Clemastine and hyperthermia enhance sensitization of osteosarcoma cells for apoptosis.

Clemastine is an antagonist of histamine H1 receptor may provide benefits in the treatment of osteosarcoma (OS). In the current study, we used hyperthermia approach to sensitize OS cells to clemastine-mediated cell death. Osteosarcoma U-2 OS and Saos-2 cells were treated with clemastine at 37°C, followed by 42°C for 2 h, and released at 37°C for 6 h. The impact of clemastine and hyperthermia on OS cell survival and autophagy-mediated cell death was investigated. Exposure of U-2 OS and Saos-2 cells to clemastine and hyperthermia (42°C) inhibited dose-dependent clemastine-mediated cell survival by increasing cell apoptosis. Hyperthermia and clemastine exposure modulated inflammatory and unfolded protein response (UPR) signaling differentially in U-2 OS and Saos-2 cells. Exposure of U-2 OS and Saos-2 cells to hyperthermia and clemastine inhibited AKT/mTOR and induced expression of the autophagy biomarkers LC3B II and LC3-positive puncta formation. The inhibition of autophagy by 3-methyladenine blocked hyperthermia and clemastine-mediated induction of LC3B II, LC3-positive puncta formation, and OS cell apoptosis. These results indicate that clemastine and hyperthermia sensitize OS cell lines by inducing increased autophagic cell death. Collectively, our data suggest that hyperthermia along with antihistamine therapy may provide an improved approach for the treatment of OS.

Gene expression analysis suggests immunosuppressive roles of endolysosomes in glioblastoma.

Targeting endolysosomes is a strategy extensively pursued for treating cancers, including glioblastomas (GBMs), on the basis that the intact function of these subcellular organelles is key to tumor cell autophagy and survival. Through gene expression analyses and cell type abundance estimation in GBMs, we showed that genes associated with the endolysosomal machinery are more prominently featured in non-tumor cells in GBMs than in tumor cells, and that tumor-associated macrophages represent the primary immune cell type that contributes to this trend. Further analyses found an enrichment of endolysosomal pathway genes in immunosuppressive (pro-tumorigenic) macrophages, such as M2-like macrophages or those associated with worse prognosis in glioma patients, but not in those linked to inflammation (anti-tumorigenic). Specifically, genes critical to the hydrolysis function of endolysosomes, including progranulin and cathepsins, were among the most positively correlated with immunosuppressive macrophages, and elevated expression of these genes is associated with worse patient survival in GBMs. Together, these results implicate the hydrolysis function of endolysosomes in shaping the immunosuppressive microenvironment of GBM. We propose that targeting endolysosomes, in addition to its detrimental effects on tumor cells, can be leveraged for modulating immunosuppression to render GBMs more amenable to immunotherapies.

Interplay between ATRX and IDH1 mutations governs innate immune responses in diffuse gliomas.

Stimulating the innate immune system has been explored as a therapeutic option for the treatment of gliomas. Inactivating mutations in ATRX, defining molecular alterations in IDH-mutant astrocytomas, have been implicated in dysfunctional immune signaling. However, little is known about the interplay between ATRX loss and IDH mutation on innate immunity. To explore this, we generated ATRX-deficient glioma models in the presence and absence of the IDH1R132H mutation. ATRX-deficient glioma cells are sensitive to dsRNA-based innate immune agonism and exhibit impaired lethality and increased T-cell infiltration in vivo. However, the presence of IDH1R132H dampens baseline expression of key innate immune genes and cytokines in a manner restored by genetic and pharmacological IDH1R132H inhibition. IDH1R132H co-expression does not interfere with the ATRX deficiency-mediated sensitivity to dsRNA. Thus, ATRX loss primes cells for recognition of dsRNA, while IDH1R132H reversibly masks this priming. This work reveals innate immunity as a therapeutic vulnerability of astrocytomas.

Heat shock factor 1 directly regulates transsulfuration pathway to promote prostate cancer proliferation and survival.

