Localization of PKCη in cell membranes as a predictor for breast cancer response to treatment.

BACKGROUND: Successful treatment of breast cancer is frequently limited by the resistance of tumors to chemotherapy. Recent studies suggested a role for protein kinase C (PKC) in the resistance to chemotherapy. Here we used retrospective analysis of breast cancer biopsies of neoadjuvantly treated patients to investigate the correlation of PKC expression with aggressiveness and resistance to chemotherapy. PATIENTS AND METHODS: Our cohort (n = 25) included patients with advanced and aggressive breast cancers, who underwent neoadjuvant therapy with the CAF regimen (cyclophosphamide, doxorubicin, fluorouracil). Core biopsies (pre-chemotherapy) and surgical biopsies of primary tumors and lymph node metastases (post-chemotherapy) were scored for PKCeta (PKCh) and PKCepsilon (PKCe) expression in the cytoplasm, cell membrane, nuclear membrane, and the nucleus. RESULTS: Our results showed increased expression of PKCh (not PKCe) in the cytoplasm and cell membranes of post-chemotherapy biopsies (p = 0.03). PKCh presence in cell membranes, indicating activation, was in correlation with poor survival (p = 0.007). CONCLUSION: PKCh staining in cell and nuclear membranes is an indicator for poor survival and a predictor for the effectiveness of neoadjuvant treatment. Other avenues of treatment should be considered for these patients. PKCh presents a target for therapy where inhibition of its activity and/or translocation to membranes could interfere with the resistance to chemotherapy.

Oral lipoma: analysis of 58 new cases and review of the literature.

Lipomas are common soft-tissue tumors but occur infrequently in the oral region. Here, we present 58 new cases of oral lipoma (OL), their clinical features, diagnostic workup, and management. The records of 58 patients with OL were reviewed. Demographic data, presenting symptoms, primary tumor site, histopathology, and clinical management were analyzed. Extraoral and intraosseous lesions were excluded. The patients were 29 men and 29 women with mean age of 59.7 years, range, 11 to 98 years. Tumor sites included the buccal mucosa (n = 31), tongue (n = 10), lip (n = 6), floor of the mouth (n = 6), and vestibule (n = 5). The mean tumor size was 2.1 cm (range, 0.3-5.0 cm). The mean tumor duration before excision was 2.6 years (31.2 months), ranging from 2 months to 12 years. Most common presenting symptom was an asymptomatic, circumscribed mass. Histologically, the tumors were classified as classic lipoma (n = 28), fibrolipoma (n = 19), intramuscular lipoma (n = 4), minor salivary gland lipoma (n = 2), angiolipoma (n = 2), and spindle cell lipoma (n = 3). A case of spindle cell lipoma affecting the lower lip that has not been described previously is presented. The provided treatment was surgical excision. Although uncommon, OL are most common in adults and very uncommon in children. No sex predilection was found. The buccal mucosa region is the most common site followed by the tongue. Classic lipomas are most common in the oral region and comprise 48% of the tumors. A case of spindle cell lipoma of the lower lip is reported. Its clinical, histologic, and cytogenetic workup is the first well-documented case. Further cytogenetic studies should be undertaken to learn more about the pathogenesis and tumorigenesis of OL.

Measles virus: evidence for association with lung cancer.

In recent years the frequency of nonsmokers among lung cancer patients has increased to 10% to 15%. The measles virus has rarely been evoked as an etiological agent in malignant tumors and its role in carcinogenesis remains doubtful. It has been suggested that measles virus phosphoprotein may inhibit ubiquitination of Pirh2, which has been reported to be overexpressed in lung carcinoma and is responsible for degrading the cell cycle regulator p53. The authors conducted a clinicopathological study of newly diagnosed patients with non-small cell lung carcinoma of all stages seen in a 10-year period. Immunohistochemical studies for measles virus antigens, p53, and Pirh2 were performed using the avidin-biotin peroxidase complex. The authors found expression of measles virus antigens in 54 of 65 cases of non-small cell lung carcinoma. This finding is associated with the older age of the patients and with expression of Pirh2. The presence of Pirh2 itself was associated with improved survival.

Ethnicity and its significance in the pathobiology of prostatic carcinoma in Southern Israel.

