Suppression of Chromosome Instability Limits Acquired Drug Resistance.

Numerical chromosome instability, or nCIN, defined as the high frequency of whole chromosome gains and losses, is prevalent in many solid tumors. nCIN has been shown to promote intratumor heterogeneity and corresponds with tumor aggressiveness, drug resistance, and tumor relapse. Although increased nCIN has been shown to promote the acquisition of genomic changes responsible for drug resistance, the potential to modulate nCIN in a therapeutic manner has not been well explored. Here we assess the role of nCIN in the acquisition of drug resistance in non-small cell lung cancer. We show that the generation of whole chromosome segregation errors in non-small cell lung cancer cells is sensitive to manipulation of microtubule dynamics and that enhancement of chromosome cohesion strongly suppresses nCIN and reduces intratumor heterogeneity. We demonstrate that suppression of nCIN has no impact on non-small cell lung cancer cell proliferation in vitro nor in tumor initiation in mouse xenograft models. However, suppression of nCIN alters the timing and molecular mechanisms that drive acquired drug resistance. These findings suggest mechanisms to suppress nCIN may serve as effective cotherapies to limit tumor evolution and sustain drug response.

Suv420 enrichment at the centromere limits Aurora B localization and function.

Centromere structure and function are defined by the epigenetic modification of histones at centromeric and pericentromeric chromatin. The constitutive heterochromatin found at pericentromeric regions is highly enriched for H3K9me3 and H4K20me3. Although mis-expression of the methyltransferase enzymes that regulate these marks, Suv39 and Suv420, is common in disease, the consequences of such changes are not well understood. Our data show that increased centromere localization of Suv39 and Suv420 suppresses centromere transcription and compromises localization of the mitotic kinase Aurora B, decreasing microtubule dynamics and compromising chromosome alignment and segregation. We find that inhibition of Suv420 methyltransferase activity partially restores Aurora B localization to centromeres and that restoration of the Aurora B-containing chromosomal passenger complex to the centromere is sufficient to suppress mitotic errors that result when Suv420 and H4K20me3 is enriched at centromeres. Consistent with a role for Suv39 and Suv420 in negatively regulating Aurora B, high expression of these enzymes corresponds with increased sensitivity to Aurora kinase inhibition in human cancer cells, suggesting that increased H3K9 and H4K20 methylation may be an underappreciated source of chromosome mis-segregation in cancer. This article has an associated First Person interview with the first author of the paper.

Live Cell Imaging to Assess the Dynamics of Metaphase Timing and Cell Fate Following Mitotic Spindle Perturbations.

Live cell time-lapse imaging is an important tool in cell biology that provides insight into cellular processes that might otherwise be overlooked, misunderstood, or misinterpreted by the fixed-cell analysis. While the fixed cell imaging and analysis is robust and sufficient to observe cellular steady-state, it can be limited in defining a temporal order of events at the cellular level and is ill-equipped to assess the transient nature of dynamic processes including mitotic progression. In contrast, live cell imaging is an eloquent tool that can be used to observe cellular processes at the single-cell level over time and has the capacity to capture the dynamics of processes that would otherwise be poorly represented in fixed cell imaging. Here we describe an approach to generate cells carrying fluorescently labeled markers of chromatin and microtubules and their use in live cell imaging approaches to monitor metaphase chromosome alignment and mitotic exit. We describe imaging-based techniques to assess the dynamics of spindle formation and mitotic progression, including the identification of cells at various stages in mitosis, identification and tracking of mitotic defects, and analysis of spindle dynamics and mitotic cell fate following the treatment with mitotic inhibitors.