Effects of Acute and One-Week Supplementation with Montmorency Tart Cherry Powder on Food-Induced Uremic Response and Markers of Health: A Proof-of-Concept Study.

Metabolic conditions, such as gout, can result from elevated uric acid (UA) levels. Consuming high-purine meals increases UA levels. Therefore, people with hyperuricemia typically must avoid ingesting such foods. Polyphenols have been shown to reduce uric acid levels and tart cherries (TCs) are a rich source of phenolic and anthocyanin compounds. This proof-of-concept study evaluated whether ingesting TCs with a purine-rich meal affects the uricemic response. Methods: A total of 25 adults (15 males and 10 females, 85.0 ± 17 kg, 40.6 ± 9 years, 29.1 ± 4.9 kg/m2) with elevated fasting UA levels (5.8 ± 1.3 mg/dL) donated a fasting blood sample. In a randomized, double-blind, crossover, placebo-controlled, counterbalanced manner, participants ingested capsules containing 960 mg of a placebo (PLA) or concentrated TC powder containing 20.7 mg of proanthocyanins with a serving of hot soup (10 g of carbohydrate, 2 g protein, and 1 g fat) containing 3 g of purines (1 g of adenosine 5'-monophosphate, 1 g of disodium 5'-guanylate, and 1 g of disodium 5'-inosinate). Blood samples were obtained at 0, 60, 120, 180, and 240 min after ingestion to assess changes in uric acid levels and pharmacokinetic profiles. Cell blood counts, a comprehensive metabolic panel, cytokines, inflammatory markers, and subjective side effects ratings were analyzed on baseline (0 min) and post-treatment (240 min) samples. Participants continued consuming two capsules/day of the assigned treatment for one week and then repeated the experiment. Participants observed a 14-day washout and then repeated the experiment while ingesting the alternate treatment. Data were analyzed using general linear model (GLM) statistics with repeated measures, pairwise comparisons, and percentage change from baseline with 95% confidence intervals (CIs). Results: No statistically significant interaction effects or differences between treatments were seen in uric acid levels or PK profiles. Analysis of percent changes from baseline revealed that TC ingestion reduced the blood glucose levels following the ingestion of the high-purine meal (-4.2% [-7.7, -0.7], p = 0017). Additionally, there was some evidence that TC ingestion attenuated the increase from baseline in IL-1ß and IL-10 and increased INF-γ. No significant differences were seen in the remaining health markers or subjective side effects ratings. Conclusions: Acute and one-week TC supplementation did not affect the uricemic response to ingesting a high-purine meal in individuals with mildly elevated UA levels. However, there was some evidence that TC supplementation may blunt the glycemic response to ingesting a meal and influence some inflammatory cytokines. Registered clinical trial NCT04837274.

Interaction of Tri-Cyclic Nucleobase Analogs with Enzymes of Purine Metabolism: Xanthine Oxidase and Purine Nucleoside Phosphorylase.

Fluorescent markers play important roles in spectroscopic and microscopic research techniques and are broadly used in basic and applied sciences. We have obtained markers with fluorescent properties, two etheno derivatives of 2-aminopurine, as follows: 1,N2-etheno-2-aminopurine (1,N2-ε2APu, I) and N2,3-etheno-2-aminopurine (N2,3-ε2APu, II). In the present paper, we investigate their interaction with two key enzymes of purine metabolism, purine nucleoside phosphorylase (PNP), and xanthine oxidase (XO), using diffraction of X-rays on protein crystals, isothermal titration calorimetry, and fluorescence spectroscopy. Crystals were obtained and structures were solved for WT PNP and D204N-PNP mutant in a complex with N2,3-ε2APu (II). In the case of WT PNP-1,N2-ε2APu (I) complex, the electron density corresponding to the ligand could not be identified in the active site. Small electron density bobbles may indicate that the ligand binds to the active site of a small number of molecules. On the basis of spectroscopic studies in solution, we found that, in contrast to PNP, 1,N2-ε2APu (I) is the ligand with better affinity to XO. Enzymatic oxidation of (I) leads to a marked increase in fluorescence near 400 nm. Hence, we have developed a new method to determine XO activity in biological material, particularly suitable for milk analysis.

Investigating the hepato-protective properties of chamomile oil and olive leaves extracts against ribociclib-induced hepatotoxicity.

