A new method for evaluating the risk of transferring leukemic cells with transplanted cryopreserved ovarian tissue

Abstract

Purpose: This study aimed to develop a method to detect ovarian residual disease by multicolor flow cytometry in acute leukemia patients.

Methods: We designed an experimental model consisting in adding acute leukemia cells to a cell suspension obtained from healthy ovarian cortex. Leukemic cell detection within the ovarian cell suspension required the development of a specific myeloid antibody panel different from that commonly used for minimal residual disease (MRD) monitoring in bone marrow. The method was then used to detect ovarian residual disease in 11 acute leukemia patients.

Results: Multicolor flow cytometry is able to evaluate the presence of viable leukemic cells in the ovarian cortex with good specificity and robust sensitivity of 10-4. We observed a good correlation between multicolor flow cytometry and quantitative polymerase chain reaction results. Ovarian residual disease detection by multicolor flow cytometry was positive in 3 out of 11 acute leukemia patients.

Conclusion: Multicolor flow cytometry can potentially be applied to ovarian tissue from all acute leukemia patients and is essential to evaluate the risk of cancer re-seeding before autograft of ovarian tissue in case of acute leukemia.

Figures

Fig. 1

a–c Serial dilutions of leukemic cells among ovarian cell suspensions detected by MFC and RT-qPCR. a Modelization results for ALL (in green), AML with hematological panel (in red), and AML with adapted panel (in blue). The X-axis represents the theoretical values of MRD dilutions and the Y-axis the experimental MRD values of the same samples quantified by multicolor flow cytometry. For each dilution level, the standard deviation is represented by a vertical bar. Axes are in log scale. b Quantification of AML cells in an ovarian cell dilution experiment. Only living nucleated cells (SYTO13+/7-AAD−) which are CD45low are represented. A MRD-positive event must be contained in gates P3 (CD11b vs CD33), P4 (CD11b vs CD43), and P5 (CD15 vs CD33) (represented in black). Living nucleated ovarian cells are indicated in pink. c Correlation of MRD levels detected by MFC and RT-qPCR. The correlation between BCR-ABL1 transcripts and MFC is represented in green and between NPM1 variant A mutations and MFC in blue. Axes are in log scale. MFC multicolor flow cytometry, RT-qPCR real-time quantitative PCR, ALL acute lymphoblastic leukemia, AML acute myeloid leukemia, MRD minimal residual disease

Fig. 2

Schematic representation of ORD results obtained by multicolor flow cytometry and quantitative PCR for 11 acute leukemia patients