There are limited therapeutic options for patients with advanced prostate cancer (PCa). We previously found that heat shock factor 1 (HSF1) expression is increased in PCa and is an actionable target. In this manuscript, we identify that HSF1 regulates the conversion of homocysteine to cystathionine in the transsulfuration pathway by altering levels of cystathionine-ß-synthase (CBS). We find that HSF1 directly binds the CBS gene and upregulates CBS mRNA levels. Targeting CBS decreases PCa growth and induces tumor cell death while benign prostate cells are largely unaffected. Combined inhibition of HSF1 and CBS results in more pronounced inhibition of PCa cell proliferation and reduction of transsulfuration pathway metabolites. Combination of HSF1 and CBS knockout decreases tumor size for a small cell PCa xenograft mouse model. Our study thus provides new insights into the molecular mechanism of HSF1 function and an effective therapeutic strategy against advanced PCa.

Targeting glutamine dependence with DRP-104 inhibits proliferation and tumor growth of castration-resistant prostate cancer.

BACKGROUND: Prostate cancer (PCa) continues to be one of the leading causes of cancer deaths in men. While androgen deprivation therapy is initially effective, castration-resistant PCa (CRPC) often recurs and has limited treatment options. Our previous study identified glutamine metabolism to be critical for CRPC growth. The glutamine antagonist 6-diazo-5-oxo-l-norleucine (DON) blocks both carbon and nitrogen pathways but has dose-limiting toxicity. The prodrug DRP-104 is expected to be preferentially converted to DON in tumor cells to inhibit glutamine utilization with minimal toxicity. However, CRPC cells' susceptibility to DRP-104 remains unclear. METHODS: Human PCa cell lines (LNCaP, LAPC4, C4-2/MDVR, PC-3, 22RV1, NCI-H660) were treated with DRP-104, and effects on proliferation and cell death were assessed. Unbiased metabolic profiling and isotope tracing evaluated the effects of DRP-104 on glutamine pathways. Efficacy of DRP-104 in vivo was evaluated in a mouse xenograft model of neuroendocrine PCa, NCI-H660. RESULTS: DRP-104 inhibited proliferation and induced apoptosis in CRPC cell lines. Metabolite profiling showed decreases in the tricarboxylic acid cycle and nucleotide synthesis metabolites. Glutamine isotope tracing confirmed the blockade of both carbon pathway and nitrogen pathways. DRP-104 treated CRPC cells were rescued by the addition of nucleosides. DRP-104 inhibited neuroendocrine PCa xenograft growth without detectable toxicity. CONCLUSIONS: The prodrug DRP-104 blocks glutamine carbon and nitrogen utilization, thereby inhibiting CRPC growth and inducing apoptosis. Targeting glutamine metabolism pathways with DRP-104 represents a promising therapeutic strategy for CRPC.

Repurposing Clemastine to Target Glioblastoma Cell Stemness.

Brain tumor-initiating cells (BTICs) and tumor cell plasticity promote glioblastoma (GBM) progression. Here, we demonstrate that clemastine, an over-the-counter drug for treating hay fever and allergy symptoms, effectively attenuated the stemness and suppressed the propagation of primary BTIC cultures bearing PDGFRA amplification. These effects on BTICs were accompanied by altered gene expression profiling indicative of their more differentiated states, resonating with the activity of clemastine in promoting the differentiation of normal oligodendrocyte progenitor cells (OPCs) into mature oligodendrocytes. Functional assays for pharmacological targets of clemastine revealed that the Emopamil Binding Protein (EBP), an enzyme in the cholesterol biosynthesis pathway, is essential for BTIC propagation and a target that mediates the suppressive effects of clemastine. Finally, we showed that a neural stem cell-derived mouse glioma model displaying predominantly proneural features was similarly susceptible to clemastine treatment. Collectively, these results identify pathways essential for maintaining the stemness and progenitor features of GBMs, uncover BTIC dependency on EBP, and suggest that non-oncology, low-toxicity drugs with OPC differentiation-promoting activity can be repurposed to target GBM stemness and aid in their treatment.

Interplay between ATRX and IDH1 mutations governs innate immune responses in diffuse gliomas.

