Search

Colorectal cancer (CRC) is type of cancer that growing in the colon or the rectum. Although CRC can be treated and managed through several means of therapy, the disease mortality is still high in Saudi Arabia. Upon treatment, cancer cells undergo different form of cell death or growth arrest such as apoptosis and senescence. Senescence is a state of proliferation arrest that is considered as a cause of drug resistance. The current study was designed to interfere with Doxorubicin-induced senescence in cancer cells, and shift the cytotoxicity of doxorubicin from proliferation arrest to apoptosis. β-galactosidase (β-gal) staining has demonstrated that Doxorubicin induces senescence in wild-type HCT116 but not in p21 knock-out cells. (DAPI)/TUNEL stating and Annexin V assay indicate that treatment with Doxorubicin promotes apoptosis in p21 knock-out HCT116 significantly more than in wild-type HCT116. β-galactosidase staining and 5-Dodecanoylaminofluorescein Di-β-D-Galactopyranosid (C12FDG) demonstrated that combination of Doxorubicin with Sorafenib, Rapamycin, or Venetoclax has significantly reduced senescence and induced more apoptosis as shown by Propidium iodide (PI)/Annexin v assay and Caspase 3/7 Assay. PCR data analysis demonstrated that not all combination of Doxorubicin with other agents significantly decreased expression of senescence protein, however, the results of this study provide findings that interference with senescence pathways may shift cytotoxicity from senescence to apoptosis. The authors extend their appreciation to the Deputyship for Research and Innovation, Ministry of Education in Saudi Arabia for partly funding this work through the project number (DRI-KSU-1273). Citation Format: Moureq Rashed Alotaibi, Homood As Sobeia, Munirah Alohaydib, Faten Alaqil, Khalid Alhazzani, Ali Alhoshani, Tareq Saleh, David Gewirtz. Studying the effects of interfering with doxorubicin-induced senescence in human colon cancer HCT116 cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2024.

Background: Acute lymphoblastic leukemia (ALL) is the most common type of childhood malignancy. Despite all the improvements in ALL treatment, cancer relapse remains a major dilemma. Immune checkpoint inhibitors are promising agents that are being evaluated in ALL clinical trials. The overall aim of our study is to evaluate the expression status of immune checkpoint genes in pediatric ALL patients as prognostic factors for the time of relapse and identify possible targets for immunotherapy. Methods: The expression of 21 regulatory immune checkpoint genes was extracted from two microarray datasets of precursor-B-ALL patients (Hogan et al. dataset (GSE28460) included 49 patients and Staal et al. dataset (GSE18497) contained 27 patients). The microarray data were run using the robust multichip average normalization method in RStudio (1.3.959). A 36 month-cutoff was determined to stratify patients into two groups, early relapse (< 36 months) and late relapse (≥ 36 months). The two datasets were normalized to a reference group (late relapse), then combined. The significant differentially expressed genes between the two groups were detected using Student's t-test. An expression score was generated for each patient based on the expression of the significant differentially expressed genes. Survival analysis and Pearson's correlation were performed to assess the impact of significant genes on the time of relapse. Results: 19 out of the 21 genes were downregulated in early relapsing patients compared to the late relapsing group, seven of which were statistically significant; BTLA4, KIR3DS, PDCD1, TIGIT, HAVCR, CTLA4, TNFRSF9. The expression of those seven genes was significantly associated with the time of relapse ( r = 0.381, P = 0.0007). Patients with a low expression profile of the significant differentially expressed genes were 2.3 times more likely to relapse early (CI 1.434-3.713, P = < 0.0001). Conclusion: The expression of immune checkpoint genes is significantly associated with the prognosis of ALL patients. BTLA4, KIR3DS, PDCD1, TIGIT, HAVCR, CTLA4, and TNFRSF9 might be prognostic factors for the time of relapse. Immunotherapy might be beneficial to the late relapsing patients where immune checkpoint genes were upregulated. The study was funded by the deputyship for Research and Innovation, Ministry of Education, Saudi Arabia (DRI-KSU-1273). Citation Format: Basil Jamal Alotaibi, Razan Saad Alshahrani, Munirah Abdulaziz Alkathiri, Ali Mohammed Alqasem, Basil Nasser Alamri, Ali Rashed Alhoshani, Moureq Rashed Alotaibi, Khalid Ahmed Alhazzani, Homood Moqbel As Sobeai. Low expression of immune checkpoint genes is significantly associated with early relapse in pediatric acute lymphoblastic leukemia [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 3031.

