Summary The histone acetyltransferase (HAT) p300/CBP is a transcriptional coactivator implicated in many gene regulatory pathways and protein acetylation events. While p300 inhibitors have been reported, a potent, selective, and readily available active-site directed small molecule inhibitor is not yet known. Here we use a structure-based, in silico screening approach to identify a commercially available pyrazolone-containing small molecule p300 HAT inhibitor, C646. C646 is a competitive p300 inhibitor with a Ki of 400 nM and is selective versus other acetyltransferases. Studies on site-directed p300 HAT mutants and synthetic modifications of C646 confirm the importance of predicted interactions in conferring potency. Inhibition of histone acetylation and cell growth by C646 in cells validate its utility as a pharmacologic probe and suggest that p300/CBP HAT is a worthy anti-cancer target.

The high risk human papillomaviruses (HPVs) are associated etiologically with the majority of human cervical carcinomas. These HPVs encode two viral oncoproteins, E6 and E7, which are expressed consistently in cervical cancers. The function of these viral oncoproteins during a productive infection is to ensure viral replication in cells that have normally withdrawn from the cell division cycle and are committed to terminal differentiation. Expression of the E7 oncoprotein has been shown to lead to the abrogation of various negative growth regulatory signals, including a p53-mediated G 1 growth arrest, TGF␤-mediated growth inhibition, and quiescence of suprabasal keratinocytes. Here we describe a novel mechanism by which E7 can uncouple cellular proliferation and differentiation. In contrast to normal, differentiating keratinocytes, HPV-16 E7-expressing keratinocytes show delayed cellular differentiation and elevated cdk2 kinase activity despite high levels of p21Cip1 and association of p21 Cip1 with cdk2. We show that the HPV E7 protein can interact with p21Cip1 and abrogate p21 Cip1-mediated inhibition of cyclin A and E-associated kinase activities. Based on these findings, we propose that this capacity of the HPV E7 oncoprotein to overcome p21 Cip1-mediated inhibition of cdk2 activity during keratinocyte differentiation contributes to the ability of E7 to allow for cellular DNA synthesis in differentiated keratinocytes.[Key Words: Human papillomavirus; cyclin dependent kinase inhibitor; cellular differentiation; cell division cycle]Received May 29, 1997; revised version accepted July 3, 1997.The papillomaviruses are small DNA viruses that have a pronounced tropism for epithelial cells where they cause benign hyperplasias or warts. Human papillomaviruses (HPVs) can be subdivided into two groups based on the propensity of the lesions to progress to malignancy. Low risk HPVs are associated with lesions with a very low frequency of malignant progression, whereas the lesions caused by the high risk HPVs can undergo carcinogenic progression (for review, see zur Hausen 1996), with more than 95% of all cervical carcinomas harboring high risk HPVs (Bosch et al. 1995). Although HPVs infect the dividing, basal cells of host epithelia, their replication is intimately tied to the cellular differentiation machinery of these cells. Expression of the viral capsid proteins and productive viral replication can only occur in the differentiated layers of the skin that are composed of cells that have withdrawn from the cell division cycle. Limited by their small genome size, with the exception of the viral E1 and E2 proteins, papillomaviruses do not encode the enzymes that are rate limiting for DNA replication. Rather, these viruses have developed multiple strategies to subvert key regulatory circuits that control host cell replication (for review, see Howley 1996). The precise mechanisms by which HPVs are able to reactivate the dormant replication machinery in these cells to replicate their own genomes are unknown. The E6 and E7 ...

Since the gene encoding Id1 was cloned in 1990, Id proteins have been implicated in regulating a variety of cellular processes, including cellular growth, senescence, differentiation, apoptosis, angiogenesis, and neoplastic transformation. The development of knockout and transgenic animal models for many members of the Id gene family has been particularly useful in sorting out the biologic relevance of these genes and their expression during normal development, malignant transformation, and tumor progression. Here we review the current understanding of Id gene function, the biologic consequences of Id gene expression, and the implications for Id gene regulation of cell growth and tumorigenesis.

BackgroundGene expression profiling has revolutionized our ability to molecularly classify primary human tumors and significantly enhanced the development of novel tumor markers and therapies; however, progress in the diagnosis and treatment of melanoma over the past 3 decades has been limited, and there is currently no approved therapy that significantly extends lifespan in patients with advanced disease. Profiling studies of melanoma to date have been inconsistent due to the heterogeneous nature of this malignancy and the limited availability of informative tissue specimens from early stages of disease.Methodology/Principle FindingsIn order to gain an improved understanding of the molecular basis of melanoma progression, we have compared gene expression profiles from a series of melanoma cell lines representing discrete stages of malignant progression that recapitulate critical characteristics of the primary lesions from which they were derived. Here we describe the unsupervised hierarchical clustering of profiling data from melanoma cell lines and melanocytes. This clustering identifies two distinctive molecular subclasses of melanoma segregating aggressive metastatic tumor cell lines from less-aggressive primary tumor cell lines. Further analysis of expression signatures associated with melanoma progression using functional annotations categorized these transcripts into three classes of genes: 1) Upregulation of activators of cell cycle progression, DNA replication and repair (CDCA2, NCAPH, NCAPG, NCAPG2, PBK, NUSAP1, BIRC5, ESCO2, HELLS, MELK, GINS1, GINS4, RAD54L, TYMS, and DHFR), 2) Loss of genes associated with cellular adhesion and melanocyte differentiation (CDH3, CDH1, c-KIT, PAX3, CITED1/MSG-1, TYR, MELANA, MC1R, and OCA2), 3) Upregulation of genes associated with resistance to apoptosis (BIRC5/survivin). While these broad classes of transcripts have previously been implicated in the progression of melanoma and other malignancies, the specific genes identified within each class of transcripts are novel. In addition, the transcription factor NF-KB was specifically identified as being a potential “master regulator” of melanoma invasion since NF-KB binding sites were identified as consistent consensus sequences within promoters of progression-associated genes.Conclusions/SignificanceWe conclude that tumor cell lines are a valuable resource for the early identification of gene signatures associated with malignant progression in tumors with significant heterogeneity like melanoma. We further conclude that the development of novel data reduction algorithms for analysis of microarray studies is critical to allow for optimized mining of important, clinically-relevant datasets. It is expected that subsequent validation studies in primary human tissues using such an approach will lead to more rapid translation of such studies to the identification of novel tumor biomarkers and therapeutic targets.