BACKGROUND: Several studies have noted ethnic differences in the natural history of prostatic carcinoma. Southern Israel has been regarded as a melting pot and, perhaps more than the rest of the country, has encouraged the ingathering of immigrants from several countries, as well as a large Bedouin community. OBJECTIVES: In an attempt to determine any differences that may exist in population groups in Israel, we have examined clinical and biologic markers in patients diagnosed with prostatic cancer in Southern Israel in 1996-2000. We wanted to demonstrate differences in the incidence and features of prostate carcinoma among the population groups in Southern Israel, and to evaluate their possible biologic significance. METHODS: Clinical parameter features, including the ethnicity origin of patients with prostatic adenocarcinoma, were reviewed in a cohort of 189 patients seen between 1996 and 2000. Tissue sections from specimens in a subset of 40 of these patients who had undergone prostatectomy were studied by immunohistochemistry for TP53, Bcl-2, and chromogranin A using the ABC peroxidase method. These markers were chosen because of their suggested impact on the biology of this tumor. Clinical correlations were examined. RESULTS: We confirm the presence of ethnic differences in the features of prostatic adenocarcinoma in our geographic area. Notably, patients of North African origin were treated surgically at a younger age than immigrants from East Europe. Higher total prostate-specific antigen levels and more robust tumor cell Bcl-2 expression were detected in the East European patients. The number of Bedouin subjects in our cohort of patients with prostatic cancer was much more limited than expected. No immigrants from Ethiopia were included in our study diagnosed with prostate carcinoma during this period. CONCLUSIONS: The proportion of patients of European, especially East European, origin was relatively high among the cohort of 189. Their older age and the lower proportion of subjects that underwent surgery, together with the tendency toward higher total prostate-specific antigen levels and higher Bcl-2 expression, suggest that this ethnic group may not differ significantly from the African-American group in the United States. The low representation of Bedouin and absence of Ethiopian immigrants among our patients with prostate cancer may point to a genuinely low incidence or it may be related to inadequate medical supervision in these population groups.

MMP-2, TIMP-1, E-cadherin, and beta-catenin expression in endometrial serous carcinoma compared with low-grade endometrial endometrioid carcinoma and proliferative endometrium.

OBJECTIVE: To investigate the expression of MMP-2, TIMP-1, E-cadherin and beta-catenin in endometrial serous carcinoma (ESC), low-grade endometrial endometrioid carcinoma (EEC), and proliferative endometrium. METHODS: We performed an immunohistochemical study on 14 cases of ESC, 15 cases of low-grade EEC, and 10 cases of proliferative endometrium. RESULTS: Compared with low-grade EEC, ESC showed significantly increased MMP-2 and TIMP-1 expression, as well as decreased membranous beta-catenin staining. E-cadherin expression was significantly lower in ESC and EEC as compared with proliferative endometrium. CONCLUSIONS: We suggest that MMP-2 and TIMP-1 expression and loss of beta-catenin have a role in the pathogenesis and progression of ESC. Decreased E-cadherin may have an important role in the development of both ESC and EEC. Furthermore, the dissimilarities in MMP-2, TIMP-1, E-cadherin and beta-catenin expressions in ESC compared with EEC may be responsible, along with other factors, for their different biological behavior.

Matrix metalloproteinases 2 and 9, E-cadherin, and beta-catenin expression in endometriosis, low-grade endometrial carcinoma and non-neoplastic eutopic endometrium.

OBJECTIVES: Endometriosis and endometrial endometrioid carcinoma are both capable of invasion and metastasis, but their biological behavior is strikingly different. Matrix metalloproteinases (MMPs) and changes in adhesion molecules have a role in the pathogenesis of various physiological and pathological processes, as well as in the development of endometriosis and endometrioid endometrial carcinoma. We hypothesized that endometriosis, being a benign process, will show different MMPs and adhesion molecules expressions, compared to endometrioid endometrial carcinoma, a disease with potential of malignant behavior. STUDY DESIGN: We performed an immunohistochemical study to investigate expression of matrix metalloproteinase-2 (MMP-2), matrix metalloproteinase-9 (MMP-9), E-cadherin and beta-catenin in endometriosis, low-grade endometrial endometrioid carcinoma, and eutopic proliferative endometrium. Endometriotic tissues (n=15), low-grade endometrial endometrioid carcinomas (n=15), and unremarkable proliferative endometrium from women without endometriosis or carcinoma (n=10) were examined. RESULTS: Endometriotic tissues showed statistically significantly stronger staining for MMP-9 and reduced beta-catenin expression when compared with proliferative endometrium. Endometrial endometrioid carcinoma showed decreased E-cadherin expression in comparison with proliferative endometrium. MMP-2 and MMP-9, and E-cadherin expressions were significantly higher and beta-catenin expression was significantly lower in endometriosis as compared to endometrioid carcinoma. CONCLUSIONS: We suggest that increased MMPs and altered beta-catenin may play a role in the pathogenesis of endometriosis. Decreased E-cadherin may be important in the development of endometrial endometrioid carcinoma. The changes in MMPs, E-cadherin and beta-catenin differ in endometriosis from those in endometrioid carcinoma, an interesting finding in view of the fact that both these diseases are capable of invasion and metastasis, but have different biological behavior.

