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Radiological differential diagnosis between fibrosis and recurrence after SBRT in early stage NSCLC
S182
Abstracts
Material and Methods: Recurrent copy number alterations of chromosome fragments and genes were analyzed by high-resolution whole-genome sequencing in DNA obtained by paraffin-embedded samples from “discovery cohort” of 26 resected MPM patients treated with pemetrexedbased chemotherapy (8 with progressive disease (PD), vs. 18 with stable disease (SD) and 8 with partial response (PR)). Prognostic markers identified by Copy Number Variation analysis with Nexus, Control-FREEC and ReadDepth software were validated by PCR gene copy number and gene expression analyses both in the “discovery” and in two “validation” cohorts of pemetrexed-treated and untreated patients (N = 45 and 40). The role of emerging genes was evaluated through siRNA and pharmacological studies using proliferation, migration and apoptosis assays in MPM cells. Results: As reported previously we observed copy number loss of CDKN2A (15.4%) and BAP1 (7.7%), while only one patient had copy number loss of NF2. Interestingly, copy number gain of NOTCH2 was observed in 50% of samples of the patients who underwent progression, whereas losses of PTP4A3 gene were associated with clinical benefit (SD+PR). The prognostic relevance of NOTCH2 was confirmed by PCR analysis, as well as in the validation cohort. Moreover, Patients with high expression levels of both NOTCH2 and PTP4A3 had the worse prognosis (OS, 6 months), while no associations were found in pemetrexed-untreated patients. NOTCH-2 silencing reduced MPM cell migration and enhanced apoptosis induction by pemetrexed, while a PTP4A3 inhibitor overcame pemetrexed resistance in MPM cells characterized by high NOTCH2 and PTP4A3 expression. Conclusions: These results support the role of NOTCH2 as a novel prognostic/predictive biomarker for MPM, prompting prospective randomized trials for its validation. Moreover, preclinical data suggest that NOTCH2 and PTP4A3 are oncogenes suitable for effective therapeutic targeting in pemetrexed-resistant MPM cells. No conflict of interest. 2054A POSTER The clinical significance of osteopontin (OPN) in non-small cell lung cancer and its biological impact on lung cancer cells C. Hao1 , Y. Cui2 , M. Hu3 , X. Zhi3 , S. Cheng1 , W.G. Jiang2 . 1 Capital Medical University, Department of Biochemistry and Molecular Biology and Beijing Key Laboratory of Cancer & Metastasis Research- School of Basic Medical Sciences, Beijing, China; 2 Cardiff University School of Medicine, CCMRC, Cardiff, United Kingdom; 3 XuanWu Hospital- Capital Medical University, Department of Thoracic Surgery, Beijing, China Background: Osteopontin (OPN) is an extracellular matrix phosphoprotein secreted by a number of cell types in the body and implicated in a variety of physiological and pathological conditions. It has been reported that expression levels of OPN significantly correlate with disease prognosis of certain tumour types. OPN has been identified as a biomarker for tumor progression in many human tumors. In NSCLC, the tumorigenic functions of OPN are incompletely understood. The current study sought to investigate the level of OPN expression in a cohort of NSCLC patients and the association with clinical implications and prognosis. The impact of OPN on the molecular and cellular functions of lung cancer cells were also evaluated Methods: OPN expression in human NSCLC tissue and plasma samples (n = 75) was analysed using IHC and ELISA, respectively. The correlation between the levels of OPN and clinical characteristics were axamined. Knockdown of OPN in NSCLC cell line A549 (positive for OPN) was carried out using shRNA targeting human OPN. An OPN overexpression cell model with the NSCLC cell line SKMES-1 (negative for OPN) was also established. The effects of OPN on the functions of in these NSCLC cell models were determined using a variety of in vitro cell function assays. Results: Significantly higher tissue levels and raised plasma levels of OPN were observed in the cancer patients when compared to normal controls (p < 0.05 and p < 0.01 respectively). The OPN expression in plasma samples from patients with distant metastasis after surgical treatment was higher than that in those from patients who remained metastasis free (p < 0.05). Knockdown of OPN suppressed cell-matrix adhesion (p < 0.05), the in vitro migration (p < 0.05) and cell invasion (p < 0.05) in A549 cells. OPN overexpression in SKMES1 increased in vitro invasiveness, matrix adhesion and cellular migration (p < 0.05), when assessed using in vitro Matrigel invasion, matrix adhesion assay and electric cell-substrate impedance sensing, respectively. Conclusion: Our study shows that OPN expression is increased in lung cancer and is associated with distant metastasis. OPN plays crucial roles in regulating the migration, adhesion and invasion of lung cancer cells. Together, these data suggest that OPN may mediate an oncogenic effect on NSCLC cells and indicates that OPN may be a potential therapeutic target. No conflict of interest.
