Inhibition of Pol I transcription treats murine and human AML by targeting the leukemia-initiating cell population.

Despite the development of novel drugs, the prospects for many patients with acute myeloid leukemia (AML) remain dismal. This study reveals that the selective inhibitor of RNA polymerase I (Pol I) transcription, CX-5461, effectively treats aggressive AML, including mixed-lineage leukemia-driven AML, and outperforms standard chemotherapies. In addition to the previously characterized mechanism of action of CX-5461 (ie, the induction of p53-dependent apoptotic cell death), the inhibition of Pol I transcription also demonstrates potent efficacy in p53null AML in vivo. This significant survival advantage in both p53WT and p53null leukemic mice treated with CX-5461 is associated with activation of the checkpoint kinases 1/2, an aberrant G2/M cell-cycle progression and induction of myeloid differentiation of the leukemic blasts. The ability to target the leukemic-initiating cell population is thought to be essential for lasting therapeutic benefit. Most strikingly, the acute inhibition of Pol I transcription reduces both the leukemic granulocyte-macrophage progenitor and leukemia-initiating cell (LIC) populations, and suppresses their clonogenic capacity. This suggests that dysregulated Pol I transcription is essential for the maintenance of their leukemia-initiating potential. Together, these findings demonstrate the therapeutic utility of this new class of inhibitors to treat highly aggressive AML by targeting LICs.

CX-5461 Preferentially Induces Top2α-Dependent DNA Breaks at Ribosomal DNA Loci.

While genotoxic chemotherapeutic agents are among the most effective tools to combat cancer, they are often associated with severe adverse effects caused by indiscriminate DNA damage in non-tumor tissue as well as increased risk of secondary carcinogenesis. This study builds on our previous work demonstrating that the RNA Polymerase I (Pol I) transcription inhibitor CX-5461 elicits a non-canonical DNA damage response and our discovery of a critical role for Topoisomerase 2α (Top2α) in the initiation of Pol I-dependent transcription. Here, we identify Top2α as a mediator of CX-5461 response in the murine Eµ-Myc B lymphoma model whereby sensitivity to CX-5461 is dependent on cellular Top2α expression/activity. Most strikingly, and in contrast to canonical Top2α poisons, we found that the Top2α-dependent DNA damage induced by CX-5461 is preferentially localized at the ribosomal DNA (rDNA) promoter region, thereby highlighting CX-5461 as a loci-specific DNA damaging agent. This mechanism underpins the efficacy of CX-5461 against certain types of cancer and can be used to develop effective non-genotoxic anticancer drugs.

Changes in long-range rDNA-genomic interactions associate with altered RNA polymerase II gene programs during malignant transformation.

The three-dimensional organization of the genome contributes to its maintenance and regulation. While chromosomal regions associate with nucleolar ribosomal RNA genes (rDNA), the biological significance of rDNA-genome interactions and whether they are dynamically regulated during disease remain unclear. rDNA chromatin exists in multiple inactive and active states and their transition is regulated by the RNA polymerase I transcription factor UBTF. Here, using a MYC-driven lymphoma model, we demonstrate that during malignant progression the rDNA chromatin converts to the open state, which is required for tumor cell survival. Moreover, this rDNA transition co-occurs with a reorganization of rDNA-genome contacts which correlate with gene expression changes at associated loci, impacting gene ontologies including B-cell differentiation, cell growth and metabolism. We propose that UBTF-mediated conversion to open rDNA chromatin during malignant transformation contributes to the regulation of specific gene pathways that regulate growth and differentiation through reformed long-range physical interactions with the rDNA.

Targeting tumors with nanobodies for cancer imaging and therapy.

The use of monoclonal antibodies has revolutionized both cancer therapy and cancer imaging. Antibodies have been used to directly inhibit tumor cell proliferation or to target drugs to tumors. Also in molecular imaging, monoclonal antibodies have found their way to the clinic. Nevertheless, distribution within tumors is hampered by their size, leading to insufficient efficacy of cancer treatment and irregular imaging. An attractive alternative for monoclonal antibodies are nanobodies or VHHs. These are the variable domain of heavy-chain antibodies from animals from the Camelidae family that were first discovered in 1993. Stimulated by the ease of nanobody selection, production, and low immunogenicity potential, a number of nanobodies specific to different disease-related targets have been developed. For cancer therapy, nanobodies have been employed as antagonistic drugs, and more recently, as targeting moieties of effector-domaINS and of drug delivery systems. In parallel, nanobodies have also been employed for molecular imaging with modalities such as nuclear and optical imaging. In this review, we discuss recent developments in the application of nanobodies as targeting moieties in cancer therapy and cancer imaging. With such a wide range of successful applications, nanobodies have become much more than simple antagonists.