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Updated: Jan 10, 2026
An Organotypic High Throughput System for Characterization of Drug Sensitivity of Primary Multiple Myeloma Cells
Published on: July 15, 2015
Enhanced cell surface enrichment and data-independent acquisition for in-depth profiling of multiple myeloma cell
Audrey Kishishita1, Szu-Ying Chen2, Abhilash Barpanda2
1Graduate Program in Chemistry and Chemical Biology, University of California San Francisco, San Francisco, CA, United States; Department of Laboratory Medicine, University of California San Francisco, San Francisco, CA, United States.
Background:
Multiple myeloma is a hematologic malignancy characterized by clonal plasma cell proliferation, with recent therapeutic advances focusing on immunotherapies targeting cell surface antigens. While several surface markers are well-characterized, there remains a critical need to identify additional specific targets for relapsed cases. Comprehensive surface proteome analysis is challenging due to the low abundance of surface proteins and limited cell numbers available from patient samples.
Methods:
We developed an optimized cell surface capture (CSC) protocol coupled with data-independent acquisition (DIA) mass spectrometry for comprehensive surfaceome profiling of multiple myeloma cell lines AMO1 and MM1.S. The method utilizes periodate-based oxidation of surface glycoproteins followed by biocytin hydrazide labeling and NeutrAvidin enrichment. Surface-labeled proteins were analyzed using DIA mode on an Orbitrap Eclipse Tribrid mass spectrometer with optimized parameters for low-input samples.
Results:
Our approach successfully identified and quantified 989 high-confidence surface proteins from minimal sample inputs (10-fold less than standard protocols), demonstrating excellent reproducibility between biological replicates (R=0.907-0.916). Comparative analysis revealed 790 shared proteins between cell lines, with 93 and 106 proteins uniquely expressed in AMO1 and MM1.S, respectively. Principal component analysis showed clear separation between cell lines (PC1=84.6% variance), highlighting distinct surface protein expression profiles. The method detected known myeloma markers including BCMA and identified additional potential therapeutic targets.
Conclusions:
CSC-DIA methodology enables comprehensive surface proteome analysis from limited cell numbers, making it suitable for rare patient samples. This provides a robust platform for discovering novel surface antigens to advance personalized myeloma immunotherapy.
Insights
A new cell surface capture method combined with mass spectrometry identifies hundreds of surface proteins, offering new targets for multiple myeloma immunotherapy, even from limited patient samples.
Area of Science:
- Proteomics
- Immunology
- Oncology
Background:
- Multiple myeloma is a plasma cell cancer with evolving immunotherapies targeting surface antigens.
- Identifying new targets is crucial for relapsed cases, but surface proteome analysis is difficult with limited patient samples.
- Challenges include low surface protein abundance and restricted cell numbers.
Purpose of the Study:
- To develop and validate an optimized cell surface capture (CSC) method coupled with data-independent acquisition (DIA) mass spectrometry.
- To enable comprehensive surfaceome profiling of multiple myeloma cell lines using minimal sample input.
- To identify novel surface antigens for potential therapeutic development in multiple myeloma.
Main Methods:
- Developed an optimized CSC protocol using periodate oxidation, biocytin hydrazide labeling, and NeutrAvidin enrichment.
- Applied DIA mass spectrometry on an Orbitrap Eclipse Tribrid mass spectrometer with parameters for low-input samples.
- Analyzed multiple myeloma cell lines AMO1 and MM1.S.
Main Results:
- Identified and quantified 989 high-confidence surface proteins from limited cell numbers with high reproducibility (R=0.907-0.916).
- Revealed distinct surface protein expression profiles between AMO1 (93 unique proteins) and MM1.S (106 unique proteins).
- Detected known markers like BCMA and identified additional potential therapeutic targets.
Conclusions:
- The CSC-DIA methodology allows comprehensive surface proteome analysis from limited cell numbers, suitable for rare patient samples.
- This robust platform facilitates the discovery of novel surface antigens.
- Enables advancements in personalized multiple myeloma immunotherapy.
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