Senescence represents a fundamental response to cancer therapy. Accumulating senescent cells contribute to the deleterious outcomes of cancer therapy including cancer relapse, effects that may be largely mediated by the Senescence-Associated Secretory Phenotype (SASP). In this work, we show that tumor cells induced into senescence by etoposide retain proliferative capacity based on their capacity to generate proliferating colonies in culture as well as giving rise to viable tumors in vivo. Using a flow cytometry-based enrichment approach based on enlarged size and expression of Senescence-Associated β-galactosidase (SA-β-gal), we were able to utilize real time imaging to establish the re-emergence of non-small cell lung cancer cells from senescence-based arrest and the generation or proliferating daughter cells (i.e. self-renewal). Moreover, we implemented High-Speed Live-Cell Interferometry (HSLCI) to provide a single-cell lineage tracking of dividing senescent cells. The recovery from senescence was accompanied by resolution of several senescence-associated hallmarks, specifically SA-β-gal activity, p21Waf1/Cip1 and several components of the SASP (IL-1β, IL-6 and CXCL1). Our data suggests that Therapy-Induced Senescence (TIS) may ultimately be a transient process in that at least a subpopulation of tumor cells can recover proliferative capacity. We further demonstrate that the senolytic agent, ABT263, which has been shown to eliminate senescent cells from aging-related animal models can also eliminate senescent tumor cells that persistent after exposure to chemotherapy by shifting the response towards apoptotic cell death. Furthermore, sequential administration of ABT263 interferes with the ability of tumor cells induced into senescence by chemotherapy to recover growth potential. These studies suggest that senescent tumor cells can potentially contribute to cancer relapse by acquiring proliferative properties. The use of senolytic agents after induction of senescence by conventional or targeted therapies allows for the clearance of residual (possibly dormant) senescent tumor cells, which could serve to suppress disease recurrence and cancer mortality. Citation Format: Tareq Saleh, Liliya Tyutyunyk-Massey, Graeme F. Murray, Moureq R. Alotaibi, Ajinkya S. Kawale, Zeinab Elsayed, Scott C. Henderson, Vasily Yakovlev, Lynne W. Elmore, Amir Toor, Hisashi Harada, Jason Reed, Joseph W. Landry, David A. Gewirtz. Elimination of senescent tumor cells by ABT263 interferes with proliferative recovery and provides a two-hit therapeutic approach [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 901.

Radiation remains a predominant treatment option for advanced, inoperable lung cancer, however, resistance represents a major barrier against effective therapy. The exact mechanisms through which cancer cells exhibit resistance to radiotherapy still remains unresolved. Traditionally, radiation-induced autophagy is considered as a mechanism of resistance which attenuates the antitumor effects of radiotherapy i.e., autophagy plays a cytoprotective role on cell survival. To examine this concept further, we employed H460 non-small cell lung cancer (NSCLC) cells that were either p53 wild-type (p53 +/+) or p53 null (p53 -/-) to study the contribution of autophagy to radiation sensitivity. While autophagy was markedly higher in p53 +/+ cells (determined by p62 degradation, LC3B conversion, and acidic vesicle formation) the p53 +/+ cells were actually more radiosensitive than their counterpart H460 p53 -/- cells, which showed lower levels of radiation-induced autophagy. Despite modest radiosensitization by chloroquine, alternative pharmacological (3-methly adenine) and genetic (ATG5 silencing) inhibition of autophagy failed to radiosensitize either p53 +/+ or p53 -/- cells, indicating that radiation induced autophagy is not cytoprotective in function in these experimental models. Furthermore, secretome analysis (nLC-MS/MS) of p53 +/+ H460 cells showed no significant association with radioresistance-related proteins in comparison with p53 -/- H460 cells. The rate and extent of apoptosis was quite low and similar in the two cells lines, essentially ruling out apoptotic cell death as the basis for differential radiation sensitivity. Finally, Senescence was more pronounced in the p53 +/+ cells compared to the p53 -/- cells, which may contribute to the greater radiation sensitivity in the p53 +/+ cells. However, the most relevant finding in this work was that when autophagy is the nonprotective form, it does not confer inherent radiation resistance in tumor cells. Citation Format: Jingwen Xu, Emmanuel Kenneth Cudjoe, Tareq Saleh, Nipa Patel, Moureq Alotaibi, Yingliang Wu, Santiago Lima, David Gewirtz. Nonprotective autophagy fails to confer resistance to radiation [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1326.

