Literature Analysis: Application of Multiplex Fluorescence IHC in Efficacy Evaluation of Tumor Immunotherapy

1.Literature Information

Journal: JAMA Oncology

DOI: 10.1001/jamaoncol.2019.1549

Published Online: 18 Jul 2019 (Online ahead of print)

PMID: 31318407 | PMCID: PMC6646995

Authors: Steve Lu, Julie E Stein, David L Rimm, Daphne Wang, J Michael Bell, Douglas Janson, Jeffrey A Sosman, Kurt A Schalper, Robert A Anders, Hao Wang, Clifford Hoyt, Drew M Pardol, Ludmila Danilova, Janis M Taube

Research Focus: A systematic review and meta-analysis to compare the accuracy of different biomarker modalities (PD-L1 IHC, TMB, GEP, mIHC/IF) in predicting the response to anti-PD-1/PD-L1 checkpoint blockade therapy

Key Technology: Systematic review and meta-analysis, integrating data from multiple studies using PD-L1 IHC, Tumor Mutation Burden (TMB), Gene Expression Profiling (GEP), and Multiplex Immunohistochemistry/Immunofluorescence (mIHC/IF)

Sample Scale: A total of 8135 cases across 10 types of solid tumors, with quantitative synthesis (meta-analysis) of 45 publications involving 56 individual analyses

Key Product Utilized: Multiplex Fluorescence IHC Staining Kits and comprehensive technical services from the Absin product line of ANT BIO PTE. LTD.

2. Research Background

Immune checkpoint blockade therapy targeting PD-1/PD-L1 has revolutionized the treatment landscape of various solid tumors. However, only a subset of patients responds to this therapy, highlighting the urgent need for reliable predictive biomarkers to identify potential responders and optimize clinical treatment strategies.

Currently, several biomarker detection modalities have been developed for evaluating pretreatment tumor tissues, including PD-L1 (Programmed Death-Ligand 1) Immunohistochemistry (IHC), Tumor Mutation Burden (TMB), Gene Expression Profiling (GEP), and Multiplex Immunohistochemistry/Immunofluorescence (mIHC/IF). However, the comparative accuracy of these methods in predicting treatment response remains unclear, which limits their rational application in clinical practice.

Traditional single-biomarker detection methods (such as PD-L1 single-index IHC) have inherent limitations in capturing the complexity of the tumor immune microenvironment. In contrast, mIHC/IF technology enables simultaneous detection of multiple immune-related markers, providing a more comprehensive assessment of the tumor immune landscape. A systematic comparison of the predictive value of these modalities is essential to confirm the clinical application value of mIHC/IF and guide the selection of optimal biomarkers for anti-PD-1/PD-L1 therapy.

3. Research Approach

The research team conducted a systematic review and meta-analysis following the PRISMA (Preferred Reporting Items for Systematic Reviews and Meta-Analyses) guidelines. A comprehensive literature search was performed to identify relevant studies, including database searches, DailyMed searches, and searches of meeting abstracts. A total of 135 records were initially identified, with 98 additional records from database searches and 25 from meeting abstracts. After removing 15 duplicate records, 243 records were screened, and 198 were excluded based on predefined criteria (e.g., non-original investigations, no PD-L1-treated population, sample size <15 patients, no reporting of Response Rate (RR) and/or Overall Survival (OS)/Progression-Free Survival (PFS), hematologic malignancies, blood-based profiling, studies on tumor lysates, and repeat presentations). Finally, 45 publications involving 56 individual analyses were included in the quantitative synthesis.

The team analyzed the correlation between different biomarker modalities (PD-L1 IHC, TMB, GEP, mIHC/IF) and patient response to anti-PD-1/PD-L1 therapy. The primary outcome measures included the Area Under the Curve (AUC) for predictive accuracy, Positive Predictive Value (PPV), Negative Predictive Value (NPV), Positive Likelihood Ratio (PLR), and Negative Likelihood Ratio (NLR). Subgroup analyses were also performed to explore the consistency of results across different tumor types and study designs.

4. Research Results

Firstly, the meta-analysis results showed that compared with single-index PD-L1 IHC, TMB, and GEP, the mIHC/IF method had a significantly higher Area Under the Curve (AUC) in pre-treatment prediction of anti-PD-1/PD-L1 therapy response. This indicates that mIHC/IF is more accurate in predicting treatment response.

