The progression and metastatic capacity of solid tumors are strongly influenced by immune cells in the tumor microenvironment. In non–small cell lung cancer (NSCLC), accumulation of anti-inflammatory tumor-associated macrophages (TAM) is associated with worse clinical outcome and resistance to therapy. Here we investigated the immune landscape of NSCLC in the presence of protumoral TAMs expressing the macrophage receptor with collagenous structure (MARCO). MARCO-expressing TAM numbers correlated with increased occurrence of regulatory T cells and effector T cells and decreased natural killer (NK) cells in these tumors. Furthermore, transcriptomic data from the tumors uncovered a correlation between MARCO expression and the anti-inflammatory cytokine IL37. In vitro studies subsequently showed that lung cancer cells polarized macrophages to express MARCO and gain an immune-suppressive phenotype through the release of IL37. MARCO-expressing TAMs blocked cytotoxic T-cell and NK-cell activation, inhibiting their proliferation, cytokine production, and tumor killing capacity. Mechanistically, MARCO+ macrophages enhanced regulatory T (Treg) cell proliferation and IL10 production and diminished CD8 T-cell activities. Targeting MARCO or IL37 receptor (IL37R) by antibody or CRISPR knockout of IL37 in lung cancer cell lines repolarized TAMs, resulting in recovered cytolytic activity and antitumoral capacity of NK cells and T cells and downmodulated Treg cell activities. In summary, our data demonstrate a novel immune therapeutic approach targeting human TAMs immune suppression of NK- and T-cell antitumor activities. Significance: This study defines tumor-derived IL37 and the macrophage scavenger receptor MARCO as potential therapeutic targets to remodel the immune-suppressive microenvironment in patients with lung cancer.

Non-small cell lung cancer (NSCLC) is a heterogeneous disease with unique combinations of somatic molecular alterations in individual patients, as well as significant differences in populations across the world with regard to mutation spectra and mutation frequencies. Here we aim to describe mutational patterns and linked clinical parameters in a population-based NSCLC cohort. Using targeted resequencing the mutational status of 82 genes was evaluated in a consecutive Swedish surgical NSCLC cohort, consisting of 352 patient samples from either fresh frozen or formalin fixed paraffin embedded (FFPE) tissues. The panel covers all exons of the 82 genes and utilizes reduced target fragment length and two-strand capture making it compatible with degraded FFPE samples. We obtained a uniform sequencing coverage and mutation load across the fresh frozen and FFPE samples by adaption of sequencing depth and bioinformatic pipeline, thereby avoiding a technical bias between these two sample types. At large, the mutation frequencies resembled the frequencies seen in other western populations, except for a high frequency of KRAS hotspot mutations (43%) in adenocarcinoma patients. Worse overall survival was observed for adenocarcinoma patients with a mutation in either TP53, STK11 or SMARCA4. In the adenocarcinoma KRAS-mutated group poor survival appeared to be linked to concomitant TP53 or STK11 mutations, and not to KRAS mutation as a single aberration. Similar results were seen in the analysis of publicly available data from the cBioPortal. In squamous cell carcinoma a worse prognosis could be observed for patients with MLL2 mutations, while CSMD3 mutations were linked to a better prognosis. Here we have evaluated the mutational status of a NSCLC cohort. We could not confirm any survival impact of isolated driver mutations. Instead, concurrent mutations in TP53 and STK11 were shown to confer poor survival in the KRAS-positive adenocarcinoma subgroup.

Cancer testis antigens (CTAs) are of clinical interest as biomarkers and present valuable targets for immunotherapy. To comprehensively characterize the CTA landscape of non–small-cell lung cancer (NSCLC), we compared RNAseq data from 199 NSCLC tissues to the normal transcriptome of 142 samples from 32 different normal organs. Of 232 CTAs currently annotated in the Caner Testis Database (CTdatabase), 96 were confirmed in NSCLC. To obtain an unbiased CTA profile of NSCLC, we applied stringent criteria on our RNAseq data set and defined 90 genes as CTAs, of which 55 genes were not annotated in the CTdatabase, thus representing potential new CTAs. Cluster analysis revealed that CTA expression is histology dependent and concurrent expression is common. IHC confirmed tissue-specific protein expression of selected new CTAs (TKTL1, TGIF2LX, VCX, and CXORF67). Furthermore, methylation was identified as a regulatory mechanism of CTA expression based on independent data from The Cancer Genome Atlas. The proposed prognostic impact of CTAs in lung cancer was not confirmed, neither in our RNAseq cohort nor in an independent meta-analysis of 1,117 NSCLC cases. In summary, we defined a set of 90 reliable CTAs, including information on protein expression, methylation, and survival association. The detailed RNAseq catalog can guide biomarker studies and efforts to identify targets for immunotherapeutic strategies.

The critical role of the immune system in controlling cancer progression has become evident and immune modulatory therapy is now approved for clinical use. However, while the majority of studies on the inflammatory tumour microenvironment have focused on the cellular immune response, in particular the prognostic and predictive role of various T cell infiltrates, the role of the humoral immune response in this context has long been overlooked. This study aimed to investigate the clinicopathological correlates and prognostic impact of B cell and plasma cell infiltration in epithelial ovarian cancer (EOC). Immunohistochemical expression of immunoglobulin kappa C (IGKC), CD20 and CD138 was analysed in tissue microarrays with tumours from 154 incident cases of EOC from two pooled prospective population-based cohorts. Subsets of corresponding benign-appearing fallopian tubes (n = 38) and omental metastases (n = 33) were also analysed. Kaplan-Meier analysis and Cox regression analysis were used to determine the impact of immune-cell specific IGKC, CD20 and CD138 expression on overall survival and ovarian cancer-specific survival. High IGKC expression correlated significantly with expression of CD20 (p = 0.001) and CD138 (p = 0.035). Expression of IGKC as well as CD138 was significantly higher in primary tumours than in fallopian tubes (p = 0.004 and p = 0.001, respectively). High CD20 and CD138 expression correlated significantly with high tumour grade (p = 0.032 and p = 0.030, respectively). CD20 and IGKC expression was not prognostic but univariable Cox regression analysis revealed high CD138 expression to correlate with a significantly reduced overall survival (HR = 2.20; 95 % CI 1.34–3.55; p–0.001) as well as ovarian cancer-specific survival (HR = 1.95; 95 % CI 1.28–2.98; p = 0.002). The prognostic impact was independent of established clinical parameters (age, grade, clinical stage) as shown in multivariable analysis (HR = 2.28; 95 % CI 1.39–3.75; p = 0.001). In conclusion, our results demonstrate that plasma cell infiltration in epithelial ovarian cancer has a significant impact on tumour progression and prognosis. The important role of the humoral immune system merits further study and may be harnessed as immune modulatory strategies in cancer therapy.

We describe a bioinformatic tool, Tumor Aberration Prediction Suite (TAPS), for the identification of allele-specific copy numbers in tumor samples using data from Affymetrix SNP arrays. It includes detailed visualization of genomic segment characteristics and iterative pattern recognition for copy number identification, and does not require patient-matched normal samples. TAPS can be used to identify chromosomal aberrations with high sensitivity even when the proportion of tumor cells is as low as 30%. Analysis of cancer samples indicates that TAPS is well suited to investigate samples with aneuploidy and tumor heterogeneity, which is commonly found in many types of solid tumors.