RESUMEN

Zoonoses such as ZIKV and SARS-CoV-2 pose a severe risk to global health. There is urgent need for broad antiviral strategies based on host-targets filling gaps between pathogen emergence and availability of therapeutic or preventive strategies. Significant reduction of pathogen titers decreases spread of infections and thereby ensures health systems not being overloaded and public life to continue. Based on previously observed interference with FGFR1/2-signaling dependent impact on interferon stimulated gene (ISG)-expression, we identified Pim kinases as promising druggable cellular target. We therefore focused on analyzing the potential of pan-Pim kinase inhibition to trigger a broad antiviral response. The pan-Pim kinase inhibitor AZD1208 exerted an extraordinarily high antiviral effect against various ZIKV isolates, SARS-CoV-2 and HBV. This was reflected by strong reduction in viral RNA, proteins and released infectious particles. Especially in case of SARS-CoV-2, AZD1208 led to a complete removal of viral traces in cells. Kinome-analysis revealed vast changes in kinase landscape upon AZD1208 treatment, especially for inflammation and the PI3K/Akt-pathway. For ZIKV, a clear correlation between antiviral effect and increase in ISG-expression was observed. Based on a cell culture model with impaired ISG-induction, activation of the PI3K-Akt-mTOR axis, leading to major changes in the endolysosomal equilibrium, was identified as second pillar of the antiviral effect triggered by AZD1208-dependent Pim kinase inhibition, also against HBV. We identified Pim-kinases as cellular target for a broad antiviral activity. The antiviral effect exerted by inhibition of Pim kinases is based on at least two pillars: innate immunity and modulation of the endolysosomal system.

Asunto(s)

Antivirales , Inmunidad Innata , Proteínas Proto-Oncogénicas c-akt , Proteínas Proto-Oncogénicas c-pim-1 , SARS-CoV-2 , Transducción de Señal , Serina-Treonina Quinasas TOR , Humanos , Inmunidad Innata/efectos de los fármacos , Antivirales/farmacología , Serina-Treonina Quinasas TOR/metabolismo , Proteínas Proto-Oncogénicas c-pim-1/antagonistas & inhibidores , Proteínas Proto-Oncogénicas c-pim-1/metabolismo , SARS-CoV-2/efectos de los fármacos , SARS-CoV-2/inmunología , Proteínas Proto-Oncogénicas c-akt/metabolismo , Transducción de Señal/efectos de los fármacos , Fosfatidilinositol 3-Quinasas/metabolismo , Lisosomas/efectos de los fármacos , Lisosomas/metabolismo , Virus Zika/efectos de los fármacos , Inhibidores de Proteínas Quinasas/farmacología , Animales , Virus de la Hepatitis B/efectos de los fármacos , Endosomas/efectos de los fármacos , Endosomas/metabolismo , Línea Celular , COVID-19/inmunología , COVID-19/virología , Tratamiento Farmacológico de COVID-19 , Replicación Viral/efectos de los fármacos , Compuestos de Bifenilo , Tiazolidinas

RESUMEN

The Proviral Integration site for the Moloney murine leukemia virus (PIM)-1 kinase and its family members (PIM-2 and PIM-3) regulate several cellular functions including survival, proliferation, and apoptosis. Recent studies showed their involvement in the pathogenesis of rheumatoid arthritis RA, while no studies are available on psoriatic arthritis (PsA) and axial spondyloarthritis (axSpA). The main objective of this study is to assess the expression of PIM kinases in inflammatory arthritides, their correlation with proinflammatory cytokines, and their variation after treatment with biologic disease-modifying anti-rheumatic drugs or JAK inhibitors. We evaluated PIM-1, -2, and -3 expression at the gene and protein level, respectively, in the peripheral blood mononuclear cells and serum of patients with RA, PsA, axSpA, and healthy individuals (CTR). All the samples showed expression of PIM-1, -2, and -3 kinases both at the gene and protein level. PIM-1 was the most expressed protein, PIM-3 the least. PIM kinase levels differed between controls and disease groups, with reduced PIM-1 protein and increased PIM-3 protein in all disease samples compared to controls. No difference was found in the expression of these molecules between the three different pathologies. PIM levels were not modified after 6 months of therapy. In conclusion, our preliminary data suggest a deregulation of the PIM pathway in inflammatory arthritides. In-depth studies on the role of PIM kinases in this field are warranted.

