. _:b24477683 "Many patients eventually developed a hormone-refractory disease due to the growth of androgen-refractory cancer cells, which leads to failure of androgen ablation therapy and leaves patients with fewer therapeutic options [>>3<<], [4]. Combination of definitive local therapies, such as radical prostatectomy together with adjuvant radiotherapy, has been demonstrated to improve the survival of PC patients [5], [6]. However, such therapy is challenged by the" . _:b530059223 . . . _:b24477713 . . . . . _:b24477705 . _:b24477699 . _:b24477707 "discussion" . _:b24477683 . _:b530059211 . _:b24477701 "For this purpose, we used ARCaPM, the mesenchymal counterpart of ARCaPE developed by Graham et al [>>21<<]. Both cells lines exhibit minimal expression of androgen receptor [22]. Consistent with previous findings by Buck et al [14], combined treatment with EGFR and IGF1R inhibitors could not synergistically inhibit mesenchymal cell growth in" . _:b24477700 . . . _:b24477703 . . _:b24477701 . . . _:b24477702 . . _:b530059215 . _:b530059233 . _:b24477703 . _:b24477700 "results" . _:b24477702 . _:b24477681 . _:b24477696 . . _:b24477697 "To detect apoptotic rate in treated cells, the cells were stained with Annexin V and PI, and then subjected to flow cytometry as described [>>16<<]. The Annexin V-positive and PI-negative cells undergoing early apoptosis were counted as apoptotic cells. All experiments were conducted in duplicates and data shown are representative of three independent experiments." . . _:b24477697 . . _:b24477698 . . _:b530059213 . _:b24477699 . _:b24477687 . _:b530059230 . _:b530059217 . . . . _:b24477708 . . _:b24477686 . _:b24477709 . _:b24477707 _:b24477708 . _:b24477707 _:b24477709 . _:b24477707 _:b24477710 . _:b24477710 . _:b24477707 _:b24477711 . _:b530059218 . _:b24477698 . _:b24477711 . _:b530059232 . _:b24477686 "play critical roles in cancer development and progression through the regulation on cell proliferation, apoptosis, anchorage-independent growth, invasion, angiogenesis, cancer immunity and resistance to chemo- and/or radiotherapy [>>7<<]. These two receptors are frequently overexpressed in a variety of human cancers including PC [8], [9], [10], and therefore could be used as candidates for targeted cancer therapy." . _:b24477704 . . _:b24477705 . _:b530059229 . _:b530059222 . _:b24477706 . _:b24477707 _:b24477712 . _:b24477701 . _:b24477707 _:b24477713 . _:b24477707 _:b24477714 . _:b24477707 . _:b24477693 . . . _:b24477680 "introduction" . . . _:b24477708 . _:b530059224 . _:b24477712 "Upon their activation, the PI3K-AKT and Ras-MAPK pathways present specific activity toward resistance to chemo- and radio-therapy, in addition to their regulatory capability on cell viability, apoptosis and cell proliferation [>>28<<], [29]: The Ras-MAPK pathway stimulates production of EGFR ligand and in turn activates EGFR in an autocrine manner, while PI3K-AKT pathway directly activates the catalytic subunit of DNA-PK, the essential component for NHEJ machinery. The" . _:b24477709 . _:b24477712 . _:b24477700 _:b24477701 . . _:b24477700 _:b24477702 . _:b24477713 . _:b24477696 "Cells were irradiated as described by Liu et al [>>16<<]. In some experiments, the cells were also transfected with IRS1 or non-silencing control (NSC) siRNAs (50 nM, Invitrogen, Shanghai, China) according to manufacturer's protocol." . . _:b24477700 _:b24477703 . _:b24477696 . . _:b24477692 . _:b24477714 . _:b24477697 . . _:b24477700 _:b24477704 . . _:b24477700 _:b24477705 . _:b24477690 "treatment of different human cancers, As expected however, the development of de novo resistance has been observed in clinic after long-term use of these medicines, suggesting the