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1.
Respir Res ; 25(1): 175, 2024 Apr 23.
Article En | MEDLINE | ID: mdl-38654248

BACKGROUND: Two isoforms of Phosphoinositide 3-kinase (PI3K), p110γ and p110δ, are predominantly expressed in leukocytes and represent attractive therapeutic targets for the treatment of allergic asthma. The study aim was to assess the impact of administration of an inhaled PI3Kγδ inhibitor (AZD8154) in a rat model of asthma. METHODS: Firstly, we checked that the tool compound, AZD8154, inhibited rat PI3K γ & δ kinases using rat cell-based assays. Subsequently, a time-course study was conducted in a rat model of asthma to assess PI3K activity in the lung and how it is temporally associated with other key transcription pathways and asthma like features of the model. Finally, the impact on lung dosed AZD8154 on target engagement, pathway specificity, airway inflammation and lung function changes was assessed. RESULTS: Data showed that AZD8154 could inhibit rat PI3K γ & δ isoforms and, in a rat model of allergic asthma the PI3K pathway was activated in the lung. Intratracheal administration of AZD8154 caused a dose related suppression PI3K pathway activation (reduction in pAkt) and unlike after budesonide treatment, STAT and NF-κB pathways were not affected by AZD8154. The suppression of the PI3K pathway led to a marked inhibition of airway inflammation and reduction in changes in lung function. CONCLUSION: These data show that a dual PI3Kγδ inhibitor suppress key features of disease in a rat model of asthma to a similar degree as budesonide and indicate that dual PI3Kγδ inhibition may be an effective treatment for people suffering from allergic asthma.


Asthma , Disease Models, Animal , Animals , Asthma/drug therapy , Asthma/metabolism , Rats , Male , Class Ib Phosphatidylinositol 3-Kinase/metabolism , Rats, Sprague-Dawley , Phosphoinositide-3 Kinase Inhibitors/pharmacology , Phosphoinositide-3 Kinase Inhibitors/therapeutic use , Lung/drug effects , Lung/metabolism , Lung/pathology , Lung/enzymology , Dose-Response Relationship, Drug , Protein Kinase Inhibitors/pharmacology , Class I Phosphatidylinositol 3-Kinases/antagonists & inhibitors , Class I Phosphatidylinositol 3-Kinases/metabolism , Anti-Asthmatic Agents/pharmacology , Ovalbumin/toxicity
2.
Nitric Oxide ; 146: 1-9, 2024 May 01.
Article En | MEDLINE | ID: mdl-38428514

BACKGROUND: Cannabidiol (CBD) is the second most abundant pharmacologically active component present in Cannabis sp. Unlike Δ-9-tetrahydrocannabinol (THC), it has no psychotomimetic effects and has recently received significant interest from the scientific community due to its potential to treat anxiety and epilepsy. CBD has excellent anti-inflammatory potential and can be used to treat some types of inflammatory and neuropathic pain. In this context, the present study aimed to evaluate the analgesic mechanism of cannabidiol administered systemically for the treatment of neuropathic pain and determine the endogenous mechanisms involved with this analgesia. METHODS: Neuropathic pain was induced by sciatic nerve constriction surgery, and the nociceptive threshold was measured using the paw compression test in mice. RESULTS: CBD produced dose-dependent antinociception after intraperitoneal injection. Selective inhibition of PI3Kγ dose-dependently reversed CBD-induced antinociception. Selective inhibition of nNOS enzymes reversed the antinociception induced by CBD, while selective inhibition of iNOS and eNOS did not alter this antinociception. However, the inhibition of cGMP production by guanylyl cyclase did not alter CBD-mediated antinociception, but selective blockade of ATP-sensitive K+ channels dose-dependently reversed CBD-induced antinociception. Inhibition of S-nitrosylation dose-dependently and completely reversed CBD-mediated antinociception. CONCLUSION: Cannabidiol has an antinociceptive effect when administered systemically and this effect is mediated by the activation of PI3Kγ as well as by nitric oxide and subsequent direct S-nitrosylation of KATP channels on peripheral nociceptors.


Analgesics , Cannabidiol , Class Ib Phosphatidylinositol 3-Kinase , KATP Channels , Neuralgia , Nitric Oxide Synthase Type I , Nitric Oxide , Signal Transduction , Animals , Cannabidiol/pharmacology , KATP Channels/metabolism , Male , Signal Transduction/drug effects , Neuralgia/drug therapy , Neuralgia/metabolism , Mice , Nitric Oxide/metabolism , Class Ib Phosphatidylinositol 3-Kinase/metabolism , Nitric Oxide Synthase Type I/metabolism , Analgesics/pharmacology , Analgesia
3.
Cancer Lett ; 584: 216615, 2024 Mar 01.
Article En | MEDLINE | ID: mdl-38199586