A rare case of enterobacter endocarditis superimposed on a mitral valve rheumatoid nodule.

We present the case of a 56-year-old man with longstanding seropositive active erosive and deforming rheumatoid arthritis with no peripheral rheumatoid nodules; he immigrated from the former Soviet Union (where he did not receive any disease-modifying antirheumatic drugs) to Israel in 1995. In February 2005, he had a buccogingival mucosal abscess on his lower lip, which was treated by surgical drainage, followed by prolonged antibiotic therapy. One and a half years later, he had 2 episodes of transient ischemic attacks characterized by speech difficulties and moderate weakness on his right side. Transesophageal echocardiogram revealed a mass on the anterior mitral valve leaflet. Repeated blood cultures were negative, and the patient was afebrile all the time. The patient underwent mitral valve replacement and the histologic findings of the mass were typical of both a rheumatoid nodule and bacterial endocarditis. The patient recovered fully after 6 weeks of antibiotic therapy. Emboli from a rheumatoid nodule should always be considered in patients with rheumatoid arthritis who present with transient ischemic attacks.

Solitary angiokeratoma of the tongue.

Angiokeratoma is a rare cutaneous lesion. It can be either a generalized systemic form, presenting as multiple asymptomatic papules on the skin, associated with metabolic diseases or a solitary cutaneous form. Oral cavity involvement is more common in the systemic form, as a part of a more generalized cutaneous disease, but very rare in the localized form of angiokeratoma. A 45-year-old female presented with a painless lesion on the tongue of one months duration, which bled occasionally. On clinical examination, a lesion of approximately 5 mm in diameter was observed on the left surface of the tongue. The lesion was purple in color with a granulomatous appearance. There were no other changes in the oral mucosa. On dermatologic examination, no angiokeratomas were found, anywhere on the skin. The lesion was excised under local anesthesia. The histologic diagnosis was angiokeratoma. A case of a solitary angiokeratoma of the tongue is reported. We report here the third intra-oral case and the second case in the tongue with solitary angiokeratoma.

Macrophage infiltration into omental versus subcutaneous fat across different populations: effect of regional adiposity and the comorbidities of obesity.

CONTEXT: Macrophage infiltration into adipose tissue has been demonstrated to accompany obesity, with a potential preferential infiltration into intraabdominal vs. sc fat. OBJECTIVE: Our objective was to determine whether this occurs across different populations with a range of body mass indexes and to assess the relationship with regional adiposity and comorbidity of obesity. SETTING AND PATIENTS: In two independent cohorts, we used paired omental (OM) and sc fat biopsies from lean controls or predominantly sc or intraabdominally obese persons with minimal comorbidity (n = 60, cohort 1), or from severely obese women with a significant rate of comorbidity (n = 29, cohort 2). RESULTS: Elevated macrophage infiltration into OM vs. sc fat was observable in lean subjects and exaggerated by obesity, particularly if predominantly intraabdominal. This was paralleled by increased monocyte chemoattractant protein-1 (MCP1) and colony-stimulating factor-1 (CSF1) mRNA levels. Level of CSF1 and MCP1 mRNA correlated with the number of OM macrophages (r = 0.521, P < 0.0001 and r = 0.258, P < 0.051, respectively). In severely obese women (mean body mass index = 43.0 +/- 1.1 kg/m(2)), higher protein expression of both MCP1 and CSF1 was detected in OM vs. sc fat. Number of OM macrophages, but not of sc macrophages, correlated with waist circumference (r = 0.636, P = 0.001 vs. r = 0.170, P = 0.427) and with the number of metabolic syndrome parameters (r = 0.385, P = 0.065 vs. r = -0.158, P = 0.472, respectively). Preferential macrophage infiltration into OM fat was mainly observed in a subgroup in whom obesity was associated with impaired glucose homeostasis. CONCLUSIONS: Preferential macrophage infiltration into OM fat is a general phenomenon exaggerated by central obesity, potentially linking central adiposity with increased risk of diabetes and coronary heart disease.