A new approach to overcome or reduce these toxicities is by using antioxidants. Ribociclib, a CDK4/6 inhibitor used in the treatment of breast cancer, has been linked to hepatotoxicity and may contribute to the development of Hepatocellular carcinoma in rats. This Study aims to assess hepatoprotective effect of chamomile oil and olive leaf extracts against ribociclib-induced Hepatotoxicity in rats. A total of 40 adult male albino rats aged 9-10 weeks were utilized in this experiment. These rats were divided into four groups, (N=10). Group A (control) comprised normal rats administered 1 ml (10 ml/kg/day) of normal saline daily. Conversely, group B rats were administered 5 mg/kg Ribociclib (n = 10), while group C was administered 5 mg/kg Ribociclib with a 500 mg/kg/day dose of chamomile oil. Group D was given 5 mg \kg Ribociclib in combination with 200 mg/kg/day of olive leaves. After the trial, the animals were sacrificed, blood samples were collected for biochemical tests, and the liver tissue was processed for histological examination. The results of histology, and biochemistry parameter analysis, indicated that co-administration of Ribociclib and chamomile oil plays a decisive role in mitigating the hepatotoxicity result from Ribociclib-induced liver injuries in rats as demonstrated by histological and biochemical parameters.The levels of cholesterol and LDL in the blood were significantly lower (P < 0.01) after administering chamomile oil compared to the control group. The results of the current study demonstrated that the simultaneous use of chamomile oil and olive leaf extract significantly reduced the liver damage caused by Ribociclib and improved the lipid profile in Albino rats. Additionally, the use of chamomile extract notably lowered urea levels (p < 0.01), indicating a protective effect on the kidneys.

Neurological and Psychiatric Adverse Events Associated with Cyclin-Dependent Kinase 4/6 Inhibitors in Breast Cancer Patients: Insights from a Pharmacovigilance Study via the FDA Adverse Event Reporting System.

BACKGROUND AND OBJECTIVE: Cyclin-dependent kinase 4/6 (CDK4/6) inhibitors have revolutionised cancer therapy, particularly breast cancer therapy. However, concerns about their potential to cause neurological and psychiatric adverse events (AEs) have emerged, and these concerns remain underexplored. This study aimed to investigate the signals related to neurological and psychiatric AEs associated with CDK4/6 inhibitor use. METHODS: A retrospective study was performed to analyse reports of AEs associated with the use of CDK4/6 inhibitors (abemaciclib, ribociclib and palbociclib) from the first quarter of 2015 to the fourth quarter of 2023 on the basis of the FDA Adverse Event Reporting System (FAERS). Both the reporting odds ratio (ROR) and the multi-item gamma Poisson shrinker (MGPS) were used for signal detection. The timing of events was assessed with the Weibull shape parameter (WSP). The management, analysis and presentation of the data were performed via Python (version 3.8) and R software (version 4.3.2). RESULTS: A total of 19,001 AE reports in which CDK4/6 inhibitors were identified as the 'primary suspect drug' were included in this study. These events were predominantly observed in patients aged 65 to 85 years. Through an ROR analysis, 85 positive signals for neurological and psychiatric AEs associated with CDK4/6 inhibitors were identified. The MGPS method revealed 61 positive AE signals for neurological and psychiatric AEs associated with CDK4/6 inhibitors. A total of 34 positive AE signals were identified by both the ROR and MGPS analyses. The WSP indicated that the onset times for AEs associated with all three CDK4/6 inhibitors tended to be early in drug therapy, suggesting a propensity for early failure type. CONCLUSION: The present study revealed neurological and psychiatric AEs associated with CDK4/6 inhibitors that often occur early in treatment. Significant signals include spinal cord herniation and cerebral microangiopathy. Close monitoring of these AEs is crucial. Further studies are necessary to verify the connection between CDK4/6 inhibitors and neurological and psychiatric AEs.

From joint pains to hair gains: baricitinib's double duty for rheumatoid arthritis and alopecia areata.

Personalised medicine is a key goal across medical specialties today: using biomarkers and knowledge of pathophysiology to ensure the right patients get the right treatment. This becomes more challenging when patients have more than one disease requiring a targeted treatment. Autoimmune diseases commonly co-occur, and thus, multidisciplinary working is important in rheumatology. We present a case where a patient with a new diagnosis of alopecia areata on a background of rheumatoid arthritis was successfully treated with baricitinib monotherapy, with improvement in both conditions.

