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Regular and Young Investigator Award Abstracts
Immune Cell Types and Biology
863 Characterization of tumor-infiltrating plasma cells in patients with NSCLC treated with neoadjuvant pembrolizumab
  1. Robert R Meyerhoff1,
  2. Alan C Chen2,
  3. Carmen Anadon-Galindo2,
  4. Gabor Kemeny2,
  5. Liliana Lyniv2,
  6. Vaibhav Jain2,
  7. Tyler Evangelous2,
  8. Madison Berry2,
  9. Katelyn Steadman2,
  10. Karen Abramson2,
  11. Kevin Wiehe2,
  12. Kent Weinhold2,
  13. Simon Gregory2,
  14. Jose Conjeo-Garcia2,
  15. Wilton B Williams2 and
  16. Scott J Antonia3
  1. 1Duke University School of Medicine, Durham, NC, USA
  2. 2Duke University, Durham, NC, USA
  3. 3Duke University Medical Center, Durham, NC, USA

  • Journal for ImmunoTherapy of Cancer (JITC) preprint. The copyright holder for this preprint are the authors/funders, who have granted JITC permission to display the preprint. All rights reserved. No reuse allowed without permission.

Abstract

Background Programmed cell death-1 (PD1) blockade is associated with development of intratumoral tertiary lymphoid structures (TLSs).1–5 These structures represent ectopic lymphoid tissue that can support development of tumor-infiltrating B cells (TIL-B), including antibody-secreting plasma cells that are associated with improved survival in patients with soft tissue sarcoma, pancreatic ductal adenocarcinoma, and non-small cell lung cancer (NSCLC).4 6–8 We hypothesized that pembrolizumab (PD1 inhibitor) is associated with expansion of TIL-Bs capable of secreting tumor antigen-reactive antibodies in patients with NSCLC.

Methods We performed high-throughput single-cell B cell receptor (BCR) sequencing on CD138-enriched plasma cells isolated from disaggregated NSCLC tumors excised from patients treated with neoadjuvant pembrolizumab (TOP1501 phase II study, NCT02818920). This study was approved by Duke University’s Ethics Board; approval number Pro000071629. Using the CellRanger software suite (10X Genomics), BCRs with identical paired HCDR3 and LCDR3 were identified and deemed to be clonally related (‘clonotypes’). B cell clonotypes were ranked from highest frequency (most expanded) to lowest frequency (least or non-expanded). Consensus BCRs for the most frequently observed clonotypes were then used to generate recombinant monoclonal antibodies (mAbs) and tested for binding specificities and effector functions.

Results We generated 20 mAbs representative of the most expanded clonotypes from three patients and found that they bound to the surface of immortalized lung tumor cell lines by immunofluorescence staining. Using a combination of co-immunoprecipitation, western blot, and mass spectrometry, we identified cognate antigens bound by our lead tumor-reactive mAb candidate (‘CL1-Ab4’), including several intracellular host proteins known to be expressed on the cell surface in stressed or malignant cells; citrullinated Vimentin, citrullinated phosphoglycerate dehydrogenase (PHGDH) and Calreticulin.9–11 Binding of CL1-Ab4 and other tumor-reactive mAbs to these antigens was confirmed with ELISA. To test the capability of CL1-Ab4 to redirect chimeric antigen receptor (CAR)-T cells to tumor cells for killing, we generated CAR-T cells bearing the CL1-Ab4 (scFv) region and found these cells able to mediate dose-dependent killing of tumor cell lines in vitro, thus highlighting an anti-tumor therapeutic promise for CL1-Ab4.

Conclusions We established a pipeline for isolation of tumor-reactive antibodies and identification of the tumor-associated antigens these antibodies recognize. That these antibodies demonstrate some level of polyspecificity may provide a beneficial feature for overcoming the diversity of tumor associated antigens. Additionally, these data highlight the potential of this pipeline for identification of tumor antibodies that may have therapeutic potential.

References

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Ethics Approval This study was approved by Duke University’s Ethics Board; approval number Pro000071629. All research participants provided their informed consent before taking part in the clinical trial from which study samples were derived (NCT02818920).

http://creativecommons.org/licenses/by-nc/4.0/

This is an open access article distributed in accordance with the Creative Commons Attribution Non Commercial (CC BY-NC 4.0) license, which permits others to distribute, remix, adapt, build upon this work non-commercially, and license their derivative works on different terms, provided the original work is properly cited, appropriate credit is given, any changes made indicated, and the use is non-commercial. See http://creativecommons.org/licenses/by-nc/4.0/.

https://doi.org/10.1136/jitc-2024-SITC2024.0863

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