Rhabdomyosarcoma (RMS) is the most common type of pediatric soft tissue sarcoma. It is classified... more Rhabdomyosarcoma (RMS) is the most common type of pediatric soft tissue sarcoma. It is classified into two main subtypes: embryonal (eRMS) and alveolar (aRMS). MYC family proteins are frequently highly expressed in RMS tumors, with the highest levels correlated with poor prognosis. A pharmacological approach to inhibit MYC in cancer cells is represented by Bromodomain and Extra-Terminal motif (BET) protein inhibitors. In this paper, we evaluated the effects of BET inhibitor (+)-JQ1 (JQ1) on the viability of aRMS and eRMS cells. Interestingly, we found that the drug sensitivity of RMS cell lines to JQ1 was directly proportional to the expression of MYC. JQ1 induces G1 arrest in cells with the highest steady-state levels of MYC, whereas apoptosis is associated with MYC downregulation. These findings suggest BET inhibition as an effective strategy for the treatment of RMS alone or in combination with other drugs.
License, which permits unrestricted use, distribution, and reproduction in any medium, provided t... more License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. The human genome is epigenetically organized through a series of modifications to the histone proteins that interact with the DNA. In cancer, many of the proteins that regulate these modifications can be altered in both function and expression. One example of this is the family of histone deacetylases (HDACs), which as their name implies remove acetyl groups from the histone proteins, allowing for more condensed nucleosomal structure. HDACs have increased expression in cancer and are also believed to promote carcinogenesis through the acetylation and interaction with key transcriptional regulators. Given this, small molecule histone deacetylases inhibitors have been identified and developed, which not only inhibit HDACs, but can also lead to growth arrest, differentiation, and/or apoptosis in tumors both in vitro and in vivo. Here, we will discuss some...
During skeletal myogenesis, muscle-regulatory factors bHLH and MEF2 promote the expression of mus... more During skeletal myogenesis, muscle-regulatory factors bHLH and MEF2 promote the expression of muscle-specific genes by recruiting several chromatin-modifying complexes on specific DNA regulatory sequences. A number of MyoD-interacting proteins have been reported, but whether they are recruited to the chromatin of myogenic loci, and the relationship with other chromatin bound proteins is unknown. We show that MyoD recruits cdk9/cyclin T2, together with the histone acetyltransferases p300 and PCAF, and the chromatin remodeling complex SWI/SNF, on promoters and enhancers of muscle-specific genes, and that this event correlates with the acetylation of histone tails, remodeling of chromatin, and phosphorylation of the C-terminal domain (CTD) of the RNA polymerase II at these elements.
Cyclin-dependent kinases (cdks) are the catalytic subunits of a large family of serine/threonine ... more Cyclin-dependent kinases (cdks) are the catalytic subunits of a large family of serine/threonine protein kinases whose best-characterized members are key regulators of eukaryotic cell cycle progression. They are activated by binding to regulatory subunits generally termed as cyclins. Cdk10 is a cdc2-related kinase that contains the canonical regulatory Tyr and Thr residues present in all protein kinases and a PSTAIRE-like motif named PISSLRE. Although little is known about this protein, human cdk10 has been shown to encode two different isoforms, each having a distinct function. They differ at both the carboxy- and amino-terminals, although most of the amino acid sequence is predicted to be identical for the two isoforms. A role at the G2/M transition has been suggested for an isoform of cdk10, while the alternative splicing form interacts with the N-terminus of the Ets2 transcription factor. Here we report the cloning and the functional characterization of a cDNA encoding the murine homologue of cdk10. Unlike its human counterpart, only one murine cdk10 protein has been identified, and this unique murine cdk10 cDNA encodes a putative protein of 360 amino acids. Comparison of the amino acid sequences of murine and human cdk10 shows high homology. Murine cdk10 binds Ets2 transcription factors in vitro, does not show a direct involvement in the G2/M transition and, therefore, does not affect the proliferation rate of the cell lines analyzed.
