CN106309415A - 卷柏素类化合物在作为抗真菌药物增效剂中的应用 - Google Patents
卷柏素类化合物在作为抗真菌药物增效剂中的应用 Download PDFInfo
- Publication number
- CN106309415A CN106309415A CN201610700369.2A CN201610700369A CN106309415A CN 106309415 A CN106309415 A CN 106309415A CN 201610700369 A CN201610700369 A CN 201610700369A CN 106309415 A CN106309415 A CN 106309415A
- Authority
- CN
- China
- Prior art keywords
- selaginellin
- drug
- antifungal
- application
- medicine
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Granted
Links
- SJSFYXIEVFIZJC-UHFFFAOYSA-N 4-[[3-(hydroxymethyl)-6-(4-hydroxyphenyl)-2-[2-(4-hydroxyphenyl)ethynyl]phenyl]-(4-hydroxyphenyl)methylidene]cyclohexa-2,5-dien-1-one Chemical class C1=CC(=O)C=CC1=C(C=1C=CC(O)=CC=1)C1=C(C#CC=2C=CC(O)=CC=2)C(CO)=CC=C1C1=CC=C(O)C=C1 SJSFYXIEVFIZJC-UHFFFAOYSA-N 0.000 title claims abstract description 39
- 239000003429 antifungal agent Substances 0.000 title claims abstract description 25
- 239000003814 drug Substances 0.000 claims abstract description 44
- QVEOXPUIBXVBOI-UHFFFAOYSA-N selaginellin Natural products OCc1ccc(c2ccccc2)c(C(=C3C=CC(=O)C=C3)c4ccc(O)cc4)c1C#Cc5ccc(O)cc5 QVEOXPUIBXVBOI-UHFFFAOYSA-N 0.000 claims abstract description 38
- XHNYXAAEIYBJSL-UHFFFAOYSA-N selaginellin A Natural products Oc1ccc(cc1)C#Cc2cccc(c3ccccc3)c2C(=C4C=CC(=O)C=C4)c5ccc(O)cc5 XHNYXAAEIYBJSL-UHFFFAOYSA-N 0.000 claims abstract description 30
- HNNJWBQSRKGCFY-UHFFFAOYSA-N 4-[[6-(4-hydroxyphenyl)-2-[2-(4-hydroxyphenyl)ethynyl]-3-methylphenyl]-phenylmethylidene]cyclohexa-2,5-dien-1-one Chemical compound C1=CC(=O)C=CC1=C(C=1C=CC=CC=1)C1=C(C#CC=2C=CC(O)=CC=2)C(C)=CC=C1C1=CC=C(O)C=C1 HNNJWBQSRKGCFY-UHFFFAOYSA-N 0.000 claims abstract description 28
- 229940121375 antifungal agent Drugs 0.000 claims abstract description 25
- 230000000843 anti-fungal effect Effects 0.000 claims abstract description 11
- 150000003852 triazoles Chemical class 0.000 claims description 13
- -1 chlorins compound Chemical class 0.000 claims description 8
- 150000002460 imidazoles Chemical class 0.000 claims description 3
- RFHAOTPXVQNOHP-UHFFFAOYSA-N fluconazole Chemical compound C1=NC=NN1CC(C=1C(=CC(F)=CC=1)F)(O)CN1C=NC=N1 RFHAOTPXVQNOHP-UHFFFAOYSA-N 0.000 abstract description 24
- KAESVJOAVNADME-UHFFFAOYSA-N Pyrrole Chemical compound C=1C=CNC=1 KAESVJOAVNADME-UHFFFAOYSA-N 0.000 abstract description 20
- 229940079593 drug Drugs 0.000 abstract description 19
- 230000000694 effects Effects 0.000 abstract description 13
- 108090000623 proteins and genes Proteins 0.000 abstract description 12
- XMAYWYJOQHXEEK-OZXSUGGESA-N (2R,4S)-ketoconazole Chemical compound C1CN(C(=O)C)CCN1C(C=C1)=CC=C1OC[C@@H]1O[C@@](CN2C=NC=C2)(C=2C(=CC(Cl)=CC=2)Cl)OC1 XMAYWYJOQHXEEK-OZXSUGGESA-N 0.000 abstract description 11
- 229960004125 ketoconazole Drugs 0.000 abstract description 11
- 206010059866 Drug resistance Diseases 0.000 abstract description 8
- 230000002441 reversible effect Effects 0.000 abstract description 6
- 238000011529 RT qPCR Methods 0.000 abstract description 4
- 238000002474 experimental method Methods 0.000 abstract description 4
- 231100000331 toxic Toxicity 0.000 abstract description 3
- 230000002588 toxic effect Effects 0.000 abstract description 3
- 230000002195 synergetic effect Effects 0.000 abstract description 2