Normalized Interferon Signatures and Clinical Improvements by IFNAR1 Blocking Antibody (Anifrolumab) in Patients with Type I Interferonopathies.

PURPOSE: A causal role of type-I interferons (IFN-I) in autoinflammatory type-I interferonopathies such as SAVI (STING-associated vasculopathy with onset in infancy) and CANDLE (chronic atypical neutrophilic dermatosis with lipodystrophy and elevated temperatures) is suggested by elevated expression of IFN-I stimulated genes (ISGs). Hitherto, the lack of specific inhibitors of IFN-I signaling has prevented the verification of a causal role for IFN-I in these conditions. Commonly used inhibitors of the JAK/STAT pathway exert broad effects on multiple signaling pathways leading to more general immunosuppression beyond IFN-I signaling. METHODS: Here we show in four patients with SAVI and one patient with CANDLE syndrome that blockade of the IFNAR1 receptor (Anifrolumab) exerts an additive effect over JAK-inhibitor alone. In two patients with SAVI, monotherapy with Anifrolumab is sufficient to retain a suppressed IFN-I signature and clinical improvement. RESULTS: Anifrolumab normalizes IFN-I signature genes and relieves symptoms beyond what is typically achieved by a JAK-inhibitor (Baricitinib) alone in patients with type-I interferonopathies. In two patients Anifrolumab was used successfully as monotherapy. Addition of Anifrolumab enabled steroid tapering and cessation with reduced overall immunosuppression and lower risks of opportunistic infections and improved metabolic states and growth which is highly beneficial in these young patients. CONCLUSION: These results verify a causal role of IFN-I signaling in type-I Interferonopathies SAVI and CANDLE and suggests Anifrolumab as an important new treatment option in autoinflammatory diseases with elevated IFN-I induced gene expression. Genia Kretzschmar, Laura Piñero Páez, and Ziyang Tan are shared-first authors. Sara Alehashemi, AnnaCarin Horne, and Petter Brodin are co-senior author.

The de novo purine synthesis enzyme Adssl1 promotes cardiomyocyte proliferation and cardiac regeneration.

There is a short window during which the neonatal heart has the proliferative capacity to completely repair damage, an ability that is lost in adulthood. Inducing proliferation in adult cardiomyocytes by reactivating cell cycle reentry after myocardial infarction (MI) improves cardiac function. De novo purine synthesis is a critical source of nucleotides for cell proliferation. Here, using loss- and gain-of-function genetic approaches, we explored the role of the muscle-specific de novo purine synthesis enzyme Adssl1 in cardiac regeneration. Deletion of Adssl1 in mouse neonatal hearts reduced cardiomyocyte proliferation and attenuated heart regeneration after apical resection. Conversely, cardiomyocyte-specific Adssl1 overexpression extended the postnatal regenerative window and induced robust cell cycle reentry after MI, which decreased fibrotic scar size and improved cardiac function. RNA sequencing analysis suggested that Adssl1 overexpression induced strong dedifferentiation and cell cycle entry. Moreover, LC-MS/MS analysis showed that Adssl1 overexpression was associated with increased amounts of purine metabolites, including inosine, which is in clinical use. Administration of exogenous inosine promoted cardiac repair after MI in adult mice. At a molecular level, the increase in purine metabolite production mediated by Adssl1 enhanced the activity of the proliferation-promoting mTORC1 pathway. Our study identifies a role for Adssl1 in supporting cardiomyocyte proliferation and cardiac regeneration.

Mutational analysis of Phanerochaete chrysosporium´s purine transporter.

We present here a mutational analysis of the purine transporter from Phanerochaete chrysosporium (PhZ), a member of the AzgA-like subfamily within the Nucleobase Ascorbate Transporters family. We identified key residues that determine its substrate specificity and transport efficiency. Thirteen PhZ mutants were generated and heterologously expressed in Aspergillus nidulans. The growth of mutant strains in the presence of purines and toxic analogues and the uptake rate of radiolabelled hypoxanthine were evaluated. Results revealed that ten mutants showed differences in transport compared to the wild-type PhZ: six mutants completely lost function, two exhibited decreased transport activity, and two showed increased hypoxanthine uptake. Subcellular localization and expression level analyses indicated that the differences in transport activity were not due to trafficking issues to the plasma membrane or protein stability. A three-dimensional model of PhZ, constructed with the artificial intelligence-based AlphaFold2 program, suggested that critical residues for transport are located in transmembrane segments and an internal helix. In the latter, the A418 residue was identified as playing a pivotal role in transport efficiency despite being far from the putative substrate binding site, as mutant A418V showed an increased initial uptake efficiency for the transporter´s physiological substrates. We also report that residue L124, which lies in the putative substrate binding site, plays a critical role in substrate transport, emerging as an additional determinant in the transport mechanism of this family of transporters. These findings underscore the importance of specific residues in AzgA-like transporters and enhance our understanding of the intricate mechanisms governing substrate specificity and transport efficiency within this family.