The biological role of B7-H1 intrinsic signal is reportedly diverse and controversial, its signal pathway remains unclear. Although B7-H1 blocking antibodies were found to have agonist capacity, their binding features and agonist mechanisms need further investigation. Here, by constructing cell strains with full-length or truncated B7-H1, we found that B7-H1 functioned as a receptor to transmit cell death signal from PD-1 protein or anti-B7-H1s through its cytoplasmic domain. Specific binding to the IgV-like domain of B7-H1 was required for the downstream signal. Upon agonists interaction, B7-H1 regulated the degradation of phosphoinositide 3-kinases (PI3Ks) subunit p110γ, subsequently inhibited the PI3K/AKT/mTOR pathway, and significantly increased autophagy. Moreover, B7-H1 agonists also suppressed ubiquitylation in B7-H1+cells by reducing ubiquitin-activating enzyme (E1), eventually leading to cell death. Finally, we validated the receptor role of B7-H1 in multiple tumor cells and demonstrated that B7-H1 agonists could suppress tumor progression independent of T cells in vivo. Our findings revealed that B7-H1 agonists functions as a PI3K inhibitor and may offer new strategies for PI3K targeting therapy.


Phosphatidylinositol 3-Kinases , Proto-Oncogene Proteins c-akt , B7-H1 Antigen/metabolism , Cell Death , Class Ib Phosphatidylinositol 3-Kinase , Histamine Agonists , Phosphatidylinositol 3-Kinases/metabolism , Proto-Oncogene Proteins c-akt/metabolism , TOR Serine-Threonine Kinases/metabolism
4.
Blood ; 143(19): 1965-1979, 2024 May 09.
Article En | MEDLINE | ID: mdl-38271660

ABSTRACT: Acute myeloid leukemia (AML) is an aggressive hematological malignancy originating from transformed hematopoietic stem or progenitor cells. AML prognosis remains poor owing to resistance and relapse driven by leukemia stem cells (LSCs). Targeting molecules essential for LSC function is a promising therapeutic approach. The phosphatidylinositol 3-kinase (PI3K)/AKT pathway is often dysregulated in AML. We found that although PI3Kγ is highly enriched in LSCs and critical for self-renewal, it was dispensable for normal hematopoietic stem cells. Mechanistically, PI3Kγ-AKT signaling promotes nuclear factor erythroid 2-related factor 2 (NRF2) nuclear accumulation, which induces 6-phosphogluconate dehydrogenase (PGD) and the pentose phosphate pathway, thereby maintaining LSC stemness. Importantly, genetic or pharmacological inhibition of PI3Kγ impaired expansion and stemness of murine and human AML cells in vitro and in vivo. Together, our findings reveal a key role for PI3Kγ in selectively maintaining LSC function by regulating AKT-NRF2-PGD metabolic pathway. Targeting the PI3Kγ pathway may, therefore, eliminate LSCs without damaging normal hematopoiesis, providing a promising therapeutic strategy for AML.


Class Ib Phosphatidylinositol 3-Kinase , Leukemia, Myeloid, Acute , Neoplastic Stem Cells , Pentose Phosphate Pathway , Leukemia, Myeloid, Acute/pathology , Leukemia, Myeloid, Acute/metabolism , Leukemia, Myeloid, Acute/genetics , Neoplastic Stem Cells/metabolism , Neoplastic Stem Cells/pathology , Animals , Humans , Mice , Class Ib Phosphatidylinositol 3-Kinase/metabolism , Class Ib Phosphatidylinositol 3-Kinase/genetics , Cell Self Renewal , NF-E2-Related Factor 2/metabolism , NF-E2-Related Factor 2/genetics , Signal Transduction , Proto-Oncogene Proteins c-akt/metabolism , Proto-Oncogene Proteins c-akt/genetics
5.
Bioorg Med Chem ; 84: 117261, 2023 04 15.
Article En | MEDLINE | ID: mdl-37011446

Targeting PI3Kγ would be a useful strategy for treating inflammatory and cancer diseases. However, the development of selective inhibitors of PI3Kγ is very challenging due to the high structural and sequence homology with other PI3K isoforms. A series of quinazolinone derivatives were designed, synthesized and biologically evaluated as PI3Kγ-selective inhibitors. Among all the 28 compounds, compound 9b was found to be the most potent selective inhibitor with IC50 values of 13.11 nM against PI3Kγ kinase. Additionally, compound 9b could generate toxicity on leukemia cells in a panel of 12 different of cancer cell lines with the IC50 value of 2.41 ± 0.11 µM on Jurkat cell. Preliminary mechanism studies indicated that compound 9b through inhibit the activity of PI3K-AKT in human and murine leukemia cells, and activated phosphorylated p38 and phosphorylated ERK presented potent antiproliferative activity, which provided a potent small molecule for further cancer therapy.


