Keiju Aokage

Predominant infiltration of macrophages and CD8(+) T Cells in cancer nests is a significant predictor of survival in stage IV nonsmall cell lung cancer.

The purpose of this study was to investigate whether tumor‐infiltrating immune cells in biopsy specimens can be used to predict the clinical outcome of stage IV nonsmall cell lung cancer (NSCLC) patients.

Molecular oncology of lung cancer.

Progress in genetic engineering has made it possible to elucidate the molecular biological abnormalities in lung cancer. Mutations in KRAS and P53 genes, loss of specific alleles, and DNA methylation of the tumor suppressor genes were the major abnormalities investigated between 1980 and the 2000s. In 2004, mutations in the epidermal growth factor receptor (EGFR) gene that cause oncogene addiction were discovered in non-small-cell lung cancers (NSCLCs), especially in adenocarcinomas. Because they are strongly associated with sensitivity to EGFR-tyrosine kinase inhibitors (EGFR-TKIs), a great deal of knowledge has been acquired in regard to both EGFR and other genes in the EGFR family and their downstream genes. Moreover, in 2007 the existence of the echinoderm microtubule-associated protein-like 4 (EML4)-anaplastic lymphoma kinase (ALK) fusion gene was discovered in NSCLC; and the same as EGFR-TKIs, ALK inhibitors are being found to be highly effective in lung cancers that have this translocation. These discoveries graphically illustrate that molecular biological findings are directly linked to the development of clinical oncology and to improving the survival rates of lung cancer patients. Here, we review the remarkable progress in molecular biological knowledge acquired thus far in regard to lung cancer, especially NSCLC, and the future possibilities.

Long-Term Outcomes of 50 Cases of Limited-Resection Trial for Pulmonary Ground-Glass Opacity Nodules

Introduction: From 1998 to 2002, we performed a trial of prospective limited resection for pulmonary ground-glass opacity lesions 2 cm or smaller. This is the second report on the long-term outcome. Methods: The enrollment criteria of the trial were as follows: pulmonary peripheral nodule less than 2 cm, diagnosis or suspected diagnosis of clinical T1N0M0 carcinoma with ground-glass opacity and lack of evident pleural indentations or vascular convergence on high-resolution computed tomography. Limited-resection (wedge or segment) specimens were intraoperatively examined by frozen section. If the nodule was confirmed as Noguchi type A or B with a resection margin of greater than 1 cm, the incision was sutured and the patient followed up. The median surveillance period was 10 years. Results: In a total of 50 enrolled participants, there were two Noguchi type A, 23 type B and 15 type C adenocarcinomas; five atypical adenomatous hyperplasias, four fibroses, and one granuloma. Although there were no patients with recurrence within the first 5 years, in four patients who underwent limited-resection pulmonary adenocarcinoma developed more than 5 years after the initial resection, of either cut-end recurrence or metachronous primary disease. Conclusions: Of 26 patients who underwent limited resection, adenocarcinoma developed in four after more than 5 years. These were possibly cut-end recurrences. We concluded that 5 years is not a sufficient period for follow-up, and that limited resection should still be done only in a trial setting, even for small ground-glass opacity lesions.

Late Recurrence of Non-Small Cell Lung Cancer More Than 5 Years After Complete Resection: Incidence and Clinical Implications in Patient Follow-up

BACKGROUND The purpose of this study was to evaluate the risk of late recurrence in patients who had undergone complete resection for non-small cell lung cancer (NSCLC) and remained recurrence-free for > or = 5 years. METHODS Between 1993 and 2002, 1,358 patients with NSCLC underwent complete primary tumor resection and systematic lymph node dissection. Of these, 819 patients remained recurrence-free for 5 years. Recurrence-free probability was estimated from the benchmark of 5 years after primary tumor resection to the date of first recurrence or last follow-up, using the Kaplan-Meier method. Multivariate Cox regression was used to test the relationship of recurrence-free probability to various clinicopathologic factors. RESULTS Of the 819 patients who were free of recurrence at 5 years, 87 (11%) developed a subsequent recurrence. The recurrence-free probabilities at 3 years and 5 years from the point of 5 years after primary tumor resection were 92% and 87%, respectively. The 5-year recurrence-free probabilities from the point of 5 years after primary tumor resection were 81% for patients with intratumoral vascular invasion (P < .001), and 89%, 84%, and 65% for patients with N0, N1, and N2 cancers, respectively (P < .001). Multivariate Cox analysis demonstrated that intratumoral vascular invasion and nodal involvement significantly influenced recurrence 5 years after complete resection (P = .030, P = .022, respectively). CONCLUSIONS Patients with NSCLC with selected tumor characteristics have a significant risk of late recurrence. Therefore, 5 years might not be a sufficient amount of time to declare that NSCLC has been cured.