- 229960004884 fluconazole Drugs 0.000 abstract 1
- 238000003753 real-time PCR Methods 0.000 abstract 1
- 230000003827 upregulation Effects 0.000 abstract 1
- 241000222122 Candida albicans Species 0.000 description 12
- 241000233866 Fungi Species 0.000 description 9
- 238000000034 method Methods 0.000 description 9
- 230000014509 gene expression Effects 0.000 description 8
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 8
- 241000894006 Bacteria Species 0.000 description 7
- 241000222173 Candida parapsilosis Species 0.000 description 6
- IAZDPXIOMUYVGZ-UHFFFAOYSA-N Dimethylsulphoxide Chemical compound CS(C)=O IAZDPXIOMUYVGZ-UHFFFAOYSA-N 0.000 description 6
- 101150106008 ERG11 gene Proteins 0.000 description 6
- 101150098080 ERG5 gene Proteins 0.000 description 6
- 101100173965 Candida albicans (strain SC5314 / ATCC MYA-2876) FLU1 gene Proteins 0.000 description 5
- 102100035361 Cerebellar degeneration-related protein 2 Human genes 0.000 description 5
- 101000737796 Homo sapiens Cerebellar degeneration-related protein 2 Proteins 0.000 description 5
- 108010047230 Member 1 Subfamily B ATP Binding Cassette Transporter Proteins 0.000 description 5
- 229940095731 candida albicans Drugs 0.000 description 5
- 102100035360 Cerebellar degeneration-related antigen 1 Human genes 0.000 description 4
- 101000737793 Homo sapiens Cerebellar degeneration-related antigen 1 Proteins 0.000 description 4
- 102100030306 TBC1 domain family member 9 Human genes 0.000 description 4
- 238000001514 detection method Methods 0.000 description 4
- 239000012153 distilled water Substances 0.000 description 4
- 238000005516 engineering process Methods 0.000 description 4
- 239000001963 growth medium Substances 0.000 description 4
- 206010007134 Candida infections Diseases 0.000 description 3
- 208000031888 Mycoses Diseases 0.000 description 3
- 230000001580 bacterial effect Effects 0.000 description 3
- 201000003984 candidiasis Diseases 0.000 description 3
- 239000012531 culture fluid Substances 0.000 description 3
- 239000000890 drug combination Substances 0.000 description 3
- 238000009413 insulation Methods 0.000 description 3
- 238000002156 mixing Methods 0.000 description 3
- 238000002360 preparation method Methods 0.000 description 3
- 102000004169 proteins and genes Human genes 0.000 description 3
- 230000001105 regulatory effect Effects 0.000 description 3
- 230000035945 sensitivity Effects 0.000 description 3
- 239000000243 solution Substances 0.000 description 3
- 239000006228 supernatant Substances 0.000 description 3
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 2
- YCKRFDGAMUMZLT-UHFFFAOYSA-N Fluorine atom Chemical compound [F] YCKRFDGAMUMZLT-UHFFFAOYSA-N 0.000 description 2
- 206010017533 Fungal infection Diseases 0.000 description 2
- 101000991061 Homo sapiens MHC class I polypeptide-related sequence B Proteins 0.000 description 2
- 101000615488 Homo sapiens Methyl-CpG-binding domain protein 2 Proteins 0.000 description 2
- 108020003285 Isocitrate lyase Proteins 0.000 description 2
- KFZMGEQAYNKOFK-UHFFFAOYSA-N Isopropanol Chemical compound CC(C)O KFZMGEQAYNKOFK-UHFFFAOYSA-N 0.000 description 2
- 102100030301 MHC class I polypeptide-related sequence A Human genes 0.000 description 2
