Crystal Structure of the Retinoblastoma Protein N Domain Provides Insight into Tumor Suppression, Ligand Interaction, and Holoprotein Architecture

Hassler, Markus; Singh, Sanchita; Yue, Wyatt W.; Luczynski, Maciej T.; Lakbir, Rachid; Sánchez-Sánchez, Francisco; Bader, Thomas; Pearl, Laurence H.; Mittnacht, Sibylle

doi:10.1016/j.molcel.2007.08.023

Cited by 70 publications

(98 citation statements)

References 53 publications

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“…Consequently, low doses of PHA-848125 strongly reduced DNA replication (as confirmed by robust inhibition of BrdUrd incorporation) and induced G 1 block. At higher drug doses, the accumulation of cells in G 2 -M, inhibition of cyclin A levels, and hypophosphorylation of pRb at Ser 249 and Thr 252 are consistent with inhibition of CDK1 activity (38).…”

Section: Discussionmentioning

confidence: 64%

Dual Targeting of CDK and Tropomyosin Receptor Kinase Families by the Oral Inhibitor PHA-848125, an Agent with Broad-Spectrum Antitumor Efficacy

Albanese

Alzani

Amboldi

et al. 2010

Molecular Cancer Therapeutics

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Altered expression and activity of cyclin-dependent kinase (CDK) and tropomyosin receptor kinase (TRK) families are observed in a wide variety of tumors. In those malignancies with aberrant CDK activation, the retinoblastoma protein (pRb) pathway is deregulated, leading to uncontrolled cell proliferation. Constitutive activation of TRKs is instead linked to cancer cell survival and dissemination. Here, we show that the novel small-molecule PHA-848125, a potent dual inhibitor of CDKs and TRKs, possesses significant antitumor activity. The compound inhibits cell proliferation of a wide panel of tumoral cell lines with submicromolar IC 50 . PHA-848125-treated cells show cell cycle arrest in G 1 and reduced DNA synthesis, accompanied by inhibition of pRb phosphorylation and modulation of other CDK-dependent markers. The compound additionally inhibits phosphorylation of TRKA and its substrates in cells, which functionally express this receptor. Following oral administration, PHA-848125 has significant antitumor activity in various human xenografts and carcinogen-induced tumors as well as in disseminated primary leukemia models, with plasma concentrations in rodents in the same range as those found active in inhibiting cancer cell proliferation. Mechanism of action was also confirmed in vivo as assessed in tumor biopsies from treated mice. These results show that the dual CDK-TRK inhibitor PHA-848125 has the potential for being a novel and efficacious targeted drug for cancer treatment. Mol Cancer Ther; 9(8); 2243-54. ©2010 AACR.

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Section: Discussionmentioning

confidence: 64%

Dual Targeting of CDK and Tropomyosin Receptor Kinase Families by the Oral Inhibitor PHA-848125, an Agent with Broad-Spectrum Antitumor Efficacy

Albanese

Alzani

Amboldi

et al. 2010

Molecular Cancer Therapeutics

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“…Indeed, nearly 20% of cancer-associated in-frame mutations in Rb are located in the Nterminal region (6). These lesions leave an intact C-terminal pocket and generate stable forms of Rb that bind E2F transcription factors and localize to the nucleus in a fashion similar to that of wild-type Rb (wt-Rb) (6)(7)(8)(9)(10). Several in-frame RbN exon deletions in familial retinoblastomas have been reported, including individual losses of exon 4 (Ex4), Ex5, Ex7, or Ex9 (11)(12)(13)(14).…”

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confidence: 99%

The N Terminus of the Retinoblastoma Protein Inhibits DNA Replication via a Bipartite Mechanism Disrupted in Partially Penetrant Retinoblastomas

Borysov

Nepon-Sixt

Alexandrow

2016

Molecular and Cellular Biology

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The N-terminal domain of the retinoblastoma (Rb) tumor suppressor protein (RbN) harbors in-frame exon deletions in partially penetrant hereditary retinoblastomas and is known to impair cell growth and tumorigenesis. However, how such RbN deletions contribute to Rb tumor-and growth-suppressive functions is unknown. Here we establish that RbN directly inhibits DNA replication initiation and elongation using a bipartite mechanism involving N-terminal exons lost in cancer. Specifically, Rb exon 7 is necessary and sufficient to target and inhibit the replicative CMG helicase, resulting in the accumulation of inactive CMGs on chromatin. An independent N-terminal loop domain, which forms a projection, specifically blocks DNA polymerase ␣ (Pol-␣) and Ctf4 recruitment without affecting DNA polymerases and ␦ or the CMG helicase. Individual disruption of exon 7 or the projection in RbN or Rb, as occurs in inherited cancers, partially impairs the ability of Rb/RbN to inhibit DNA replication and block G 1 -to-S cell cycle transit. However, their combined loss abolishes these functions of Rb. Thus, Rb growth-suppressive functions include its ability to block replicative complexes via bipartite, independent, and additive N-terminal domains. The partial loss of replication, CMG, or Pol-␣ control provides a potential molecular explanation for how N-terminal Rb loss-offunction deletions contribute to the etiology of partially penetrant retinoblastomas. Mutational inactivation or deletion of the retinoblastoma (Rb) tumor suppressor gene occurs in multiple cancer types, including retinoblastoma, osteosarcoma, and breast and small cell lung cancers, and deregulation or inactivation of regulatory components of the Rb pathway is a hallmark of human cancers (1). The Rb protein functions to harness a variety of cellular processes important in tumorigenesis, including regulation of the cell cycle, apoptosis, differentiation, stress responses, and DNA replication. The role of Rb in these processes derives to a large extent from interactions of proteins with the C terminus of Rb that contains a large pocket domain (1-5), and most Rb loss-of-function mutations compromise pocket structure and/or function and are highly penetrant alleles of inherited cancer in humans and mice (6).Multiple observations indicate that the N-terminal domain of Rb (RbN) (residues 1 to 400) also plays an important role in growth suppression and tumorigenesis. Indeed, nearly 20% of cancer-associated in-frame mutations in Rb are located in the Nterminal region (6). These lesions leave an intact C-terminal pocket and generate stable forms of Rb that bind E2F transcription factors and localize to the nucleus in a fashion similar to that of wild-type Rb (wt-Rb) (6-10). Several in-frame RbN exon deletions in familial retinoblastomas have been reported, including individual losses of exon 4 (Ex4), Ex5, Ex7,. Inframe deletions and mutations have also been found in Rb exons 6 and 8 in prostate cancers and astrocytomas, respectively (15, 16). Furthermore, in contrast to pock...

