Mutagenesis, Vol. 14, No. 5, 483-490,
September 1999
© 1999 UK Environmental Mutagen Society/Oxford University Press
Previous loss of chromosome 11 containing a suppressor locus increases radiosensitivity, neoplastic transformation frequency and delayed death in HeLa x fibroblast human hybrid cells
Radiation and Cancer Biology Laboratory, Department of Radiation Oncology, 975 West Walnut Street, IB-346, Indiana University School of Medicine, Indianapolis, IN 46202, USA
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Abstract |
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CGL1 (HeLa x fibroblast) hybrid cells have been utilized to study mechanisms of radiation-induced neoplastic transformation of human cells in vitro. Previous analysis has shown that loss of active tumor suppressor alleles on fibroblast chromosomes 11 and 14 may be required for radiation-induced neoplastic transformation of CGL1 cells. Loss of chromosome 11 alone was, therefore, found to be necessary but not sufficient for neoplastic transformation. We postulated that the loss of chromosome 11 may make the hybrid cells more susceptible to radiation-induced neoplastic transformation, since these cells have already undergone one of the required tumor suppressor loss events. Hybrid cells which have lost one copy of chromosome 11 were designated CON104(–11). CON104(–11) hybrid cells were found to have increased X-ray sensitivity and susceptibility to radiation-induced neoplastic transformation when compared with the parental CGL1 cells. In addition, the neoplastically transformed foci appear to arise earlier after radiation exposure in CON104(–11) versus CGL1 cells. Furthermore, the plating efficiency (PE) of the progeny of the irradiated CON104(–11) cells, growing in transformation flasks, is persistently lower than parental CGL1 cells during the 21 day assay period. The lower PE of the progeny of irradiated cells was attributed to the expression of delayed death/lethal mutations post-irradiation, a reflection of genomic instability. Taken together, the data indicate that previous loss of chromosome 11 may increase the radiation-induced genomic instability of the hybrid cells, leading to increased radiation sensitivity and neoplastic transformation potential. The data suggest that one possible function of the chromosome 11 tumor suppressor gene may be to help maintain genome stability after radiation damage.
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Introduction |
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Current models of multistage carcinogenesis indicate that loss of several tumor suppressor loci may be required for neoplastic progression (Boyd and Barrett, 1990
; Weinberg, 1995). The concepts of a mutator phenotype and genomic instability have been proposed to account for the multiple genetic changes required in carcinogenesis (Cheng and Loeb, 1993; Morgan et al., 1996; Lengauer et al., 1997; Mendonca et al., 1998a,b). The relevance of the order of gene loss in these models is only beginning to be appreciated. For example, loss of function of the p53 tumor suppressor gene may allow genomically unstable cells to acquire additional mutations required for neoplastic transformation without signaling removal of these cells via apoptosis (Smith and Fornace, 1995; Anthoney et al., 1996; Wahl et al., 1997).
Studies into the mechanism of radiation-induced neoplastic transformation in vitro, utilizing the HeLa x skin fibroblast hybrid cell CGL1 model, have shown that loss of tumor suppressor alleles on chromosomes 11 and 14 appear to be required for radiation-induced neoplastic transformation (Stanbridge et al., 1981; Mendonca et al., 1991, 1995, 1998a, , b; Redpath et al., 1992). These chromosomes are of fibroblast origin and are responsible for the observed tumor suppression in this system (Stanbridge et al., 1981). While these data support our proposal that tumor suppressor gene loss plays a role in radiation-induced human cancer, the individual functions of the chromosome 11 and 14 tumor suppressor loci remain unknown. We propose that studies of the effect of previous chromosome 11 or 14 loss on radiation-induced neoplastic transformation might give us insight into how these tumor suppressor loci control tumorigenicity.
Hybrid cell lines from irradiated populations of CGL1 cells were isolated which clearly demonstrated that loss of fibroblast chromosome 11 was necessary, but not sufficient, for neoplastic transformation (Mendonca et al., 1998a). We demonstrate here that CON104(–11) hybrid cells, which have lost a complete copy of fibroblast chromosome 11, are more sensitive to X-ray-induced cell killing and to radiation-induced neoplastic transformation when compared with the parental cell line CGL1. Furthermore, the loss of chromosome 11 results in an increase in the expression of delayed death or lethal mutations post-irradiation during the 21 day neoplastic transformation assay period. All three of these observations are interpreted to be a reflection of increased genomic instability in CON104(–11) cells after radiation exposure. The data suggest that one possible function of the chromosome 11 tumor suppressor gene may be to help maintain genome stability after radiation damage.
