Phosphatidylinositol 3-kinase Pathway Research Articles

Sodium-glucose cotransporter-2 inhibitors for hypergycemia in phosphoinositide 3-kinase pathway inhibition.

Phosphoinositide 3-kinase (PI3K) inhibition is used for the treatment of certain cancers, but can cause profound hyperglycemia and insulin resistance, for which sodium-glucose cotransporter-2 (SGLT2) inhibitors have been proposed as a preferred therapy. The objective of this research is to assess the effectiveness and safety of SGLT2 inhibitors for hyperglycemia in PI3K inhibition. We conducted a single-center retrospective review of adults initiating the PI3K inhibitor alpelisib. Exposure to different antidiabetic drugs and adverse events including diabetic ketoacidosis (DKA) were assessed through chart review. Plasma and point-of-care blood glucoses were extracted from the electronic medical record. Change in serum glucose and the rate of DKA on SGLT2 inhibitor versus other antidiabetic drugs were examined as co-primary outcomes. We identified 103 patients meeting eligibility criteria with median follow-up of 92days after starting alpelisib. When SGLT2 inhibitors were used to treat hyperglycemia, they were associated with a decrease in mean random glucose by -46mg/dL (95% CI -77 to -15) in adjusted linear modeling. Five cases of DKA were identified, two occurring in patients on alpelisib plus SGLT2 inhibitor. Estimated incidence of DKA was: alpelisib plus SGLT2 inhibitor, 48 DKA cases per 100 patient-years (95% CI 6, 171); alpelisib with non-SGLT2 inhibitor antidiabetic drugs, 15 (95% CI 2, 53); alpelisib only, 4 (95% CI 0.1, 22). SGLT2 inhibitors are effective treatments for hyperglycemia in the setting of PI3K inhibition.

FBXO21 mediated degradation of p85α regulates proliferation and survival of acute myeloid leukemia

Acute myeloid leukemia (AML) is a heterogeneous disease characterized by clonal expansion of myeloid blasts in the bone marrow (BM). Despite advances in therapy, the prognosis for AML patients remains poor, and there is a need to identify novel molecular pathways regulating tumor cell survival and proliferation. F-box ubiquitin E3 ligase, FBXO21, has low expression in AML, but expression correlates with survival in AML patients and patients with higher expression have poorer outcomes. Silencing FBXO21 in human-derived AML cell lines and primary patient samples leads to differentiation, inhibition of tumor progression, and sensitization to chemotherapy agents. Additionally, knockdown of FBXO21 leads to up-regulation of cytokine signaling pathways. Through a mass spectrometry-based proteomic analysis of FBXO21 in AML, we identified that FBXO21 ubiquitylates p85α, a regulatory subunit of the phosphoinositide 3-kinase (PI3K) pathway, for degradation resulting in decreased PI3K signaling, dimerization of free p85α and ERK activation. These findings reveal the ubiquitin E3 ligase, FBXO21, plays a critical role in regulating AML pathogenesis, specifically through alterations in PI3K via regulation of p85α protein stability.

P10.19.A COMBINED TREATMENT USING MAPK AND PI3K SIGNALING PATHWAYS INHIBITORS EFFECTIVELY REDUCES GROWTH OF MELANOMA BRAIN METASTASIS CELLS IN VITRO