Antineoplastic Agents , Leukemia , Neoplasms , Protein Kinase Inhibitors , Quinazolinones , Animals , Humans , Mice , Antineoplastic Agents/chemistry , Cell Proliferation , Drug Design , Phosphatidylinositol 3-Kinases/metabolism , Protein Kinase Inhibitors/chemistry , Protein Kinase Inhibitors/pharmacology , Quinazolinones/chemistry , Quinazolinones/pharmacology , Structure-Activity Relationship , Class Ib Phosphatidylinositol 3-Kinase
6.
Cell Immunol ; 380: 104573, 2022 10.
Article En | MEDLINE | ID: mdl-36031460

Considering the possible interaction between mesenchymal stem cells (MSCs) and PI3Kγ-associated drugs, we evaluated the efficacy and action mechanism of MSCs in the treatment of colitis in PI3Kγ-/- mice. Trinitro-benzene-sulfonic acid enema was used to create a colitis model, and MSCs were transplanted through the caudal vein to treat colitis in wild-type and PI3Kγ-/- mice. We sequenced microbial 16S rRNA genes in the colonic mucosa of PI3Kγ-/- and wild-type mice and quantified colonic IgA, IL-2, IL-10, IL-17A, occludin, and serum IgA. MSC transplantation led to a more serious reduction in the weight of trinitro-benzene-sulfonic acid-administered PI3Kγ-/- mice than that in wild-type mice. The disease activity index, pathological scoring, number of taxa in the colon, Berger-Parker index, I-index, proportion of Proteobacteria, and IgA level in the blood were higher in PI3Kγ-/- mice than in wild-type mice after MSC transplantation. The occludin and IL-10 levels in the colon tissues decreased before and after MSC transplantation in PI3Kγ-/- mice, whereas they were increased in wild-type mice The IL-17 level decreased in both wild-type and PI3Kγ-/- mice, with knockout mice showing a greater decrease. Therefore, MSC transplantation in PI3Kγ-/- mice led to increased numbers of exogenous pathogenic microorganisms and enhanced colitis that was difficult to relieve.


Class Ib Phosphatidylinositol 3-Kinase/metabolism , Colitis , Mesenchymal Stem Cell Transplantation , Animals , Benzene , Colitis/chemically induced , Cytokines , Disease Models, Animal , Immunoglobulin A , Inflammation , Interleukin-10/genetics , Mice , Mice, Inbred C57BL , Mice, Knockout , Occludin , RNA, Ribosomal, 16S , Trinitrobenzenesulfonic Acid
7.
Immunobiology ; 227(5): 152242, 2022 09.
Article En | MEDLINE | ID: mdl-35870262

Single nucleotide polymorphisms (SNPs) that do not change the composition of amino acids and cause synonymous mutations (sSNPs) were previously considered to lack any functional roles. However, sSNPs have recently been shown to interfere with protein expression owing to a myriad of factors related to the regulation of transcription, mRNA stability, and protein translation processes. In patients with Chagas disease, the presence of the synonymous mutation rs1129293 in phosphatidylinositol-4,5-bisphosphate 3-kinase gamma (PIK3CG) gene contributes to the development of the chronic Chagas cardiomyopathy (CCC), instead of the digestive or asymptomatic forms. In this study, we aimed to investigate whether rs1129293 is associated with the transcription of PIK3CG mRNA and its activity by quantifying AKT phosphorylation in the heart samples of 26 chagasic patients with CCC. Our results showed an association between rs1129293 and decreased PIK3CG mRNA expression levels in the cardiac tissues of patients with CCC. The phosphorylation levels of AKT, the protein target of PI3K, were also reduced in patients with this mutation, but were not correlated with PI3KCG mRNA expression levels. Moreover, bioinformatics analysis showed that rs1129293 and other SNPs in linkage disequilibrium (LD) were associated with the transcriptional regulatory elements, post-transcriptional modifications, and cell-specific splicing expression of PIK3CG mRNA. Therefore, our data demonstrates that the synonymous SNP rs1129293 is capable of affecting the PIK3CG mRNA expression and PI3Kγ activation.


Chagas Cardiomyopathy , Chagas Cardiomyopathy/genetics , Class Ib Phosphatidylinositol 3-Kinase/genetics , Humans , Phosphatidylinositol 3-Kinases , Polymorphism, Single Nucleotide , Proto-Oncogene Proteins c-akt , RNA, Messenger/genetics , RNA, Messenger/metabolism , Silent Mutation
8.
Ann N Y Acad Sci ; 1515(1): 196-207, 2022 09.
Article En | MEDLINE | ID: mdl-35725890

Phosphoinositide 3-kinases (PI3Ks) are a family of enzymes phosphorylating phospholipids in the membrane, thereby, promoting the PI3K/AKT signaling cascade. PI3Ks are involved in a variety of fundamental cellular functions, including tumor necrosis factor α (TNFα)-induced tight junction (TJ) impairment-a hallmark of inflammatory bowel diseases. Most of the studies analyzing the role of class I PI3K signaling in epithelial barrier maintenance did not decipher which of the isoforms are responsible for the observed effects. By using wild-type and PI3Kγ-deficient HT-29/B6 cells, we characterized the functional role of PI3Kγ in these cells under inflammatory conditions. Measurement of the transepithelial electrical resistance and the paracellular flux of macromolecules revealed that monolayers of PI3Kγ-deficient cells, compared with wild-type cells, were protected against TNFα-induced barrier dysfunction. This effect was independent of any PI3K activity because treatment with a pan-PI3K inhibitor did not alter this observation. By immunostaining, we found correlative changes in the distribution of the TJ marker ZO-1. Furthermore, the absence of PI3Kγ reduced the basal level of the pore-forming TJ protein claudin-2. Our study suggests a novel noncanonical, kinase-independent scaffolding function of PI3Kγ in TNFα-induced barrier dysfunction.