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Materials and methods |
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Parental human hybrid cell line CGL1
The parental cell line, CGL1, is a non-tumorigenic hybrid cell line obtained from the third serial subclone of ESH5 in methylcellulose. It was originally isolated from a fusion of the tumorigenic HeLa cell line D98/AH-2 and a non-tumorigenic normal human skin fibroblast cell (GM00077). This is described in detail elsewhere (Der and Stanbridge, 1981
; Stanbridge et al., 1981). CGL1 is non-tumorigenic when inoculated s.c. into nude mice, negative for expression of the HeLa tumor-associated antigen intestinal alkaline phosphatase (IAP) and is genetically stable (Mendonca et al., 1995, 1998b). It contains, on average, four copies of each chromosome, two of fibroblast and two of HeLa origin.
Isolation of IAP-negative irradiated control cells (CONs) and CON104 subclones
The CON cell lines were isolated from irradiated populations (7 Gy of 
-rays) after 10 days growth. They appear morphologically similar to the original CGL1 cell population. Subsequent chemical measurement of alkaline phosphatase (AP) activity and flow cytometry using a monoclonal antibody to p75-IAP confirmed them to be IAP-negative. CONs were used as irradiated controls and designated CONs 1–5 (Mendonca et al., 1991). Later passages of the original CON1, described previously (Mendonca et al., 1991, 1995, 1998a) were further subcloned to isolate hybrids which had lost one copy of fibroblast chromosome 11. Cytogenetic and molecular analyses were performed to identify subclones which had undergone loss of an entire fibroblast chromosome 11 but remained IAP-negative (Mendonca et al., 1995, 1998a). Three independent isolates of the CON1 cell line were selected and tested for X-ray sensitivity and radiation-induced neoplastic transformation frequency. For clarity, only data from one representative subclone, denoted CON104(–11), will be shown and discussed.
Cell culture, growth curves and plating efficiency determination
All cell culture stocks were maintained in Corning T-75 flasks in 15 ml of medium. Cell lines were grown in Eagle's modified minimum essential medium (Flow Laboratories) supplemented with 5% calf serum (JRH), 2 mM glutamine (Sigma), non-essential amino acids (Sigma), 20 mM sodium bicarbonate and 100 IU/ml penicillin (Sigma). Cells were incubated in a 37°C incubator in an atmosphere of 5% CO2 in air. Cell numbers for growth curves and plating efficiencies were performed as follows. Cells were removed from the flasks by incubating in 1x trypsin. When the cells had detached, they were resuspended in complete medium and an aliquot was counted using a Coulter counter. Appropriate cell numbers were plated for survival after 0–7 Gy of X-irradiation (see below). In vitro plating efficiencies, population doubling times and surviving fractions were determined by standard methods (Mendonca et al., 1989a, 1990b, 1991a,, b).
Determination of survival and neoplastic transformation frequencies for CON104(–11) versus parental CGL1
The non-tumorigenic, IAP-negative cell line CON104(–11) and the parental cell hybrid CGL1 were irradiated with 0, 1, 2, 3, 5 or 7 Gy of 250 KvP X-rays at a dose rate of 68.7 cGy/min and plated for cell survival and neoplastic transformation. The standard doses of 1–7 Gy of X-rays were utilized so that the complete dose–response relationship for neoplastic transformation of CON104(–11) and CGL1 could be compared. The cell numbers for each cell line were adjusted so that at the time of radiation exposure there are 1–1.5x106 cells/T-25 flask. After irradiation, the cells were replaced in the incubator for 6 h at 37°C to allow repair of potentially lethal damage. The cells were then trypsinized, counted and plated in six T-25 flasks for survival determination. After 7–10 days growth the colonies were fixed and stained with a solution containing 0.35% crystal violet in 35% ethanol. Only colonies with >50 cells were scored.