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“…Micro-RnAs (miRnAs) are small, noncoding RnAs that regulate gene expression by reducing the stability of conventional mRnAs and/or inhibiting their translation into proteins [1]. Over the past few years, characteristic miRNA expression profiles have been identified that enable tumor classification and prognostication and various individual miRnAs have been described as oncogenes or tumor suppressors [2][3][4]. Several recent studies have identified specific miRnAs that are central to metastatic progression.…”

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confidence: 99%

miRNA Expression in a Human Papillary Thyroid Carcinoma Cell Line Varies with Invasiveness

et al. 2010

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Abstract. The purpose of these studies was to explore the expression pattern of miRnAs associated with invasion and metastasis of human papillary thyroid carcinoma (pTC). A transwell invasion chamber was used to select progressively more invasive cancer cell populations from a clonal cell line of human pTC with cervical lymph node metastasis, ihh-4. Three sublines with progressive invasiveness, designated ihh-4M-1, ihh-4M-2 and ihh-4M-3, were obtained through this in vitro selection process. The sublines manifested an increase in colony-forming ability on soft agar and metastatic potency in nude mice. Then metastasis-related miRnAs differentially expressed between them were analyzed utilizing the miRnA microarray technique. We found that 11 metastasis-related miRnAs were differentially expressed between the invasive subpopulations and their control subpopulations; these miRNAs may account for their significant difference in the invasion capabilities and favor lymphatic metastasis of pTC.Key words: miRnA, papillary thyroid carcinoma, invasiveness PTC is the most common type of thyroid malignancy; it accounts for approximately 80% of all thyroid cancers. Micro-RnAs (miRnAs) are small, noncoding RnAs that regulate gene expression by reducing the stability of conventional mRnAs and/or inhibiting their translation into proteins [1]. Over the past few years, characteristic miRNA expression profiles have been identified that enable tumor classification and prognostication and various individual miRnAs have been described as oncogenes or tumor suppressors [2][3][4]. Several recent studies have identified specific miRnAs that are central to metastatic progression. however, little is known about the expression of miRnAs in pTC, or about their utility as biomarkers for predicting metastatic behavior and clinical outcome.invasion and metastasis are closely correlated during the development and progression of tumors. The migration and metastasis of cancer cells takes place in three steps: adherence, degradation, and movement. Tumor cells must initiate and successfully complete all of these steps in order to enter the circulatory and lymphatic systems to form distant metastases [5,6]. in order to study mechanisms accompanying each invasive step, several in vitro invasion models have been developed and can be used to assess the invasive activity of various tumor cells [7,8]. To gain insight into alterations in metastasis related miRnA expression that might govern lymphatic metastasis of pTC, we previously established highly metastatic pTC cell lines (designated ihh-4M-1, ihh-4M-2 and ihh-4M-3) from a clonal cell line of human pTC (ihh-4) through in vitro selection using a transwell invasion chamber. We compared the in vitro tumorigenicity and in vivo metastatic capabilities of these sublines by colony-formation assay and xenograft nude mouse model and found that metastatic tumor cells showed signifi-

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Crystal Structure of the Retinoblastoma Protein N Domain Provides Insight into Tumor Suppression, Ligand Interaction, and Holoprotein Architecture

Cited by 70 publications

References 53 publications

Dual Targeting of CDK and Tropomyosin Receptor Kinase Families by the Oral Inhibitor PHA-848125, an Agent with Broad-Spectrum Antitumor Efficacy

Dual Targeting of CDK and Tropomyosin Receptor Kinase Families by the Oral Inhibitor PHA-848125, an Agent with Broad-Spectrum Antitumor Efficacy

The N Terminus of the Retinoblastoma Protein Inhibits DNA Replication via a Bipartite Mechanism Disrupted in Partially Penetrant Retinoblastomas

miRNA Expression in a Human Papillary Thyroid Carcinoma Cell Line Varies with Invasiveness

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