Histopathologic Features of the Tumor Budding in Adenocarcinoma of the Lung: Tumor Budding As an Index to Predict the Potential Aggressiveness

Introduction: The term tumor budding has been applied to single cells or small clusters of up to four cells within the stromal tissue at the invasive margin of colorectal cancers. This morphologic feature is increasingly being recognized as an adverse prognostic factor. The purpose of this study was to evaluate the clinicopathologic significance of tumor budding in adenocarcinomas of the lung. Methods: We investigated the relationship between tumor budding and clinicopathologic parameters of adenocarcinomas of the lung and the prognostic significance of tumor budding by reviewing the cases of 201 consecutive patients who had undergone complete resection of adenocarcinoma of the lung measuring 30 mm or less in diameter. We examined immunohistochemical profile of budding cells (BCs) by immunohistochemical staining with 14 antibodies. Results: Tumor budding was observed in 78 (43.1%) of the 181 cases with invasive adenocarcinoma. The presence of tumor budding was significantly associated with lymph node metastasis (p = 0.005), pathologic stage (p < 0.001), vascular invasion (p = 0.003), lymphatic invasion (p = 0.009), and pleural invasion (p = 0.029). Examination of the relation between the presence of tumor budding and the predominant histologic subtype revealed that the predominant papillary subtype was significantly associated with the presence of tumor budding (p = 0.0023), whereas the predominant bronchioloalveolar carcinoma subtype was significantly associated with the absence of tumor budding (p < 0.001). The overall 5-year survival rates of the group with budding and the group without budding was 67.5% and 88.3%, respectively, and difference was significant (p = 0.0057). Compared with cancer cells forming nests, BCs displayed reduced expression of cellular adhesion molecule, E-cadherin, and β-catenin (p < 0.05 and p < 0.05, respectively) and increased expression of laminin5-γ2 (p < 0.05). However, BCs displayed reduced expression of differentiation marker, surfactant protein A (p < 0.05). Multivariate analysis revealed that tumor budding was significant independent prognostic factor of the small-sized adenocarcinoma of the lung. Conclusions: Our data showed that tumor budding in adenocarcinoma of the lung is a distinct morphologic feature that has biologic and prognostic significance.

Dynamic molecular changes associated with epithelial-mesenchymal transition and subsequent mesenchymal-epithelial transition in the early phase of metastatic tumor formation

Metastatic tumor formation via vessel route begins with cancer cell extravasation from vessel lumen, migration into the connective tissue surrounding vessels, and invasion into target organ parenchyma. Epithelial–mesenchymal transition (EMT) and mesenchymal–epithelial transition (MET) have been recognized to play an important role in metastatic process, however, how and where these biological changes take place in the early phase of metastatic tumor development has never been clarified. We morphologically evaluated 34 small intrapulmonary metastases formed after cancer cell extravasation from lymphatics (lymphogenic metastasis) and 40 formed in the absence of extravasation (aerogenous metastasis) in human specimens and found that isolated or small clusters of invasive cancer cells (tumor budding) were frequently observed in lymphogenic metastasis (24/34; 71%), but were never observed within aerogenous metastasis. We immunostained 34 lymphogenic metastases for 13 molecular markers of EMT and MET and scored the immunostaining intensity of cancer cells floating in lymphatic vessels (LVs), migrating into the connective tissue surrounding vessels [bronchovascular bundle (BVB)], and growing in lung parenchyma (LP). Cancer cells within BVBs stained more weakly for E‐cadherin (p < 0.001), β‐catenin (p < 0.001), and Geminin (p < 0.001) and more strongly for MMP‐7 (p = 0.046) and Laminin‐5 γ2 (p = 0.037) than tumor cells in LVs. However, cancer cells in LP exhibited resurgent E‐cadherin (p = 0.011), β‐catenin (p < 0.001), and Geminin (p = 0.037) expression and reduced MMP‐7 (p = 0.038) and Laminin‐5 γ2 (p = 0.001) expression in comparison with cancer cells in BVBs. Our results suggested that in the early phase of metastatic tumor formation cancer cells undergo dynamic phenotypic change associated with EMT and subsequent MET.

Possible Delayed Cut-End Recurrence After Limited Resection for Ground-Glass Opacity Adenocarcinoma, Intraoperatively Diagnosed as Noguchi Type B, in Three Patients

Introduction: In our limited resection trial of pulmonary peripheral ground-glass opacity (GGO) lesions from 1998 to 2002, limited resection of Noguchi type A and B carcinomas seemed to have a positive outcome. However, recently three of the 24 patients, with mixed GGO lesions intraoperatively diagnosed as type B, developed a solid lesion at the cut-end scar. Methods: Medical records and radiology and pathology findings of the three patients were reviewed. We also analyzed epidermal growth factor receptor gene mutations when possible. Results: Radiologically, these three second tumors were clearly cut-end scar area recurrences. However, other pathologic and mutation findings suggest metachronous primary cancers developed in Case 1, cut-end recurrence in Case 2, and needle biopsy implantation in Case 3. It is difficult to definitively conclude whether the second tumors were recurrences or metachronous primaries. Conclusions: These second tumors have convinced us that our initial caution in concluding GGO lesions can be cured by limited resection was very appropriate. The recurrences definitely indicate that continuing follow-up attention for more than 5 years is needed after limited resection even for GGO bronchioloalveolar carcinomas.

