Single-cell RNA-seq uncovers that plerixafor can mobilize more monocyte progenitor cells and enhance the activating graft immune status in multiple myeloma autologous transplantation. Free

Background:

Autologous Hematopoietic Stem Cell Transplantation (Auto-HSCT) is one of the most effective treatments for patients with Multiple Myeloma (MM), significantly improving survival rates and quality of life. Adequate mobilization and collection of peripheral blood Hematopoietic Stem Cells are crucial for the success of Auto-HSCT. The CXCR4 antagonist Plerixafor (AMD3100), in combination with G-CSF, has been widely used for clinical mobilization, especially when G-CSF alone fails to mobilize sufficient CD34+ cells. Previous studies have shown that Plerixafor enhances hematopoietic recovery after myeloablative allogeneic hematopoietic stem cell transplantation. However, despite the well-documented efficacy of Plerixafor in mobilizing CD34+ cells, its impact on other cell types and states within the graft, as well as the overall immune status of the graft, remains underexplored. Therefore, this study aims to comprehensively investigate the effects of Plerixafor mobilization on graft cell composition and immune status using single-cell RNA sequencing (scRNA-seq) technology.

Methods:

Clinical Data Collection:

We conducted a single-center retrospective analysis by analyzing data from 123 MM patients who underwent mobilization at Sun Yat-sen University Cancer Center from January 1, 2021, to May 1, 2025.

Single-Cell RNA Sequencing: Fresh peripheral blood samples were collected from 4 patients at three critical time points: before Plerixafor administration(Plerixafor_before), after Plerixafor administration(Plerixafor_after), and immune reconstitution after autologous transplantation(immune_reconstitution).(a) Single-cell RNA sequencing data were subjected to quality control, where low-quality cells and lowly expressed genes were removed; (b) Data were integrated using robust principal component analysis (rPCA) to remove batch effect.

Results:

1,123 mobilized MM patients were included in this study, and 38.2% (47/123) of patients were female, and 61.8% (76/123) of patients were male, with a mean age of 53.5 years and a mean weight of 63.2kg. As for monoclonal immunoglobulin type, IgG (60/123,48.8%), Light Chain Myeloma (24/123,19.5%), and IgA(18/123,14.6%) are the top three, and 51.2% patients express Kappa light chain and 34.1% patients express Lambda light chain.There was a higher proportion of CD34 + cells (0.39 vs 0.20, p = 0.022) in the collection after Plerixafor compared to the mobilized harvest without Plerixafor, and the percentage of CD34 + cells in the collection after Plerixafor was significantly higher than that after Plerixafor (p = 0.022), more CD34 + cells could be mobilized on the first day of drying (3.009 * 10 ^ 6 cells/kg vs 4.449 * 10 ^ 6 cells/kg, p = 0.0425). The percentage of neutrophil lymphocyte decreased more (0.02933 vs-0.05352, p < 0.0001) and increased more (- 0.02905 vs 0.009634, p < 0.0001) after the use of Plerixafor compared with the mobilization harvesting without Plerixafor. The proportion of monocytes increased more (0.002014 vs 0.03811, P = 0.0032).

2、After integrating the single-cell data from the three time points using rPCA, a total of 101,955 high-quality cells were obtained, which could be annotated into 12 subtypes (HSC, Pro-Monocyte, Pro-Neutrophil, Pro-TNK, Neutrophil, Monocyte, T cell, NK cell, B cell, EBM, platelet, and DC. Plerixafor_after data has a higher proportion of HSCs, Pro-Monocyte, and Monocyte compared with Plerixafor_before data.Interestingly, CD16+ Monocyte has high expression of complement 1q (C1Q genes, including C1QA, C1QB, and C1QC) and macrophage markers (CD68).

In the immune_reconstitution data, a higher proportion than Plerixafor_before data and the Plerixafor_after data of conventional DCs was observed, which represents higher antigen presentation capacity. Compared with Plerixafor_before data, neutrophils of Plerixafor_after data are down-regulated in NF-kappa B signaling pathway, which is important in the formation and activation of Myeloid-derived suppressor cells.

Conclusion:This finding suggests that Plerixafor mobilization not only significantly increases the proportion of CD34+ cells and monocytic progenitor cells but also alters the overall immune status of the graft, shifting it towards an immune-activated state, providing a new perspective for optimizing mobilization strategies in MM autologous transplantation and may have significant implications for immune reconstitution and transplantation outcomes.