Search Results (30)

Background/Objective: Oncogenic tyrosine kinases (TKs) such as ALK and SRC promote cancer progression, but their effects on metabolic enzymes are still not well understood. This study examines how TK signaling regulates inosine monophosphate dehydrogenase 2 (IMPDH2), a rate-limiting enzyme in purine biosynthesis, and assesses its potential as a therapeutic target. Methods: Phosphoproteomic screening and in vitro kinase assays were used to identify phosphorylation sites on IMPDH2. Lipid-binding assays explored the role of phosphatidylinositol 3-phosphate (PI3P) in IMPDH2 regulation. Structure-based virtual screening discovered small-molecule allosteric inhibitors, which were tested in lymphoma cell models, including ALK and BTK-inhibitor resistant lines. Results: Here, we identify Inosine monophosphate dehydrogenase-2 (IMPDH2), a rate-limiting enzyme in purine biosynthesis, as a novel substrate of ALK and SRC. We show that phosphorylation at the conserved Y233 residue within the allosteric domain enhances IMPDH2 activity, linking TK signaling to metabolic reprogramming in cancer cells. We further identify PI3P as a natural lipid inhibitor that binds IMPDH2 and suppresses its enzymatic function. Using structure-based virtual screening, we developed Comp-10, a first-in-class allosteric IMPDH inhibitor. Unlike classical active-site inhibitors such as mycophenolic acid (MPA), Comp-10 decreases IMPDH1/2 protein levels, blocks filament (rod/ring) formation, and inhibits the growth of ALK and BTK inhibitor-resistant lymphoma cells. Comp-10 acts post-transcriptionally and avoids compensatory IMPDH upregulation observed with MPA (rod/ring) formation, and inhibited growth in TKI-resistant lymphoma cells. Notably, Comp-10 avoided the compensatory IMPDH upregulation observed with MPA. Conclusion: These findings uncover a novel TK–IMPDH2 signaling axis and provide mechanistic and therapeutic insight into the allosteric regulation of IMPDH2. Comp-10 represents a promising therapeutic candidate for targeting metabolic vulnerabilities in tyrosine kinase driven cancers. Full article

Uveal melanoma is a melanocyte-derived malignancy of the eye with a high propensity for liver metastasis. Metastatic uveal melanoma is associated with high mortality and is poorly responsive to currently available therapies. Most uveal melanoma cases are driven by activating mutations in GNAQ and GNA11 genes, which convey oncogenic signaling through the mitogen-activated protein kinase (MAPK) pathway. Despite promising early results, safe doses of pharmacological inhibitors of the MAPK cascade failed to effectively control uveal melanoma in human trials. Considering the role of the RAC/PAK signaling axis as a co-regulator of the MAPK cascade, we set forth to investigate whether the efficacy of MAPK cascade inhibitors in pre-clinical models may be enhanced by direct inhibition of RAC and PAK proteins, or by indirect control of RAC via inhibition of guanylate biosynthesis. We observed that pharmacological inhibition of RAC, PAK and the key guanylate biosynthesis enzyme IMPDH significantly synergized with various inhibitors of the MAPK cascade in suppressing oncogenic signaling and the growth of uveal melanoma cells. In a mouse model, the addition of an IMPDH inhibitor to the treatment regimen significantly enhanced the ability of a MAPK cascade inhibitor to improve the survival of tumor-bearing animals. Targeting of the RAC/PAK axis provides a new strategy to increase the efficacy of targeted therapies in uveal melanoma. While RAC and PAK inhibitors are still undergoing pre-clinical development, clinically available inhibitors of IMPDH offer an opportunity to test the efficacy of this novel synergistic combination in the context of human disease. Full article

