Clinical Trial Of Preoperative Radiotherapy Research Articles

High expression of cytoplasmic FOXO3 protein associated with poor prognosis of rectal cancer patients: A study from Swedish clinical trial of preoperative radiotherapy to big database analysis

IntroductionAccumulating evidence has implicated a pivotal role for FOXO3, FOXM1 and SIRT6 in cancer progression. The majority of researches focused on the functions of these proteins in drug resistance, but their relationships with radiotherapy (RT) response remain unclear. In this study, we examined protein expression of FOXO3, FOXM1 and SIRT6 and their clinical significance in a Swedish rectal cancer trial of preoperative RT. MethodsExpression of FOXO3, FOXM1 and SIRT6 protein was examined by immunohistochemistry in patient samples. Genetic analysis of FOXO3, FOXM1 and SIRT6 were performed by cBioportal and MEXPRESS database. Gene-gene network analysis was conducted using GeneMANIA. Functional enrichment analysis was performed based on LinkedOmics and Metascape online software. ResultsFOXO3 and FOXM1were mainly expressed in the cytoplasm in both normal and tumour tissues, and SIRT6 in both the cytoplasm and nucleus in normal and tumour tissues. FOXO3 and FOXM1 expression increased from normal mucosa to primary cancer (P < 0.001), while SIRT6 expression decreased from normal mucosa to primary cancer (P < 0.001). High FOXO3 expression correlated with late TNM stage (P = 0.040), distant metastasis (P = 0.032) and independently with disease free survival (DFS) in the RT patients (HR = 7.948; P = 0.049; 95% CI = 1.002–63.032) but not in non-RT patients (P > 0.05). Genetic analysis indicated that DNA methylation status contributed to FOXO3 overexpression. Functional enrichment analysis demonstrated that FOXO3 was closely related to metabolism-related signalling pathway which in turn associated with cancer radioresistance. Moreover, there were strong gene–gene interactions between FOXO3 and metabolism-related signalling. ConclusionsOur findings suggest that FOXO3 may be a prognostic factor in rectal cancer patients with RT.

High Cytoplasmic FOXO3 Expression Associated with Poor Prognosis in Rectal Cancer Patients Received Preoperative Radiotherapy: A Study from Swedish Clinical Trial of Preoperative Radiotherapy to Big Database Analysis

High Cytoplasmic FOXO3 Expression Associated with Poor Prognosis in Rectal Cancer Patients Received Preoperative Radiotherapy: A Study from Swedish Clinical Trial of Preoperative Radiotherapy to Big Database Analysis

FBI-1 expression in relation to clinicopathological variables in rectal cancers with a Swedish clinical trial of preoperative radiotherapy.

e15102 Background:FBI-1 is a recently characterized proto-oncoprotein of the POZ domain Krüppel-like (POK) family of transcription factors. Although several studies provide the evidence that FBI-1 is an important gene regulator in CRC, no analysis of any correlation between FBI-1 expression and preoperative radiotherapy (RT) has been studied in rectal cancers. Methods: This study included the patients with rectal cancer that participated in a Swedish clinical trial of preoperative RT between 1987 and 1990. Patients were divided into preoperative RT (62) and non-RT (77) groups. Applying immunohistochemstry, we detected FBI-1 expression in 118 normal mucosa, 139 primary rectal cancers, and 45 lymph node metastases, and analyzed its relationship with clinicopathological features and RT response. Results: FBI-1 was detected both in the cytoplasm and nucleus, and the cytoplasmic staining was up-regulated compared with normal mucosa both in non-RT and RT groups (74.0% vs. 17.7%, p &lt; 0.001; 69.4% vs. 41.1%, p = 0.002), however, the nuclear staining was down-regulated compared with normal mucosa both in non-RT and RT groups (22.1% vs. 75.8%, p &lt; 0.001; 35.5% vs. 83.9% p &lt; 0.001). Both cytoplasmic and nuclear staining were no difference between the non-RT and RT groups (74.0% vs. 69.4%, p = 0.542; 22.1% vs. 35.5%, p = 0.080, respectively). So we combined the non-RT and RT group together for further analysis. Nuclear staining of FBI-1 was positively with TNM stage and distance recurrence, it showed higher expression in III+ IV stage than that in I+II stage (41.0% vs. 17.9%, p = 0.003). The patients with distance recurrence showed higher FBI-1 expression than that with no distance recurrence (39.7% vs. 19.8%, p = 0.010). In stage I, II and III patients, higher nuclear FBI-1 in primary cancer showed worse disease free survival (HR: 1.934; 95%CI: 1.055-3.579, p = 0.033) and overall survival (HR: 2.174; 95%CI: 1.102-4.290; p = 0.025) independent of gender, age, growth pattern, differentiation and RT. Conclusions: Higher nuclear FBI-1 is related with later TNM stage, distance recurrence, and worse prognosis, it can be used as a potential diagnostic and prognostic biomarker in rectal cancer.