Poster Session, Sunday 29 January 2017 2055 POSTER Efficacy of postoperative radiotherapy in patients with pathological stage N2 EGFR wild type adenocarcinoma & squamous cell carcinoma lung cancer S.-Y. Wu1 . 1 Taipei Medical University − Wan Fang Medical Center, Department of Radiation Oncology, Taipei, Taiwan Background: There is a lack of large, prospective, randomized studies comparing postoperative radiotherapy (PORT) in pathological N2 (pN2) and surgical resection alone in terms of long-term survival in the setting of NSCLC. This national cohort study clarifies the role of PORT in the survival of pN2 non-small-cell lung cancer patients. Material and Methods: We analyzed data of patients with adenocarcinoma (adenoCA; wild-type [WT] epidermal growth factor receptor [EGFR]) and squamous cell carcinoma (squCA) collected from the Taiwan Cancer Registry database. The patients were categorized into five groups according to the treatment modality: Group 1, comprising those undergoing surgery alone; Group 2, comprising those undergoing adjuvant chemotherapy (CT) alone; Group 3, comprising those undergoing adjuvant radiotherapy (RT) alone; Group 4, comprising those receiving adjuvant concurrent chemoradiotherapy; and Group 5, comprising those receiving adjuvant sequential CT and intensity modulation RT. Results: We enrolled 588 lung adenoCA (WT EGFR) and squCA patients without distant metastasis. After adjustments for age at surgery, surgical years, and Charlson comorbidity index scores, the multivariate Cox regression analysis demonstrated that adjusted HRs (aHRs; 95% confidence intervals [CIs]) for the overall mortality of female lung adenoCA (WT EGFR) patients were 0.257 (0.111–0.594), 0.530 (0.226–1.243), 0.192 (0.069–0.534), and 0.399 (0.172–0.928) in Groups 2, 3, 4, and 5, respectively. For male lung squCA patients, the aHRs (95% CIs) for overall mortality were 0.269 (0.160–0.451), 0.802 (0.458–1.327), 0.597 (0.358– 0.998), and 0.456 (0.265–0.783) in Groups 2, 3, 4, and 5, respectively. Conclusions: Adjuvant CCRT or sequential CT and IMRT 5000 cGy significantly reduced the mortality rate of female lung adenoCA (WT EGFR) and male squCA pN2 patients. No conflict of interest. 2056 POSTER Radiological differential diagnosis between fibrosis and recurrence after SBRT in early stage NSCLC R. Frakulli1 , F. Salvi2 , D. Balestrini2 , M. Palombarini3 , V. Panni1 , V. Dionisi1 , A. Zamagni1 , M. Ferioli1 , E. Farina1 , F. Deodato4 , I. Mascia1 , S. Ciabatti1 , S. Cammelli1 , A. Arcelli1 , A.G. Morganti1 , M. Zompatori5 , G. Frezza2 . 1 University of Bologna- Ospedale S. Orsola Malpighi, Radiation Oncology Center- Department of Experimental- Diagnostic and Specialty MedicineDIMES, Bologna, Italy; 2 Bellaria Hospital, Radiation Oncology Unit, Bologna, Italy; 3 Bellaria Hospital, Physic Unit, Bologna, Italy; 4 Fondazione di Ricerca e Cura “Giovanni Paolo II”- Catholic University of Sacred Heart, Radiotherapy Unit, Campobasso, Italy; 5 University of Bologna- Ospedale S. Orsola Malpighi, Radiology Unit- Department of ExperimentalDiagnostic and Specialty Medicine- DIMES, Bologna, Italy Background: Parenchymal changes after Stereotactic body radiation therapy (SBRT) make differential diagnosis