colostrum, transitional, and mature milk using an untargeted GC-MS approach. Additionally, it explores potential correlations between the identified metabolites and maternal nutritional factors. Methods: This was a longitudinal, prospective, and observational study. We included human milk samples from 113 Mexican women who practiced exclusive breastfeeding. Partial least squares-discriminant analysis (PLS-DA) was performed to assess differences among milk types. Metabolites showing significant variation across lactation stages were further analyzed using Friedman tests with post hoc Wilcoxon tests and Bonferroni correction. Correlations with maternal anthropometric were evaluated. Results: Twenty-three metabolites were identified, including amino acids and derivatives, sugars, fatty acids, and energetic metabolites. Alanine and creatinine decreased during lactation, while aspartate, serine, and valine increased. Rhamnose was higher in colostrum, whereas decanoic, dodecanoic, and tetradecanoic acids increased over time, and 11,14-eicosadienoic acid decreased. Lactic acid declined across stages. Positive correlations were found between several amino acids and maternal anthropometric variables, while lactic acid correlated negatively. Conclusions: Human milk metabolomic profiles display distinct, stage-specific variations shaped by maternal characteristics, reflecting the dynamic physiological and nutritional demands of the developing infant.

Protein tyrosine phosphatase 1B (PTP1B) is crucial for negatively regulating the canonical insulin and leptin signaling pathways. This enzyme is a validated target for treating various disorders, including diabetes and obesity. However, to date, no PTP1B inhibitors have been approved for use. In earlier studies, we developed two modified versions of 18β‐glycyrrhetinic acid (18β‐GA) called FC‐114 and FC‐122 , which showed better inhibitory PTP1B activity than ursolic acid, a well‐known inhibitor. To develop even stronger inhibitors, we looked at another compound, 18α‐glycyrrhetinic acid (18α‐GA), which is more potent than 18β‐GA. Thus, in this study, we aimed to synthesize the analogs 18epi‐FC114 ( 3c ) and 18epi‐FC‐122 ( 5c ). These compounds were prepared with and without the carbonyl group at C11. The results showed that converting 18β‐H to 18α‐H, as well as the absence of the 11‐carbonyl group, negatively impacted the PTP1B inhibitory activity. However, the synthesized compounds exhibited an uncompetitive type of inhibition toward PTP1B and did not inhibit the TCPTP enzyme. Molecular docking and dynamics simulations suggest that the inversion of 18β‐H pushes the 30‐COOH group away, disrupting interactions at the C‐terminal site of PTP1B _1–400 . Additionally, the absence of the 11‐carbonyl group positions the compounds unfavorably, limiting critical interactions in the same region.

Background/Objectives: COVID-19 is a systemic viral infection that may lead to serious complications including acute kidney injury that requires kidney replacement therapy. The primary aim of this study was to evaluate urinary SerpinA3 (uSerpinA3) excretion as a biomarker of kidney recovery at 90 days, and the mortality in patients with critical COVID-19 and AKI requiring kidney replacement therapy (KRT). Methods: The study included patients with critical COVID-19 on invasive mechanical ventilation (IMV) requiring KRT. Blood and urine samples were obtained when KRT was initiated (day zero), and thereafter on days 1, 3, 7, and 14 post-replacement. uSerpinA3, kidney injury molecule-1 (uKIM-1), and neutrophil gelatinase-associated lipocalin (uNGAL) were measured in urine, and interleukin-6 (IL-6), interleukin-10 (IL-10), and tumor necrosis factor alpha (TNF-α) in peripheral blood. In addition, metabolomics in sample days zero and 3, and in the survivors on sample day 90 was performed by employing gas chromatography coupled with mass spectrometry. Results: A total of 60 patients were recruited, of whom 29 (48%) survived hospitalization and recovered kidney function by day 90. In the survivors, 79% presented complete recovery (CRR) and the remaining (21%) recovered partially (PRR). In terms of uSerpinA3, levels on days 7 and 14 predicted CRR, with AUC values of 0.68 (p = 0.041) and 0.71 (p = 0.030), respectively, as well as mortality, with AUC values of 0.75 (p = 0.007) and 0.76 (p = 0.015), respectively. Among the other biomarkers, the excretion of uKIM-1 on day zero of KRT had a superior performance as a CRR predictor [(AUC, 0.71 (p = 0.017)], and as a mortality predictor [AUC, 0.68 (p = 0.028)]. In the metabolomics analysis, we identified four distinct profiles; the metabolite that maintained statistical significance in predicting mortality was p-cresol glucuronide. Conclusions: This study strongly suggests that uSerpinA3 and uKIM-1 can predict CRR and mortality in patients with critical COVID-19 and AKI requiring KRT. Metabolic analysis appears promising for identifying affected pathways and their clinical impact in this population.

