Radiosensitization and apoptosis - Nature

Oncogene (1998) 17, 3359 ± 3363 ã 1998 Stockton Press All rights reserved 0950 ± 9232/98 $12.00 http://www.stockton-press.co.uk/onc

Radiosensitization and apoptosis Ruth J Muschel*,1, Daniel E Soto2, WG McKenna2 and Eric J Bernhard2 1

Department of Pathology and Lab Medicine, University of Pennsylvania, Philadelphia, Pennsylvania 19104, USA; 2Department of Radiation Oncology, University of Pennsylvania, Philadelphia, Pennsylvania 19104, USA

The toxicity of radiation to living tissues was discovered soon after the discovery of radioactivity itself and this toxicity is the basis for cancer therapy with radiation. Although this mode of therapy is often e€ective, its success is far from assured. One major diculty in the implementation of radiotherapy is that normal tissues are also sensitive to killing by radiation so that treatment is often limited by the tolerance of normal tissues for radiation. Thus methods that sensitize tumor cells while sparing normal tissues could potentially lead to greater success with radiation as a therapy. Oncogenes are frequently altered in tumors, but are not in normal tissue making them potential targets for altering radiosensitivity and apoptosis in tumors. Keywords: radiation; X-rays; cell cycle; radiosensitization; apoptosis

E€ect of X-rays on cells Perhaps surprisingly the characteristics of radiation induced cell death are not the same for all cells. Since Kerr and Wyllie coined the term apoptosis, the distinctions between necrosis and apoptosis have been highlighted as two distinct forms of cell death (Kerr et al., 1972; Wyllie et al., 1980). Necrosis may occur after radiation although this is relatively rare. After irradiation some cell types indeed do die predominantly by apoptosis. Others however show little evidence of apoptosis. Even among cells in which apoptosis is readily observed the pattern or timing of apoptosis is dramatically di€erent between cell types. Highly radiosensitive tissues containing cells such as thymocytes, intestinal crypt cells, or germ cells undergo apoptosis within 8 ± 24 h of exposure to radiation (Clarke et al., 1993; Hasegawa et al., 1997; Hendry et al., 1982, 1996; Ijiri and Potten, 1983; Ohyama et al., 1985; Yamada and Ohyama, 1988). Before the general use of the term apoptosis, radiation biologists termed this form of death interphase death because the loss of viability preceded any cell division. Interphase death has the characteristics of apoptotic cell death including caspase activation, formation of DNA ladders, DNA cleavage, and nuclear fragmentation. For thymocytes and for colonic crypt cells, but not for crypt cells of the small intestine, the induction of apoptosis after irradiation is dependent on p53 and the striking radiosensitivity of these cells is also dependent upon p53. Both small intestinal crypt cells and more mature