To measure the neoplastic transformation frequency for a particular dose for each cell line, we plated 30–60 T-75 flasks with 5000–50 000 cells/flask depending on the expected cell survival for each dose. The exact cell numbers plated were adjusted to achieve a viable cell density in the transformation flasks of 50 cells/cm2, which has become standard in this system for -ray and X-ray work (Redpath et al., 1987; Sun et al., 1988; M.S.Mendonca, unpublished results). After 21 days, the flasks were fixed and stained for neoplastically transformed foci by the Western Blue method (Mendonca et al., 1992) and the transformation frequencies determined for each dose. Briefly, on day 21 the medium was removed, the cultures were rinsed twice in phosphate-buffered saline (PBS), fixed with 2% formaldehyde in PBS for 20 min and then rinsed with PBS four times. The cells were stained with 2 ml of Western Blue (alkaline phosphatase detection reagent; Promega) for 7 min to stain foci of cells expressing the tumor-associated antigen p75/150 IAP. The reagent was removed and the flasks were rinsed with PBS and the blue foci were visually scored using a stereomicroscope. Neoplastic transformation frequencies were expressed as the total number of foci per surviving cells (Mendonca et al., 1993).
Kinetics of the appearance and number of cells per neoplastically transformed foci
To determine the onset and kinetics of IAP-expressing, neoplastically transformed foci after 7 Gy of X-rays for CGL1 and CON 104(–11), a standard transformation assay was performed. One hundred and fifty T-75 flasks were plated of each cell line so that 15 flasks of each could be fixed and stained by the Western Blue technique approximately every other day from day 4 to day 21. Groups of flasks from unirradiated cells for each cell line were also fixed and stained to determine the spontaneous background on the designated days. The transformation flasks were screened for foci with a stereomicroscope so that small foci were not missed. We have previously shown that neoplastically transformed foci with 10 cells or less can be consistently detected by this method (Mendonca et al., 1993). The number of cells in each focus was also counted with an inverted phase contrast microscope (Leitz). Results of this experiment were expressed as neoplastic transformation frequency over time. The experiments described above were performed three times except where noted.
Plating efficiencies of irradiated and non-irradiated CON104(–11) and parental CGL1 cells during the 21 day neoplastic transformation assay (delayed death assays)
Irradiated and non-irradiated plating efficiencies were measured during the 21 day neoplastic transformation assays (Mendonca et al., 1989a, 1990b, 1992). Changes in clonogenic plating efficiencies (PEs) of control and irradiated CGL1 and CON104(–11) cells in the transformation flasks on days 4, 6, 8, 11, 13, 15,18, 20 and 21 were performed as described (Mendonca et al., 1989, 1990, 1992). Briefly, the cells in 75 cm2 transformation flasks were trypsinized and counted. Irradiated and non-irradiated cells were plated at 100–500 cells/25 cm2 tissue culture flask, which contained 5 ml of pre-equilibrated medium (pH 7.2), six flasks per point. The PE flasks were incubated for 7–10 days to assess colony-forming ability. Colonies were stained and scored as described above. The experiments described were performed a minimum of three times, using six replicates per condition. Plating efficiencies for irradiated and control samples were calculated by dividing the average number of colonies in the six flasks by the number of cells initially plated (Redpath et al., 1987; Mendonca et al., 1991).
Data presentation and statistical analysis
Radiation-induced neoplastic transformation data for CGL1 and CON104(–11) were scored by counting the total number of foci within the T-75 flasks, as well as the total number of cells surviving each treatment protocol. Transformation frequency is expressed as the total number of foci/total number of surviving cells. When necessary a 
2 analysis was performed to compare the accumulated data for CGL1 with CON104(–11) to test for statistical significance. For transformation frequency, this was done using the total number of foci and the total number of cells surviving treatment (Mendonca et al., 1990, 1998b). Plating efficiencies or survival fractions at various dose points for CON104(–11) versus parental CGL1 cells were considered significantly different if the 95% confidence intervals (±2 SD) did not overlap.
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X-ray induced cell killing of CON104(–11) versus the parental CGL1 hybrid
Extensive subcloning and cytogenetic studies of the nontumorigenic control hybrid cell line (CON1) allowed us to isolate three cell lines which were missing an entire copy of human fibroblast chromosome 11 (Mendonca et al., 1998). The X-irradiation response of CON104(–11) versus CGL1 is presented in Figure 1
. CON104(–11) was more radiosensitive than the parental CGL1. The dose for 50% survival (D50) for Con104(–11) was 3.5 Gy, versus 5.0 Gy for CGL1. The difference in radiation response appeared to be due to a difference in the shoulder size of the survival curves. Graphical analyses confirmed that the D0 values of the curves were equivalent (2.8 ± 0.2 Gy) and the Dq values for CON104(–11) and CGL1 were significantly different at 1.5 ± 0.2 and 3.0 ± 0.2 Gy, respectively (P < 0.05). CON104(–11) was one of the three subclones isolated, but the data for this cell line are representative of all three isolates.