Stimulating the innate immune system has been explored as a therapeutic option for the treatment of gliomas. Inactivating mutations in ATRX , defining molecular alterations in IDH -mutant astrocytomas, have been implicated in dysfunctional immune signaling. However, little is known about the interplay between ATRX loss and IDH mutation on innate immunity. To explore this, we generated ATRX knockout glioma models in the presence and absence of the IDH1 R 132 H mutation. ATRX-deficient glioma cells were sensitive to dsRNA-based innate immune agonism and exhibited impaired lethality and increased T-cell infiltration in vivo . However, the presence of IDH1 R 132 H dampened baseline expression of key innate immune genes and cytokines in a manner restored by genetic and pharmacological IDH1 R132H inhibition. IDH1 R132H co-expression did not interfere with the ATRX KO-mediated sensitivity to dsRNA. Thus, ATRX loss primes cells for recognition of dsRNA, while IDH1 R132H reversibly masks this priming. This work reveals innate immunity as a therapeutic vulnerability of astrocytoma.

Rewiring of the N-Glycome with prostate cancer progression and therapy resistance.

An understanding of the molecular features associated with prostate cancer progression (PCa) and resistance to hormonal therapy is crucial for the identification of new targets that can be utilized to treat advanced disease and prolong patient survival. The glycome, which encompasses all sugar polymers (glycans) synthesized by cells, has remained relatively unexplored in the context of advanced PCa despite the fact that glycans have great potential value as biomarkers and therapeutic targets due to their high density on the cell surface. Using imaging mass spectrometry (IMS), we profiled the N-linked glycans in tumor tissue derived from 131 patients representing the major disease states of PCa to identify glycosylation changes associated with loss of tumor cell differentiation, disease remission, therapy resistance and disease recurrence, as well as neuroendocrine (NE) differentiation which is a major mechanism for therapy failure. Our results indicate significant changes to the glycosylation patterns in various stages of PCa, notably a decrease in tri- and tetraantennary glycans correlating with disease remission, a subsequent increase in these structures with the transition to therapy-resistant PCa, and downregulation of complex N-glycans correlating with NE differentiation. Furthermore, both nonglucosylated and monoglucosylated mannose 9 demonstrate aberrant upregulation in therapy-resistant PCa which may be useful therapeutic targets as these structures are not normally presented in healthy tissue. Our findings characterize changes to the tumor glycome that occur with hormonal therapy and the development of castration-resistant PCa (CRPC), identifying several glycan markers and signatures which may be useful for diagnostic or therapeutic purposes.

Oncofetal protein glypican-3 is a biomarker and critical regulator of function for neuroendocrine cells in prostate cancer.

Neuroendocrine (NE) cells comprise ~1% of epithelial cells in benign prostate and prostatic adenocarcinoma (PCa). However, they become enriched in hormonally treated and castration-resistant PCa (CRPC). In addition, close to 20% of hormonally treated tumors recur as small cell NE carcinoma (SCNC), composed entirely of NE cells, which may be the result of clonal expansion or lineage plasticity. Since NE cells do not express androgen receptors (ARs), they are resistant to hormonal therapy and contribute to therapy failure. Here, we describe the identification of glypican-3 (GPC3) as an oncofetal cell surface protein specific to NE cells in prostate cancer. Functional studies revealed that GPC3 is critical to the viability of NE tumor cells and tumors displaying NE differentiation and that it regulates calcium homeostasis and signaling. Since our results demonstrate that GPC3 is specifically expressed by NE cells, patients with confirmed SCNC may qualify for GPC3-targeted therapy which has been developed in the context of liver cancer and displays minimal toxicity due to its tumor-specific expression. © 2023 The Pathological Society of Great Britain and Ireland.

Cancer-associated SMARCAL1 loss-of-function mutations promote alternative lengthening of telomeres and tumorigenesis in telomerase-negative glioblastoma cells.