Abstract 4897: Genetic variants and expression of AEG-1 in relation to clinical outcome and radiotherapy response in colorectal cancer patients and cell lines

Abstract Background: Colorectal cancer (CRC) is the third most common cancer in men and the second in woman worldwide. The therapy of CRC has been critically improved during the past two decades, but the treatment response varies significantly between different treatments and patients. Therefore it is of need to search for biomarkers for more suitable prognosis and treatment. The aim of this study was to investigate genetic variants of the astrocyte elevated gene-1 (AEG-1), its expression in CRC patient samples and colon cancer cell lines and the potential correlation to clinicopathological variables and treatment response. Materials and Methods: We analysed genetic variants and the mRNA and protein expression of AEG-1 in 593 CRC patient samples by direct Sanger sequencing, qPCR and immunohistochemistry. Of the patients, 158 rectal cancer patients were enrolled in the Swedish clinical trial of preoperative radiotherapy (RT). AEG-1 expression in response to γ-irradiation was analysed by Western blot in 5 colon cancer cell lines. We inhibited the AEG-1 expression by siRNA in the cell lines, and their survival was analysed after γ-irradiation. Results: By direct sequencing we found 29 variants, of which 12 were novel. Six of the variants were exonic and 23 intronic. The variant c.1353G&amp;gt;A, rs2331652 was found only in 4 samples and the variant c.1679-6 T&amp;gt;C in only one sample. In the cell lines KM12C, KM12SM and KM12L4a we found a deletion in exon 4 (c.595_598delAGAG). AEG-1 mRNA and protein expression revealed a significant increased AEG-1 expression in the primary tumors and metastases compared to the normal mucosa in all patient cohorts (p&amp;lt;0.006) and on the mRNA level a higher expression in tumors located in the rectum compared to tumors in the colon (p=0.047). In the rectal cancer patients from the Swedish trail of preoperative RT, a high AEG-1 expression correlated with a higher risk of distant recurrence and disease free survival (p=0.001 respectively) independently of the tumor stage, only in patients receiving preoperative RT. AEG-1 knock-down in the radioresistant cell lines KM12L4a, SW480 and SW620 resulted in reduction of the survival after radiation, but not in the radiosensitive cell lines KM12C and HCT116. The AEG-1 expression was up-regulated shortly after 2 Gy γ-radiation in KM12C, KM12L4a and SW480, but not in SW620 and HCT116. Conclusion: We found several novel AEG-1 variants. The AEG-1 expression at the mRNA and/or protein level was up-regulated in the primary tumour and metastases compared to the normal mucosa. Furthermore we showed for the first time that the AEG-1 expression is an independent prognostic factor for distant recurrence and disease-free survival in rectal cancer patients after treatment with preoperative RT. The cell line studies suggest AEG-1 as a promising radiosensitizing target for rectal cancer. Citation Format: Sebastian Gnosa, Veronika Patcha Brodin, Ivana Ticha, Gunnar Adell, Gunnar Arbman, Hong Zhang, Xiao-Feng Sun. Genetic variants and expression of AEG-1 in relation to clinical outcome and radiotherapy response in colorectal cancer patients and cell lines. [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 4897. doi:10.1158/1538-7445.AM2014-4897