Derivatives of mycophenolic acid (MPA) and 5-aminolevulinic acid photodynamic therapy (ALA-PDT) have been used separately to treat psoriasis, a chronic, inflammatory skin disease that is characterized by the unregulated hyperproliferation of epidermal keratinocytes and a T-cell-mediated immune response. However, the combination of these two therapies has not previously been explored. This study investigated the in vitro effects of combining MPA with ALA-PDT to suppress keratinocytes and the in vitro inhibition of inosine monophosphate dehydrogenase, a key enzyme. The effects of ALA, MPA, and their combination on protoporphyrin IX (PpIX) generation and cell viability in HaCaT cells, as well as the inhibition of IMPDH, were evaluated. Treatment of HaCaT cells with ALA, MPA, and their 1:1 molar combination showed that ALA alone induced PpIX production, with concentrations increasing from 5.25 ng/mL at 10 μM to 157.5 ng/mL at 1 mM. MPA did not increase PpIX on its own but had a modest synergistic effect with ALA at low concentrations (10 μM and 50 μM). The impact of blue light irradiation (465 nm) on cell viability was also assessed, revealing that ALA and ALA + MPA treatment led to significant reductions in HaCaT cell viability at higher concentrations (500 μM–1 mM), while MPA alone with blue light irradiation showed no cytotoxicity. The reduction in skin cell viability was enhanced when ALA was combined with MPA. Additionally, MPA effectively inhibited IMPDH activity in a dose-dependent manner, with 94–96% inhibition at concentrations of 100 μM and above. Interestingly, ALA weakly inhibited IMPDH, with a peak inhibition of 46% at 5 μM. At higher ALA concentrations, its inhibitory effect diminished, and it interfered with the potency of MPA’s IMPDH2 inhibition, suggesting that ALA could modulate MPA’s therapeutic action. These findings suggest that the combination of MPA with ALA-PDT may be a viable new treatment for psoriasis. Full article

Tea production and quality are largely determined by the many genetic and biochemical characteristics that occur in tea plant cultivars. Worldwide, tea is consumed for its pleasing and refreshing effects due to its caffeine content. The present study performed transcriptomics analyses of two tea species (Camellia sinensis var. Shuchazao (SCZ) and Camellia ptilophylla (CAF)) and identified diversity in the gene expression levels and major regulatory transcription factors (TFs) for the characterization of purine alkaloids and phenylpropanoid biosynthesis pathways. The RNA-seq analysis of two species (SCZ and CAF) revealed the differences in caffeine and catechins synthesis. In the purine alkaloid biosynthesis pathway, the S-adenosyl methionine (SAM) and adenosine monophosphate (AMP) pathway genes were significantly related to xanthosine synthesis in contrasting purine alkaloids among (Camellia sinensis var. Shuchazao (SCZ) and Camellia ptilophylla (CAF)). The significant expression of SAMS-5, PPAT-2, IMPDH-2, TCS-2, TCS-3, XMT-1, XMT-13, and XDH-4 in the xanthosine degradation pathway in CAF is attributed to higher theobromine content as compared to SCZ. Moreover, the transcription factors (TFs) AP2/ERF (20%), WRKY (12%), NAC (11%), and MYB (8%) were significantly correlated. The upregulated expression of caffeine synthesis genes in SCZ was correlated with MYB and AP2/ERF transcription factors. This study provides the basis for differences in the genetic mechanism in purine alkaloids, phenylpropanoid, and flavonoid biosynthesis pathways, which would be helpful in the development and selection of tea plant species with high or low caffeine concentrations. This study also provides a road map for future genetic improvement in tea species and cultivars. Full article

Borrelia burgdorferi’s inosine-5′-monophosphate dehydrogenase (IMPDH, GuaB encoded by the guaB gene) is a potential therapeutic target. GuaB is necessary for B. burgdorferi replication in mammalian hosts but not in standard laboratory culture conditions. Therefore, we cannot test novel GuaB inhibitors against B. burgdorferi without utilizing mammalian infection models. This study aimed to evaluate modifications to a standard growth medium that may mimic mammalian conditions and induce the requirement of GuaB usage for replication. The effects of two GuaB inhibitors (mycophenolic acid, 6-chloropurine riboside at 125 μM and 250 μM) were assessed against B. burgdorferi (guaB+) grown in standard Barbour–Stoenner–Kelly-II (BSK-II) medium (6% rabbit serum) and BSK-II modified to 60% concentration rabbit serum (BSK-II/60% serum). BSK-II directly supplemented with adenine, hypoxanthine, and nicotinamide (75 μM each, BSK-II/AHN) was also considered as a comparison group. In standard BSK-II, neither mycophenolic acid nor 6-chloropurine riboside affected B. burgdorferi growth. Based on an ANOVA, a dose-dependent increase in drug effects was observed in the modified growth conditions (F = 4.471, p = 0.001). Considering higher drug concentrations at exponential growth, mycophenolic acid at 250 μM reduced spirochete replication by 48% in BSK-II/60% serum and by 50% in BSK-II/AHN (p < 0.001 each). 6-chloropurine riboside was more effective in both mediums than mycophenolic acid, reducing replication by 64% in BSK-II/60% serum and 65% in BSK-II/AHN (p < 0.001 each). These results demonstrate that modifying BSK-II medium with physiologically relevant levels of mammalian serum supports replication and induces the effects of GuaB inhibitors. This represents the first use of GuaB inhibitors against Borrelia burgdorferi, building on tests against purified B. burgdorferi GuaB. The strong effects of 6-chloropurine riboside indicate that B. burgdorferi can salvage and phosphorylate these purine derivative analogs. Therefore, this type of molecule may be considered for future drug development. Optimization of this culture system will allow for better assessment of novel Borrelia-specific GuaB inhibitor molecules for Lyme disease interventions. The use of GuaB inhibitors as broadcast sprays or feed baits should also be evaluated to reduce spirochete load in competent reservoir hosts. Full article