BACKGROUND: Telomere maintenance mechanisms are required to enable the replicative immortality of malignant cells. While most cancers activate the enzyme telomerase, a subset of cancers uses telomerase-independent mechanisms termed alternative lengthening of telomeres (ALT). ALT occurs via homology-directed-repair mechanisms and is frequently associated with ATRX mutations. We previously showed that a subset of adult glioblastoma (GBM) patients with ATRX-expressing ALT-positive tumors harbored loss-of-function mutations in the SMARCAL1 gene, which encodes an annealing helicase involved in replication fork remodeling and the resolution of replication stress. However, the causative relationship between SMARCAL1 deficiency, tumorigenesis, and de novo telomere synthesis is not understood. METHODS: We used a patient-derived ALT-positive GBM cell line with native SMARCAL1 deficiency to investigate the role of SMARCAL1 in ALT-mediated de novo telomere synthesis, replication stress, and gliomagenesis in vivo. RESULTS: Inducible rescue of SMARCAL1 expression suppresses ALT indicators and inhibits de novo telomere synthesis in GBM and osteosarcoma cells, suggesting that SMARCAL1 deficiency plays a functional role in ALT induction in cancers that natively lack SMARCAL1 function. SMARCAL1-deficient ALT-positive cells can be serially propagated in vivo in the absence of detectable telomerase activity, demonstrating that the SMARCAL1-deficient ALT phenotype maintains telomeres in a manner that promotes tumorigenesis. CONCLUSIONS: SMARCAL1 deficiency is permissive to ALT and promotes gliomagenesis. Inducible rescue of SMARCAL1 in ALT-positive cell lines permits the dynamic modulation of ALT activity, which will be valuable for future studies aimed at understanding the mechanisms of ALT and identifying novel anticancer therapeutics that target the ALT phenotype.

Joint gene network construction by single-cell RNA sequencing data.

In contrast to differential gene expression analysis at the single-gene level, gene regulatory network (GRN) analysis depicts complex transcriptomic interactions among genes for better understandings of underlying genetic architectures of human diseases and traits. Recent advances in single-cell RNA sequencing (scRNA-seq) allow constructing GRNs at a much finer resolution than bulk RNA-seq and microarray data. However, scRNA-seq data are inherently sparse, which hinders the direct application of the popular Gaussian graphical models (GGMs). Furthermore, most existing approaches for constructing GRNs with scRNA-seq data only consider gene networks under one condition. To better understand GRNs across different but related conditions at single-cell resolution, we propose to construct Joint Gene Networks with scRNA-seq data (JGNsc) under the GGMs framework. To facilitate the use of GGMs, JGNsc first proposes a hybrid imputation procedure that combines a Bayesian zero-inflated Poisson model with an iterative low-rank matrix completion step to efficiently impute zero-inflated counts resulted from technical artifacts. JGNsc then transforms the imputed data via a nonparanormal transformation, based on which joint GGMs are constructed. We demonstrate JGNsc and assess its performance using synthetic data. The application of JGNsc on two cancer clinical studies of medulloblastoma and glioblastoma gains novel insights in addition to confirming well-known biological results.

MOR promotes epithelial-mesenchymal transition and proliferation via PI3K/AKT signaling pathway in human colorectal cancer.

The mu-opioid receptor (MOR), a membrane-bound G protein-coupled receptor, is implicated in progression and long-term outcome of several types of tumors. However, the expression and clinical significance of MOR in colorectal cancer (CRC) remain unclear. In this study, a total of 180 paraffin-embedded samples of paired tumors and normal tissues from CRC patients are used to explore expression levels of MOR by immunohistochemistry (IHC). Results show that MOR is highly expressed in tumors compared with that in paired normal tissues ( P<0.0001). MOR expression levels are associated with the degree of differentiation ( P<0.001) and the regional lymph node metastasis ( P<0.001). In addition, a significant difference is also found in the overall survival (OS) between MOR low- and high-expression groups ( P=0.002), especially in patients with TNM stage III or IV CRC ( P=0.007). Both univariate ( P=0.002) and multivariate ( P=0.013) analyses indicated that MOR is an independent risk factor associated with CRC prognosis. We further investigate the mechanism in MOR-positive CRC cell line HCT116. The results show that silencing of MOR significantly suppresses epithelial-mesenchymal transition (EMT), in addition to suppressing cell proliferation, migration, and invasion. In addition, the expression of downstream p-AKT is also significantly downregulated, and the above suppression effect could be rescued by PI3K/AKT signaling agonist. We conclude that MOR mediates EMT via PI3K/AKT signaling, facilitating lymph node metastasis and resulting in poor survival of CRC patients. Our findings suggest that MOR is a novel prognostic indicator and the application of opioid receptor antagonists may be a novel therapeutic strategy for CRC patients with high MOR expression.

Rubicon promotes the M2 polarization of Kupffer cells via LC3-associated phagocytosis-mediated clearance to improve liver transplantation.