While cellular senescence has long been recognized as an irreversible form of growth arrest, evidence in the literature has suggested that subpopulations of senescent tumor cells may retain proliferative capacity. To directly address this question, H460 non-small cell lung cancer cells induced into senescence by exposure to etoposide, and enriched based on β-galactosidase staining and size, were shown to recover reproductive capacity, which was accompanied by resolution of the DNA-damage-response (downregulation of p53 and p21Waf1/Cip1 induction), attenuation of the senescence-associated secretory phenotype (SASP) as well as downregulation of miRNA34 expression and an increase in c-myc. The senescence-enriched lung cancer cells were also capable of tumor growth upon transplantation into immunodeficient mice. Re-emergence from etoposide-induced senescence was also observed using HCT116 colon cancer cells, as evidenced by concomitant downregulation of a senescence-associated reporter (BTG1-RFP). Collectively, these findings indicate that therapy-induced senescence (TIS) may ultimately be a transient process in that at least a subpopulation of tumor cells can resume proliferation. We propose that some forms of tumor dormancy that lead to disease recurrence may reflect cells that enter into and ultimately escape from senescence. Citation Format: Tareq Saleh, Moureq R. Alotaibi, Sarah L. Kyte, Emmanuel K. Cudjoe, Ajinkya Kawale, Jingwen Xu, Zeinab Elsayed, Joseph W. Landry, Scott C. Henderson, Vasily Yakovlev, Lynne W. Elmore, David A. Gewirtz. Therapy-induced senescence is reversible in vitro and in vivo [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 459.

Lung cancer is the leading cause of cancer-related death in both men and women in the United States. Most lung cancer cases are diagnosed in advanced, inoperable stages and are treated with chemoradiation; while chemoradiation is effective in suppressing tumor progression, recurrence following treatment is not infrequent. As there is no well-accepted preclinical model for tumor dormancy and disease recurrence, we hypothesize that the promotion of transient autophagy and senescence could be developed as a model of tumor growth and recovery following treatment. Studies were performed utilizing H460 NSCLC cells as well as a primary NSCLC cell line isolated and grown in our laboratory from a stage IV adenocarcinoma. After treatment with etoposide (1 uM), growth arrest was accompanied by the robust induction of autophagy and senescence. This growth arrest was transient and was followed by proliferative recovery in the course of several days. Genetic and pharmacological inhibition of autophagy failed to interfere with the ability of the cells to regrow, indicating a non-cytoprotective function of autophagy in response to etoposide (in contrast to the cytoprotective function of autophagy exhibited in response to radiation). Quantification of senescence over time based on C12FDG staining and flow cytometry demonstrated that the reversal of growth arrest coincided with a decline in the extent of senescence. To more precisely define the source of the recovered cells, senescent and non-senescent but growth arrested cells were separated by flow cytometry based on their relative β-galactosidase expression and replated. Both cell populations demonstrated the ability to re-emerge from the growth-arrested state and recover proliferative capacity. These observations suggest that senescence is ultimately a transient process in that at least a subpopulation of tumor cells can and will recover proliferative capacity. Furthermore, the reversibility of therapy-induced senescence (TIS) might prove to be a useful model both in cell culture and in vivo to study tumor dormancy and disease recurrence. Studies are currently in progress to determine the impact of senescence inhibition on tumor recovery as well as sensitivity to subsequent therapy given that recurrent disease tends to be relatively refractory to further treatment. Citation Format: Tareq Saleh, Theresa Thekkudan, Moureq R. Alotaibi, David A. Gewirtz. Proliferative recovery after chemotherapy-induced senescence in non-small cell lung cancer (NSCLC) cells as a model of tumor dormancy and disease recurrence. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 2846.