Phosphatidylinositol 3-Kinases , Tumor Necrosis Factor-alpha , Class Ib Phosphatidylinositol 3-Kinase , Claudin-2/metabolism , Colon , HT29 Cells , Humans , Intestinal Mucosa/metabolism , Phosphatidylinositol 3-Kinases/metabolism , Phosphatidylinositol 3-Kinases/pharmacology , Phosphatidylinositols/metabolism , Phosphatidylinositols/pharmacology , Proto-Oncogene Proteins c-akt/metabolism , Tight Junctions/metabolism , Tumor Necrosis Factor-alpha/metabolism
9.
Nat Commun ; 13(1): 1768, 2022 04 01.
Article En | MEDLINE | ID: mdl-35365657

Myeloid cells play key roles in cancer immune suppression and tumor progression. In response to tumor derived factors, circulating monocytes and granulocytes extravasate into the tumor parenchyma where they stimulate angiogenesis, immune suppression and tumor progression. Chemokines, cytokines and interleukins stimulate PI3Kγ-mediated Rap1 activation, leading to conformational changes in integrin α4ß1 that promote myeloid cell extravasation and tumor inflammation Here we show that PI3Kγ activates a high molecular weight form of myosin light chain kinase, MLCK210, that promotes myosin-dependent Rap1 GTP loading, leading to integrin α4ß1 activation. Genetic or pharmacological inhibition of MLCK210 suppresses integrin α4ß1 activation, as well as tumor inflammation and progression. These results demonstrate a critical role for myeloid cell MLCK210 in tumor inflammation and serve as basis for the development of alternative approaches to develop immune oncology therapeutics.


Class Ib Phosphatidylinositol 3-Kinase/metabolism , Myosin-Light-Chain Kinase , Neoplasms , Cell Adhesion/physiology , Humans , Inflammation , Molecular Weight , Myeloid Cells/metabolism , Myosin-Light-Chain Kinase/metabolism , Neoplasms/genetics
10.
Dis Markers ; 2022: 8707061, 2022.
Article En | MEDLINE | ID: mdl-35308138

Objective: To investigate the role of leptin in regulating cell inflammation and protecting myocardium after myocardial ischemia-reperfusion injury in rats through signaling pathway at tissue and molecular protein levels. Methods: Healthy female SD rats were randomly divided into 4 groups, which were sham, I/R group, leptin low-dose intervention group, and high-dose intervention group (40 µg/kg and 80 µg/kg, respectively). Cardiac hemodynamics, myocardial enzymology, inflammatory indices, and pathological changes were observed. Western blot was used to observe the expression of PI3K, AKT, and NFκB protein by leptin. Results: Leptin can improve the hemodynamics of cardiac ischemia-reperfusion rats, improve the expression of myocardial enzymology, reduce the release of cardiac and serum inflammatory factors, increased PI3k, AKT, and NFκB expression, and reduce the occurrence of inflammation from the perspective of gross pathology, thus protecting the body. Conclusion: Leptin pretreatment can reduce MIRI injury, and the protective mechanism may be that leptin upregulates PI3K-AKT-NFκB expression in myocardial tissue to reduce inflammation and promote repair of I/R injury.


Inflammation/metabolism , Leptin , Myocardial Reperfusion Injury , Protective Agents , Animals , Class Ib Phosphatidylinositol 3-Kinase , Female , Leptin/immunology , Leptin/metabolism , Myocardial Reperfusion Injury/drug therapy , Myocardial Reperfusion Injury/metabolism , Myocardium/enzymology , Phosphatidylinositol 3-Kinases/metabolism , Proto-Oncogene Proteins c-akt/metabolism , Rats , Rats, Sprague-Dawley , Signal Transduction
11.
Nat Rev Immunol ; 22(11): 687-700, 2022 11.
Article En | MEDLINE | ID: mdl-35322259

Over the past two decades, new insights have positioned phosphoinositide 3-kinase-γ (PI3Kγ) as a context-dependent modulator of immunity and inflammation. Recent advances in protein structure determination and drug development have allowed for generation of highly specific PI3Kγ inhibitors, with the first now in clinical trials for several oncology indications. Recently, a monogenic immune disorder caused by PI3Kγ deficiency was discovered in humans and modelled in mice. Human inactivated PI3Kγ syndrome confirms the immunomodulatory roles of PI3Kγ and strengthens newly defined roles of this molecule in modulating inflammatory cytokine release in macrophages. Here, we review the functions of PI3Kγ in the immune system and discuss how our understanding of its potential as a therapeutic target has evolved.