Livin expression is an independent factor in rectal cancer patients with or without preoperative radiotherapy

BackgroundThis study was aimed to investigate the expression significance of Livin in relation to radiotherapy (RT), clinicopathological and biological factors of rectal cancer patients.MethodsThis study included 144 primary rectal cancer patients who participated in a Swedish clinical trial of preoperative radiotherapy. Tissue microarray samples from the excised primary rectal cancers, normal mucosa and lymph node metastases were immunostained with Livin antibody. The proliferation of colon cancer cell lines SW620 and RKO was assayed after Livin knock-down.ResultsThe expression of Livin was significantly increased from adjacent (P = 0.051) or distant (P = 0.028) normal mucosa to primary tumors. 15.4% (2/13) and 39.7% (52/131) patients with Livin-negative and positive tumors died at 180 months after surgery, and the difference tended to be statistically significant (P = 0.091). In multivariate analyses, the difference achieved statistical significance, independent of TNM stage, local and distant recurrence, grade of differentiation, gender, and age (odds ratio = 5.09, 95% CI: 1.01-25.64, P = 0.048). The in vitro study indicated colon cancer cells with Livin knock-down exhibited decreased proliferation compared with controls after RT.ConclusionsThe expression of Livin was was independently related to survival in rectal cancer patients, suggesting Livin as a useful prognostic factor for rectal cancer patients.

WRAP53 is an independent prognostic factor in rectal cancer- a study of Swedish clinical trial of preoperative radiotherapy in rectal cancer patients

BackgroundExpression of WRAP53 protein has oncogenic properties and it is up regulated in several types of tumors.MethodsWe examined expression of WRAP53 protein in rectal cancers and analyzed its relationship to the response to preoperative radiotherapy and patient survival. The WRAP53 protein was examined by immunohistochemistry in normal mucosa, primary tumors and lymph node metastases from 143 rectal cancer patients participated in a Swedish clinical trial of preoperative radiotherapy.ResultsFrequency of WRAP53 protein expression was increased in primary rectal cancer compared to the normal mucosa (p < 0.05). In non-radiotherapy group positive WRAP53 in primary tumors (p = 0.03, RR, 3.73, 95% CI, 1.13-11.89) or metastases (p = 0.01, RR, 4.11, 95% CI, 1.25-13.14), was associated with poor prognosis independently of stages and differentiations. In radiotherapy group, positive WRAP53 in the metastasis correlated with better survival (p = 0.04). An interaction analysis showed that the correlations of WRAP53 with the prognostic significance with and without radiotherapy in the metastasis differed (p = 0.01). In the radiotherapy group, expression of WRAP53 in metastases gave a better outcome (p = 0.02, RR, 0.32, 95% CI, 0.13-0.84), and an interaction analysis showed significance between the two groups (p = 0.01).ConclusionWRAP53 may be a new biomarker used to predict prognosis and to select suitable patients for preoperative radiotherapy.

Biological Function and Prognostic Significance of Peroxisome Proliferator-Activated Receptor δ in Rectal Cancer

To investigate the expression significance of PPAR β/δ in relation to radiotherapy (RT), clinicopathologic, and prognostic variables of rectal cancer patients. We included 141 primary rectal cancer patients who participated in a Swedish clinical trial of preoperative RT. Tissue microarray samples from the excised rectal cancers and the adjacent or distant normal mucosa and lymph node metastases were stained with PPAR δ antibody. Survival probability was computed by the Kaplan-Meier method and Cox regression model. The proliferation of colon cancer cell lines KM12C, KM12SM, and KM12L4a was assayed after PPAR δ knockdown. PPAR δ was increased from adjacent or distant normal mucosa to primary cancers, whereas it decreased from primary cancers to lymph node metastases. After RT, PPAR δ was increased in normal mucosa, whereas it decreased in primary cancers and lymph node metastases. In primary cancers, the high expression of PPAR δ was related to higher frequency of stage I cases, lower lymph node metastasis rate, and low expression of Ki-67 in the unirradiated cases, and related to favorable survival in the cases either with or without RT. The proliferation of the KM12C, KM12SM, or KM12L4a cells was significantly accelerated after PPAR δ knockdown. RT decreases the PPAR δ expression in primary rectal cancers and lymph node metastases. PPAR δ is related to the early development of rectal cancer and inhibits the proliferation of colorectal cancer cells. Increase of PPAR δ predicts favorable survival in the rectal cancer patients either with or without preoperative RT.

