High Content Assay for Compounds that inhibit the Assembly of the Perinucleolar Compartment: Summary
Assay Submitter (PI): Sui Huang, Northwestern University, Feinberg School of Medicine, Cell and Molecular Biology, Chicago, Illinois. ..more
High Content Assay for Compounds that inhibit the Assembly of the Perinucleolar Compartment
NIH Chemical Genomics Center [NCGC]
NIH Molecular Libraries Probe Centers Network [MLPCN]
MLPCN Grant: MH082371-01A2
Assay Submitter (PI): Sui Huang, Northwestern University, Feinberg School of Medicine, Cell and Molecular Biology, Chicago, Illinois.
The perinucleolar compartment (PNC) is a non-membrane-bound nuclear subdomain that is associated with but structurally distinct from the nucleolus. The PNC is a heritable trait, in which the number of PNCs per cell in daughter cells often mimics that of their mother cells. The PNC is heterogeneous in shape, is stable through interphase, disassembles during mitosis, and reassembles in early G1. The PNC is concentrated with newly synthesized RNA polymerase III (pol III) RNAs (MRP RNA, RNAse P H1 RNA, hY RNAs[hY1, 2, 5], Alu RNA, and SRP [7SL] RNA) and RNA binding proteins (nucleolin, PTB, CUG-BP, KSRP, Raver1, Raver2, and Rod). Continuous transcription by pol III is necessary for the structural integrity of the PNC and some protein components have been shown to bind RNAs within the PNC implicating involvement of PNCs in pol III RNA metabolism. Chemical biology studies have demonstrated that PNC maintenance is dependent on the integrity of DNA, specifically DNA base pairing, and cell biology investigations demonstrated that the PNC is nucleated upon a DNA locus, indicating that the components of the PNC are most likely directly interacting with the DNA locus.
Extensive in vitro studies have shown that the PNC is unique to tumor cells and preferentially forms in tumor cells derived from solid tissues. Examination of cancer cell lines from various origins and malignant capacities has shown that PNC prevalence correlates with the malignancy of tumors and has the potential to be developed as a pan-cancer prognostic marker. In vivo investigations using human breast tissue samples demonstrated that PNC prevalence was 0% in normal breast epithelium, increases in parallel with disease progression (as determined by staging), and reaches nearly 100% in distant metastases, demonstrating that PNC prevalence associates with the malignancy of breast cancer in vivo. Multivariate analysis further showed that a high PNC prevalence is associated with poor patient outcomes independent of current prognostic factors for stage I breast cancer patients. These studies demonstrate that the presence of the PNC reflects an advanced transformation state of cancer cells that are capable of metastasis.
Although the PNC is associated with a DNA locus and a handful of components have been identified, much remains to be investigated regarding the structure and function of the PNC and its mechanistic link to cancer. We have previously shown that PNC prevalence reduction is a valid strategy to identify compounds that can be used to study the structure and function of the PNC. It is necessary, however, to create a screening platform that is able to test a large number of compounds in an automated fashion. Here we report the development and robustness of an automated whole-cell phenotypic image-based screening assay that is capable of identifying such compounds.
Only the initial primary screening has been completed for this project, and no probes have yet been declared.
Categorized Comment - additional comments and annotations
From MLP Probe Report:
Probe count: 1
MLP Probe ML# for probe 1: ML246
PubChem Substance ID (SID) for probe 1: 117695978
PubChem Compound ID (CID) for probe 1: 50985821
Probe type for probe 1: Inhibitor
IC50/EC50 (nM) for probe 1: 397
Target for probe 1: PNC (gi: 4506243)
Disease relevance for probe 1: Cancer
Anti-target for probe 1: Cytotoxicity
Fold selectivity for probe 1: 25
NCBI Book chapter link for probe 1: http://www.ncbi.nlm.nih.gov/books/NBK143533/ (ID: 3036108)
Grant number for probe 1: MH082371-01
NCBI Book chapter title for probe 1: Discovery and Development of Small Molecules That Reduce PNC Prevalence