Mitochondrial Transporter ABCB10 Protects Against Doxorubicin-Induced Respiratory Muscle Dysfunction Independent of Changes to Diaphragm Accumulation.

BACKGROUND: Doxorubicin (DOX) is a highly effective chemotherapeutic agent whose use can cause respiratory toxicity, increasing patient fatigue and negatively impacting quality of life and survival. These adverse effects occur due to diaphragm muscle mitochondrial accumulation of DOX, where it causes reactive oxygen species production and iron dysregulation. ABCB10 is a mitochondria-localized ATP-binding cassette transporter hypothesized to play a role in the maintenance of mitochondrial redox balance and iron homeostasis, and potentially the mitochondrial export of DOX. This study investigated potential therapeutic effects of ABCB10 to prevent DOX-induced respiratory muscle dysfunction. METHODS: DOX respiratory muscle toxicity was modelled in rats using both single (20 mg/kg, once) and multicycle (5.7 mg/kg, 3 cycles) administration. The effects of overexpression and knockdown of ABCB10 on DOX-induced diaphragm dysfunction, mitochondrial DOX accumulation and markers of mitochondrial iron homeostasis were evaluated via administration of rAAV9-MHCK7-ABCB10 or an antisense oligonucleotide targeting ABCB10, respectively. RESULTS: ABCB10 significantly improved diaphragm rate of fatigue (138.2 ± 11.66 s vs. 104.6 ± 8.79 s in DOX), specific force production (22.12 ± 0.70 N/cm2 vs. 18.31 ± 1.65 N/cm2 at 160 Hz in DOX) and fibre area (Type I: 1309.05 ± 56.86 μm2 vs. 1027.04 ± 50.53 μm2 in DOX; Type IIa: 1389.13 ± 47.72 μm2 vs. 1027.04 ± 50.07 μm2 in DOX; Type IIb/x: 2590.81 ± 103.21 μm2 vs. 2302.13 ± 138.62 μm2 in DOX) following a single injection of DOX. These improvements did not occur as a result of ABCB10-induced efflux of DOX (1006.03 ± 214.30 pg/μg vs. 1008.69 ± 195.62 pg/μg in DOX) but were associated with reduced mitochondrial iron (0.17 ± 0.02 nmol/mg vs. 0.23 ± 0.02 nmol/mg in DOX). The beneficial effects on diaphragm rate of fatigue (136.5 ± 11.93 s vs. 121.5 ± 9.47 s in DOX), specific force production (23.36 ± 1.40 N/cm2 vs. 19.26 ± 1.21 N/cm2 at 160 Hz in DOX) and fibre area (Type I: 1124.68 ± 63.02 μm2 vs. 914.57 ± 63.09 μm2 in DOX; Type IIa: 1244.67 ± 106.18 μm2 vs. 950.02 ± 62.38 μm2 in DOX; Type IIb/x: 2548.37 ± 235.69 μm2 vs. 2222.17 ± 234.61 μm2 in DOX) were also present in rats that received multiple cycles of DOX. Diaphragm rescue with ABCB10 was attendant with reduced mitoferrin 1 gene expression (+1.62 ± 0.47 fold vs. +3.85 ± 0.99 fold in DOX), preservation of mitochondrial function and a reduction in markers of heme synthesis, including Fech (+0.78 ± 0.17 fold vs. +0.24 ± 0.09 fold in DOX), ALAS1 (+0.54 ± 0.18 fold vs. +0.19 ± 0.04 fold in DOX), ALAS2 (+0.68 ± 0.23 fold vs. +0.33 ± 0.11 fold in DOX) and CPOX (+0.84 ± 0.16 fold vs. +0.38 ± 0.09 fold in DOX). While increasing ABCB10 in the diaphragm prevented DOX respiratory toxicity, reducing its expression did not exacerbate diaphragm dysfunction or mitochondrial DOX and iron accumulation. CONCLUSIONS: These results suggest that ABCB10 can preserve mitochondrial and diaphragm muscle function following DOX treatment by regulating iron redox-cycling and heme synthesis, independent of changes to DOX accumulation.