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802 Circulating Tumor Cells After Neoadjuvant Chemotherapy Predict Survival in Non-metastatic Breast Cancer
Poster Sessions
european journal of cancer 48, suppl. 5 (2012) S25–S288
800 Role of HERG1 Potassium Channel in Both Malignant Transformation and Disease Progression in Head and Neck Carcinomas 1 1 ´ S. Alvarez-Teijeiro , S. Tirados Menendez ´ , J.P. Rodrigo1 , M.A. Villaronga1 , 1 E. Allonca1 , A. Vallina1 , A. Astudillo1 , F. Barros2 , C. Suarez ´ , J.M. Garc´ıaPedrero1 . 1 Hospital Universitario Central de Asturias (HUCA), Instituto ´ Universitario Oncologico del Principado de Asturias (IUOPA), Oviedo − Asturias, Spain, 2 Universidad de Oviedo, Departamento de Bioqu´ımica, Oviedo − Asturias, Spain Introduction: Evidences indicate that HERG1 voltage-gated potassium channels could represent new valuable membrane therapeutic targets and diagnostic/prognostic biomarkers in various cancers. This study is the first to investigate the expression pattern of HERG1 potassium channel subunit in both primary tumors and precancerous lesions to establish its clinical and biological role during the development and progression of head and neck squamous cell carcinomas. Material and Method: HERG1 protein expression was evaluated by immunohistochemistry in paraffin-embedded tissue specimens from 133 patients with laryngeal/hypopharyngeal squamous cell carcinomas and 75 patients with laryngeal dysplasia, and correlated with clinical data. In vitro functional studies in HNSCC-derived cell lines further contributed to clarify the pathobiological role of HERG1 potassium channel subunit. Results and Discussion: Our findings demonstrate that HERG1 is frequently aberrantly expressed in a high percentage of primary tumors (87%), whereas expression was negligible in both stromal cells and normal-adjacent epithelia. HERG1 expression increased during head and neck squamous cell carcinoma progression and was significantly associated with lymph node metastasis (P = 0.04), advanced disease stages (P < 0.001), regional tumor recurrence (P = 0.004), distant metastasis (P = 0.03), and reduced diseasespecific survival (P = 0.012, log-rank test). HERG1-positive expression was also detected in 31 (41%) of 75 laryngeal dysplasias. Interestingly, HERG1 expression increased with the grade of dysplasia; however, HERG1 expression but not histology correlated significantly with increased laryngeal cancer risk (P = 0.007). In addition, functional studies in head and neck squamous cell carcinoma-derived cell lines further revealed that HERG1 expression promotes anchorage-dependent and -independent cell growth and invasive capability, although independently of its ion-conducting function. Conclusion: Our data demonstrate that HERG1 expression is a biological and clinical relevant feature in head and neck squamous cell carcinoma progression and also during malignant transformation and a promising candidate as cancer risk marker and therapeutic target for head and neck squamous cell carcinoma prevention and treatment. 801 Frequent Aberrant Expression of the Human Ether a Go-go (hEAG1) Potassium Channel in Head and Neck Cancer − Pathobiological Mechanisms and Clinical Implications 1 ´ M.A. Villaronga Torres1 , S. Tirados Menendez ´ , J.P. Rodrigo1 , S. AlvarezTeijeiro1 , D. Garc´ıa-Carracedo1 , R.G. Urdinguio2 , M.F. Fraga2 , L.A. Pardo3 , 1 C. Suarez ´ , J.M. Garc´ıa-Pedrero1 . 1 Hospital Universitario Central de Asturias (HUCA), Instituto Universitario de Oncolog´ıa del Principado de Asturias (IUOPA), Oviedo − Asturias, Spain, 2 Instituto Universitario de Oncolog´ıa del Principado de Asturias (IUOPA), Epigenetic Unit, Oviedo − Asturias, Spain, 3 Max-Planck-Institute of Experimental Medicine, Department of Molecular ¨ Biology of Neuronal Signals, Gottingen, Germany
