Identification of new microtubule small-molecule inhibitors and microtubule-associated genes against triple negative breast cancer.

Breast cancer represents the leading cancer type and leading cause of cancer-related death among women in the world. Triple-negative breast cancer (TNBC) is a subset of breast cancer with the poorest prognosis and still lacking of effective therapeutic options. We recently screened a natural product library and identified 3 new hit compounds with selective and prominent anti-TNBC activities on different subtype of TNBC cell lines. Interestingly, all of these 3 hit compounds belong to "cytoskeletal drugs" that target tubulin and microtubule function. Our data also showed that these hit compounds showed consistently effective on TNBC cells which are resistant to those currently used antimicrotubule agents such as Paclitaxel. RNA-Sequencing analyses revealed the anti-TNBC mechanisms of these hit compounds and identified a subset of new cellular factors commonly affected by hit compounds in different subtypes of TNBC cells. Among them, we demonstrated AHCYL1 and SPG21 as new microtubule-associated proteins, which were required for TNBC cell survival with clinical implication through tissue array analysis. Our studies provide new insights into the mechanisms of TNBC pathogenesis and offer promising therapeutic directions for this aggressive breast cancer.

Roles of Human Endogenous Retrovirus-K-Encoded Np9 in Human Diseases: A Small Protein with Big Functions.

Human Endogenous Retrovirus Sequences (HERVs) constitute up to 8% of the human genome, yet not all HERVs remain silent passengers within our genomes. Some HERVs, especially HERV type K (HERV-K), have been found to be frequently transactivated in a variety of inflammatory diseases and human cancers. Np9, a small protein translated from the HERV-K env reading frame, has been reported as an oncogenic protein and is present in a variety of tumors and transformed cells. The Np9 protein can crosstalk with many cellular factors and is involved in the pathogenicity of various diseases, including some oncogenic virus infections. In the current review, we summarize recent findings about Np9 clinical relevance/implications, its mediated cellular functions/mechanisms, and potential targeted therapies in development.

Development of human endogenous retrovirus type K- related treatments for human diseases.

Human endogenous retroviruses (HERVs) constitute approximately 8% of the human genome and have long been regarded as silent passengers within our genomes. However, the reactivation of HERVs has been increasingly linked to a range of human diseases, particularly the HERV-K (HML-2) family. Many studies are dedicated to elucidating the potential role of HERV-K in pathogenicity. While the underlying mechanisms require further investigation, targeting HERV-K transactivation emerges as a promising avenue for treating human diseases, including cancer, autoimmune disorders, neurodegenerative conditions, and infectious diseases. In this review, we summarize recent advancements in the development of HERV-K-targeted therapeutic strategies against various human diseases, including antiretroviral drugs, immunotherapy, and vaccines.

Human endogenous retrovirus type K encoded Np9 oncoprotein induces DNA damage response.

Human endogenous retrovirus sequences (HERVs) constitute up to 8% of the human genome, yet not all HERVs remain silent passengers within our genomes. Some HERVs, especially the HERV type K (HERV-K), have been found to be frequently transactivated in a variety of inflammatory diseases and human cancers. Np9, a 9-kDa HERV-K encoded protein, has been reported as an oncoprotein and found present in a variety of tumors and transformed cells. In the current study, we for the first time reported that ectopic expression of Np9 protein was able to induce DNA damage response from host cells especially through upregulation of γH2AX. Furthermore, we found that direct knockdown of Np9 by RNAi in Kaposi's Sarcoma-associated herpesvirus (KSHV) infected cells effectively reduced LANA expression, the viral major latent oncoprotein in vitro and in vivo, which may represent a novel strategy against virus-associated malignancies.

TP53RK Drives the Progression of Chronic Kidney Disease by Phosphorylating Birc5.