Abstract BACKGROUND Brain metastasis is associated with poor prognosis, and median survival time from the time of diagnosis for cancer patients with brain metastases is only a few months. Brain metastases are estimated to occur in 20-40% of cancer patients. All solid tumors may spread to the brain, but the risk of developing brain metastases is higher in breast cancer, lung cancer and melanoma. Standard treatment of melanoma brain metastases (MBM) includes radiation therapy and surgery. In melanoma, the mitogen activated protein kinase (MAPK) and the phosphoinositide 3-kinase (PI3K) signaling pathways are often activated by genetic alterations. An activation mutation (BRAFV600E) in the MAPK pathway is commonly found in melanoma refractory to therapy. This mutation leads to increased proliferation and tumor cell survival. Previous studies show that BRAF-inhibitors block the increase in cell proliferation induced by the activated BRAF gene, however, many of the patients develop drug resistance within a few months. Objectives: In this project, we aimed to show that drugs targeting BRAF (vemurafenib) and PI3K pathways (buparlisib) used in combination effectively inhibit growth of treatment resistant MBM cells. MATERIAL AND METHODS Our research lab has previously developed human MBM cell lines, which have been made resistant towards vemurafenib treatment. To study the potential antitumoral effects of combined treatment we used these cells in several in vitro assays such as cell viability and proliferation, cell migration, colony forming assay, spheroid growth, apoptosis by IncuCyte and western blot to confirm effects on signaling pathways. RESULTS We found that combination treatment reduced the viability of resistant MBM cells, as well as their ability to proliferate. Further, the cells showed less ability to migrate after therapy. Altered cell morphology was also seen. Initial results show effects on colony formation and increased apoptosis after combined treatment. CONCLUSION A combination of MAPK and PI3K inhibitors effectively inhibits growth and viability of treatment resistant human MBM cells, compared to single drug treatment alone. The results support clinical use of combined therapy for melanoma patients with metastatic disease.

Myricetin inhibits Micropterus salmoides rhabdovirus infection by damping inflammation through the NF-κB pathway

Largemouth bass have been plagued by diseases elicited by bacteria, parasites, and viruses, particularly viral diseases, which have caused significant economic losses in recent years. And there is a lack of approved antiviral drugs in the aquaculture industry. Micropterus salmoides Rhabdovirus (MSRV) is the causative agent of Micropterus salmoides Rhabdovirus disease, which mostly affects largemouth bass fry. By using experimental infection, transcriptome sequencing, and inhibitor treatment, the inflammatory response mediated by mitogen-activated protein kinase (MAPK), phosphatidylinositol3-kinase (PI3K), and nuclear factor kappa-B (NF-κB) pathways was verified as one of the pathological processes of MSRV infection. Further, Myricetin, arteannuin, naringenin, ursolic acid, andrographolide, and resveratrol were selected to evolute their anti-inflammatory activity based on the promoter of NF-κB and found myricetin, naringenin, andrographolide, and resveratrol significantly inhibited NF-κB 1 promoter activity. Myricetin was found to damp MSRV caused-inflammation and inhibit MSRV infection by suppressing NF-κB signaling. Besides, myricetin improves the survival rate of largemouth bass after MSRV infection. In conclusion, myricetin presented anti-MSRV activity by targeting the NF-κB inflammatory pathway and can be further developed as a potential preventative drug of rhabdovirus disease.

Clinical and mutational profile of AT-rich interaction domain 1A-mutated cancers.

AT-rich interaction domain 1A (ARID1A) encodes a key component of the SWItch/Sucrose Non-Fermentable (SWI/SNF) chromatin remodeling complex that participates in gene expression. ARID1A alterations are quite common among cancer patients, although their role remains debated. The aim of this article was to study ARID1A-mutated cancer patients. Molecular and clinical data of cancer patients evaluated at Phase 1 Unit of Fondazione Policlinico Universitario A. Gemelli IRCCS were collected. Molecular analyses were performed using FoundationOne® CDx (Foundation Medicine Inc., Cambridge, MA, United States). Cancer patients with at least one molecular alteration in ARID1A gene were identified as ARID1A+. Among the 270 patients undergoing molecular analysis, we found 25 (9%) with at least one pathogenic alteration in ARID1A. The vast majority of these patients were female (84%). The median age at diagnosis was 59; most of the cancers (15, 60%) were gynecological (especially endometrioid endometrial cancers and clear cell ovarian cancers), diagnosed at an early stage. Frameshift alterations in ARID1A were the most common (19/31, 61%) alterations. The median number of mutations in ARID1A+ population was higher compared to ARID1A- population (6 vs. 4), as well as tumor mutational burden (TMB) [20 mutations/megabase (mut/Mb) vs. 1.26 mut/Mb]. Phosphatidylinositol-4,5-bisphosphate 3-kinase catalytic subunit alpha (PIK3CA), phosphatase and tensin homolog (PTEN), catenin beta 1 (CTNNB1), and lysine methyltransferase 2D (MLL2) mutations were enriched in ARID1A+ population. In this cohort, ARID1A did not display any relation with response to platinum chemotherapy. Cancers with double alterations in ARID1A (ARID1A2+) were all gynecological cancers (83% endometrioid endometrial cancers). This analysis provides clinical and molecular details about the phenotypes of ARID1A+ cancers, in particular the subgroup of gynecologic cancers. The high frequency of concurrent mutations in the phosphoinositide 3-kinase (PI3K) pathway among endometrioid endometrial cancers may support the proposal of a new treatment strategy based on the combination of ataxia telangiectasia and Rad3-related (ATR) inhibitor and PIK3CA inhibitor.