Asunto(s)

Artritis Psoriásica , Espondiloartritis Axial , Proteínas Serina-Treonina Quinasas , Proteínas Proto-Oncogénicas c-pim-1 , Humanos , Artritis Psoriásica/tratamiento farmacológico , Artritis Psoriásica/genética , Leucocitos Mononucleares , Inhibidores de Proteínas Quinasas/farmacología , Inhibidores de Proteínas Quinasas/uso terapéutico , Proteínas Serina-Treonina Quinasas/metabolismo , Proteínas Proto-Oncogénicas/metabolismo

RESUMEN

PIM kinases have important pro-tumorigenic roles and mediate several oncogenic traits, including cell proliferation, survival, and chemotherapeutic resistance. As a result, multiple PIM inhibitors have been pursued as investigational new drugs in cancer; however, response to PIM inhibitors in solid tumors has fallen short of expectations. We found that inhibition of PIM kinase activity stabilizes protein levels of all three PIM isoforms (PIM1/2/3), and this can promote resistance to PIM inhibitors and chemotherapy. To overcome this effect, we designed PIM proteolysis targeting chimeras (PROTACs) to target PIM for degradation. PIM PROTACs effectively downmodulated PIM levels through the ubiquitin-proteasome pathway. Importantly, degradation of PIM kinases was more potent than inhibition of catalytic activity at inducing apoptosis in prostate cancer cell line models. In conclusion, we provide evidence of the advantages of degrading PIM kinases versus inhibiting their catalytic activity to target the oncogenic functions of PIM kinases.

Asunto(s)

Resistencia a Antineoplásicos , Neoplasias de la Próstata , Masculino , Humanos , Neoplasias de la Próstata/tratamiento farmacológico , Neoplasias de la Próstata/patología , Fosforilación , Apoptosis , Proliferación Celular , Inhibidores de Proteínas Quinasas/farmacología , Inhibidores de Proteínas Quinasas/uso terapéutico , Proteínas Proto-Oncogénicas c-pim-1

RESUMEN

The serine/threonine-specific Moloney murine leukemia virus (PIM) kinase family (i.e., PIM1, PIM2, and PIM3) has been extensively studied in tumorigenesis. PIM kinases are downstream of several cytokine signaling pathways that drive immune-mediated diseases. Uncontrolled T helper 17 (Th17) cell activation has been associated with the pathogenesis of autoimmunity. However, the detailed molecular function of PIMs in human Th17 cell regulation has yet to be studied. In the present study, we comprehensively investigated how the three PIMs simultaneously alter transcriptional gene regulation during early human Th17 cell differentiation. By combining PIM triple knockdown with bulk and scRNA-seq approaches, we found that PIM deficiency promotes the early expression of key Th17-related genes while suppressing Th1-lineage genes. Further, PIMs modulate Th cell signaling, potentially via STAT1 and STAT3. Overall, our study highlights the inhibitory role of PIMs in human Th17 cell differentiation, thereby suggesting their association with autoimmune phenotypes.

Asunto(s)

Proteínas Serina-Treonina Quinasas , Proteínas Proto-Oncogénicas c-pim-1 , Animales , Ratones , Humanos , Proteínas Serina-Treonina Quinasas/metabolismo , Proteínas Proto-Oncogénicas c-pim-1/genética , Proteínas Proto-Oncogénicas c-pim-1/metabolismo , Transducción de Señal , Hematopoyesis , Diferenciación Celular , Células Th17/metabolismo

RESUMEN

Following the publication of this paper, it was drawn to the Editor's attention by a concerned reader that (in addition to overlapping data panels internally within the figure, suggesting that some of the data had been derived from the same original sources where the results of differently performed experiments were intended to have been portrayed) certain of the data featured in Fig. 5A on p. 2123 had already been published in another article written by different authors at different research institutes [Tian F, Ding D and Li D: Fangchinoline targets PI3K and suppresses PI3K/AKT signaling pathway in SGC7901 cells. Int J Oncol 46: 2355­2363, 2015]. Owing to the fact that the contentious data in the above article had already been published prior to its submission to International Journal of Oncology, the Editor has decided that this paper should be retracted from the Journal. The authors were asked for an explanation to account for these concerns, but the Editorial Office did not receive a reply. The Editor apologizes to the readership for any inconvenience caused. [International Journal of Oncology 49: 2116­2126, 2016; DOI: 10.3892/ijo.2016.3708].