Introduction: Compelling evidence indicates that the human ether-a-go-go voltage-gated potassium channels (hEAG1) may represent new valuable membrane therapeutic targets and diagnostic/prognostic biomarkers in various cancers. This study is the first to investigate the expression of hEAG1 potassium channel subunit in both primary tumors and HNSCC-derived cell lines to ascertain its clinical and biological role in the progression of head and neck squamous cell carcinomas. Material and Method: hEAG1 mRNA expression was analyzed by QRTPCR in a prospective series of 54 fresh HNSCC tissue specimens and 44 patient-matched normal epithelia. Molecular alterations were associated with clinicopathological parameters and disease outcome. To identify the mechanisms responsible for the aberrant expression of hEAG1 in HNSCC we studied the possible contribution of hEAG1 gene copy gain by Q-PCR in 88 HNSCC tissue specimens and also epigenetic transcriptional regulatory mechanisms (DNA methylation by pyrosequencing and histone acetylation by chromatin immunoprecipitation). Results and Discussion: Our findings demonstrate that hEAG1 is frequently aberrantly expressed in a high percentage of primary tumors (83%, 45/54 cases) and HNSCC-derived cell lines (83%, 10/12 cell lines). hEAG1 expression increased during HNSCC progression and was more frequent in advanced tumors. Strikingly, hEAG1 expression was also detected in a notable proportion (39%, 17/44 cases) of patient-matched normal adjacent mucosa, whereas no expression was detected in normal epithelia from nononcologic patients without exposure to tobacco carcinogens. In an attempt to
S191
identify the underlying mechanisms of aberrant hEAG1 expression in HNSCC, we found that hEAG1 gene copy gain occurred at a low frequency (15%, 13/88 cases) in primary tumors but was not observed in early stages of HNSCC tumorigenesis. Furthermore, this study provides original evidence supporting the involvement of histone acetylation (i.e. H3Ac and H4K16Ac activating marks) in the regulation of hEAG1 expression in HNSCC. In addition, functional studies in HNSCC cells further revealed that hEAG1 expression is a biologically relevant feature that promotes cell proliferation and invasion, although independently of its ion-conducting function. Conclusion: Our findings strongly support the notion that hEAG1 may represent a promising candidate as tumor marker and membrane therapeutic target for HNSCC treatment. 802 Circulating Tumor Cells After Neoadjuvant Chemotherapy Predict Survival in Non-metastatic Breast Cancer A. Lucci1 , A. Lodhi1 , A. Bhattacharyya1 , C. Hall1 , S. Jackson2 , B. Singh1 , S. Krishnamurthy3 , H. Kuerer1 . 1 UT MD Anderson Cancer Center, Surgical Oncology, Houston TX, USA, 2 UT MD Anderson Cancer Center, Breast Medical Oncology, Houston TX, USA, 3 UT MD Anderson Cancer Center, Pathology, Houston TX, USA Introduction: Circulating tumor cells (CTCs) predict outcome in metastatic breast cancer, but their significance is unclear in non-metastatic patients. Furthermore, it is unclear if the presence of CTCs after completion of neoadjuvant chemotherapy (NACT) predicts worse outcome. The purpose of this study was to determine if CTCs after NACT predict worse outcome. Methods: Clinical stage I-III breast cancer patients seen at a single tertiary cancer center provided informed consent to participate in an IRB-approved study involving collection of blood (7.5 ml x 2 tubes) at the time of surgery for their primary breast cancer. CTCs were detected using the CellSearch® system. A positive result was defined as the presence of one or more cells per 7.5 ml blood since the threshold for positivity has not been established in nonmetastatic breast cancer. Statistical analyses used chi-square and Fischer’s exact test. Results: One hundred and thirty seven patients were prospectively enrolled. Median age was 52 years and median follow-up was 34 months. Nine percent of patients had T1 disease, 36% T2, 20% T3, and 35% T4. Fifty-four percent of patients (73/137) had ER positive and 38% (52/137) had PR positive disease. Thirty percent of patients (41/137) were HER-2 positive. Twenty eight percent (38/137) had triple-negative tumors. Sixty-eight percent (93/137) had lymph node positive disease. One CTC was found in 27% (37/137) of patients postNACT, but its presence did not predict worse outcome (p = NS). Two or more CTCs were present in 9% (12/137) of patients. Of the 20 patients who relapsed, 6 had 2 or more CTCs (P = 0.002), while of the 14 patients who died, 4 had 2 or more CTCs (P = 0.001). Conclusions: Presence of two or more CTCs after NACT predicted worse relapse-free and overall survival in patients with stage I-III breast cancer. 