Biochemical pharmacology最新文献

Breast cancer is the most common malignant tumor endangering women's life and health. Tamoxifen citrate (TAM) is the first-line drug of adjuvant endocrine therapy for estrogen receptor-positive (ER⁺) breast cancer patients. Some sporadic cases have described rare adverse reactions of TAM with potentially life-threatening dermatological manifestations, which were associated with skin allergy. Mas related G protein-coupled receptor X2 (MRGPRX2) on human mast cells is the key target for skin allergy. We aimed to investigate the mechanism of TAM-induced allergic reactions and their potential effects on TAM treatment for breast cancer. In our study, TAM can specifically bind with MRGPRX2, which was mainly driven by hydrophobic force. TAM formed hydrogen bonds with TRP243, TRP248, and GLU164 residues in MRGPRX2. TAM induced calcium mobilization and degranulation of mast cells via MRGPRX2. Besides, TAM induced passive cutaneous anaphylaxis and active systemic anaphylaxis in C57BL/6 mice. The release of β-hexosaminidase, histamine, tumor necrosis factor-α, monocyte chemoattractant protein 1, and interleukin-8 were increased by TAM in vitro and in vivo. Furthermore, we found that MCF-7 and T-47D breast cancer cells can recruit mast cells to adjacent cancerous tissues. Besides, mast cell activation induced by TAM via MRGPRX2 significantly promoted the proliferation and migration of MCF-7 and T-47D cells, which can be effectively reversed by mast cell membrane stabilizer clarithromycin and MRGPRX2 silencing. This study proposed an anti-allergic therapeutic strategy for breast cancer treatment with TAM, while also the potential of MRGPRX2 as an adjunctive target.

PAS domain-containing serine/threonine-protein kinase (PASK) is a nutrient and energy sensor regulated by fasting/refeeding conditions in hypothalamic areas involved in controlling energy balance. In this sense, PASK plays a role in coordinating the activation/inactivation of AMP-activated protein kinase (AMPK) and mechanistic target of rapamycin (mTOR) in response to fasting. PASK deficiency protects against the development of diet-induced obesity. This has prompted an investigation into the potential role of PASK on energy expenditure through thermogenesis in adipose tissue. Our results indicate that PASK-deficient male mice exhibited higher brown adipose tissue (BAT) thermogenic activity and heat production. The inhibition of PASK function induces the expression of Uncoupling Protein 1 (UCP1) and the adipogenic marker peroxisome proliferator-activated receptor gamma (PPARγ) in BAT. In addition, PASK deficiency promotes the expression of UCP1 and other browning markers such as PR/SET Domain 16 (PRDM16) in inguinal white adipose tissue (WAT). PASK-deficient mice record an enhanced thermogenic response, even under stimuli such as β-3adrenergic receptor agonist or cold. This evidence reveals PASK as a new mechanism modulating BAT thermogenesis.

Temozolomide is universally used to treat glioblastoma due to its unique ability to cross the blood-brain barrier and inhibit tumor growth through DNA alkylation. However, over time, the inevitable emergence of resistance to temozolomide impedes successful treatment of this cancer. As a result, there is an urgent need to identify new therapeutic targets to improve treatment outcomes for this malignancy. In this work, acquired temozolomide-resistant glioblastoma cell lines LN18 (LN18-TR) and T98G (T98G-TR) exhibited stronger aggressiveness and lower endoplasmic reticulum (ER) stress than their parental cells.. Besides, temozolomide resistance was associated with elevated proteasome activity that suppressed ER stress, which was restored upon inhibition of the proteasome with MG132. Specifically, our study revealed that the 19S proteasomal regulatory subunit PSMC2, which was overexpressed in adapted temozolomide-resistant glioblastoma cells, reduced pro-death autophagy and decreased temozolomide sensitivity in parental cells when overexpressed. While autophagy increased in parental cells following temozolomide treatment, it was not elevated in temozolomide-resistant glioblastoma cells. Genetic suppression of PSMC2 triggered the JNK signalling pathway causing phosphorylation of BCL2, allowing Beclin1 to be released from the BCL2-Beclin1 complex. This boosted autophagosome nucleation, increased pro-death autophagy, and restored apoptosis in temozolomide-resistant glioblastoma cells. Finally, targeting PSMC2 provided a unique method for interrupting autophagy-mediated ER stress maintenance and temozolomide resistance in glioblastoma.

