Activity and Immune Correlates of Programmed Death-1 Blockade Therapy in Patients With Advanced Large Cell Neuroendocrine Carcinoma.

BACKGROUND: The efficacy of anti-programmed death receptor 1 (PD-1) therapy in patients with large cell neuroendocrine carcinoma (LCNEC) remains unclear. We investigated the outcome of anti-PD-1 therapy and its predictive markers by evaluating the immune-related tumor microenvironment. PATIENTS: We retrospectively reviewed patients with advanced LCNEC treated with systemic chemotherapy. We also evaluated PD ligand 1 (PD-L1) expression (clone: 22C3), CD8-positive tumor-infiltrating lymphocytes (TILs), and the mutational profiles. RESULTS: Seventy patients were enrolled, and 13 of 70 patients received anti-PD-1 therapy. The progression-free survival (PFS) and objective response rate (ORR) of the anti-PD-1 therapy were 4.2 months and 39%, respectively. The overall survival of patients treated with anti-PD-1 therapy (n = 13) was significantly better than those treated without anti-PD-1 therapy (n = 57) (25.2 months vs 10.9 months; P = .02). Among the 13 patients treated with anti-PD-1 therapy, 10 patients (90%) had PD-L1-negative tumors. Patients with a high density of tumoral CD8-positive TILs (≥38/mm2) had a significantly better ORR and PFS than those with a low density of tumoral CD8-positive TILs (ORR: P = .02; PFS: P = .003). Additionally, all 3 patients with TP53 mutation co-occurring with PIK3CA mutation (2 of 8 patients) or RB1 mutation (1 of 8 patients) responded to anti-PD-1 therapy. CONCLUSIONS: Anti-PD-1 therapy was effective regardless of PD-L1 positivity in patients with advanced LCNEC. Our investigation might suggest that the density of tumoral CD8-positive TILs and the presence of co-occurring mutations are predictors of the efficacy of anti-PD-1 therapy in patients with advanced LCNEC.

Real-world survival outcomes with immune checkpoint inhibitors in large-cell neuroendocrine tumors of lung.

BACKGROUND: Little is known regarding the efficacy of immune checkpoint inhibitors (ICI) in patients with advanced large-cell neuroendocrine lung carcinoma (aLCNEC). METHODS: 125 consecutive patients with aLCNEC were identified in the electronic databases of 4 participating cancer centers. The patients were divided into group A (patients who received ICI, n=41) and group B (patients who did not receive ICI, n=84). Overall survival since advanced disease diagnosis (OS DX) and OS since ICI initiation (OS ICI) were captured. RESULTS: With a median follow-up of 11.8 months (mo) (IQR 7.5-17.9) and 6.0mo (IQR 3.1-10.9), 66% and 76% of patients died in groups A and B, respectively. Median OS DX was 12.4mo (95% CI 10.7 to 23.4) and 6.0mo (95% CI 4.7 to 9.4) in groups A and B, respectively (log-rank test, p=0.02). For ICI administration, HR for OS DX was 0.59 (95% CI 0.38 to 0.93, p=0.02-unadjusted), and 0.58 (95% CI 0.34 to 0.98, p=0.04-adjusted for age, Eastern Cooperative Oncology Group (ECOG) performance status (PS), presence of liver metastases and chemotherapy administration). In a propensity score matching analysis (n=74; 37 patients in each group matched for age and ECOG PS), median OS DX was 12.5 mo (95% CI 10.6 to 25.2) and 8.4 mo (95% CI 5.4 to 16.9) in matched groups A and B, respectively (log-rank test, p=0.046). OS ICI for patients receiving ICI as monotherapy (n=36) was 11.0 mo (95% CI 6.1 to 19.4). CONCLUSIONS: With the limitations of retrospective design and small sample size, the results of this real-world cohort analysis suggest a positive impact of ICI on OS in aLCNEC.

Prognostic impact of peripheral blood neutrophil to lymphocyte ratio in advanced-stage pulmonary large cell neuroendocrine carcinoma and its association with the immune-related tumour microenvironment.

