In vivo elucidation of PIM kinase role in tumourigenesis

Tesis doctoral inédita realizada en el Centro Nacional de Investigaciones Oncológicas y leída en la Universidad Autónoma de Madrid. Facultad de Medicina, Departamento de Bioquímica. Fecha de lectura: 18 de Junio de 201

The PIM Family of Serine/Threonine Kinases in Cancer

The proviral insertion site in Moloney murine leukemia virus, or PIM proteins, are a family of serine/threonine kinases composed of three different isoforms (PIM1, PIM2, and PIM3) that are highly evolutionarily conserved. These proteins are regulated primarily by transcription and stability through pathways that are controlled by Janus kinase/Signal transducer and activator of transcription, JAK/STAT, transcription factors. The PIM family proteins have been found to be overexpressed in hematological malignancies and solid tumors, and their roles in these tumors were confirmed in mouse tumor models. Furthermore, the PIM family proteins have been implicated in the regulation of apoptosis, metabolism, cell cycle, and homing and migration, which has led to the postulation of these proteins as interesting targets for anticancer drug discovery. In the present work, we review the importance of PIM kinases in tumor growth and as drug targets. © 2013 Wiley Periodicals, Inc.This work was supported by a fellowship from the Foundation for Research in Oncology (FERO), Fundació Josep Botet and grants from the Spanish Ministry of Science and Innovation (SAF2009–08605) and FIS (PI12/00137), Consejeria de Ciencia e Innovacion and Consejeria de Salud of the Junta de Andalucia (CTS-6844 and PI-0142).Peer Reviewe

Pim1 Kinase Cooperates with Hormone Treatment to Promote Bladder and Ureteral Urothelial Hyperplasia

This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.The Pim serine/threonine kinases have been shown to be overexpressed in cancer. Elevated levels of Pim1 kinase were demonstrated in human leukemia and lymphomas, as well as in solid tumors such as pancreatic, prostate and bladder cancers, and have been proposed as a prognostic marker. Although the Pim kinases have been identified as oncogenes in transgenic mouse models, they have only weak transforming abilities on their own. However, they have been shown to greatly enhance the ability of other genes or chemical carcinogens to induce tumors. To explore the role of Pim1 in bladder and ureteral urothelial cancer, we generated a conditional Pim1 transgenic mouse model and found that prostate specific antigen-(PSA)-driven Cre expression lead to transgene expression in the bladder upon (testosterone/estrogen) hormone treatment. We then explored the effect of Pim1 overexpression on hormone treatment, either alone or in combination with Pten haploinsufficiency. We found that Pim1 overexpression increased the severity of bladder and ureteral urothelial hyperplasias in both backgrounds, leading to pyelonephritis in transgenic animals. Our data suggest that Pim1 might contribute to progression, rather than initiation, and that the hyperplasias also contribute to the development of pyelonephritis.This work was supported by grants from Spanish Ministry of Economy and Competitivity, ISCIII (Fis: PI12/00137, RTICC: RD12/0036/0028), Consejeria de Ciencia e Innovacion (CTS-6844) and Consejeria de Salud of the Junta de Andalucia (PI-0135-2010 and PI-0306-2012). MN-G was funded by a fellowship from the Spanish Ministry of Science and Innovation.Spanish Ministry of Economy and CompetitivityISCIIISpanish Ministry of Science and InnovationConsejeria de Ciencia e Innovacion of the Junta de AndaluciaConsejeria de Salud of the Junta de AndaluciaPeer reviewe

Conditional Transgenic Expression of PIM1 Kinase in Prostate Induces Inflammation-Dependent Neoplasia