existence of bypass mechanisms within tumor cells [>>11<<]. Mechanistic studies on the cellular and molecular events revealed that extensive crosstalk between EGFR and IGF1R signaling occurs at multiple levels, and that blockage of EGFR signaling leads to enhanced responses to the IGF1R ligand," . . _:b24477700 _:b24477706 . _:b530059225 . _:b530059213 "2"^^ . _:b24477682 "As a consequence, androgen ablation was considered the standard treatment for PC for over 50 years [>>2<<]. Many patients eventually developed a hormone-refractory disease due to the growth of androgen-refractory cancer cells, which leads to failure of androgen ablation therapy and leaves patients with fewer therapeutic options [3], [4]." . _:b24477704 "For this analysis, the nuclear extract from R503 cells was used as the positive control, as demonstrated by Yang et al [>>23<<]. In addition, expression of NHEJ-related DNA repair proteins, including DNA-PK, Ku70, Ku80 and XRCC4, were not affected by AG1024, Erlotinib or both in DU145 cells (Figure 2E), suggesting that inhibition of these two signaling pathways" . _:b530059213 . "PMC0" . _:b530059235 . _:b530059212 "3"^^ . _:b530059212 . _:b24477705 "Previous studies revealed extensive crosstalk between EGFR and IGF1R signaling pathway at multiple levels [>>14<<]. Our data above have demonstrated that co-inhibition of EGFR and IGF1R could additively radio-sensitize PC cells through the impairment of HRR DSB repair." . _:b530059215 "2"^^ . _:b530059215 . . _:b530059214 "2"^^ . . _:b530059214 . _:b530059209 . _:b530059209 "5"^^ . _:b24477702 "Both cells lines exhibit minimal expression of androgen receptor [>>22<<]. Consistent with previous findings by Buck et al [14], combined treatment with EGFR and IGF1R inhibitors could not synergistically inhibit mesenchymal cell growth in response to irradiation, as it did for epithelial growth (Figure 1A)." . . _:b24477706 . _:b24477703 "Consistent with previous findings by Buck et al [>>14<<], combined treatment with EGFR and IGF1R inhibitors could not synergistically inhibit mesenchymal cell growth in response to irradiation, as it did for epithelial growth (Figure 1A)." . _:b530059211 "3"^^ . . _:b530059211 . _:b530059210 "3"^^ . _:b530059210 . _:b24477680 _:b24477684 . _:b24477711 . _:b24477680 _:b24477685 . _:b530059221 "2"^^ . _:b24477680 _:b24477686 . _:b24477713 "Upon their activation, the PI3K-AKT and Ras-MAPK pathways present specific activity toward resistance to chemo- and radio-therapy, in addition to their regulatory capability on cell viability, apoptosis and cell proliferation [28], [>>29<<]: The Ras-MAPK pathway stimulates production of EGFR ligand and in turn activates EGFR in an autocrine manner, while PI3K-AKT pathway directly activates the catalytic subunit of DNA-PK, the essential component for NHEJ machinery." . _:b24477680 _:b24477687 . _:b530059221 . _:b530059210 . _:b530059220 "2"^^ . _:b24477680 _:b24477681 . _:b24477680 _:b24477682 . _:b24477680 _:b24477683 . _:b530059220 . _:b24477680 _:b24477692 . _:b24477680 _:b24477693 . _:b530059223 "2"^^ . _:b24477680 _:b24477694 . _:b24477693 "Consistently, studies have shown that dual targeting of both receptors blocks their reciprocal hyperphosphorylation, inhibits the proliferation and induces apoptosis in multiple cancer cells including PC and colorectal cancer [>>14<<], [15]." . _:b530059223 . _:b24477680 _:b24477688 . _:b24477695 . _:b530059222 "2"^^ . _:b530059220 . _:b24477680 _:b24477689 . _:b24477680 _:b24477690 . _:b24477714 "However, in IGF1R pathway, Trojanek et al. demonstrated that upon IGF-I stimulation, IRS1, the major IGF1R substrate, binds with Rad51, stimulates nuclear localization of the latter to the site of damaged