Chemical proteomics using biotin probes of natural products have significantly advanced our understanding of molecular targets and therapeutic potential. This review highlights recent progress in the application of biotin probes of homoisoflavonoids for identifying binding proteins and elucidating mechanisms of action. Notably, homoisoflavonoids exhibit antiangiogenic, anti-inflammatory, and antidiabetic effects. A combination of biotin probes, pull-down assays, mass spectrometry, and molecular modeling has revealed how natural products and their derivatives interact with several proteins such as ferrochelatase (FECH), soluble epoxide hydrolase (sEH), inosine monophosphate dehydrogenase 2 (IMPDH2), phosphodiesterase 4 (PDE4), and deoxyhypusine hydroxylase (DOHH). These target identification approaches pave the way for new therapeutic avenues, especially in the fields of oncology and ophthalmology. Future research aimed at expanding the repertoire of target identification using biotin probes of homoisoflavonoids promises to further elucidate the complex mechanisms and develop new drug candidates. Full article

IMP dehydrogenase (IMPDH) inhibition has emerged as a new target therapy for glioblastoma multiforme (GBM), which remains one of the most refractory tumors to date. TCGA analyses revealed distinct expression profiles of IMPDH isoenzymes in various subtypes of GBM and low-grade glioma (LGG). To dissect the mechanism(s) underlying the anti-tumor effect of IMPDH inhibition in adult GBM, we investigated how mycophenolic acid (MPA, an IMPDH inhibitor) treatment affected key oncogenic drivers in glioblastoma cells. Our results showed that MPA decreased the expression of telomerase reverse transcriptase (TERT) in both U87 and U251 cells, and the expression of O⁶-methylguanine-DNA methyltransferase (MGMT) in U251 cells. In support, MPA treatment reduced the amount of telomere repeats in U87 and U251 cells. TERT downregulation by MPA was associated with a significant decrease in c-Myc (a TERT transcription activator) in U87 but not U251 cells, and a dose-dependent increase in p53 and CCCTC-binding factor (CTCF) (TERT repressors) in both U87 and U251 cells. In U251 cells, MPA displayed strong cytotoxic synergy with BCNU and moderate synergy with irinotecan, oxaliplatin, paclitaxel, or temozolomide (TMZ). In U87 cells, MPA displayed strong cytotoxic synergy with all except TMZ, acting primarily through the apoptotic pathway. Our work expands the mechanistic potential of IMPDH inhibition to TERT/telomere regulation and reveals a synthetic lethality between MPA and anti-GBM drugs. Full article

The goal of this study was to examine commonalities in the molecular basis of learning in mice and humans. In previous work we have demonstrated that the anterior cingulate cortex (ACC) and hippocampus (HC) are involved in learning a two-choice visuospatial discrimination task. Here, we began by looking for candidate genes upregulated in mouse ACC and HC with learning. We then determined which of these were also upregulated in mouse blood. Finally, we used RT-PCR to compare candidate gene expression in mouse blood with that from humans following one of two forms of learning: a working memory task (network training) or meditation (a generalized training shown to change many networks). Two genes were upregulated in mice following learning: caspase recruitment domain-containing protein 6 (Card6) and inosine monophosphate dehydrogenase 2 (Impdh2). The Impdh2 gene product catalyzes the first committed step of guanine nucleotide synthesis and is tightly linked to cell proliferation. The Card6 gene product positively modulates signal transduction. In humans, Card6 was significantly upregulated, and Impdh2 trended toward upregulation with training. These genes have been shown to regulate pathways that influence nuclear factor kappa B (NF-κB), a factor previously found to be related to enhanced synaptic function and learning. Full article