RESUMEN

Lactate dehydrogenase A (LDHA) is a glycolytic enzyme catalysing the reversible conversion of pyruvate to lactate. It has been implicated as a substrate for PIM kinases, yet the relevant target sites and functional consequences of phosphorylation have remained unknown. Here, we show that all three PIM family members can phosphorylate LDHA at serine 161. When we investigated the physiological consequences of this phosphorylation in PC3 prostate cancer and MCF7 breast cancer cells, we noticed that it suppressed ubiquitin-mediated degradation of nuclear LDHA and promoted interactions between LDHA and 14-3-3 proteins. By contrast, in CRISPR/Cas9-edited knock-out cells lacking all three PIM family members, ubiquitination of nuclear LDHA was dramatically increased followed by its decreased expression. Our data suggest that PIM kinases support nuclear LDHA expression and activities by promoting phosphorylation-dependent interactions of LDHA with 14-3-3ε, which shields nuclear LDHA from ubiquitin-mediated degradation.

Asunto(s)

Lactato Deshidrogenasa 5 , Proteínas Serina-Treonina Quinasas , Serina , Humanos , Línea Celular Tumoral , Lactato Deshidrogenasa 5/metabolismo , Serina/genética , Serina/metabolismo , Ubiquitinación , Ubiquitinas/metabolismo , Proteínas Serina-Treonina Quinasas/metabolismo

RESUMEN

Introduction: Multiple myeloma (MM) is still an incurable plasma cell malignancy. The efficacy of immunotherapy on MM remains unsatisfactory, and the underlying molecular mechanisms still are not fully understood. Methods: In this study, we delineated the dynamic features of immune cell in MM bone marrow (BM) along with elevated tumor cell infiltration by single-cell RNA sequencing (scRNA-seq), and investigated the underlying mechanisms on dysfunction of immune cells associated with myelomagenesis. Results: We found that immune cells were activated in those patients with low infiltration of tumor cells, meanwhile suppressed with elevated infiltration of MM cells, which facilitated MM escaping from immune surveillance. Besides PD-1, abnormal expression of PIM kinases, KLRB1 and KLRC1 were involved in the defect of immune cells in MM patients. Importantly, we found aberrant metabolic processes were associated with the immunosuppressive microenvironment in MM patients. Disordered amino acid metabolism promoted the dysfunction of cytotoxicity CD8 T cells as well as lipid metabolism disorder was associated with the dysregulation of NK and DCs in MM. As metabolic checkpoints, PIM kinases would be potential effective strategies for MM immunotherapy. Discussion: In summary, redressing the disordered metabolism should be the key points to get promising effects in immune-based therapies.

Asunto(s)

Mieloma Múltiple , Humanos , Inmunoterapia , Células Plasmáticas/patología , Médula Ósea , Vigilancia Inmunológica , Microambiente Tumoral

RESUMEN

We previously demonstrated that natural product-inspired 3,4-dihydropyrrolo[1,2-a]pyrazin-1(2H)-ones derivatives delivered potent and selective PIM kinases inhibitors however with non-optimal ADME/PK properties and modest oral bioavailability. Herein, we describe a structure-based scaffold decoration and a stereoselective approach to this chemical class. The synthesis, structure-activity relationship studies, chiral analysis, and pharmacokinetic data of compounds from this inhibitor class are presented herein. Compound 20c demonstrated excellent potency on PIM1 and PIM2 with exquisite kinases selectivity and PK properties that efficiently and dose-dependently promoted c-Myc degradation and appear to be promising lead compounds for further development.

Asunto(s)

Alcaloides , Antineoplásicos , Alcaloides/farmacología , Antineoplásicos/farmacología , Línea Celular Tumoral , Inhibidores de Proteínas Quinasas/química , Inhibidores de Proteínas Quinasas/farmacología , Proteínas Proto-Oncogénicas c-pim-1/química , Proteínas Proto-Oncogénicas c-pim-1/metabolismo , Estereoisomerismo , Relación Estructura-Actividad