DNA, and promotes HRR [>>24<<]." . _:b24477690 . _:b24477680 _:b24477691 . _:b530059222 . _:b530059217 "2"^^ . _:b530059217 . . _:b24477710 "This explains why the combination of these two inhibitors could synergistically inhibit HRR, but only showing additive effect on DSB repair inhibition, consistent with previous studies that NHEJ is the main mechanism of DSB repair [>>26<<]." . _:b530059216 "2"^^ . _:b24477714 . . _:b24477685 . _:b24477680 . _:b530059216 . _:b24477681 "Prostate cancer (PC) is the most common malignancy and the second leading cause of cancer-related deaths among male patients [>>1<<]. During cancer progression, the initial growth of PC cells is androgen-dependent, and these cells undergo apoptosis upon androgen depletion." . _:b530059212 . . _:b530059219 "2"^^ . _:b24477682 . _:b530059219 . . _:b530059218 "2"^^ . _:b530059218 . _:b530059229 "2"^^ . _:b530059229 . . _:b530059228 "2"^^ . _:b530059228 . . _:b530059231 "2"^^ . _:b530059231 . _:b24477684 . _:b530059230 "2"^^ . . _:b24477694 . . _:b24477699 "The quantitative in vitro homologous recombination (HR) assay was performed using the pDR-GFP recombination reporter system [>>19<<]. Briefly, the pDR-GFP plasmid expressing two nonfunctional GFP genes was stably transfected into DU145 cells." . _:b24477685 . _:b530059230 . _:b530059225 "2"^^ . _:b530059225 . _:b24477704 . _:b24477686 . _:b530059238 . _:b24477710 . . _:b530059224 "2"^^ . _:b530059224 . _:b24477687 . _:b530059227 "2"^^ . _:b530059234 . . . _:b530059227 . _:b24477680 . _:b530059236 . _:b24477689 . _:b530059226 "2"^^ . . _:b530059214 . _:b530059237 . . _:b530059238 . _:b24477681 . _:b530059226 . . _:b530059237 "2"^^ . _:b530059232 . _:b530059233 . _:b530059231 . _:b530059234 . _:b24477682 . _:b530059237 . _:b24477695 "materials and methods" . _:b24477684 "Many patients eventually developed a hormone-refractory disease due to the growth of androgen-refractory cancer cells, which leads to failure of androgen ablation therapy and leaves patients with fewer therapeutic options [3], [>>4<<]. Combination of definitive local therapies, such as radical prostatectomy together with adjuvant radiotherapy, has been demonstrated to improve the survival of PC patients [5], [6]. However, such therapy is challenged by the emergence of" . _:b530059235 . _:b530059236 "2"^^ . _:b24477695 _:b24477696 . _:b24477695 _:b24477697 . _:b24477683 . _:b530059236 . . _:b24477695 _:b24477698 . _:b24477695 _:b24477699 . _:b24477706 "Our current data on AG1024 and Erlotinib regulating IRS1 phosphorylation/activation implicated the potential involvement of IRS1/Rad51-mediated HRR in response to \u03B3 irradiation in PC cells, as previous study also indicated [>>24<<]. Indeed, our data showed that IRS1 phosphorylation was induced by \u03B3-irradiation with peak activation achieved at 4 h after \u03B3-irradiation in DU145 and PC3 cells, while the peak of IRS1 phosphorylation was delayed to 8 h after \u03B3-irradiation" . _:b24477692 . _:b530059209 . _:b530059238 "2"^^ . _:b24477692 "studies on the cellular and molecular events revealed that extensive crosstalk between EGFR and IGF1R signaling occurs at multiple levels, and that blockage of EGFR signaling leads to enhanced responses to the IGF1R ligand, IGF [12], [>>13<<]. These data imply that targeting both receptors at the same time could provide better efficacy in cancer treatment and overcome tumor resistance to an individual inhibitor, while improving the sensitivity of individual inhibitors to" . _:b530059238 . _:b24477693 . _:b24477688 . . _:b530059227 . _:b24477707 . _:b530059233 "2"^^ . _:b24477700 . _:b530059233 . _:b24477694 . _:b530059212 . _:b24477712 . _:b530059232 "2"^^ . _:b530059213 . _:b530059228 . _:b530059214 . _:b530059232 . _:b24477695 . _:b530059215 . _:b530059235 "2"^^ . _:b530059209 . _:b530059216 . _:b530059210 . _:b24477688 . _:b530059235 . _:b530059211 . _:b24477708 "Without proper DNA repair capacity, cells undergo apoptosis, mitotic catastrophe, autophagy, cellular senescence, or even carcinogensis [>>25<<]. However, in tumor cells after radiation therapy, enhanced DNA repair capacity such as HRR or NHEJ leads to resistance to radiotherapy. HRR is a template-driven and thus error-free DNA repair mechanism that involves many proteins" . _:b530059234 "2"^^ . . _:b530059234 . _:b24477709 "NHEJ is a more error-prone mechanism that operates with DNA repair proteins such as Ku70/80, XRCC4, XRCC1, DNA-PK, and XLF [>>25<<]. In the current study, we showed that inhibition of both EGFR and IGF1R was able to suppress HRR repair of damaged DNA after \u03B3-irradiation, but not NHEJ repair. This explains why the combination of these two inhibitors could" . _:b24477689 . . _:b24477687 "These two receptors are frequently overexpressed in a variety of human cancers including PC [>>8<<], [9], [10], and therefore could be used as candidates for targeted cancer therapy." . . _:b24477689 "These two receptors are frequently overexpressed in a variety of human cancers including PC [8], [9], [>>10<<], and therefore could be used as candidates for targeted cancer therapy." . . _:b24477690 . "10.1371%2Fjournal.pone.0068784" . _:b530059228 . _:b530059229 . _:b530059230 . _:b24477691 . _:b530059231 . _:b530059221 . _:b530059224 . _:b530059225 . _:b24477711 "Both signaling pathways including the downstream signaling cascades are activated in response to ionizing radiation [>>27<<]. Upon their activation, the PI3K-AKT and Ras-MAPK pathways present specific activity toward resistance to chemo- and radio-therapy, in addition to their regulatory capability on cell viability, apoptosis and cell proliferation [28], [29]:" . _:b24477684 . _:b530059226 . _:b530059219 . . _:b530059227 . . _:b530059220 . _:b530059221 . _:b530059237 . _:b530059222 . _:b530059223 . . _:b530059216 . _:b530059217 . _:b530059218 . _:b24477698 "Immunoprecipitation and Western-blot assay were performed as described by Liu et al [>>17<<]. The following antibodies were from Cell Signaling:" . _:b530059219 . _:b24477691 "studies on the cellular and molecular events revealed that extensive crosstalk between EGFR and IGF1R signaling occurs at multiple levels, and that blockage of EGFR signaling leads to enhanced responses to the IGF1R ligand, IGF [>>12<<], [13]. These data imply that targeting both receptors at the same time could provide better efficacy in cancer treatment and overcome tumor resistance to an individual inhibitor, while improving the sensitivity of individual inhibitors to" . _:b530059226 . _:b24477691 . _:b530059236 . . _:b24477685 "Combination of definitive local therapies, such as radical prostatectomy together with adjuvant radiotherapy, has been demonstrated to improve the survival of PC patients [>>5<<], [6]. However, such therapy is challenged by the emergence of resistance in tumor cells. It is, therefore, of paramount importance to develop novel therapeutic strategies to overcome radioresistance and improve radio-sensitivity by" . . . _:b24477694 "Consistently, studies have shown that dual targeting of both receptors blocks their reciprocal hyperphosphorylation, inhibits the proliferation and induces apoptosis in multiple cancer cells including PC and colorectal cancer [14], [>>15<<]." . _:b24477688 "These two receptors are frequently overexpressed in a variety of human cancers including PC [8], [>>9<<], [10], and therefore could be used as candidates for targeted cancer therapy." . .