We have previously demonstrated that DEC1 promotes osteoblast differentiation. This study aims to evaluate the impact of DEC1 knockout on osteopenic activities, such as osteoclast differentiation and the expression of bone-degrading genes. To gain mechanistic insights, we employed both in vivo and in vitro experiments, utilizing cellular and molecular approaches, including osteoclast differentiation assays and RNA-seq in combination with ChIP-seq. Our results showed that NFATc1, a master regulator of osteoclast differentiation, and PPP3CB, a member of the calcineurin family, were significantly upregulated in DEC1^-/- mice. In vitro experiments revealed that osteoclast differentiation significantly increased both the number and size of osteoclasts in DEC1^-/- bone marrow macrophages (BMMs) compared to DEC1^+/+ BMMs. Additionally, NFATc1 expression was notably higher in DEC1^-/- BMMs than in DEC1^+/+ BMMs. Overexpression of DEC1 reduced NFATc1 promoter activity, while knockout increased it. Furthermore, intracellular free Ca²⁺ levels and calcineurin activity were elevated (∼150 %) in DEC1^-/- BMMs compared to DEC1^+/+ BMMs. Importantly, the use of calcineurin inhibitors and calcium channel blockers effectively abolished the increased osteoclast differentiation observed in DEC1^-/- BMMs. In summary, DEC1 deficiency promotes osteoclast differentiation by enhancing NFATc1 signaling through transcriptional regulation and the Ca²⁺/calcineurin pathway. Clinically, the mRNA levels of DEC1 were reduced by up to 75 % in patients with osteoporosis. The findings of this study establish that inducing DEC1 expression, alongside attenuators of the Ca²⁺/calcineurin pathway, offers a molecular basis for preventing and treating osteoporosis associated with DEC1 deficiency.

PTEN, a tumor suppressor phosphatase, regulates cellular functions by antagonizing the growth promoting PI3K/Akt/mTOR pathway through the dephosphorylation of the second messenger PIP₃. Many preclinical cellular and animal studies have used PTEN inhibitors to highlight specific disease contexts where acute activation of PI3K/Akt/mTOR pathway might offer therapeutic advantages. In the present study we have re-evaluated first-generation PTEN inhibitors, including established bisperoxo-vanadium^(V) complexes (bpVs). In vitro, all compounds tested inhibited PTEN with IC₅₀ values between 0.2-0.8 μM, although their activity diminished under reducing conditions. bpV(phen) and bpV(HΟpic) significantly increased pSer473Akt levels in PTEN wild-type cells while bpV(phen) induced phosphorylation in PTEN null cells upon re-expression of functional PTEN. bpV(ΗΟpic) was less specific since it also triggered PTEN-independent Erk1/2 phosphorylation. In vivo, bpV(phen) administration in Wistar rats enhanced pS6 levels in kidney and liver tissues, but not in several CNS tissues, and led to reduced locomotion and exploratory behaviour in the open field test. The consensus mechanism of action of first generation PTEN inhibitors appears to be oxidative inhibition, however bpV(phen) does not induce oxidation of cellular endogenous PTEN. Instead, our findings suggest that the inhibition of PTEN by bpV(phen) in cells and in vivo may proceed through a mechanism involving non-specific S-nitrosylation of PTEN. Our study highlights the complexity of PTEN inhibition by first-generation compounds and their limitations, such as low specificity, adverse effects and non-specific mechanisms of action, and emphasizes the need for developing more selective and potent PTEN inhibitors with improved efficacy and well-defined mechanisms of actions.

Glucocorticoid-induced osteoporosis (GIOP) is the most common type of secondary osteoporosis, marked by reduced bone density and impaired osteoblast function. Current treatments have serious side effects, highlighting the need for new drug candidates. Pyrimidine derivatives have been noted for their potential in suppressing osteoclastogenesis, but their effects on osteogenesis and GIOP remain underexplored. Our recent study identified a novel pyrimidine derivative, Pym-18a, which enhances osteoblast functions. In this study, Pym-18a was found to mitigate the detrimental effects of Dexamethasone (Dex) in osteoblast cells and in GIOP in Balb/C mice. Pretreatment with Pym-18a followed by Dex (100 µM) for 24 h restored osteoblast alkaline phosphatase activity and viability. Pym-18a reduced Dex-induced apoptosis and reactive oxygen species (ROS) generation at cellular and mitochondrial levels and preserved mitochondrial membrane potential. Dex impaired autophagy and mitophagy, however but Pym-18a pretreatment increased expression of autophagy markers (LC3II) and mitophagy markers (PINK1, Parkin, TOM20) while decreasing P62 expression. The osteogenic effects of Pym-18a were diminished in the presence of 3-MA (an autophagy inhibitor). In silico studies showed mTOR inhibition by Pym-18a, corroborated by its suppression of Dex-induced mTOR activation. In vivo, Pym-18a (10 mg/kg) significantly improved bone microarchitecture, trabecular connectivity, and strength, and corrected P1NP and CTX levels altered by Dex. Pym-18a also promoted autophagy, mitophagy, and suppressed mTOR activation in GIOP mice. Overall, Pym-18a mitigates detrimental effect of Dex by modulating autophagy and PINK/Parkin-mediated mitophagy through mTOR inhibition, suggesting it as a potential novel therapeutic option for GIOP.