Pathogenic variants in IMPDH1 are associated with autosomal dominant retinitis pigmentosa 10 (RP10), and Leber congenital amaurosis 11. This case report of a 13-year-old girl with Down’s syndrome and keratoglobus is aimed at linking the novel variant IMPDH1 c.134A>G, p.(Tyr45Cys), a variant of uncertain significance, to a clinical phenotype and to provide grounds for the objective assignment of its benign features. RP10 is characterized by the early onset and rapid progression of ocular symptoms, beginning with nyctalopia in childhood, accompanied by typical RP fundus changes. As evidenced via thorough clinical examination and testing, none of the RP10 characteristics were present in our patient. On the contrary, our patient who was heterozygous for IMPDH1 c.134A>G, p.(Tyr45Cys) showed no signs of peripheral retinal dystrophy, and did not manifest any disease characteristics typical of the IMPDH1 gene mutation. Consequently, we conclude that the variant did not contribute to the phenotype. According to standards and guidelines for the interpretation of sequence variants, IMPDH1 c.134A>G, p.(Tyr45Cys) revealed likely benign features. Full article

Lung cancer is one of the most common cancers worldwide, and despite improvements in treatment regimens, patient prognosis remains poor. Lung adenocarcinomas develop from the lung epithelia and understanding how specific genetic and environmental factors lead to oncogenic transformation in these cells is of great importance to define the pathways that contribute to tumorigenesis. The recent rise in the use of immunotherapy to treat different cancers has prompted the exploration of immune modulators in tumour cells that may provide new targets to manipulate this process. Of these, the B7 family of cell surface receptors, which includes PD-1, is of particular interest due to its role in modulating immune cell responses within the tumour microenvironment. B7-H3 (CD276) is one family member that is upregulated in many cancer types and suggested to contribute to tumour–immune interactions. However, the function and ligand(s) for this receptor in normal lung epithelia and the mechanisms through which the overexpression of B7-H3 regulate cancer progression in the absence of immune cell interactions remain unclear. Here, we present evidence that B7-H3 is associated with one of the key rate-limiting metabolic enzymes IMPDH2, and the localisation of this complex is altered in human lung cancer cells that express high levels of B7-H3. Mechanistically, the IMPDH2:B7-H3 complex provides a protective role in cancer cells to escape oxidative stress triggered by chemotherapy, thus leading to cell survival. We further demonstrate that the loss of B7-H3 in cancer cells has no effect on growth or migration in 2D but promotes the expansion of 3D spheroids in an IMPDH2-dependent manner. These findings provide new insights into the B7-H3 function in the metabolic homeostasis of normal and transformed lung cancer cells, and whilst this molecule remains an interesting target for immunotherapy, these findings caution against the use of anti-B7-H3 therapies in certain clinical settings. Full article

Mycophenolate mofetil (MMF) is part of the standard immunosuppressive treatment after transplantation and usually given as “one-dose-fits-all” together with a calcineurin inhibitor (CNI). Although drug concentrations are frequently monitored, there is still a group of patients who experience side effects related to excessive or insufficient immune suppression. We therefore aimed to identify biomarkers that reflect the overall immune status of the patient and might support individualized dosing. We previously studied immune biomarkers for CNIs and aimed to investigate whether these are also suitable to monitor MMF activity. Healthy volunteers received a single dose of MMF or placebo, after which IMPDH enzymatic activity, T cell proliferation, and cytokine production were measured and compared to MPA (MMF’s active metabolite) concentration in three different matrices (plasma, peripheral blood mononuclear cells, and T cells). MPA concentrations in T cells exceeded those in PBMCs, but all intracellular concentrations correlated strongly with plasma concentrations. At clinically relevant MPA concentrations, IL-2 and IFN-γ production was mildly suppressed, while MPA T cell proliferation was strongly inhibited. Based on these data, it is expected that monitoring of T cell proliferation in MMF-treated transplantation patients may be a valid strategy to avoid excessive immune suppression. Full article