Endosialin expression in relation to clinicopathological and biological variables in rectal cancers with a Swedish clinical trial of preoperative radiotherapy

BackgroundThe importance of changes in tumour-associated stroma for tumour initiation and progression has been established. Endosialin is expressed in fibroblasts and pericytes of blood vessels in several types of tumours, and is involved in the progression of colorectal cancer. In order to see whether endosialin was related to radiotherapy (RT) response, and clinicopathological and biological variables, we investigated endosialin expression in rectal cancers from the patients who participated in a Swedish clinical trial of preoperative RT.MethodsEndosialin was immunohistochemically examined in normal mucosa, including distant (n = 72) and adjacent (n = 112) normal mucosa, and primary tumours (n = 135). Seventy-three of 135 patients received surgery alone and 62 received additional preoperative RT.ResultsEndosialin expression in the stroma increased from normal mucosa to tumour (p < 0.0001) both in RT and non-RT group. In the RT group, endosialin expression in the stroma was positively associated with expression of cyclooxygenase-2 (Cox-2) (p = 0.03), p73 (p = 0.01) and phosphates of regenerating liver (PRL) (p = 0.002). Endosialin expression in the tumour cells of both in the RT group (p = 0.01) and the non-RT group (p = 0.06) was observed more often in tumours with an infiltrative growth pattern than in tumours with an expansive growth pattern. In the RT group, endosialin expression in tumour cells was positively related to PRL expression (p = 0.02), whereas in the non-RT group, endosialin expression in tumour cells was positively related to p73 expression (p = 0.01).ConclusionsEndosialin expression may be involved in the progression of rectal cancers, and was related to Cox-2, p73 and PRL expression. However, a direct relationship between endosialin expression and RT responses in patients was not found.

The location of lymphangiogenesis is an independent prognostic factor in rectal cancers with or without preoperative radiotherapy

BackgroundLymphangiogenesis and angiogenesis are essential for tumour development and progression. The lymphatic vessel density (LVD) and blood vessel density (BVD) and their relationship to outcome have been studied extensively, however the clinical significance of the location of LVD/BVD in tumour is not known. In the present study, the location and degree of LVD/BVD and their relationship to preoperative radiotherapy (RT), clinicopathological, histopathological and biological factors were studied in rectal cancer patients participating in a Swedish clinical trial of preoperative RT. Patients and methodsThe location and degree of LVD/BVD were analysed in primary tumours (n=138/140) and in their subgroups of non-RT (n=74) and RT (n=64/66). Further, the degree of LVD/BVD was examined in the corresponding distant normal mucosa (n=35/31) and adjacent normal mucosa (n=72/91). All sections were immunohistochemically examined by using D2-40 and CD34 antibodies. ResultsIn the whole series of the patients, a higher LVD at the periphery was related to negative p53 expression (P=0.03) and favourable survival independent of tumour–node–metastasis stage, differentiation and p53 expression (P=0.03). LVD was increased in p53-negative tumours after RT (P=0.01). ConclusionLVD at the periphery of the tumour was an independent prognostic factor in rectal cancer patients.