Osteoporosis is a chronic disease distinguished by decreased bone density and degradation of bone microstructure, frequently linked with inflammation and oxidative stress, both of which contribute to the acceleration of bone resorption. The compound 5,7-Dihydroxy-4-methylcoumarin (D4M) present in Artemisia dracunculus exhibits significant antioxidant and anti-inflammatory properties. Nonetheless, the potential anti-osteoporotic effects of D4M, along with the molecular targets and mechanisms responsible for these effects, have not been studied. This study aims to assess the impact of D4M on osteoblastogenesis and glucocorticoid-induced osteoporosis while examining the potential underlying mechanisms. We examined the effects of varying concentrations of D4M on the proliferation and differentiation of MC3T3-E1 cells. Additionally, in vivo experiments were carried out using a glucocorticoid-induced zebrafish osteoporosis model to evaluate the effects of D4M on vertebral bone density and osteogenic markers. Target prediction and molecular docking analyses were conducted to investigate the binding interactions between D4M and its target proteins. D4M showed a significant enhancement of MC3T3-E1 cell proliferation and differentiation within the concentration range of 10 to 40 μM, with the greatest increase in mineralization noted at 20 μM. Furthermore, in the zebrafish osteoporosis model, treatment with 20 μM D4M resulted in a significant improvement in vertebral bone density and the restoration of osteoblast-specific marker expression. Ligand-based target prediction identified AKT1 as a potential target for D4M, and molecular docking highlighted the binding interactions between D4M and AKT1 phosphorylation sites. Co-treatment with the AKT1 inhibitor A-443654 abolished the anti-osteoporotic effects of D4M. These findings demonstrate that D4M enhances osteoblast differentiation and mitigates osteoporosis through its interaction with AKT1, suggesting its potential as a therapeutic agent for treating osteoporosis.

Dermal papilla cells (DPCs) are a crucial subset of mesenchymal cells in the skin responsible for regulating hair follicle development and growth, making them invaluable for cell-based therapies targeting hair loss. However, obtaining sufficient DPCs with potent hair-inducing abilities remains a persistent challenge. In this study, the Food and Drug Administration (FDA)-approved drug library was utilized to screen small molecules capable of reprogramming readily accessible human skin fibroblasts into functional DPCs. In the initial screening, five candidate small molecules were identified from a pool of 1,817 compounds, and the small molecule peficitinib was further identified by the further hair follicle regeneration experiments. Following peficitinib treatment, fibroblasts derived from primary human foreskin and scalp exhibited the capability to induce hair growth and possessed a molecular profile highly similar to that of primary DPCs. We refer to these cells as dermal papilla cell-like cells (DPC-LCs). Furthermore, transcriptome analysis showed that the wingless/integrated (Wnt) signaling pathway and the transforming growth factor β (TGF-β) signaling pathway, both of which play crucial roles in hair follicle morphogenesis, are upregulated and enriched in these DPC-LCs. These functional DPC-LCs offer a promising avenue for obtaining a plentiful supply of hair-inducing cells, thereby advancing the development of therapeutic strategies for hair loss treatment.

Colorectal cancer (CRC) is a malignancy with high global incidence and mortality rates, posing a serious threat to human health. Despite favorable outcomes following early detection and surgical intervention, the asymptomatic nature of CRC often results in delayed diagnoses, limiting surgical treatment options. Furthermore, effective therapeutic drugs for CRC remain lacking in clinical practice, highlighting an urgent need to identify novel therapeutic targets. In this study, we identified that prolyl 4-hydroxylase subunit alpha 3 (P4HA3) is significantly upregulated in CRC and is associated with poor prognosis in patients. Both in vitro and in vivo experiments demonstrated that knockdown of P4HA3 induces ferroptosis, thereby inhibiting tumor growth. This ferroptosis induction is closely linked to increased lipid peroxidation, and P4HA3 knockdown promotes ferroptosis by upregulating acyl-CoA synthetase long-chain family member 4 (ACSL4), which regulates polyunsaturated fatty acid-containing phospholipids (PUFA-PLs) biosynthesis. Mechanistically, P4HA3 knockdown stabilizes ACSL4 mRNA by downregulating AUF1, an important RNA-binding protein (RBP) that binds to AU-rich elements (AREs) in the ACSL4 mRNA 3' untranslated region (UTR), thereby preventing its degradation. Additionally, given the lack of research on P4HA3 inhibitors, we employed virtual screening and identified Tubuloside A as a potential therapeutic agent. Tubuloside A promotes the ubiquitin-proteasome degradation of P4HA3, exerting anti-CRC effects. In summary, our findings demonstrate that P4HA3 protects CRC cells from ferroptosis by regulating ACSL4 mRNA stability via AUF1, and Tubuloside A serves as a potential P4HA3 degrader, offering a promising therapeutic strategy for CRC treatment.