Multispectral Imaging Simulation

1987 ◽  
Author(s):  
Gene R. Loefer ◽  
Ken Q. Lao
Author(s):  
Sui-Xian Li ◽  
Da-Qi Xu ◽  
Li-Yan Zhang ◽  
Jun Lin

Previous research has shown that the effectiveness of selecting filter set from a large set of commercial broadband filters by vector analyzing method based on maximum linear independence (MLI). However, the traditional MLI is suboptimal due to predefining the first filter of the selected filter set being the maximum ℓ2 norm among all those of the filters. An exhaustive imaging simulation is conducted to investigate the features of the most competent filter set. In the simulation, every filter in a subset of all the filters is selected serving as the first filter in turn. From the results of the simulation, we found there are remarkable characteristics for the most competent filter set. Besides smaller condition number, the geometric features of the best-performed filter set comprise the distinct peak of the transmittance of the first filter, the generally uniform distributing of the peaks of the transmittance curve of the filters, the substantial overlapping of the transmittance curves with those of the adjacent filer sets. Therefore, the best-performed filter sets can be decided intuitively by simple vector analyzing and just a few experimental verifications. This work would be useful for optimizing spectral sensitivity of broadband multispectral imaging sensors or SFA sensors.


2020 ◽  
Author(s):  
T Ritz ◽  
J Baues ◽  
O Krenkel ◽  
P Schirmacher ◽  
T Longerich

2021 ◽  
Vol 9 (7) ◽  
pp. e002844
Author(s):  
Alexander Stein ◽  
Donjete Simnica ◽  
Christoph Schultheiß ◽  
Rebekka Scholz ◽  
Joseph Tintelnot ◽  
...  

BackgroundIn patients with microsatellite stable (MSS) metastatic colorectal cancer (mCRC), immune checkpoint blockade is ineffective, and combinatorial approaches enhancing immunogenicity need exploration.MethodsWe treated 43 patients with predominantly microsatellite stable RAS/BRAF wild-type mCRC on a phase II trial combining chemotherapy with the epidermal growth factor receptor antibody cetuximab and the programmed cell death ligand 1 (PD-L1) antibody avelumab. We performed next-generation gene panel sequencing for mutational typing of tumors and liquid biopsy monitoring as well as digital droplet PCR to confirm individual mutations. Translational analyses included tissue immunohistochemistry, multispectral imaging and repertoire sequencing of tumor-infiltrating T cells. Detected PD-L1 mutations were mechanistically validated in CRISPR/Cas9-generated cell models using qRT-PCR, immunoblotting, flow cytometry, complement-dependent cytotoxicity assay, antibody-dependent cytotoxicity by natural killer cell degranulation assay and LDH release assay as well as live cell imaging of T cell mediated tumor cell killing.ResultsCirculating tumor DNA showed rapid clearance in the majority of patients mirroring a high rate of early tumor shrinkage. In 3 of 13 patients expressing the high-affinity Fcγ receptor 3a (FcγR3a), tumor subclones with PD-L1 mutations were selected that led to loss of tumor PD-L1 by nonsense-mediated RNA decay in PD-L1 K162fs and protein degradation in PD-L1 L88S. As a consequence, avelumab binding and antibody-dependent cytotoxicity were impaired, while T cell killing of these variant clones was increased. Interestingly, PD-L1 mutant subclones showed slow selection dynamics reversing on avelumab withdrawal and patients with such subclones had above-average treatment benefit. This suggested that the PD-L1 mutations mediated resistance to direct antitumor effects of avelumab, while at the same time loss of PD-L1 reduced biological fitness by enhanced T cell killing limiting subclonal expansion.ConclusionThe addition of avelumab to standard treatment appeared feasible and safe. PD-L1 mutations mediate subclonal immune escape to avelumab in some patients with mCRC expressing high-affinity FcγR3a, which may be a subset experiencing most selective pressure. Future trials evaluating the addition of avelumab to standard treatment in MSS mCRC are warranted especially in this patient subpopulation.Trial registration numberNCT03174405.


Author(s):  
Dhananjaya Jayasundara ◽  
Lakshitha Ramanayake ◽  
Neranjan Senarath ◽  
Sanjaya Herath ◽  
Roshan Godaliyadda ◽  
...  

2020 ◽  
Vol 8 (Suppl 3) ◽  
pp. A795-A795
Author(s):  
Hyeonbin Cho ◽  
Jae-Hwan Kim ◽  
Ji-Hyun Kim

