| 1. |
- Backman, Max, 1987-
(författare)
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Spatial immune analyses in clinical cancer tissue
- 2022
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Cancer is a leading cause of premature death and lung cancer is the deadliest cancer type, with non-small cell lung cancer (NSCLC) representing 85% of lung cancer cases. Despite promising development in cancer treatment in recent decades, overall prognosis is poor. The aim of this thesis was to explore novel techniques in protein visualization in clinical cancer tissue to better our understanding of cancer immunity and to discover new biomarkers for improved cancer diagnostics.In Paper I traditional immunohistochemistry (IHC) was compared to the in-situ proximity ligation assay (isPLA). Both techniques were applied to stain 12 proteins in 39 cell lines and 37 tissue types. Two different antibodies were used in the IHC assay and in the isPLA, where binding by both antibodies is required to generate detection signals. The comparison of staining patterns showed that the isPLA presents a valuable alternative to traditional IHC.In Paper II cancer tissue from 357 NSCLC patients was immunophenotyped through IHC annotations of 11 different immune markers. A distinct group of cases with a signature of NK cells and/or plasma cells had favorable prognosis despite significantly lower T-cell activation signatures. This study provides a detailed description of the immune landscape in NSCLC, extending previous concepts, and highlights plasma and NK-cells as potential biomarkers for further validation.In Paper III a multiplex-multispectral pipeline was established to explore three immune marker panels in a NSCLC cohort, spatially quantifying 13 immune cell types. The immune composition of NSCLC was analyzed for the prognostic relevance of immune cell coordination. Cell densities and distances were found to contribute independently to prognosis, indicating that spatial information on local immune cell infiltration is crucial for understanding tumor immunity.In Paper IV an extensive characterization of the immune cell landscape of colon cancer identified a prognostic signature based on the ratio of CD8+ lymphocytes to CD68+CD163+ macrophages. This signature was superior to the state-of-the-art ‘Immunoscore’, and was also associated with longer survival when analyzed in other common cancer types. This presents a promising immunological biomarker that warrants further validation as a prognostic and predictive signature in common cancers.In summary, this thesis presents an in-depth study of immune cell infiltration in several cancer types to better understand cancer immunity. Through novel techniques and spatial metrics, we describe immunophenotypes that might contribute to cancer classification and prognostication. The identified immune phenomena may also present alternative treatment targets to overcome resistance to immunotherapy.
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| 2. |
- Backman, Max, et al.
(författare)
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Infiltration of NK and plasma cells is associated with a distinct immune subset in non‐small cell lung cancer
- 2021
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Ingår i: Journal of Pathology. - : John Wiley & Sons. - 0022-3417 .- 1096-9896. ; 255:3, s. 243-256
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Tidskriftsartikel (refereegranskat)abstract
- Immune cells of the tumor microenvironment are central but erratic targets for immunotherapy. The aim of this study was to characterize novel patterns of immune cell infiltration in non-small cell lung cancer (NSCLC) in relation to its molecular and clinicopathologic characteristics. Lymphocytes (CD3+, CD4+, CD8+, CD20+, FOXP3+, CD45RO+), macrophages (CD163+), plasma cells (CD138+), NK cells (NKp46+), PD1+, and PD-L1+ were annotated on a tissue microarray including 357 NSCLC cases. Somatic mutations were analyzed by targeted sequencing for 82 genes and a tumor mutational load score was estimated. Transcriptomic immune patterns were established in 197 patients based on RNA sequencing data. The immune cell infiltration was variable and showed only poor association with specific mutations. The previously defined immune phenotypic patterns, desert, inflamed, and immune excluded, comprised 30, 13, and 57% of cases, respectively. Notably, mRNA immune activation and high estimated tumor mutational load were unique only for the inflamed pattern. However, in the unsupervised cluster analysis, including all immune cell markers, these conceptual patterns were only weakly reproduced. Instead, four immune classes were identified: (1) high immune cell infiltration, (2) high immune cell infiltration with abundance of CD20+ B cells, (3) low immune cell infiltration, and (4) a phenotype with an imprint of plasma cells and NK cells. This latter class was linked to better survival despite exhibiting low expression of immune response-related genes (e.g. CXCL9, GZMB, INFG, CTLA4). This compartment-specific immune cell analysis in the context of the molecular and clinical background of NSCLC reveals two previously unrecognized immune classes. A refined immune classification, including traits of the humoral and innate immune response, is important to define the immunogenic potency of NSCLC in the era of immunotherapy. © 2021 The Authors. The Journal of Pathology published by John Wiley & Sons, Ltd. on behalf of The Pathological Society of Great Britain and Ireland.
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| 3. |
- Backman, Max, 1987-, et al.