- 102100030300 MHC class I polypeptide-related sequence B Human genes 0.000 description 2
- 102100021299 Methyl-CpG-binding domain protein 2 Human genes 0.000 description 2
- 101710163270 Nuclease Proteins 0.000 description 2
- 239000001888 Peptone Substances 0.000 description 2
- 108010080698 Peptones Proteins 0.000 description 2
- 150000003851 azoles Chemical class 0.000 description 2
- 210000000170 cell membrane Anatomy 0.000 description 2
- 230000008859 change Effects 0.000 description 2
- 150000001875 compounds Chemical class 0.000 description 2
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 2
- 230000000857 drug effect Effects 0.000 description 2
- 229910052731 fluorine Inorganic materials 0.000 description 2
- 239000011737 fluorine Substances 0.000 description 2
- 230000036541 health Effects 0.000 description 2
- 230000006872 improvement Effects 0.000 description 2
- 239000004615 ingredient Substances 0.000 description 2
- 239000007788 liquid Substances 0.000 description 2
- 238000012986 modification Methods 0.000 description 2
- 230000004048 modification Effects 0.000 description 2
- 235000019319 peptone Nutrition 0.000 description 2
- 238000004321 preservation Methods 0.000 description 2
- 238000012360 testing method Methods 0.000 description 2
- OILXMJHPFNGGTO-UHFFFAOYSA-N (22E)-(24xi)-24-methylcholesta-5,22-dien-3beta-ol Natural products C1C=C2CC(O)CCC2(C)C2C1C1CCC(C(C)C=CC(C)C(C)C)C1(C)CC2 OILXMJHPFNGGTO-UHFFFAOYSA-N 0.000 description 1
- RQOCXCFLRBRBCS-UHFFFAOYSA-N (22E)-cholesta-5,7,22-trien-3beta-ol Natural products C1C(O)CCC2(C)C(CCC3(C(C(C)C=CCC(C)C)CCC33)C)C3=CC=C21 RQOCXCFLRBRBCS-UHFFFAOYSA-N 0.000 description 1
- 238000005160 1H NMR spectroscopy Methods 0.000 description 1
- FRXSZNDVFUDTIR-UHFFFAOYSA-N 6-methoxy-1,2,3,4-tetrahydroquinoline Chemical compound N1CCCC2=CC(OC)=CC=C21 FRXSZNDVFUDTIR-UHFFFAOYSA-N 0.000 description 1
- OQMZNAMGEHIHNN-UHFFFAOYSA-N 7-Dehydrostigmasterol Natural products C1C(O)CCC2(C)C(CCC3(C(C(C)C=CC(CC)C(C)C)CCC33)C)C3=CC=C21 OQMZNAMGEHIHNN-UHFFFAOYSA-N 0.000 description 1
- 229920001817 Agar Polymers 0.000 description 1
- 241000004633 Bulbine glauca Species 0.000 description 1
- 108010078791 Carrier Proteins Proteins 0.000 description 1
- 102000014914 Carrier Proteins Human genes 0.000 description 1
- 206010011409 Cross infection Diseases 0.000 description 1
- 108090000790 Enzymes Proteins 0.000 description 1
- 102000004190 Enzymes Human genes 0.000 description 1
- DNVPQKQSNYMLRS-NXVQYWJNSA-N Ergosterol Natural products CC(C)[C@@H](C)C=C[C@H](C)[C@H]1CC[C@H]2C3=CC=C4C[C@@H](O)CC[C@]4(C)[C@@H]3CC[C@]12C DNVPQKQSNYMLRS-NXVQYWJNSA-N 0.000 description 1
- 102100031181 Glyceraldehyde-3-phosphate dehydrogenase Human genes 0.000 description 1
- 238000005481 NMR spectroscopy Methods 0.000 description 1
- 102000006382 Ribonucleases Human genes 0.000 description 1
- 108010083644 Ribonucleases Proteins 0.000 description 1
- 241000195974 Selaginella Species 0.000 description 1
- 239000008272 agar Substances 0.000 description 1
- 238000000137 annealing Methods 0.000 description 1
- 230000008485 antagonism Effects 0.000 description 1
- 230000000844 anti-bacterial effect Effects 0.000 description 1
- 230000001716 anti-fugal effect Effects 0.000 description 1
