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- Backman, Max, 1987-
(author)
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Spatial immune analyses in clinical cancer tissue
- 2022
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Doctoral thesis (other academic/artistic)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. |
- Ericson Lindquist, Kajsa, et al.
(author)
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Difficulties in diagnostics of lung tumours in biopsies : an interpathologist concordance study evaluating the international diagnostic guidelines
- 2022
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In: Journal of Clinical Pathology. - : BMJ Publishing Group Ltd. - 0021-9746 .- 1472-4146. ; 75:5, s. 302-309
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Journal article (peer-reviewed)abstract
- AIMS: Accurate and reliable diagnosis is essential for lung cancer treatment. The study aim was to investigate interpathologist diagnostic concordance for pulmonary tumours according to WHO diagnostic criteria.METHODS: Fifty-two unselected lung and bronchial biopsies were diagnosed by a thoracic pathologist based on a broad spectrum of immunohistochemical (IHC) stainings, molecular data and clinical/radiological information. Slides stained with H&E, thyroid transcription factor-1 (TTF-1) clone SPT24 and p40 were scanned and provided digitally to 20 pathologists unaware of reference diagnoses. The pathologists independently diagnosed the cases and stated if further diagnostic markers were deemed necessary.RESULTS: In 31 (60%) of the cases, ≥80% of the pathologists agreed with each other and with the reference diagnosis. Lower agreement was seen in non-small cell neuroendocrine tumours and in squamous cell carcinoma with diffuse TTF-1 positivity. Agreement with the reference diagnosis ranged from 26 to 45 (50%-87%) for the individual pathologists. The pathologists requested additional IHC staining in 15-44 (29%-85%) of the 52 cases. In nearly half (17 of 36) of the malignant cases, one or more pathologist advocated for a different final diagnosis than the reference without need of additional IHC markers, potentially leading to different clinical treatment.CONCLUSIONS: Interpathologist diagnostic agreement is moderate for small unselected bronchial and lung biopsies based on a minimal panel of markers. Neuroendocrine morphology is sometimes missed and TTF-1 clone SPT24 should be interpreted with caution. Our results suggest an intensified education need for thoracic pathologists and a more generous use of diagnostic IHC markers.
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| 3. |
- Thurfjell, Viktoria, et al.
(author)
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Comparison of ROS1-rearrangement detection methods in a cohort of surgically resected non-small cell lung carcinomas
- 2022
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In: Translational Lung Cancer Research (TLCR). - : AME Publishing. - 2218-6751 .- 2226-4477. ; 11:12, s. 2477-2494
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Journal article (peer-reviewed)abstract
- Background: Patients with non-small cell lung cancer (NSCLC) harboring a ROS proto-oncogene 1 (ROS1)-rearrangement respond to treatment with ROS1 inhibitors. To distinguish these rare cases, screening with immunohistochemistry (IHC) for ROS1 protein expression has been suggested. However, the reliability of such an assay and the comparability of the antibody clones has been debated. Therefore we evaluated the diagnostic performance of current detection strategies for ROS1-rearrangement in two NSCLC-patient cohorts.Methods: Resected tissue samples, retrospectively collected from consecutive NSCLC-patients surgically treated at Uppsala University Hospital were incorporated into tissue microarrays [all n=676, adenocarcinomas (AC) n=40 1, squamous cell carcinomas (SCC) n=2 13, other NSCLC n=62]. ROS1rearrangements were detected using fluorescence in situ hybridization (FISH) (Abbott Molecular; ZytoVision). In parallel, ROS1 protein expression was detected using IHC with three antibody clones (D4D6, SP384, EPMGHR2) and accuracy, sensitivity, and specificity were determined. Gene expression microarray data (Affymetrix) and RNA-sequencing data were available for a subset of patients. NanoString analyses were performed for samples with positive or ambiguous results (n=21).Results: Using FISH, 2/630 (0.3% all NSCLC; 0.5% non-squamous NSCLC) cases were positive for ROS1 fusion. Additionally, nine cases demonstrated ambiguous FISH results. Using IHC, ROS1 protein expression was detected in 24/665 (3.6% all NSCLC; 5.1% non-squamous NSCLC) cases with clone D4D6, in 18/639 (2.8% all NSCLC; 3.9% non-squamous NSCLC) cases with clone SP384, and in 1/593 (0.2% all NSCLC; 0.3% non-squamous NSCLC) case with clone EPMGHR2. Elevated RNA-levels were seen in 19/369 (5.1%) cases (Affymetrix and RNA-sequencing combined). The overlap of positive results between the assays was poor. Only one of the FISH-positive cases was positive with all antibodies and demonstrated high RNA-expression. This rearrangement was confirmed in the NanoString-assay and also in the RNA sequencing data. Other cases with high protein/RNA-expression or ambiguous FISH were negative in the NanoString-assay.Conclusions: The occurrence of ROS1 fusions is low in our cohorts. The IHC assays detected the fusions, but the accuracy varied depending on the clone. The presumably false-positive and uncertain FISH results questions this method for detection of ROS1-rearrangements. Thus, when IHC is used for screening, transcript-based assays are preferable for validation in clinical diagnostics.
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