BACKGROUND: Acute rejection (AR) after liver transplantation (LT) is closely related to the survival of patients after surgery. Enhancement of the ability of Kupffer cells (KCs) to eliminate apoptotic cells can effectively alleviate AR. METHODS: Rubicon lentivirus (LV) and Rubicon small interfering RNA (siRNA) were transfected into KCs extracted from the liver tissue of mice. Primary KCs were extracted and cocultured with zymosan and apoptotic T lymphocytes. The levels of CD86, CD163, IL-10, TNF-α, TGF-ß, JAK1, STAT6, AKT1, mTOR and peroxisome proliferator-activated receptor-γ (PPARγ) were assessed via Western blotting (WB) and q-PCR. The levels of CD86 and CD163 were assessed via flow cytometry. mCherry-GFP-LC3 adenovirus (AV) was transfected into KCs. The recruitment of LC3II and the fusion of phagosomes and lysosomes were detected using immunofluorescence. Rubicon adeno-associated virus (AAV) was transfected into the liver tissue of mice via the portal vein, and models of immune tolerance (IT) and AR following LT were established. Pathological changes in the liver tissue were detected using HE staining. Apoptotic cells were assessed via TUNEL staining. The polarization state of KCs was detected via immunohistochemical staining. RESULTS: Rubicon-mediated LC3-associated phagocytosis (LAP) promotes the ability of KCs to degrade and clear apoptotic T lymphocytes. Polyunsaturated fatty acids (PUFAs), the product of apoptotic T lymphocyte degradation, activate PPARγ, which further promotes the M2 polarization of KCs. Enhanced degradation mediated by Rubicon contributes to promoting the M2 polarization of KCs and a microenvironment supportive of IT. CONCLUSIONS: Rubicon-mediated LAP promotes the clearance capability and M2 polarization of KCs via PUFA-dependent PPARγ activation to improve LT.

A study protocol of a randomized phase II trial of perioperative chemoimmunotherapy verses perioperative chemoimmunotherapy plus preoperative chemoradiation for locally advanced gastric (G) or gastroesophageal junction (GEJ) adenocarcinoma: the NeoRacing study.

BACKGROUND: Perioperative chemotherapy (ChT) and preoperative chemoradiation (CRT) are both the standard treatments for locally advanced gastric cancer (LAGC). CRT can achieve a higher pathological complete regression (pCR) rate, but whether this higher pCR rate can be transformed into a long-term survival benefit remains inconclusive. Therefore, relevant studies are in progress. On the other hand, immunotherapy has been established for the first-line treatment of advanced gastric cancer (AGC) and has been widely explored in the perioperative setting. The combination of chemotherapy/radiotherapy and immunotherapy may have a synergistic effect, which will lead to a better antitumor effect. The preliminary reports of ongoing studies show promising results, including a further improved pCR rate. However, the preferred treatment combination for LAGC is still not established. To solve this problem, we are carrying out this randomized phase II trial, which aims to evaluate the efficacy and safety of perioperative chemotherapy plus the use of PD-1 antibody with or without preoperative chemoradiation for LAGC. METHODS: Eligible patients with LAGC or gastroesophageal junction (GEJ) adenocarcinoma were randomized to receive perioperative ChT, PD-1 antibody, surgery with (Arm A) or without preoperative CRT (Arm B), and PD-1 antibody maintenance until one year after surgery. The primary endpoint of this study is that the pCR rate of Arm A will be significantly higher than that of Arm B. The secondary endpoints include the pathological partial regression (pPR) rate, R0 resection rate, objective response rate (ORR), event-free survival (EFS), overall survival (OS), safety and surgical complications. Moreover, several explorative endpoints will be evaluated to find and validate the predictive biomarkers of immunotherapy. DISCUSSION: The results of the NeoRacing study will provide important information concerning the application of PD-1 antibody in LAGC patients during the perioperative setting. Meanwhile, the two treatment protocols will be compared in terms of efficacy and safety. TRIAL REGISTRATION: ClinicalTrials.gov , NCT05161572 . Registered 17 December 2021 - Retrospectively registered.

Purine Synthesis Inhibitor L-Alanosine Impairs Mitochondrial Function and Stemness of Brain Tumor Initiating Cells.

