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84 Deciphering the immune microenvironment in castration-resistant prostate cancer (CRPC) through hyperplex immunofluorescence and AI-assisted image analysis: Unveiling potential immunotherapy biomarkers
  1. Bora Gurel,
  2. Ines Figueiredo,
  3. Mateus Crespo,
  4. Christina Guo,
  5. George Seed,
  6. Ana Ferreira,
  7. Ruth Riisnaes,
  8. Wei Yuan,
  9. Suzanne Carreira and
  10. Johann De Bono
  1. The Institute of Cancer Research, London, UK
  • 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.


Background Prostate Cancer (PCa) is a major health concern in developed countries, with rising incidence and progression to advanced stages, necessitating effective treatments. Chronic inflammation is identified as a key factor in PCa pathogenesis and its advancement, creating a complex dynamic between inflammation, genetic changes, and the tumor microenvironment. Under chronic inflammation, the risk of mutation increases due to increased DNA damage and cell regeneration. Inflammation also transforms the tumor microenvironment, promoting tumor growth and potential immune evasion.

Methods Our study used single-stain CD3 IHC, Hyperplex Immunofluorescence (IF), and AI-assisted image analysis to explore the landscape in castration-resistant prostate cancer (CRPC). Evaluating a cohort of 360 hormone-sensitive prostate cancer (HSPC) and 465 CRPC samples, we discovered significant inflammation in a substantial subset, challenging the traditional view of prostate cancer as a ‘cold’ tumor.

Results We leveraged density-based clustering algorithms to decode distinctive spatial patterns of CD3+ cell distribution in CRPC, finding a significant positive correlation with patient survival. We noted diffuse and nodular patterns of CD3+ inflammation, with the nodular pattern associated with worse survival outcomes, possibly indicating immature tertiary lymphoid structures (TLSs). Through IF data, we charted immune landscapes in a smaller CRPC cohort, revealing distinct immune responses in inflamed tumors. We identified high, diffuse inflammation with a prevalence of various immune cells, and another characterized by nodules formation by specific immune cells, hinting at emerging immature TLSs. We observed a correlation between mature TLSs and intense inflammation, suggesting potential NLRP3 inflammasome activation or Senescence Associated Secretory Phenotype (SASP).

Conclusions Our work provides insights into the complex immune microenvironment in CRPC, offering a new understanding of its role in disease progression. The potential biomarkers identified may enhance the effectiveness of immunotherapy, helping to select and monitor patients for immune checkpoint inhibition therapy, with the goal of improving patient outcomes.

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

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