*Correspondence: RJ Muschel

T cells undergo a rapid apoptotic death after irradiation, yet in these cases apoptosis appears to be independent of p53 (Clarke et al., 1993; Hendry et al., 1996, 1997; Lowe et al., 1993b; Merritt et al., 1997). The situation is more complex for other cell types including many tumor cells. After radiation apoptosis often occurs rapidly (within 24 h) in tumors derived from tissues that are highly radiosensitive including lymphomas or seminomas (derived from germ cells). However for the majority of tumor cells in culture, apoptosis does not occur soon after irradiation (Dewey et al., 1995; Harms-Ringdahl et al., 1996; Meyn, 1997; Radford, 1991; Radford et al., 1994). In these cases apoptosis may occur, but only after cell division and often after multiple cell divisions. These cells may undergo delayed apoptosis after radiation yet not be particularly radiosensitive. Primary ®broblasts do not undergo apoptosis within 48 h of irradiation. Instead they arrest in G1, an arrest that is p53 dependent (Di Leonardo et al., 1994; Linke et al., 1997). It has been suggested that this arrest is characteristic of terminal di€erentiation by ®broblasts (Bayreuther et al., 1988; Lara et al., 1996; Rodemann et al., 1991, 1996). Other cell types may show a division delay with a pronounced prolongation of G2, but subsequently continue to divide for several cycles. This pattern occurs in a variety of tumor cells including Hela cells. For up to 48 h after irradiation very little if any apoptosis may be seen in marked contrast to the e€ect irradiation has on thymocytes or many lymphoma cell lines that undergo apoptosis rapidly at 8 ± 10 h after irradiation. Following a pronounced prolongation of G2, these cells resume cycling for several divisions, even as many as ®ve generations after exposure to ionizing radiation (Hurwitz and Tolmach, 1969; Thompson and Suit, 1967, 1969). Eventually proliferation ceases resulting in a reproductive death that can sterilize a tumor, but not in a rapid metabolic death (Sinclair, 1964). This e€ect is seen as a small or abortive colony. The blockade of proliferation without loss of viability should be sucient for cancer therapy. Although apoptosis is not seen immediately, it may occur later (Radford et al., 1994; Tauchi and Sawada, 1994). Often the peak of apoptosis in vitro will be 48 h after irradiation of tumor cell lines (Bernhard et al., 1996b; Guo et al., 1997; McKenna et al., 1996; Vidair et al., 1996). However apoptosis can be detected at even later times. Vidair et al. (1996) tracked immortalized rat embryo ®broblasts by intravital microscopy for 6 days after irradiation. Some of the daughter cells were seen to undergo apoptosis after four divisions and up to 6 days after irradiation. The continuation of cell division for several cycles after the lethal injury and the possibility in some cases of radiation inducing terminal di€erentiation (a state that

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results in blocked proliferation, but not cell death) means that metabolic viability is not an adequate assay to judge radiation survival. Hence radiation survival is usually evaluated in assays that depend upon full colony formation such as clonogenic survival assays or dilution assays in microtiter plates; both assays that require colony formation and in which abortive colonies are not scored. Radiation survival The extent of apoptosis after radiation and of radiation survival has been shown to be in¯uenced by a variety of genes, but few have been used in strategies aimed at altering radiation survival. While the expression of p53 clearly a€ects apoptosis and survival in radiosensitive cell types such as thymocytes, its e€ect on less sensitive, more apoptosis resistant cells types is variable. Some tumor cells are in¯uenced in their response to radiation by the presence of wild type p53 while others are not (Bristow et al., 1996, 1998; Chang et al., 1997; DeWeese et al., 1997; Haas-Kogan et al., 1996; Lee and Bernstein, 1993; Slichenmyer et al., 1993; Tsang et al., 1995; Wang et al., 1996; Yount et al., 1996). The e€ects of Rb are also disparate with di€erent results obtained in di€erent settings (Bowen et al., 1998; HaasKogan et al., 1995; Tsang and Little, 1994). Evaluation of the e€ects of bcl2 on survival would appear to be useful for judging the e€ects of apoptosis experimentally since overexpression of bcl2 will abolish the immediate induction of apoptosis. In the case of thymocytes that undergo early apoptosis after irradiation bcl2 enhances radiation survival (Reed, 1995). In tumor cells induction of bcl2 abolishes apoptosis within 72 h of radiation and after treatment with chemotherapeutic agents, but survival may be una€ected (Yin and Schimke, 1995). Apoptosis may merely be delayed by the high levels of bcl2 (Milner et al., 1997). With the later occurrence of apoptosis, bcl2 might not be expected to alter survival after treatment with DNA damaging agents or with radiation. The family of ras genes are also frequently mutated in cancers. Expression of the mutant oncogenic rasH inhibits the extent of apoptosis induced by radiation in transformed fibroblasts (McKenna et al., 1996). The ras oncogene induces radioresistance in those cells suggesting that the ras oncogene might a€ect radioresistance (Ling and Endlich, 1989; McKenna et al., 1990; Samid et al., 1991; Sklar, 1988). Inhibition of rasH action leads to radiosensitization and restores radiation induced apoptosis (Bernhard, 1996a; Miller et al., 1993). There is also evidence that rasK a€ects radiation survival in that inhibition of rasK using prenylation inhibitors in cells with mutated rasK, but not with wild type rasK leads to radiation sensitization (Bernhard et al., 1998). The inhibition of the neu oncogene through the use of dominant negative receptor components also leads to radiosensitization. Since neu signals through the ras pathway it is interesting to speculate that in some cases activation of autocrine growth factors and their receptors leads to signaling through ras that may also contribute to radioresistance in tumor cells (O'Rourke et al., 1998). Radiation survival in vivo may also depend upon apoptosis, although studies in vivo are limited in