The Pim proteins are a family of highly homologous protein serine/threonine kinases that have been found to be overexpressed in cancer. Elevated levels of Pim1 kinase were first discovered in human leukemia and lymphomas. However, more recently Pim1 was found to be increased in solid tumors, including pancreatic and prostate cancers, and has been proposed as a prognostic marker. Although the Pim kinases have been identified as oncogenes in transgenic models, they have weak transforming abilities on their own. However, they have been shown to greatly enhance the ability of other genes or chemical carcinogens to induce tumors. To explore the role of Pim1 in prostate cancer, we generated conditional Pim1 transgenic mice, expressed Pim1 in prostate epithelium, and analyzed the contribution of PIM1 to neoplastic initiation and progression. Accordingly, we explored the effect of PIM1 overexpression in 3 different settings: upon hormone treatment, during aging, and in combination with the absence of one Pten allele. We have found that Pim1 overexpression increased the severity of mouse prostate intraepithelial neoplasias (mPIN) moderately in all three settings. Furthermore, Pim1 overexpression, in combination with the hormone treatment, increased inflammation surrounding target tissues leading to pyelonephritis in transgenic animals. Analysis of senescence induced in these prostatic lesions showed that the lesions induced in the presence of inflammation exhibited different behavior than those induced in the absence of inflammation. While high grade prostate preneoplastic lesions, mPIN grades III and IV, in the presence of inflammation did not show any senescence markers and demonstrated high levels of Ki67 staining, untreated animals without inflammation showed senescence markers and had low levels of Ki67 staining in similar high grade lesions. Our data suggest that Pim1 might contribute to progression rather than initiation in prostate neoplasia. © 2013 Narlik-Grassow et al.This work was supported by grants from the Spanish Ministry of Science and Innovation and Feder Funds (SAF2009-08605), Consejeria de Ciencia e Innovacion and Consejeria de Salud of the Junta de Andalucia (CTS-6844 and PI-0142). AC's laboratory is also funded by a fellowship from the Fundacion Oncologica FERO, supported by the Fundació Josep Botet. MN-G was funded by a fellowship from the Spanish Ministry of Science and Innovation.Peer Reviewe

The essential role of PIM kinases in sarcoma growth and bone invasion

This is a pre-copy-editing, author-produced PDF of an article accepted for publication in Carcinogenesis following peer review. The definitive publisher-authenticated version is available online at: http://dx.doi.org/10.1093/carcin/bgs176PIM kinases are a family of serine/threonine kinases composed of three different isoforms (PIM1, PIM 2 and PIM 3) that are highly homologous. Their expression is mediated by the JAK/STAT signalling pathway, providing survival and cell cycle transition signals. PIM kinases are heavily targeted for anticancer drug discovery. However, very little is known about the relative contribution of the different isoforms to the tumourigenesis process in vivo, and how their individual inhibition might affect tumour growth. Taking advantage of genetically modified mice, we explored whether the inhibition of specific isoforms is required to prevent sarcomas induced by 3-methylcholanthrene carcinogenic treatment. We found that absence of Pim2 and Pim3 greatly reduced sarcoma growth to a similar extent to the absence of all three isoforms. This model of sarcoma generally produces bone invasion by the tumour cells. Lack of Pim2 and Pim3 reduced tumour-induced bone invasion by 70%, which is comparable with the reduction of tumour-induced bone invasion in the absence of all three isoforms. Similar results were obtained in mouse embryonic fibroblasts (MEFs) derived from these knockout (KO) mice, where double Pim2/3 KO MEFs already showed reduced proliferation and were resistant to oncogenic transformation by the RAS oncogene. Our data also suggest an important role of Gsk3β phosphorylation in the process of tumourigenesis. Abbreviations: KOknockout. MEFsmouse embryonic fibroblasts. 3MCmethylcholanthrene. WTwild-type. © The Author 2012. Published by Oxford University Press. All rights reserved.Peer Reviewe