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Radiation-induced neoplastic transformation of CON104(–11) and the parental CGL1 hybrid versus X-ray dose
To examine whether a previous loss of either fibroblast chromosome 11 influenced the number of neoplastically transformed foci produced 21 days after radiation exposure, standard neoplastic transformation assays were performed with CON104(–11) and the parental CGL1. In Figure 2
, accumulated data of neoplastic transformation frequency versus X-ray dose for CON104(–11) and CGL1 are shown. More foci were produced at each dose and the transformation frequency of CON104(–11) was consistently higher than the parental cell line on day 21 post-irradiation. In Table I, the data from three independent neoplastic transformation experiments for CGL1 versus CON104(–11) after 7 Gy are shown. 2 analysis on the accumulated data at 7 Gy demonstrated that the neoplastic transformation frequency was significantly higher for CON104(–11) versus CGL1 (1.0x10–3 versus 4.7x10–4, respectively; P < 0.05). The increase in radiation-induced neoplastic transformation frequency at 5 Gy is also statistically significant (P < 0.05). The data indicate that CON104(–11) is more susceptible to X-ray-induced neoplastic transformation than the parental cell line CGL1 from which it was derived.
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Radiation-induced neoplastic transformation of CON104(–11) and the parental CGL1 hybrid versus time after 7 Gy of X-rays
To investigate whether the increased sensitivity of CON104 (–11) to radiation-induced neoplastic transformation seen on day 21 was also evident at earlier time points, a study of neoplastic transformation frequency versus time post-irradiation for CGL1 and CON104(–11) after 7 Gy is shown in Figure 3
. For CGL1 cells, the majority of neoplastically transformed foci did not begin to arise until after day 12 or 13 post-irradiation and were fully expressed by day 21. This delayed appearance of foci after X-ray exposure is in agreement with the previously published -ray foci kinetic data for CGL1 (Sun et al., 1988; Mendonca et al., 1993). In CON104(–11), increased neoplastic transformation frequencies (more foci) were evident by as early as day 6 post-irradiation and this increase in neoplastic transformation frequency above CGL1 cells persisted through day 18. The data indicate that radiation-induced foci arise earlier in CON104(–11) than in the parental CGL1 cells by at least 3–4 days (Figure 3).
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Another method to show the kinetics of foci development is to quantify the size distribution of foci after 7 Gy of X-rays at various times during the 21 day assay period (Mendonca et al., 1993
, 1998b). This was done by microscopically counting the number of neoplastically transformed IAP-positive cells per focus on specified days post-irradiation. In this system the foci develop from one or two IAP-positive neoplastically transformed cells and progressively grow in size during the 21 day assay (Mendonca et al., 1993, 1998b). IAP-positive cells per focus as a function of time post-irradiation for CON104(–11) and CGL1 are shown in Figure 4. Consistent with the results in Figure 3
, the data demonstrate that irradiated CON104(–11) cells developed more foci at earlier days than CGL1. Comparison of foci size on day 11 shows that in CON104(–11) there are six foci with >100 cells but there are only two foci with >100 cells in CGL1. On day 13 there are 12 CON104(–11) foci with >1000 cells (seven of which contain >2500 cells), but in CGL1 there are only four foci with >1000 cells, none of them with >1200 cells (Figure 4). Both the increase in numbers of foci and the tendency towards larger foci size on days 11 and 13 would be consistent with foci arising a few days earlier in CON104(–11). The data also demonstrate that the differences between CGL1 and CON104(–11) are not due to small foci being missed in either cell line since this method is capable of detecting foci containing <10 cells (Figure 4; Mendonca et al., 1993).
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Growth curves of CON104(–11) and CGL1 cells after 7 Gy of X-rays
To rule out that large differences in cell population doubling times of CON104(–11) versus CGL1 could be the reason for the difference in the timing of appearance of foci post-irradiation, growth curves of the irradiated (7 Gy) cells in the transformation flasks are shown in Figure 5
. The growth curves overlap during the exponential growth period through day 10, indicating similar population doubling times for CON104(–11) and CGL1 (20 ± 1 h) (Mendonca et al., 1991a). After day 10, the irradiated cells reach a steady-state plateau which is typical for hybrid cells growing under these conditions. However, the average density of CON104(–11) cells at the steady-state plateau, from days 10–20 post-irradiation, is significantly lower at 7.3x106 ± 0.9x106 cells/flask versus CGL1 cells, which average 13x106 ± 1.2x106 cells/flask (P < 0.05) during the same time period.