The human genome is epigenetically organized through a series of modifications to the histone pro... more The human genome is epigenetically organized through a series of modifications to the histone proteins that interact with the DNA. In cancer, many of the proteins that regulate these modifications can be altered in both function and expression. One example of this is the family of histone deacetylases (HDACs), which as their name implies remove acetyl groups from the histone proteins, allowing for more condensed nucleosomal structure. HDACs have increased expression in cancer and are also believed to promote carcinogenesis through the acetylation and interaction with key transcriptional regulators. Given this, small molecule histone deacetylases inhibitors have been identified and developed, which not only inhibit HDACs, but can also lead to growth arrest, differentiation, and/or apoptosis in tumors bothin vitroandin vivo. Here, we will discuss some of the recent developments in clinical trials utilizing HDACs inhibitors for the treatment of both hematological malignancies as well a...
Purpose: RB2/p130, a member of the retinoblastoma gene family, maps to human chromosome 16q12.2, ... more Purpose: RB2/p130, a member of the retinoblastoma gene family, maps to human chromosome 16q12.2, a region in which deletions have been found in several human neoplasms including breast, prostatic, and ovarian carcinoma. We sought to evaluate pRb2/p130 protein expression and function in ovarian carcinoma. Experimental Design: pRb2/p130 expression was detected by immunohistochemical and Western blot analyses in 45 primary ovarian carcinoma samples. Results: Immunohistochemical analysis revealed loss or decrease of pRb2/p130 expression in 18 cases (40%). pRb2/p130 expression was mostly nuclear and inversely correlated to the tumor grade (P < 0.05). Western blot analysis correlated with immunohistochemical expression. Reverse transcription-PCR followed by Southern blot analysis was performed on a representative set of 20 ovarian carcinomas. RB2/p130 mRNA levels were consistent with protein expression. We found a significant increase in the percentage of G1-phase-arrested cells in CAO...
Proceedings: AACR 103rd Annual Meeting 2012‐‐ Mar 31‐Apr 4, 2012; Chicago, IL It is well known th... more Proceedings: AACR 103rd Annual Meeting 2012‐‐ Mar 31‐Apr 4, 2012; Chicago, IL It is well known that alterations in the pathway controlling pRb phosphorylation account for more than 50% of tumors. Here, we report a striking experimental demonstration that, in cancer cells, pRb phosphorylation by CDK/cyclin proteins depends on Pin1 activity. We propose, for the first time, a detailed mechanism regarding pRb phosphorylation and activity, thereby significantly advancing the basic understanding of cell cycle control. We conclude that at the beginning-middle of G1, CDKs phosphorylate pRb pocket domain, thus allowing its interaction with Pin1. Next, in mid-late G1, the new complex pRb-Pin1 allows CDKs to complete pRb phosphorylation, and therefore allows the expression of S phase entry genes (Cell Death & Differentiation, in press). We believe our finding will have broad implications on target therapy for cancer patients. In fact, cancer cells can inactive pRb by overexpressing only Pin1, without affecting the well-known CDK pathways. Since Pin1 is overexpressed in most tumors, this suggests a new approach to treating cancer patients by utilizing Pin1 inhibitors in combination with or without CDK inhibitors. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr LB-284. doi:1538-7445.AM2012-LB-284
Although innumerable investigations regarding the biology of lung cancer have been carried out, m... more Although innumerable investigations regarding the biology of lung cancer have been carried out, many aspects thereof remain to be addressed, including the role played by the retinoblastoma-related protein Rb2/ p130 during the evolution of this disease. Here we report novel findings on the mechanisms that control Rb2/ p130 gene expression in lung fibroblasts and characterize the effects of Rb2/p130 deregulation on the proliferative features of lung cancer cells. We revealed for the first time that in lung fibroblasts the expression of Rb2/p130 gene is directly controlled by the chromatin insulator CCCTC-binding factor, CTCF, which by binding to the Rb2/p130 gene promoter induces, and/or maintains, a specific local chromatin organization that in turn governs the transcriptional activity of Rb2/p130 gene. However, in lung cancer cells the activity of CTCF in controlling Rb2/p130 gene expression is impaired by BORIS, a CTCF-paralogue, which by binding to the Rb2/p130 gene could trigger ...
