E. Lin

Epidermal Growth Factor Receptor Variant III Status Defines Clinically Distinct Subtypes of Glioblastoma

PURPOSE The clinical significance of epidermal growth factor receptor variant III (EGFRvIII) expression in glioblastoma multiforme (GBM) and its relationship with other key molecular markers are not clear. We sought to evaluate the clinical significance of GBM subtypes as defined by EGFRvIII status. PATIENTS AND METHODS The expression of EGFRvIII was assessed by immunohistochemistry in 649 patients with newly diagnosed GBM. These data were then examined in conjunction with the expression of phospho-intermediates (in a subset of these patients) of downstream AKT and Ras pathways and YKL-40 as well as with known clinical risk factors, including the Radiation Therapy Oncology Groups recursive partitioning analysis (RTOG-RPA) class. RESULTS The RTOG-RPA class was highly predictive of survival in EGFRvIII-negative patients but much less predictive in EGFRvIII-positive patients. These findings were seen in both an initial test set (n = 268) and a larger validation set (n = 381). Similarly, activation of the AKT/MAPK pathways and YKL-40 positivity were predictive of poor outcome in EGFRvIII-negative patients but not in EGFRvIII-positive patients. Pair-wise combinations of markers identified EGFRvIII and YKL-40 as prognostically important. In particular, outcome in patients with EGFRvIII-negative/YKL-40-negative tumors was significantly better than the outcome in patients with the other three combinations of these two markers. CONCLUSION Established prognostic factors in GBM were not predictive of outcome in the EGFRvIII-positive subset, although this requires confirmation in independent data sets. GBMs negative for both EGFRvIII and YKL-40 show less aggressive behavior.

Preoperative Bevacizumab Does Not Significantly Increase Postoperative Complication Rates in Patients Undergoing Hepatic Surgery for Colorectal Cancer Liver Metastases

PURPOSE Although bevacizumab (BV) increases survival rates when used with chemotherapy (CTX) in patients who have metastatic colorectal cancer (CRC), an increase in wound complications has been observed in patients who undergo surgery while receiving BV. We therefore evaluated whether neoadjuvant BV is associated with an increase in postoperative complications in patients undergoing surgery for CRC liver metastases. PATIENTS AND METHODS Two subgroups of patients who received neoadjuvant CTX + BV (n = 81) or CTX alone (n = 44) were identified from a database of patients who underwent surgery for CRC liver metastases. Univariate and multivariate logistic regression models were used to evaluate the association of patient and tumor characteristics, neoadjuvant therapy, and operative factors with postoperative complications. RESULTS Postoperative complications developed in 40 patients (49%) who received CTX + BV and 19 patients (43%) who received CTX. The median time from BV discontinuation to surgery was 58 days (range, 31 to 117 days). No significant associations were identified between BV use and timing of BV discontinuation and postoperative complications. On multivariate analysis, lower serum albumin and concomitant surgical procedures were associated with an increased risk of developing any complication (P = .035 and .023, respectively), and lower serum albumin was associated with hepatobiliary complications (P = .016). CONCLUSION Neither the use of BV nor timing of BV administration was associated with an increase in complication rates. These data suggest that the combination of BV with neoadjuvant CTX in patients who have CRC liver metastases does not increase surgical complications. To determine the optimal timing of surgery in patients receiving neoadjuvant BV, confirmatory prospective studies are required.

Prognostic Associations of Activated Mitogen-Activated Protein Kinase and Akt Pathways in Glioblastoma