comparison to those done in culture. Transformed ®broblasts derived from animals with inactivation of p53 through homologous recombination are more resistant in cell culture, and tumors formed by these cells are more sensitive to radiation in vivo (Lowe et al., 1994; Lowe et al., 1993a). The radiation response of tumor cells di€ers in vivo from that in tissue culture. The time course of apoptosis induction after irradiation is much more rapid in tumor xenografts than in the same cells grown in culture. Meyn's group noted the peak number of apoptotic bodies 4 h after irradiation in cells lines that in culture would undergo apoptosis at times from 24 ± 72 h after irradiation (Stephens et al., 1991, 1993). Radiation survival can di€er when measured in culture in comparison to measurements in tumor xenograpfts. Waldman et al. (1997) found that colon tumor cells with p21 Waf1/ Cip1 eliminated by homologous recombination had the same radiation survival in culture in clonogenic assays as the parental cells, but the p21 de®cient tumors were more readily cured by radiation and they regrew with a greater lag in vivo. The di€erence appears to be through apoptosis independent pathways (Wouters et al., 1997). In other instances suppression of apoptosis in culture translated into altered survival in vivo. Wang et al. (1997) noted that genetic elimination of p21 Waf1/Cip1 increased the radiosensitivity of intestinal crypt cells in ATM de®cient mice and in this case they provided some evidence that the extent of apoptosis was the signi®cant factor. Targeted expression of bcl2 in the lymphocytes of transgenic mice altered their radiation survival in vivo (Van Houten et al., 1997). Rupnow et al. (1998) altered the radiation response of transformed rat embryo ®broblasts through inducible myc expression and found alterations in radiation survival both in vivo and in culture. Radiosensitization of tumors Since many of the oncogenes that a€ect the radiation response are also altered in human tumors, it is possible that these oncogenes might be targets for a€ecting radiosensitivity. This approach would target the tumor and spare the normal cells. Many methods of radiosensitization do not discriminate between normal and tumor cells and could result in severe damage to normal tissues without being able to substantially increase the lethality to the tumor. For example the use of gemcitabine a nucleoside analog that sensitizes both normal and tumor tissue as a radiosensitizer has resulted in major toxicity due to the sensitization of normal tissues (Gregoire et al., 1997; Shewach and Lawrence, 1996). Several targeting strategies focus upon p53. Ca€eine results in radiosensitization of tumor cells and also abolishes the radiation perturbations of G2 (Busse et al., 1978; Tolmach et al., 1977). Ca€eine can alter the cellular response to damage into an apoptotic pathway. Hela cells, which normally do not undergo apoptosis within 72 h of radiation and which in general are quite resistant to induction of apoptosis by a variety of means, undergo apoptosis by 24 h as well as radiosensitization after exposure to ca€eine (Bernhard et al., 1996b). What has been appreciated recently is that ca€eine radiosensitizes cells with inactive p53; cells