Activation of the EGFR/PI3K/AKT pathway limits the efficacy of trametinib treatment in head and neck cancer.

Blocking the mitogen-activated protein kinase (MAPK) pathway with the MEK1/2 inhibitor trametinib has produced promising results in patients with head and neck squamous cell carcinoma (HNSCC). In the current study, we showed that trametinib treatment leads to overexpression and activation of the epidermal growth factor receptor (EGFR) in HNSCC cell lines and patient-derived xenografts. Knockdown of EGFR improved trametinib treatment efficacy both in vitro and in vivo. Mechanistically, we demonstrated that trametinib-induced EGFR overexpression hyperactivates the phosphatidylinositol 3-kinase (PI3K)/AKT pathway. In vitro, blocking the PI3K pathway with GDC-0941 (pictilisib), or BYL719 (alpelisib), prevented AKT pathway hyperactivation and enhanced the efficacy of trametinib in a synergistic manner. In vivo, a combination of trametinib and BYL719 showed superior antitumor efficacy vs. the single agents, leading to tumor growth arrest. We confirmed our findings in a syngeneic murine head and neck cancer cell line in vitro and in vivo. Taken together, our findings show that trametinib treatment induces hyperactivation of EGFR/PI3K/AKT; thus, blocking of the EGFR/PI3K pathway is required to improve trametinib efficacy in HNSCC.

Vemurafenib activates the sonic hedgehog pathway and promotes thyroid cancer stem cell self-renewal.

B-Raf kinase inhibitors such as vemurafenib (PLX4032) and dabrafenib have limited therapeutic efficacy on BRAF-mutated thyroid cancer. Cancer stem cells (CSCs) play important roles in tumor recurrence, drug resistance, and metastasis. Whether CSCs play a role in dampening the antitumor activity of B-Raf kinase inhibitors remains unknown. Here, we report that vemurafenib (PLX4032) induced the expression of several stemness-related genes including Gli1, Snail, BMI1, and SOX2 in two anaplastic thyroid cancer cell lines, SW1736 and 8505C, but decreased the expression of these genes in A375 cells, a human melanoma cell line. PLX4032 promoted thyroid cancer stem cell self-renewal, as evidenced by increased numbers of aldehyde dehydrogenase-positive cells and thyrospheres. Mechanistically, PLX4032 activates the PI-3 and mitogen-activated protein kinase pathways through HER3 to cross-activate Gli1, a transcription factor of the sonic hedgehog (Shh) pathway. GANT61, a specific inhibitor of Gli1, blocked the expression of the stemness-related genes in PLX4032-treated thyroid cancer cells in vitro and in vivo in two thyroid cancer xenograft models. GANT61 treatment alone weakly inhibited SW1736 tumor growth but enhanced the antitumor activity of PLX4032 when used in combination. Our study provides mechanistic insights into how thyroid cancer poorly responds to B-Raf kinase inhibitors and suggests that targeting B-Raf and the Shh pathway in combination may overcome thyroid cancer drug resistance.