- 230000003064 anti-oxidating effect Effects 0.000 description 1
- 229940088710 antibiotic agent Drugs 0.000 description 1
- 239000008346 aqueous phase Substances 0.000 description 1
- 238000004500 asepsis Methods 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 230000008901 benefit Effects 0.000 description 1
- 230000001851 biosynthetic effect Effects 0.000 description 1
- 230000015572 biosynthetic process Effects 0.000 description 1
- 239000008280 blood Substances 0.000 description 1
- 210000004369 blood Anatomy 0.000 description 1
- 230000037396 body weight Effects 0.000 description 1
- 229940041514 candida albicans extract Drugs 0.000 description 1
- 229940055022 candida parapsilosis Drugs 0.000 description 1
- 239000003153 chemical reaction reagent Substances 0.000 description 1
- 230000002301 combined effect Effects 0.000 description 1
- 230000007812 deficiency Effects 0.000 description 1
- 230000007850 degeneration Effects 0.000 description 1
- 238000013461 design Methods 0.000 description 1
- 238000011161 development Methods 0.000 description 1
- 238000010586 diagram Methods 0.000 description 1
- 238000009792 diffusion process Methods 0.000 description 1
- 238000010790 dilution Methods 0.000 description 1
- 239000012895 dilution Substances 0.000 description 1
- 201000010099 disease Diseases 0.000 description 1
- 238000004090 dissolution Methods 0.000 description 1
- 238000002330 electrospray ionisation mass spectrometry Methods 0.000 description 1
- DNVPQKQSNYMLRS-SOWFXMKYSA-N ergosterol Chemical compound C1[C@@H](O)CC[C@]2(C)[C@H](CC[C@]3([C@H]([C@H](C)/C=C/[C@@H](C)C(C)C)CC[C@H]33)C)C3=CC=C21 DNVPQKQSNYMLRS-SOWFXMKYSA-N 0.000 description 1
- 230000008686 ergosterol biosynthesis Effects 0.000 description 1
- 239000000284 extract Substances 0.000 description 1
- 238000000605 extraction Methods 0.000 description 1
- JFUIHGAGFMFNRD-UHFFFAOYSA-N fica Chemical compound FC1=CC=C2NC(C(=O)NCCS)=CC2=C1 JFUIHGAGFMFNRD-UHFFFAOYSA-N 0.000 description 1
- 239000012530 fluid Substances 0.000 description 1
- 230000002538 fungal effect Effects 0.000 description 1
- 244000053095 fungal pathogen Species 0.000 description 1
- 108020004445 glyceraldehyde-3-phosphate dehydrogenase Proteins 0.000 description 1
- 230000002401 inhibitory effect Effects 0.000 description 1
- 238000007689 inspection Methods 0.000 description 1
- 230000007154 intracellular accumulation Effects 0.000 description 1
- 238000011835 investigation Methods 0.000 description 1
- 101150044508 key gene Proteins 0.000 description 1
- 210000004185 liver Anatomy 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 230000007246 mechanism Effects 0.000 description 1
- 239000002609 medium Substances 0.000 description 1
- 239000010445 mica Substances 0.000 description 1
- 229910052618 mica group Inorganic materials 0.000 description 1
- IDSXLJLXYMLSJM-UHFFFAOYSA-N morpholine;propane-1-sulfonic acid Chemical compound C1COCCN1.CCCS(O)(=O)=O IDSXLJLXYMLSJM-UHFFFAOYSA-N 0.000 description 1
- 230000036457 multidrug resistance Effects 0.000 description 1
- 230000035772 mutation Effects 0.000 description 1
- 230000000324 neuroprotective effect Effects 0.000 description 1
- 239000002547 new drug Substances 0.000 description 1
