An apoptosis-enhancing drug overcomes platinum resistance in a tumour-initiating subpopulation of ovarian cancer
Authors: J Lu
S Memarzadeh
D M Janzen
J A Salehi
M Pellegrini
E Tiourin
D Y Paik
Journal: Nature communications (2015)
Discipline: Multidisciplinary
Keywords: High-grade serous ovarian cancers; carboplatin resistance; CA125-negative population; cancer initiating cells; homologous recombination DNA repair
Affiliations: Department of Obstetrics and Gynecology, David Geffen School of Medicine, University of California, Los Angeles, Los Angeles, California 90095, USA., Department of Molecular, Cell and Developmental Biology, University of California, Los Angeles, Los Angeles, California 90095, USA., Eli and Edythe Broad Center of Regenerative Medicine and Stem Cell Research, University of California, Los Angeles, Los Angeles, California 90095, USA., The VA Greater Los Angeles Health Care System, Los Angeles, California 90073, USA.

Abstract:  High-grade serous ovarian cancers (HGSCs) are deadly malignancies that relapse despite carboplatin chemotherapy. Here we show that 16 independent primary HGSC samples contain a CA125-negative population enriched for carboplatin-resistant cancer initiating cells. Transcriptome analysis reveals upregulation of homologous recombination DNA repair and anti-apoptotic signals in this population. While treatment with carboplatin enriches for CA125-negative cells, co-treatment with carboplatin and birinapant eliminates these cells in HGSCs expressing high levels of the inhibitor of apoptosis protein cIAP in the CA125-negative population. Birinapant sensitizes CA125-negative cells to carboplatin by mediating degradation of cIAP causing cleavage of caspase 8 and restoration of apoptosis. This co-therapy significantly improves disease-free survival in vivo compared with either therapy alone in tumour-bearing mice. These findings suggest that therapeutic strategies that target CA125-negative cells may be useful in the treatment of HGSC. 

Retracted: No

PubPeer comments:
 Comment on bioRxiv: Comments from Bruce Wang, Ludan Zhao, Matt Fish, Catriona Logan, Roel Nusse In this manuscript, the authors raise a number of issues related to our publication from 2015 [1]. Most importantly, they report varying degrees of labeling of Axin2 expressing cells in the liver as well as dominant phenotypes in mice in which one or both Axin2 alleles are disrupted by insertion of the CreERT2 cassette. May et al. report that in their hands, Axin2-CreERT2 homozygous mice could not be generated at all and that Axin2-CreERT2 heterozygous mice are born at sub-Mendelian ratios. In various labs, including our own, mice that are homozygous mutant for Axin2 have been used and characterized. The first such mice were generated by the Birchmeier lab [2], which have a LacZ insert at the exact same nucleotide position where we later inserted the CreERT2 cassette [3]. Both Axin2-CreERT2 and Axin2-LacZ, as well as other 5u2019 Axin2 knock-in strains, are considered to be null alleles of Axin2. Nevertheless, homozygous Axin2-LacZ animals are viable and fertile with relatively minor phenotypes reported in the literature (including a shortened head, abnormal eye morphology and enhanced osteoblast differentiation; see http: www.informatics.jax..... In our lab, all records from previous personnel spanning 2012-2016 have demonstrated that Axin2-CreERT2 homozygotes could be obtained despite mild phenotypes, and these animals were generated at expected frequencies. Axin2-CreERT2 heterozygotes bred to wild type mice also gave progeny at expected Mendelian ratios. Current crosses of Axin2-heterozygotes to wild-type continue to give expected ratios. That being said, it is conceivable that rederivation and backcrossing of Axin2-CreERT2 animals to different backgrounds and their maintenance in different facilities might change the penetrance of Axin2 loss-of-function phenotypes via as of yet unknown modifier loci or other, non-genetic variables. This was recently described by the van Amerongen lab [4], which reported non-Mendelian offspring ratios from Axin2-CreERT2 mice (two rederivations removed from the line at our facility and three rederivations removed from the mice originally described in [3] or the mice deposited at Jackson labs) in heterozygous backcrosses to C57Bl 6 but not FVB N (see Supplementary Table 2 in [4]). In August of 2017, we added this information to the description of the Axin2-CreERT2 mouse line on the Jackson Laboratory website and high (5 x 4mg tamoxifen 25gm body weight) tamoxifen dosing aimed at low and higher frequencies of labelling. With both doses we continued to see long-term expansion of labeled cells more than 300 days after initial tamoxifen dosing. Unlike May, with the high dose labeling we saw no displacement of pericentral labeled cells by unlabeled cells and this was the dosing used to quantify expansion of labeled cells. We interpret this as evidence that the Axin2 labeled cells in zone 3 expand, more so than other cells, and that lingering tamoxifen (which should be long gone from the system) was not the cause of the increased labeling over time. Moreover, in our original publication [1] we wondered whether Axin2 heterozygosity affected our results but found no evidence that this was the case. First, we measured cell cycle parameters by EdU incorporation in heterozygous Axin2-CreERT2 mice compared to wild type and found no difference in proliferation rate with EdU labeling for 7 days. While we did not discriminate between male and female animals in Wang et al. [1], the results by May et al. suggest that this could be important. This is also part of a larger debate and movement towards appreciating sex as a biological variable. Particularly in the case of stem cells, a difference between males and females has been reported earlier [5], and it is tempting to speculate that this might actually be directly due to a difference in Wnt beta-catenin pathway activity, as the results by May et al. would seem to suggest. In addition to our lineage tracing experiments with Axin2-CreERT2, we also used the Axin2-rtTA transgenic mouse in which rtTA expression is driven by an Axin2 promoter fragment and where the wildtype Axin2 gene locus is kept intact. Using this model, we found a similar pattern of initial labeling of pericentral hepatocytes followed by expansion of labeled cells as with Axin2-CreERT2 mice, further supporting the importance of this cell population in particular, and arguing against reduced gene dosage of Axin2 as the reason for the observed proliferation of zone 3 cells and their progeny. However, we acknowledge that these were relatively short-term experiments. Cre lox mediated lineage tracing is a powerful method, but there are challenges and limitations in its design and interpretation [6, 7] which could affect our own as well as May et al.u2019s experiments. We emphasize that apart from our data, independent DNA label dilution experiments performed almost twenty years ago also indicate that the pericentral area (or zone 3) has a higher rate of DNA replication compared to other zones [8]. In the adult liver, both Wnt signaling and expression of the Wnt target gene Tbx3, known to be essential for embryonic hepatocyte proliferation [9, 10], continue to be active in pericentral cells [11]. Moreover, in adult liver tissue, pericentral cells express relatively high levels of c-Myc and CyclinD, hallmarks of cells that are active in the cell cycle [11]. In experimental models of mouse hepatocellular carcinomas, the cells of origin of the tumors are located pericentrally [12, 13]. We therefore believe our findings are consistent with the important role of Wnt beta-catenin signaling in postnatal growth of the liver [14] and for the repair of liver tissue after chemical injury [15, 16]. Whether pericentral cells in adult livers are quiescent or divide and at which rate will depend on the specific context the cells are in and this will likely be affected by uncontrolled variables and environmental parameters. References 1. 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