Radiosensitization and apoptosis RJ Muschel et al

with wild type p53 are resistant to this action of ca€eine. Drugs similar to ca€eine such as pentoxyphylline produce similar e€ects (Fan et al., 1995; Powell et al., 1995; Russell et al., 1995). The clinical applicability of these drugs is limited since the doses of ca€eine and of pentoxyphylline required to produce radiosensitization are far above the amounts that can be safely administered in vivo. The possibility remains that a safer drug with a similar action could sensitize tumor cells with inactive p53 leaving the normal cells with wild type p53 una€ected. Ca€eine has been shown to a€ect the activation of chk-1, a kinase that phosphorylates cdc25C (Kumagai et al., 1998). Cdc25C is a phosphatase required to activate p34cdc2 during the normal cell cycle. The activity of this phosphatase is reduced following radiation of cells (Barth et al., 1996, Barratt et al., 1998) These experiments were performed in Xenopus oocyte extracts which in analogy to other embryonic tissues might not display p53 activity and hence may mimic tumor cells with mutant forms of p53. Many labs have also attempted to radiosensitize tumor cells by transfection with p53 containing retroviruses or adenoviruses. Whether this strategy is selective to cells with mutant p53 is not entirely established. And the problem with gene therapy in regard to cancer is the diculty in delivery to the majority of cells in a tumor. Nonetheless there are promising results in this regard (Badie et al., 1998; Gallardo et al., 1996; Geng et al., 1998; Gjerset et al., 1995; Lang et al., 1998; Ogawa et al., 1997; Pirollo et al., 1997). Lawrence's lab has developed a strategy based upon di€erential perturbation of the cell cycle although this may be independent of p53 (Lawrence, personal communication). This group noted that treatment of tumor cells with halogenated pyrimidines resulted in a G1 prolongation in some tumor cells, but not in others. Radiation resulted in increased toxicity in cells that continued to progress into S. Thus treatment with halogenated pyrimidines may delay the entry into S in wild type cells limiting the radiation toxicity since halogenated pyrimidines are especially toxic to cells in S (Davis et al., 1995; Lawrence et al., 1996a,b; McGinn et al., 1996). The controls on the perturbation of cycling cells by halogenated pyrimidines are not currently understood.

As was indicated above, ras genes with oncogeneic mutations confer radioresistance both on human tumor cells and on transformed rat ®broblasts. Since the ras gene family are frequently mutated in human cancers, it has been proposed that targeting ras could allow radiosensitization of tumor cells without a€ecting normal tissues (Bernhard et al., 1996a; 1998; Miller et al., 1993). Preliminary experiments would suggest that one result of blocking ras action is to increase radiation induced apoptosis (Bernhard et al., 1996a; Lebowitz et al., 1997c). A variety of laboratories have attempted to block ras action by attacking the posttranslational processing of the ras proteins (Barrington et al., 1998; Cox and Der, 1997; Gelb et al., 1998; Gibbs et al., 1997; Sebti and Hamilton, 1997). RasH and rasK are farnesylated at their C terminal ends. Farnesyl transferase inhibitors prevent this posttranslational processing and block ras action. The problem with this approach is twofold. First ras proteins are not the only proteins that are farnesylated. Indeed farnesyl transferase inhibitors can block the growth of tumor cells, but this action is independent of oncogenic ras (Prendergast et al., 1996; Sepp-Lorenzino et al., 1995). It has been suggested that rhoB is targeted by this drug and that inhibition of rhoB leads to the morphological changes induced by farnesyl transferase inhibitors (Lebowitz et al., 1997a,b). The target for growth inhibition has not been conclusively identi®ed. The second problem is that while RasH is readily inactivated by farnesyl transferase inhibitors, RasK is not. This may be because RasK can be modi®ed by geranyl geranyl transferases when farnesyl transferases are inhibited (Rowell et al., 1997; Whyte et al., 1997). Indeed rasK can be inhibited by using a combination of farnesyl transferase inhibitors and geranyl geranyl transferase inhibitors (Bernhard et al., 1998; Lerner et al., 1997; Sun et al., 1998) While the ideal drug has certainly not yet been designed for inhibition of ras, the concept of using mutated ras as a target for tumor speci®c radiosensitization remains valid. Optimal sensitization to radiation requires that the tumor be targeted while the normal tissue is spared. Many of the genes that are mutated in cancer including p53 and ras have signi®cant e€ects on apoptosis and radiosensitization making them conceptually ideal targets for manipulating the radiosensitivity of tumors.

