Pancreatic cancer-initiating cell exosome message transfer into noncancer-initiating cells: the importance of CD44v6 in reprogramming

Abstract

BackgroundCancer-initiating cell (CIC) exosomes (CIC-TEX) are suggested reprogramming Non-CIC. Mode of message transfer and engagement of CIC-markers being disputed, we elaborated the impact of CD44v6 and Tspan8 on the response of Non-CIC.MethodsNon-metastasizing CD44v6- and Tspan8-knockdown (kd) pancreatic cancer cells served as Non-CIC. CIC-TEX coculture-induced changes were evaluated by deep-sequencing and functional assays. Tumor progression was surveyed during in vivo CIC-TEX treatment.ResultsDeep-sequencing of CIC-TEX-cocultured CD44v6kd-Non-CIC revealed pronounced mRNA changes in signaling, transport, transcription and translation; altered miRNA affected metabolism, signaling and transcription. CIC-TEX coculture-induced changes in Tspan8kd-Non-CIC mostly relied on CIC-TEX-Tspan8 being required for targeting. CIC-TEX transfer supported apoptosis resistance and significantly promoted epithelial mesenchymal transition, migration, invasion and (lymph)angiogenesis of the kd Non-CIC in vitro and in vivo, deep-sequencing allowing individual mRNA and miRNA assignment to altered functions. Importantly, CIC-TEX act as a hub, initiated by CD44v6-dependent RTK, GPCR and integrin activation and involving CD44v6-assisted transcription and RNA processing. Accordingly, a kinase inhibitor hampered CIC-TEX-fostered tumor progression, which was backed by an anti-Tspan8 blockade of CIC-TEX binding.ConclusionsThis in depth report on the in vitro and in vivo impact of CIC-TEX on CD44v6kd and Tspan8kd Non-CIC unravels hub CIC-TEX activity, highlighting a prominent contribution of the CIC-markers CD44v6 to signaling cascade activation, transcription, translation and miRNA processing in Non-CIC and of Tspan8 to CIC-TEX targeting. Blocking CIC-TEX binding/uptake and uptake-initiated target cell activation significantly mitigated the deleterious CIC-TEX impact on CD44v6kd and Tspan8kd Non-CIC.Electronic supplementary materialThe online version of this article (10.1186/s13046-019-1129-8) contains supplementary material, which is available to authorized users.

Tspan8-Tumor Extracellular Vesicle-Induced Endothelial Cell and Fibroblast Remodeling Relies on the Target Cell-Selective Response

“…The reverse, a lower signal strength in both ASML-Tspan8kd-TEX and AS-TEX accounted only for 10 miRNA ( Figure S2E,F). Though confirming a slight impact of Tspan8 on miRNA recovery in TEX, we recently elaborated that we are dealing with an indirect effect due to Tspan8 associating with proteins that are directly engaged in miRNA recruitment into ILV [54].…”

“…TEX received much attention as transferring tumor-growth and -progression promoting activities into non-cancer-initiating cells (Non-CIC) [88] as well as host cells promoting (lymph)angiogenesis [7,89], premetastatic niche establishment [8][9][10], and deviation of hematopoietic cells towards immunosuppression [90]. We recently elaborated the mode whereby CIC-TEX affect Non-CIC, which revealed that an impact of TEX-Tspan8 predominantly relies on the engagement of Tspan8 complexes in TEX biogenesis [5,30,54,57,91] including the transfer of Tspan8-associated CIC-markers into TEX and, most importantly, on Tspan8 facilitating TEX binding and uptake by target cells via ligands for Tspan8-associated molecules [5,27,38,52,55,68]. Uptaken TEX stimulating non-CIC inherent programs [5,30,54] and Tspan8 being particularly engaged in TEX binding and uptake, we asked whether non-transformed cells respond alike oncogene-transformed cells.…”

“…We recently elaborated the mode whereby CIC-TEX affect Non-CIC, which revealed that an impact of TEX-Tspan8 predominantly relies on the engagement of Tspan8 complexes in TEX biogenesis [5,30,54,57,91] including the transfer of Tspan8-associated CIC-markers into TEX and, most importantly, on Tspan8 facilitating TEX binding and uptake by target cells via ligands for Tspan8-associated molecules [5,27,38,52,55,68]. Uptaken TEX stimulating non-CIC inherent programs [5,30,54] and Tspan8 being particularly engaged in TEX binding and uptake, we asked whether non-transformed cells respond alike oncogene-transformed cells. We selected for Fb that contribute to premetastatic niche generation [9,92] and EC responding to TEX after 1h to 5d [50], which we considered a hint for a target cell autonomous response.…”

“…293T cells (Cell bank, China) were cultured in high-glucose Dulbecco's modified Eagle's medium/10%FBS/antibiotics (Gibco, CA, USA). The human PaCa line A818.4 and an A818.4-Tspan8kd are described [53,54]. All cultures were kept in a humidified atmosphere at 37 • C, 5%CO 2 in air.…”

“…AS-Tspan8-TEX promote EC growth, spriting and tube formation ( Figure 5A) [46,50,54,[68][69][70]. Because distinct to Fb, expression of only few mRNA became upregulated by coculture with AS-TEX, we searched only for clusters of upregulated genes in AS-Tspan8-TEX-treated EC.…”