BACKGROUND: The prognostic value of the neutrophil-to-lymphocyte ratio (NLR) with large cell neuroendocrine carcinoma (LCNEC) patients remains unclear. Thus, we performed a retrospective study to examine the relationship between the pretreatment NLR and clinical outcome in advanced LCNEC patients and the impact of the immune-related tumour microenvironment (TME). METHODS: This retrospective study included 63 advanced LCNEC patients who had received chemotherapy. We collected clinical data and investigated the TME status (CD4, CD8, CD20 and FOXP3). RESULTS: The overall survival of the patients with a low NLR (<5) was significantly longer than those with a high NLR (≥5) (14.9 vs. 5.2 months; p < 0.001). A multivariate analysis identified a high NLR as a predictor of a poor prognosis (HR, 3.43; 95% CI, 1.73-6.79; p < 0.001). The NLR was inversely correlated with tumoural and stromal CD8-positive tumour-infiltrating lymphocytes (tumoural: r = -0.648, p = 0.005, stromal: r = -0.490, p = 0.046). CONCLUSIONS: A high NLR was associated with a poor prognosis in advanced LCNEC patients. Our study revealed that the NLR can reflect the TME, at least in part, suggesting that the NLR plays an important role not only as a clinical outcome predictor but also as a tumour immune status indicator.

Phase II Study of Immunotherapy With Tecemotide and Bevacizumab After Chemoradiation in Patients With Unresectable Stage III Non-Squamous Non-Small-Cell Lung Cancer (NS-NSCLC): A Trial of the ECOG-ACRIN Cancer Research Group (E6508).

INTRODUCTION: Although chemoradiotherapy (CRT) is the standard of care for patients with unresectable stage III non-small-cell lung cancer (LA-NSCLC), most patients relapse. Tecemotide is a MUC1 antigen-specific cancer immunotherapy vaccine. Bevacizumab improves survival in advanced nonsquamous (NS)-NSCLC and has a role in immune modulation. This phase II trial tested the combination of tecemotide and bevacizumab following CRT in patients with LA-NSCLC. PATIENTS AND METHODS: Subjects with stage III NS-NSCLC suitable for CRT received carboplatin/paclitaxel weekly + 66 Gy followed by 2 cycles of consolidation carboplatin/paclitaxel ≤ 4 weeks of completion of CRT (Step 1). Patients with partial response/stable disease after consolidation therapy were registered onto step 2, which was 6 weekly tecemotide injections followed by every 6 weekly injections and bevacizumab every 3 weeks for up to 34 doses. The primary endpoint was to determine the safety of this regimen. RESULTS: Seventy patients were enrolled; 68 patients (median age, 63 years; 56% male; 57% stage IIIA) initiated therapy, but only 39 patients completed CRT and consolidation therapy per protocol, primarily owing to disease progression or toxicity. Thirty-three patients (median age, 61 years; 58% male; 61% stage IIIA) were registered to step 2 (tecemotide + bevacizumab). The median number of step 2 cycles received was 11 (range, 2-25). Step 2 worst toxicity included grade 3, N = 9; grade 4, N = 1; and grade 5, N = 1. Grade 5 toxicity in step 2 was esophageal perforation attributed to bevacizumab. Among the treated and eligible patients (n = 32) who were treated on step 2, the median overall survival was 42.7 months (95% confidence interval, 21.7-63.3 months), and the median progression-free survival was 14.9 months (95% confidence interval, 11.0-20.9 months) from step 1 registration. CONCLUSIONS: This cooperative group trial met its endpoint, demonstrating tolerability of bevacizumab + tecemotide after CRT and consolidation. In this selected group of patients, the median progression-free survival and overall survival are encouraging. Given that consolidation immunotherapy is now a standard of care following CRT in patients with LA-NSCLC, these results support a role for continued investigation of antiangiogenic and immunotherapy combinations in LA-NSCLC.

Efficacy of immune check-point inhibitors (ICPi) in large cell neuroendocrine tumors of lung (LCNEC).

OBJECTIVES: Little is known regarding the ICPi efficacy in LCNEC. We explored the efficacy and safety of ICPi in LCNEC and assessed its impact on OS. MATERIALS AND METHODS: Thirty-seven consecutive patients with advanced LCNEC were selected from the Davidoff Cancer Center database. These were divided into groups A1 (patients treated with ICPi, n-23) and A2 (patients not treated with ICPi, n-14). Additionally, group A1* was introduced (patients treated with ICPi as a monotherapy, n-21). Another cohort of advanced non-LCNEC lung cancer patients treated with nivolumab at five Israeli cancer centers was chosen as a comparator (group B, n-270). ORR, PFS with ICPi in group A1* were assessed (RECIST 1.1), OS with ICPi was compared between groups A1* and B. OS since advanced disease diagnosis (OSDx) was compared between groups A1 and A2. RESULTS: In group A1*, ORR and median PFS with ICPi were 33 %, and 4.2 months (95 % CI, 2.4-8.1), respectively. With median follow-up since start of ICPi of 6.2 months [IQR 2.2-12.1] and 4.9 months [IQR 2.3-8.9] in groups A1* and B, respectively, 52 % and 64 % of patients died in groups A1* and B, respectively. Median OS with ICPi comprised 11.8 months (95 % CI, 3.7-NR) and 6.9 months (95 % CI, 5.5-8.1) in groups A1* and B, respectively (p-0.23). Median OSDx was 14.5 months (95 % CI, 10.1-38.9) and 10.3 months (95 % CI, 2.6-NR), in groups A1 and A2, respectively (p-0.54). CONCLUSION: In advanced LCNEC, ICPi outcomes are comparable to the outcomes observed in advanced NSCLC. Future research is needed to clarify the impact of ICPi on OS, and to correlate its benefit with tumor mutational landscape.