Class Ib Phosphatidylinositol 3-Kinase , Phosphatidylinositol 3-Kinases , Humans , Mice , Animals , Class Ib Phosphatidylinositol 3-Kinase/metabolism , Phosphatidylinositol 3-Kinases/metabolism , Phosphatidylinositol 3-Kinase , Phosphoinositide-3 Kinase Inhibitors , Macrophages/metabolism
12.
Sci Transl Med ; 14(638): eabl6328, 2022 03 30.
Article En | MEDLINE | ID: mdl-35353541

Cyclic adenosine 3',5'-monophosphate (cAMP)-elevating agents, such as ß2-adrenergic receptor (ß2-AR) agonists and phosphodiesterase (PDE) inhibitors, remain a mainstay in the treatment of obstructive respiratory diseases, conditions characterized by airway constriction, inflammation, and mucus hypersecretion. However, their clinical use is limited by unwanted side effects because of unrestricted cAMP elevation in the airways and in distant organs. Here, we identified the A-kinase anchoring protein phosphoinositide 3-kinase γ (PI3Kγ) as a critical regulator of a discrete cAMP signaling microdomain activated by ß2-ARs in airway structural and inflammatory cells. Displacement of the PI3Kγ-anchored pool of protein kinase A (PKA) by an inhaled, cell-permeable, PI3Kγ mimetic peptide (PI3Kγ MP) inhibited a pool of subcortical PDE4B and PDE4D and safely increased cAMP in the lungs, leading to airway smooth muscle relaxation and reduced neutrophil infiltration in a murine model of asthma. In human bronchial epithelial cells, PI3Kγ MP induced unexpected cAMP and PKA elevations restricted to the vicinity of the cystic fibrosis transmembrane conductance regulator (CFTR), the ion channel controlling mucus hydration that is mutated in cystic fibrosis (CF). PI3Kγ MP promoted the phosphorylation of wild-type CFTR on serine-737, triggering channel gating, and rescued the function of F508del-CFTR, the most prevalent CF mutant, by enhancing the effects of existing CFTR modulators. These results unveil PI3Kγ as the regulator of a ß2-AR/cAMP microdomain central to smooth muscle contraction, immune cell activation, and epithelial fluid secretion in the airways, suggesting the use of a PI3Kγ MP for compartment-restricted, therapeutic cAMP elevation in chronic obstructive respiratory diseases.


Cystic Fibrosis Transmembrane Conductance Regulator , Phosphatidylinositol 3-Kinase , Animals , Class Ib Phosphatidylinositol 3-Kinase , Cystic Fibrosis Transmembrane Conductance Regulator/metabolism , Humans , Inflammation , Mice , Peptides/metabolism , Phosphatidylinositol 3-Kinase/metabolism , Phosphatidylinositol 3-Kinases/metabolism
13.
Cells ; 11(5)2022 02 23.
Article En | MEDLINE | ID: mdl-35269396

Inflammatory cells contribute to the pathogenesis of renal ischemia-reperfusion injury (IRI). However, the signaling mechanisms underlying the infiltration of inflammatory cells into the kidney are not well understood. In this study, we examined the effects of phosphoinositide 3 kinase γ (PI3Kγ) on inflammatory cells infiltration into the kidney in response to ischemia-reperfusion injury. Compared with wild-type mice, PI3Kγ knockout mice displayed less IRI in the kidney with fewer tubular apoptotic cell. Furthermore, PI3Kγ deficiency decreased the number of infiltrated neutrophils, macrophages, and T cells in the kidney, which was accompanied by a decrease in the expression of pro-inflammatory cytokines in the kidney. Moreover, wild-type mice treated with AS-605240, a selective PI3Kγ inhibitor, displayed less tubular damage, accumulated fewer inflammatory cells, and expressed less proinflammatory molecules in the kidney following IRI. These results demonstrate that PI3Kγ has a critical role in the pathogenesis of kidney damage in IRI, indicating that PI3Kγ inhibition may serve as a potential therapeutic strategy for the prevention of ischemia-reperfusion-induced kidney injury.


Acute Kidney Injury , Class Ib Phosphatidylinositol 3-Kinase/metabolism , Reperfusion Injury , Acute Kidney Injury/metabolism , Animals , Mice , Mice, Inbred C57BL , Phosphatidylinositol 3-Kinase , Phosphatidylinositol 3-Kinases , Reperfusion Injury/metabolism
14.
Eur J Pain ; 26(4): 825-834, 2022 04.
Article En | MEDLINE | ID: mdl-35044019

BCKGROUND: Aripiprazole is an antipsychotic drug used to treat schizophrenia and bipolar disorder. Recently, its peripheral analgesic component was evaluated, however, the mechanism involved in this effect is not fully established. Therefore, the aim of the study was to obtain pharmacological evidence for the involvement of the nitric oxide system in the peripheral antinociceptive effect induced by aripiprazole. METHODS: The hyperalgesia was induced via intraplantar injection of prostaglandin E2 in mice and the nociceptive thresholds were evaluated using the paw pressure test. All drugs were injected locally into the right hind paw. RESULTS: The PI3K inhibitor (AS605240), but not rapamycin (mTOR kinase inhibitor), reversed the peripheral antinociceptive effect induced by Aripiprazole. Antinociception was antagonized by the non-selective inhibitor of the nitric oxide synthase (L-NOarg). The same response was observed using the selective iNOS, but not with the selective nNOS inhibitors. The selective guanylyl cyclase enzyme inhibitor (ODQ) and the non-selective potassium channel blocker tetraethylammonium were able to reverse the antinociceptive effect of aripiprazole. The same was seen using glibenclamide, an ATP-dependent K+ channel blocker. However, calcium-activated potassium channel blockers of small and high conductance, dequalinium chloride and paxilline, respectively, did not reverse this effect. The injection of cGMP-specific phosphodiesterase type 5 inhibitor zaprinast, potentiated the antinociceptive effect induced by a low dose of aripiprazole. CONCLUSION: The results provide evidence that aripiprazole induces peripheral antinociceptive effects via PI3K/NO/cGMP/KATP pathway activation.