Rhabdomyosarcoma (RMS) is the most common type of pediatric soft tissue sarcoma. It is classified... more Rhabdomyosarcoma (RMS) is the most common type of pediatric soft tissue sarcoma. It is classified into two main subtypes: embryonal (eRMS) and alveolar (aRMS). MYC family proteins are frequently highly expressed in RMS tumors, with the highest levels correlated with poor prognosis. A pharmacological approach to inhibit MYC in cancer cells is represented by Bromodomain and Extra-Terminal motif (BET) protein inhibitors. In this paper, we evaluated the effects of BET inhibitor (+)-JQ1 (JQ1) on the viability of aRMS and eRMS cells. Interestingly, we found that the drug sensitivity of RMS cell lines to JQ1 was directly proportional to the expression of MYC. JQ1 induces G1 arrest in cells with the highest steady-state levels of MYC, whereas apoptosis is associated with MYC downregulation. These findings suggest BET inhibition as an effective strategy for the treatment of RMS alone or in combination with other drugs.
License, which permits unrestricted use, distribution, and reproduction in any medium, provided t... more License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. The human genome is epigenetically organized through a series of modifications to the histone proteins that interact with the DNA. In cancer, many of the proteins that regulate these modifications can be altered in both function and expression. One example of this is the family of histone deacetylases (HDACs), which as their name implies remove acetyl groups from the histone proteins, allowing for more condensed nucleosomal structure. HDACs have increased expression in cancer and are also believed to promote carcinogenesis through the acetylation and interaction with key transcriptional regulators. Given this, small molecule histone deacetylases inhibitors have been identified and developed, which not only inhibit HDACs, but can also lead to growth arrest, differentiation, and/or apoptosis in tumors both in vitro and in vivo. Here, we will discuss some...
During skeletal myogenesis, muscle-regulatory factors bHLH and MEF2 promote the expression of mus... more During skeletal myogenesis, muscle-regulatory factors bHLH and MEF2 promote the expression of muscle-specific genes by recruiting several chromatin-modifying complexes on specific DNA regulatory sequences. A number of MyoD-interacting proteins have been reported, but whether they are recruited to the chromatin of myogenic loci, and the relationship with other chromatin bound proteins is unknown. We show that MyoD recruits cdk9/cyclin T2, together with the histone acetyltransferases p300 and PCAF, and the chromatin remodeling complex SWI/SNF, on promoters and enhancers of muscle-specific genes, and that this event correlates with the acetylation of histone tails, remodeling of chromatin, and phosphorylation of the C-terminal domain (CTD) of the RNA polymerase II at these elements.
Cyclin-dependent kinases (cdks) are the catalytic subunits of a large family of serine/threonine ... more Cyclin-dependent kinases (cdks) are the catalytic subunits of a large family of serine/threonine protein kinases whose best-characterized members are key regulators of eukaryotic cell cycle progression. They are activated by binding to regulatory subunits generally termed as cyclins. Cdk10 is a cdc2-related kinase that contains the canonical regulatory Tyr and Thr residues present in all protein kinases and a PSTAIRE-like motif named PISSLRE. Although little is known about this protein, human cdk10 has been shown to encode two different isoforms, each having a distinct function. They differ at both the carboxy- and amino-terminals, although most of the amino acid sequence is predicted to be identical for the two isoforms. A role at the G2/M transition has been suggested for an isoform of cdk10, while the alternative splicing form interacts with the N-terminus of the Ets2 transcription factor. Here we report the cloning and the functional characterization of a cDNA encoding the murine homologue of cdk10. Unlike its human counterpart, only one murine cdk10 protein has been identified, and this unique murine cdk10 cDNA encodes a putative protein of 360 amino acids. Comparison of the amino acid sequences of murine and human cdk10 shows high homology. Murine cdk10 binds Ets2 transcription factors in vitro, does not show a direct involvement in the G2/M transition and, therefore, does not affect the proliferation rate of the cell lines analyzed.