Case Report: Therapeutic Response to Chemo-Immunotherapy in an Advanced Large Cell Lung Carcinoma Patient With Low Values of Multiple Predictive Biomarkers.

Immune checkpoint inhibitors have revolutionized the treatments of lung cancers, and multiple predictive biomarkers alone or in combination help clinicians with the appropriate therapeutic selections. Recently, chemo-immunotherapy has been recommended for treating advanced non-small cell lung cancers in patients without driver mutations. However, the clinical relevance of predictive biomarkers and the treatment efficacy of chemo-immunotherapy in large cell lung carcinoma (LCLC) remain unclear. Here, we reported a rare case of LCLC with none driver gene mutations and low values of multiple predictive biomarkers. These biomarkers included a low PD-L1 expression of 5-10%, a low tumor mutational burden (TMB) of 2.5 muts/mb, a low CD8(+) tumor-infiltrating lymphocyte density of 147.91 psc/mm². After one-cycle chemotherapy, the patient progressed rapidly and then was switched to pembrolizumab combining paclitaxel plus cisplatin. Interestingly, he achieved a partial response after two cycles of chemo-immunotherapy, showing multiple lymph nodes obviously shrunk on CT scan, and other clinical symptoms were relieved when compared with the baseline findings. After five cycles of chemo-immunotherapy, this advanced patient still benefited and was changed to maintenance immunotherapy monotherapy. This case suggests that chemo-immunotherapy may provide an effective therapeutic option for those LCLC patients with low values of multiple predictive biomarkers, particularly for those who progressed from first-line classical treatments.

Predictive impact of antibiotics in patients with advanced non small-cell lung cancer receiving immune checkpoint inhibitors : Antibiotics immune checkpoint inhibitors in advanced NSCLC.

PURPOSE: In this study, we test the hypothesis that the use of ATB reduces the efficacy of PD(L)1-targeting mAb. METHODS: We included patients with locally advanced, inoperable or metastatic, EGFR wildtype and ALK-negative non-small-cell lung cancer (NSCLC) who received a PD(L)1 targeting mAb (immune checkpoint inhibitor, ICI) between January 2013 and December 2017. The primary study objective was to assess the predictive impact of ATB use within 2 months prior to starting ICI treatment on overall survival from the time of starting ICI treatment (OS-ICI). RESULTS: 33 out of 218 evaluable patients (15.1%) received ATB within 2 months prior to starting ICI treatment. The use of ATB prior to starting ICI was associated with a lower rate of radiological response (18.2 vs. 28.3%, respectively, P = 0.02). PFS was significantly shorter in patients receiving ATB within 2 months prior to ICI compared to those not receiving ATB (median PFS 1.4 vs. 5.5 months, HR = 2.22, P < 0.01). OS-ICI was significantly shorter in NSCLC patients receiving ATB within 2 months prior to ICI compared to those not receiving ATB (median OS-ICI 1.8 vs. 15.4 months, HR = 2.61, P < 0.01; adjusted HR = 3.73, P < 0.01). CONCLUSION: The results of this study suggest that ATB may have a deleterious effect in patients with advanced NSCLC receiving ICI treatment, and more research seems to be justified to explore potential mechanisms.

Large cell neuroendocrine lung carcinoma induces peripheral T-cell repertoire alterations with predictive and prognostic significance.