IDENTIFICATION AND SCREENING OF PLANT-BASED POTENT INHIBITORS AGAINST nSH2 DOMAIN OF PI3K OF BREAST CANCER USING DOCKING AND SIMULATION STUDIES

Breast cancer (BC) is a critical health issue that affects countless women, and it is the second leading reason of death worldwide. The phosphatidylinositol 3 kinases (PI3Ks) constitute a group of lipid kinases that play a role in tumorigenesis, development, migration, infiltration, programmed cell death, glycogen synthesis, DNA correction and viability by the PI3K/Akt cascade. The PI3K pathway has been linked to a variety of malignancies and increases the activation rate of cancer. Here, focus was given to the study of PI3K pathway involved in BC and emphasis was given on a particular nSH2 domain that resides in the regulatory subunit of PI3K to find a potent inhibitor. A detailed pathway and interaction study was performed from KEGG pathway database and from the cystoscope. A total list of 60 compounds, comprises phytochemicals, and herbal compounds were screened based on structural similarity and eight FDA-approved drugs were considered. The docking analysis was carried over through the AutoDock software and Ligplot analysis was performed to investigate the interaction between the nSH2 domain and the potent inhibitors. To ensure the complex stability, 20 ns of simulation run was also performed on the best complexes using GROMACS. From this study, it can be concluded that Evodia fruit has the maximum stability in the catalytic region among all the listed inhibitors against the target proteins and can act as a potent inhibitor among the others.

Inhibitors of the PI3K/AKT/mTOR pathway in human malignancies; trend of current clinical trials.

The phosphoinositide 3-kinase (PI3K)/AKT/mammalian target of rapamycin (mTOR) signaling pathway regulates proliferation, survival and metabolism, and its dysregulation is one of the most frequent oncogenic events across human malignancies. Over the last two decades, there has been significant focus on the clinical development of PI3K pathway inhibitors. More than 40 different inhibitors of this axis have reached various stages of clinical trials, but only a fewof themhave been approved by the Food and Drug Administration (FDA) for cancer treatment. These clinical results, however, could be improved given the importance of PI3K signaling in cancer and its role in linking cancer growth with metabolism. In this systematic review, after a glance at PI3K/AKT/mTOR pathway and its different inhibitors, we retrieved registered clinical trials evaluating the efficacy and safety of PI3K/AKT/mTOR inhibitors on Clinicaltrials.gov. Following the extraction of the data, finally we analyzed 2250 included studies in multiple steps, beginning with an overview and moving on to the details about type of malignancies, inhibitors, and treatment strategies. We also took a closer look at more than 100 phase III-IV clinical trials to pinpoint promising therapies, hoping that presenting a comprehensive picture of current clinical trials casts a flash of light on what remains to be done in future clinical trials of PI3K/AKT/mTOR inhibitors in human malignancies.

Regulation of Ras Signaling by S-Nitrosylation.

Ras are a family of small GTPases that function as signal transduction mediators and are involved in cell proliferation, migration, differentiation and survival. The significance of Ras is further evidenced by the fact that Ras genes are among the most mutated oncogenes in different types of cancers. After translation, Ras proteins can be targets of post-translational modifications (PTM), which can alter the intracellular dynamics of the protein. In this review, we will focus on how S-nitrosylation of Ras affects the way these proteins interact with membranes, its cellular localization, and its activity. S-Nitrosylation occurs when a nitrosyl moiety of nitric oxide (NO) is covalently attached to a thiol group of a cysteine residue in a target protein. In Ras, the conserved Cys118 is the most surface-exposed Cys and the preferable residue for NO action, leading to the initiation of transduction events. Ras transduces the mitogen-activated protein kinases (MAPK), the phosphoinositide-3 kinase (PI3K) and the RalGEF cellular pathways. S-Nitrosylation of elements of the RalGEF cascade remains to be identified. On the contrary, it is well established that several components of the MAPK and PI3K pathways, as well as different proteins associated with these cascades, can be modified by S-nitrosylation. Overall, this review presents a better understanding of Ras S-nitrosylation, increasing the knowledge on the dynamics of these proteins in the presence of NO and the underlying implications in cellular signaling.

APP96-110 Elicits Neuroprotective Effects Following Ischemic Insult in Animal Models.