Glioblastoma (GBM) is a lethal brain cancer exhibiting high levels of drug resistance, a feature partially imparted by tumor cell stemness. Recent work shows that homozygous MTAP deletion, a genetic alteration occurring in about half of all GBMs, promotes stemness in GBM cells. Exploiting MTAP loss-conferred deficiency in purine salvage, we demonstrate that purine blockade via treatment with L-Alanosine (ALA), an inhibitor of de novo purine synthesis, attenuates stemness of MTAP-deficient GBM cells. This ALA-induced reduction in stemness is mediated in part by compromised mitochondrial function, highlighted by ALA-induced elimination of mitochondrial spare respiratory capacity. Notably, these effects of ALA are apparent even when the treatment was transient and with a low dose. Finally, in agreement with diminished stemness and compromised mitochondrial function, we show that ALA sensitizes GBM cells to temozolomide (TMZ) in vitro and in an orthotopic GBM model. Collectively, these results identify purine supply as an essential component in maintaining mitochondrial function in GBM cells and highlight a critical role of mitochondrial function in sustaining GBM stemness. We propose that purine synthesis inhibition can be beneficial in combination with the standard of care for MTAP-deficient GBMs, and that it may be feasible to achieve this benefit without inflicting major toxicity.

MTAP loss correlates with an immunosuppressive profile in GBM and its substrate MTA stimulates alternative macrophage polarization.

Glioblastoma (GBM) is a lethal brain cancer known for its potent immunosuppressive effects. Loss of Methylthioadenosine Phosphorylase (MTAP) expression, via gene deletion or epigenetic silencing, is one of the most common alterations in GBM. Here we show that MTAP loss in GBM cells is correlated with differential expression of immune regulatory genes. In silico analysis of gene expression profiles in GBM samples revealed that low MTAP expression is correlated with an increased proportion of M2 macrophages. Using in vitro macrophage models, we found that methylthioadenosine (MTA), the metabolite that accumulates as a result of MTAP loss in GBM cells, promotes the immunosuppressive alternative activation (M2) of macrophages. We show that this effect of MTA on macrophages is independent of IL4/IL3 signaling, is mediated by the adenosine A2B receptor, and can be pharmacologically reversed. This study suggests that MTAP loss in GBM cells may contribute to the immunosuppressive tumor microenvironment, and that MTAP status should be considered for characterizing GBM immune states and devising immunotherapy-based approaches for treating MTAP-null GBM.

Angiogenin and MMP-2 as potential biomarkers in the differential diagnosis of gestational trophoblastic diseases.

BACKGROUND: Gestational trophoblastic diseases (GTDs) are characterized by vascular abnormalities of the trophoblast, but their pathogenesis is unknown. Angiogenin (ANG) and matrix metalloproteinase (MMP)-2, which are molecules implicated in the angiogenic process, may play some role in this process. MATERIAL AND METHODS: We determined ANG and MMP-2 in the placental tissues of 26 patients who had a benign mole (BM), 12 patients with gestational trophoblast neoplasia (GTN) (1 invasive hydatidiform mole, 10 choriocarcinomas, and 1 placental-site trophoblastic tumor), and 28 normal chorionic villi (NCV) subjects using immunohistochemistry staining. We obtained the serum samples from 20 patients with GTDs and 20 early pregnant women and evaluated them by the enzyme linked immunosorbent assay. RESULTS: ANG expression in GTN (66.7%) and BM (100%) samples were both significantly higher (strong/intermediate staining) than in NCV (60.7%) samples (P < .001). Similarly, the immunoreactivities of MMP-2 in the GTN (66.7%) and BM (80.8%) samples were significantly elevated compared to that of the NCV (57.1%) samples (P < .001). The levels of ANG and MMP-2 in the maternal serum of the GTN group were both significantly higher than those of the control group (P < .001). ANG and MMP-2 expressions were associated with gestation age, clinical stage, and FIGO stage. A positive correlation between ANG and MMP-2 expression was observed (rs = 0.725; P < .01). CONCLUSION: ANG and MMP-2 levels were significantly elevated in the placental tissues and maternal serum from patients with GTDs. Further studies with more patients may clarify the vascular abnormalities in GTDs and determine potential biomarkers in the differential diagnosis of GTDs.

Targeting glutamine metabolism network for the treatment of therapy-resistant prostate cancer.