The human genome is epigenetically organized through a series of modifications to the histone pro... more The human genome is epigenetically organized through a series of modifications to the histone proteins that interact with the DNA. In cancer, many of the proteins that regulate these modifications can be altered in both function and expression. One example of this is the family of histone deacetylases (HDACs), which as their name implies remove acetyl groups from the histone proteins, allowing for more condensed nucleosomal structure. HDACs have increased expression in cancer and are also believed to promote carcinogenesis through the acetylation and interaction with key transcriptional regulators. Given this, small molecule histone deacetylases inhibitors have been identified and developed, which not only inhibit HDACs, but can also lead to growth arrest, differentiation, and/or apoptosis in tumors bothin vitroandin vivo. Here, we will discuss some of the recent developments in clinical trials utilizing HDACs inhibitors for the treatment of both hematological malignancies as well a...
Purpose: RB2/p130, a member of the retinoblastoma gene family, maps to human chromosome 16q12.2, ... more Purpose: RB2/p130, a member of the retinoblastoma gene family, maps to human chromosome 16q12.2, a region in which deletions have been found in several human neoplasms including breast, prostatic, and ovarian carcinoma. We sought to evaluate pRb2/p130 protein expression and function in ovarian carcinoma. Experimental Design: pRb2/p130 expression was detected by immunohistochemical and Western blot analyses in 45 primary ovarian carcinoma samples. Results: Immunohistochemical analysis revealed loss or decrease of pRb2/p130 expression in 18 cases (40%). pRb2/p130 expression was mostly nuclear and inversely correlated to the tumor grade (P < 0.05). Western blot analysis correlated with immunohistochemical expression. Reverse transcription-PCR followed by Southern blot analysis was performed on a representative set of 20 ovarian carcinomas. RB2/p130 mRNA levels were consistent with protein expression. We found a significant increase in the percentage of G1-phase-arrested cells in CAO...
Proceedings: AACR 103rd Annual Meeting 2012‐‐ Mar 31‐Apr 4, 2012; Chicago, IL It is well known th... more Proceedings: AACR 103rd Annual Meeting 2012‐‐ Mar 31‐Apr 4, 2012; Chicago, IL It is well known that alterations in the pathway controlling pRb phosphorylation account for more than 50% of tumors. Here, we report a striking experimental demonstration that, in cancer cells, pRb phosphorylation by CDK/cyclin proteins depends on Pin1 activity. We propose, for the first time, a detailed mechanism regarding pRb phosphorylation and activity, thereby significantly advancing the basic understanding of cell cycle control. We conclude that at the beginning-middle of G1, CDKs phosphorylate pRb pocket domain, thus allowing its interaction with Pin1. Next, in mid-late G1, the new complex pRb-Pin1 allows CDKs to complete pRb phosphorylation, and therefore allows the expression of S phase entry genes (Cell Death & Differentiation, in press). We believe our finding will have broad implications on target therapy for cancer patients. In fact, cancer cells can inactive pRb by overexpressing only Pin1, without affecting the well-known CDK pathways. Since Pin1 is overexpressed in most tumors, this suggests a new approach to treating cancer patients by utilizing Pin1 inhibitors in combination with or without CDK inhibitors. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr LB-284. doi:1538-7445.AM2012-LB-284
Although innumerable investigations regarding the biology of lung cancer have been carried out, m... more Although innumerable investigations regarding the biology of lung cancer have been carried out, many aspects thereof remain to be addressed, including the role played by the retinoblastoma-related protein Rb2/ p130 during the evolution of this disease. Here we report novel findings on the mechanisms that control Rb2/ p130 gene expression in lung fibroblasts and characterize the effects of Rb2/p130 deregulation on the proliferative features of lung cancer cells. We revealed for the first time that in lung fibroblasts the expression of Rb2/p130 gene is directly controlled by the chromatin insulator CCCTC-binding factor, CTCF, which by binding to the Rb2/p130 gene promoter induces, and/or maintains, a specific local chromatin organization that in turn governs the transcriptional activity of Rb2/p130 gene. However, in lung cancer cells the activity of CTCF in controlling Rb2/p130 gene expression is impaired by BORIS, a CTCF-paralogue, which by binding to the Rb2/p130 gene could trigger ...
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