Radiotherapy continues to be a primary modality in the treatment of cancer. DNA damage induced by radiation can promote apoptosis as well as both autophagy and senescence, where autophagy and senescence can theoretically function to prolong tumor survival. A primary aim of this work was to investigate the hypothesis that autophagy and/or senescence could be permissive for DNA repair, thereby facilitating tumor cell recovery from radiation-induced growth arrest and/or cell death. In addition, studies were designed to elucidate the involvement of autophagy and senescence in radiation sensitization by PARP inhibitors and the re-emergence of a proliferating tumor cell population. In the context of this work, the relationship between radiation-induced autophagy and senescence was also determined. Studies were performed using DNA repair proficient HCT116 colon carcinoma cells and a repair deficient Ligase IV (-/-) isogenic cell line. Irradiation promoted a parallel induction of autophagy and senescence that was strongly correlated with the extent of persistent H2AX phosphorylation in both cell lines; however inhibition of autophagy failed to suppress senescence, indicating that the two responses were dissociable. Irradiation resulted in a transient arrest in the HCT116 cells while arrest was prolonged in the Ligase IV (-/-) cells; however, both cell lines ultimately recovered proliferative function, which may reflect maintenance of DNA repair capacity. The PARP inhibitors (Olaparib) and (Niraparib) increased the extent of persistent DNA damage induced by radiation as well as the extent of both autophagy and senescence; neither cell line underwent significant apoptosis by radiation alone or in the presence of the PARP inhibitors. Inhibition of autophagy failed to attenuate radiation sensitization, indicating that autophagy was not involved in the action of the PARP inhibitors. As with radiation alone, despite sensitization by PARP inhibition, proliferative recovery was evident within a period of 10-20 days. While inhibition of DNA repair via PARP inhibition may initially sensitize tumor cells to radiation via the promotion of senescence, this strategy does not appear to interfere with proliferative recovery, which could ultimately contribute to disease recurrence. Citation Format: Moureq R. Alotaibi, Khushboo Sharma, Tareq Saleh, Lawrence Povirk, Eric Hendrickson, David Gewirtz. Autophagy, senescence and proliferative recovery subsequent to DNA damage in radiation sensitization by PARP inhibition. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 2848.

Radiotherapy is used along with other modalities such as surgery, chemotherapy, and immunotherapy to either shrink tumors before surgery or kill surviving tumor cells post surgery. Although radiation (and chemotherapy) induce different modes of cell death, recurrence of cancer may occur. It has been shown that DNA damage can induce cells to undergo both autophagy and senescence where both responses can function either as a pro-survival mechanism or in promoting loss of self-renewal capacity in the tumor cell. The relationship between autophagy, senescence, and the DNA repair system has not been well-characterized. The primary aims of this work were to understand whether autophagy and senescence might be permissive for DNA repair, thereby allowing the tumor cells to overcome the persistent DNA damage and recover using HCT116 WT and HCT116 Ligase IV (-/-) human colon carcinoma cells. Irradiation caused HCT116 Ligase IV (-/-) cells to lose their capacity to proliferate followed by delayed cell death, whereas HCT116 WT cells showed proliferative recovery 72 hours post-treatment, consistent with deficiencies in DNA repair in the Ligase IV (-/-) cells. Irradiation of both cell lines resulted in enlargement in cell size and accumulation of acidic vacuoles, indicative of the induction of autophagy. Inhibition of autophagy by Bafilomycin and Choroquine resulted in radio-sensitization in HCT116 WT cells but not in HCT116 Ligase IV (-/-) cells, indicating that radiation-induced autophagy in HCT116 WT cells is likely to be cytoprotective. Both repair competent HCT116 colon carcinoma cells and Ligase IV deficient cells demonstrated significant increases in senescent populations upon irradiation based on β-galactosidase staining and enlargement of treated cells. The PARP inhibitor, Olaparib, enhanced the efficacy of radiation through interference with DNA repair and the promotion of both autophagy and senescence but not apoptosis. Taken together, it appears that both autophagy and senescence may facilitate DNA repair during exposure to genotoxic stress, and that autophagy and senescence are closely linked responses in irradiated tumor cells. Citation Format: Moureq R. Alotaibi, Lawrence Povirk, Daivd Gewirtz. The relationship between autophagy, senescence, and DNA repair. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 993. doi:10.1158/1538-7445.AM2015-993