between treatment outcomes and disease recurrence often difficult. The purpose of our study is to identify the radiographic features detectable at computed tomography (CT) scan (High Risk Features: HRFs) that allow enough specificity and sensitivity for early detection of recurrence. Material and Methods: We retrospectively evaluated patients who underwent SBRT for inoperable early stage NSCLC. The median delivered dose was 50 Gy in 5 fractions prescribed to 80% isodose. All patients underwent chest CT scan before SBRT and at 3, 6, 12, 18, 24 months after, and then annually. Each CT scan was evaluated and benign and high risk features were recorded. 18 F FDG PET/CT was not used routinely. Results: Forty-five patients were included (34 males, 11 females; median age: 77 years; stage IA: 77.8%, stage IB: 22.2%, median follow-up: 21.7 months). Two-year and actuarial local control was 77%. HRFs were identified in 20 patients. The most significant predictor of relapse was an enlarging opacity at 12 months (p < 0001) with 84.6% sensitivity and 71.8% specificity. The presence of >2 HRFs demonstrated a high sensitivity (92.3%) and specificity (71.9%) (p < 00001). Conclusions: Detection of HRFS is predictive of relapse with a sensitivity that increases with the number of HRF’s observed. This observation may allow to better define the diagnostic follow up algorithm suggesting to perform further exams only in patients with >2 HRFs. No conflict of interest.
Abstracts
Material and Methods: Recurrent copy number alterations of chromosome fragments and genes were analyzed by high-resolution whole-genome sequencing in DNA obtained by paraffin-embedded samples from “discovery cohort” of 26 resected MPM patients treated with pemetrexedbased chemotherapy (8 with progressive disease (PD), vs. 18 with stable disease (SD) and 8 with partial response (PR)). Prognostic markers identified by Copy Number Variation analysis with Nexus, Control-FREEC and ReadDepth software were validated by PCR gene copy number and gene expression analyses both in the “discovery” and in two “validation” cohorts of pemetrexed-treated and untreated patients (N = 45 and 40). The role of emerging genes was evaluated through siRNA and pharmacological studies using proliferation, migration and apoptosis assays in MPM cells. Results: As reported previously we observed copy number loss of CDKN2A (15.4%) and BAP1 (7.7%), while only one patient had copy number loss of NF2. Interestingly, copy number gain of NOTCH2 was observed in 50% of samples of the patients who underwent progression, whereas losses of PTP4A3 gene were associated with clinical benefit (SD+PR). The prognostic relevance of NOTCH2 was confirmed by PCR analysis, as well as in the validation cohort. Moreover, Patients with high expression levels of both NOTCH2 and PTP4A3 had the worse prognosis (OS, 6 months), while no associations were found in pemetrexed-untreated patients. NOTCH-2 silencing reduced MPM cell migration and enhanced apoptosis induction by pemetrexed, while a PTP4A3 inhibitor overcame pemetrexed resistance in MPM cells characterized by high NOTCH2 and PTP4A3 expression. Conclusions: These results support the role of NOTCH2 as a novel prognostic/predictive biomarker for MPM, prompting prospective randomized trials for its validation. Moreover, preclinical data suggest that NOTCH2 and PTP4A3 are oncogenes suitable for effective therapeutic targeting in pemetrexed-resistant MPM cells. No conflict of interest. 