References Badie B, Kramar MH, Lau R, Boothman DA, Economou JS and Black KL. (1998). J. Neuro-Oncology, 37, 217 ± 222. Barrington RE, Subler MA, Rands E, Omer CA, Miller PJ, Hundley JE, Koester SK, Troyer DA, Bearss DJ, Conner MW, Gibbs JB, Hamilton K, Koblan KS, Mosser SD, O'Neill TJ, Schaber MD, Senderak ET, Windle JJ, Oli€ A and Kohl NE. (1998). Mol. & Cell. Biol., 18, 85 ± 92. Barratt R, Kao G, McKenna W, Kuang J and Muschel R. (1998). Cancer Res., 58, 2639 ± 2645. Barth H, Ho€man I and Kinzel V. (1996). Cancer Res., 56, 2268 ± 2272. Bayreuther K, Rodemann HP, Hommel R, Dittmann K, Albiez M and Francz PI. (1988). Proc. Natl. Acad. Sci. USA, 85, 5112 ± 5116. Bernhard EJ, Kao G, Cox AD, Sebti SM, Hamilton AD, Muschel RJ and McKenna WG. (1996a). Cancer Res., 56, 1727 ± 1730.

Bernhard EJ, McKenna WG, Hamilton AD, Sebti SM, Qian Y, Wu JM and Muschel RJ. (1998). Cancer Res., 58, 1754 ± 1761. Bernhard EJ, Muschel RJ, Bakanauskas VJ and McKenna WG. (1996b). Internl. J. Radi. Biol., 69, 575 ± 584. Bowen C, Spiegel S and Gelmann EP. (1998). Cancer Res., 58, 3275 ± 3281. Bristow RG, Benchimol S and Hill RP. (1996). Radiotherapy & Oncology, 40, 197 ± 223. Bristow RG, Hu Q, Jang A, Chung S, Peacock J, Benchimol S and Hill R. (1998). Oncogene, 16, 1789 ± 1802. Busse PM, Bose SK, Jones RW and Tolmach LJ. (1978). Radiat. Res., 76, 292 ± 307. Chang EH, Jang YJ, Hao Z, Murphy G, Rait A, Fee Jr. WE, Sussman HH, Ryan P, Chiang Y and Pirollo KF. (1997). Archives of Otolaryngology ± Head & Neck Surgery, 123, 507 ± 512.