RESUMEN

Proviral integration sites for Moloney murine leukemia virus (PIM) kinases are upregulated at the protein level in response to hypoxia and have multiple protumorigenic functions, promoting cell growth, survival, and angiogenesis. However, the mechanism responsible for the induction of PIM in hypoxia remains unknown. Here, we examined factors affecting PIM kinase stability in normoxia and hypoxia. We found that PIM kinases were upregulated in hypoxia at the protein level but not at the mRNA level, confirming that PIMs were upregulated in hypoxia in a hypoxia inducible factor 1-independent manner. PIM kinases were less ubiquitinated in hypoxia than in normoxia, indicating that hypoxia reduced their proteasomal degradation. We identified the deubiquitinase ubiquitin-specific protease 28 (USP28) as a key regulator of PIM1 and PIM2 stability. The overexpression of USP28 increased PIM protein stability and total levels in both normoxia and hypoxia, and USP28-knockdown significantly increased the ubiquitination of PIM1 and PIM2. Interestingly, coimmunoprecipitation assays showed an increased interaction between PIM1/2 and USP28 in response to hypoxia, which correlated with reduced ubiquitination and increased protein stability. In a xenograft model, USP28-knockdown tumors grew more slowly than control tumors and showed significantly lower levels of PIM1 in vivo. In conclusion, USP28 blocked the ubiquitination and increased the stability of PIM1/2, particularly in hypoxia. These data provide the first insight into proteins responsible for controlling PIM protein degradation and identify USP28 as an important upstream regulator of this hypoxia-induced, protumorigenic signaling pathway.

Asunto(s)

Hipoxia , Proteínas Proto-Oncogénicas c-pim-1 , Línea Celular Tumoral , Enzimas Desubicuitinizantes , Humanos , Hipoxia/metabolismo , Ratones , Proteínas Proto-Oncogénicas c-pim-1/genética , Ubiquitina Tiolesterasa

RESUMEN

Mayaro virus (MAYV) manipulates cell machinery to successfully replicate. Thus, identifying host proteins implicated in MAYV replication represents an opportunity to discover potential antiviral targets. PIM kinases are enzymes that regulate essential cell functions and also appear to be critical factors in the replication of certain viruses. In this study we explored the consequences of PIM kinase inhibition in the replication of MAYV and other arboviruses. Cytopathic effects or viral titers in samples from MAYV-, Chikungunya-, Una- or Zika-infected cells treated with PIM kinase inhibitors were evaluated using an inverted microscope or plaque-forming assays. The expression of viral proteins E1 and nsP1 in MAYV-infected cells was assessed using an immunofluorescence confocal microscope or Western blot. Our results revealed that PIM kinase inhibition partially prevented MAYV-induced cell damage and also promoted a decrease in viral titers for MAYV, UNAV and ZIKV. The inhibitory effect of PIM kinase blocking was observed for each of the MAYV strains tested and also occurred as late as 8 h post infection (hpi). Finally, PIM kinase inhibition suppressed the expression of MAYV E1 and nsP1 proteins. Taken together, these findings suggest that PIM kinases could represent an antiviral target for MAYV and other arboviruses.

Asunto(s)

Alphavirus/efectos de los fármacos , Inhibidores de Proteínas Quinasas/farmacología , Proteínas Proto-Oncogénicas c-pim-1/antagonistas & inhibidores , Replicación Viral/efectos de los fármacos , Animales , Antivirales/farmacología , Línea Celular , Virus Chikungunya/efectos de los fármacos , Humanos , Virus Zika/efectos de los fármacos

RESUMEN

Pim kinases (proviral integration site for Moloney murine leukemia virus kinases) are overexpressed in various types of hematological malignancies and solid carcinomas, and promote cell proliferation and survival. Thus, Pim kinases are validated as targets for antitumor therapy. In this context, our combined efforts in natural product-inspired library generation and screening furnished very promising dibenzo[b,d]furan derivatives derived from cercosporamide. Among them, lead compound 44 was highlighted as a potent Pim-1/2 kinases inhibitor with an additional nanomolar IC50 value against CLK1 (cdc2-like kinases 1) and displayed a low micromolar anticancer potency towards the MV4-11 (AML) cell line, expressing high endogenous levels of Pim-1/2 kinases. The design, synthesis, structure-activity relationship, and docking studies are reported herein and supported by enzyme, cellular assays, and Galleria mellonella larvae testing for acute toxicity.