OBJECTIVES: This study was performed to evaluate for a potentially important role of T cells in the pathophysiology and treatment sensitivity of large cell neuroendocrine lung carcinoma (LCNEC), an orphan disease with poor prognosis and scarce data to guide novel therapeutic strategies. MATERIALS AND METHODS: We performed T-cell receptor (TCR) ß-chain spectratyping on blood samples of patients treated within the CRAD001KDE37 trial (n = 35) using age-matched current or former (n = 11) and never smokers (n = 10) as controls. The data were analyzed in conjunction with the complete blood counts of the probands as well as the data about response to treatment and overall survival in the clinical trial. RESULTS AND CONCLUSION: Untreated stage IV LCNEC patients had significant T-cell repertoire alterations (p < 0.001) compared to age-matched smokers. These changes correlated positively with blood lymphocyte counts (r = 0.49, p < 0.01), suggesting antigen-induced T-cell proliferation as the causative mechanism. At the same time, LCNEC patients showed mild lymphopenia (1.54 vs. 2.51/nl in median, p < 0.01), which reveals a second, antigen-independent mechanism of systemic immune dysregulation. More pronounced T-cell repertoire alterations and higher blood lymphocyte counts at diagnosis were associated with a better treatment response by RECIST and with a longer overall survival (441 vs. 157 days in median, p = 0.019). A higher degree of T-cell repertoire normalization after 3 months of therapy also distinguished a patient group with more favourable prognosis (median overall survival 617 vs. 316 days, p = 0.036) independent of radiological response. Thus, LCNEC induces clinically relevant changes of the T-cell repertoire, which are measurable in the blood and could be exploited for prognostic, predictive and therapeutic purposes. Their pathogenesis appears to involve antigen-induced oligoclonal T-cell expansions superimposed on TCR-independent lymphopenia.

Phase I clinical trial of a novel autologous modified-DC vaccine in patients with resected NSCLC.

BACKGROUND: The primary aim of this study was to evaluate the safety of a novel dendritic cell (DC) vaccine pulsed with survivin and MUC1, silenced with suppressor of cytokine signaling 1 (SOCS1), and immune stimulated with flagellin for patients with stage I to IIIA non-small cell lung cancer (NSCLC) in a phase I open-label, uncontrolled, and dose-escalation trial. Moreover, we evaluate the potential efficacy of this modified DC vaccine as secondary aim. METHODS: The patients were treated with the vaccine at 1 × 106, 1 × 107and the maximum dose 8 × 107 at day 7, 14, and 21 after characterization of the vaccine phenotype by flow cytometry. The safety of the vaccine was assessed by adverse events, and the efficacy by the levels of several specific tumor markers and the patient quality of life. RESULTS: The vaccine was well tolerated without dose-limiting toxicity even at higher doses. The most common adverse event reported was just grade 1 flu-like symptoms without unanticipated or serious adverse event. A significant decrease in CD3 + CD4 + CD25 + Foxp3+ T regulatory (Treg) cell number and increase in TNF-α and IL-6 were observed in two patients. Two patients showed 15% and 64% decrease in carcino-embryonic antigen and CYFRA21, respectively. The vaccination with the maximum dose significantly improved the patients'quality of life when administered at the highest dose. More importantly, in the long-term follow-up until February 17, 2017, 1 patient had no recurrence, 1 patients had a progressive disease (PD), and 1 patient was died in the low dose group. In the middle dose group, all 3 patients had no recurrence. In the high dose group, 1 patient was died, 1 patient had a PD, and the other 7 patients had no recurrence. CONCLUSIONS: We provide preliminary data on the safety and efficacy profile of a novel vaccine against non-small cell lung cancer, which was reasonably well tolerated, induced modest antitumor activity without dose-limiting toxicity, and improved patients' quality of life. Further more, the vaccine maybe a very efficacious treatment for patients with resected NSCLC to prevent recurrence. Our findings on the safety and efficacy of the vaccine in this phase I trial warrant future phase II/III clinical trial.

Rovalpituzumab tesirine, a DLL3-targeted antibody-drug conjugate, in recurrent small-cell lung cancer: a first-in-human, first-in-class, open-label, phase 1 study.