Nonclinical evaluations of deucravacitinib and Janus kinase inhibitors in homeostatic and inflammatory pathways.

Translational medicine provides insight into novel drugs and predicts unwanted effects. In well-characterized pathways (e.g., cytokine-Janus kinase [JAK]-signal transducers and activators of transcription [STAT]), a variety of in vitro assessments were used to estimate selectivity of effects on different potential targets (i.e., JAK1, JAK2, JAK3, and tyrosine kinase 2 [TYK2]). Several approved drugs were characterized as selective for the JAK family. These assessments are challenged by a lack of compounds that only inhibit one JAK family member. Deucravacitinib is a first-in-class, oral, selective, allosteric inhibitor of TYK2, a kinase required for IL-12, IL-23, and Type I interferon signaling. Unlike deucravacitinib, which selectively binds to the TYK2 regulatory domain, JAK1,2,3 inhibitors target the catalytic domain, contributing to nonselective targeting of JAK1,2,3. Cytokines associated with JAK1,2,3 signaling are required for both immune and nonimmune functions. A similar laboratory abnormality profile was observed in clinical trials using JAK1,2,3 inhibitors that has not been observed with deucravacitinib. In vitro testing of JAK1,2,3 inhibitors has relied upon assays of signal transduction, such as those measuring STAT phosphorylation, for estimates of potency and selectivity. These assay systems can be effective in estimating in vivo efficacy; however, they may not provide insight into downstream outcomes of receptor signaling, which may be more relevant for evaluating safety aspects. Assay systems assessing functional outcomes from cells may yield a more useful translational evaluation. Here, deucravacitinib was assessed for potency and selectivity versus three representatives of the JAK inhibitor class (tofacitinib, baricitinib, and upadacitinib) based on functional assays. JAK inhibitors had suppressive activity against JAK2-dependent hematopoietic colony-forming assays modeling thrombopoiesis, erythropoiesis, and myelopoiesis; however, deucravacitinib did not. Deucravacitinib had limited potency against NK cells, cytotoxic T cells, T-helper cells, and regulatory T cells activated by JAK1/JAK3-dependent common gamma chain cytokines. These data are consistent with the biologic role of JAK1,2,3 and pharmacodynamic changes in clinical laboratory abnormalities. Against TYK2-dependent cytokines, deucravacitinib selectively inhibited Type I interferon stimulation of monocytes and dendritic cells and was a more potent inhibitor than JAK inhibitors. IL-12 and IL-23 functional outputs were similarly potently inhibited by deucravacitinib. Results are consistent with deucravacitinib selectively inhibiting TYK2.

Quantification of Letrozole, Palbociclib, Ribociclib, Abemaciclib, and Metabolites in Volumetric Dried Blood Spots: Development and Validation of an LC-MS/MS Method for Therapeutic Drug Monitoring.

Therapeutic drug monitoring (TDM) may be beneficial for cyclin-dependent kinase 4/6 inhibitors (CDK4/6is), such as palbociclib, ribociclib, and abemaciclib, due to established exposure-toxicity relationships and the potential for monitoring treatment adherence. Developing a method for quantifying CDK4/6is, abemaciclib metabolites (M2, M20), and letrozole in dried blood spots (DBS) could be useful to enhance the feasibility of TDM. Thus, an optimized LC-MS/MS method was developed using the HemaXis DB10 device for volumetric (10 µL) DBS collection. Chromatographic separation was achieved using a reversed-phase XBridge BEH C18 column. Detection was performed with a triple quadrupole mass spectrometer, utilizing ESI source switching between negative and positive ionization modes and multiple reaction monitoring acquisition. Analytical validation followed FDA, EMA, and IATDMCT guidelines, demonstrating high selectivity, adequate sensitivity (LLOQ S/N ≥ 30), and linearity (r ≥ 0.997). Accuracy and precision met acceptance criteria (between-run: accuracy 95-106%, CV ≤ 10.6%). Haematocrit independence was confirmed (22-55%),with high recovery rates (81-93%) and minimal matrix effects (ME 0.9-1.1%). The stability of analytes under home-sampling conditions was also verified. Clinical validation supports DBS-based TDM as feasible, with conversion models developed for estimating plasma concentrations (the reference for TDM target values) of letrozole, abemaciclib, and its metabolites. Preliminary data for palbociclib and ribociclib are also presented.