803 Integrative Marker Analysis and Risk Assessment in Stage II Colon Cancer U. Nitsche1 , R. Rosenberg1 , T. Schuster2 , J. Slotta-Huspenina3 , F.G. Bader1 , H. Friess1 , P.M. Schlag4 , U. Stein5 , K.P. Janssen1 . 1 Klinikum Rechts der Isar, Chirurgische Klinik und Poliklinik, Munchen, ¨ Germany, 2 Klinikum Rechts der Isar, Institute of Statistics and Epidemiology, Munchen, ¨ Germany, 3 Klinikum Rechts der Isar, Institute of Pathology, Munchen, ¨ Germany, 4 Charite´ Medical Faculty and Max-Delbruck-Center ¨ for Molecular Medicine and Charite´ Comprehensive Cancer Center, Experimental and Clinical Research Center, Berlin, Germany, 5 Charite´ Medical Faculty and Max-Delbruck-Center ¨ for Molecular Medicine, Experimental and Clinical Research Center, Berlin, Germany Background: Colorectal cancer progression is governed by genetic and epigenetic alterations that can occur in multiple parallel pathways. However, with the exception of KRAS mutations, molecular markers currently play no role in risk stratification and therapy decision. E.g., risk assessment based on clinico-pathological parameters is not reliable for patients with stage II colon cancer (locally restricted disease). We hypothesize that the individual recurrence risk is defined by tumor genetics, and have therefore analyzed a panel of molecular genetic markers for risk prediction. Material and Methods: Non-microdissected fresh frozen tissue from 232 patients (T3−4 N0 M0) with complete tumor resection and median followup of 97 months was analyzed for microsatellite stability by multiplex PCR, KRAS exon 2, and BRAF exon 15 mutations by high-resolution melting analysis. Moreover, gene expression of the WNT-pathway surrogate marker osteopontin and the metastasis-associated genes SASH1 and MACC1 was determined for 179 patients. The results were correlated with metachronous distant metastasis as primary endpoint (n = 22 patients). Results: Mutations of KRAS were detected in 30% of patients, mutations of BRAF in 15%, and microsatellite instability in 26%, in good accordance with reported findings. Risk of recurrence was associated with KRAS mutation
european journal of cancer 48, suppl. 5 (2012) S25–S288
800 Role of HERG1 Potassium Channel in Both Malignant Transformation and Disease Progression in Head and Neck Carcinomas 1 1 ´ S. Alvarez-Teijeiro , S. Tirados Menendez ´ , J.P. Rodrigo1 , M.A. Villaronga1 , 1 E. Allonca1 , A. Vallina1 , A. Astudillo1 , F. Barros2 , C. Suarez ´ , J.M. Garc´ıaPedrero1 . 1 Hospital Universitario Central de Asturias (HUCA), Instituto ´ Universitario Oncologico del Principado de Asturias (IUOPA), Oviedo − Asturias, Spain, 2 Universidad de Oviedo, Departamento de Bioqu´ımica, Oviedo − Asturias, Spain Introduction: Evidences indicate that HERG1 voltage-gated potassium channels could represent new valuable membrane therapeutic targets and diagnostic/prognostic biomarkers in various cancers. This study is the first to investigate the expression pattern of HERG1 potassium channel subunit in both primary tumors and precancerous lesions to establish its clinical and biological role during the development and progression of head and neck squamous cell carcinomas. Material and Method: HERG1 protein expression was evaluated by immunohistochemistry in paraffin-embedded tissue specimens from 133 patients with laryngeal/hypopharyngeal squamous cell carcinomas and 75 patients with laryngeal dysplasia, and correlated with clinical data. In vitro functional studies in HNSCC-derived cell lines further contributed to clarify the pathobiological role of HERG1 potassium channel subunit. Results and Discussion: Our findings demonstrate that HERG1 is frequently aberrantly expressed in a high percentage of primary tumors (87%), whereas expression was negligible in both stromal cells and normal-adjacent epithelia. HERG1 expression increased during head and neck squamous cell carcinoma progression and was significantly associated with lymph node metastasis (P = 0.04), advanced disease stages (P < 0.001), regional tumor recurrence (P = 0.004), distant metastasis (P = 0.03), and reduced diseasespecific survival (P = 0.012, log-rank test). HERG1-positive