Chemotherapy resistance in acute myeloid leukemia is associated with decreased anti-tumor immune response through MHC molecule and B7 family members

Abstract

Acute myeloid leukemia (AML) remains challenging due to chemotherapeutic drug-resistance (CDR). Aberrant expression B7 family proteins are involved in tumors evasion. We wonder whether B7 family protein alteration in AML CDR further supports tumor escape. Here, we establish AML cytarabine-resistant cell line U937/Ara-C and report on the expression MHC molecule and B7 family member. HLA-ABC was highly expressed similarly on both cell lines. MIC (MHC class I chain related) A/B and B7-H6 was moderately expressed on the surface of U937 and decreased dramatically by U937/Ara-C. In contrast, enhanced expression of B7-H1 and B7-H7 by U937/Ara-C was observed. HLA-DR and other B7 family members including CD80, CD86, B7-DC, B7-H2, B7-H3, B7-H4, and B7-H5 were not detected by both cell lines. Compared co-cultured with U937, peripheral blood mononuclear cells showed a decreased cytotoxicity when incubated with U937/Ara-C, as indicated by decreased levels of granzyme B and perforin production, accompanied with less TNF-α and lactate dehydrogenase secretion. In conclusion, AML CDR further evades the anti-tumor immune response which may through MHC molecule and B7 family members.

Keywords: AML; B7 family; Chemotherapeutic drug-resistant; MHC molecules.

Fig. 1

Growth curve and doubling time of U937 and U937/Ara-C. U937 and U937/Ara-C were seeded at the same density, and cells were counted on day 1, day 2, and day 3. Growth curve (A) and doubling time (B) were determined. A representative result of at least two experiments is shown, and the data are the mean ± SD of triplicate determinations. **P < 0.01, U937/Ara-C was compared with U937 under similar conditions

Fig. 2

Cytarabine sensitivity of U937 and U937/Ara-C. U937 and U937/Ara-C were cultured without (A) or with (B-D) Cytarabine at the indicated concentration. The absorbency was measured and OD value was detected at different time point. A representative result of at least two experiments is shown, and the data are the mean ± SD of triplicate determinations. *P < 0.05, ****P < 0.0001, U937/Ara-C was compared with U937 under similar conditions

Fig. 3

Cell cycle distributions of U937 and U937/Ara-C as detected by flow cytometry. The cell cycle diagrams of U937 (A) and U937/Ara-C (B). The total population of cell cycle proportion (C) in G0/G1 phase (D), S phase (E), and G2/M phase (F) were compared respectively between U937 and U937/Ara-C. A representative result of at least three experiments is shown, and the data for (C–F) are the mean ± SD of triplicate determinations. *P < 0.05, ****P < 0.0001, U937/Ara-C was compared with U937 under similar conditions

Fig. 4

MHC molecule and B7 family member expression on the surface of U937 and U937/Ara-C as detected by flow cytometry. U937 and U937/Ara-C were harvested for immunofluorescence staining and flow cytometry. The Black in histograms denotes fluorescence when stained with the PE-conjugated antibody to the indicated antigen, and the gray line represents fluorescence when stained with the isotype control mAb. Images are representative of at least three experiments

Fig. 5

Cytotoxic induction in U937 and U937/Ara-C by PBMC. PBMC were incubated with U937 or U937/Ara-C at an E:T ratio of 10:1 for 24 h, and the supernatant was collected. Then, the levels of secreted perforin (A), granzyme B (B), TNF-α (C), and LDH (D) in the supernatant were measured with specific ELISA kits. A representative result of at least three experiments is shown, and the data are the mean ± SD of triplicate determinations. **P < 0.01, ***P < 0.001, PBMC-U937/Ara-C coculture was compared with PBMC-U937 coculture under similar conditions

Fig. 6

The schematic outline of AML chemotherapeutic drug-resistance further evades the anti-tumor immune response