Analgesics , Antipsychotic Agents , Aripiprazole , Adenosine Triphosphate , Analgesics/therapeutic use , Animals , Antipsychotic Agents/pharmacology , Aripiprazole/pharmacology , Class Ib Phosphatidylinositol 3-Kinase/metabolism , Cyclic GMP/metabolism , Mice , Nitric Oxide/metabolism
15.
Bioengineered ; 13(1): 1491-1506, 2022 01.
Article En | MEDLINE | ID: mdl-34986757

This paper probes the mechanisms underlying miR-142-3p's modulation of hepatocellular carcinoma (HCC) invasion and apoptosis. Quantitative real-time PCR and Western blot monitored the miR-142-3p profile in HCC tissues and non-tumor tissues. The correlation between miR-142-3p expression and HCC patients' clinicopathological indicators was analyzed. miR-142-3p overexpression and knockdown models were established in HCC cell lines. Cell proliferation was gauged by the colony formation assay and BrdU staining. For measuring apoptosis, flow cytometry and Western blot were implemented. Transwell assay tested cell migration and invasion. miR-142-3p mimics or inhibitors were transfected in Huh7 and HCCLM3 cells. The targeting association between miR-142-3p and PIK3CG was predicted through bioinformatics and further verified by related experiments. The influence of PIK3CG overexpression on miR-142-3p's role in HCC was assayed. A xenografted tumor model was built in mice to validate miR-142-3p knockdown's influence on HCC in vivo. As a result, miR-142-3p exhibited a decreased profile in HCC tissues and cells. Overexpressing miR-142-3p accelerated apoptosis and suppressed the PI3K/AKT/HIF-1α signal. Knocking down miR-142-3p presented opposite effects. PIK3CG overexpression dampened the anti-tumor effect of miR-142-3p. miR-142-3p repressed HCC invasion and intensified apoptosis to restrain HCC by abating the PIK3CG-mediated PI3K/AKT/HIF-1α pathway.


Carcinoma, Hepatocellular/drug therapy , Class Ib Phosphatidylinositol 3-Kinase/genetics , Liver Neoplasms/drug therapy , MicroRNAs/genetics , RNA, Small Interfering/administration & dosage , Animals , Apoptosis , Carcinoma, Hepatocellular/genetics , Carcinoma, Hepatocellular/pathology , Cell Line, Tumor , Cell Movement , Cell Proliferation , Class Ib Phosphatidylinositol 3-Kinase/drug effects , Female , Gene Expression Regulation, Neoplastic/drug effects , Gene Knockdown Techniques , Humans , Liver Neoplasms/genetics , Liver Neoplasms/pathology , Male , Mice , RNA, Small Interfering/pharmacology , Xenograft Model Antitumor Assays
16.
Blood ; 139(4): 523-537, 2022 01 27.
Article En | MEDLINE | ID: mdl-35084470

Current limitations in using chimeric antigen receptor T(CART) cells to treat patients with hematological cancers include limited expansion and persistence in vivo that contribute to cancer relapse. Patients with chronic lymphocytic leukemia (CLL) have terminally differentiated T cells with an exhausted phenotype and experience low complete response rates after autologous CART therapy. Because PI3K inhibitor therapy is associated with the development of T-cell-mediated autoimmunity, we studied the effects of inhibiting the PI3Kδ and PI3Kγ isoforms during the manufacture of CART cells prepared from patients with CLL. Dual PI3Kδ/γ inhibition normalized CD4/CD8 ratios and maximized the number of CD8+ T-stem cell memory, naive, and central memory T-cells with dose-dependent decreases in expression of the TIM-3 exhaustion marker. CART cells manufactured with duvelisib (Duv-CART cells) showed significantly increased in vitro cytotoxicity against CD19+ CLL targets caused by increased frequencies of CD8+ CART cells. Duv-CART cells had increased expression of the mitochondrial fusion protein MFN2, with an associated increase in the relative content of mitochondria. Duv-CART cells exhibited increased SIRT1 and TCF1/7 expression, which correlated with epigenetic reprograming of Duv-CART cells toward stem-like properties. After transfer to NOG mice engrafted with a human CLL cell line, Duv-CART cells expressing either a CD28 or 41BB costimulatory domain demonstrated significantly increased in vivo expansion of CD8+ CART cells, faster elimination of CLL, and longer persistence. Duv-CART cells significantly enhanced survival of CLL-bearing mice compared with conventionally manufactured CART cells. In summary, exposure of CART to a PI3Kδ/γ inhibitor during manufacturing enriched the CART product for CD8+ CART cells with stem-like qualities and enhanced efficacy in eliminating CLL in vivo.