Asunto(s)

Antineoplásicos/farmacología , Benzofuranos/química , Benzofuranos/farmacología , Inhibidores de Proteínas Quinasas/farmacología , Proteínas Serina-Treonina Quinasas/antagonistas & inhibidores , Proteínas Tirosina Quinasas/antagonistas & inhibidores , Proteínas Proto-Oncogénicas c-pim-1/antagonistas & inhibidores , Animales , Antineoplásicos/síntesis química , Antineoplásicos/química , Benzofuranos/síntesis química , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Ensayos de Selección de Medicamentos Antitumorales , Humanos , Modelos Moleculares , Estructura Molecular , Mariposas Nocturnas , Inhibidores de Proteínas Quinasas/síntesis química , Inhibidores de Proteínas Quinasas/química , Proteínas Serina-Treonina Quinasas/metabolismo , Proteínas Tirosina Quinasas/metabolismo , Proteínas Proto-Oncogénicas c-pim-1/metabolismo , Células Tumorales Cultivadas

RESUMEN

We performed an X-ray crystallographic study of complexes of protein kinase PIM-1 with three inhibitors comprising an adenosine mimetic moiety, a linker, and a peptide-mimetic (d-Arg)6 fragment. Guided by the structural models, simplified chemical structures with a reduced number of polar groups and chiral centers were designed. The developed inhibitors retained low-nanomolar potency and possessed remarkable selectivity toward the PIM kinases. The new inhibitors were derivatized with biotin or fluorescent dye Cy5 and then applied for the detection of PIM kinases in biochemical solutions and in complex biological samples. The sandwich assay utilizing a PIM-2-selective detection antibody featured a low limit of quantification (44 pg of active recombinant PIM-2). Fluorescent probes were efficiently taken up by U2OS cells and showed a high extent of co-localization with PIM-1 fused with a fluorescent protein. Overall, the developed inhibitors and derivatives represent versatile chemical tools for studying PIM function in cellular systems in normal and disease physiology.

Asunto(s)

Colorantes Fluorescentes , Imagen Molecular , Peptidomiméticos , Inhibidores de Proteínas Quinasas , Proteínas Proto-Oncogénicas c-pim-1 , Carbocianinas/química , Carbocianinas/farmacología , Línea Celular Tumoral , Cristalografía por Rayos X , Colorantes Fluorescentes/química , Colorantes Fluorescentes/farmacología , Humanos , Peptidomiméticos/química , Peptidomiméticos/farmacología , Inhibidores de Proteínas Quinasas/química , Inhibidores de Proteínas Quinasas/farmacología , Proteínas Proto-Oncogénicas c-pim-1/antagonistas & inhibidores , Proteínas Proto-Oncogénicas c-pim-1/metabolismo

RESUMEN

BACKGROUND: The oncogenic PIM kinases and the tumor-suppressive LKB1 kinase have both been implicated in the regulation of cell growth and metabolism, albeit in opposite directions. Here we investigated whether these kinases interact with each other to influence AMPK activation and tumorigenic growth of prostate and breast cancer cells. METHODS: We first determined how PIM and LKB1 kinases affect AMPK phosphorylation levels. We then used in vitro kinase assays to demonstrate that LKB1 is phosphorylated by PIM kinases, and site-directed mutagenesis to identify the PIM target sites in LKB1. The cellular functions of PIM and LKB1 kinases were evaluated using either pan-PIM inhibitors or CRISPR/Cas9 genomic editing, with which all three PIM family members and/or LKB1 were knocked out from PC3 prostate and MCF7 breast cancer cell lines. In addition to cell proliferation assays, we examined the effects of PIM and/or LKB1 loss on tumor growth using the chick embryo chorioallantoic membrane (CAM) xenograft model. RESULTS: We provide both genetic and pharmacological evidence to demonstrate that inhibition of PIM expression or activity increases phosphorylation of AMPK at Thr172 in both PC3 and MCF7 cells, but not in their derivatives lacking LKB1. This is explained by our observation that all three PIM family kinases can phosphorylate LKB1 at Ser334. Wild-type LKB1, but not its phosphodeficient derivative, can restore PIM inhibitor-induced AMPK phosphorylation in LKB1 knock-out cells. In the CAM model, loss of LKB1 enhances tumorigenicity of PC3 xenografts, while cells lacking both LKB1 and PIMs exhibit slower proliferation rates and form smaller tumors. CONCLUSION: PIM kinases are novel negative regulators of LKB1 that affect AMPK activity in an LKB1-dependent fashion. The impairment of cell proliferation and tumor growth in cells lacking both LKB1 and PIMs indicates that these kinases possess a shared signaling role in the context of cancer. These data also suggest that PIM inhibitors may be a rational therapeutic option for LKB1-deficient tumors. Video Abstract.