BACKGROUND: Rovalpituzumab tesirine is a first-in-class antibody-drug conjugate directed against delta-like protein 3 (DLL3), a novel target identified in tumour-initiating cells and expressed in more than 80% of patients with small-cell lung cancer. We aimed to assess the safety and activity of rovalpituzumab tesirine in patients who progressed after one or more previous regimen. METHODS: We conducted a phase 1 open-label study at ten cancer centres in the USA. Eligible patients were aged 18 years or older and had histologically or cytologically confirmed small-cell lung cancer or large-cell neuroendocrine tumours with progressive measurable disease (according to Response Evaluation Criteria in Solid Tumors [RECIST], version 1.1) previously treated with one or two chemotherapeutic regimens, including a platinum-based regimen. We assigned patients to dose-escalation or expansion cohorts, ranging from 0·05 mg/kg to 0·8 mg/kg rovalpituzumab tesirine intravenously every 3 weeks or every 6 weeks, followed by investigation of the dose schedules 0·3 mg/kg and 0·4 mg/kg every 6 weeks and 0·2 mg/kg every 3 weeks. Primary objectives were to assess the safety of rovalpituzumab tesirine, including the maximum tolerated dose and dose-limiting toxic effects. The primary activity endpoint was objective response by intention-to-treat analysis. This study is registered with ClinicalTrials.gov, number NCT01901653. The study is closed to enrolment; this report focuses on the cohort with small-cell lung cancer. FINDINGS: Between July 22, 2013, and Aug 10, 2015, 82 patients were enrolled, including 74 patients with small-cell lung cancer and eight with large-cell neuroendocrine carcinoma, all of whom received at least one dose of rovalpituzumab tesirine. Dose-limiting toxic effects of rovalpituzumab tesirine occurred at a dose of 0·8 mg/kg every 3 weeks, including grade 4 thrombocytopenia (in two of two patients at that dose level) and grade 4 liver function test abnormalities (in one patient). The most frequent grade 3 or worse treatment-related adverse events in 74 patients with small-cell lung cancer were thrombocytopenia (eight [11%]), pleural effusion (six [8%]), and increased lipase (five [7%]). Drug-related serious adverse events occurred in 28 (38%) of 74 patients. The maximum tolerated dose of rovalpituzumab tesirine was 0·4 mg/kg every 3 weeks; the recommended phase 2 dose and schedule is 0·3 mg/kg every 6 weeks. At active doses of rovalpituzumab tesirine (0·2 mg/kg or 0·4 mg/kg every 3 weeks or 0·3 mg/kg or 0·4 mg/kg every 6 weeks), 11 (18%) of 60 assessable patients had a confirmed objective response. 11 (18%) of 60 assessable patients had a confirmed objective response, including ten (38%) of 26 patients confirmed to have high DLL3 expression (expression in 50% or more of tumour cells). INTERPRETATION: Rovalpituzumab tesirine shows encouraging single-agent antitumour activity with a manageable safety profile. Further development of rovalpituzumab tesirine in DLL3-expressing malignant diseases is warranted. FUNDING: Stemcentrx Inc.

HHLA2, a New Immune Checkpoint Member of the B7 Family, Is Widely Expressed in Human Lung Cancer and Associated with EGFR Mutational Status.

PURPOSE: Immunotherapy with antibodies against B7/CD28 family members, including PD-1, PD-L1, and CTLA-4 has shifted the treatment paradigm for non-small cell lung carcinoma (NSCLC) with improved clinical outcome. HHLA2 is a newly discovered member of the family. By regulating T-cell function, HHLA2 could contribute to tumor immune suppression and thus be a novel target for cancer immunotherapy. There is limited information and critical need to characterize its expression profile and clinical significance in NSCLC. EXPERIMENTAL DESIGN: We performed IHC with an HHLA2-specific antibody (clone 566.1) using tissue microarrays constructed from 679 NSCLC tumor tissues, including 392 cases in the discovery set and 287 cases in the validation cohort. We also studied clinicopathologic characteristics of these patients. RESULTS: Overall, HHLA2 was not detected in most of normal lung tissue but expressed in 66% of NSCLC across different subtypes. In particular, EGFR-mutated NSCLC was significantly associated with higher tumor HHLA2 expression in both discovery (EGFR vs. WT: 76% vs. 53%, P = 0.01) and validation cohorts (89% vs. 69%, P = 0.01). In one of the two cohorts, HHLA2 expression was higher in lung adenocarcinoma as compared with squamous and large cell histology, non-Hispanic White versus Hispanics, and tumors with high tumor-infiltrating lymphocyte (TIL) density. In the multivariate analysis, EGFR mutation status and high TIL intensity were independently associated with HHLA2 expression in lung adenocarcinoma. CONCLUSIONS: HHLA2 is widely expressed in NSCLC and is associated with EGFR mutation and high TILs in lung adenocarcinoma. It is potentially a novel target for lung cancer immunotherapy. Clin Cancer Res; 23(3); 825-32. ©2016 AACR.

Serum Protein Markers for the Early Detection of Lung Cancer: A Focus on Autoantibodies.

Lung cancer has the highest mortality rate among cancer patients in the world, in particular because most patients are only diagnosed at an advanced and noncurable stage. Computed tomography (CT) screening on high-risk individuals has shown that early detection could reduce the mortality rate. However, the still high false-positive rate of CT screening may harm healthy individuals because of unnecessary follow-up scans and invasive follow-up procedures. Alternatively, false-negative and indeterminate results may harm patients due to the delayed diagnosis and treatment of lung cancer. Noninvasive biomarkers, complementary to CT screening, could lower the false-positive and false-negative rate of CT screening at baseline and thereby reduce the number of patients that need follow-up and diagnose patients at an earlier stage of lung cancer. Lung cancer tissue generates lung cancer-associated proteins to which the immune system might produce high-affinity autoantibodies. This autoantibody response to tumor-associated antigens starts during early stage lung cancer and may endure over years. Identification of tumor-associated antigens or the corresponding autoantibodies in body fluids as potential noninvasive biomarkers could thus be an effective approach for early detection and monitoring of lung cancer. We provide an overview of differentially expressed protein, antigen, and autoantibody biomarkers that combined with CT imaging might be of clinical use for early detection of lung cancer.