Immunotherapy, Adoptive/methods , Isoquinolines/therapeutic use , Leukemia, Lymphocytic, Chronic, B-Cell/therapy , Phosphoinositide-3 Kinase Inhibitors/therapeutic use , Purines/therapeutic use , Animals , Cells, Cultured , Cellular Reprogramming Techniques/methods , Class I Phosphatidylinositol 3-Kinases/antagonists & inhibitors , Class I Phosphatidylinositol 3-Kinases/metabolism , Class Ib Phosphatidylinositol 3-Kinase/metabolism , Epigenesis, Genetic , Humans , Leukemia, Lymphocytic, Chronic, B-Cell/genetics , Mice
17.
Sci Rep ; 12(1): 347, 2022 01 10.
Article En | MEDLINE | ID: mdl-35013447

Cancer stem cells (CSCs) are capable of continuous proliferation, self-renewal and are proposed to play significant roles in oncogenesis, tumor growth, metastasis and cancer recurrence. We have established a model of CSCs that was originally developed from mouse induced pluripotent stem cells (miPSCs) by proposing miPSCs to the conditioned medium (CM) of cancer derived cells, which is a mimic of carcinoma microenvironment. Further research found that not only PI3K-Akt but also EGFR signaling pathway was activated during converting miPSCs into CSCs. In this study, we tried to observe both of PI3Kγ inhibitor Eganelisib and EGFR inhibitor Gefitinib antitumor effects on the models of CSCs derived from miPSCs (miPS-CSC) in vitro and in vivo. As the results, targeting these two pathways exhibited significant inhibition of cell proliferation, self-renewal, migration and invasion abilities in vitro. Both Eganelisib and Gefitinib showed antitumor effects in vivo while Eganelisib displayed more significant therapeutic efficacy and less side effects than Gefitinib on all miPS-CSC models. Thus, these data suggest that the inhibitiors of PI3K and EGFR, especially PI3Kγ, might be a promising therapeutic strategy against CSCs defeating cancer in the near future.


Class Ib Phosphatidylinositol 3-Kinase/metabolism , ErbB Receptors/antagonists & inhibitors , Gefitinib/pharmacology , Induced Pluripotent Stem Cells/drug effects , Neoplastic Stem Cells/drug effects , Phosphoinositide-3 Kinase Inhibitors/pharmacology , Protein Kinase Inhibitors/pharmacology , Animals , Apoptosis/drug effects , Cell Line, Tumor , Cell Movement/drug effects , Cell Proliferation/drug effects , Cell Self Renewal/drug effects , ErbB Receptors/metabolism , Female , Induced Pluripotent Stem Cells/enzymology , Induced Pluripotent Stem Cells/pathology , Mice , Mice, Inbred BALB C , Mice, Nude , Neoplasm Invasiveness , Neoplastic Stem Cells/enzymology , Neoplastic Stem Cells/pathology , Signal Transduction , Tumor Burden/drug effects
18.
J Med Chem ; 65(2): 1418-1444, 2022 01 27.
Article En | MEDLINE | ID: mdl-34672584

Phosphoinositide-3-kinase γ (PI3Kγ) is highly expressed in immune cells and promotes the production and migration of inflammatory mediators. The inhibition of PI3Kγ has been shown to repolarize the tumor immune microenvironment to a more inflammatory phenotype, thereby controlling immune suppression in cancer. Herein, we report the structure-based optimization of an early lead series of pyrazolopyrimidine isoindolinones, which culminated in the discovery of highly potent and isoform-selective PI3Kγ inhibitors with favorable drug-like properties. X-ray cocrystal structure analysis, molecular docking studies, and detailed structure-activity relationship investigations resulted in the identification of the optimal amide and isoindolinone substituents to achieve a desirable combination of potency, selectivity, and metabolic stability. Preliminary in vitro studies indicate that inhibition of PI3Kγ with compound 56 results in a significant immune response by increasing pro-inflammatory cytokine gene expression in M1 macrophages.


Amides/chemistry , Class Ib Phosphatidylinositol 3-Kinase/chemistry , Drug Design , Drug Discovery , Phosphoinositide-3 Kinase Inhibitors/pharmacology , Pyrimidines/chemistry , Animals , Humans , Male , Molecular Docking Simulation , Rats , Rats, Sprague-Dawley , Structure-Activity Relationship
19.
Eur J Cancer ; 157: 450-463, 2021 11.
Article En | MEDLINE | ID: mdl-34601286