Asunto(s)

Quinasas de la Proteína-Quinasa Activada por el AMP/metabolismo , Proteínas Quinasas Activadas por AMP/metabolismo , Carcinogénesis/metabolismo , Carcinogénesis/patología , Proteínas Proto-Oncogénicas c-pim-1/metabolismo , Proteínas Quinasas Activadas por AMP/antagonistas & inhibidores , Línea Celular Tumoral , Proliferación Celular , Activación Enzimática , Humanos , Fosforilación , Unión Proteica , Especificidad por Sustrato

RESUMEN

The three oncogenic PIM family kinases have been implicated in the development of prostate cancer (PCa). The aim of this study was to examine the mRNA and protein expression levels of PIM1, PIM2, and PIM3 in PCa and their associations with the MYC and ERG oncogenes. We utilized prostate tissue specimens of normal, benign prostatic hyperplasia (BPH), prostatic intraepithelial neoplasia (PIN), untreated PCa, and castration-resistant prostate cancer (CRPC) for immunohistochemical (IHC) analysis. In addition, we analyzed data from publicly available mRNA expression and chromatin immunoprecipitation sequencing (ChIP-Seq) datasets. Our data demonstrated that PIM expression levels are significantly elevated in PCa compared to benign samples. Strikingly, the expression of both PIM1 and PIM2 was further increased in CRPC compared to PCa. We also demonstrated a significant association between upregulated PIM family members and both the ERG and MYC oncoproteins. Interestingly, ERG directly binds to the regulatory regions of all PIM genes and upregulates their expression. Furthermore, ERG suppression with siRNA reduced the expression of PIM in PCa cells. These results provide evidence for cooperation of PIM and the MYC and ERG oncoproteins in PCa development and progression and may help to stratify suitable patients for PIM-targeted therapies.

Asunto(s)

Neoplasias de la Próstata/genética , Proteínas Proto-Oncogénicas c-pim-1/genética , Anciano , Regulación Neoplásica de la Expresión Génica/genética , Humanos , Masculino , Persona de Mediana Edad , Próstata/patología , Hiperplasia Prostática/genética , Hiperplasia Prostática/patología , Neoplasias de la Próstata/patología , ARN Mensajero/genética , ARN Interferente Pequeño/genética , Regulador Transcripcional ERG/genética , Regulación hacia Arriba/genética

RESUMEN

Dysregulation of the developmentally important Notch signaling pathway is implicated in several types of cancer, including breast cancer. However, the specific roles and regulation of the four different Notch receptors have remained elusive. We have previously reported that the oncogenic PIM kinases phosphorylate Notch1 and Notch3. Phosphorylation of Notch1 within the second nuclear localization sequence of its intracellular domain (ICD) enhances its transcriptional activity and tumorigenicity. In this study, we analyzed Notch3 phosphorylation and its functional impact. Unexpectedly, we observed that the PIM target sites are not conserved between Notch1 and Notch3. Notch3 ICD (N3ICD) is phosphorylated within a domain, which is essential for formation of a transcriptionally active complex with the DNA-binding protein CSL. Through molecular modeling, X-ray crystallography, and isothermal titration calorimetry, we demonstrate that phosphorylation of N3ICD sterically hinders its interaction with CSL and thereby inhibits its CSL-dependent transcriptional activity. Surprisingly however, phosphorylated N3ICD still maintains tumorigenic potential in breast cancer cells under estrogenic conditions, which support PIM expression. Taken together, our data indicate that PIM kinases modulate the signaling output of different Notch paralogs by targeting distinct protein domains and thereby promote breast cancer tumorigenesis via both CSL-dependent and CSL-independent mechanisms.