3361

Radiosensitization and apoptosis RJ Muschel et al

3362

Clarke AR, Purdie CA, Harrison DJ, Morris RG, Bird CC, Hooper ML and Wyllie AH. (1993). Nature, 362, 849 ± 852. Cox AD and Der CJ. (1997). Biochimica et Biophysica Acta, 1333, F51 ± F71. Davis MA, Tang HY, Maybaum J and Lawrence TS. (1995). Internl. J. Radi. Biol., 67, 509 ± 517. DeWeese TL, Walsh JC, Dillehay LE, Kessis TD, Hedrick L, Cho KR and Nelson WG. (1997). Intern. J. Rad. Oncology, Biology, Physics, 37, 145 ± 154. Dewey WC, Ling CC and Meyn RE. (1995). Intern. J. Rad. Oncology, Biology, Physics, 33, 781 ± 796. Di Leonardo A, Linke SP, Clarkin K and Wahl GM. (1994). Genes & Development, 8, 2540 ± 2551. Fan SJ, Smith ML, Rivet DJ, Duba D, Zhan QM, Kohn KW, Fornace AJ and O'Connor PM. (1995). Cancer Res., 55, 1649 ± 1654. Gallardo D, Drazan KE and McBride WH. (1996). Cancer Res., 56, 4891 ± 4893. Gelb MH, Scholten JD and Sebolt-Leopold JS. (1998). Curr. Opin. Chem. Biol., 2, 40 ± 48. Geng L, Walter S, Melian E and Vaughan AT. (1998). Radiation Res., 150, 31 ± 37. Gibbs JB, Graham SL, Hartman GD, Koblan KS, Kohl NE, Omer CA and Oli€ A. (1997). Curr. Opin. Chem. Biol., 1, 197 ± 203. Gjerset RA, Turla ST, Sobol RE, Scalise JJ, Mercola D, Collins H and Hopkins PJ. (1995). Molec. Carcinogenesis, 14, 275 ± 285. Gregoire V, Beauduin M, Rosier JF, De Coster B, Bruniaux M, Octave-Prignot M and Scalliet P. (1997). Br. J. Cancer, 76, 1315 ± 1321. Guo M, Chen C, Vidair C, Marino S, Dewey WC and Ling CC. (1997). Radiation Res., 147, 295 ± 303. Haas-Kogan DA, Kogan SC, Levi D, Dazin P, T'Ang A, Fung YK and Israel MA. (1995). EMBO J., 14, 461 ± 472. Haas-Kogan DA, Yount G, Haas M, Levi D, Kogan SS, Hu L, Vidair C, Deen DF, Dewey WC and Israel MA. (1996). Intern. J. Rad. Oncology, Biology, Physics, 36, 95 ± 103. Harms-Ringdahl M, Nicotera P and Radford IR. (1996). Mutation Res., 366, 171 ± 179. Hasegawa M, Wilson G, Russell LD and Meistrich ML. (1997). Radiation Res., 147, 457 ± 467. Hendry JH, Adeeko A, Potten CS and Morris ID. (1996). Intern. J. Rad. Biol., 70, 677 ± 682. Hendry JH, Cai WB, Roberts SA and Potten CS. (1997). Radiation Res., 148, 254 ± 259. Hendry JH, Potten CS, Chadwick C and Bianchi M. (1982). Intern. J. Rad. Biol. & Related Studies in Physics, Chemistry & Medicine, 42, 611 ± 620. Hurwitz C and Tolmach LJ. (1969). Biophys. J, 9, 607 ± 633. Ijiri K and Potten CS. (1983). Br. J. Cancer, 47, 175 ± 185. Kerr JFK, Wyllie AH and Currie AH. (1972). Br. J. Cancer, 26, 239 ± 245. Kumagai A, Guo Z, Emami K, Wang S and Dunphy W. (1998). J. Cell Biol., 142, 1559 ± 1569. Lang FF, Yung WK, Raju U, Libunao F, Terry NH and To®lon PJ. (1998). J. Neurosurg., 89, 125 ± 132. Lara PC, Russell NS, Smolders IJ, Bartelink H, Begg AC and Coco-Martin JM. (1996). Intern. J. Rad. Biol., 70, 683 ± 692. Lawrence TS, Davis MA and Loney TL. (1996a). Cancer Res., 56, 3203 ± 3206. Lawrence TS, Davis MA, Tang HY and Maybaum J. (1996b). Intern. J. Rad. Biol., 70, 273 ± 280. Lebowitz PF, Casey PJ, Prendergast GC and Thissen JA. (1997a). J. Biol. Chem., 272, 15591 ± 15594. Lebowitz PF, Du W and Prendergast GC. (1997b). J. Biol. Chem., 272, 16093 ± 16095. Lebowitz PF, Sakamuro D and Prendergast GC. (1997c). Cancer Res., 57, 708 ± 713.