Secondly, even compared with the combined evaluation of the three modalities (PD-L1 single-index IHC, TMB, and GEP), the mIHC/IF method still achieved a higher AUC. This demonstrates the unique advantage of mIHC/IF in predictive accuracy, surpassing the combined performance of traditional single biomarkers.

Thirdly, in terms of clinical practical value, mIHC/IF showed superior performance in both positive and negative prediction compared to the other three modalities and their combination. This means that mIHC/IF is more effective in identifying patients who are likely to respond (positive prediction) and those who are not (negative prediction), providing reliable guidance for clinical patient selection.

Collectively, the analysis of 8135 cases across 10 solid tumor types confirmed that mIHC/IF has higher accuracy than PD-L1 single-index IHC, TMB, GEP, and their combination in predicting the efficacy of anti-PD-1/PD-L1 therapy, indicating significant application value in clinical diagnosis and treatment.

5. Product Empowerment

The outstanding performance of mIHC/IF in tumor immunotherapy efficacy prediction highlights the importance of high-quality mIHC solutions. ANT BIO PTE. LTD. provides comprehensive Multiplex Fluorescence IHC services and products through its Absin product line, which fully supports research and clinical applications in this field.

Absin Multiplex Fluorescence IHC services cover the entire process from pre-experiment verification, staining, imaging to data analysis. The service supports up to 6 indicators with 7 colors (including DAPI) for multi-color IHC staining (up to 4 indicators with 5 colors for tissue microarrays). Applicable sample types include paraffin sections, frozen sections, and tissue microarrays (TMA); for large tissue sections, FFPE samples with a thickness of 3-5 μm are recommended, and it is advisable to provide 2 sections per case (1 for backup). The number of pre-experiment samples is determined according to the pre-experiment protocol (all antibody indicators need to be provided by the customer).

The service results provided include: 1) For 5 colors or less, full-slide 200X original images (equivalent to 20X objective and 10X eyepiece) are provided; for 6-7 colors, 200X images of designated fields are provided (3 fields by default, additional fields are charged separately). A viewing software compatible with client terminals is provided, and data is uploaded to a cloud disk for samples ≤1G or sent via USB flash drive for samples >1G. 2) Single-cell analysis results of customer-specified regions: percentage and density of single-positive cells relative to nucleated cells; percentage and density of double-positive cells relative to nucleated cells; calculation of tissue area and specific pathological morphology area; for TMA, the above quantitative results for each core are provided.

Absin Multiplex Fluorescence IHC Staining Kits (4-color to 7-color) are composed of high-quality components, including TSA fluorescent dyes, signal amplification reaction solution, mouse/rabbit universal or anti-rabbit HRP-labeled secondary antibodies, anti-fluorescence quenching mounting medium, and DAPI. These components ensure high signal intensity, stable fluorescence, and clear imaging results, laying a solid foundation for accurate detection and analysis of multiple immune markers.

6. Brand Mission

ANT BIO PTE. LTD. is dedicated to empowering life science research through the provision of high-quality, reliable reagents and comprehensive solutions. Our specialized sub-brands (Absin, Starter, UA) are strategically positioned to cover a full spectrum of research needs: Absin focuses on general reagents and kits, Starter specializes in antibodies, and UA is dedicated to recombinant proteins. We strive to support researchers in unlocking scientific mysteries, advancing medical progress, and addressing unmet clinical needs through continuous innovation and customer-centric services.

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8. Disclaimer

This article is AI-compiled and interpreted based on the original work in DOI: 10.1001/jamaoncol.2019.1549 and document 1226.docx (Application of mIHC in Efficacy Evaluation of Tumor Immunotherapy). All intellectual property (e.g., images, data) of the original publication shall belong to the journal and the research team. For any infringement, please contact us promptly and we will take immediate action.

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At ANTBIO, we are committed to advancing life science research through high-quality, reliable reagents and comprehensive solutions. Our specialized sub-brands (Absin, Starter, UA) cover a full spectrum of research needs, from general reagents and kits to antibodies and recombinant proteins. With a focus on innovation, quality, and customer-centricity, we strive to be your trusted partner in unlocking scientific mysteries and driving medical progress. Explore our product portfolio today and elevate your research to new heights.