- 238000010606 normalization Methods 0.000 description 1
- 230000002018 overexpression Effects 0.000 description 1
- 239000012071 phase Substances 0.000 description 1
- 239000002504 physiological saline solution Substances 0.000 description 1
- 239000002244 precipitate Substances 0.000 description 1
- 230000008569 process Effects 0.000 description 1
- 239000011535 reaction buffer Substances 0.000 description 1
- 230000004044 response Effects 0.000 description 1
- 238000010839 reverse transcription Methods 0.000 description 1
- 239000003161 ribonuclease inhibitor Substances 0.000 description 1
- 238000000926 separation method Methods 0.000 description 1
- 210000002966 serum Anatomy 0.000 description 1
- 230000001954 sterilising effect Effects 0.000 description 1
- 238000004659 sterilization and disinfection Methods 0.000 description 1
- 238000005728 strengthening Methods 0.000 description 1
- 239000000126 substance Substances 0.000 description 1
- 238000003786 synthesis reaction Methods 0.000 description 1
- 238000002560 therapeutic procedure Methods 0.000 description 1
- 238000001262 western blot Methods 0.000 description 1
- 239000012138 yeast extract Substances 0.000 description 1
Classifications
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K31/00—Medicinal preparations containing organic active ingredients
- A61K31/12—Ketones
- A61K31/122—Ketones having the oxygen directly attached to a ring, e.g. quinones, vitamin K1, anthralin
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K31/00—Medicinal preparations containing organic active ingredients
- A61K31/33—Heterocyclic compounds
- A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
- A61K31/41—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having five-membered rings with two or more ring hetero atoms, at least one of which being nitrogen, e.g. tetrazole
- A61K31/4196—1,2,4-Triazoles
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K31/00—Medicinal preparations containing organic active ingredients
- A61K31/33—Heterocyclic compounds
- A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
- A61K31/495—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with two or more nitrogen atoms as the only ring heteroatoms, e.g. piperazine or tetrazines
- A61K31/496—Non-condensed piperazines containing further heterocyclic rings, e.g. rifampin, thiothixene or sparfloxacin
Landscapes
- Health & Medical Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Medicinal Chemistry (AREA)
- Pharmacology & Pharmacy (AREA)
- Epidemiology (AREA)
- Life Sciences & Earth Sciences (AREA)
- Animal Behavior & Ethology (AREA)
- General Health & Medical Sciences (AREA)
- Public Health (AREA)
- Veterinary Medicine (AREA)
- Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
Abstract
本发明公开了selaginellin、selaginellin A等卷柏素类化合物作为抗真菌增效剂的用途。实验结果表明，卷柏素selaginellin、selaginellin A与氟康唑、酮康唑等抗真菌药物合用时，可达到协同增效的作用，可降低唑类药物的用药剂量，从而减轻药物的毒副作用。并且通过实时荧光定量PCR(qRT‑PCR)实验表明，卷柏素selaginellin、selaginellin A可显著逆转唑类药物耐药基因的上调，有逆转唑类药物耐药的作用，具有重要的临床应用价值。
Description
技术领域
本发明涉及一种抗真菌增效剂，具体涉及selaginellin或selaginellin A等卷柏素类化合物在作为抗真菌药物增效剂中的应用，属医药技术领域。
背景技术
近年来，临床真菌感染的发病率和死亡率逐年上升。念珠菌是人体重要的条件致病性真菌，是引起医院感染性疾病主要病原菌之一。唑类药物是临床上常用的能有效治疗深部及浅表部真菌感染的抗真菌药，如咪唑类中的酮康唑；三唑类中的氟康唑等。但是，随着药物长期、大量使用，使得真菌对唑类药物的敏感性降低，且耐药性显著增高，导致不得不加大药物剂量，某些抗真菌药物甚至加大用药量也无效。目前通行的做法为、加快新药研发、联合用药。加大药物剂量的危害是显而易见的，比如副作用增大、加速真菌耐药、