2054A POSTER The clinical significance of osteopontin (OPN) in non-small cell lung cancer and its biological impact on lung cancer cells C. Hao1 , Y. Cui2 , M. Hu3 , X. Zhi3 , S. Cheng1 , W.G. Jiang2 . 1 Capital Medical University, Department of Biochemistry and Molecular Biology and Beijing Key Laboratory of Cancer & Metastasis Research- School of Basic Medical Sciences, Beijing, China; 2 Cardiff University School of Medicine, CCMRC, Cardiff, United Kingdom; 3 XuanWu Hospital- Capital Medical University, Department of Thoracic Surgery, Beijing, China Background: Osteopontin (OPN) is an extracellular matrix phosphoprotein secreted by a number of cell types in the body and implicated in a variety of physiological and pathological conditions. It has been reported that expression levels of OPN significantly correlate with disease prognosis of certain tumour types. OPN has been identified as a biomarker for tumor progression in many human tumors. In NSCLC, the tumorigenic functions of OPN are incompletely understood. The current study sought to investigate the level of OPN expression in a cohort of NSCLC patients and the association with clinical implications and prognosis. The impact of OPN on the molecular and cellular functions of lung cancer cells were also evaluated Methods: OPN expression in human NSCLC tissue and plasma samples (n = 75) was analysed using IHC and ELISA, respectively. The correlation between the levels of OPN and clinical characteristics were axamined. Knockdown of OPN in NSCLC cell line A549 (positive for OPN) was carried out using shRNA targeting human OPN. An OPN overexpression cell model with the NSCLC cell line SKMES-1 (negative for OPN) was also established. The effects of OPN on the functions of in these NSCLC cell models were determined using a variety of in vitro cell function assays. Results: Significantly higher tissue levels and raised plasma levels of OPN were observed in the cancer patients when compared to normal controls (p < 0.05 and p < 0.01 respectively). The OPN expression in plasma samples from patients with distant metastasis after surgical treatment was higher than that in those from patients who remained metastasis free (p < 0.05). Knockdown of OPN suppressed cell-matrix adhesion (p < 0.05), the in vitro migration (p < 0.05) and cell invasion (p < 0.05) in A549 cells. OPN overexpression in SKMES1 increased in vitro invasiveness, matrix adhesion and cellular migration (p < 0.05), when assessed using in vitro Matrigel invasion, matrix adhesion assay and electric cell-substrate impedance sensing, respectively. Conclusion: Our study shows that OPN expression is increased in lung cancer and is associated with distant metastasis. OPN plays crucial roles in regulating the migration, adhesion and invasion of lung cancer cells. Together, these data suggest that OPN may mediate an oncogenic effect on NSCLC cells and indicates that OPN may be a potential therapeutic target. No conflict of interest.