Cytogenetic and fluorescence in situ hybridization analysis of a basal cell adenocarcinoma of the mandible.

Basal cell adenocarcinoma (BCAC) of the salivary glands is rare. Distant metastasis to the mandible from a salivary gland tumor is also considered rare. The cytogenetic finding of a case of metastatic BCAC of the mandible is described. We are unaware of earlier reports regarding cytogenetic findings of BCAC either at the primary site or at a distant metastasis site. An 80-year-old female with primary BCAC of the parotid salivary gland underwent parotidectomy and chemotherapy. One year later, a metastatic lesion in the mandible was found. Tissue specimens from the mandibular lesion were tested by the following pathologic methods: hematoxylin-eosin and immunohistochemistry for CK8/18, CK/903, vimentin, and smooth muscle actin. The characteristic histologic architecture of BCAC found in the mandible was similar to that of the earlier findings of the tumor in the parotid gland. A fresh sample from the mandibular lesion was examined by cytogenetics and fluorescence in situ hybridization (FISH), using centromeric probes for chromosomes 4, 8, 10, 18, and 22. A paraffin-embedded sample of the primary tumor was also examined by FISH. Cytogenetic and FISH analyses of the mandibular metastatic lesion revealed a clone with a pericentric inversion of chromosome 17 and a clone with trisomy 4, respectively. Trisomy 4 was also found in the paraffin-embedded samples of the primary parotid tumor.

Metastatic tumors to the jaws: a report of eight new cases.

PURPOSE: The purpose of the article is to present 8 new cases of metastatic tumors occurring in the jawbones, their clinical features , diagnostic workup and management. PATIENTS AND METHODS: The records of 8 patients with metastatic jaw lesions were reviewed. Demographic data, presenting symptoms, primary tumor site, radiographic findings, bone scintigraphy , histopathology and clinical management were analyzed. RESULTS: The patients , ranged in age from 44 to 80 years, with a mean of 64.5 years. The primary malignant sites were: the lung , the breast , the rectum, the thyroid, the uterus and the parotid gland . The mandible was the site of oral involvement in seven cases and the maxilla in one. There was no gender difference with respect to the oral site affected. The clinical jaw presentations were: exophytic soft tissue mass, paresthesia of the lower lip and a periapical lesion The provided treatment protocols were: chemotherapy , radiotherapy and chemotherapy, surgery and chemotherapy and supportive care only. In one case the jaw lesion was the first indication of an unknown malignancy at a distant primary site. CONCLUSIONS: Metastatic jaw lesions are uncommon. Paresthesia of the lower lip and the chin is a sinister sign for patients with a metastatic jaw lesion. In view of these cases it can be said that meticulous work-up of of jaw lesions suspected of being metastatic, may be life saving or extend the patient s survival period.

Adherence of Hodgkin's disease-derived cell lines to the various lymph node compartments.

This adherence study was performed to clarify the trafficking of Hodgkin-Reed-Sternberg (HRS) cells in Hodgkin's disease (HD) and thus try to unravel the peculiar pathways of dissemination in the early stages of this malignant neoplasm. Using non-neoplastic human necropsy or biopsy lymph node as well as tonsillar tissue sections, we have studied the adherence of the KMH-2 and L-428 HD-derived cell lines and have compared it to that of peripheral blood lymphocytes (PBLs). In necropsy lymph nodes, cell lines predominantly adhered to sinuses and paracortex, whilst adhered PBLs were distributed more widely. In biopsy lymph nodes, adhesion to high endothelial venules (HEVs) was rarely observed, whilst cell lines were found to adhere to sinuses. Inhibition by EDTA pretreatment affected adherence to HEVs as well as to sinuses and paracortex to a similar degree. Our findings point to the possible importance of the lymph node sinuses and paracortex in relation to homing of the HRS cells and their dissemination during the early stages of HD. The results suggest a significant primary role of the extracellular matrix of the paracortex and sinuses in the homing of HRS cells, with the HEVs of only secondary importance.

