Abstract 3637: Age-dependent loss of HAPLN1 erodes vascular integrity via upregulation of endothelial ICAM1 in melanoma
Authors: Marino Gloria E. , Chhabra Yash , Kaur Amanpreet , Almeida Filipe , Liu Ying , Eisinger-Mathason Karin T. S. , Weeraratna Ashani T. 
Published: June 2022

Abstract
The 5-year survival rate for metastatic melanoma is only 23%, and patients over 55 years old have higher incidence of metastasis than younger patients independent of other prognostic factors. Our lab has shown that age-dependent changes in the tumor microenvironment drive this disparity. In particular, loss of a secreted extracellular matrix protein, HAPLN1, in the aged dermis confers increased metastasis via matrix breakdown and resulting cellular dysfunction. In this study we found that HAPLN1-mediated matrix changes impede the barrier function of blood vessels and upregulate vascular ligands which in turn increase the propensity for melanoma intravasation and metastasis via this route. VE-cadherin is a key endothelial cell junctional protein whose expression is directly correlated with vascular integrity. We showed via IHC that intradermal treatment of aged mice with recombinant HAPLN1 (rHAPLN1) is sufficient to significantly increase VE-cadherin levels in intratumoral blood vessels compared to untreated aged mice, equal to levels seen in young mice. Further, we isolated the contribution of the matrix by producing in vitro fibroblast derived matrices (FDMs) from the dermal fibroblasts of healthy young and aged donors. An impedance-based assay showed that resistance of human endothelial cell (HUVEC) monolayers is significantly decreased when cultured on aged matrix compared to young or aged+rHAPLN1 matrices. We found via qRT-PCR screen that HUVECs cultured on HAPLN1-low FDMs (aged, young+shHAPLN1) have upregulated ICAM1 compared those in HAPLN1-high environments (young, aged+rHAPLN1). ICAM1 is an endothelial cell receptor whose upregulation initiates a signaling cascade to downregulate VE-cadherin and increase vascular permeability. We confirmed the HAPLN1-dependent expression of ICAM1 in vivo via IHC of murine tumors. ICAM1 expression is known to be modulated in response to ECM stiffness. We have shown that matrices formed by aged fibroblasts are stiffer than those formed by young, and that this is due in part to the absence of HAPLN1. We created artificially stiffened FDMs via chemical cross-linking of otherwise pliable young FDMs to determine if stiffness alone was sufficient to upregulate ICAM1 on endothelial cells. We found via qRT-PCR that artificially stiffened young matrices were sufficient to increase endothelial ICAM1 levels on par with those in aged and young+shHAPLN1 conditions. This is critical, as endothelial ICAM1 is upregulated in melanoma and other cancers and is used as a ligand for cancer cells to move through the vasculature. In sum, we have found that age-dependent loss of HAPLN1 in the dermal ECM confers increased matrix stiffness which leads to the upregulation of endothelial ICAM1. This in turn increases vascular permeability to allow melanoma cells to more easily intravasate and metastasize via the hematogenous pathway in the aged system.
Citation Format: Gloria E. Marino, Yash Chhabra, Amanpreet Kaur, Filipe Almeida, Ying Liu, Karin T. S. Eisinger-Mathason, Ashani T. Weeraratna. Age-dependent loss of HAPLN1 erodes vascular integrity via upregulation of endothelial ICAM1 in melanoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3637.