Competitive amyloidogenic pathways play an important role in many neurological diseases such as the onset of various degenerative diseases and ischemic stroke. Overexpression of amyloid precursor protein (APP) and amyloid-beta is modulated via the amyloidogenic pathway, which plays a crucial role in neuroinflammation. During ischemic conditions, the activity of the anti-inflammatory non-amyloidogenic pathway decreases, thus increasing the activity of amyloidogenic pathway. The soluble alpha form of APP (sAPPα), formed via the non-amyloidogenic pathway, exhibits neuroprotective effects against neurological diseases. sAPPα is thought to have a modulatory effect on several cell survival pathways, including its ability to inhibit the phosphoinositide 3-kinases (PI3K) pathway, thereby inhibiting the inflammatory response. The APP derivative, APP96-110, could act as a functional substitute for native sAPPα. Herein, we investigated whether APP96-110 has neuroprotective effects against neuroinflammation and damage following cerebral ischemic stroke. Treatment with diluted APP96-110 (0.005mg/kg) in mice after 30min of transient middle cerebral artery occlusion (tMCAO) showed improved motor function and reduced expression of the inflammatory marker CD86. APP96-110 decreased the infarct size and induced an anti-inflammatory response by inhibiting the PI3K pathway. These results suggest that the treatment of APP96-110 is efficacious in reducing neuroinflammation and infarct size in ischemic stroke.

Unraveling the molecular landscape: a comparative analysis of PI3K and MAPK signaling pathways in plasmablastic lymphoma and diffuse large B-cell lymphoma with therapeutic implications

Plasmablastic lymphoma (PBL) is a rare and aggressive subtype of non-Hodgkin lymphoma that shares features with diffuse large B-cell lymphoma (DLBCL). While significant progress has been made in treating DLBCL, the prognosis for PBL remains poor, highlighting the need to identify new therapeutic targets. Using RNA expression analysis, we compared the expression of genes involved in the phosphatidylinositol-3-kinase (PI3K) and mitogen-activated protein kinase (MAPK) signaling pathways between PBL and DLBCL. We used critical PI3K (n=201) and MAPK (n=57) signaling probe sets to achieve this objective. Our results demonstrate unique molecular mechanisms underlying PBL pathogenesis compared to DLBCL, particularly within the PI3K and MAPK signaling pathways. We found that elevated STAT3 expression in PBL correlates with hyperactive MAPK and PI3K pathways, unlike DLBCL. Additionally, the hyperactivation of the PI3K signaling axis in PBL is unrelated to B-cell receptoror phosphatase and tensin homologactivity, indicating a distinct mechanism compared to DLBCL. Furthermore, we observed unique activation patterns in MAPK pathways between PBL and DLBCL, with PBL exhibiting high expression of the neurotrophic tyrosine kinase receptor (NTKR) family, specifically NTRK1 and NTRK2 genes, which have therapeutic potential. We also found that neither human immunodeficiency virus nor Epstein-Barr virus infection influences gene expression profiles linked to PI3K and MAPK signaling in PBL. These findings could lead to adapting targeted therapies developed for DLBCL to address the specific needs of PBL patients better and contribute to developing novel, targeted therapeutic strategies to improve patient outcomes.

Social memory in female mice is rapidly modulated by 17β-estradiol through ERK and Akt modulation of synapse formation

Social memory is essential to the functioning of a social animal within a group. Estrogens can affect social memory too quickly for classical genomic mechanisms. Previously, 17β-estradiol (E2) rapidly facilitated short-term social memory and increased nascent synapse formation, these synapses being potentiated following neuronal activity. However, what mechanisms underlie and coordinate the rapid facilitation of social memory and synaptogenesis are unclear. Here, the necessity of extracellular signal-regulated kinase (ERK) and phosphoinositide 3-kinase (PI3K) signaling for rapid facilitation of short-term social memory and synaptogenesis was tested. Mice performed a short-term social memory task or were used as task-naïve controls. ERK and PI3K pathway inhibitors were infused intradorsal hippocampally 5 min before E2 infusion. Forty minutes following intrahippocampal E2 or vehicle administration, tissues were collected for quantification of glutamatergic synapse number in the CA1. Dorsal hippocampal E2 rapid facilitation of short-term social memory depended upon ERK and PI3K pathways. E2 increased glutamatergic synapse number (bassoon puncta positive for GluA1) in task-performing mice but decreased synapse number in task-naïve mice. Critically, ERK signaling was required for synapse formation/elimination in task-performing and task-naïve mice, whereas PI3K inhibition blocked synapse formation only in task-performing mice. While ERK and PI3K are both required for E2 facilitation of short-term social memory and synapse formation, only ERK is required for synapse elimination. This demonstrates previously unknown, bidirectional, rapid actions of E2 on brain and behavior and underscores the importance of estrogen signaling in the brain to social behavior.