Renal fibrosis is a common characteristic of various chronic kidney diseases (CKDs) driving the loss of renal function. During this pathological process, persistent injury to renal tubular epithelial cells and activation of fibroblasts chiefly determine the extent of renal fibrosis. In this study, the role of tumor protein 53 regulating kinase (TP53RK) in the pathogenesis of renal fibrosis and its underlying mechanisms is investigated. TP53RK is upregulated in fibrotic human and animal kidneys with a positive correlation to kidney dysfunction and fibrotic markers. Interestingly, specific deletion of TP53RK either in renal tubule or in fibroblasts in mice can mitigate renal fibrosis in CKD models. Mechanistic investigations reveal that TP53RK phosphorylates baculoviral IAP repeat containing 5 (Birc5) and facilitates its nuclear translocation; enhanced Birc5 displays a profibrotic effect possibly via activating PI3K/Akt and MAPK pathways. Moreover, pharmacologically inhibiting TP53RK and Birc5 using fusidic acid (an FDA-approved antibiotic) and YM-155(currently in clinical phase 2 trials) respectively both ameliorate kidney fibrosis. These findings demonstrate that activated TP53RK/Birc5 signaling in renal tubular cells and fibroblasts alters cellular phenotypes and drives CKD progression. A genetic or pharmacological blockade of this axis serves as a potential strategy for treating CKDs.

Mitochondrial proton leak in cardiac aging.

Age-associated diseases are becoming progressively more prevalent, reflecting the increased lifespan of the world's population. However, the fundamental mechanisms of physiologic aging are poorly understood, and in particular, the molecular pathways that mediate cardiac aging and its associated dysfunction are unclear. Here, we focus on certain ion flux abnormalities of the mitochondria that may contribute to cardiac aging and age-related heart failure. Using oxidative phosphorylation, mitochondria pump protons from the matrix to the intermembrane space to generate a proton gradient across the inner membrane. The protons are returned to the matrix by the ATPase complex within the membrane to generate ATP. However, a portion of protons leak back to the matrix and do not drive ATP production, and this event is called proton leak or uncoupling. Accumulating evidence suggests that mitochondrial proton leak is increased in the cardiac myocytes of aged hearts. In this mini-review, we discuss the measurement methods and major sites of mitochondrial proton leak with an emphasis on the adenine nucleotide transporter 1 (ANT1), and explore the possibility of inhibiting augmented mitochondrial proton leak as a therapeutic intervention to mitigate cardiac aging.

LONP1 targets HMGCS2 to protect mitochondrial function and attenuate chronic kidney disease.

Mitochondria comprise the central metabolic hub of cells and their imbalance plays a pathogenic role in chronic kidney disease (CKD). Here, we studied Lon protease 1 (LONP1), a major mitochondrial protease, as its role in CKD pathogenesis is unclear. LONP1 expression was decreased in human patients and mice with CKD, and tubular-specific Lonp1 overexpression mitigated renal injury and mitochondrial dysfunction in two different models of CKD, but these outcomes were aggravated by Lonp1 deletion. These results were confirmed in renal tubular epithelial cells in vitro. Mechanistically, LONP1 downregulation caused mitochondrial accumulation of the LONP1 substrate, 3-hydroxy-3-methylglutaryl-CoA synthase 2 (HMGCS2), which disrupted mitochondrial function and further accelerated CKD progression. Finally, computer-aided virtual screening was performed, which identified a novel LONP1 activator. Pharmacologically, the LONP1 activator attenuated renal fibrosis and mitochondrial dysfunction. Collectively, these results imply that LONP1 is a promising therapeutic target for treating CKD.

A novel 3-phenylglutaric acid derivative (84-B10) alleviates cisplatin-induced acute kidney injury by inhibiting mitochondrial oxidative stress-mediated ferroptosis.