The metabolic profile of the Aspergillus sp. 1901NT-1.2.2 sponge-associated fungal strain was investigated using the HPLC MS technique, and more than 23 peaks in the HPLC MS chromatogram were detected. Only two minor peaks were identified as endocrocin and terpene derivative MS data from the GNPS database. The main compound was isolated and identified as known anthraquinone derivative vismione E. The absolute stereochemistry of vismione E was established for the first time using ECD and quantum chemical methods. Vismione E showed high cytotoxic activity against human breast cancer MCF-7 cells, with an IC₅₀ of 9.0 µM, in comparison with low toxicity for normal human breast MCF-10A cells, with an IC₅₀ of 65.3 µM. It was found that vismione E inhibits MCF-7 cell proliferation and arrests the cell cycle in the G1 phase. Moreover, the negative influence of vismione E on MCF-7 cell migration was detected. Molecular docking of vismione E suggested the IMPDH2 enzyme as one of the molecular targets for this anthraquinone derivative. Full article

There is considerable interest in urine as a non-invasive liquid biopsy to detect prostate cancer (PCa). PCa-specific transcripts such as the TMPRSS2:ERG fusion gene can be found in both urine extracellular vesicles (EVs) and urine cell-sediment (Cell) but the relative usefulness of these and other genes in each fraction in PCa detection has not been fully elucidated. Urine samples from 76 men (PCa n = 40, non-cancer n = 36) were analysed by NanoString for 154 PCa-associated genes-probes, 11 tissue-specific, and six housekeeping. Comparison to qRT-PCR data for four genes (PCA3, OR51E2, FOLH1, and RPLP2) was strong (r = 0.51–0.95, Spearman p < 0.00001). Comparing EV to Cells, differential gene expression analysis found 57 gene-probes significantly more highly expressed in 100 ng of amplified cDNA products from the EV fraction, and 26 in Cells (p < 0.05; edgeR). Expression levels of prostate-specific genes (KLK2, KLK3) measured were ~20× higher in EVs, while PTPRC (white-blood Cells) was ~1000× higher in Cells. Boruta analysis identified 11 gene-probes as useful in detecting PCa: two were useful in both fractions (PCA3, HOXC6), five in EVs alone (GJB1, RPS10, TMPRSS2:ERG, ERG_Exons_4-5, HPN) and four from Cell (ERG_Exons_6-7, OR51E2, SPINK1, IMPDH2), suggesting that it is beneficial to fractionate whole urine prior to analysis. The five housekeeping genes were not significantly differentially expressed between PCa and non-cancer samples. Expression signatures from Cell, EV and combined data did not show evidence for one fraction providing superior information over the other. Full article

Autoimmunity and cancer rates have both been on the rise in Western civilization prompting many to investigate the link between the two entities. This review will investigate the complex interactions between the activation and deactivation of the immune system and the development of malignancy. Additional focus will be placed on the main classes of immune inhibitor therapy utilized in transplant patients and in autoimmune disease including TNF-alpha, Calcineurin, mTOR, purine synthesis antagonists and IMPDH inhibitors. Full article

Mycophenolic acid (MPA) is a widely used immunosuppressive agent and exerts its effect by inhibiting inosine 5′-monophosphate dehydrogenase (IMPDH), the main regulating enzyme of purine metabolism. However, significant unexplained differences in the efficacy and tolerability of MPA therapy pose a clinical challenge. Therefore, broad pharmacogenetic, pharmacokinetic, and pharmacodynamic approaches are needed to individualize MPA therapy. In this prospective cohort study including 277 renal transplant recipients, IMPDH2 rs11706052 SNP status was assessed by genetic sequencing, and plasma MPA trough levels were determined by HPLC and IMPDH enzyme activity in peripheral blood mononuclear cells (PBMCs) by liquid chromatography–mass spectrometry. Among the 277 patients, 84 were identified with episodes of biopsy-proven rejection (BPR). No association was found between rs11706052 SNP status and graft rejection (OR 1.808, and 95% CI, 0.939 to 3.479; p = 0.076). Furthermore, there was no association between MPA plasma levels and BPR (p = 0.69). However, the patients with graft rejection had a significantly higher predose IMPDH activity in PBMCs compared to the controls without rejection at the time of biopsy (110.1 ± 50.2 vs. 95.2 ± 45.4 pmol/h; p = 0.001), and relative to the baseline IMPDH activity before transplantation (p = 0.042). Our results suggest that individualization of MPA therapy, particularly through pharmacodynamic monitoring of IMPDH activity in PBMCs, has the potential to improve the clinical outcomes of transplant patients. Full article