Interferon-γ regulates immunosuppression in septic mice by promoting the Warburg effect through the PI3K/AKT/mTOR pathway

BackgroundThe main cause of high mortality from sepsis is that immunosuppression leads to life-threatening organ dysfunction, and reversing immunosuppression is key to sepsis treatment. Interferon γ (IFNγ) is a potential therapy for immunosuppression of sepsis, promoting glycolysis to restore metabolic defects in monocytes, but the mechanism of treatment is unclear.MethodsTo explore the immunotherapeutic mechanism of IFNγ, this study linked the Warburg effect (aerobic glycolysis) to immunotherapy for sepsis and used cecal ligation perforation (CLP) and lipopolysaccharide (LPS) to stimulate dendritic cells (DC) to establish in vivo and in vitro sepsis models, Warburg effect inhibitors (2-DG) and PI3K pathway inhibitors (LY294002) were used to explore the mechanism by which IFNγ regulates immunosuppression in mice with sepsis through the Warburg effect.ResultsIFNγ markedly inhibited the reduction in cytokine secretion from lipopolysaccharide (LPS)-stimulated splenocytes. IFNγ-treated mice had significantly increased the percentages of positive costimulatory receptor CD86 on Dendritic cells expressing and expression of splenic HLA-DR. IFNγ markedly reduced DC-cell apoptosis by upregulating the expression of Bcl-2 and downregulating the expression of Bax. CLP-induced formation of regulatory T cells in the spleen was abolished in IFNγ -treated mice. IFNγ treatment reduced the expression of autophagosomes in DC cells. IFNγ significant reduce the expression of Warburg effector-related proteins PDH, LDH, Glut1, and Glut4, and promote glucose consumption, lactic acid, and intracellular ATP production. After the use of 2-DG to suppress the Warburg effect, the therapeutic effect of IFNγ was suppressed, demonstrating that IFNγ reverses immunosuppression by promoting the Warburg effect. Moreover, IFNγ increased the expression of phosphoinositide 3-kinases (PI3K), protein kinase B (Akt), rapamycin target protein (mTOR), hypoxia-inducible factor-1 (HIF-1α), pyruvate dehydrogenase kinase (PDK1) protein, the use of 2-DG and LY294002 can inhibit the expression of the above proteins, LY294002 also inhibits the therapeutic effect of IFNγ.ConclusionsIt was finally proved that IFNγ promoted the Warburg effect through the PI3K/Akt/mTOR pathway to reverse the immunosuppression caused by sepsis. This study elucidates the potential mechanism of the immunotherapeutic effect of IFNγ in sepsis, providing a new target for the treatment of sepsis.

Combined Aurora Kinase A and CHK1 Inhibition Enhances Radiosensitivity of Triple-Negative Breast Cancer Through Induction of Apoptosis and Mitotic Catastrophe Associated With Excessive DNA Damage

There is an urgent need for biomarkers and new actionable targets to improve radiosensitivity of triple-negative breast cancer (TNBC) tumors. We characterized the radiosensitizing effects and underlying mechanisms of combined Aurora kinase A (AURKA) and CHK1 inhibition in TNBC. Different TNBC cell lines were treated with AURKA inhibitor (AURKAi, MLN8237) and CHK1 inhibitor (CHK1i, MK8776). Cell responses to irradiation (IR) were then evaluated. Cell apoptosis, DNA damage, cell cycle distribution, and mitogen-activated protein kinase (MAPK)/extracellular signal-regulated kinase (ERK) and Phosphoinositide 3-Kinase (PI3K) pathways were evaluated in vitro. Transcriptomic analysis was performed to facilitate the identification of potential biomarkers. Xenograft and immunohistochemistry were carried out to investigate the radiosensitizing effects of dual inhibition in vivo. Finally, the prognostic effect of CHEK1/AURKA in TNBC samples in the The Cancer Genome Atlas (TCGA) database and our center were analyzed. AURKAi (MLN8237) induced overexpression of phospho-CHK1 in TNBC cells. The addition of MK8776 (CHK1i) to MLN8237 greatly reduced cell viability and increased radiosensitivity compared with either the control or MLN8237 alone in vitro. Mechanistically, dual inhibition resulted in inducing excessive DNA damage by prompting G2/M transition to cells with defective spindles, leading to mitotic catastrophe and induction of apoptosis after IR. We also observed that dual inhibition suppressed the phosphorylation of ERK, while activation of ERK with its agonist or overexpression of active ERK1/2 allele could attenuate the apoptosis induced by dual inhibition with IR. Additionally, dual inhibition of AURKA and CHK1 synergistically enhanced radiosensitivity in MDA-MB-231 xenografts. Moreover, we detected that both CHEK1 and AURKA were overexpressed in patients with TNBC and negatively correlated with patient survival. Our findings suggested that AURKAi in combination with CHK1i enhanced TNBC radiosensitivity in preclinical models, potentially providing a novel strategy of precision treatment for patients with TNBC.

