7080

NKX2-1

NK2 homeobox 1;NKX2-1;NK2 homeobox 1

NK-2 homolog A|homeobox protein NK-2 homolog A|homeobox protein Nkx-2.1|thyroid nuclear factor 1|thyroid transcription factor 1|thyroid-specific enhancer-binding protein

14q13

protein-coding

Despite the high prevalence and poor outcome of patients with metastatic lung cancer the mechanisms of tumour progression and metastasis remain largely uncharacterized. Here we modelled human lung adenocarcinoma, which frequently harbours activating point mutations in KRAS and inactivation of the p53 pathway, using conditional alleles in mice. Lentiviral-mediated somatic activation of oncogenic Kras and deletion of p53 in the lung epithelial cells of Kras(LSL-G12D/+);p53(flox/flox) mice initiates lung adenocarcinoma development. Although tumours are initiated synchronously by defined genetic alterations, only a subset becomes malignant, indicating that disease progression requires additional alterations. Identification of the lentiviral integration sites allowed us to distinguish metastatic from non-metastatic tumours and determine the gene expression alterations that distinguish these tumour types. Cross-species analysis identified the NK2-related homeobox transcription factor Nkx2-1 (also called Ttf-1 or Titf1) as a candidate suppressor of malignant progression. In this mouse model, Nkx2-1 negativity is pathognomonic of high-grade poorly differentiated tumours. Gain- and loss-of-function experiments in cells derived from metastatic and non-metastatic tumours demonstrated that Nkx2-1 controls tumour differentiation and limits metastatic potential in vivo. Interrogation of Nkx2-1-regulated genes, analysis of tumours at defined developmental stages, and functional complementation experiments indicate that Nkx2-1 constrains tumours in part by repressing the embryonically restricted chromatin regulator Hmga2. Whereas focal amplification of NKX2-1 in a fraction of human lung adenocarcinomas has focused attention on its oncogenic function, our data specifically link Nkx2-1 downregulation to loss of differentiation, enhanced tumour seeding ability and increased metastatic proclivity. Thus, the oncogenic and suppressive functions of Nkx2-1 in the same tumour type substantiate its role as a dual function lineage factor.

The thyroid transcription factor 1 gene (TTF-1 or NKX2-1) is essential to lung development; however, it is also a critical factor in lung cancer. TTF-1 is amplified in lung cancers, suggesting that it is a gain-of-function lung oncogene. Conversely, TTF-1 counters epithelial to mesenchymal transition in cell-based studies and inhibits progression of primary lung adenocarcinomas to metastases in an animal model of lung adenocarcinomas. The unifying theory regarding TTF-1 is that it exhibits both pro-oncogenic and anti-metastatic function depending on the cellular context. Occludin is the first discovered constituent of the epithelial tight junction; in recent years, a functional role of occludin as a tumor suppressor has begun to emerge. Here, we demonstrate that TTF-1 transactivated the expression of the epithelial tight junction molecules occludin (OCLN) and claudin-1 (CLDN1). We show that transcriptional activation occurred through a direct interaction of TTF-1 with the OCLN and CLDN1 promoters. Furthermore, in cells that lack TTF-1, exogenous TTF-1 expression dampened the inhibitory effect of TGF-beta on occludin and claudin-1 content. Using cells derived from a genetically engineered mouse model of lung adenocarcinomas, we observed that silenced TTF-1 expression down-regulated occludin, which we supported with additional siRNA experiments. Finally, TTF-1 knockdown conferred human lung cancer cells resistance to anoikis, and expression of occludin restored cellular sensitivity to anoikis. Overexpression of occludin impeded migration and induced anoikis in lung carcinoma cells. Collectively, these data suggest that TTF-1 transcriptionally regulates occludin, which represents another avenue of TTF-1-mediated metastasis suppression.

BACKGROUND: Thyroid transcription factor-1 (TTF-1) is a homeodomain-containing transcription factor selectively expressed in thyroid, lung and diencephalon. It has been shown to label pulmonary adenocarcinoma, thyroid tumors, and small cell carcinoma (pulmonary and extrapulmonary) with relatively high sensitivity and specificity. The usefulness of this immunostain in cytology specimens has not been thoroughly discussed in the literature. METHODS: The authors evaluated 36 effusion cytology cases (17 pleural effusion, 18 ascitic fluid, and 1 pericardial effusion) diagnosed as metastatic adenocarcinoma and with cell blocks prepared from the file of Pamela Youde Nethersole Eastern Hospital, Hong Kong, during a three-year period from 1998 to early 2001. The clinical, radiologic, cytologic, and histologic (if any) findings were reviewed. A provisional diagnosis of the primary site was deduced for each of the 36 cases by clinical, radiologic, and/or histologic correlation. Immunohistochemical study was performed on the cell block sections of the effusion cytology specimens using mouse monoclonal antibody against TTF-1, after microwave heat-antigen retrieval. The results were correlated with the primary origin of the metastatic adenocarcinoma. RESULTS: Among the 17 cases of metastatic pulmonary adenocarcinoma, 15 cases showed nuclear staining for TTF-1 in most of the tumor cells (sensitivity, 88.2%). None of the 19 cases of metastatic extrapulmonary adenocarcinoma expressed TTF-1 (specificity, 100%). CONCLUSIONS: The current study validates TTF-1 as a highly sensitive and specific immunomarker for distinguishing between metastatic pulmonary and extrapulmonary adenocarcinoma in effusion cytology specimens, which are known to be associated with intrinsic artifact due to less than ideal cellular preservation.