Assessing PDL-1 and PD-1 in Non-Small Cell Lung Cancer: A Novel Immunoscore Approach.

INTRODUCTION: Immune checkpoint inhibitors targeting programmed cell death protein 1 (PD-1) or its ligand, PD-L1, have gained momentum in the treatment of non-small cell lung cancer (NSCLC). However, their prognostic significance remains controversial. The present study evaluated the expression of PD-L1 and PD-1 and their potential role in an Immunoscore, supplementing the TNM classification of NSCLC. MATERIALS AND METHODS: Tissue microarrays constructed from tumor tissue samples from 2 cohorts of a total of 536 patients (University Hospital of North Norway, n = 285; Nordland Hospital, n = 251) with primary resected stage I to IIIA NSCLC. PD-L1 and PD-1 were evaluated by immunohistochemistry in the primary tumor and metastatic lymph node tissue. RESULTS: In univariate analysis, a high density of PD-L1+ immune cells in the stromal compartment (S-PD-L1) and PD-1+ intraepithelial tumor infiltrating lymphocytes (T-PD-1) was associated with favorable disease-specific survival (DSS; S-PD-L1, P = .004; T-PD-1, P = .012), both limited to the squamous cell carcinoma histologic subgroup (S-PD-L1, P = .002; T-PD-1, P = .034). A combined low S-PD-L1 and T-PD-1 was associated with poor survival in all patients (DSS: hazard ratio [HR], 1.81; 95% confidence interval [CI], 1.37-2.40; P < .001) at both centers and for all pathologic stages. In multivariate analysis, S-PD-L1 and T-PD-1 were independent positive prognostic factors, and combined low scores remained an independent prognosticator for poor survival (DSS: HR, 1.72; 95% CI, 1.29-2.28; P < .001; disease-free survival, P = .001; overall survival, P = .005). CONCLUSION: Our study identified S-PD-L1 and T-PD-1 as independent positive prognostic factors for NSCLC patients. Their combination added significant prognostic impact within each pathologic stage and hence are feasible to include in a TNM Immunoscore.

Prognostic value of tumor-infiltrating lymphocytes for patients with completely resected stage IIIA(N2) non-small cell lung cancer.

BACKGROUND: The patient prognosis after complete resection for pathologic stage IIIA(N2) non-small cell lung cancer (NSCLC) remains a significant concern. The clinical relevance of the host immune response to NSCLC has yet to be established. We aimed to investigate the prognostic value of tumor-infiltrating lymphocytes (TILs) in a uniform cohort of patients with completely resected stage IIIA(N2) NSCLC. METHODS: From 2005 to 2012, consecutive patients with pathologic stage IIIA(N2) NSCLC who underwent complete resection at our institution were reviewed. For each case, full-face hematoxylin and eosin-stained sections from surgical specimens were evaluated for the TIL density. A published, recommended TIL scoring scale was followed. The patients were stratified into the TIL- or TIL+ group based on pathologic evaluation. RESULTS: Data from 320 patients were included in the analysis. Based on a median follow-up duration of 30.8 months, a higher density of TILs was associated with an improved postoperative survival time (P = 0.06). Subgroup analyses indicated that this positive effect was the greatest for patients with squamous cell carcinoma (SCC; P = 0.03). Among those with SCC, the TIL+ patients experienced a significantly increased 3-year distant metastasis-free survival (DMFS) compared to the TIL- patients (60.6% versus 42.7%, P = 0.02). Multivariate analyses of the 93 patients with SCC tumors confirmed that TIL+ was an independent prognostic factor for an increased DMFS (HR = 0.39, 95%CI 0.17-0.87, P = 0.02) and a prolonged overall survival (OS; HR = 0.47, 95%CI 0.22-1.00, P = 0.05). CONCLUSIONS: Our data suggest a potential role of TILs in predicting the survival of patients with completely resected stage IIIA(N2) NSCLC. The beneficial effects of TILs were more pronounced in the prediction of the DMFS and the OS in patients with SCC. This parameter should be considered for prospective inclusion in clinical trials.