Cisplatin is one of the most effective chemotherapy drugs and is widely used for cancer treatment. However, its clinical use is limited by nephrotoxicity. Emerging findings suggested that both ferroptosis and mitochondrial dysfunction mediate cisplatin-induced nephrotoxicity. In the current study, a novel 3-phenylglutaric acid derivative 5-[[2-(4-methoxyphenoxy)-5-(trifluoromethyl)phenyl]amino]-5-oxo-3-phenylpentanoic acid (referred to as 84-B10) was found to play a protective role in cisplatin-induced acute kidney injury with no tumor promoting effects. A genome-wide transcriptome analysis indicated that the protective effect of 84-B10 might be dependent on antagonizing ferroptosis. In accordance, lipid peroxide accumulation and downregulation of key ferroptosis suppressors were reversed using 84-B10 treatment both in vivo and in vitro. In addition, 84-B10 inhibited cisplatin-induced mitochondrial damage and mitochondrial reactive oxygen species (mtROS) production and restored superoxide dismutases (SODs). Furthermore, 84-B10 showed similar therapeutic effects to MnTBAP (a cell-permeable SOD mimetic) in eliminating mtROS, restoring mitochondrial homeostasis, and inhibiting ferroptosis under cisplatin challenge. Comparable effects of 84-B10 and liproxstatin-1 in ameliorating cisplatin-induced ferroptosis were observed. However, liproxstatin-1 failed to prevent mitochondrial dysfunction. These data indicated that mtROS might act upstream of cisplatin-induced tubular ferroptosis. Taken together, the novel 3-phenylglutaric acid derivative 84-B10 showed therapeutic potential against cisplatin-induced nephrotoxicity possibly by restoring mitochondria homeostasis and inhibiting mtROS-induced ferroptosis, which suggests the potential use of 84-B10 in preventing and treating cisplatin-nephrotoxicity.

Anti-anemia drug FG4592 retards the AKI-to-CKD transition by improving vascular regeneration and antioxidative capability.

Acute kidney injury (AKI) is a known risk factor for the development of chronic kidney disease (CKD), with no satisfactory strategy to prevent the progression of AKI to CKD. Damage to the renal vascular system and subsequent hypoxia are common contributors to both AKI and CKD. Hypoxia-inducible factor (HIF) is reported to protect the kidney from acute ischemic damage and a novel HIF stabilizer, FG4592 (Roxadustat), has become available in the clinic as an anti-anemia drug. However, the role of FG4592 in the AKI-to-CKD transition remains elusive. In the present study, we investigated the role of FG4592 in the AKI-to-CKD transition induced by unilateral kidney ischemia-reperfusion (UIR). The results showed that FG4592, given to mice 3 days after UIR, markedly alleviated kidney fibrosis and enhanced renal vascular regeneration, possibly via activating the HIF-1α/vascular endothelial growth factor A (VEGFA)/VEGF receptor 1 (VEGFR1) signaling pathway and driving the expression of the endogenous antioxidant superoxide dismutase 2 (SOD2). In accordance with the improved renal vascular regeneration and redox balance, the metabolic disorders of the UIR mice kidneys were also attenuated by treatment with FG4592. However, the inflammatory response in the UIR kidneys was not affected significantly by FG4592. Importantly, in the kidneys of CKD patients, we also observed enhanced HIF-1α expression which was positively correlated with the renal levels of VEGFA and SOD2. Together, these findings demonstrated the therapeutic effect of the anti-anemia drug FG4592 in preventing the AKI-to-CKD transition related to ischemia and the redox imbalance.

Novel sulfonamide porphyrin TBPoS-2OH used in photodynamic therapy for malignant melanoma.

The application of photodynamic therapy (PDT) for the treatment of skin diseases has been receiving much attention. Here, we examined the anti-tumor effect of a novel porphyrin-based photosensitizer TBPoS-2OH in the malignant melanoma A375 and B16 cells. TBPoS-2OH has obvious cell photo-cytotoxicity, but it has low cell dark-cytotoxicity. Further research showed that TBPoS-2OH is enriched in lysosomes after being taken up by cells. Subsequently, the apoptotic rates were significantly increased in TBPoS-2OH-treated A375 and B16 cells. The specific mechanism may be that after receiving light stimulation, TBPoS-2OH could effectively increase the level of intracellular reactive oxygen species (ROS), thereby activating mitochondrial apoptosis pathway-related proteins in A375 and B16 cells. We found an increase in the content of cytochrome C in the cytoplasm, and the levels of related proteins, such as cleaved caspase-3, cleaved caspase-9, and cleaved PARP1, were significantly increased in TBPoS-2OH-treated cells. These results indicated that the new compound TBPoS-2OH could be developed and become an alternative drug for the treatment of melanoma. Some reference ideas for the development of new photosensitizers are also provided.