(författare)
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Spatial immunophenotyping of the tumour microenvironment in non-small cell lung cancer
- 2023
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Ingår i: European Journal of Cancer. - : Elsevier. - 0959-8049 .- 1879-0852. ; 185, s. 40-52
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Tidskriftsartikel (refereegranskat)abstract
- Introduction: Immune cells in the tumour microenvironment are associated with prognosis and response to therapy. We aimed to comprehensively characterise the spatial im-mune phenotypes in the mutational and clinicopathological background of non-small cell lung cancer (NSCLC).Methods: We established a multiplexed fluorescence imaging pipeline to spatially quantify 13 immune cell subsets in 359 NSCLC cases: CD4 effector cells (CD4-Eff), CD4 regulatory cells (CD4-Treg), CD8 effector cells (CD8-Eff), CD8 regulatory cells (CD8-Treg), B-cells, natural killer cells, natural killer T-cells, M1 macrophages (M1), CD163 thorn myeloid cells (CD163), M2 macrophages (M2), immature dendritic cells (iDCs), mature dendritic cells (mDCs) and plasmacytoid dendritic cells (pDCs).Results: CD4-Eff cells, CD8-Eff cells and M1 macrophages were the most abundant immune cells invading the tumour cell compartment and indicated a patient group with a favourable prognosis in the cluster analysis. Likewise, single densities of lymphocytic subsets (CD4-Eff, CD4-Treg, CD8-Treg, B-cells and pDCs) were independently associated with longer survival. However, when these immune cells were located close to CD8-Treg cells, the favourable impact was attenuated. In the multivariable Cox regression model, including cell densities and distances, the densities of M1 and CD163 cells and distances between cells (CD8-Treg-B-cells, CD8-Eff-cancer cells and B-cells-CD4-Treg) demonstrated positive prognostic impact, whereas short M2-M1 distances were prognostically unfavourable.Conclusion: We present a unique spatial profile of the in situ immune cell landscape in NSCLC as a publicly available data set. Cell densities and cell distances contribute independently to prognostic information on clinical outcomes, suggesting that spatial information is crucial for diagnostic use.
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| 4. |
- La Fleur, Linnéa
(författare)
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Mutation and immune profiling of non-small cell lung cancer
- 2019
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Several novel therapies that target molecular alterations and immune checkpoints in lung cancer have been introduced in the last decade. Still, only a minority of patients obtain long term disease control and overall survival remains poor. The aim of this thesis was to characterize the landscape of genetic alterations and immune cell infiltrates in tumor tissues from a large representative patient cohort of non-small cell lung cancer (NSCLC).The mutational status of 82 genes related to lung cancer development were evaluated, in paper I, by a targeted re-sequencing approach adapted to work on “real-life” samples of mixed quality. We observed a remarkably high prevalence of activating KRAS mutations. Otherwise, the mutation spectrum resembled other western lung cancer populations. Poor survival was linked to subgroups of lung adenocarcinoma with mutations in TP53, STK11 and SMARCA4, independent of concomitant KRAS mutations. In lung squamous cell carcinoma, patients with mutations in CSMD3 had better survival.The infiltration of tumor-associated immune cells was assessed by immunohistochemical analysis in paper II. Previously described immune response patterns termed “inflamed” and “desert” were confirmed in our dataset. In addition, we discovered a new immune phenotype characterized by overall sparse presence of most immune cell types except for a distinct infiltration of NK and plasma cells. This novel immune class displayed a favorable prognosis and was therefore designated “oasis”.In paper III, infiltration of macrophage subtypes was evaluated by immunohistochemical analysis of CD68, CD163, MSR1 and MARCO. The majority of macrophages exhibited a tumor promoting phenotype and expression of MARCO, a targetable scavenger receptor, was detected in a distinct subset of NSCLC patients. Further investigation of the functional roles of MARCO in a human NSCLC setting was carried out in paper IV. Here, MARCO expression on cultured myeloid cells could be induced by NSCLC cell lines. The MARCO+ cells displayed an immunosuppressive phenotype and could effectively suppress the cytolytic effect of NK cells and CD8+ T cells. A monoclonal antibody targeting MARCO removed these inhibitory effects of the MARCO+ cells.In summary, this thesis contributes knowledge on the genetic and immunologic underpinning of lung cancer that forms the basis for current and future treatment strategies in the evolving era of personalized oncology and pathology.
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| 5. |
- Elfving, Hedvig, et al.
(författare)
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Spatial distribution of tertiary lymphoid structures in the molecular and clinical context of non-small cell lung cancer.
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Annan publikation (övrigt vetenskapligt/konstnärligt)abstract
- Tertiary lymphoid structures (TLS) are lymphocyte aggregates resembling secondary lymphoid organs and are pivotal in cancer immunity. The ambiguous morphological definition of TLS makes it challenging to ascertain their clinical impact on patient survival and response to immunotherapy. This study aimed to characterize TLS in hematoxylin-eosin tissue sections from lung cancer patients, assessing their occurrence in relation to the local immune environment, mutational background, and patient outcome.Two pathologists evaluated one whole tissue section from each resection specimen of 680 NSCLC patients. TLS were spatially quantified within the tumor area or periphery and further categorized based on the presence of germinal centers (mature TLS). Metrics were integrated with immune cell counts, genomic and transcriptomic data, and correlated with clinical parameters.Out of 536 evaluable cases, TLS were present in 86% of tumor samples, predominantly in the tumor periphery, with a median of eight TLS per case. TLS with germinal centers were found in 24% of cases. TLS presence correlated positively with increased plasma cell (CD138+) and lymphocytic cell (CD3+, CD8+, FOXP3+) infiltration. Tumors with higher tumor mutational burden (TMB) exhibited higher periphery TLS numbers. The overall TLS quantity was associated with improved patient survival, irrespective of TLS maturation status. This prognostic association held true for periphery TLS but not for tumor TLS.In conclusion, TLS occurrence in NSCLC is common and its correlation with a specific immune phenotype suggests biological relevance in the local immune reaction. The prognostic significance of this scoring system on routine hematoxylin-eosin sections has the potential to augment diagnostic algorithms for NSCLC patients.
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