KR20160101944A - 알파-MSH의 ClpB 단백질 모방을 통한 식욕의 조절에 대한 세균 영향 - Google Patents
알파-MSH의 ClpB 단백질 모방을 통한 식욕의 조절에 대한 세균 영향 Download PDFInfo
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Abstract
Description
도면 1: 대장균 (E. coli) K12 단백질 및 α-MSH 사이에 분자 모방의 프로테오믹 확인.
(a) 대장균 (E. coli) 세포질 단백질의 2D GE. (b, c) α-MSH로 전흡착된 (c) 또는 전흡착되지 않은 (b) Rb 항-α-MSH IgG로 검출된 대장균 (E. coli) 단백질의 면역블롯. 적색의 원은 단백질 확인에 이용되었던 α-MSH IgG에 의해 특이적으로 인식된 스팟을 둘러싼다. 청색의 원은 비특이적 스팟을 지시한다. 스팟 1-4에서 확인된 단백질은 ClpB의 동종형이다. (d) Stretcher 프로그램을 이용한 α-MSH 및 ClpB 아미노산 서열 정렬. (e) 항-α-MSH IgG로 드러난 재조합 ClpB의 웨스턴 블롯. 레인 1 및 2, 각각 20 및 10 μg의 ClpB.
도면 2. 생쥐에서 ClpB 면역화.
ClpB-면역화된 생쥐 (ClpB+Adj)는 어쥬번트 (Adj), PBS 또는 대조 (Ctr)를 제공받는 생쥐와 비교되었다. (a) 연구의 32 일 동안 체중 변화. 식품 섭취 및 급식 패턴은 BioDAQ 케이지에서 최종 2 주 동안 연구되었다. 연구의 최종 10 일 동안 평균 일일 식품 섭취 (b), 식사 크기 (c) 및 식사 횟수 (d). (e) α-MSH (100 μg kg- 1 체중, i.p.) 또는 PBS의 주사 후 24 시간 동안 식품 섭취. (f) 10-6M α-MSH로 흡착 전후에 ClpB-반응성 IgG의 혈장 수준. (g) 해리 평형 상수 (KD 값)로서 도시된 항-ClpB IgG의 친화성. (h) α-MSH-반응성 전체 IgG의 혈장 수준. (i) 항-α-MSH IgG의 친화성 (KD). (j) 단독으로, 또는 ClpB-면역화된 생쥐로부터 또는 Adj-주사된 생쥐로부터 모아진 IgG (0.5 mg ml-1)와 함께, α-MSH에 의한 자극 후 MC4R을 과다발현하는 인간 배아 신장-293 세포에서 cAMP 검정. (k) cAMP 검정은 항-α-MSH IgG로부터 고갈된 IgG로 수행되었다. (a) α-MSH 주사 (100 μg kg-1 체중, i.p.) 전에 2-원 반복된 계측 분산 분석 (ANOVA), Po0.0001, 본페로니 사후 검증 *a 최소한, Po0.05 ClpB 군 대 Ctr; *b 최소한, Po0.05 Adj 군 대 Ctr.; *c, Po0.05, 스튜던트 t 검증 ClpB 군 대 PBS; 및 *d, Po0.05, 스튜던트 t 검증 ClpB 군 대 Ctr. (b) ANOVA P=0.0002, Tukey 사후 검증 ***Po0.001, **Po0.01, #Po0.05, 스튜던트 t 검증. (c) ANOVA P=0.007, Tukey 사후 검증 **Po0.01. (e) 스튜던트 t 검증, *Po0.05. (f, g) ANOVA Po0.0001, Tukey 사후 검증 ***Po0.001 ClpB +Adj 대 다른 군, 짝비교 t 검증 ##Po0.01, ###Po0.001. (h) ANOVA P =0.0002, Tukey 사후 검증 ***Po0.001, *Po0.05; (i) 크러스칼 왈리스 검증 P =0.003, 던 사후 검증 **Po0.01, (평균 ±s.e.m., n=8). (j) ANOVA P= 0.005, Tukey 사후 검증 *Po0.05; ANOVA P =0.04, 스튜던트 t 검증 #Po0.05, aClpB 대 α-MSH, bClpB 대 Adj. (평균 ± s.e.m.; j, n =6, k, n=3).
도면 3: 생쥐에서 대장균 (E. coli) 보충.
생쥐에서 체중 (a), 식품 섭취 (b), 식사 크기 (c) 및 식사 횟수 (d)에 대한 대장균 (E. coli) K12 야생형 (WT), ClpBdeficient (△ClpB) 대장균 (E. coli) K12 또는 LB 배지로 위내 일일 위관영양 (1-21 일자)의 효과. (e) 세균 ClpB DNA의 180-염기쌍 단편의 PCR 검출, 첫 번째 레인, 분자량 마커, 두 번째 레인 대장균 (E. coli) K12 WT의 시험관내 배양액으로부터 DNA, 세 번째 레인 대장균 (E. coli) K12 △ClpB의 시험관내 배양액으로부터 DNA, 그리고 나머지 레인 21 일자에 수집된 생쥐 대변으로부터 DNA. 10-6M α-MSH로 흡착 전후에 항-ClpB IgM (f) 및 IgG (g)의 효소 결합 면역흡착 검정에서 흡광도에서 혈장 수준.