Poster Session, Sunday 29 January 2017 2055 POSTER Efficacy of postoperative radiotherapy in patients with pathological stage N2 EGFR wild type adenocarcinoma & squamous cell carcinoma lung cancer S.-Y. Wu1 . 1 Taipei Medical University − Wan Fang Medical Center, Department of Radiation Oncology, Taipei, Taiwan Background: There is a lack of large, prospective, randomized studies comparing postoperative radiotherapy (PORT) in pathological N2 (pN2) and surgical resection alone in terms of long-term survival in the setting of NSCLC. This national cohort study clarifies the role of PORT in the survival of pN2 non-small-cell lung cancer patients. Material and Methods: We analyzed data of patients with adenocarcinoma (adenoCA; wild-type [WT] epidermal growth factor receptor [EGFR]) and squamous cell carcinoma (squCA) collected from the Taiwan Cancer Registry database. The patients were categorized into five groups according to the treatment modality: Group 1, comprising those undergoing surgery alone; Group 2, comprising those undergoing adjuvant chemotherapy (CT) alone; Group 3, comprising those undergoing adjuvant radiotherapy (RT) alone; Group 4, comprising those receiving adjuvant concurrent chemoradiotherapy; and Group 5, comprising those receiving adjuvant sequential CT and intensity modulation RT. Results: We enrolled 588 lung adenoCA (WT EGFR) and squCA patients without distant metastasis. After adjustments for age at surgery, surgical years, and Charlson comorbidity index scores, the multivariate Cox regression analysis demonstrated that adjusted HRs (aHRs; 95% confidence intervals [CIs]) for the overall mortality of female lung adenoCA (WT EGFR) patients were 0.257 (0.111–0.594), 0.530 (0.226–1.243), 0.192 (0.069–0.534), and 0.399 (0.172–0.928) in Groups 2, 3, 4, and 5, respectively. For male lung squCA patients, the aHRs (95% CIs) for overall mortality were 0.269 (0.160–0.451), 0.802 (0.458–1.327), 0.597 (0.358– 0.998), and 0.456 (0.265–0.783) in Groups 2, 3, 4, and 5, respectively. Conclusions: Adjuvant CCRT or sequential CT and IMRT 5000 cGy significantly reduced the mortality rate of female lung adenoCA (WT EGFR) and male squCA pN2 patients. No conflict of interest. 2056 POSTER Radiological differential diagnosis between fibrosis and recurrence after SBRT in early stage NSCLC R. Frakulli1 , F. Salvi2 , D. Balestrini2 , M. Palombarini3 , V. Panni1 , V. Dionisi1 , A. Zamagni1 , M. Ferioli1 , E. Farina1 , F. Deodato4 , I. Mascia1 , S. Ciabatti1 , S. Cammelli1 , A. Arcelli1 , A.G. Morganti1 , M. Zompatori5 , G. Frezza2 . 1 University of Bologna- Ospedale S. Orsola Malpighi, Radiation Oncology Center- Department of Experimental- Diagnostic and Specialty MedicineDIMES, Bologna, Italy; 2 Bellaria Hospital, Radiation Oncology Unit, Bologna, Italy; 3 Bellaria Hospital, Physic Unit, Bologna, Italy; 4 Fondazione di Ricerca e Cura “Giovanni Paolo II”- Catholic University of Sacred Heart, Radiotherapy Unit, Campobasso, Italy; 5 University of Bologna- Ospedale S. Orsola Malpighi, Radiology Unit- Department of ExperimentalDiagnostic and Specialty Medicine- DIMES, Bologna, Italy Background: Parenchymal changes after Stereotactic body radiation therapy (SBRT) make differential diagnosis between treatment outcomes and disease recurrence often difficult. The purpose of our study is to identify the radiographic features detectable at computed tomography (CT) scan (High Risk Features: HRFs) that allow enough specificity and sensitivity for early detection of recurrence. Material and Methods: We retrospectively evaluated patients who underwent SBRT for inoperable early stage NSCLC. The median delivered dose was 50 Gy in 5 fractions prescribed to 80% isodose. All patients underwent chest CT scan before SBRT and at 3, 6, 12, 18, 24 months after, and then annually. Each CT scan was evaluated and benign and high risk features were recorded. 18 F FDG PET/CT was not used routinely. Results: Forty-five patients were included (34 males, 11 females; median age: 77 years; stage IA: 77.8%, stage IB: 22.2%, median follow-up: 21.7 months). Two-year and actuarial local control was 77%. HRFs were identified in 20 patients. The most significant predictor of relapse was an enlarging opacity at 12 months (p < 0001) with 84.6% sensitivity and 71.8% specificity. The presence of >2 HRFs demonstrated a high sensitivity (92.3%) and specificity (71.9%) (p < 00001). Conclusions: Detection of HRFS is predictive of relapse with a sensitivity that increases with the number of HRF’s observed. This observation may allow to better define the diagnostic follow up algorithm suggesting to perform further exams only in patients with >2 HRFs. No conflict of interest.