Recent advances in the treatment of pulmonary adenocarcinoma have increased the need for accurate typing of non-small cell carcinomas. Immunohistochemistry for thyroid transcription factor-1 is widely used in the diagnosis of pulmonary adenocarcinomas because it marks approximately 75% of lung adenocarcinomas and is negative in most squamous cell carcinomas and adenocarcinomas of other organs. Napsin A is an aspartic proteinase involved in the maturation of surfactant protein B. It is detected in the cytoplasm of type 2 pneumocytes and alveolar macrophages and is a putative marker for pulmonary adenocarcinomas. We performed immunohistochemistry for napsin A and thyroid transcription factor-1 using tissue microarrays of 95 adenocarcinomas, 48 squamous cell carcinomas, 6 neuroendocrine tumors of the lung, as well as 5 colonic, 31 pancreatic, and 17 breast adenocarcinomas, 38 malignant mesotheliomas, 118 renal cell carcinomas, and 81 thyroid tumors. The tissue microarrays also included 15 different benign tissues. Pulmonary adenocarcinomas were napsin A positive in 79 (83%) of 95 cases compared with 69 (73%) of 95 cases that were thyroid transcription factor-1 positive. There were 13 napsin A-positive/thyroid transcription factor-1-negative and 2 thyroid transcription factor-1-positive/napsin A-negative tumors, increasing the number of cases that were positive with at least one of the markers to 81 (85%) of 95. The limited number of neuroendocrine tumors tested was napsin A negative. All squamous cell carcinomas, adenocarcinomas of the colon, pancreas and breast, and mesotheliomas were negative for both markers. Of the renal tumors, napsin A was positive in most of papillary renal cell carcinomas (79%), about one third (34%) of clear cell renal cell carcinomas, and in a single case of chromophobe renal cell carcinoma (3%). In the thyroid, only 2 cases of papillary thyroid carcinoma (5%), both with tall cell morphology, were positive for napsin A, whereas all other papillary and follicular carcinomas were negative. As expected, all renal tumors were thyroid transcription factor-1 negative, and all thyroid tumors, except for one papillary carcinoma, were thyroid transcription factor-1 positive. Napsin A is a sensitive marker for pulmonary adenocarcinoma and is also expressed in a subset of renal cell carcinomas, particularly of the papillary type, as well as in rare cases of papillary thyroid carcinomas. The combined use of napsin A and thyroid transcription factor-1 results in improved sensitivity and specificity for identifying pulmonary adenocarcinoma in primary lung tumors and in a metastatic setting.

OBJECTIVE: To evaluate the usefulness of thyroid transcription factor-1 (TTF-1) and CDX-2 in determining the primary tumor site of metastatic adenocarcinomas (ACs) in serous effusions. STUDY DESIGN: Cell blocks were constructed from cells in metastatic AC effusion fluids (n = 97) that had been previously stained with a panel of antibodies against MOC-31, D2-40 and calretinin. Primary tumor sites included the lungs (n = 52), ovaries (n = 6), pancreas (n = 4), breasts (n = 3), bile duct (n = 2) and gastrointestinal (GI) tract (30), including stomach (n = 28) and colon (n = 2). Primary sites were determined by tissue confirmation of the original tumor or review of the clinical charts. Immunocytochemical staining was performed with antibodies against TTF-1 and CDX-2. RESULTS: The lung ACs showed TTF-1 positivity in 58% (30/52) of cases. All nonpulmonary ACs lacked TTF-1 staining. Among the 30 GI ACs, 9 (30%) (7 from the stomach and 2 from the colon) showed CDX-2 positivity. All non-GI ACs lacked CDX-2 staining. Specificities and positive predictive values for TTF-1 and CDX-2 equaled 100% for metastatic pulmonary and GI ACs, respectively. CONCLUSION: Our results suggested that TTF-1 and CDX-2 are specific markers to separate metastatic pulmonary and GI ACs, respectively, from other metastatic ACs in serous effusions. However, sensitivity values of these markers were low.

Thyroid transcription factor-1 (TTF-1) is a 38-kd homeodomain containing DNA-binding protein, identified in thyroid and lung as a regulator of thyroid-specific genes and surfactant and Clara cell secretory protein gene expression. TTF-1 has been used as a reliable lineage marker for lung adenocarcinoma and thyroid carcinoma in surgical pathology. However, TTF-1 expression has been recently reported in carcinomas of other origins including female genital tract. We evaluated TTF-1 expression with 3 primary different antibodies (8G7G3/1, SPT24, and BGX-397A) and 2 secondary automated detection systems (Envision+/Dako autostainer versus Refine/Bond Max) in 104 ovarian and endometrial tumors on routine surgical specimens and 108 ovarian tumors on tissue microarray (TMA) specimens. SPT24 and Refine/Bond Max autostainer was the most sensitive system among the primary antibodies and secondary detection/autostainers tested. By using SPT24/Refine/Bond Max, TTF-1 reactivity could be detected in all major histologic subtypes of gynecologic tumors and up to 26% of all cases tested on routine surgical specimens and 6.4% on TMA. TTF-1 was most frequently detected in uterine malignant mixed Mullerian tumor (82%), more common in uterine tumors than ovarian tumors, and more common in surgical specimen than TMA. When present, tumor cells can be rarely positive or diffusely positive for TTF-1 reactivity. In addition to malignant tumors, TTF-1 was also detected in benign tumors and benign tubal and endometrial epithelia. TTF 1 immunostaining has the potential to misguide a pathologist to conclude an ovarian or endometrial tumor being a lung metastasis. However, the role of TTF-1 in female genital tract and its tumors is unknown.