Characterization of Upregulated Adhesion GPCRs in Acute Myeloid Leukemia

Background: Adhesion G-protein coupled receptor (aGPCR) is one of the five GPCRs families that are named after their large extracellular regions linked to the seven-transmembrane moiety (TM7) and are found to have a dual role in signaling and cell adhesion. In normal tissues, aGPCRs play important roles in neuronal development and the immune system. The aGPCR GPR56 has been reported to be involved in the hematopoiesis and the interaction between hematopoietic stem cells and the bone marrow. In malignant tissues, aGPCRs were reported to play a role in tumor cell migration and invasion of melanoma and prostate cancers, and are relevant to the regulation of angiogenesis in glioblastomas. Limited data are available related to the expression and role of aGPCRs in hematological malignancies, particularly in acute myeloid leukemia. Here, we aim to characterize the expression of aGPCRs in patients with AML and examine whether upregulation of genes in this family is associated with clinical and molecular patient's characteristics.

Methods: We analyzed the TCGA data of 200 AML patients for which 173 have complete clinical and expression data available. We examined gene upregulation of the 33 aGPCRs and assessed the association between each gene and patient's overall survival (OS); genes that were significantly associated with shorter overall survival were combined in one signature (poor survival genes) for further analysis to test their association with patient's clinical and molecular characteristics and clinical outcome. To gain an insight into the mechanistic role of aGPCRs in AML, we conducted Ingenuity Pathway Analysis (IPA) to identify potential pathways common among the poor survival aGPCR genes, and between the aGPCRs genes and AML frequently mutated genes.

Results: Among the 33 aGPCRs; eight were found to be associated with worse clinical outcome. Patients with high expression (Z≥1) in one of the eight genes have significantly shorter median overall survival and disease-free survival (DFS) compared with patients with low (Z<1) expression of that gene (ADGRB1: OS 8.20 vs 21.50 months, p=0.0157; CELSR2: OS 10.00 vs 21.50 months, p=0.0488; ADGRD1: OS 11.10 vs 20.50 months, p=0.0085; ADGRE1: OS 10.60 vs 20.50 months, p=0.0198; ADGRE2: DFS 12.10 vs 18.20 months, p=0.0374; ADGRE5: OS 7.35 vs 24.10 months, p=0.0015; ADGRG1: OS 6.80 vs 22.30 months, p=0.0057; ADGRG3: OS 10.00 vs 21.50 months, p=0.0177). Among the 200 samples in TCGA data; 118 (72.8%) of 162 sequenced patients have an alteration in at least one of these eight genes with a minimum 10% alternation in ADGRD1 to a maximum 33% alternation in ADGRE2, most of them are upregulation. When we combined all eight genes in one signature for further analysis; we found that patients with high expression (Z≥1) in any of the eight genes (ADGRB1, CELSR2, ADGRD1, ADGRE1, ADGRE2, ADGRE5, ADGRG1, ADGRG3), have significantly shorter overall survival or disease-free survival compared with patients with low (Z<1) expression of all eight aGPCR genes (median OS:11.8 vs 55.4 months; p< 0.0001). Next, we examined the association between the expression of eight aGPCRs with patient primary and molecular characteristics. High expression of the eight aGPCRs was significantly associated with older age (≥60; p=0.011). Patients with high aGPCRs are more frequently classified in the poor risk status group and less in the good risk group compared with patients with low aGPCRs levels (31% vs 17% p=0.049 and 14% vs 28% p=0.027, respectively). Multivariate analyses showed that high (Z≥1) aGPCRs expression was associated with shorter overall survival in patients with AML (HR:1.73, 95% CI 1.11-2.69; p=0.015) after adjusting by age, molecular risk, and transplant status. Using IPA analysis, we identified the SNX27 gene, to connect three of the eight aGPCRs. SNX27 is a member of sorting nexin protein family and is implicated in endocytic trafficking. Importantly, we found that patients with low (Z≤-1) expression of SNX27 have a significantly shorter overall survival compared with patients with Z>-1 (median OS: 8.10 vs 20.50 months, p=0.0251).

Conclusion: Overall, our data suggest that particular aGPCRs are frequently upregulated in AML and that their overexpression is associated with poor clinical outcome. Future functional and mechanistic analysis are needed to address the role of aGPCRs in AML.