Size Does Matter: Why Polyploid Tumor Cells are Critical Drug Targets in the War on Cancer

Abstract

Tumor evolution presents a formidable obstacle that currently prevents the development of truly curative treatments for cancer. In this perspective, we advocate for the hypothesis that tumor cells with significantly elevated genomic content (polyploid tumor cells) facilitate rapid tumor evolution and the acquisition of therapy resistance in multiple incurable cancers. We appeal to studies conducted in yeast, cancer models, and cancer patients, which all converge on the hypothesis that polyploidy enables large phenotypic leaps, providing access to many different therapy-resistant phenotypes. We develop a flow-cytometry based method for quantifying the prevalence of polyploid tumor cells, and show the frequency of these cells in patient tumors may be higher than is generally appreciated. We then present recent studies identifying promising new therapeutic strategies that could be used to specifically target polyploid tumor cells in cancer patients. We argue that these therapeutic approaches should be incorporated into new treatment strategies aimed at blocking tumor evolution by killing the highly evolvable, therapy-resistant polyploid cell subpopulations, thus helping to maintain patient tumors in a drug sensitive state.

Keywords: aneuploidy; cancer stem cell; chromosomal instability; hyperdiploidy; polyploidy; therapy resistance; tumor evolution; tumor initiation.

Figure 1

An improved flow-cytometry assay for measuring the prevalence of polyploidy in tumors cell populations. (A) Tumor cell samples are spiked with carboxyfluorescein diacetate succinimidylester (CFSE) stained primary neonatal foreskin fibroblasts (NFF) diploid control. The CFSE-negative tumor cells shown in blue are readily gated from the CFSE-high NFF diploid controls, shown in red. (B) DNA content of the Glioblastoma tumor cells (blue histogram) overlayed onto the NFF diploid control histogram, shown in red. Most Glioblastoma cell lines that we have studied are aneuploid with a slightly hyperdiploid DNA content, and contain a small sub-population of cells that are near-tetraploid with respect to the tumor bulk population (i.e., pseudo-polyploid). (C) A typical polyploidy flow-cytometry assessment utilized by many cancer cell biologists, who use the proportion of live single cells with greater than 4n DNA (shown within the red gate) as being representative of the total pseudo-polyploid population. In this example, 4.5% of the total cells are classed as pseudo-polyploid. (D) The same tumor sample assessed for pseudo-polyploidy using Cyclin-B1 staining to discriminate between the G₂/M (the Cyclin-B1 high cells with a 4n DNA content) population of the pseudo-diploid bulk, from the pseudo-diploid G₀/G₁ population (the Cyclin-B1 low cells with a 4n DNA content). The pseudo-polyploid gate (shown in red) identifies both the cycling and the non-cycling pseudo-polyploid tumor cells, which make up approximately 22% of the total tumor cell population. (E) Ten low-passage primary patient glioblastoma cell lines, grown under serum-free tumorsphere conditions, assessed for pseudo-polyploidy using the Cyclin-B1 gating strategy from (D). In eight lines, the dominant cell population was aneuploidy with a near-diploid DNA content, with a sub-population of pseudo-polyploid cells that made up 5–38% of the total cell population. In contrast, two glioblastoma lines were pre-dominantly pseudo-polyploid (65 and 78%), with a small of near-diploid sub-population. Detailed staining protocols are provided in Ref. (110).

Figure 2

An integrated model of tumor evolution highlighting potential roles of polyploidy during cellular transformation. Here, we present a simplified view of disease progression, highlighting the role of polyploidy in overcoming selection pressures to drive the evolution of cellular transformation. The fist selections pressures pre-malignant lesions must overcome are those of apoptosis, senescence and terminal differentiation. Polyploidy enables adaptation to these barriers by silencing p53 remove p53-dependent pro-apoptotic and senescence signaling, rewiring the DNA-damage response to suppress p53-independent senescence programs, and enabling acquisition of primitive stem-cell phenotypes. Once a proliferative state is reached, polyploidy increases the acquisition of transforming mutations by increasing chromosomal instability and buffering the proliferative cells against the effects of deleterious mutations. Polyploidy also increases glycolysis, enabling survival in low oxygen environments, and enables EMT and the generation of invasive and metastatic phenotypes. Polyploid cells provide multiple mechanisms of therapy resistance, buffer the cancer genome against deleterious mutation resulting from genotoxic therapies, and generate primitive tumor-initiating phenotypes that are capable of driving disease recurrence. Throughout this process, tumor heterogeneity and karyotypic complexity increases, which in turn increase the heterogeneity and evolutionary capacity of the tumor.