VIP Expression Drives an Immunosuppressive Tumor Microenvironment in TP53-Mutated AML
Authors: Zeng Fanyuan , Chaudagar Kiranj , Wang Yuou , Li Jian-Ming , Waller Edmund K. 
Published: December 2024

Introduction: Intensive chemotherapy regimens are a standard of care in AML patients. However, around 10% of AML patients harbor de novo or acquired TP53 mutations and are resistant to chemotherapy. The TP53-mutated sub-group has a median survival of 2-10 months. Recent clinical trials combining hypomethylating agents with phagocytosis checkpoint inhibitors have yielded only modest improvements in the survival of TP53-mutated AML patients, achieving complete remission rates of 32%. Therefore, identifying definitive therapeutic targets in TP53-mutated AML patients is an unmet need. Vasoactive Intestinal Polypeptide (VIP) is a 28 amino acid peptide over-expressed in CD34hi and TP53-mutated AML. VIP interacts with VPAC1 and VPAC2 receptors on immune cells to suppress adaptive and innate immunity in T cells and monocytes. We postulate that the VIP-VPAC signaling axis may serve as a targetable immune checkpoint for TP53-mutated AML.

Methods: Publicly accessible AML patient datasets from cBioPortal were used to investigate the immune characteristics of TP53-mutated AML vs. non-mutated AML and their relationship with the VIP/VIP-R pathway. Confirmatory immune profiling was performed on AML specimens using multi-parameter flow cytometry. Mice engrafted with transplantable AML cell lines were treated with 10-14 daily injections of a VIP-receptor antagonist.

Results: Principal component analysis of gene expression in the AML database demonstrated an enrichment of TP53-mutated AML cluster relative to non-mutated patients. An unbiased analysis of the expression of VIP-signaling and immune checkpoints on T cells and phagocytotic cells showed changes in genes down-stream of the VIP-receptor but without a relationship with PD-L1/PD-1, CTLA4/CD80, CD47/SIRPα pathways. TP53-mutated AML patients had significantly higher VIP expression and lower VPAC2 (vipr2) expression than patients with TP53-non-mutated AML, without significant changes in VPAC1 (vipr1) expression. Correlation analyses across multiple gene pathways identified positive correlations between VIP with CD34, VPAC1 with CD33, and VPAC2 with CD8 in TP53-mutated and non-mutated AML patients. Notably, vipr1 and vipr2 mRNA levels were negatively correlated, suggesting an association of VPAC1 with immunosuppressive myeloid cells and VPAC2 with immunostimulatory lymphoid cells. Differential gene expression and pathway enrichment analysis showed inflammation and innate immunity activation in the TP53-mutated AML tumor microenvironment (TME) vs. the TME in non-mutated AML. Strikingly, CD15 expression was positively correlated with VPAC1 expression in TP53-mutated vs. non-mutated patients, highlighting the potential contributions of myeloid-derived suppressor cells (MDSC) to the TP53-mutated AML TME. To further understand the effect of VIP expression on innate immunity in the TP53-mutated AML TME, a bioinformatics analysis of high VIP-expressing TP53-mutated AML vs. non-mutated AML patients demonstrated suppression of interferon-gamma responses in the TP53-mutated subset. Moreover, we found a positive correlation of PD-L1 expression with VPAC2 expression in VIPhi AML patients. Non-mutated AML samples had a positive correlation between CD16 and VPAC1 compared to mutated cases, suggesting that, in the absence of a TP53 mutation, VIP signaling may enhance the opsonization of AML in response to antibody therapies. These data suggest that the context-specificity of VIP-receptor signaling may affect the susceptibility of AML to novel immune checkpoint therapies. In line with this hypothesis, treatment of leukemic mice with a VIP-receptor antagonist led to an immune-cell-mediated eradication of leukemia, resulting in 40% and 75% long-term survival in VIP (-) P815 and VIP (+) C1498 syngeneic murine AML models, respectively.

Conclusions: The TP53 mutation is a potential driver of immunosuppression in AML patients via a feedforward loop involving VIP and PD-L1 signaling pathways on T cells and monocytes. Monocytes/macrophages that have lost CD16 expression contribute to this immunosuppressive phenotype. Re-inducing the expression of CD16 in conjunction with a VIP-receptor antagonist may be an effective novel treatment strategy for TP53-mutated AML.