The role of PIM1/PIM2 kinases in tumors of the male reproductive system

The PIM family of serine/threonine kinases has three highly conserved isoforms (PIM1, PIM2 and PIM3). PIM proteins are regulated through transcription and stability by JAK/STAT pathways and are overexpressed in hematological malignancies and solid tumors. The PIM kinases possess weak oncogenic abilities, but enhance other genes or chemical carcinogens to induce tumors. We generated conditional transgenic mice that overexpress PIM1 or PIM2 in male reproductive organs and analyzed their contribution to tumorigenesis. We found an increase in alterations of sexual organs and hyperplasia in the transgenic mice correlating with inflammation. We also found that PIM1/2 are overexpressed in a subset of human male germ cells and prostate tumors correlating with inflammatory features and stem cell markers. Our data suggest that PIM1/2 kinase overexpression is a common feature of male reproductive organs tumors, which provoke tissue alterations and a large inflammatory response that may act synergistically during the process of tumorigenesis. There is also a correlation with markers of cancer stem cells, which may contribute to the therapy resistance found in tumors overexpressing PIM kinases.The AC lab was supported by grants from the Spanish Ministry of Economy and Competitiveness, Plan Estatal de I+D+I 2013–2016, ISCIII (FIS: PI15/00045) co-funded by FEDER from the Regional Development European Funds (European Union), Consejeria de Ciencia e Innovacion (CTS-1848), and Consejeria de Salud of the Junta de Andalucia (PI-0306-2012, PI-0096-2014 and ECAIF2-0176-2013). This work has also been made possible by the Fundacion BBVA. The Spanish Ministry of Education (FPU13/00426) funded MPJG.Peer reviewe

Conditional Transgenic Expression of PIM1 Kinase in Prostate Induces Inflammation-Dependent Neoplasia

<div><p>The Pim proteins are a family of highly homologous protein serine/threonine kinases that have been found to be overexpressed in cancer. Elevated levels of Pim1 kinase were first discovered in human leukemia and lymphomas. However, more recently Pim1 was found to be increased in solid tumors, including pancreatic and prostate cancers, and has been proposed as a prognostic marker. Although the Pim kinases have been identified as oncogenes in transgenic models, they have weak transforming abilities on their own. However, they have been shown to greatly enhance the ability of other genes or chemical carcinogens to induce tumors. To explore the role of Pim1 in prostate cancer, we generated conditional Pim1 transgenic mice, expressed Pim1 in prostate epithelium, and analyzed the contribution of PIM1 to neoplastic initiation and progression. Accordingly, we explored the effect of PIM1 overexpression in 3 different settings: upon hormone treatment, during aging, and in combination with the absence of one <i>Pten</i> allele. We have found that Pim1 overexpression increased the severity of mouse prostate intraepithelial neoplasias (mPIN) moderately in all three settings. Furthermore, Pim1 overexpression, in combination with the hormone treatment, increased inflammation surrounding target tissues leading to pyelonephritis in transgenic animals. Analysis of senescence induced in these prostatic lesions showed that the lesions induced in the presence of inflammation exhibited different behavior than those induced in the absence of inflammation. While high grade prostate preneoplastic lesions, mPIN grades III and IV, in the presence of inflammation did not show any senescence markers and demonstrated high levels of Ki67 staining, untreated animals without inflammation showed senescence markers and had low levels of Ki67 staining in similar high grade lesions. Our data suggest that Pim1 might contribute to progression rather than initiation in prostate neoplasia.</p> </div

Correction: Conditional Transgenic Expression of PIM1 Kinase in Prostate Induces Inflammation-Dependent Neoplasia.

The Pim proteins are a family of highly homologous protein serine/threonine kinases that have been found to be overexpressed in cancer. Elevated levels of Pim1 kinase were first discovered in human leukemia and lymphomas. However, more recently Pim1 was found to be increased in solid tumors, including pancreatic and prostate cancers, and has been proposed as a prognostic marker. Although the Pim kinases have been identified as oncogenes in transgenic models, they have weak transforming abilities on their own. However, they have been shown to greatly enhance the ability of other genes or chemical carcinogens to induce tumors. To explore the role of Pim1 in prostate cancer, we generated conditional Pim1 transgenic mice, expressed Pim1 in prostate epithelium, and analyzed the contribution of PIM1 to neoplastic initiation and progression. Accordingly, we explored the effect of PIM1 overexpression in 3 different settings: upon hormone treatment, during aging, and in combination with the absence of one Pten allele. We have found that Pim1 overexpression increased the severity of mouse prostate intraepithelial neoplasias (mPIN) moderately in all three settings. Furthermore, Pim1 overexpression, in combination with the hormone treatment, increased inflammation surrounding target tissues leading to pyelonephritis in transgenic animals. Analysis of senescence induced in these prostatic lesions showed that the lesions induced in the presence of inflammation exhibited different behavior than those induced in the absence of inflammation. While high grade prostate preneoplastic lesions, mPIN grades III and IV, in the presence of inflammation did not show any senescence markers and demonstrated high levels of Ki67 staining, untreated animals without inflammation showed senescence markers and had low levels of Ki67 staining in similar high grade lesions. Our data suggest that Pim1 might contribute to progression rather than initiation in prostate neoplasia. © 2013 Narlik-Grassow et al.This work was supported by grants from the Spanish Ministry of Science and Innovation and Feder Funds (SAF2009-08605), Consejeria de Ciencia e Innovacion and Consejeria de Salud of the Junta de Andalucia (CTS-6844 and PI-0142). AC's laboratory is also funded by a fellowship from the Fundacion Oncologica FERO, supported by the Fundació Josep Botet. MN-G was funded by a fellowship from the Spanish Ministry of Science and Innovation.Peer reviewe