Radiotherapy is used along with other modalities such as surgery, chemotherapy, and immunotherapy to either shrink tumors before surgery or kill surviving tumor cells post surgery. Although radiation may induce different modes of cell death, recurrence of cancer may occur. It has been shown that DNA damage can induce cells to undergo both autophagy and senescence where both responses can function either as a pro-survival mechanism or in promoting loss of self-renewal capacity in the tumor cell. The relationship between autophagy, senescence, and the DNA repair system has not been well-characterized. The primary aims of this work were to understand whether autophagy and senescence might be permissive for DNA repair thereby allowing the tumor cells to overcome the persistent DNA damage and recover. Studies were performed using M059K (DNA-PK proficient) and M059J (DNA-PK deficient) human glioma cells to assess the kinetics of DNA repair after radiation in the absence and presence of autophagy inhibitors. M059J cells were observed to lose their capacity to proliferate as they undergo growth arrest followed by delayed cell death, whereas M059K cells showed proliferative recovery 72 hours post-treatment. Both irradiated cells showed an enlargement in cell size and accumulation of acidic vacuoles, indicating the induction of autophagy. Both cell lines also demonstrated reduced p62 level after radiation, indicative of autophagic flux. Inhibition of autophagy by 3-MA showed a radio-sensitization in M059K cells but not in M059J cells, indicating that radio-induced autophagy in M059K cells might be cytoprotective. Additional studies were performed in HCT116 cells that lack ligase IV to demonstrate a correspondence between the extent of drug or radiation induced senescence and the persistence of DNA damage foci. Ligase IV deficient cells were more sensitive to low radiation doses (2 Gy), whereas the ligase proficient cells showed recovery 96 hrs post-treatment. Both parental HCT116 colon carcinoma cells and Ligase IV deficient cells demonstrated significant increases in senescent populations at low doses of radiation based on β-galactosidase staining and enlargement and extensions of treated cells; senescence was more pronounced in the Ligase IV deficient cells. HCT116 cells were then treated with a dose of radiation (8 Gy) that induces ∼75% of senescence after 96 hrs and then reirradiated with half of that dose (4 Gy) to monitor the ability of senescent cells to repair the DNA damage. Although the majority of the cells were in a state of senescence, we found that these cells were able to repair the DNA damage induced by the second dose.Taken together, it seems that autophagy has an important function in the enhancement of DNA repair during exposure to genotoxic stress, but the role of autophagy may differ according to the status of DNA repair. Also, persistent DNA damage could be a determinant for senescence, and senescent cells might be able to repair these breaks. Citation Format: Moureq R. Alotaibi, Daivd Gewirtz, Lawrence Povirk. Involvement of autophagy and senescence in DNA repair in irradiated tumor cells. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2385. doi:10.1158/1538-7445.AM2014-2385