Advanced and aggressive prostate cancer (PCa) depends on glutamine for survival and proliferation. We have previously shown that inhibition of glutaminase 1, which catalyzes the rate-limiting step of glutamine catabolism, achieves significant therapeutic effect; however, therapy resistance is inevitable. Here we report that while the glutamine carbon is critical to PCa survival, a parallel pathway of glutamine nitrogen catabolism that actively contributes to pyrimidine assembly is equally important for PCa cells. Importantly, we demonstrate a reciprocal feedback mechanism between glutamine carbon and nitrogen pathways which leads to therapy resistance when one of the two pathways is inhibited. Combination treatment to inhibit both pathways simultaneously yields better clinical outcome for advanced PCa patients.

Yunnan Black Tea Flavonoids Can Improve Cognitive Dysfunction in Septic Mice by Activating SIRT1.

This study explored the effect and mechanism of Yunnan black tea flavonoids (YBTF) on cognitive dysfunction in septic mice. The mice were induced sepsis, the serum was determined using kits, and the tissue was determined by qPCR assay. The Yunnan black tea flavonoids were checked using HPLC. The test results showed that compared with the model group, YBTF could increase the survival rate of the mice; meanwhile, YBTF could also increase the total distance travelled, number of stands, and number of groomings, as well as the number of times crossing the area in the target quadrant. Detection of nerve cells showed that YBTF could reduce the rate of nerve cell apoptosis caused by sepsis. YBTF also reduced the levels of tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), interleukin-1 beta (IL-1ß), and malondialdehyde (MDA) in the hippocampus of septic mice and increased the activity of superoxide dismutase (SOD) and catalase (CAT) enzymes. YBTF could also upregulate the mRNA expression of SOD1, SOD2, CAT, and forkhead box O1 (FOXO1) and downregulate the mRNA expression of TNF-α, IL-1ß, nuclear factor kappa-B (NF-κB), p53, and SIRT1 in the hippocampus of septic mice. The animal experiment results showed that YBTF could improve the cognitive dysfunction of septic mice. The effect of YBTF was weaker than that of dexamethasone, but it could enhance the improvement effect when used in conjunction with dexamethasone. The component analysis results showed that YBTF contained 9 compounds, including catechin, gallocatechin gallate, rutin, hyperoside, epicatechin gallate, dihydroquercetin, quercetin, myricetin, and sulphuretin. From these results, YBTF could activate SIRT1 through its active compound components to improve the cognitive dysfunction of septic mice.

TP53 wild-type/PPM1D mutant diffuse intrinsic pontine gliomas are sensitive to a MDM2 antagonist.

Diffuse intrinsic pontine gliomas (DIPGs) are high-grade tumors of the brainstem that often occur in children, with a median overall survival of less than one year. Given the fact that DIPGs are resistant to chemotherapy and are not amenable to surgical resection, it is imperative to develop new therapeutic strategies for this deadly disease. The p53 pathway is dysregulated by TP53 (~ 60%) or PPM1D gain-of-function mutations (~ 30%) in DIPG cases. PPM1D gain-of-function mutations suppress p53 activity and result in DIPG tumorigenesis. While MDM2 is a major negative regulator of p53, the efficacy of MDM2 inhibitor has not been tested in DIPG preclinical models. In this study, we performed a comprehensive validation of MDM2 inhibitor RG7388 in patient-derived DIPG cell lines established from both TP53 wild-type/PPM1D-mutant and TP53 mutant/PPM1D wild-type tumors, as well in TP53 knockout isogenic DIPG cell line models. RG7388 selectively inhibited the proliferation of the TP53 wild-type/PPM1D mutant DIPG cell lines in a dose- and time-dependent manner. The anti-proliferative effects were p53-dependent. RNA-Seq data showed that differential gene expression induced by RG7388 treatment was enriched in the p53 pathways. RG7388 reactivated the p53 pathway and induced apoptosis as well as G1 arrest. In vivo, RG7388 was able to reach the brainstem and exerted therapeutic efficacy in an orthotopic DIPG xenograft model. Hence, this study demonstrates the pre-clinical efficacy potential of RG7388 in the TP53 wild-type/PPM1D mutant DIPG subgroup and may provide critical insight on the design of future clinical trials applying this drug in DIPG patients.