Asunto(s)

Neoplasias de la Mama/patología , Carcinogénesis , Proteínas Proto-Oncogénicas c-pim-1/metabolismo , Receptor Notch3/metabolismo , Transporte Activo de Núcleo Celular , Animales , Línea Celular Tumoral , Núcleo Celular/metabolismo , Humanos , Proteína de Unión a la Señal Recombinante J de las Inmunoglobulinas/metabolismo , Ratones , Modelos Moleculares , Proteínas Musculares/metabolismo , Fosforilación , Dominios Proteicos , Receptor Notch3/química

RESUMEN

BACKGROUND: Novel therapies are needed for patients with hepatoblastoma because of an increasing incidence of disease and poor prognosis for advanced, refractory, and recurrent disease. PIM kinases promote tumorigenesis in hepatoblastoma. A novel PIM inhibitor, PIM447, has shown promise in inhibiting oncogenesis in hematologic and lymphoid malignancies. We hypothesized that PIM inhibition with PIM447 would result in decreased tumorigenesis in hepatoblastoma. METHODS: The effects of PIM447 on hepatoblastoma viability, proliferation, motility, apoptosis, and tumor cell stemness were assessed in HuH6, a human hepatoblastoma cell line, and COA67, a human hepatoblastoma patient-derived xenograft. RESULTS: PIM447 significantly decreased the viability, proliferation, and motility of HuH6 and COA67 cells. Apoptosis significantly increased following PIM447 treatment. PIM447 had a significant impact on tumor cell stemness as evidenced by decreased expression of CD133 and reduced ability of HuH6 and COA67 cells to form tumorspheres. Furthermore, combining PIM447 with cisplatin resulted in a significant decrease in cell viability compared to either treatment alone. CONCLUSION: We showed that PIM447 inhibits oncogenesis and potentiates the effects of cisplatin in hepatoblastoma and, therefore, warrants further investigation as a potential therapeutic agent for hepatoblastoma.

Asunto(s)

Hepatoblastoma , Neoplasias Hepáticas , Apoptosis , Línea Celular Tumoral , Proliferación Celular , Transformación Celular Neoplásica , Cisplatino/farmacología , Hepatoblastoma/tratamiento farmacológico , Hepatoblastoma/genética , Humanos , Neoplasias Hepáticas/tratamiento farmacológico

RESUMEN

Proviral integration site for Moloney murine leukemia virus (Pim)-1/2 kinase overexpression has been identified in a variety of hematologic (e.g., multiple myeloma or acute myeloid leukemia (AML)) and solid (e.g., colorectal carcinoma) tumors, playing a key role in cancer progression, metastasis, and drug resistance, and is linked to poor prognosis. These kinases are thus considered interesting targets in oncology. We report herein the design, synthesis, structure-activity relationships (SAR) and in vitro evaluations of new quinoxaline derivatives, acting as dual Pim1/2 inhibitors. Two lead compounds (5c and 5e) were then identified, as potent submicromolar Pim-1 and Pim-2 inhibitors. These molecules were also able to inhibit the growth of the two human cell lines, MV4-11 (AML) and HCT-116 (colorectal carcinoma), expressing high endogenous levels of Pim-1/2 kinases.

Asunto(s)

Inhibidores de Proteínas Quinasas/síntesis química , Inhibidores de Proteínas Quinasas/farmacología , Proteínas Serina-Treonina Quinasas/antagonistas & inhibidores , Proteínas Proto-Oncogénicas c-pim-1/antagonistas & inhibidores , Proteínas Proto-Oncogénicas/antagonistas & inhibidores , Quinoxalinas/síntesis química , Quinoxalinas/farmacología , Técnicas de Química Sintética , Humanos , Simulación del Acoplamiento Molecular , Conformación Proteica , Inhibidores de Proteínas Quinasas/química , Inhibidores de Proteínas Quinasas/metabolismo , Proteínas Serina-Treonina Quinasas/química , Proteínas Serina-Treonina Quinasas/metabolismo , Proteínas Proto-Oncogénicas/química , Proteínas Proto-Oncogénicas/metabolismo , Proteínas Proto-Oncogénicas c-pim-1/química , Proteínas Proto-Oncogénicas c-pim-1/metabolismo , Quinoxalinas/química , Quinoxalinas/metabolismo

RESUMEN

N-substituted azaindoles were discovered as potent pan-PIM inhibitors. Lead optimization, guided by structure and focused on physico-chemical properties allowed us to solve inherent hERG and permeability liabilities, and provided compound 27, which subsequently impacted KG-1 tumor growth in a mouse model.