The stilbene derivative, cis-3, 4’, 5-trimethoxy-3’-aminostilbene (stilbene 5c), is a potentially potent antitumor agent that acts via binding to the colchicine-binding pocket in microtubules. Earlier studies have shown that stilbene 5c induces cell death in ovarian cancer cells and leukemic cells (Cao TM et al. Am J Hematol. 2008;83:390-7; Durrant D et al. Gynecol Oncol. 2008; 110:110-7).). The present study was designed to investigate the effectiveness of this microtubule poison against the HCT-116 human colon cancer cell line as well as B16F10 melanoma cells and its mechanisms of action. Time course studies demonstrated that stilbene 5c produces a decrease in cell viability in both cell lines. The capacity of the cells to proliferate was not restored upon removal of the drug after 5 days of exposure. Consistent with the results of the time course studies, β-galactosidase staining indicated that treatment with stilbene 5c also promotes senescence. In addition to senescence, HCT-116 cells and B16F10 melanoma cells treated with stilbene 5c displayed formation of autophagic vesicles by acridine orange staining, which was supported by fluorescence-activated cell sorting (FACS). Further evidence of autophagy was derived from treatment of HCT116 cells expressing an RFP-LC3 construct with stilbene 5c, in which LC3 puncta formation increased in a time-dependent manner. DAPI staining, TUNEL, and Annexin 5 staining indicated that apoptosis is also occurring in stilbene 5c-treated HCT-116 cells. Cell cycle analysis of HCT116 cells demonstrated growth arrest at both G1 and G2/M, and an increase in the subG1 population at days 3 and 5, which correspond to senescence and apoptosis respectively. Interestingly, DAPI and Hoechst staining of HCT116 cells revealed morphological changes in the cell nuclei (binucleated and micronucleated cells), which suggest that mitotic catastrophe may also serve as a mode of cell death after treatment with stilbene 5c. Our studies also indicated that stilbene 5c works in a p53-independent manner. p53-null HCT116 cells demonstrated similar sensitivity to stilbene as p53-wild type HCT116 cells. Consistent with previous studies in other experimental cancer models, this work indicates that stilbene 5c could potentially be effective against melanoma and colon cancer through the promotion of multiple modes of cell death. Citation Format: Moureq R. Alotaibi. Cell death and growth arrest pathways mediating the actions of stilbene 5c in HCT-116 colon cancer cells and B16F10 melanoma cells. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1658. doi:10.1158/1538-7445.AM2013-1658

The stilbene derivative, cis-3, 4′, 5-trimethoxy-3′-aminostilbene (stilbene 5c), is a potentially potent antitumor agent that acts via binding to the colchicine-binding pocket in microtubules. Earlier studies have shown that stilbene 5c induces cell death in ovarian cancer cells and leukemic cells (Lee RM et al, 2008; Lee RM et al, 2008). The present study was designed to investigate the effectiveness of this microtubule poison against the HCT-116 human colon cancer cell line and its mechanisms of action. Time course studies demonstrated that stilbene 5c produces a biphasic decrease in cell viability. The capacity of the cells to proliferate was not restored upon removal of the drug after 6 days of exposure. Consistent with the results of the time course studies, β-galactosidase staining indicated that treatment with stilbene 5c also promotes senescence. In addition to senescence, stilbene 5c-treated HCT-116 cells displayed formation of autophagic vesicles by acridine orange staining, which was supported by fluorescence-activated cell sorting (FACS). Further evidence of autophagy was derived from treatment of HCT116 cells carrying an RFP-LC3 construct with stilbene 5c, in which LC3 puncta formation increased in a time-dependent manner. DAPI staining, TUNEL, and Annexin 5 staining indicated that apoptosis is also occurring in stilbene 5c-treated HCT-116 cells. Cell cycle analysis demonstrated growth arrest at both G1 and G2/M, and an increase in the subG1 population at days 3 and 5, which correspond to senescence and apoptosis respectively. Interestingly, DAPI and Hoechst staining revealed morphological changes in the cell nuclei (binucleated and micronucleated cells), which suggest that mitotic catastrophe may also serve as a mode of cell death after treatment with stilbene 5c. Consistent with previous studies in other experimental cancer models, this work indicates that stilbene 5c could potentially be effective against colon cancer through the promotion of multiple modes of cell death. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3957. doi:1538-7445.AM2012-3957