A red-light-activated sulfonamide porphycene for highly efficient photodynamic therapy against hypoxic tumor.

Photodynamic therapy (PDT) is an emerging alternative cancer treatment modality that utilizes photo-sensitivity to cause cell death upon photo-irradiation. However, PDT efficiency has been hampered by tumor hypoxia, blue-shifted excitation wavelengths, and the high dark toxicity of photo-sensitizers. We designed and synthesized two novel porphycene-based photosensitizers (TBPoS-OH and TBPoS-2OH) with potent photo-cytotoxicity and a LD50 in the nM range under both normoxic and hypoxic conditions in a variety of cell types after photo-irradiation (λ = 640 ± 15 nm). Further studies showed fast-cellular uptake for TBPoS-OH that localized lysosomes and subsequently induced cell apoptosis via the lysosomal-mitochondrial pathway. Moreover, TBPoS-OH significantly reduced tumor growth in two xenografted mouse models bearing melanoma A375 and B16 cells. Finally, TBPoS-OH exhibited no obvious immunogenicity and toxicity to blood cells and major organs in mice. These data demonstrated that these two porphycene-based photosensitizers, especially TBPoS-OH, could be developed as a potential PDT modality.

Genetic variations in the CXCR5 gene decrease the risk of clinical relapse after discontinuation of nucleos(t)ide analogue therapy in patients with chronic hepatitis B.

Discontinuation of nucleos(t)ide analogue (NA) therapy in patients with chronic hepatitis B (CHB) remains a global but controversial problem. Clinical outcomes of NA cessation depend on the interplay between viral factors and host immunity. Recent studies have shown that genetic polymorphisms might influence the immune response in chronic HBV infection. A total of 33 single-nucleotide polymorphisms (SNPs) from 16 genes (BCL6, CD40, CD40L, CTLA-4, CXCL13, CXCR5, ICOS, IL-21, HLA-C, NTCP, UBE2L3, STAT4, IFN-λ3, CYP27B1, INST10, and IPS1) were selected and analyzed in 106 CHB patients enrolled in an off-treatment cohort. Significantly unbalanced distributions between patients who experienced clinical relapse and those who did not were found regarding two SNPs, rs676925 in CXCR5 and rs733618 in CTLA-4. Furthermore, the genotype 'GC' of rs676925 were associated with decreased risk of clinical relapse, implicating that rs676925 may serve as a protective factor for HBV control and facilitate a virus-specific immune response. We also compared the expression of CXCR5 in lymphocytes and its ligand CXCL13 in plasma between different genotypes of rs676925. However, no significant differences were observed. In conclusion, this study suggested that the rs676925 'GC' genotype of the CXCR5 gene were associated with decreased risk of clinical relapse after discontinuation of long-term NA therapy in CHB patients.

Potential target-related proteins in rabbit platelets treated with active monomers dehydrocorydaline and canadine from Rhizoma corydalis.