PRb2/p130 protein in relation to clinicopathological and biological variables in rectal cancers with a clinical trial of preoperative radiotherapy

pRb2/p130 plays a key role in cell proliferation and is a considerable progress about expression patterns of pRb2/p130 in number of malignancies. However, pRb2/p130 expression and its significance in rectal cancer remain unknown. The purpose of the present study was to investigate pRb2/p130 protein patterns and their correlations with clinicopathological and biological factors in rectal cancer patients with or without preoperative radiotherapy (RT). pRb2/p130 protein was examined by immunohistochemistry in 130 primary tumors, along with the corresponding 61 distant normal mucosa specimens, 85 adjacent normal mucosa specimens, 34 lymph node metastases, and 93 primary tumor biopsies from rectal cancer patients who participated in a Swedish clinical trial of preoperative RT. The pRb2/p130 protein was mainly localized in the cytoplasm of tumor cells. In nonradiated cases, the lack of pRb2/p130 was related to advanced tumor-node-metastases stage, poorer differentiation, weak fibrosis, less inflammatory infiltration, higher Ki-67, and positive Cox-2 expression (p < 0.05). In radiated cases, the lack of pRb2/p130 was related to nonstaining of Cox-2 and survivin (p < 0.05). pRb2/p130 protein in primary tumors tended to be increased after RT (27% vs 16%, p = 0.07). pRb2/p130 was mainly localized in the cytoplasm rather than in the nucleus in rectal cancer. After RT, pRb2/p130 protein seems to be increased in primary tumors, and further the relationship of the pRb2/p130 with the clinicopathological and biological variables changed compared to the nonradiated cases. However, we did not find that the pRb2/p130 was directly related to RT, tumor recurrence, and patients' survival.

Expression of FXYD-3 is an Independent Prognostic Factor in Rectal Cancer Patients With Preoperative Radiotherapy

FXYD-3 (MAT-8) is overexpressed in several types of cancers; however, its clinical relevance in rectal cancers has not been studied. Therefore, we examined FXYD-3 expression in rectal cancers from the patients who participated in a Swedish clinical trial of preoperative radiotherapy (RT) to determine whether FXYD-3 was overexpressed in rectal cancers and correlated with RT, survival, and other clinicopathologic variables. The study included 140 rectal cancer patients who participated in a clinical trial of preoperative RT, 65 with and 75 without RT before surgery. FXYD-3 expression was immunohistochemically examined in distant (n = 70) and adjacent (n = 101) normal mucosa, primary tumors (n = 140), and lymph node metastasis (n = 36). In the whole cohort, strong FXYD-3 expression was correlated with infiltrative tumor growth (p = 0.02). In the RT group, strong FXYD-3 expression alone (p = 0 .02) or combined with phosphatase of regenerating liver was associated with an unfavorable prognosis (p = 0.02), independent of both TNM stage and tumor differentiation. In tumors with strong FXYD-3 expression, there was less tumor necrosis (p = 0.02) and a trend toward increased incidence of distant metastasis (p = 0.08) after RT. None of these effects was seen in the non-RT group. FXYD-3 expression in the primary tumors tended to be increased compared with normal mucosa regardless of RT. FXYD-3 expression was a prognostic factor independent of tumor stage and differentiation in patients receiving preoperative RT for rectal cancer.

Expression of MAC30 in Rectal Cancers with or without Preoperative Radiotherapy

Objective: Meningioma-associated protein (MAC30) is overexpressed in several types of cancers, but its therapeutic implication in the patients has not been studied. We examined the relationship of MAC30 with clinicopathological and biological factors in rectal cancer patients with or without radiotherapy (RT). Methods: MAC30 was immunohistochemically examined in 75 distant and 91 adjacent normal mucosa specimens, 132 primary tumours and 39 lymph node metastases from rectal cancer patients participating in a clinical trial of preoperative RT. Results: In the RT group, MAC30 was or tended to be positively correlated with infiltrated growth pattern (p = 0.02), PRL (phosphatase of regenerating liver, p = 0.01) and Ki-67 expression (p = 0.06). MAC30 at the invasive margin of the metastasis was related to poor survival (p = 0.02) in the whole group of patients. MAC30 in primary tumours was not related to recurrence and survival in the non-RT or RT group. Conclusions:MAC30 expression in metastasis was an indicator for poor survival. After RT, MAC30 seemed to be more related to aggressive morphological and biological factors; however, we did not find direct evidence that MAC30 expression was related to the outcome of patients with or without RT.