Preclinical and clinical evaluation of Buparlisib (BKM120) in recurrent/refractory Central Nervous System Lymphoma

Central Nervous System (CNS) Lymphomas are aggressive brain tumors with limited treatment options. Targeting the phosphoinositide 3-kinase (PI3K) pathway yields promising responses across B-cell malignancies, but its therapeutic potential in CNS lymphomas remains unexplored. We present pre-clinical and clinical data on the pan-PI3K inhibitor Buparlisib in CNS lymphomas. In a primary CNS lymphoma-patient-derived cell line, we define the EC50. Four patients with recurrent CNS lymphoma were enrolled in a prospective trial. We evaluated Buparlisib plasma and cerebrospinal fluid pharmacokinetics, clinical outcomes, and adverse events. Treatment was well tolerated. Common toxicities include hyperglycemia, thrombocytopenia, and lymphopenia. The presence of Buparlisib in plasma and CSF was confirmed 2h post-treatment with a median CSF concentration below the EC50 defined in the cell line All four patients were evaluated for response and the median time to progression was 39 days. Buparlisib monotherapy did not lead to meaningful responses and the trial was prematurely stopped.Clinical Trial Registration: NCT02301364

Stimulation of ectopically expressed muscarinic receptors induces IFN-γ but suppresses IL-2 production by inhibiting activation of pAKT pathways in primary T cells

T cell antigen receptor stimulation induces tyrosine phosphorylation of downstream signaling molecules and the phosphatidylinositol, Ras, MAPK, and PI3 kinase pathways, leading to T cell activation. Previously, we reported that the G-protein-coupled human muscarinic receptor could bypass tyrosine kinases to activate the phosphatidylinositol pathway and induce interleukin-2 production in Jurkat leukemic T cells. Here, we demonstrate that stimulating G-protein-coupled muscarinic receptors (M1 and synthetic hM3Dq) can activate primary mouse T cells if PLCβ1 is coexpressed. Resting peripheral hM3Dq+PLCβ1 (hM3Dq/β1) T cells did not respond to clozapine, an hM3Dq agonist, unless they were preactivated by TCR and CD28 stimulation which increased hM3Dq and PLCβ1 expression. This permitted large calcium and phosphorylated ERK responses to clozapine. Clozapine treatment induced high IFN-γ, CD69, and CD25 expression, but surprisingly did not induce substantial IL-2 in hM3Dq/β1 T cells. Importantly, costimulation of both muscarinic receptors plus the TCR even led to reduced IL-2 expression, suggesting a selective inhibitory effect of muscarinic receptor costimulation. Stimulation of muscarinic receptors induced strong nuclear translocation of NFAT and NFκB and activated AP-1. However, stimulation of hM3Dq led to reduced IL-2 mRNA stability which correlated with an effect on the IL-2 3'UTR activity. Interestingly, stimulation of hM3Dq resulted in reduced pAKT and its downstream pathway. This may explain the inhibitory impact on IL-2 production in hM3Dq/β1T cells. Moreover, an inhibitor of PI3K reduced IL-2 production in TCR-stimulated hM3Dq/β1 CD4 T cells, suggesting that activating the pAKT pathway is critical for IL-2 production in T cells.