BackgroundCancer immunotherapy (CIT) has substantially improved the survival of cancer patients. However, according to recent studies, liver metastasis was reported to predict worse outcomes for CIT. The main objective of the study is to evaluate the differences in the immune microenvironment (IME) between the primary lung cancer (PL) and synchronous liver metastasis (LM) using a multispectral imaging system.MethodsSix immune markers (CD4, CD8, CTLA-4, granzyme B (GZB), Foxp3 and PD-L1) were analyzed using a multiplex IHC system and inForm program (Akoya) on paired lung-liver samples of 10 patients. Cells were categorized into tumor nest and stroma, and cell counts per unit area were measured for comparison.ResultsThe number of tumor-infiltrating cytotoxic T cells (TIL) in PL (262.5 cells/mm2) was higher than that of LM (113.3 cells/mm2). Additionally, the ratio between the number of TIL and non-TIL was greater in PL (0.31) compared to that of LM (0.26). A similar trend appeared for Helper T cells and regulatory T cells (Treg), as PL consisted of higher numbers of T cells (791.8 Helper T cells/mm2, 195.7 Treg/mm2) than LM (626.3 Helper T cells/mm2, 121.3 Treg/mm2). However, cytotoxic T cells exhibiting GZB+ and CTLA-4- were fewer in PL (140.2 cells/mm2) than in LM (203.3 cells/mm2), and the ratio is 0.69. The mean number of GZB+ TIL in PL (32.5 cells/mm2) was lower than in LM (35.3 cells/mm2), and their proportions among total TIL counts were 0.12 and 0.31, respectively. In PL, GZB+: GZB- ratio is 0.16 while the ratio is 1.91 for LM. A fewer number of TILs exhibiting GZB suggests that PL has lower efficiency in immune response than LM. Another crucial checkpoint receptor that inhibits immune response, CTLA-4, was more prevalent in PL, with CTLA-4+: CTLA-4- ratio in Treg being 0.36 in PL, compared to 0.11 in LM. The tumor proportion score (TPS) of PD-L1 was higher in PL than LM (40.0 vs. 6.6).ConclusionsIn our study, we showed the differences in the numbers of TIL or regulatory T cells and expressions of immune checkpoint receptors (PD-L1, CTLA-4), which significantly influence outcomes for CIT. The study is ongoing to confirm different IME between the PL and LM groups in a larger tumor cohort.ReferencesPeng, Jianhong, et al., Immune Cell Infiltration in the Microenvironment of Liver Oligometastasis from Colorectal Cancer: Intratumoural CD8/CD3 Ratio Is a Valuable Prognostic Index for Patients Undergoing Liver Metastasectomy. Cancers 2019 Dec; 11(12): 1922. https://doi.org/10.3390/cancers11121922Tumeh, Paul C., et al., Liver Metastasis and treatment outcome with Anti-PD-1 monoclonal antibody in patients with melanoma and NSCLC. Cancer Immunol Res 2017 May; 5(5): 417–424. doi: 10.1158/2326-6066.CIR-16-0325Parra, E.R., Immune Cell Profiling in Cancer Using Multiplex Immunofluorescence and Digital Analysis Approaches; Streckfus, C.F., Ed.; IntechOpen: London, UK, 2018; pp. 1–13. doi: 10.5772/intechopen.80380Ribas, A., Hu-Lieskovan, S., What does PD-L1 positive or negative mean?. The Journal of Experimental Medicine 2016;213(13):2835–2840. https://doi.org/10.1084/jem.20161462


2020 ◽  
pp. 1-1
Author(s):  
Mohammad Reza Mohebbian ◽  
Md Hanif Ali Sohag ◽  
Seyed Shahim Vedaei ◽  
Khan A. Wahid

2021 ◽  
Vol 9 (7) ◽  
pp. e002197
Author(s):  
Janis M Taube ◽  
Kristin Roman ◽  
Elizabeth L Engle ◽  
Chichung Wang ◽  
Carmen Ballesteros-Merino ◽  
...  

BackgroundEmerging data suggest predictive biomarkers based on the spatial arrangement of cells or coexpression patterns in tissue sections will play an important role in precision immuno-oncology. Multiplexed immunofluorescence (mIF) is ideally suited to such assessments. Standardization and validation of an end-to-end workflow that supports multisite trials and clinical laboratory processes are vital. Six institutions collaborated to: (1) optimize an automated six-plex assay focused on the PD-1/PD-L1 axis, (2) assess intersite and intrasite reproducibility of staining using a locked down image analysis algorithm to measure tumor cell and immune cell (IC) subset densities, %PD-L1 expression on tumor cells (TCs) and ICs, and PD-1/PD-L1 proximity assessments.MethodsA six-plex mIF panel (PD-L1, PD-1, CD8, CD68, FOXP3, and CK) was rigorously optimized as determined by quantitative equivalence to immunohistochemistry (IHC) chromogenic assays. Serial sections from tonsil and breast carcinoma and non-small cell lung cancer (NSCLC) tissue microarrays (TMAs), TSA-Opal fluorescent detection reagents, and antibodies were distributed to the six sites equipped with a Leica Bond Rx autostainer and a Vectra Polaris multispectral imaging platform. Tissue sections were stained and imaged at each site and delivered to a single site for analysis. Intersite and intrasite reproducibility were assessed by linear fits to plots of cell densities, including %PDL1 expression by TCs and ICs in the breast and NSCLC TMAs.ResultsComparison of the percent positive cells for each marker between mIF and IHC revealed that enhanced amplification in the mIF assay was required to detect low-level expression of PD-1, PD-L1, FoxP3 and CD68. Following optimization, an average equivalence of 90% was achieved between mIF and IHC across all six assay markers. Intersite and intrasite cell density assessments showed an average concordance of R2=0.75 (slope=0.92) and R2=0.88 (slope=0.93) for breast carcinoma, respectively, and an average concordance of R2=0.72 (slope=0.86) and R2=0.81 (slope=0.68) for NSCLC. Intersite concordance for %PD-L1+ICs had an average R2 value of 0.88 and slope of 0.92. Assessments of PD-1/PD-L1 proximity also showed strong concordance (R2=0.82; slope=0.75).ConclusionsAssay optimization yielded highly sensitive, reproducible mIF characterization of the PD-1/PD-L1 axis across multiple sites. High concordance was observed across sites for measures of density of specific IC subsets, measures of coexpression and proximity with single-cell resolution.


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