Asunto(s)

Antineoplásicos/farmacología , Compuestos Aza/farmacología , Indoles/farmacología , Inhibidores de Proteínas Quinasas/farmacología , Animales , Antineoplásicos/síntesis química , Antineoplásicos/metabolismo , Compuestos Aza/síntesis química , Compuestos Aza/metabolismo , Línea Celular Tumoral , Cristalografía por Rayos X , Humanos , Indoles/síntesis química , Indoles/metabolismo , Ratones , Piperidinas/síntesis química , Piperidinas/metabolismo , Piperidinas/farmacología , Unión Proteica , Inhibidores de Proteínas Quinasas/síntesis química , Inhibidores de Proteínas Quinasas/metabolismo , Proteínas Proto-Oncogénicas c-pim-1/antagonistas & inhibidores , Proteínas Proto-Oncogénicas c-pim-1/metabolismo , Pirrolidinas/síntesis química , Pirrolidinas/metabolismo , Pirrolidinas/farmacología , Ratas , Estereoisomerismo , Ensayos Antitumor por Modelo de Xenoinjerto

RESUMEN

BACKGROUND: The PIM family kinases promote cancer cell survival and motility as well as metastatic growth in various types of cancer. We have previously identified several PIM substrates, which support cancer cell migration and invasiveness. However, none of them are known to regulate cellular movements by directly interacting with the actin cytoskeleton. Here we have studied the phosphorylation-dependent effects of PIM1 on actin capping proteins, which bind as heterodimers to the fast-growing actin filament ends and stabilize them. METHODS: Based on a phosphoproteomics screen for novel PIM substrates, we have used kinase assays and fluorescence-based imaging techniques to validate actin capping proteins as PIM1 substrates and interaction partners. We have analysed the functional consequences of capping protein phosphorylation on cell migration and adhesion by using wound healing and real-time impedance-based assays. We have also investigated phosphorylation-dependent effects on actin polymerization by analysing the protective role of capping protein phosphomutants in actin disassembly assays. RESULTS: We have identified capping proteins CAPZA1 and CAPZB2 as PIM1 substrates, and shown that phosphorylation of either of them leads to increased adhesion and migration of human prostate cancer cells. Phosphorylation also reduces the ability of the capping proteins to protect polymerized actin from disassembly. CONCLUSIONS: Our data suggest that PIM kinases are able to induce changes in actin dynamics to support cell adhesion and movement. Thus, we have identified a novel mechanism through which PIM kinases enhance motility and metastatic behaviour of cancer cells. Video abstract.

Asunto(s)

Proteínas de Capping de la Actina/metabolismo , Movimiento Celular , Neoplasias de la Próstata/metabolismo , Neoplasias de la Próstata/patología , Proteínas Proto-Oncogénicas c-pim-1/metabolismo , Actinas/metabolismo , Animales , Adhesión Celular , Línea Celular Tumoral , Extensiones de la Superficie Celular/metabolismo , Citoplasma/metabolismo , Humanos , Masculino , Ratones , Fosforilación , Multimerización de Proteína , Subunidades de Proteína/metabolismo , Proteínas Proto-Oncogénicas c-pim-1/antagonistas & inhibidores

RESUMEN

The PIM Kinases belong to the family of a proto-oncogene that essentially phosphorylates the serine/threonine residues of the target proteins. They are primarily categorized into three types PIM-1, PIM-2, PIM-3 which plays an indispensable regulatory role in signal transduction cascades, by promoting cell survival, proliferation, and drug resistance. These kinases are overexpressed in several solid as well as hematopoietic tumors which supports in vitro and in vivo malignant cell growth along with survival by regulating cell cycle and inhibiting apoptosis. They lack regulatory domain which makes them constitutively active once transcribed. PIM kinases usually appear to be important downstream effectors of oncoproteins which overexpresses and helps in mediating drug resistance to available agents, such as rapamycin. Structural studies of PIM kinases revealed that they have unique hinge regions where two Proline resides and makes ATP binding unique, by offering a target for an increasing number of potent PIM kinase inhibitors. Preclinical studies of those inhibitory compounds in various cancers indicate that these novel agents show promising activity and some of them currently being under examination. In this review, we have outlined PIM kinases molecular mechanism and signaling pathways along with matriculation in various cancer and list of inhibitors often used.

Asunto(s)

Neoplasias/tratamiento farmacológico , Inhibidores de Proteínas Quinasas/farmacología , Proteínas Proto-Oncogénicas c-pim-1/antagonistas & inhibidores , Animales , Antineoplásicos/farmacología , Biomarcadores de Tumor/metabolismo , Humanos , Neoplasias/enzimología , Fosforilación , Proto-Oncogenes Mas , Proteínas Proto-Oncogénicas c-pim-1/metabolismo , Transducción de Señal/efectos de los fármacos