Inhibition of the NLRP3 Inflammasome Activation/Assembly through the Activation of the PI3K Pathway by Naloxone Protects Neural Stem Cells from Ischemic Condition.

Naloxone is a well-known opioid antagonist and has been suggested to have neuroprotective effects in cerebral ischemia. We investigated whether naloxone exhibits anti-inflammatory and neuroprotective effects in neural stem cells (NSCs) injured by oxygen-glucose deprivation (OGD), whether it affects the NOD-like receptor protein 3 (NLRP3) inflammasome activation/assembly, and whether the role of the phosphatidylinositol 3-kinase (PI3K) pathway is important in the control of NLRP3 inflammasome activation/assembly by naloxone. Primary cultured NSCs were subjected to OGD and treated with different concentrations of naloxone. Cell viability, proliferation, and the intracellular signaling proteins associated with the PI3K pathway and NLRP3 inflammasome activation/assembly were evaluated in OGD-injured NSCs. OGD significantly reduced survival, proliferation, and migration and increased apoptosis of NSCs. However, treatment with naloxone significantly restored survival, proliferation, and migration and decreased apoptosis of NSCs. Moreover, OGD markedly increased NLRP3 inflammasome activation/assembly and cleaved caspase-1 and interleukin-1β levels in NSCs, but naloxone significantly attenuated these effects. These neuroprotective and anti-inflammatory effects of naloxone were eliminated when cells were treated with PI3K inhibitors. Our results suggest that NLRP3 inflammasome is a potential therapeutic target and that naloxone reduces ischemic injury in NSCs by inhibiting NLRP3 inflammasome activation/assembly mediated by the activation of the PI3K signaling pathway.

PTPN22 Activates PI3K Pathway via 14-3-3τ in T Cells.

The protein tyrosine phosphatase PTPN22 inhibits T cell activation by dephosphorylating some essential proteins in the TCR-mediated signaling pathway, and its negative regulatory function protects organisms from autoimmune disease. 14-3-3τ is an adaptor protein that regulates target protein function through its intracellular localization. In this study, we identified that PTPN22 binds to 14-3-3τ via the PTPN22-Ser640 phosphorylation side. PTPN22 binding to 14-3-3τ resulted in a 14-3-3τ-Tyr179 dephosphorylation, and reduced the association between 14-3-3τ and Shc, which competitively increased 14-3-3ζ binding to Shc and activated PI3K by bringing it to membrane. In addition, PTPN22 decreases the tyrosine phosphorylation of p110 to activate PI3K. These two pathways cooperatively affect PI3K activity and the expression of PI3K downstream proteins, such as phosphorylated Akt, mTOR and FoxO1, which inhibited the expression of some proinflammatory factors such as IL-1β, IL-2, IL-6, IFN-γ and TNF-α. Our research provides a preliminary theory for PTPN22 regulating T cell activation, development and immune response via the PI3K/Akt/mTOR pathway and brings a new clue to clarify the functions of PTPN22 in autoimmune diseases.

Effectiveness of Polyphenols on Perinatal Brain Damage: A Systematic Review of Preclinical Studies.

Polyphenol supplementation during early life has been associated with a reduction of oxidative stress and neuroinflammation in diseases caused by oxygen deprivation, including cerebral palsy, hydrocephaly, blindness, and deafness. Evidence has shown that perinatal polyphenols supplementation may alleviate brain injury in embryonic, fetal, neonatal, and offspring subjects, highlighting its role in modulating adaptative responses involving phenotypical plasticity. Therefore, it is reasonable to infer that the administration of polyphenols during the early life period may be considered a potential intervention to modulate the inflammatory and oxidative stress that cause impairments in locomotion, cognitive, and behavioral functions throughout life. The beneficial effects of polyphenols are linked with several mechanisms, including epigenetic alterations, involving the AMP-activated protein kinase (AMPK), nuclear factor kappa B (NF-κB), and phosphoinositide 3-kinase (PI3K) pathways. To highlight these new perspectives, the objective of this systematic review was to summarize the understanding emerging from preclinical studies about polyphenol supplementation, its capacity to minimize brain injury caused by hypoxia-ischemia in terms of morphological, inflammatory, and oxidative parameters and its repercussions for motor and behavioral functions.