新型細菌細胞膜標靶藥物SC5005與特定天然產物對抗甲氧苯青黴素金黄色葡萄球菌展現協同殺菌效果

Abstract

細菌的抗藥性演化已造成全球公共衛生上巨大的挑戰。根據統計，抗甲氧苯青黴素之金黃色葡萄球菌的感染每年約造成11,300的美國人死亡。此外，此細菌也是人類皮膚感染的主要致病菌。由於此細菌具有對抗多數抗生素之能力，導致治療上非常棘手，因此發展新穎的抗葡萄球菌藥物是迫切需要的。我們過去的研究中證明一個小分子藥物SC5005對革蘭氏陽性菌展現良好抗菌能力，且會影響細菌細胞膜的完整性。在這裡我們進一步發現低濃度SC5005會增加細菌細胞膜的通透性，且在抑菌濃度會明顯造成細胞膜去極化的現象，進而導致大規模的細菌細胞膜破損。根據小鼠皮膚感染實驗的結果顯示，1 %的SC5005與臨床用藥的效果一樣好，都可有效降低皮膚組織內的細菌數量。 另外，先前的研究發現SC5005在有特定天然物(NP7)的存在下， 其殺菌力可加強達8,000倍以上。故我們也進一步去探討SC5005及特定天然物之間對於金黃色葡萄球菌的協同殺菌的效果以及其協同作用機制。Time- Kill Assay結果顯示，SC5005與NP7的合併可以在極短的時間內便殺死試管中所有的MRSA和懸浮狀態的休眠細菌。然而，NP7並不會加強SC5005對於人類上皮細胞的毒性，證明此NP7有很高的特異性去加強SC5005的抗菌能力。後續實驗也顯示，細菌經過SC5005的作用後可增加對於NP7的感受性，且這個藥物組合的協同抗菌能力也會隨著抗氧化劑濃度的提高而受到抑制。這表示NP7在與SC5005合併殺菌的機制上很有可能扮演氧化劑的角色。另外， NP7和SC5005的合併也會對金黃色葡萄球菌的細胞膜去極化有協同性的增強效果。以上結果顯示該組合在對MRSA的協同殺菌作用中具有多模式的作用機制。 上述實驗結果指出，SC5005可以有效降低生物體內細菌數目，且與NP7有高度協同殺菌的能力。若多加以研究及探討，這個藥物組合很有潛力成為臨床上抗甲氧苯青黴素之金黃色葡萄球菌和革蘭氏陽性細菌的皮膚感染的新藥。

The emergence of antibiotic resistance has imposed a serious challenge to public health worldwide. In the United States, methicillin-resistant Staphylococcus aureus (MRSA) infections claim nearly 11,300 lives each year. Besides, MRSA also is the leading cause of skin infections in the United States. As conventional antibiotics are not able to treat MRSA infections, a new antibiotic with a novel action mechanism is highly needed. Previously, our team identified a small-molecule compound, SC5005, exhibiting potent activity against Gram-positives, including clinical isolated MRSAs. At present study, we aimed to further investigate SC5005’s mode of action. Our results indicated that SC5005 at low concentration can specifically increase permeability of S. aureus membrane followed by causing membrane depolarization at 1 × MIC, leading to a complete membrane disruption at higher concentration. Furthermore, topical treatment of 1 % SC5005 was as effective as commercial antibiotic in eradicating MRSAs in skin infection mouse model. Previously, in looking for a vehicle for SC5005’s delivery, we unexpectedly identified a nature product (NP7), which dramatically potentiated SC5005’s antibacterial activity for more than 8,000 ×. Here, we further uncovered the synergistic bactericidal mechanism of this combination and assessed their efficacy in treating MRSA skin infection. In the Time-Kill assays, this combination displayed a fast-killing activity against MRSA and planktonic persister cells as demonstrated by a complete eradication of bacteria within 3 minutes and 2 hours, respectively. In addition, NP7 had no effect on SC5005’s cytotoxicity toward human epithelial cells, indicating its synergistic effect is specific for SC5005’s antibacterial activity. Subsequent results showed that short exposure of SC5005 increased MRSA susceptibility to NP7, and the synergistic effect of this combination can be reduced by antioxidants in a dose-dependent manner, indicating NP7’s pro-oxidant activity played a crucial role in the synergistic bactericidal effect with SC5005. Also, NP7 can synergistically enhanced depolarization of S. aureus’ membrane in combined with SC5005. Above findings indicated that this combination displayed multimodal mechanism of action in the synergistic bactericidal effect against MRSA. Overall, SC5005 exhibited a non-inferior efficacy to commercial antibiotic ointment in skin-infection mouse model. As NP7 and SC5005 showed a highly synergistic effect on killing bacteria, this combination could be further developed as a topical medication for skin infections of MRSA and other Gram-positives.