The coronavirus disease 2019 (COVID-19) survivors are frequently observed to present persistent symptoms constituting what has been called “post-acute COVID-19 syndrome” (PACS) or “long COVID-19”. Some clinical risk factors have been identified to be associated with PACS development; however, specific mechanisms responsible for PACS pathology remain unknown. This study investigates clinical, immunological, and metabolomic risk factors associated with post-acute COVID-19 syndrome (PACS) in 51 patients, assessed 7–19 months after acute infection. Among the participants, 62.7% were male and 37.2% were female, with an average age of 47.8 years. At the follow-up, 37.2% met the criteria for PACS, revealing significant differences in immunological and metabolomic profiles at the time of acute infection. Patients with PACS were characterized by elevated levels of mature low-density granulocytes (LDGs), interleukin-8 (IL-8), pyruvate, pseudouridine, and cystine. Baseline multivariate analysis showed increased pyruvate and decreased alpha tocopherol levels. At follow-up, there was a decrease in absolute B lymphocytes and an increase in non-classical monocytes and 3-hydroxyisovaleric acid levels. These findings suggest that specific immunological and metabolomic markers during acute infection can help identify patients at higher risk of developing persistent PACS.

The relevance of the fluorinated thiolate 2,3,5,6-S(C₆F₄H-4) to serve as a reliable motif for the construction of π_F–π_F stacking interactions in a series of metal complexes in the solid state for its application in crystal engineering is Illustrated.

Coamorphous salt in a 1:1 ratio prepared by ball milling from Fluvastatin sodium (FLV) and Pioglitazone hydrochloride (PGZ·HCl) can be selectively formed by neat grinding (NG). Furthermore, the salt–cocrystal continuum was preferably formed by employing liquid-assisted grinding (LAG) using ethanol (EtOH). Attempts to prepare the coamorphous salt starting from the salt–cocrystal continuum by NG were unsuccessful. Interestingly, through ball milling by NG or LAG, a great diversity of solid forms (PGZ·HCl-FLV 1:1) could be accessed: NG and hexane (coamorphous); ethyl acetate (physical mixture); EtOH (salt–cocrystal continuum); and water (which presents two T_g, indicating immiscibility of the components). An exploration was performed at different drug-to-drug ratios by NG. By differential scanning calorimetry (DSC), the presence of two endothermic events was observed in this screening: incongruous melting point (solidus) and excess of one of the components (liquidus), except in the 1:1 solid form. From these results, eutectic behavior was observed. Through the construction of a binary phase diagram, it was determined that the 1:1 molar ratio gives rise to the formation of the most stable coamorphous composition. Dissolution profile studies of these solid forms were carried out, specifically on pure FLV and the solid forms of PGZ⋅HCl-FLV (1:2; 1:4; and 1:6), together with the coamorphous 1:1 salt. By itself, pure FLV presented the highest K_int (13.6270 ± 0.8127 mg/cm²⋅min). On the other hand, the coamorphous 1:1 showed a very low K_int (0.0220 ± 0.0014 mg/cm²·min), indicating very fast recrystallization by the FLV, which avoids observing a sudden release of this drug in the solution. This same behavior was observed in the eutectic composition 1:2. In the other solid forms, the value of K_int increases along with the %w of FLV. From the mechanochemical point of view, ball milling by NG or LAG became an important synthetic tool since it allows obtaining a great variety of solid forms to explore the solid-state reactivity of the drug–drug solid-form PGZ HCl-FLV.

Ball-milling using neat grinding (NG) or liquid-assisted grinding (LAG) by varying the polarity of the solvents allowed access to various drug–drug solid forms of pioglitazone hydrochloride (PGZ·HCl) and rosuvastatin calcium (RSV). Using NG, the coamorphous form was formed from the reaction of pioglitazone hydrochloride (PGZ·HCl) and rosuvastatin calcium (RSV) in a 2:1 molar ratio. The formation of the expected coamorphous salt could not be corroborated by FT-IR, but DSC data showed that it was indeed a single-phase amorphous mixture. By varying the molar ratios of the reactants, either keeping PGZ·HCl constant and varying RSV or vice versa, another coamorphous form was obtained when a 1:1 molar ratio was employed. In the case of the other outcomes, it was observed that they were a mixture of solid forms coexisting simultaneously with the coamorphous forms (1:1 or 2:1) together with the drug that was in excess. When RSV was in excess, it was in an amorphous form. In the case of PGZ·HCl, it was found in a semicrystalline form. The intrinsic dissolution rates (IDRs) of the solid forms of PGZ·HCl-RSV in stoichiometric ratios (1:1, 2:1, 1:4, 6:1, and 1:10) were evaluated. Interestingly, a synchronized release of both drugs in the dissolution medium was observed. In the case of the release of RSV, there were no improvements in the dissolution profiles, because the acidic media caused the formation of degradation products, limiting any probable modification in the dissolution processes. However, the coamorphous 2:1 form exhibited an improvement of 1.03 times with respect to pure PGZ·HCl. It is proposed that the modification of the dissolution process of the coamorphous 2:1 form was limited by changes in the pH of the media as RSV consumes protons from the media due to degradation processes.