CIP2A coordinates phosphosignaling, mitosis, and the DNA damage response.

Human cancers share requirements for phosphorylation-dependent signaling, mitotic hyperactivity, and survival after DNA damage. The oncoprotein CIP2A (cancerous inhibitor of PP2A) can coordinate all these cancer cell characteristics. In addition to controlling cancer cell phosphoproteomes via inhibition of protein phosphatase PP2A, CIP2A directly interacts with the DNA damage protein TopBP1 (topoisomerase II-binding protein 1). Consequently, CIP2A allows DNA-damaged cells to enter mitosis and is essential for mitotic cells that are defective in homologous recombination (HR)-mediated DNA repair (e.g., BRCA mutants). The CIP2A-TopBP1 complex is also important for clustering fragmented chromosomes at mitosis. Clinically, CIP2A is a disease driver for basal-like triple-negative breast cancer (BL-TNBC) and a promising cancer therapy target across many cancer types.

CIP2A Interacts with TopBP1 and Drives Basal-Like Breast Cancer Tumorigenesis.

Basal-like breast cancers (BLBC) are characterized by defects in homologous recombination (HR), deficient mitotic checkpoint, and high-proliferation activity. Here, we discover CIP2A as a candidate driver of BLBC. CIP2A was essential for DNA damage-induced initiation of mouse BLBC-like mammary tumors and for survival of HR-defective BLBC cells. CIP2A was dispensable for normal mammary gland development and for unperturbed mitosis, but selectively essential for mitotic progression of DNA damaged cells. A direct interaction between CIP2A and a DNA repair scaffold protein TopBP1 was identified, and CIP2A inhibition resulted in enhanced DNA damage-induced TopBP1 and RAD51 recruitment to chromatin in mammary epithelial cells. In addition to its role in tumor initiation, and survival of BRCA-deficient cells, CIP2A also drove proliferative MYC and E2F1 signaling in basal-like triple-negative breast cancer (BL-TNBC) cells. Clinically, high CIP2A expression was associated with poor patient prognosis in BL-TNBCs but not in other breast cancer subtypes. Small-molecule reactivators of PP2A (SMAP) inhibited CIP2A transcription, phenocopied the CIP2A-deficient DNA damage response (DDR), and inhibited growth of patient-derived BLBC xenograft. In summary, these results demonstrate that CIP2A directly interacts with TopBP1 and coordinates DNA damage-induced mitotic checkpoint and proliferation, thereby driving BLBC initiation and progression. SMAPs could serve as a surrogate therapeutic strategy to inhibit the oncogenic activity of CIP2A in BLBCs. SIGNIFICANCE: These results identify CIP2A as a nongenetic driver and therapeutic target in basal-like breast cancer that regulates DNA damage-induced G2-M checkpoint and proliferative signaling.

Discovery of a Novel CIP2A Variant (NOCIVA) with Clinical Relevance in Predicting TKI Resistance in Myeloid Leukemias.

PURPOSE: Cancerous inhibitor of protein phosphatase 2A (CIP2A) is an oncoprotein that inhibits the tumor suppressor PP2A-B56α. However, CIP2A mRNA variants remain uncharacterized. Here, we report the discovery of a CIP2A splicing variant, novel CIP2A variant (NOCIVA). EXPERIMENTAL DESIGN: Characterization of CIP2A variants was performed by both 3' and 5' rapid amplification of cDNA ends from cancer cells. The function of NOCIVA was assessed by structural and molecular biology approaches. Its clinical relevance was studied in an acute myeloid leukemia (AML) patient cohort and two independent chronic myeloid leukemia (CML) cohorts. RESULTS: NOCIVA contains CIP2A exons 1 to 13 fused to 349 nucleotides from CIP2A intron 13. Intriguingly, the first 39 nucleotides of the NOCIVA-specific sequence are in the coding frame with exon 13 of CIP2A and code for a 13-amino acid peptide tail nonhomologous to any known human protein sequence. Therefore, NOCIVA translates to a unique human protein. NOCIVA retains the capacity to bind to B56α, but, whereas CIP2A is predominantly a cytoplasmic protein, NOCIVA translocates to the nucleus. Indicative of prevalent alternative splicing from CIP2A to NOCIVA in myeloid malignancies, AML and CML patient samples overexpress NOCIVA, but not CIP2A mRNA. In AML, a high NOCIVA/CIP2A mRNA expression ratio is a marker for adverse overall survival. In CML, high NOCIVA expression is associated with inferior event-free survival among imatinib-treated patients, but not among patients treated with dasatinib or nilotinib. CONCLUSIONS: We discovered a novel variant of the oncoprotein CIP2A and its clinical relevance in predicting tyrosine kinase inhibitor therapy resistance in myeloid leukemias.

Targeting of Micelles and Liposomes Loaded with the Pro-Apoptotic Drug, NCL-240, into NCI/ADR-RES Cells in a 3D Spheroid Model.

PURPOSE: To develop transferrin (Tf)-targeted delivery systems for the pro-apoptotic drug, NCL-240, and to evaluate the efficacy of this delivery system in ovarian cancer NCI/ADR-RES cells, grown in vitro in a 3D spheroid model. METHODS: Tf-targeted PEG-PE-based micellar and ePC/CHOL-based liposomal delivery systems for NCL-240 were prepared. NCI/ADR-RES cells were used to generate spheroids by a non-adhesive liquid overlay technique. Spheroid growth and development were monitored by size (diameter) analysis and H&E staining. The targeted formulations were compared to untargeted ones in terms of their degree of spheroid association and penetration. A cell viability analysis with NCL-240-loaded micelles and liposomes was performed to assess the effectiveness of Tf-targeting. RESULTS: Tf-targeted polymeric micelles and Tf-targeted liposomes loaded with NCL-240 were prepared. NCI/ADR-RES cells generated spheroids that demonstrated the presence of a distinct necrotic core along with proliferating cells in the spheroid periphery, partly mimicking in vivo tumors. The Tf-targeted micelles and liposomes had a deeper spheroid penetration as compared to the untargeted delivery systems. Cell viability studies using the spheroid model demonstrated that Tf-mediated targeting markedly improved the cytotoxicity profile of NCL-240. CONCLUSION: Transferrin targeting enhanced delivery and effectiveness of micelles and liposomes loaded with NCL-240 against NCI/ADR-RES cancer cells in a 3D spheroid model.