항-α-MSH IgM (h) 및 IgG (i)의 혈장 수준. (j) 항-α-MSH IgG의 친화성 (평형 상수). (a) 2-원 반복된 계측 분산 분석 (ANOVA), P =0.3, 2 일자 본페로니 사후 검증, **Po0.01 대조 (Ctr) 대 대장균 (E. coli) WT. (b) 1-2 일자 ANOVA, P =0.0006, Tukey 사후 검증 ***Po0.001, *Po0.05, 대장균 (E. coli) WT 대 aCtr 및 bLB. (c) 세 번째 주 크러스칼 왈리스 (K-W) 검증 P=0.0001, 던 사후 검증, ***Po0.001, **Po0.01, 대장균 (E. coli) WT 대 aCtr, bLB 및 C△ClpB. (d) 1-2 일자 ANOVA, P =0.006, Tukey 사후 검증 **Po0.01, *Po0.05, 세 번째 주 K-W 시험 Po0.0001, 던 사후 검증, ***Po0.001, **Po0.01, 대장균 (E. coli) WT 대 aCtr, bLB 및 C△ClpB. (f) 흡착 전 K-W 시험 P =0.02, 던 사후 검증 *Po0.05, 흡착 후 ANOVA, Po0.0001, Tukey 사후 검증 **Po0.01, 대장균 (E. coli) WT 대 다른 군. (g) 흡착 전 ANOVA, P=0.01, Tukey 사후 검증 *Po0.05, 대장균 (E. coli) WT 대 다른 군, 짝비교 t 검증 ##Po0.01. (h) 스튜던트 t 검증, 대장균 (E. coli) WT 대 다른 군 *Po0.05. (j) K-W 시험 P =0.02, 던 사후 검증 *Po0.05, 만-휘트니 검증, #Po0.05. (평균 ± s.e.m., n=8).
도면 4. ED 환자에서 항-ClpB 항체.
건강한 여성 (대조, Ctr)에서 및 AN, BN 및 BED를 앓는 환자에서 항-ClpB IgG (a) 및 IgM (b)의 혈장 수준. 10-6M α-MSH로 흡착 전후에 ClpB IgG (c) 및 IgM (d)의 혈장 수준. α-MSH 교차반응성 항-ClpB IgG (e) 및 IgM (f)의 백분율. (b) 스튜던트 t 검증 *Po0.05. (c, d) 짝비교 t 검증, ***Po0.001, **Po0.01. (e) 크러스칼 왈리스 검증 Po0.0001, 던 사후 검증, **Po0.01, 만-휘트니 검증 #Po0.05. (f) 분산 분석 P =0.02, Tukey 사후 검증 *Po0.05. (평균 ±s.e.m., Ctr, n =65, AN, n=27 BN, n =32 및 BED, n=14).
도면 5. 식사 장애를 앓는 환자 및 건강한 대조 (Ctr)에서 세균 ClpB 단백질의 혈장 농도.
AN, 신경성 식욕부진, BN, 신경성 폭식증, BED, 폭식 장애. *p<0.05 평균 ClpB 농도 대 Ctr의 스튜던트 t 검증. 대조의 평균+2 표준 편차 (SD)보다 높은 ClpB 농도를 갖는 환자의 백분율 (%).
ClpB IgG (대조) | 성장 공포 r= -0.31 * |
충동 조절 r= -0.26 * |
사회적 불안정 r= -0.26 * |
ClpB IgG (AN) | 신체 불만족 r= 0.4 * |
날씬해지려는 욕구 r= 0.35 * |
완벽주의 r= 0.38 * |
ClpB IgM (AN) | 무력함 r= -0.42 * |
대인간 불신 r= -0.58 ** |
사회적 불안정 무쾌감증 r= -0.52 ** r= -0.35 * |
ClpB IgM (BED) | 폭식증 r= 0.53 * |
완벽주의 r= 0.6 * |
연령 r= -0.74 ** |
Claims (13)
- 식사 장애의 치료 또는 예방에서 이용을 위한 최소한 하나의 ClpB 단백질 발현 세균에 대해 지향된 최소한 하나의 항균제를 포함하는 조성물.
- 개체에서 식욕을 조절하는 비-치료 방법에 있어서, 상기 방법은 최소한 하나의 ClpB 단백질 발현 세균에 대해 지향된 최소한 하나의 항균제를 포함하는 조성물의 효과량을 상기 개체에 투여하는 것을 포함하는 것을 특징으로 하는 방법.