BACKGROUND: Dehydrocorydaline (DHC) and canadine (THB) are two active alkaloid compounds in Corydalis yanhusuo (Y.H. Chou & Chun C. Hsu) W.T. Wang ex Z.Y. Su & C.Y. Wu (Papaveraceae) (Rhizoma Corydalis). DHC and THC were previously shown to exert anti-platelet aggregation effect dose-dependently, but their exact mechanisms had not yet been addressed. Therefore, it is essential to study the mechanisms of DHC and THB affecting on platelet's function. PURPOSE: To investigate the anti-platelet effects and corresponding signal cascades of DHC and THB on platelet aggregation. METHODS: Firstly, in vitro anti-platelet aggregation of DHC and THB induced by different agonists including thrombin (THR), adenosine diphosphate (ADP) and arachidonic acid (AA) were determined through turbidimetry method. Then the possible target-related platelet proteins after treated with DHC/THB were separated and identified by two dimensional gel electrophoresis (2-DE) and MALDI-TOF-MS/MS analysis, respectively. Finally, the signal cascades network induced by DHC/THB were predicted through functional analysis of these proteins along with the determination of platelet DAG concentration. RESULTS: The platelet aggregation stimulated by THR, ADP and AA were inhibited by DHC and THB dose-dependently to a certain degree. Meanwhile, DHC and THB had the strongest effect on ADP- and THR-induced platelet aggregation respectively. In addition, treatment of these two compounds caused regulations of about sixty proteins in platelet, including cytoskeleton proteins, cell signaling proteins, proteins related to material energy metabolism, etc. CONCLUSIONS: Using proteomic analysis combined with platelet aggregation test and ELISA, this study was successful in exploring the possible mechanisms of DHC/THB on platelet aggregation. DHC might inhibit platelet aggregation by a mechanism involving the ADP receptors P2Y1 and P2Y12, and the effect of THB on platelet function may be related to its binding to THR receptor PAR1 for mediated Gi signaling pathway. These results provide fundamental information for the anti-thrombotic effect of RC.

Nicotinamide mononucleotide alleviates Aluminum induced bone loss by inhibiting the TXNIP-NLRP3 inflammasome.

Aluminum (Al) recognized as a persistent environmental contaminant is associated with bone diseases. Nicotinamide mononucleotide (NMN) is an intermediate of nicotinamide adenine dinucleotide (NAD+) biosynthesis widely used to replenish NAD+. Increasing evidences demonstrated that replenishment of NAD+ can protect against bone loss. However, the potentially protective effects of NMN against Al-induced bone impairment and the underlying mechanisms remain unknown. In the present study, we sought to investigate the protective effects of NMN on Al-induced bone damages and elucidate the potential mechanisms. We orally exposed AlCl3 (10 mg/L) to Sprague-Dawley rats in drinking water for 12 weeks while NMN (20 mg/kg) were intraperitoneally injected in last 4 weeks. We found that Al could induce bone damages, bone loss and oxidative stress. In addition, we showed that Al triggered inflammatory responses, which is mediated by the NOD-like receptor pyrin domain containing 3 (NLRP3) inflammasome activation. However, NMN treatment significantly alleviated Al-induced bone injuries by decreasing bone loss, suppressing oxidative stress as well as inhibiting Thioredoxin-interacting protein (TXNIP)-NLRP3 inflammasome pathway and pro-inflammatory cytokine production in vivo and in vitro. Meanwhile, treatment with TXNIP siRNA performed the same protective effects as NMN in Al-treated MC3T3-E1 cells. Collectively, our results suggest that NMN may reduce Al-induced bone loss partly by suppression of the TXNIP-NLRP3 inflammasome pathway.

A case report of CRB2 mutation identified in a Chinese boy with focal segmental glomerulosclerosis.

RATIONALE: Focal segmental glomerulosclerosis (FSGS) is a common disease resulting in end-stage renal disease. The incidence of FSGS is increasing in Western countries. The clinical manifestations include proteinuria, hypoproteinemia, oedema, and hypertension. Single-gene heritable mutations are considered to be the source of FSGS pathogenicity according to recent in-depth studies on the pathogenesis. Here, we first reported the case of a Chinese boy whose histology presented with FSGS caused by a compound heterozygous mutation. PATIENT CONCERNS: A 7-year-old Chinese boy was repeatedly admitted to our hospital for fever, cough, and proteinuria since he was 1.6 years old. DIAGNOSES: FSGS was identified by renal biopsy. Whole exome sequencing (WES) showed that a novel mutation of crumbs homolog 2 (CRB2) was identified in a Chinese boy with FSGS. INTERVENTIONS: Patient was treated with low-dose corticosteroid and mycophenolate mofetil for maintenance therapy. OUTCOMES: At last follow-up, protein (+∼++) was observed in his urinalysis. LESSONS: We identified a novel mutation of CRB2 in a Chinese boy with FSGS that had never been described in a previous report. These findings suggested that mutations in recessive disease genes are more frequent among early-onset disease.