Lee JM and Bernstein A. (1993). Proc. Natl Acad. Sci. USA, 90, 5742 ± 5746. Lerner EC, Zhang TT, Knowles DB, Qian Y, Hamilton AD and Sebti SM. (1997). Oncogene, 15, 1283 ± 1288. Ling CC and Endlich B. (1989). Radiat. Res., 120, 267 ± 279. Linke SP, Clarkin KC and Wahl GM. (1997). Cancer Res., 57, 1171 ± 1179. Lowe SW, Bodis S, McClatchey A, Remington L, Ruley HE, Fisher DE, Housman DE and Jacks T. (1994). Science, 266, 807 ± 810. Lowe SW, Ruley HE, Jacks T and Housman DE. (1993a). Cell, 74, 957 ± 967. Lowe SW, Schmitt EM, Smith SW, Osborne BA and Jacks T. (1993b). Nature, 362, 847 ± 849. McGinn CJ, Shewach DS and Lawrence TS. (1996). J. Natl Cancer Instit., 88, 1193 ± 1203. McKenna WG, Bernhard EJ, Markiewicz DA, Rudoltz MS, Maity A and Muschel RJ. (1996). Oncogene, 12, 237 ± 245. McKenna WG, Weiss MC, Endlich B, Ling CC, Bakanauskas VJ, Kelsten ML and Muschel RJ. (1990). Cancer Res., 50, 97 ± 102. Merritt AJ, Allen TD, Potten CS and Hickman JA. (1997). Oncogene, 14, 2759 ± 2766. Meyn RE. (1997). Oncology, 11, 349 ± 356; discussion 356, 361, 365. Miller AC, Kariko K, Myers CE, Clark EP and Samid D. (1993). Int. J. Cancer, 53, 302 ± 307. Milner AE, Grand RJ, Vaughan AT, Armitage RJ and Gregory CD. (1997). Oncogene, 15, 1815 ± 1822. O'Rourke DM, Kao GD, Singh N, Park BW, Muschel RJ, Wu CJ and Greene MI. (1998). Proc. Natl Acad. Sci. USA, 95, 10842 ± 10847. Ogawa N, Fujiwara T, Kagawa S, Nishizaki M, Morimoto Y, Tanida T, Hizuta A, Yasuda T, Roth JA and Tanaka N. (1997). Internl. J. Cancer, 73, 367 ± 370. Ohyama H, Yamada T, Ohkawa A and Watanabe I. (1985). Radiation Res., 101, 123 ± 130. Pirollo KF, Hao Z, Rait A, Jang YJ, Fee Jr. WE, Ryan P, Chiang Y and Chang EH. (1997). Oncogene, 14, 1735 ± 1746. Powell SN, DeFrank JS, Connell P, Eogan M, Pre€er F, Dombkowski D, Tang W and Friend S. (1995). Cancer Res., 55, 1643 ± 1648. Prendergast GC, Davide JP, Lebowitz PF, Wechsler-Reya R and Kohl NE. (1996). Cancer Res., 56, 2626 ± 2632. Radford IR. (1991). Intern. J. Rad. Biol., 59, 1353 ± 1369. Radford IR, Murphy TK, Radley JM and Ellis SL. (1994). Int. J. Rad. Biol., 65, 217 ± 227. Reed JC. (1995). Curr. Opin. Oncol., 7, 541 ± 546. Rodemann HP, Binder A, Burger A, Guven N, Lo‚er H and Bamberg M. (1996). Kidney International ± Supplement, 54, S32 ± S36. Rodemann HP, Peterson HP, Schwenke K and von Wangenheim KH. (1991). Scanning Microscopy, 5, 1135 ± 1142; discussion 1142 ± 1143. Rowell CA, Kowalczyk JJ, Lewis MD and Garcia AM. (1997). J. Biol. Chem., 272, 14093 ± 14097. Rupnow BA, Murtha AD, Alarcon RM, Giaccia AJ and Knox SJ. (1998). Cancer Res., 58, 1779 ± 1784. Russell KJ, Wiens LW, Demers GW, Galloway DA, Plon SE and Groudine M. (1995). Cancer Res., 55, 1639 ± 1642. Samid D, Miller AC, Rimoldi D, Gafner J and Clark EP. (1991). Radiat. Res., 126, 244 ± 250. Sebti S and Hamilton AD. (1997). Curr. Opin. Oncol., 9, 557 ± 561. Sepp-Lorenzino L, Ma Z, Rands E, Kohl NE, Gibbs JB, Oli€ A and Rosen N. (1995). Cancer Res., 55, 5302 ± 5309. Shewach DS and Lawrence TS. (1996). Investigational New Drugs, 14, 257 ± 263. Sinclair W. (1964). Radiat. Res., 21, 584 ± 611. Sklar MD. (1988). Science, 239, 645 ± 647.

