A novel canine kidney cell line model for the evaluation of neoplastic development: karyotype evolution associated with spontaneous immortalization and tumorigenicity

Abstract

The molecular mechanisms underlying spontaneous neoplastic transformation in cultured mammalian cells remain poorly understood, confounding recognition of parallels with the biology of naturally occurring cancer. The broad use of tumorigenic canine cell lines as research tools, coupled with the accumulation of cytogenomic data from naturally occurring canine cancers, makes the domestic dog an ideal system in which to investigate these relationships. We developed a canine kidney cell line, CKB1-3T7, which allows prospective examination of the onset of spontaneous immortalization and tumorigenicity. We documented the accumulation of cytogenomic aberrations in CKB1-3T7 over 24 months in continuous culture. The majority of aberrations emerged in parallel with key phenotypic changes in cell morphology, growth kinetics, and tumor incidence and latency. Focal deletion of CDKN2A/B emerged first, preceding the onset and progression of tumorigenic potential, and progressed to a homozygous deletion across the cell population during extended culture. Interestingly, CKB1-3T7 demonstrated a tumorigenic phenotype in vivo prior to exhibiting loss of contact inhibition in vitro. We also performed the first genome-wide characterization of the canine tumorigenic cell line MDCK, which also exhibited CDKN2A/B deletion. MDCK and CKB1-3T7 cells shared several additional aberrations that we have reported previously as being highly recurrent in spontaneous canine cancers, many of which, as with CDKN2A/B deletion, are evolutionarily conserved in their human counterparts. The conservation of these molecular events across multiple species, in vitro and in vivo, despite their contrasting karyotypic architecture, is a powerful indicator of a common mechanism underlying emerging neoplastic activity. Through integrated cytogenomic and phenotypic characterization of serial passages of CKB1-3T7 from initiation to development of a tumorigenic phenotype, we present a robust and readily accessible model (to be made available through the American Type Culture Collection) of spontaneous neoplastic transformation that overcomes many of the limitations of earlier studies.Electronic supplementary materialThe online version of this article (doi:10.1007/s10577-015-9474-8) contains supplementary material, which is available to authorized users.

Circulating let‐7g is down‐regulated in Bernese Mountain dogs with disseminated histiocytic sarcoma and carcinomas – a prospective study

“…These databases allow searching for experimentally validated miRNA–mRNA interactions based on the miRNA binding sites (by reporter assay, western blot, microarray and next‐generation sequencing experiments). The CDKN2A gene encodes multiple tumour suppressors, such as p16 INK4A and p14 ARF and down‐regulation of these suppressors has previously been suggested in different types of canine malignant neoplasia, including DHS . CDKN2A deletions have also been associated with histiocytic neoplastic disorders in humans and it is considered likely that loss of p16 INK4A and p14 ARF is important for the pathogenesis of human histiocytic sarcoma .…”

Evolution of Cancer Progression in the Context of Darwinism

“…The notion of a chromosomal role in cancer development was postulated by Theodore Boveri over 100 years ago (41). Current progress in molecular genetics and biotechnology such as fluorescent in situ hybridization, whole-genome profiling, comparative chromosomal mapping and synthetic chromosome approach, have allowed us a better understanding of the role of chromosomes in cancer evolution (41)(42)(43)(44). The arrangement of genetic materials in eukaryotic cells is significantly different from that of prokaryotic organisms in that it reflects the evolution-dependent higher stability of eukaryotic chromosomes and more complex regulation of gene activity (45).…”

Novel cell lines derived from Chinese hamster kidney tissue

“…In addition, cells have been efficiently immortalized in combination with cell cycle regulators [ 11 , 12 ]. Immortalization also alters cell properties, with changes in gene expression profiles and karyotypes [ 13 ]. Although spontaneous immortalization of renal cells such as MDCK [ 14 ] and Vero cells [ 15 ] has been reported, the mechanism of this process in such cells remains poorly understood, and their biological differences to naturally occurring cancer cells are also unclear.…”