Central Function for JAM-a in Multiple Myeloma Patients with Extramedullary Disease
Authors: Solimando Antonio Giovanni , Da Via' Matteo Claudio , Borrelli Paola , Leone Patrizia , Di Lernia Giuseppe , Tabares Gaviria Paula , Brandl Andreas , Pedone Giacinto Luca , Rauert-Wunderlich Hilka , Lapa Constantin , Tafuri Silvio , Bianchi Francesco Paolo , Ria Roberto , Bittrich Max , Ribatti Domenico , Rosenwald Andreas , Kortüm K. Martin , Racanelli Vito , Vacca Angelo , Einsele Hermann , Beilhack Andreas 
Published: October 2019

Abstract
During the last years, multiple myeloma (MM) has been recognized to present in some patients with extra-medullary disease (EMD) manifestation with different pathophysiological and clinical characteristics. Recently, we reported junctional adhesion molecule-A (JAM-A) as a novel target and a clinical biomarker in MM [Solimando et al. Leukemia 32(3):736-743, 2018]. Here, we hypothesized that alterations in the adhesion molecules, particularly JAM-A expression, on MM plasma cells (MM-PCs) may play a pivotal role in high-risk EMD patients.
Therefore, we evaluated JAM-A expression in 60 MM patient bone marrow (BM) aspirates and biopsies at different disease stages with flow cytometry and immunohistochemistry. We compared these results with RNA-Seq data from 647 newly diagnosed MM (NDMM) patients collected in the MMRF (MM Research Foundation) CoMMpass study. Using bioinformatics, we investigated JAM-A related pathways with hierarchical cluster analysis. Subsequently, we functionally tested these JAM-A related pathways regarding epithelial mesenchymal transition, invasion and MM dissemination in vitro.
The median OS differed significantly in subjects with elevated membrane JAM-A expression levels, dividing the patients in EMD JAM-Ahighgroup with a median OS of 84.1 months, whereas median OS in JAM-Alowpatients was not reached, irrespective from the EMD status (log-rank=4.19, P=0.04). Immunohistochemistry analysis of BM biopsies confirmed these findings. RNA-Seq analysis corroborated the prognostic impact of JAM-A (log-rank= 3.8, P= 0.051).High-risk myeloma patients with EMD and JAM-A expression displayed a unique gene-expression signature. Additionally, we constructed a novel expression cluster subgroup MM model, revealing complex molecular disease patterns and associations between clinical traits and expression markers. MM patients clustering according to JAM-A expression levels and EMD manifestation revealed a clear link to the epithelial-mesenchymal-transition and focal adhesion pathways. Notably, functional knockdown of JAM-A in vitro not only reduced cell viability, but also diminished MM adhesion molecules and CD138 surface expression. Furthermore, mTOR/PI3K in vitro inhibition with BEZ235 significantly down regulated surface expression in JAM-Ahigh MM cell lines. Interestingly, when we inhibited mTOR/PI3K in JAM-AlowMM cells, we observed JAM-A induction.
In the present study we shed more light on the EMD pathophysiology. Conclusively, our results support that JAM-A can serve as promising prognostic marker for high risk EMD MM patients. Our data provide a biological rationale for a JAM-A guided decision-making process to better combine molecular and immunochemo-therapy modalities in the EMD MM patient subset.

Disclosures
No relevant conflicts of interest to declare.