INTRODUCTION: We hypothesised that the combined use of radiation therapy and a phosphoinositide 3-kinaseγδ inhibitor to reduce immune suppression would enhance the efficacy of an immune checkpoint inhibitor. METHODS: Murine breast cancer cells (4T1) were grown in both immune-competent and -deficient BALB/c mice, and tumours were irradiated by 3 fractions of 24 Gy. A PD-1 blockade and a phosphoinositide 3-kinase (PI3K)γδ inhibitor were then administered every other day for 2 weeks. The same experiments were performed in humanised patient-derived breast cancer xenograft model and its tumour was sequenced to identify immune-related pathways and profile infiltrated immune cells. Transcriptomic and clinical data were acquired from The Cancer Genome Atlas pan-cancer cohort, and the deconvolution algorithm was used to profile immune cell repertoire. RESULTS: Using a PI3Kγδ inhibitor, radiation therapy (RT) and PD-1 blockade significantly delayed primary tumour growth, boosted the abscopal effect and improved animal survival. RT significantly increased CD8+cytotoxic T-cell fractions, immune-suppressive regulatory T cells (Tregs), myeloid-derived suppressor cells and M2 tumour-associated macrophages (TAMs). However, the PI3Kγδ inhibitor significantly lowered the proportions of Tregs, myeloid-derived suppressor cells and M2 TAMs, achieving dramatic gains in splenic, nodal, and tumour CD8+ T-cell populations after triple combination therapy. In a humanised patient-derived breast cancer xenograft model, triple combination therapy significantly delayed tumour growth and decreased immune-suppressive pathways. In The Cancer Genome Atlas cohort, high Treg/CD8+ T cell and M2/M1 TAM ratios were associated with poor overall patient survival. CONCLUSION: These findings indicate PI3Kγ and PI3Kδ are clinically relevant targets in an immunosuppressive TME, and combining PI3Kγδ inhibitor, RT and PD-1 blockade may overcome the therapeutic resistance of immunologically cold tumours. SYNOPSIS: Combining PI3Kγδ inhibitor, RT, and PD-1 blockade may be a viable clinical approach, helping to overcome the therapeutic resistance of immunologically cold tumours such as breast cancer.


Antineoplastic Combined Chemotherapy Protocols/pharmacology , Breast Neoplasms/therapy , Chemoradiotherapy/methods , Immune Checkpoint Inhibitors/pharmacology , Phosphoinositide-3 Kinase Inhibitors/pharmacology , Animals , Antineoplastic Combined Chemotherapy Protocols/therapeutic use , Breast Neoplasms/genetics , Breast Neoplasms/immunology , Breast Neoplasms/pathology , Cell Line, Tumor , Class I Phosphatidylinositol 3-Kinases/metabolism , Class Ib Phosphatidylinositol 3-Kinase/metabolism , Disease Models, Animal , Drug Resistance, Neoplasm/drug effects , Drug Resistance, Neoplasm/immunology , Drug Resistance, Neoplasm/radiation effects , Drug Synergism , Female , Humans , Immune Checkpoint Inhibitors/therapeutic use , Mice , Phosphoinositide-3 Kinase Inhibitors/therapeutic use , Programmed Cell Death 1 Receptor/antagonists & inhibitors , T-Lymphocytes, Cytotoxic/drug effects , T-Lymphocytes, Cytotoxic/immunology , T-Lymphocytes, Cytotoxic/radiation effects , Tumor Escape/drug effects , Tumor Escape/radiation effects , Tumor Microenvironment/drug effects , Tumor Microenvironment/immunology , Tumor Microenvironment/radiation effects , Xenograft Model Antitumor Assays
20.
Cells ; 10(10)2021 09 24.
Article En | MEDLINE | ID: mdl-34685514

(1) Background: Rapid microglial proliferation contributes to the complex responses of the innate immune system in the brain to various neuroinflammatory stimuli. Here, we investigated the regulatory function of phosphoinositide 3-kinase γ (PI3Kγ) and reactive oxygen species (ROS) for rapid proliferation of murine microglia induced by LPS and ATP. (2) Methods: PI3Kγ knockout mice (PI3Kγ KO), mice expressing catalytically inactive PI3Kγ (PI3Kγ KD) and wild-type mice were assessed for microglial proliferation using an in vivo wound healing assay. Additionally, primary microglia derived from newborn wild-type, PI3Kγ KO and PI3Kγ KD mice were used to analyze PI3Kγ effects on proliferation and cell viability, senescence and cellular and mitochondrial ROS production; the consequences of ROS production for proliferation and cell viability after LPS or ATP stimulation were studied using genetic and pharmacologic approaches. (3) Results: Mice with a loss of lipid kinase activity showed impaired proliferation of microglia. The prerequisite of induced microglial proliferation and cell viability appeared to be PI3Kγ-mediated induction of ROS production. (4) Conclusions: The lipid kinase activity of PI3Kγ plays a crucial role for microglial proliferation and cell viability after acute inflammatory activation.


Cell Proliferation/physiology , Cell Survival/physiology , Class Ib Phosphatidylinositol 3-Kinase/metabolism , Microglia/metabolism , Animals , Brain/metabolism , Cell Proliferation/genetics , Cell Survival/genetics , Class Ib Phosphatidylinositol 3-Kinase/genetics , Cyclic AMP/metabolism , Mice, Knockout , Neurogenesis/physiology , Reactive Oxygen Species/metabolism
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