Abstract 4675: Aryl hydrocarbon receptor antagonist enhances cord blood-derived human hematopoietic stem cell expansion and platelet formation
Authors: Kim Dong Chan , Yoon Sung-Soo , Shin Dong-Yeop 
Published: August 2019

Background: Cord blood transplantation is one of the alternatives for hematopoietic stem cell transplantation (HSCT). However, cord blood transplantation has a crucial problem with the small number of hematopoietic stem cells (HSC) in a single cord blood unit which results in graft failure. To overcome this problem with the paucity of HSCs, here we tested the effect of two aryl hydrocarbon receptor (AHR) antagonists on the HSC ex vivo expansion and explored the mechanism of different effect by analyzing the gene expression profiles related with lineage-specific commitment in expanded human HSPCs.

Methods & Results: For ex vivo expansion culture, we enriched CD34 positive cell proportion up to 90% from cord blood units. We demonstrated ex vivo expansion culture with two AHR antagonist, StemRegenin 1 (SR1) and CH223191. Lin-/CD34+/CD38-, Lin-/CD34+/CD90+ and Lin-/CD34+/CD45RA+cell proportions are increased by SR1 (x6.5, 4.1, 2.9) and CH223191 (x3.6, 2.5, 2.6) at 14 culture days. With repeated analysis, the absolute number of Lin-/CD34+/CD38-/CD90+/CD45RA-cells were increased more than initial seeding cell numbers. We further performed the colony forming assay (CFA) with expanded HSPCs to functionally validate of expanded HSPCs’ stemness. The number of each colony distinguished by their morphologies were counted; BFU, CFU-G, -M, -GM, -GEMM. SR1 and CH223191 treatment increased the total number of colonies compared to control, and the number of progenitor cells was increased by antagonists. Fourteen days after CFA culture, colonies were harvested and analyzed by FACS with various hematopoietic lineage cell markers (CD45, CD33, CD235a, and CD19). There were differences between SR1- and CH223191-expanded HSPC by FACS analysis and CFA, though two molecules had the similar ability to expand hematopoietic stem/progenitor cell number with the differentiation capacity. To explore the putative cause of the different effect on HSPC expansion capacities of SR1 and CD223191, we performed total-RNA sequencing (Illumina NextSeq) using expanded progenitor cells and examined results in various ways. By differential expressed gene (DEG) analysis, we selected genes significantly changed their expression. Comparing with control, SR1 and CH223191 decreased the expression of genes which are known to be related with the AhR pathway. On the other hand, we found that CH223191 treatment increased gene expression related to platelet activation and formation, coagulation cascade and complement.

Conclusions: In this study, we show that the blockade of aryl hydrocarbon receptor results in HSPCs expansion. CH223191 induces the expansion of HSPCs and the gene expression which are related to platelet formation. On this point, further investigation for the therapeutic use of CH223191 for expanded cord blood transplant and the treatment amegakaryocytic thrombocytopenia is needed.

Citation Format: Dong Chan Kim, Sung-Soo Yoon, Dong-Yeop Shin. Aryl hydrocarbon receptor antagonist enhances cord blood-derived human hematopoietic stem cell expansion and platelet formation [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 4675.