增加社会经济负担等等；如酮康唑等对肝脏或心脏有毒副作用的药物，从而限制了这些药物在临床上的应用。寻找唑类药物的增效剂日益受到重视，已成为抗真菌治疗的主要发展方向之一。所谓增效剂是指对受试生物无药效或药效很低，但与某种药物混合使用时，能明显提高该药物效果的辅助化合物
卷柏素为从卷柏属植物中提取的系列有效成分。具有抗微生、抗氧化、抗衰老，及神经保护等活性，但未见其作为抗真菌药物增效剂与抗菌药物联合使用治疗真菌病的报道。
其中代表性成分selaginellin及selaginellin A的结构式如下；
发明内容
发明目的：本发明所要解决的技术问题是克服现有技术的不足，对selaginellin及selaginellin A的活性进行深入研究，提供selaginellin或selaginellin A卷柏素类化合物在作为抗真菌药物增效剂中的应用。
技术方案：为解决以上技术问题，本发明采取如下技术方案：
卷柏素类化合物在作为抗真菌药物增效剂中的应用
selaginellin或selaginellin A在作为抗真菌药物增效剂中的应用。
作为优选方案，selaginellin或selaginellin A与唑类抗真菌药物合用在制备抗真菌药物中的应用。
selaginellin或selaginellin A与唑类抗真菌药物合用在制备抗真菌药物中的应用，作为优选方案，所述的selaginellin或selaginellin A在总药物中的浓度为0.5～256μg/mL。
selaginellin或selaginellin A与唑类抗真菌药物合用在制备抗真菌药物中的应用，作为优选方案，所述的selaginellin或selaginellin A在总药物中的浓度为2～64μg/mL。
本发明所述的selaginellin或selaginellin A与唑类抗真菌药物合用在制备抗真菌药物中的应用，所述的唑类抗真菌药物为咪唑类或三唑类抗真菌药物。如酮康唑或氟康唑。所述的真菌为念珠菌。
唑类药物发挥抗真菌作用主要是抑制了与细胞膜麦角甾醇合成有关的酶，如:由ERG11编码的14α-羊毛甾醇-去甲基化酶(14-DM)和由ERG5编码的Δ222去饱和酶。ERG11、ERG5发生突变或过表达可以降低真菌对唑类药物的敏感性，并产生耐药。真菌外排作用的增加，导致药物在细胞内蓄积减少，从而降低了菌株对药物的敏感性，因此编码外排蛋白的基因过度表达是真菌对唑类药物耐药的主要机制之一。目前已知与耐药有关的外排蛋白包括含ABC转运蛋白超家族(ABCT)的CDR1、CDR2，多药耐药转运蛋白MDR1，以及易化扩散超家族转运蛋白FLU1。本研究表明白念珠菌暴露于FLC后，ERG11、ERG5、CDR1、CDR2、MDR1与FLU1表达上调，而卷柏素与氟康唑合用可以显著逆转这一趋势，说明两药联用，有可能逆转耐药。
有益效果：本发明和现有技术相比具有如下优点：
本发明通过大量实验深入研究selaginellin或selaginellin A的活性，实验结果表明白念珠菌暴露于氟康唑后，ERG11、ERG5、CDR1、CDR2、MDR1与FLU1表达上调，而卷柏素selaginellin或selaginellin A与氟康唑合用可以显著逆转这一趋势，表明两药联用，可逆转耐药。selaginellin和selaginellin A可用作抗真菌药物的增效剂，从而可减轻药物的毒副作用，特别是氟康唑、酮康唑；并有可能逆转真菌耐药，有效治疗真菌感染，具有重要的临床应用价值。
附图说明
图1为selaginellin或selaginellin A联合氟康唑对白念株菌相关基因表达相对变化差异的结果图。
图2为selaginellin或selaginellin A联合氟康唑对白念株菌相关基因表达水平变化趋势图。
具体实施方式
下面结合具体实施例，进一步阐明本发明，应理解这些实施例仅用于说明本发明而不用于限制本发明的范围，在阅读了本发明之后，本领域技术人员对本发明的各种等价形式的修改均落于本申请所附权利要求所限定的范围。
实施例1 Selaginellin和氟康唑合用对不同临床真菌株的作用
一、材料和方法
试药：
selaginellin及selaginellin A卷柏素类化合物：实验室自制，使用1H-NMR,13C-NMR及ESI-MS对其进行了对其进行结构解析，与文献对照确定其化学结构。经HPLC-UV面积归一化法计算，纯度均在98％以上。氟康唑(FLC)，酮康唑(KEC)：辉瑞制药有限公司。
菌株：
分别为美国模式培养物集存库(American Type Culture Collection,ATCC)标准菌株白念珠菌(Candida albicans)ATCC90028，近平滑念珠菌(C.parapsilosis)ATCC22019，白念珠菌临床株(62342)。。
培养液：
RMPI-1640培养基：RMPI-1640(Gibco公司)10.0g，吗啡啉丙磺酸(Sigma)34.5g，加双蒸水900mL溶解，定容置1000mL，滤过消毒，-20℃保存，使用前检查培养基是否无菌。
SDA培养基：蛋白胨10g，琼脂15g，加双蒸水1000mL，高压灭菌，4℃备用。
YEPD培养液：酵母浸膏10g，蛋白胨20g，加双蒸水1000mL，高压灭菌，4℃备用。
菌液制备：
念珠菌菌液：从4℃保存的SDA培养基上挑取菌少量，接种至1mL YEPD培养液，于30℃以100r/min振荡培养活化，使真菌处于指数生长期。吸取菌液至适量无菌生理盐水中，经血球计数板计数，孢子数约为1.0×106～5.0×106cfu/ml。用倍量RMPI-1640液体培养基稀释成菌1.0×103cfu/ml。
药液的配制：
氟康唑以灭菌蒸馏水配成2560μg·mL-1，selaginellin、selaginellinA以DMSO配成药物原液(浓度为2560μg·mL-1)，置于-20℃冰箱冻存。实验前再用灭菌蒸馏水依次进行倍比稀释。联合药敏孔内药物终浓度范围如下，FLC:16～0.03μg·mL-1，KEC：4～0.008μg·mL-1，selaginellin：128～2μg·mL-1；selaginellinA：64～1μg·mL-1。96孔平板孔内DMSO终浓度≤1％，对菌株生长的影响可忽略不计。于96孔板中每孔加入100μL不同浓度的药液备用，注意避光。
MIC值判定：
100％MIC：以溶媒对照为参照，肉眼观察药基孔中无菌生长为最低抑菌浓度，100％MIC值。
联合药敏试验：
参照M27-A3方案。将制备完毕的平板置于恒温培养箱中，35℃培养48h。(n＝3)。
以FICI法评价唑类药物与selaginellin、selaginellinA的体外联合效果，其公式为∑FIC＝FICA+FICB＝MICAB/MICA+MICBA/MICB，式中的MICA和MICB分别表示A和B单用时的MIC值，MICAB和MICBA分别表示两种药物联用时各自的MIC值。FICI≤0.5为协同作用，FICI＞4为拮抗作用，0.5＜FICI≤4为无关作用。
实验分组：
qRT-PCR和Western Blot实验各指标检测均为4个组别：空白对照组，FLC单用组(0.25μg·mL-1)，selaginellin、selaginellinA单用组(64μg·mL-1)，两药联用组(0.25μg·mL-1 FLC+64μg·mL-1selaginellin)。
qRT-PCR检测相关基因的表达水平