Prescription trends in Japanese advanced Parkinson's disease patients with non-motor symptoms: J-FIRST.

BACKGROUND: Non-motor symptoms (NMS) are important factors when selecting treatments for patients with advanced Parkinson's disease (PD). We sought to elucidate the prescribing practices for advanced PD patients with NMS in Japanese clinical practice. METHODS: We examined the prescription rates and doses of anti-PD drugs, and the use of non-steroidal anti-inflammatory drugs (NSAIDs) in post hoc analyses of a 52-week observational study of 996 PD patients with wearing-off on levodopa-containing therapy and ≥1 NMS. RESULTS: Dopamine agonists were the most frequently prescribed drugs combined with levodopa-containing drugs, followed by entacapone, zonisamide, istradefylline, selegiline, and amantadine. The daily dose of levodopa-containing drugs, rotigotine, entacapone, istradefylline, and droxidopa, and the levodopa-equivalent dose increased during the observation period. In a subgroup analysis of patients stratified by NMS status (improved/unchanged/deteriorated), the deteriorated group had higher prescription rates of entacapone and istradefylline, whereas the improved group had higher prescription rates of NSAIDs and zonisamide at Week 52. Prescriptions varied by geographical region for anti-PD drugs and by NMS status for NSAIDs. CONCLUSIONS: There were significant changes in the prescriptions and dosing of selected anti-PD drugs, especially newer drugs. Anti-PD drug and NSAID prescriptions also varied by changes in NMS status and geographic region.

Enhancing Cellular Homeostasis: Targeted Botanical Compounds Boost Cellular Health Functions in Normal and Premature Aging Fibroblasts.

The human skin, the body's largest organ, undergoes continuous renewal but is significantly impacted by aging, which impairs its function and leads to visible changes. This study aimed to identify botanical compounds that mimic the anti-aging effects of baricitinib, a known JAK1/2 inhibitor. Through in silico screening of a botanical compound library, 14 potential candidates were identified, and 7 were further analyzed for their effects on cellular aging. The compounds were tested on both normal aged fibroblasts and premature aging fibroblasts derived from patients with Hutchinson-Gilford Progeria Syndrome (HGPS). Results showed that these botanical compounds effectively inhibited the JAK/STAT pathway, reduced the levels of phosphorylated STAT1 and STAT3, and ameliorated phenotypic changes associated with cellular aging. Treatments improved cell proliferation, reduced senescence markers, and enhanced autophagy without inducing cytotoxicity. Compounds, such as Resveratrol, Bisdemethoxycurcumin, Pinosylvin, Methyl P-Hydroxycinnamate, cis-Pterostilbene, and (+)-Gallocatechin, demonstrated significant improvements in both control and HGPS fibroblasts. These findings suggest that these botanical compounds have the potential to mitigate age-related cellular alterations, offering promising strategies for anti-aging therapies, particularly for skin health. Further in vivo studies are warranted to validate these results and explore their therapeutic applications.

One-carbon-mediated purine synthesis underlies temozolomide resistance in glioblastoma.

Glioblastoma accounts for nearly half of all primary malignant brain tumors in adults, and despite an aggressive standard of care, including excisional surgery and adjuvant chemoradiation, recurrence remains universal, with an overall median survival of 14.6 months. Recent work has revealed the importance of passenger mutations as critical mediators of metabolic adaptation in cancer progression. In our previous work, we identified a role for the epigenetic modifier ID-1 in temozolomide resistance in glioblastoma. Here, we show that ID-1-mediated glioblastoma tumourigenesis is accompanied by upregulation of one-carbon (1-C) mediated de novo purine synthesis. ID-1 knockout results in a significant reduction in the expression of 1-C metabolism and purine synthesis enzymes. Analysis of glioblastoma surgical specimens at initial presentation and recurrence reveals that 1-C purine synthesis metabolic enzymes are enriched in recurrent glioblastoma and that their expression correlates with a shorter time to tumor recurrence. Further, we show that the 1-C metabolic phenotype underlies proliferative capacity and temozolomide resistance in glioblastoma cells. Supplementation with exogenous purines restores proliferation in ID-1-deficient cells, while inhibition of purine synthesis with AICAR sensitizes temozolomide-resistant glioblastoma cells to temozolomide chemotherapy. Our data suggest that the metabolic phenotype observed in treatment-resistant glioma cells is a potential therapeutic target in glioblastoma.