expression was also detected in 31 (41%) of 75 laryngeal dysplasias. Interestingly, HERG1 expression increased with the grade of dysplasia; however, HERG1 expression but not histology correlated significantly with increased laryngeal cancer risk (P = 0.007). In addition, functional studies in head and neck squamous cell carcinoma-derived cell lines further revealed that HERG1 expression promotes anchorage-dependent and -independent cell growth and invasive capability, although independently of its ion-conducting function. Conclusion: Our data demonstrate that HERG1 expression is a biological and clinical relevant feature in head and neck squamous cell carcinoma progression and also during malignant transformation and a promising candidate as cancer risk marker and therapeutic target for head and neck squamous cell carcinoma prevention and treatment. 801 Frequent Aberrant Expression of the Human Ether a Go-go (hEAG1) Potassium Channel in Head and Neck Cancer − Pathobiological Mechanisms and Clinical Implications 1 ´ M.A. Villaronga Torres1 , S. Tirados Menendez ´ , J.P. Rodrigo1 , S. AlvarezTeijeiro1 , D. Garc´ıa-Carracedo1 , R.G. Urdinguio2 , M.F. Fraga2 , L.A. Pardo3 , 1 C. Suarez ´ , J.M. Garc´ıa-Pedrero1 . 1 Hospital Universitario Central de Asturias (HUCA), Instituto Universitario de Oncolog´ıa del Principado de Asturias (IUOPA), Oviedo − Asturias, Spain, 2 Instituto Universitario de Oncolog´ıa del Principado de Asturias (IUOPA), Epigenetic Unit, Oviedo − Asturias, Spain, 3 Max-Planck-Institute of Experimental Medicine, Department of Molecular ¨ Biology of Neuronal Signals, Gottingen, Germany
Introduction: Compelling evidence indicates that the human ether-a-go-go voltage-gated potassium channels (hEAG1) may represent new valuable membrane therapeutic targets and diagnostic/prognostic biomarkers in various cancers. This study is the first to investigate the expression of hEAG1 potassium channel subunit in both primary tumors and HNSCC-derived cell lines to ascertain its clinical and biological role in the progression of head and neck squamous cell carcinomas. Material and Method: hEAG1 mRNA expression was analyzed by QRTPCR in a prospective series of 54 fresh HNSCC tissue specimens and 44 patient-matched normal epithelia. Molecular alterations were associated with clinicopathological parameters and disease outcome. To identify the mechanisms responsible for the aberrant expression of hEAG1 in HNSCC we studied the possible contribution of hEAG1 gene copy gain by Q-PCR in 88 HNSCC tissue specimens and also epigenetic transcriptional regulatory mechanisms (DNA methylation by pyrosequencing and histone acetylation by chromatin immunoprecipitation). Results and Discussion: Our findings demonstrate that hEAG1 is frequently aberrantly expressed in a high percentage of primary tumors (83%, 45/54 cases) and HNSCC-derived cell lines (83%, 10/12 cell lines). hEAG1 expression increased during HNSCC progression and was more frequent in advanced tumors. Strikingly, hEAG1 expression was also detected in a notable proportion (39%, 17/44 cases) of patient-matched normal adjacent mucosa, whereas no expression was detected in normal epithelia from nononcologic patients without exposure to tobacco carcinogens. In an attempt to
S191
identify the underlying mechanisms of aberrant hEAG1 expression in HNSCC, we found that hEAG1 gene copy gain occurred at a low frequency (15%, 13/88 cases) in primary tumors but was not observed in early stages of HNSCC tumorigenesis. Furthermore, this study provides original evidence supporting the involvement of histone acetylation (i.e. H3Ac and H4K16Ac activating marks) in the regulation of hEAG1 expression in HNSCC. In addition, functional studies in HNSCC cells further revealed that hEAG1 expression is a biologically relevant feature that promotes cell proliferation and invasion, although independently of its ion-conducting function. Conclusion: Our findings strongly support the notion that hEAG1 may represent a promising candidate as tumor marker and membrane therapeutic target for HNSCC treatment. 802 Circulating Tumor Cells After Neoadjuvant Chemotherapy Predict Survival in Non-metastatic Breast Cancer A. Lucci1 , A. Lodhi1 , A. Bhattacharyya1 , C. Hall1 , S. Jackson2 , B. Singh1 , S. Krishnamurthy3 , H. Kuerer1 . 