提取总RNA：将上述菌体于-80℃冻干过夜。用1mL Total RNA Extractor重悬，震荡混匀，在4℃12000r/min×10min离心；上清液加入200μL氯仿，震荡15s，离心；抽取上层无色水相加入等体积异丙醇，混匀，离心；去除上清，加入1ml 70％的乙醇，离心；吸尽上清，加入30μL的无RNase水溶解RNA沉淀，待完全溶解后于-70℃保存。
引物设计与合成：从NCBI基因库中查得所需基因序列，并用Primer 5软件设计所需引物，委托南京金斯瑞科技有限公司合成引物。
逆转录：每个组份轻轻混匀，然后2000rpm离心20s；取灭过菌且无核酸酶的0.2mlPCR管，依次加入如下组份：RNA(2μg)1μL，Oligo dT(18)(50μM)2μL，无核酸酶的双蒸水至总体积12.5μL；65℃保温5min，冰浴5min；依次加入如下组份：RNase抑制剂(40u/μL)0.5μL，5×Reaction Buffer 4.0μL，dNTPs(10mM)2.0uL，M-MuLV 1.0μL；混匀后，2000rpm离心20s；42℃保温1h，70℃保温10min，置冰上5min。
qRT-PCR检测目的基因表达：反应程序：95℃×15s(变性)，60℃×20s(退火)，72℃×15s(延伸)。以GAPDH作为内参标准，一个样本基因做3个复孔，结果应用软件GraphpadPrism 5.0分析，基因水平采用倍数变化来表示(2(△△Ct)法)。
实验结果：
1)以棋盘法检测两类药物合用后对C.albicans ATCC90028和临床分离株62342，以及C.parapsilosis ATCC22019的MIC值(见表1、2)，计算对应的FICI指数。结果显示，selaginellin、selaginellin A与FLC、KEC对C.albicans ATCC90028和临床株62342，C.parapsilosis ATCC22019显示出较好的协同增效作用。
表1.棋盘法检测selaginellin与FLC、KEC联用对C.albicans、C.parapsilosis的MIC(μg·mL-1)
SEL表示selaginellin化合物。
表2.棋盘法检测selaginellin A与FLC、KEC联用对C.albicans、C.parapsilosis的MIC(μg·mL-1)
SELA表示selaginellin A化合物。
2)qRT-PCR考察SEL和FLC联用后耐药相关基因的表达差异
采用qRT-PCR技术检测了真菌细胞膜麦角固醇生物合成的关键基因ERG5、ERG11，外排泵基因CDR1、CDR2、MDR1、FLU1的表达水平。结果显示，单用FLC显著上调ERG5、ERG11、CDR1、CDR2、MDR1、FLU1的表达，单用selaginellin、selaginellinA也有一定的上调作用，但氟康唑(FLC)与selaginellin、selaginellinA合用后，selaginellin、selaginellinA可逆转氟康唑(FLC)引起的上述基因的表达上调，分别下调52％、86％、71％、62％、67％、56％、36％(如图1和图2)。从基因表达方面证实selaginellin、selaginellinA可降低真菌对氟康唑的耐药性，取得了非常好的预料不到的技术效果。
以上所述仅是本发明的优选实施方式，应当指出，对于本技术领域的普通技术人员来说，在不脱离本发明原理的前提下，还可以做出若干改进和润饰，这些改进和润饰也应视为本发明的保护范围。
Claims (6)
1.卷柏素类化合物在作为抗真菌药物增效剂中的应用。
2.selaginellin或selaginellin A在作为抗真菌药物增效剂中的应用。
3.selaginellin或selaginellin A与唑类抗真菌药物合用在制备抗真菌药物中的应用。
4.根据权利要求2所述的selaginellin或selaginellin A与唑类抗真菌药物合用在制备抗真菌药物中的应用，其特征在于，selaginellin或selaginellin A在总药物中的浓度为0.5～256μg/mL。
5.根据权利要求4所述的selaginellin或selaginellin A与唑类抗真菌药物合用在制备抗真菌药物中的应用，其特征在于，selaginellin或selaginellin A在总药物中的浓度为2～64μg/mL。
6.根据权利要求3所述的selaginellin或selaginellin A与唑类抗真菌药物合用在制备抗真菌药物中的应用，其特征在于，所述的唑类抗真菌药物为咪唑类或三唑类抗真菌药物。
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201610700369.2A CN106309415B (zh) | 2016-08-22 | 2016-08-22 | 卷柏素类化合物在作为抗真菌药物增效剂中的应用 |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201610700369.2A CN106309415B (zh) | 2016-08-22 | 2016-08-22 | 卷柏素类化合物在作为抗真菌药物增效剂中的应用 |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN106309415A true CN106309415A (zh) | 2017-01-11 |
| CN106309415B CN106309415B (zh) | 2019-08-20 |
Family
ID=57742605
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201610700369.2A Expired - Fee Related CN106309415B (zh) | 2016-08-22 | 2016-08-22 | 卷柏素类化合物在作为抗真菌药物增效剂中的应用 |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN106309415B (zh) |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN114814057A (zh) * | 2022-04-28 | 2022-07-29 | 湖南省药品检验检测研究院 | 一种非靶向代谢组学区分卷柏品种真伪的方法及应用 |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101711779A (zh) * | 2008-10-08 | 2010-05-26 | 中南大学 | 垫状卷柏提取物在制备抗阿尔茨海默病药物中的用途 |
| CN102838464A (zh) * | 2011-06-24 | 2012-12-26 | 沈阳药科大学 | 一种新的抗癌活性化合物selaginellin N及其用途 |
-
2016
- 2016-08-22 CN CN201610700369.2A patent/CN106309415B/zh not_active Expired - Fee Related
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101711779A (zh) * | 2008-10-08 | 2010-05-26 | 中南大学 | 垫状卷柏提取物在制备抗阿尔茨海默病药物中的用途 |
