The tumors were histologically subtyped and graded according to the third edition of the World Health Organization guidelines. The patients were classified according to gender, and their ages ranged from 28 to 78 years (median = 56 years). Clinical characteristics were retrieved from available clinical records. The clinico-pathological factors were

retrospectively assessed and are listed in Table 1. The normal control tissues consisted of two parts. Twenty-four matched adjacent non-malignant tissues were collected at sites at least 3 cm away from the edge of tumor mass. Efforts were done to avoiding contamination by the tumor cells. Twenty-two non-malignant tissues were obtained from the benign lung disease patients during lung volume reduction surgery. Table 1 Clinico-pathological features of lung cancer cases (N =96) Group Characteristics Number (%) Sex       Male 73(76.04%)   Female 23(23.96%) Selonsertib order Age       <60 54(56.25%)   ≥60 42(43.75%) Pack years of smoking

      >40 47(48.96%)   20.1–40 4(4.17%)   0.1–20 8(8.33%)   0 37(38.54%) LCZ696 nmr Histology       LAC 41(42.71%)   LSCC 39(40.63%)   SCLC 11(11.46%)   LCLC 3(3.13%)   Undifferentiated 2(2.83%) Pathologic grade       Poorly differentiated 26(27.08%)   Moderately differentiated 33(34.38%)   Well-differentiated 21(21.88%)   Others 16(16.67%) Clinical staging       IB 3(3.1%)   IIA-IIB 53(55.3%)   IIIA-IIIB 25(26.04%)   IV 4(4.1%)   Unavailable 11(11.46%) next Pleural invasion       Absent 82(85.42%)   Present 14(14.58%) Lymphatic invasion       Positive 55(57.29%)   Negative 41(42.71%) LAC, lung

adenocarcinoma; LSCC, lung squamous cell carcinoma; SCLC, small cell lung cancer; LCLC, large cell lung cancer. Preparation and identification of cell protein samples The cells were dissolved in a lysis buffer, and then centrifuged at 12,000 rpm for 30 min at 4°C. The supernatant was transferred to a fresh tube, and the selleck chemical cellular protein concentration was measured by the Bradford method. Trypsin (Promega, USA) was added to each of the groups, and equal amounts of proteins from each sample was added according to the protocol of the isobaric tags for relative and absolute quantization kit. The protein lysates of cells were labeled with the corresponding labeled reagent. The proteins were identified by 2D LC-MS /MS according to a method previously described [10]. The MS/MS spectra were collected in a data-dependent manner, in which up to four precursor ions above an intensity threshold of seven counts/s were selected for MS/MS analysis from each survey “scan.” In the tandem MS data database query, the peptide sequence tag (PKL) format files that were generated from MS/MS were imported into the Mascot search engine with an MS/MS tolerance of ± 0.05 Da to search the NCBInr database.