Mechanisms of immunosuppression triggered by checkpoint blockade immunotherapy

Checkpoint blockade immunotherapy (CBI) by inhibiting PD1 has provided durable benefits in ~40% of patients with metastatic melanoma and several patients with Merkel cell, non-small cell lung, squamous head and neck, and clear cell kidney cancer. However, PD1 blockade alone and in combination with blockade of other checkpoints (“dual” CBI) still fails in most patients and tumor types. There is an urgent need to understand the underlying reasons; otherwise, CBI will remain ineffective in most cancer patients. One poorly appreciated aspect of PD-1 blockade is that it triggers not only anti-tumor but also immunomodulatory responses, at least partly mediated by T regulatory cells (Tregs). Based on existing literature and extensive preliminary data, we hypothesize that the effect of PD-1 monotherapy on tumor Treg biology depends on the type of antigens expressed by the cancer (CD8-stimulating neoantigens or Treg-supporting dominant autoantigens), the type of DC presenting the antigens to Tregs, and various Treg-supporting pathways instructed by other cells in the tumor immune environment. Moreover, we posit that CBI modulates the spatial organization of the tumor environment and instructs immunosuppression, for instance, by causing the localization of effector T cells to areas of suppressive cytokine or metabolic signaling. The rationale of our study is that understanding the mechanisms of immunosuppression triggered by CBI is a prerequisite to disabling them and increasing the therapeutic efficacy of immunotherapy. Aim 1 will investigate the mechanisms of Treg support enabled by PD-1 blockade. We will study the role of different classes of antigens, and of the DC subtypes presenting them, using functional intravital microscopy. We will also use cell communication analysis of various RNA sequencing data to unbiasedly identify the signaling pathways that support Treg numbers or function after PD-1 blockade. Aim 2 will systematically explore the resistance mechanisms to PD-1 blockade associated with the inhibition of CTLA-4, Lag-3, and ICOSL, specifically focusing on the immunosuppressive spatial reorganizations of the tumor environment that occur after dual CBI. We will stratify animals based on response (or lack thereof) to dual CBI and examine differences in immune cell numbers, phenotype, and spatial organization relative to various intratumor niches via high-resolution spatial sequencing. Finally, we will block the identified mechanisms of adaptive immunosuppression to ameliorate the therapeutic outcome of dual CBI in human-relevant preclinical models. At the conclusion of the proposed project, we would have obtained a mechanistic understanding of the immunosuppressive reactions that cause resistance to PD-1-based CBI. Such knowledge will be instrumental in optimizing clinical immunotherapy and extending its benefits to otherwise resistant patients. Project Number: 1R01CA287116-01A1 | Fiscal Year: 2026 | NIH Institute/Center: National Cancer Institute (NCI) | Principal Investigator: Francesco Marangoni | Institution: UNIVERSITY OF CALIFORNIA-IRVINE, IRVINE, CA | Award Amount: $648,626 | Activity Code: R01 | Study Section: Therapeutic Immune Regulation Study Section[TIR] View on NIH RePORTER: https://reporter.nih.gov/project-details/11298878