| CN102838464A (zh) * | 2011-06-24 | 2012-12-26 | 沈阳药科大学 | 一种新的抗癌活性化合物selaginellin N及其用途 |
Non-Patent Citations (5)
| Title |
|---|
| CAO YUAN，ET AL.: "Cytotoxic Constituents from Selaginella tamariscina", 《NAT PROD RES DEV》 * |
| XIAN-LONG CHENG, ET AL.: "Selaginellin A and B, Two Novel Natural Pigments Isolated from Selaginella tamariscina", 《CHEM. PHARM. BULL》 * |
| XIN LIU, ET AL.: "Selaginpulvilins A−D, New Phosphodiesterase‑4 Inhibitors with an Unprecedented Skeleton from Selaginella pulvinata", 《ORGANIC LETTERS》 * |
| YUAN CAO, ET AL.: "Antimicrobial selaginellin derivatives from Selaginella pulvinata", 《BIOORGANIC & MEDICINAL CHEMISTRY LETTERS》 * |
| 俞森洋等主编: "《呼吸内科主治医生660问》", 30 April 2009, 中国协和医科大学出版社 * |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN114814057A (zh) * | 2022-04-28 | 2022-07-29 | 湖南省药品检验检测研究院 | 一种非靶向代谢组学区分卷柏品种真伪的方法及应用 |
| CN114814057B (zh) * | 2022-04-28 | 2024-01-16 | 湖南省药品检验检测研究院 | 一种非靶向代谢组学区分卷柏品种真伪的方法及应用 |
Also Published As
| Publication number | Publication date |
|---|---|
| CN106309415B (zh) | 2019-08-20 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| Pulcrano et al. | Ambroxol influences voriconazole resistance of Candida parapsilosis biofilm | |
| Dogra et al. | Mucormycosis amid COVID-19 crisis: pathogenesis, diagnosis, and novel treatment strategies to combat the spread | |
| CN105193808A (zh) | Bdsf与伊曲康唑联用对临床耐药白念珠菌的协同药效 | |
| CN102106851B (zh) | 鸦胆苦醇作为化疗药增效剂的应用 | |
| CN101301289B (zh) | 小檗碱及其结构类似物在逆转多药耐药泵中的应用 | |
| Su et al. | Antifungal activity and potential mechanism of 6, 7, 4′-O-triacetylscutellarein combined with fluconazole against drug-resistant C. albicans | |
| CN115120602A (zh) | 一种抗真菌药物增效剂 | |
| An et al. | Synergistic Effect of the Combination of Deferoxamine and Fluconazole In Vitro and In Vivo against Fluconazole-Resistant Candida Spp. | |
| Shin et al. | Pichia ohmeri fungemia associated with phlebitis: successful treatment with amphotericin B | |
| CN106309415A (zh) | 卷柏素类化合物在作为抗真菌药物增效剂中的应用 | |
| CN109646447B (zh) | 重楼皂苷在抗糠秕马拉色菌和白假丝酵母菌制品中的应用 | |
| CN109820887A (zh) | 具有治疗脚气作用的组合物及其应用 | |
| CN110755456A (zh) | 美洲大蠊提取物与抗真菌药物组成的抗真菌药物组合物 | |
| CN104188962A (zh) | 厚朴酚与唑类药物在制备抗真菌联合用药物中的应用 | |
| CN110859950B (zh) | 一种用于白念珠菌棘白菌素类药物耐药菌的药物组合物及其应用 | |
| CN103040852B (zh) | 赖氨酸作为增效剂在制备抗真菌药物中的应用 | |
| McGinnis et al. | Fatal cerebritis caused by a new species of Cladophialophora | |
| CN104474550A (zh) | 一种杀灭耐药烟曲霉的药物组合物及其方法 | |
| CN101816646B (zh) | 茶黄素用作抗真菌药物增效剂的用途 | |
| CN110548029B (zh) | 甲磺酸奥希替尼在制备治疗真菌感染药物中的应用 | |
| CN108186617A (zh) | 香叶醇及其衍生物在制备mrsa感染性疾病药物中的新用途 | |
| EGBE | Antifungal effects of Nigella sativa L.(Black cumin) seed extracts and seed oil on selected Candida albicans strains | |
| CN104069128A (zh) | Pvp修饰的纳米银与唑类药物在制备抗耐药真菌活性联合用药物中的应用 | |
| Jia et al. | In Vitro and In Vivo Evaluation of Synergistic Effects of Everolimus in Combination with Antifungal Agents on Exophiala dermatitidis | |
| CN101816666A (zh) | 连翘酯苷用作抗真菌药物增效剂的用途 |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| GR01 | Patent grant | ||
| GR01 | Patent grant | ||
| CF01 | Termination of patent right due to non-payment of annual fee |
Granted publication date: 20190820 Termination date: 20210822 |
|
| CF01 | Termination of patent right due to non-payment of annual fee |