The Avoidance of Purine Stretches by Cancer Mutations.

Purine stretches, sequences of adenine (A) and guanine (G) in DNA, play critical roles in binding regulatory protein factors and influence gene expression by affecting DNA folding. This study investigates the relationship between purine stretches and cancer development, considering the aromaticity of purines, quantified by methods like Hückel's rule and NICS calculations, and the importance of the flanking sequence context. A pronounced avoidance of long purine stretches by typical cancer mutations was observed in public data on the intergenic regions of cancer patients, suggesting a role of intergenic sequences in chromatin reorganization and gene regulation. A statistically significant shortening of purine stretches in cancerous tumors (p value < 0.0001) was found. The insights into the aromatic nature of purines and their stacking energies explain the role of purine stretches in DNA structure, contributing to their role in cancer progression. This research lays the groundwork for understanding the nature of purine stretches, emphasizing their importance in gene regulation and chromatin restructuring, and offers potential avenues for novel cancer therapies and insights into cancer etiology.

Rheumatoid Arthritis and COVID-19 at the Intersection of Immunology and Infectious Diseases: A Related PRISMA Systematic Literature Review.

Rheumatoid arthritis (RA) patients face different health challenges when infected with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) than the general population, due to both their immunocompromised state and the immunosuppressive therapies they receive. This systematic literature review, which follows the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) paradigm, explores the interactions between RA and SARS-CoV-2 infection, focusing on immunologic issues, disease management, vaccination, and adverse outcomes. In order to obtain the most relevant information, we systematically reviewed the specific literature from 1 January 2021 to 31 December 2023, based on the PRISMA method, by which we eventually selected 35 eligible articles, to which we added other ISI-indexed studies to enrich our results further. Consequently, we performed a funnel analysis to evaluate the potential for publication bias. Firstly, the data collected revealed the impact of the pandemic on RA diagnoses and the fear of face-to-face medical consultations that delayed adequate treatment. Secondly, cardiovascular and metabolic comorbidities increase the risk of prolonged COVID-19 symptoms, hospitalization, and severe COVID-19 outcomes for RA patients. With respect to immunosuppressive treatment used to control RA, it was observed that glucocorticoids (especially high-dose usage) and Rituximab (RTX) predispose the patients to poor SARS-CoV-2 outcomes, as opposed to Baricitinib and interleukin-6 (IL-6) and tumor necrosis factor-alpha (TNF-α) inhibitors. COVID-19 vaccination has proven effective and generally safe for RA patients in some studies, although therapies with Methotrexate (MTX), Abatacept (ABA), and RTX have been associated with impaired vaccine immune response. This systematic literature review brings updated and thorough information with respect to the immunological, clinical, and management of a complex immune-mediated inflammatory disease (IMID) like RA in the setting of COVID-19 and underlines the challenges faced by this group of patients. The lessons learned can be extended beyond the pandemic in shaping a more informed and compassionate healthcare system and offering long-term medical care for patients with RA.

Combinatorial Anti-Cancer Effect of Polypurine Reverse Hoogsteen Hairpins against KRAS and MYC Targeting in Prostate and Pancreatic Cancer Cell Lines.