Augmented expression of chromogranin A and serotonin in peri-malignant benign prostate epithelium as compared to adenocarcinoma.

There is a growing body of evidence that the occurrence of neuroendocrine (NE) differentiation in prostate carcinoma correlates with poor prognosis, tumor progression, and androgen-independence. In the present study, the expression of common NE markers, i.e., chromogranin A (ChGA), serotonin (5HT), neuron-specific enolase (NSE) and adrenomedullin (AM), was retrospectively examined in formalin-fixed, paraffin-embedded prostate tissue samples obtained from patients with adenocarcinoma and from patients with nodular hyperplasia of the prostatic gland (NHPG) (33 and 28, respectively). The statistical analysis of the results (tested the equality of matched pairs of observations using the Wilcoxon matched-pairs signed ranks test) revealed a more prominent expression of ChGA in benign epithelial cells adjacent to adenocarcinomatic lesions (Peri-PAC) than in the adenocarcinoma (PAC) (p = 0.0049). A similar pattern of expression was detected for 5HT (p = 0.000). When comparing the expression of ChGA and 5HT in tissue samples originating in cancer patients with those obtained from NHPG samples, more ChGA and 5HT were expressed in Peri-PAC than in NHPG (p = 0.0004 and 0.002, respectively). The results obtained raise the possibility that adenocarcinoma cells urge some adjacent benign epithelial cells to differentiate into NE cells, which, in turn, may promote tumor growth and invasion.

EML4-ALK-positive non-small cell lung cancer in a patient treated with azathioprine for ulcerative colitis.

EML4-ALK-positive lung cancer is a novel cancer entity associated with light or never smoking, younger age, and adenocarcinoma with acinar or signet-ring cell type histology. Another mutation of ALK with NPM, resulting in NPM-ALK fusion mutation, was described in patients with anaplastic large cell lymphoma (ALCL). It was subsequently reported in organ transplant recipients and patiens undergoing immunosuppressive therapy. We describe a case of lung cancer in a 36-year-old nonsmoking woman with ulcerative colitis treated with azathioprine, who was diagnosed with EML4-ALK-positive, metastatic lung cancer two months postpartum. Crizotinib 300 mg/day has been effective in maintaining response after chemotherapy failed. The resemblance of this case to ALK-positive ALCL in organ transplant recipients suggests that similar mechanisms may be responsible for the development of both ALK-positive lung cancer and ALCL in patients receiving immunosuppressive therapy.

Solitary fibrous tumor of the buccal vestibule: report of two cases.

Solitary fibrous tumor (SFT) is a rare benign tumor that occurs most frequently in the pleura. It is considered rare in the maxillofacial area. Two new cases of SFT of the buccal vestibule are reported. The previously reported cases of oral SFT are reviewed. The tumors were composed of spindle-shaped cells that were arranged haphazardly and were positive for CD-34, BCL-2, CD-99 and vimentin. Although rare, SFT should be included in the differential diagnosis of oral soft tissue tumors. The clinical presentation and imaging can provide the clinician a better tool for preoperative diagnosis.

PKCη is a novel prognostic marker in non-small cell lung cancer.

BACKGROUND: Novel biomarkers which may serve as therapeutic targets are essential for lung cancer treatment. Here we investigated the prognostic significance of protein kinase Cη (PKCη), a cell cycle regulator involved in tumorigenesis and chemotherapy resistance, in patients diagnosed with non-small cell lung cancer (NSCLC). PATIENTS AND METHODS: Sixty-three chemotherapy-naïve patients were examined for PKCη by immunohistochemistry and divided into PKCη H-Score tertiles (low, intermediate and high). Time until event (relapse or mortality) within one year was determined using Cochran-Armitage test and Cox proportional hazards regression model. RESULTS: The distribution of patients according to clinical stage 1-4 was: 27%, 5%, 26% and 42%, respectively. PKCη overexpression was associated with advanced stage (p=0.03) and the risk for an event (p=0.045). Patients of the lowest tertile were less likely to experience an event. CONCLUSION: PKCη is a novel prognostic marker in NSCLC that may predict poor prognosis. The use of PKCη-specific inhibitors in NSCLC may prove valuable.

