Myeloid-Derived Suppressor Cells As Potential Contributors to Pregnancy-Associated Lymphoma Progression

Introduction: Non-Hodgkin lymphoma (NHL), the most frequent hematological malignancy observed during pregnancy, is reported to be associated with advanced disease stage at diagnosis, essential involvement of reproductive organs, and a turbulent course. Immunosuppression is one of the key players in maternal-fetal tolerance that may have an impact on the tumor microenvironment (TME), thus contributing to cancer progression in this setting. Immunomodulatory cells, such as regulatory T cells (Tregs) and myeloid-derived suppressor cells (MDSCs), recruited by TME, are capable of inhibiting anti-tumor immune responses. The current study was designed to explore potential contribution of the pregnancy milieu to induction of immunosuppressive microenvironment supporting lymphoma growth and progression.

Methods: A20 murine lymphoma B cells were injected subcutaneously to pregnant and non-pregnant female wild type Balb/C mice. The tumor volume was measured twice weekly and a multiple comparison was done with Bonferroni correction. Following delivery, mice from both groups were sacrificed and tumor cells were harvested. RNA was extracted from tumors of 5 control and 6 pregnant mice and RNAseq data were generated. Then, statistical analysis was performed for differentially expressed genes in these two groups. Tregs (CD4⁺CD25⁺) and MDSCs (CD11b⁺Gr1⁺) within tumors were estimated by FACS analysis.

Results: Pregnant mice demonstrated significantly accelerated tumor growth compared to non-pregnant animals. The tumor volume in mice sacrificed after parturition reached 752.3 ± 492.7 mm³ versus 499.3 ± 373.5 mm³ in the control group (n = 41 and 29, respectively; p<0.05). Similarly, a significant intergroup difference in tumor weight at time of sacrifice was observed (1.73 ± 0.94 vs. 1.3 ± 0.65 g; n = 32 and 22, respectively; p=0.05). Gene expression profiling of tumor samples revealed a significant difference between the groups in the expression pattern of 180 out of 25,000 examined genes. Eighty-four of these genes can be categorized according to specific cellular pathways into those associated with T cell differentiation, cytokine-cytokine receptor interaction and adhesion molecules, all potentially involved in immunosuppressive cell migration to TME. Examination for the presence of inhibitory cell subpopulations in TME did not demonstrate significant difference in percentage of Tregs between pregnant and non-pregnant mice (1.18% ± 1.35 vs. 1.15% ± 1.09; n = 16 & 13, respectively). However, pregnant mice exhibited a greater granulocytic MDSC subpopulation (CD11b⁺Gr1^high) within the tumor compared with their control counterparts (0.41% ± 0.3 vs. 0.22% ± 0.13; p<0.05; n = 17 & 14, respectively). To evaluate whether this finding was related to pregnancy alone or to a cumulative effect of tumor and pregnancy, spleen peripheral blood mononuclear cells (PBMCs) were immunophenotyped for these markers. The results obtained showed that the expansion of the CD11b⁺Gr1⁺ subpopulation in pregnant mice was limited to the TME only, while spleen PBMCs failed to exhibit this effect (2% ± 2 in pregnant mice vs. 1.88% ± 1.16 in the controls; n = 8 & 6, respectively). A parallel analysis of spleen samples for Treg percentage also did not reveal a difference between the two groups (0.27% ± 0.07 vs. 0.25% ± 0.09, n = 8 & 6, respectively).

Conclusions: Pregnant mice demonstrated accelerated tumor growth following lymphoma cell inoculation. The tumor gene expression profile in pregnant mice is likely to reflect the cellular pathways that could be associated with differentiation and recruitment of immunosuppressive cells to the TME. In this regard, MDSCs emerge as potential candidates contributing to pregnancy-associated lymphoma progression. Further studies are warranted to define the kinetics of this cell subpopulation and identify biological effects of MDSCs in such setting.