Inflammatory infiltration, fibrosis, necrosis and mucinous content in relation to clinicopathological and molecular factors in rectal cancers with or without preoperative radiotherapy

The association between inflammatory infiltration, fibrosis, necrosis and mucinous content in rectal cancers, and their relationship to preoperative radiotherapy (RT) clinicopathological and biological factors (p53, apoptosis and Cox-2) is not fully characterised. We analysed these histopathological parameters and their relationships in rectal cancer patients who participated in a clinical trial of preoperative RT. One hundred and forty-eight preoperative biopsies and 153 surgically resected tumours were examined. Of the surgical specimens, 81 had surgery alone and 72 received RT before surgery. A higher grade of inflammatory infiltration was related to favourable survival in the whole group of patients (p=0.004, for multivariate analysis p=0.01) as well as in the subgroups of patients with (p=0.04) or without RT (p=0.01). After RT, tumours showed a decreased infiltration (p=0.0003) and increased necrosis (p=0.006), strong necrosis was related to favourable survival (p=0.046). Necrosis (p=0.054) and fibrosis (p=0.06) tended to be increased in p53-negative tumours after RT. Inflammatory infiltration was a strong prognostic factor in rectal cancer patients, regardless of RT. RT tended to induce necrosis and fibrosis in p53-negative tumours.

Expression of the p73 protein in rectal cancers with or without preoperative radiotherapy

To investigate p73 expression in normal mucosa, primary tumor, and metastasis in relation to radiotherapy (RT) response and clinicopathologic/biologic variables in rectal cancers. p73 was immunohistochemically examined on biopsies (unirradiated, n=102), distant (from the large bowel, n=82), and adjacent (adjacent to primary tumor, n=89) normal mucosa samples, primary tumors (n=131), and lymph node metastasis (n=32) from rectal cancer patients participating in a clinical trial of preoperative RT. Seventy-four patients received surgery alone and 57 received additional RT. Cytoplasmic p73 was increased in the primary tumor compared with the distant or adjacent mucosa (p<or=0.0001). Nuclear (p=0.02) and cytoplasmic (p=0.003) p73 was higher in irradiated distant mucosa samples than in unirradiated ones, and nuclear p73 tended to be increased in irradiated primary tumors compared with unirradiated ones (p=0.06). p73 was positively related to cyclooxygenase-2 expression in irradiated tumors (p=0.03). p73-negative tumors tended to have a lower local recurrence after RT compared with unirradiated cases (p=0.06). Normal epithelial cells seem more sensitive to RT than tumor cells regarding p73 expression. Patients with p73-negative rectal tumors may have a lower risk of local recurrence after RT.

Preoperative radiotherapy in combination with metronidazole for resectable gastric cancer: long-term results of a phase 2 study

Aim: Surgery remains the main treatment of stomach carcinoma. Poor long-term survival mandates the study of other techniques. An original treatment scheme with preoperative radiotherapy and metronidazole as a radiosensitizer is reported here.Methods: Between 1982 and 1988, MRRC RAMS carried out a prospective clinical trial of preoperative radiotherapy (20Gy/5 days) in combination with metronidazole (given orally, 3 times, 5gms per metre2). Of 91 patients who received preoperative radiotherapy, 67 patients were operated on with curative intent and were eligible for further analysis.Results: Acute gastro-intestinal toxicity was significant but manageable without surgery delay in most cases. There were 4 postoperative deaths. Overall 5-year, 10-year survival and median survival were 46%, 36% and 46 months. Serosal and nodal involvement were the most significant adverse prognostic factors. Tumours confined to the gastric wall, node negative cases, middle and distal location, differentiated tumours, female sex and age more than 50 years were associated with relatively good long-term results: overall 5-year survival was 50% or better, overall 10-year survival was 40% or better.Conclusion: The combined treatment showed significant but manageable acute toxicity. The long-term results seem encouraging and support further investigations in multimodal treatment of gastric cancer.