1 UT MD Anderson Cancer Center, Surgical Oncology, Houston TX, USA, 2 UT MD Anderson Cancer Center, Breast Medical Oncology, Houston TX, USA, 3 UT MD Anderson Cancer Center, Pathology, Houston TX, USA Introduction: Circulating tumor cells (CTCs) predict outcome in metastatic breast cancer, but their significance is unclear in non-metastatic patients. Furthermore, it is unclear if the presence of CTCs after completion of neoadjuvant chemotherapy (NACT) predicts worse outcome. The purpose of this study was to determine if CTCs after NACT predict worse outcome. Methods: Clinical stage I-III breast cancer patients seen at a single tertiary cancer center provided informed consent to participate in an IRB-approved study involving collection of blood (7.5 ml x 2 tubes) at the time of surgery for their primary breast cancer. CTCs were detected using the CellSearch® system. A positive result was defined as the presence of one or more cells per 7.5 ml blood since the threshold for positivity has not been established in nonmetastatic breast cancer. Statistical analyses used chi-square and Fischer’s exact test. Results: One hundred and thirty seven patients were prospectively enrolled. Median age was 52 years and median follow-up was 34 months. Nine percent of patients had T1 disease, 36% T2, 20% T3, and 35% T4. Fifty-four percent of patients (73/137) had ER positive and 38% (52/137) had PR positive disease. Thirty percent of patients (41/137) were HER-2 positive. Twenty eight percent (38/137) had triple-negative tumors. Sixty-eight percent (93/137) had lymph node positive disease. One CTC was found in 27% (37/137) of patients postNACT, but its presence did not predict worse outcome (p = NS). Two or more CTCs were present in 9% (12/137) of patients. Of the 20 patients who relapsed, 6 had 2 or more CTCs (P = 0.002), while of the 14 patients who died, 4 had 2 or more CTCs (P = 0.001). Conclusions: Presence of two or more CTCs after NACT predicted worse relapse-free and overall survival in patients with stage I-III breast cancer. 803 Integrative Marker Analysis and Risk Assessment in Stage II Colon Cancer U. Nitsche1 , R. Rosenberg1 , T. Schuster2 , J. Slotta-Huspenina3 , F.G. Bader1 , H. Friess1 , P.M. Schlag4 , U. Stein5 , K.P. Janssen1 . 1 Klinikum Rechts der Isar, Chirurgische Klinik und Poliklinik, Munchen, ¨ Germany, 2 Klinikum Rechts der Isar, Institute of Statistics and Epidemiology, Munchen, ¨ Germany, 3 Klinikum Rechts der Isar, Institute of Pathology, Munchen, ¨ Germany, 4 Charite´ Medical Faculty and Max-Delbruck-Center ¨ for Molecular Medicine and Charite´ Comprehensive Cancer Center, Experimental and Clinical Research Center, Berlin, Germany, 5 Charite´ Medical Faculty and Max-Delbruck-Center ¨ for Molecular Medicine, Experimental and Clinical Research Center, Berlin, Germany Background: Colorectal cancer progression is governed by genetic and epigenetic alterations that can occur in multiple parallel pathways. However, with the exception of KRAS mutations, molecular markers currently play no role in risk stratification and therapy decision. E.g., risk assessment based on clinico-pathological parameters is not reliable for patients with stage II colon cancer (locally restricted disease). We hypothesize that the individual recurrence risk is defined by tumor genetics, and have therefore analyzed a panel of molecular genetic markers for risk prediction. Material and Methods: Non-microdissected fresh frozen tissue from 232 patients (T3−4 N0 M0) with complete tumor resection and median followup of 97 months was analyzed for microsatellite stability by multiplex PCR, KRAS exon 2, and BRAF exon 15 mutations by high-resolution melting analysis. Moreover, gene expression of the WNT-pathway surrogate marker osteopontin and the metastasis-associated genes SASH1 and MACC1 was determined for 179 patients. The results were correlated with metachronous distant metastasis as primary endpoint (n = 22 patients). Results: Mutations of KRAS were detected in 30% of patients, mutations of BRAF in 15%, and microsatellite instability in 26%, in good accordance with reported findings. Risk of recurrence was associated with KRAS mutation