- 개체에서 식사 장애를 치료하거나 또는 이들 장애의 발생의 기회를 감소시키는 방법에 있어서, 상기 방법은 최소한 하나의 ClpB 발현 세균에 대해 지향된 최소한 하나의 항균제를 포함하는 조성물을 치료가 필요한 개체에 투여하는 것을 포함하는 것을 특징으로 하는 방법.
- 식사 장애의 치료 또는 예방에서 이용을 위해, ClpB 단백질을 발현하지 않는 프로바이오틱스를 포함하는 조성물.
- 개체에서 식욕을 조절하는 비-치료 방법에 있어서, 상기 방법은 ClpB 단백질을 발현하지 않는 프로바이오틱스를 포함하는 조성물의 효과량을 상기 개체에 투여하는 것을 포함하는 것을 특징으로 하는 방법.
- 백신으로서 또는 면역원성 조성물로서 이용을 위한 ClpB 단백질을 포함하는 조성물.
- 식사 장애에 대항하여 면역화를 위해 청구항 5에 따라 이용되는 조성물.
- 식사 장애를 예방하기 위해 청구항 5 또는 6에 따라 이용되는 조성물.
- 개체에서 식사 장애를 진단하기 위한 시험관내 방법에 있어서, 상기 방법은 다음을 포함하는 것을 특징으로 하는 시험관내 방법:
a) 상기 개체로부터 생물학적 표본에서 ClpB 단백질 및/또는 항-ClpB 항체의 수준을 계측하고;
b) ClpB 단백질 및/또는 항-ClpB 항체의 상기 계측된 수준을 참조값에 비교하고; 그리고
c) 개체가 식사 장애를 겪는 지를 추론함. - 식사 장애로 고통받는 개체를, ClpB 단백질 및/또는 항-ClpB 항체의 수준을 감소시키는 치료에 반응할 개연성이 있는 것으로 선별하는 시험관내 방법에 있어서, 상기 방법은 다음을 포함하는 것을 특징으로 하는 시험관내 방법:
a) 상기 개체로부터 생물학적 표본에서 ClpB 단백질 및/또는 항-ClpB 항체의 수준을 계측하고;
b) ClpB 단백질 및/또는 항-ClpB 항체의 상기 계측된 수준을 참조값에 비교하고; 그리고
c) ClpB 단백질 및/또는 항-ClpB 항체의 수준을 감소시키는 치료에 대해 개체를 선별하고, 여기서 ClpB 단백질 및/또는 항-ClpB 항체의 수준을 감소시키는 상기 치료는 최소한 하나의 ClpB 발현 세균에 대해 지향된 한 항균제를 포함하는 조성물의 효과량을 상기 개체에 투여하고 및/또는 ClpB를 발현하지 않는 프로바이오틱스 및/또는 이들의 조합을 포함하는 조성물의 효과량을 상기 개체에 투여함. - 청구항 9 내지 10 중 어느 한 항에 있어서, 단계 a)에서 ClpB 단백질의 수준은 상기 개체의 대변에서 ClpB DNA를 정량함으로써 계측되는 것을 특징으로 하는 시험관내 방법.
- 식사 장애가 신경성 식욕부진 (AN), 신경성 폭식증 (BN), 폭식 장애 (BED), 과식, 과식증, 소모병, 예를 들면, 악액질로 구성된 군에서 선택되는 것을 특징으로 하는, 청구항 1, 4 또는 6 내지 8 중 어느 한 항에 따라 이용되는 조성물, 청구항 2 또는 5에 따른 비-치료 방법 또는 청구항 9 내지 10 중 어느 한 항에 따른 시험관내 방법.
- 비만의 치료 또는 예방에서 이용을 위한 ClpB 단백질을 과다발현하는 프로바이오틱스를 포함하는 조성물.
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WO1997003359A1 (en) * | 1995-07-07 | 1997-01-30 | Oravax, Inc. | Helicobacter clpb |
JP2009524640A (ja) * | 2006-01-27 | 2009-07-02 | ダニスコ エー/エス | 肥満及びそれに関連する疾患の治療及び予防のためのプロバイオティック微生物の使用 |
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Also Published As
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CA2932488A1 (en) | 2015-06-11 |
HRP20241279T1 (hr) | 2024-12-06 |
EP3076985B1 (en) | 2024-07-24 |
EP3076985C0 (en) | 2024-07-24 |
CN106061491B (zh) | 2021-01-08 |
US20160313348A1 (en) | 2016-10-27 |
KR102332078B1 (ko) | 2021-11-26 |
PL3076985T3 (pl) | 2024-12-16 |
JP2019089815A (ja) | 2019-06-13 |
JP2017503768A (ja) | 2017-02-02 |
JP6876732B2 (ja) | 2021-05-26 |
WO2015082633A1 (en) | 2015-06-11 |
EP3076985A1 (en) | 2016-10-12 |
CN106061491A (zh) | 2016-10-26 |
ES2994101T3 (en) | 2025-01-17 |
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