Tumour islet Foxp3+ T-cell infiltration predicts poor outcome in nonsmall cell lung cancer.

The impact of host immunity on outcome in nonsmall cell lung cancer (NSCLC) is controversial. We examined the relationship between lymphoid infiltration patterns in NSCLC and prognosis.Tumour- and stroma-infiltrating CD3(+), CD8(+) and forkhead box P3 (Foxp3)(+) T-lymphocytes were identified using immunohistochemistry and a novel image analysis algorithm to assess total, cytotoxic and regulatory T-lymphocyte counts, respectively, in 196 NSCLC cases. The median cell count was selected as a cut-point to define patient subgroups and the ratio of the corresponding tumour islet:stroma (TI/S) counts was determined.There was a positive association between overall survival and increased CD8(+) TI/S ratio (hazard ratio (HR) for death 0.44, p<0.001) but an inverse relationship between Foxp3(+) TI/S ratio and overall survival (HR 4.86, p<0.001). Patients with high CD8(+) islet (HR 0.48, p<0.001) and Foxp3(+) stromal (HR 0.23, p<0.001) counts had better survival, whereas high CD3(+) and CD8(+) stromal counts and high Foxp3(+) islet infiltration conferred a worse survival (HR 1.55, 2.19 and 3.14, respectively). By multivariate analysis, a high CD8(+) TI/S ratio conferred an improved survival (HR 0.48, p=0.002) but a high Foxp3(+) TI/S ratio was associated with worse survival (HR 3.91, p<0.001).Microlocalisation of infiltrating T-lymphocytes is a powerful predictor of outcome in resected NSCLC.

SPLUNC1 Is a Significant Marker in Pleural Effusion from Lung Cancer Compared to Tuberculosis.

SPLUNC1 (Short palate, lung and nasal epithelium clone1) protein is an abundant secretory product of epithelia present throughout the conducting airways. Although its function is still not fully known, most studies have focused on its defensive effect in the infection of human airways and its potential to serve as a molecular marker for lung cancer. In this study, we further evaluated the SPLUNC1 expression in patients with lung disease to explore its role in cancer or tuberculosis at the protein level. We generated a panel of antibodies by using protein from a eukaryotic expression system as the immunogen to mice. It was the panel of SPLUNC1 monoclonal antibodies that allowed us to comparatively determine SPLUNC1 protein in lung cancer and tuberculosis infection by detecting sera and pleural effusion other than airway surface. The results showed that the SPLUNC1 level was not significantly changed either from sera of lung cancer or control. There was a significant increase in pleural effusion from lung cancer when compared to tuberculosis. These results indicate that SPLUNC1 may be a useful marker for tracing lung cancer cells, based on its epithelial origin property in pleural effusion.

Primary Central Nervous System (CNS) Lymphoma B Cell Receptors Recognize CNS Proteins.

Primary lymphoma of the CNS (PCNSL) is a diffuse large B cell lymphoma confined to the CNS. To elucidate its peculiar organ tropism, we generated recombinant Abs (recAbs) identical to the BCR of 23 PCNSLs from immunocompetent patients. Although none of the recAbs showed self-reactivity upon testing with common autoantigens, they recognized 1547 proteins present on a large-scale protein microarray, indicating polyreactivity. Interestingly, proteins (GRINL1A, centaurin-α, BAIAP2) recognized by the recAbs are physiologically expressed by CNS neurons. Furthermore, 87% (20/23) of the recAbs, including all Abs derived from IGHV4-34 using PCNSL, recognized galectin-3, which was upregulated on microglia/macrophages, astrocytes, and cerebral endothelial cells upon CNS invasion by PCNSL. Thus, PCNSL Ig may recognize CNS proteins as self-Ags. Their interaction may contribute to BCR signaling with sustained NF-κB activation and, ultimately, may foster tumor cell proliferation and survival. These data may also explain, at least in part, the affinity of PCNSL cells for the CNS.

Receptor for hyaluronic acid-mediated motility (RHAMM, CD168) expression is prognostically important in both nodal negative and nodal positive large cell lung cancer.