Introduction: KRAS and MYC are proto-oncogenes that are strictly regulated in healthy cells that have key roles in several processes such as cell growth, proliferation, differentiation, or apoptosis. These genes are tightly interconnected, and their dysregulation can lead to cancer progression. We previously individually targeted these oncogenes using Polypurine Reverse Hoogsteen (PPRH) hairpins, mostly targeting the complementary strand of G-quadruplex-forming sequences. We validated them in vitro in different cancer cell lines with deregulated KRAS and/or MYC. In this work we focused on our understanding of the cooperative dynamics between these oncogenes, by investigating the combined impact of PPRHs targeting KRAS and MYC in pancreatic and prostate cancer cells. Results: The combinations had a modulatory impact on the expression of both oncogenes, with transcriptional and translational downregulation occurring five days post-treatment. Out of the four tested PPRHs, MYC-targeting PPRHs, especially HpMYC-G4-PR-C directed against the promoter, showed a greater cytotoxic and expression modulation effect. When both KRAS- and MYC-targeting PPRHs were applied in combination, a synergistic reduction in cell viability was observed. Conclusion: The simultaneous targeting of KRAS and MYC demonstrates efficacy in gene modulation, thus in decreasing cell proliferation and viability.

AB036. Targeting glioblastoma de-novo purine metabolism to overcome chemoradiation resistance: an interim result of phase 0/1 clinical trial in newly diagnosed and recurrent glioblastoma.

BACKGROUND: Glioblastoma cells preferentially use de-novo purine synthesis pathway, whereas normal brain prefers salvage pathway. Mycophenolate mofetil (MMF), a commonly used oral immunosuppressant that inhibits inosine-5'-monophosphate dehydrogenase (IMPDH), a key enzyme in the de-novo purine pathway. Pre-clinical suggested MMF can improve radiation and temozolomide efficacy in glioblastoma which led to this phase 0/1 trial (NCT04477200) to assess MMF's tolerability with chemoradiation in glioblastoma, mycophenolic acid accumulation, and purine synthesis inhibition in tumor. METHODS: In the phase 0 study, eight recurrent glioblastoma patients received MMF at doses ranging 500-2,000 mg BID for 1-week before surgery. The tissues were analyzed using mass spectrometry for drug accumulation and purine synthesis inhibition. In the phase 1 study, adult patients were given MMF starting at 1,000 mg orally (PO) twice daily (BID), with the possible dose ranging 500-2,000 PO BID. Nineteen recurrent glioblastoma patients (target N=30) received MMF 1-week prior to and concurrently with re-irradiation (40.5 Gy). Thirty newly diagnosed glioblastoma patients received MMF 1-week prior to and concurrently with chemoradiation, followed by MMF 1-day before and during 5 days of each adjuvant temozolomide cycle. RESULTS: Both enhancing and non-enhancing tumors from phase 0 subjects yielded >1 µM active drug metabolite, and the guanosine triphosphate: inosine monophosphate ratio was decreased by 75% in enhancing tumors in MMF-treated patients compared to untreated controls (P=0.009), indicating effective target engagement and inhibition of purine synthesis. In the phase 1 study, no dose-limiting toxicities (DLTs) were observed at the interim analysis at MMF 1,000-1,500 mg BID combined with chemoradiation. At 2,000 mg BID, there was no DLT combined with temozolomide alone, however, there were four DLTs noted (hemiparesis, cognitive disturbance, fatigue, thrombocytopenia) when combined with radiotherapy and temozolomide together, though all were reversible. Interim median overall survival in recurrent phase 1 is 15.6 months, and not reached yet in newly diagnosed phase 1. CONCLUSIONS: MMF with chemoradiation has been reasonably well tolerated and showed promising evidence of brain tumor target engagement and drug accumulation. This study led to a recommended phase 2 dose of MMF 1,500 mg BID and will provide a preliminary efficacy estimate for a randomized phase 2/3 trial through the Alliance for Clinical Trials in Oncology.

In situ chemoproteomic profiling reveals itaconate inhibits de novo purine biosynthesis in pathogens.

Itaconate serves as an immune-specific metabolite that regulates gene transcription and metabolism in both host and pathogens. S-itaconation is a post-translational modification that regulates immune response; however, its antimicrobial mechanism under the physiological condition remains unclear. Here, we apply a bioorthogonal itaconate probe to perform global profiling of S-itaconation in living pathogens, including S. Typhimurium, S. aureus, and P. aeruginosa. Some functional enzymes are covalently modified by itaconate, including those involved in the de novo purine biosynthesis pathway. Further biochemical studies demonstrate that itaconate suppresses this specific pathway to limit Salmonella growth by inhibiting the initiator purF to lower de novo purine biosynthesis and simultaneously targeting the guaABC cluster to block the salvage route. Our chemoproteomic study provides a global portrait of S-itaconation in multiple pathogens and offers a valuable resource for finding susceptible targets to combat drug-resistant pathogens in the future.