Purpose: Activation of mitogen-activated protein kinase (MAPK) and members of the Akt pathway have been shown to promote cell proliferation, survival, and resistance to radiation. This study was conducted to determine whether any of these markers are associated with survival time and response to radiation in glioblastoma. Experimental Design: The expression of phosphorylated (p-)Akt, mammalian target of rapamycin (p-mTOR), p-p70S6K, and p-MAPK were assessed by immunohistochemical staining in 268 cases of newly diagnosed glioblastoma. YKL-40, a prognostic marker previously examined in these tumors, was also included in the analysis. Expression data were tested for correlations with response to radiation therapy in 131 subtotally resected cases and overall survival (in all cases). Results were validated in an analysis of 60 patients enrolled in clinical trials at a second institution. Results: Elevated p-MAPK expression was most strongly associated with poor response to radiotherapy, a finding corroborated in the validation cohort. For survival, higher expressions of p-mTOR, p-p70S6K, and p-MAPK were associated with worse outcome (all P < 0.03). YKL-40 expression was associated with the expressions of p-MAPK, p-mTOR, and p-p70S6K (all P < 0.02), with a trend toward association with p-Akt expression (P = 0.095). When known clinical variables were added to a multivariate analysis, only age, Karnofsky performance score, and p-MAPK expression emerged as independent prognostic factors. Conclusions: p-MAPK and activated members of the Akt pathway are markers of outcome in glioblastoma. Elevated expression of p-MAPK is associated with increased radiation resistance and represents an independent prognostic factor in these tumors.

Integrated Molecular and Clinical Analysis of AKT Activation in Metastatic Melanoma

Purpose: Activation of the phosphoinositide 3-kinase (PI3K)-AKT pathway has been implicated in melanoma based primarily on the prevalence of mutations in PTEN and NRAS. To improve our understanding of the regulation and clinical significance of the PI3K-AKT pathway in melanoma, we quantitatively measured the levels of phosphorylated AKT, its substrate GSK3α/β, and its negative regulator PTEN in clinical metastases. Results were compared with mutational status, clinical outcomes, and sites of metastasis. Experimental Design: DNA and protein were isolated from dissected frozen melanoma metastases (n = 96). Activating mutations of BRAF, NRAS, AKT, PIK3CA, and KIT were detected by mass spectroscopy genotyping. Phosphorylated AKT (Ser473 and Thr308), P-GSK3α/β, and PTEN protein expression were measured by reverse-phase protein array. A panel of human melanoma cells lines (n = 58) was analyzed for comparison. Results: BRAF-mutant tumors had higher levels of P-AKT-Ser473 (P = 0.01), P-AKT-Thr308 (P = 0.002), and P-GSK3α/β (P = 0.08) than NRAS-mutant tumors. Analysis of individual tumors showed that almost all tumors with elevated P-AKT had low PTEN levels; NRAS-mutant tumors had normal PTEN and lower P-AKT. Similar results were observed in melanoma cell lines. Stage III melanoma patients did not differ in overall survival based on activation status of the PI3K-AKT pathway. Brain metastases had significantly higher P-AKT and lower PTEN than lung or liver metastases. Conclusions: Quantitative interrogation of the PI3K-AKT pathway in melanoma reveals unexpected significant differences in AKT activation by NRAS mutation and PTEN loss, and hyperactivation of AKT in brain metastases. These findings have implications for the rational development of targeted therapy for this disease. (Clin Cancer Res 2009;15(24):7538–46)

Dermatofibrosarcoma protuberans treated with wide local excision and followed at a cancer hospital: prognostic significance of clinicopathologic variables.

Dermatofibrosarcoma protuberans (DFSP) is a relatively rare low-grade sarcoma. Local control can usually be achieved by wide local excision, but some patients still develop recurrences. The aim of this study was to investigate the correlation between clinicopathologic factors and recurrence-free survival (RFS)/overall survival (OS) in a large series of DFSP patients from a single institution. The study group included sections and medical records of 122 patients (63 women and 59 men, median age of 43) with primary DFSP from UT-MD Anderson Cancer Center between 1976 and 2005. Fibrosarcomatous change was detected in 24 (20.9%) patients. Thirty-eight of 120 patients (31.7%) recurred with a median RFS of 10.2 years. The 5-year RFS rate was 64.2%. Based on univariate analyses, fibrosarcomatous change, mitotic count, metastasis at time of diagnosis, and acral location were significantly associated with shorter RFS. On multivariate analysis, acral location and fibrosarcomatous change remained significant for shorter RFS. Five-year OS was 95.5% (95% confidence interval: 75.42%–99.3%). On univariate analysis, mitotic count per square millimeter, presence of necrosis, and metastasis at time of diagnosis were significantly associated with lower OS. On multivariate analysis, only presence of metastasis remained significantly associated with shorter OS. DFSP-FS variant and acral site are associated with shorter recurrence-free interval after wide local excision. Therefore, patients with tumors on acral sites or those with a fibrosarcomatous component may benefit from aggressive therapies other than wide local excision. The only factor that remains significantly associated with decreased OS is detection of metastasis.