OBJECTIVE: Despite advances in therapy, lung cancer is still the leading cause of cancer-related mortality in the world. Further prognostic tools are warranted for risk-adapted therapeutic decisions. We analysed a cohort of primary surgically treated non-small cell lung cancers (NSCLCs) to determine the prognostic role of CD44 and associated molecules (receptor for hyaluronic acid-mediated motility (RHAMM), CD95, osteopontin (OPN), P-glycoprotein (P-gp) and caspase 3 (Casp3)). CD44 is a cell adhesion molecule. While the standard form (CD44s) is ubiquitously expressed, its variant isoforms are claimed to play an important role in invasion and metastasis in various cancers. METHODS: Three-hundred and eighty-three primary surgically resected NSCLC specimens were brought into a standardised tissue microarray platform. Immunohistochemistry for CD44, CD95, RHAMM, OPN, P-gp and Casp3 was performed. The clinical correlation was made with known histopathological, phenotypical and genotypical variables; clinical data were available for a postoperative follow-up period of up to 15 years. RESULTS: RHAMM expression in the subgroup of large cell carcinomas (LCC) was associated with inferior survival (p=0.000223). Median overall survival was 92 versus 18 months for RHAMM-negative and positive patients, respectively. This survival difference remained significant in both nodal negative and positive patients (pN0: p=0.013 and pN≥1: p=0.007, respectively). P-gp expression was associated with inferior survival in adenocarcinomas (ACA; p=0.013) and appeared to be a postsurgical Union International Contre le Cancer (pUICC)- stage and gender-independent prognostic factor, irrespective of adjuvant chemotherapy, in the multivariable analysis; considering nodal status, this survival difference applied to pN0 cancers (p=0.026). CONCLUSIONS: Analysis of RHAMM expression is a valuable predictor of survival in LCC. RHAMM-positive patients may benefit from a targeted therapy even in early nodal negative stages. Expression of P-gp identifies a subset of pN0 ACA patients with poor outcome independent of stage, gender and adjuvant chemotherapy.

Surgery decreases number of cells secreting cytotoxic mediators and increases secretion of interleukin 10 in patients with lung cancer.

AIMS: The phenomenon of immunosuppression induced by surgery is widely described as the adverse impact of surgical interventions on leukocytes' populations and secretion of several cytokines. Best of our knowledge, we present the first report evaluating the effect of surgical treatment on the specific anti-cancer immune response against tumour antigens. METHODS: The study included 30 patients operated on for lung cancer. Specific secretion of IFN-γ, Granzyme B, perforines, IL-4, IL-5, IL-10, IL-17a was assessed by ELISPOT (Enzyme-Linked Immunosorbent Spot Assay). RESULTS: Number of cells secreting IFN-γ, Granzyme B and perforines under the influence of autologous tumour antigens or mitogens was significantly decreased on the first day after surgery. During the postoperative recovery we observed an increase in the number of cells secreting IFN-γ, but on the 7th day it still remained lower than before the operation. On the 28th postoperative day it reached a level which was not significantly higher than before the surgery. On the 1st and 7th postoperative day we discovered a significant increase in IL-10 secretion, in response to autologous tumour antigens. CONCLUSIONS: Our results suggest an immunosuppressive effect of surgery on the specific and nonspecific immune stimulation. This effect is particularly expressed in relation to Th1-type immunological response which is associated with direct elimination of cancer cells. Another unfavourable observation is elevated secretion of immunosuppressive IL-10 in response to cancer antigens. These phenomena are associated with shorter survival of the patients.

Impact of dendritic cell vaccines pulsed with Wilms' tumour-1 peptide antigen on the survival of patients with advanced non-small cell lung cancers.

PURPOSE: Dendritic cell (DC)-based vaccines have been expected to serve as new therapeutic approaches for advanced non-small cell lung cancers (NSCLCs); however, their clinical outcomes have not been fully elucidated. We report a single-centre clinical study analysing factors affecting the survival of patients with advanced NSCLCs who received DC vaccines pulsed with or without Wilms' tumour protein-1 (WT1) peptide. METHODS: Among 62 patients with previously treated inoperable or postoperatively relapsed NSCLCs who met the inclusion criteria, DCs from 47 (76%) patients who showed HLA-A2402/0201/0206 were pulsed with one or more corresponding WT1 peptide antigens. DC vaccines were intradermally injected biweekly. RESULTS: Clinical responses based on response evaluation criteria in solid tumours (RECIST) were found in 31 (50%) patients at 3 months after the first DC vaccine (complete response: 1 (1.6%), partial response: 4 (6.5%), stable disease: 26 (41.9%)). Median survival time was 27 months (82% in 1 year and 54% in 2 years) from initial diagnosis, and that was 12 months (48% in 1 year and 22% in 2 years) from the first DC vaccination. Importantly, multivariate analyses revealed that only two factors, blood haemoglobin and the use of WT1 peptides, significantly affected the overall survival of patients from both initial diagnosis and first vaccination. CONCLUSIONS: This study is the first to suggest that DC vaccines pulsed with WT1 may hold a significant impact to prolong the overall survival of patients with advanced NSCLCs.