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Plating efficiency of CON104(–11) and CGL1 cells after 7 Gy of X-rays
A lower steady-state plateau of the irradiated hybrid cells in this transformation system has been attributed to increased delayed death due to genomic instability (Mendonca et al., 1993
, 1998b). This delayed death or lethal mutation has been previously described in irradiated CGL1 cells and is also manifested as a persistent reduction in the PE of replated cells from the irradiated transformation flasks during the 21 assay period (Mendonca et al., 1989a, 1993, 1998b). In Figure 6, the PE of X-irradiated (7 Gy) CON104(–11) and CGL1 cells from transformation flasks are shown. The data indicate that after 7 Gy of X-rays, the PE of irradiated CON104(–11) is consistently lower than that observed for the parental CGL1 during the 21 day assay period. In contrast, the replated PE of the unirradiated control CON104(–11) cells in the transformation flasks was 0.85 ± 0.07, which is within the range consistently observed for CGL1 (Mendonca et al., 1989, 1991, 1993).
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Discussion |
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Understanding the basis of radiation-induced neoplastic transformation of human cells requires identification of the potential gene targets for mutagenesis and an understanding of the mechanisms by which the genes are either activated or inactivated after X-ray exposure. Analysis of HeLa x fibroblast hybrid cell lines suggested the presence of functional tumor suppressor loci on chromosomes 11 and 14 of fibroblast origin (Stanbridge et al., 1981
; Kaelbling and Klinger, 1986; Saxon et al., 1986; Srivatsan et al., 1986). We have recently shown that the loss of alleles on both fibroblast chromosomes 11 and 14 is evident after radiation-induced neoplastic transformation of the human hybrid cells (Mendonca et al., 1995, 1998a). These data represented one of the first direct demonstrations that radiation-induced loss of chromosomes containing active tumor suppressor loci may be involved in radiation-induced human cancer (Mendonca et al., 1995, 1998a). In this study we investigated the consequence of the previous loss of a fibroblast chromosome 11 on X-irradiation sensitivity and neoplastic transformation potential in the hybrid neoplastic transformation system to gain some insight into how this tumor suppressor locus may control tumorigenicity in these cells.
Loss of chromosome 11 increases X-ray-induced cell killing
We first investigated the effect of the previous loss of a fibroblast chromosome 11 on X-irradiation sensitivity in the hybrid neoplastic transformation system. The data indicate that the loss of a fibroblast chromosome 11 increases the X-ray sensitivity of the CON104(–11) cells versus CGL1 (Figure 1). The difference is due to a decrease in the shoulder of the CON104(–11) survival curve. This has been interpreted to be a reflection of decreased repair capacity in many mammalian cells in vitro and in vivo (Elkind, 1984; Iliakis, 1988; Mendonca et al., 1989b, 1990a). Recently, some of the genes involved in repair and X-irradiation sensitivity have been identified and a partial list include p53, p21, ATM, Ku70/80 and DNA-PK (reviewed in Rosen et al., 1999). Interestingly, the ATM gene is located on chromosome 11q22–23. Mutation of this gene has been implicated in the DNA repair deficiency syndrome ataxia telangiectasia (reviewed in Meyn, 1995). The HeLa/cervical cancer tumor suppressor locus, which we are currently in the process of cloning with our collaborators, is located on chromosome 11q13 (Mendonca et al., in preparation). Whether the ATM gene, the HeLa/cervical cancer gene or changes in other genes on chromosome 11 can explain the increased radiosensitivity of CON104(–11) will require additional studies. However, our data do suggest that the loss of chromosome 11 alleles increases the radiation sensitivity of the hybrid cells.
Loss of chromosome 11 increases the number of X-ray-induced neoplastically transformed foci and shortens the lag time for their appearance
Standard neoplastic transformation assays with CON104(–11) and CGL1 were performed to investigate whether previous loss of chromosome 11 also increased the sensitivity to X-ray-induced neoplastic transformation. The data show that more foci are produced at each X-ray dose and the transformation frequency is significantly higher in CON104(–11) than CGL1 after 7 Gy of X-rays (Figure 2 and Table I; P < 0.05). In addition, a study of neoplastic transformation frequency versus time post-irradiation for CGL1 and CON104(–11) after 7 Gy of X-rays indicates that foci in CON104(–11) appear a few days earlier than in CGL1 (Figure 3). Analysis of the size distribution of foci after 7 Gy of X-rays confirmed that foci arise a few days earlier in CON104(–11) cells than foci in CGL1 cells (Figure 4). Growth curves indicate that large differences in cell population doubling times for the two cell lines are not evident and therefore cannot explain the difference in the timing of the appearance of foci post-irradiation (Figure 5).
