Association of Multi-Drug Resistance-1 (MDR1) Gene Polymorphism with Leukocytopenia in Breast Cancer Patients treated with Chemotherapy

Abstract It has been well established that γδ T-cells play a role in innate anti-tumor immunity. However, the exact mechanism has not yet been fully elucidated. Most of these responses have been ascribed to Vγ9Vδ2 cells, which represent a major subset of the circulating γδ T-cells in humans (1–10%). IFN-γ and granzyme B are important molecules in the anti-tumor immune responses. Upon stimulation, γδ T-cells rapidly produce IFN-γ and cytotoxic molecules. In the present study we analyzed the immune responses by γδ T-cells in 30 newly diagnosed breast cancer patients and 30 normal controls before and after expansion with zoledronic acid. We also scanned the granzyme B gene polymorphism in breast cancer patients and controls. Our result revealed that γδ-T cells in PBMC were reduced in both frequency and function in breast cancer patients compared with the normal controls. Ex-vivo stimulation of γδ T-cells with zoledronic acid and IL-2 partially compensated for this deficiency, as it stimulates production of IFN- γ and release of cytotoxic molecules by these cells. However, the IFN- γ and granzyme B and cytotoxicity of the expanded γδ T-cells from breast cancer patients remained significantly below normal control. Genotypic analysis of granzyme B gene revealed significantly higher frequency of the RAH haplotype in breast cancer patients compared with normal controls. The prevalence of the wild genotype QPY/QPY was significantly higher in normal controls compared with the breast cancer patients. Cytotoxicity by γδ T-cells against various targets was reduced in breast cancer patients compared to normal controls. In conclusion, our analysis shows a defective immune function of γδ T-cells and granzyme B gene polymorphism in breast cancer patients. The γδ T-cell function defect in these patients can be partially corrected by zoledronic acid. Further studies of γδ T-cell function and granzyme B gene polymorphism in other cancers, as well as the therapeutic use of zoldedronic acid is warranted.

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ELOVL2: a novel tumor suppressor attenuating tamoxifen resistance in breast cancer

10.21203/rs.3.rs-69357/v1 ◽

2020 ◽

Author(s):

Dawoon Jeong ◽

Juyeon Ham ◽

Hyeon Woo Kim ◽

Heejoo Kim ◽

Hwee Won Ji ◽

...

Keyword(s):

Breast Cancer ◽

Drug Resistance ◽

Cancer Patients ◽

Tumor Tissue ◽

Tamoxifen Resistance ◽

Colony Formation Assay ◽

Breast Cancer Patients ◽

Methylation Analysis ◽

Genome Wide ◽

Mcf 7

Abstract Background To comprehensively understand the molecular mechanism of tamoxifen resistance (TamR) acquisition by epigenetically regulated genes, it is essential to identify pivotal genes by genome-wide methylation analysis and verify their function in xenograft animal model and cancer patients. Methods The MCF-7/TamR breast cancer cell line was developed and a genome-wide methylation array was performed. The methylation and expression of ELOVL2 was validated in cultured cells, xenografted tumor tissue, and breast cancer patients by methylation-specific PCR, qRT-PCR, Western blot analysis, and immunohistochemistry. Deregulation of ELOVL2 and THEM4 was achieved using siRNA or generating stable transfectants. Tam sensitivity, cell growth, and apoptosis were monitored by colorimetric and colony formation assay and flow cytometric analysis. Pathway analysis was performed to generate networks for the differentially methylated genes in the MCF-7/TamR cells and for the differentially expressed genes in the ELOVL2-overexpressing cells. Results Genome-wide methylation analysis in the MCF-7/TamR cells identified elongation of very-long chain fatty acid protein 2 (ELOVL2) to be significantly hypermethylated and downregulated, which was further verified in the tumor tissues from TamR breast cancer patients (n = 28) compared with those from Tam-sensitive (TamS) patients (n = 33) (P < 0.001). Immunohistochemical analysis of tissues from cancer patients showed lower expression of ELOVL2 in the TamR than TamS tissues. Growth of the MCF-7/TamR cells overexpressing ELOVL2 was retarded in cell culture and also in xenograft tumor tissue. Strikingly, ELOVL2 attenuated resistance to Tam up to 70% judged by the colorimetric and colony formation assay and xenograft mouse model. ELOVL2 contributed to the recovery of Tam sensitivity by regulating a group of genes in the AKT and ERα signaling pathways, e.g., THEM4, which plays crucial roles in drug resistance. Conclusions ELOVL2 was hypermethylated and downregulated in TamR breast cancer patients compared with TamS patients. ELOVL2 is responsible for the recovery of Tam sensitivity. AKT- and ERα-hubbed networks are pivotal in ELOVL2 signaling, where THEM4 contributes to the relaying ELOVL2 signaling. This study implies that deregulation of a gene in fatty acid metabolism can lead to drug resistance, giving insight into the development of a new therapeutic strategy for drug-resistant breast cancer.

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