Long-term survival and risk factors for recurrence in stage I non-small cell lung cancer patients with tumors up to 3 cm in maximum dimension.

BACKGROUND The purpose of this study was to evaluate patients with stage I non-small cell lung cancer (NSCLC) and tumors up to 3 cm in maximum dimension who underwent surgical resection on the revised TNM classification and to investigate the risk factors for recurrence. METHODS Between 1994 and 2003, 713 consecutive stage I NSCLC patients with tumors up to 3 cm in maximum dimension underwent complete resection. Recurrence-free probability was estimated from the date of the primary tumor resection to the date of the first recurrence or the last follow-up using the Kaplan-Meier method. RESULTS The recurrence-free probability of stage I NSCLC patients with tumors up to 3 cm in maximum dimension was 87% at 5 years. On multivariate analyses, three variables were shown to be independently significant recurrence risk factors: histologic differentiation (hazard ratio, 2.3), intratumoral vessel invasion (hazard ratio, 2.9), and visceral pleural invasion (VPI) (hazard ratio, 1.8). According to subgroup analyses combining these three risk factors, the 5-year recurrence-free probability was 94% for patients with zero or one factor (n = 492) and 71% for patients with two or three factors (n = 221), respectively (P < .001). CONCLUSION In stage I NSCLC patients with tumors up to 3 cm in maximum dimension, we identified three risk factors for recurrence that independently increase their risk of recurrence. In addition to VPI, histologic differentiation and intratumoral vessel invasion should be examined and their data collected for the next revision of the TNM staging system.

Prognostic Impact of CD204-Positive Macrophages in Lung Squamous Cell Carcinoma: Possible Contribution of Cd204-Positive Macrophages to the Tumor-Promoting Microenvironment

Introduction: Tumor-associated macrophages (TAMs) are recruited into cancer-induced stroma and produce a specific microenvironment for cancer progression. CD204 (+) TAMs are reportedly related to tumor progression and clinical outcome in some tumors. The aim of this study was to clarify the correlation between CD204 (+) TAMs and the clinicopathological features of lung squamous cell carcinoma. Methods: We investigated the relationships between the numbers of CD204 (+) TAMs and clinicopathological factors, microvessel density, and the numbers of Foxp3 (+) lymphocytes in 208 consecutively resected cases. We also examined the relationships between the numbers of CD204 (+) TAMs and the expression levels of cytokines involved in the migration and differentiation of CD204 (+) TAMs. Results: A high number of CD204 (+) TAMs in the stroma was significantly correlated with an advanced p-stage, T factor, N factor, and the presence of vascular and pleural invasion. A high number of CD204 (+) TAMs in the stroma was also a significant prognostic factor for all p-stages and p-stage I. Moreover, the numbers of CD204 (+) TAMs were correlated with the microvessel density and the numbers of Foxp3 (+) lymphocytes. A high number of CD204 (+) TAMs was strongly correlated with the tissue expression level of monocyte chemoattractant protein-1. CD204 (+) TAMs were shown to be significant independent prognostic factors in a multivariate analysis. Conclusions: CD204 (+) TAMs were an independent prognostic factor in lung squamous cell carcinoma. CD204 (+) TAMs, along with other tumor-promoting stromal cells such as regulatory T cells and endothelial cells, may create tumor-promoting microenvironments.

Forkhead box P3 regulatory T cells coexisting with cancer associated fibroblasts are correlated with a poor outcome in lung adenocarcinoma

Recently, an association between tumor infiltrating Forkhead box P3 regulatory T cells (Treg) and an unfavorable prognosis has been clinically shown in some cancers, but the mechanism of Treg induction in the tumor microenvironment remains uncertain. The aims of the present study were to examine the relationship between Treg and patient outcome and to investigate whether Treg induction is influenced by the characteristics of cancer‐associated fibroblasts (CAF) in lung adenocarcinoma. The numbers of Treg in both the tumor stroma and the tumor nest were counted in 200 consecutive pathological stage I lung invasive adenocarcinoma specimens. To examine whether the characteristics of CAF influence Treg induction, we selected and cultured CAF from low Treg and high Treg adenocarcinoma. The number of Treg was much higher in the stroma than in the nest (P < 0.01). Patients with high Treg had a significantly poorer prognosis than those with low Treg (overall survival: P = 0.03; recurrence‐free survival: P = 0.02; 5‐year overall survival: 85.4% vs 93.0%). Compared with the CAF from low Treg adenocarcinoma, culture supernatant of the CAF from high Treg adenocarcinoma induced more Treg (P = 0.01). Also, CAF from high Treg adenocarcinoma expressed significantly higher mRNA levels of transforming growth factor‐β (P = 0.01) and vascular endothelial growth factor (P = 0.01), both of which are involved in Treg induction. Our studies suggest the possibility that CAF expressing immunoregulatory cytokines may induce Treg in the stroma, creating a tumor‐promoting microenvironment in lung adenocarcinoma that leads to a poor outcome.