Measles virus antigens in breast cancer.

INTRODUCTION: Breast cancer (BC) is the most common cancer in women in Western countries, showing a bimodal age distribution with peaks at 50 and 70 years. Multiple factors are encountered in the etiology of BC, including hereditary and hormonal causes. A few viruses, including MMTV, EBV and HPV have been reported to be associated with BC. This group previously described the presence of the measles virus (MV) in biopsies of patients with Hodgkin's lymphoma. The present study explored MV antigens in BC patients. PATIENTS AND METHODS: A total of 131 patients with invasive BC diagnosed during the years 1998-2005 were studied using immunohistochemistry (IHC) for the presence of MV antigens, hemagglutinin and nucleoprotein. Clinicopathological parameters included age, stage, histological grade and the expression of estrogen and progesterone receptors, HER2/neu, p53, and Ki67. RESULTS: Hormone receptors and HER2/neu were positive in 54% and 18% of the tumors, respectively. Both MV antigens were detected in 64% of the tumors. All biopsies containing a DCIS component showed MV in DCIS in addition to invasive BC. In univariate analysis, MV correlated with estrogen receptor (p=0.018), low Ki67 index (<40%, p=0.029), low or intermediate grade (p=0.037), age under 50 years (p=0.039), progesterone receptor (p=0.043) and overexpression of p53 (p=0.049). In multivariate analysis, only grade (p=0.011), p53 (p=0.03) and age (p=0.041) remained associated with MV. CONCLUSION: This study provides evidence for the presence of MV antigens in a relatively large proportion of BC patients. MV was associated with younger age, lower histological grade and overexpression of p53, suggesting that it may play a role in the development of BC.

The prognostic importance of cathepsin D and E-cadherin in early breast cancer: A single-institution experience.

Molecular tools have increasingly been used for decision-making in patients with early breast cancer (EBC). Nevertheless, simple tools such as immunohistochemistry may still be required in particular cases to complement traditional and molecular prognosticators. In this study, the prognostic significance of three well-known immunohistochemical biomarkers, cathepsin D, E-cadherin and Ki67, was studied in 270 patients with EBC, followed by a median time of 126 months in a single institution. Histological examination was performed to confirm the histopathological diagnosis and select specimens. The specimens were evaluated using immunohistochemistry and survival curves were plotted. Results revealed the following patient characteristics: node-negative/1-3 lymph nodes in 228 (86%) patients, hormone receptor-positive in 217 (80%); triple-negative in 31 (11%), and Her2-overexpression in 23 (9%) patients. Breast cancer-related events occurred in 37 patients (14%). A total of 217 patients (80%) survived. Receiver operating characteristic analysis for breast cancer-specific survival showed an area under curve for the clinicopathological model of 0.75 (95% CI, 0.64-0.86), 0.79 (95% CI, 0.68-0.90) for the three-biomarker model, and 0.82 (95% CI, 0.72-0.92) for the E-cadherin and cathepsin D only model. We propose that a simple prognostic model based on combined scores of E-cadherin and cathepsin D may aid treatment decisions in patients with EBC.

p27kip1 Expression in non-small cell lung cancer is not an independent prognostic factor.

BACKGROUND: p27(Kip1) (p27) plays an important role in cancer cell cycle regulation. Recent evidence however, suggests that p27 may function as an oncogene rather than a tumour suppressor gene. PATIENTS AND METHODS: Ninety-two patients with previously untreated non-small cell lung cancer (NSCLC) were studied for the association between the immunohistochemical localization of p27 through a semi-quantitative method and time-to-progression (TTP) and overall survival (OS). RESULTS: The relationship between p27 H-Score and both TTP and OS was polynomial. Short TTP in patients with metastasis or whose tumors progressed during the eight-month period after diagnosis was statistically associated with overexpression of PIRH2 (p<0.001). In patients whose tumours progressed later, long TTP was associated with NSCLC of the non-adenocarcinoma type (p=0.027), p27 H-Score (p=0.032) and well-differentiated adenocarcinoma (p=0.047). None of the parameters correlated with duration of OS. CONCLUSIONS: This study showed that p27 H-Score may not appear to be an independent prognostic factor in patients with NSCLC.