Reactive Oxygen Species Generation in Human Cells by a Novel Magnetic Resonance Imaging Contrast Agent

The novel positive-contrast magnetic resonance imaging (MRI) marker C4 consists of an aqueous solution of cobalt chloride (CoCl2) complexed with the chelator N-acetylcysteine (NAC). We evaluated whether the presence of C4 or its components would produce reactive oxygen species (ROS, including hydroxyl, peroxyl, or other reactive oxygen species) in cultured cells. Human cancer or normal cells were incubated with 1% (w/v) CoCl2·6H2O or 2% NAC or a combination of both (1% CoCl2·6H2O : 2% NAC in an aqueous solution, abbreviated as Co : NAC) in the presence or absence of H2O2. Intracellular ROS levels were measured and quantified by change in relative fluorescence units. Students t-tests were used. In all cell lines exposed to 1000 μM H2O2, the Co : NAC led to ≥94.7% suppression of ROS at 5 minutes and completely suppressed ROS at 60 and 90 minutes; NAC suppressed ROS by ≥76.6% at 5 minutes and by ≥94.5% at 90 minutes; and CoCl2·6H2O suppressed ROS by ≥37.2% at 30 minutes and by ≥48.6% at 90 minutes. These results demonstrate that neither Co : NAC nor its components generated ROS; rather, they suppressed ROS production in cultured cells, suggesting that C4 would not enhance ROS production in clinical use.

Abstract 69: Proton versus photon irradiation induced cell death in head and neck cancer cells with different human papillomavirus status

Proton radiotherapy (PRT) has shown to be less toxic in the treatment of head and neck cancers (HNSCC) than photon radiotherapy (XRT). XRT causes multiple types of cancer cell death by inducing DNA double-strand breaks. The DNA damage induced by PRT is more complex and has more severe biological consequences versus XRT. The present study was undertaken to uncover the cell death caused by PRT versus XRT in human papillomavirus positive (HPV+, frequently p53 wild-type, p16 positive; p53, p16 are key factors for cell death) and HPV-negative (HPV-, frequently p53 mutant, p16 negative) HNSCC cells. HNSCC cell lines HN5, SqCC/Y1 (HPV-) and UMSCC-47, UPCI-SCC-154 (HPV+) were used. Single doses of 4 Gy irradiation induced cell apoptosis, necrosis, mitotic catastrophe, and senescence were determined. Clinical 200-MeV proton beams (18 cm × 18 cm field) or 6-MV X-ray beams (25 cm × 25 cm field) were used. Cells were positioned in the centers of the irradiation fields (in the middle of the spread-out Bragg peak for PRT). At 4 hours (h), 24h and 48h after PRT or XRT, cell necrosis and apoptosis were determined by Annexin VFITC-conjugated staining. XRT and PRT caused increased cell necrosis in the HPV– SqCC/Y1 (moderate at 24h, significant at 48h [p all In conclusion, mitotic catastrophe and senescence are the major types of cell death induced by both XRT and PRT, in which PRT inducing higher levels of both types of cell death as compared to XRT. Citation Format: Li Wang, Shichao Han, Jinming Zhu, Xiaochun Wang, Yuting Li, Zeming Wang, Eric Lin, Xiaofang Wang, David P. Molkentine, Pierre Blanchard, Yining Yang, Ruiping Zhang, Narayan Sahoo, Michael Gillin, Xiaorong Ronald Zhu, Xiaodong Zhang, Jeffrey N. Myers, Steven J. Frank. Proton versus photon irradiation induced cell death in head and neck cancer cells with different human papillomavirus status [abstract]. In: Proceedings of the AACR-AHNS Head and Neck Cancer Conference: Optimizing Survival and Quality of Life through Basic, Clinical, and Translational Research; April 23-25, 2017; San Diego, CA. Philadelphia (PA): AACR; Clin Cancer Res 2017;23(23_Suppl):Abstract nr 69.