Loss of chromosome 11 increases the expression of delayed death post-irradiation in CON104(–11) cells
Our initial hypothesis was that an increased sensitivity of CON104(–11) to radiation-induced neoplastic transformation may be due to fewer genetic loss events being required. However, that explanation is difficult to reconcile with the increased X-ray-induced cell killing observed in CON104(–11) cells. We have previously shown that the delayed appearance of foci in CGL1 correlates with a persistently reduced PE of irradiated cells during the 21 day assay period (Mendonca et al., 1993, 1995, 1998b). A number of investigators have observed this lower PE in the progeny of irradiated cells weeks to months after radiation exposure, which we and others attribute to the expression of delayed death or lethal mutations (Seymour et al., 1986; Mendonca et al., 1989a; Chang and Little, 1991; Seymour and Mothersill, 1992). We have proposed that the expression of delayed death in our system is a result of genomic instability (Mendonca et al., 1993). Work from a number of laboratories has shown that the expression of chromosome and chromatid aberrations after radiation exposure can be delayed and last for several generations post-irradiation. This is also believed to be a consequence of radiation-induced genomic instability (reviewed in Morgan et al., 1996; Mothersill et al., 1996; Murnane, 1996).
The lower steady-state plateau for irradiated CON104(–11) versus irradiated CGL1, seen in the growth curves in Figure 5, suggests reduced clonogenic potential in the CON104(–11) cells. In Figure 6, a plating efficiency versus time post-irradiation study with CON104(–11) and CGL1 indicates that after 7 Gy of X-rays the PE of CON104(–11) is consistently lower than that observed for CGL1. This would suggest an increased expression of delayed death in the CON104(–11) progeny, which correlates with the higher radiation-induced transformation frequency. Therefore, another possible explanation for the increased susceptibility of CON104(–11) to X-irradiation is that the lower irradiated PE of CON104(–11) is a reflection of increased genomic instability post-irradiation, which increases the probability of chromosome loss leading either to neoplastic transformation or, in many cases, to increased cell killing.
Tumor suppressor loci and genomic instability
The concepts of a mutator phenotype and associated genomic instability being involved in neoplastic progression have been proposed to account for the multiple genetic changes required in carcinogenesis (Cheng and Loeb, 1993; Lengauer et al., 1997). We have recently shown a correlation between the expression of delayed death and the onset of delayed apoptosis in CGL1 cells during the 21 day neoplastic transformation assay period (Mendonca et al., 1999). Limoli and co-workers have recently shown a link between reproductive failure, apoptosis and compromised genomic integrity in a human/hamster cell line (Limoli et al., 1998).
Our long-term goal is to clone both the chromosome 11 and 14 tumor suppressor loci in these cells. However, until that time analysis of cells which have undergone previous chromosome 11 and 14 loss may allow investigation of their individual phenotypes. We have shown here that previous loss of chromosome 11 increases the sensitivity of CON104(–11) to radiation-induced cell killing and neoplastic transformation. The significance of previous chromosome 14 loss on radiation sensitivity and neoplastic transformation is currently under investigation.
Data from studies of both the retinoblastoma and p53 tumor suppressor genes indicate that regulation of gene expression involved in cell growth may be an important function of tumor suppressor genes (Hinds, 1995; Ko and Prives, 1996). A number of tumor suppressor genes are involved in the regulation of DNA damage checkpoints and therefore in the control of genomic instability (Baylin, 1997; O'Connor, 1997; Wahl et al., 1997). The data reported here suggest that the HeLa/cervical cancer tumor suppressor locus on chromosome 11 may be involved in the control of genomic instability post-irradiation.
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Acknowledgments |
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This work was supported by start-up funds awarded to M.S.M. from the Department of Radiation Oncology, Indiana University School of Medicine, and in part by grant IRG-84-002-14 from the American Cancer Society awarded to M.S.M.
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Notes |
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1 To whom correspondence should be addressed. Tel: +1 317 278 0404; Fax: +1 317 278 0405; Email: mmendonc{at}iupui.edu
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Received on January 27, 1999; accepted on May 14, 1999.
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