Inflammation and stem markers association to PIM1/PIM2 kinase-induced tumors in breast and uterus

The PIM family of Ser/Thr kinase proteins has been implicated in tumorigenesis at different levels. PIM proteins are overexpressed in several tumor types and have been associated with chemoresistance. However, their role in hormone-dependent female tissues has not been explored, especially in the uterus, breast and ovary. We generated conditional transgenic mice with confined expression of human PIM1 or PIM2 genes in these tissues. We characterized the tumoral response to these genetic alterations corroborating their role as oncogenes since they induce hyperproliferation in all tissues and tumors in mammary gland and uterus. Furthermore, we observed a high degree of inflammatory infiltration in these tissues of transgenic mice accompanied by NFAT and mTOR activation and IL6 expression. Moreover, PIM1/2 were overexpressed in human breast, uterine and ovarian tumors, correlating with inflammatory features and stem cell markers. Our data suggest that PIM1/2 kinase overexpression provoke tissue alterations and a large IL6-dependent inflammatory response that may act synergistically during the process of tumorigenesis. The possible end-point is an increased percentage of cancer stem cells, which may be partly responsible for the therapy resistance found in tumors overexpressing PIM kinases.This work was funded by grants from the Spanish Ministry of Economy and Competitivity, Plan Estatal de I+D+I 2013–2016, ISCIII (Fis: PI15/00045) and CIBER de Cáncer (CD16/12/00275), co-funded by FEDER from Regional Development European Funds (European Union), Consejeria de Ciencia e Innovacion (CTS-1848) and Consejeria de Salud of the Junta de Andalucia (PI-0096-2014). Especial thanks to the AECC Foundation for also supporting this work. This work has also been made possible by the Fundacion BBVA. The Spanish Ministry of Education (FPU13/00426) funded MPJG.Peer reviewe

Senescence in prostate lesions.

<p>To determine senescence in prostate lesions, immunohistochemistry for several senescence markers, such as p21, p16 and p19, was performed in the prostate tissues of mice showing high grade and low grade mPIN lesions (10-month-old untreated mice and 16-week-old hormone treated mice). <b>A</b>) Senescence markers in prostate lesions. To determine senescence in prostate lesions from 16-week-old mice before and after hormone treatment, as well in 10-month-old untreated mice, immunohistochemistry for several senescence markers, namely p21, p16 and p19, was performed on the prostate tissues of mice of each genotype using the following grading scale for senescence grade (s-grade): s-grade 1 - few cells in 1 lesion (1–5% positive cells); s-grade 2 - few cells in more than one lesion; s-grade 3 - several cells (5–20%) in more than one lesion; s-grade 4 - more than 20% positive cells in more than one lesion; only concluding true senescence if s-grades 3 or 4 were reached in the same animal for at least 2 senescence markers preferably p16 and p19. <b>B</b>) Representative picture showing that senescence markers appear only in high grade mPIN in untreated mice. <b>C</b>) Senescence markers in high grade lesions – aging vs. hormone treatment. Immunohistochemistry for p16 and p19 in high grade mPIN lesions (mPINIV) in a 10-month-old untreated PTEN-Het mouse and a 16-week-old hormone treated PTEN-het mouse.</p