Radiosensitization and apoptosis RJ Muschel et al

Slichenmyer WJ, Nelson WG, Slebos RJ and Kastan MB. (1993). Cancer Res., 53, 4164 ± 4168. Stephens LC, Ang KK, Schultheiss TE, Milas L and Meyn RE. (1991). Radiation Res., 127, 308 ± 316. Stephens LC, Hunter NR, Ang KK, Milas L and Meyn RE. (1993). Radiation Res., 135, 75 ± 80. Sun J, Qian Y, Hamilton AD and Sebti SM. (1998). Oncogene, 16, 1467 ± 1473. Tauchi H and Sawada S. (1994). Int. J. Radiat. Biol., 65, 449 ± 455. Thompson LH and Suit HD. (1967). Int. J. Radiat. Biol. & Related Studies in Physics, Chemistry & Medicine, 13, 391 ± 397. Thompson LH and Suit HD. (1969). Int. J. Radiat. Biol. & Related Studies in Physics, Chemistry & Medicine, 15, 347 ± 362. Tolmach LJ, Jones RW and Busse PM. (1977). Radiat. Res., 71, 653 ± 665. Tsang NM and Little JB. (1994). Radiat. Res., 140, 172 ± 179. Tsang NM, Nagasawa H, Li C and Little JB. (1995). Oncogene, 10, 2403 ± 2408. Van Houten N, Blake SF, Li EJ, Hallam TA, Chilton DG, Gourley WK, Boise LH, Thompson CB and Thompson EB. (1997). Internl. Immunol., 9, 945 ± 953.

Vidair CA, Chen CH, Ling CC and Dewey WC. (1996). Cancer Res., 56, 4116 ± 4118. Waldman T, Zhang Y, Dillehay L, Yu J, Kinzler K, Vogelstein B and Williams J. (1997). Nature Med., 3, 1034 ± 1036. Wang L, Cui Y, Lord BI, Roberts SA, Potten CS, Hendry JH and Scott D. (1996). Radiation Res., 146, 259 ± 266. Wang YA, Elson A and Leder P. (1997). Proc. Natl Acad. Sci. USA, 94, 14590 ± 14595. Whyte DB, Kirschmeier P, Hockenberry TN, Nunez-Oliva I, James L, Catino JJ, Bishop WR and Pai JK. (1997). J. Biol. Chem., 272, 14459 ± 14464. Wouters BG, Giaccia AJ, Denko NC and Brown JM. (1997). Cancer Res., 57, 4703 ± 4706. Wyllie AH, Kerr JFR and Currie AR. (1980). Int. Rev. Cytol., 68, 251 ± 305. Yamada T and Ohyama H. (1988). Intern. J. Radiat. Biol. & Related Studies in Physics, Chemistry & Medicine, 53, 65 ± 75. Yin DX and Schimke RT. (1995). Cancer Res., 55, 4922 ± 4928. Yount GL, Haas-Kogan DA, Vidair CA, Haas M, Dewey WC and Israel MA. (1996). Cancer Res., 56, 500 ± 506.

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