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Methods and compositions for treating inflammatory skin disease with recombinant microorganisms

A technology for recombining microorganisms and compositions, applied in skin diseases, drug combinations, chemical instruments and methods, etc.

Pending Publication Date: 2020-06-12
AZITRA INC
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Therefore, given the enormous burden that atopic dermatitis places on our healthcare system, there is a large unmet need

Method used

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  • Methods and compositions for treating inflammatory skin disease with recombinant microorganisms
  • Methods and compositions for treating inflammatory skin disease with recombinant microorganisms
  • Methods and compositions for treating inflammatory skin disease with recombinant microorganisms

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0145] Example 1: Development of a nucleic acid construct that encodes a protein that can be exported from S. epidermidis cells and then imported into human keratinocytes

[0146] In one embodiment, the present invention describes the production of recombinant S. epidermidis strains capable of secreting heterologous proteins, thus overcoming the intractability of genetic modification of S. epidermidis. Previous functional gene analyzes of the common skin colonizers S. aureus and S. epidermidis have been limited by the presence of type I and type IV restriction systems in nearly all strains of these bacteria. These restriction systems recognize methylated cytosine bases in DNA from standard clonal expansion systems such as DH10B E. coli. However, several constructs have been created in S. epidermidis strain ATCC12228 using the methylation-deficient E. coli strain DC10B 5, the strain is a commensal non-pathogenic isolate that lacks the ica operon. Thus, the present invention...

Embodiment 2

[0151] Example 2: Determination of Persistence and Localization of Topically Applied Staphylococcus Epidermidis Using an In Vitro Model System

[0152] Materials and methods

[0153] Build reporter bacteria

[0154] To facilitate tracking of topically applied bacteria, a S. epidermidis (SE) strain expressing sGFP was used. remove SecA and RMR peptide so that sGFP protein does not shuttle into the secretion system and free sGFP does not penetrate the stratum corneum. This construct is called SE-sGFP .

[0155] Quantify and compare the growth characteristics of transformed bacteria in liquid media

[0156] A basic understanding of the ability of transformed (recombinant) S. epidermidis to compete with wild-type S. epidermidis is required. To understand the growth characteristics of transformed bacteria as well as the growth kinetics of recombinant protein producing bacteria, colony forming units (CFU) in liquid media were quantified using standard techniques. Staphylo...

Embodiment 3

[0164] Example 3: Characterization of delivery of bacterially secreted sGFP to skin using an in vitro model system

[0165] Characterization of sGFP production in SE

[0166] Characterization of delivery of bulk purified sGFP and sGFP+RMR to RHE

[0167] Data on the localization of purified sGFP and sGFP+RMR will help understand: (i) whether sGFP+RMR penetrates the cuticle; (ii) if so, how deep penetration can be detected; and (iii) Penetration dynamics. Here, 5.0 μg / μL of GFP + / - RMR was applied at time points 0, 2, 6, 12, 18, and 24 h to determine the effect of dose on breakthrough and time. The results showed that GFP was detected as deep as the epidermal-dermal junction within 30 minutes of application.

[0168] Table of cellular compartments and penetration depths of in situ secreted sGFP proteins from SE-GFP reporter and control strains sign

[0169] The goals of this assay are similar to those described above, but with the key difference that the protein is ...

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Abstract

The present invention provides isolated plasmids, recombinant microorganisms, kits, and methods for the treatment of inflammatory skin disease.

Description

[0001] related application [0002] This application claims priority to US Provisional Application 62 / 554,271, filed September 5, 2017, and US Provisional Application 62 / 685,687, filed June 15, 2018, the entire contents of both of which are incorporated herein by reference in their entirety. [0003] Background of the invention [0004] Atopic dermatitis (AD) or eczema is a chronic, pruritic, inflammatory skin disease that affects 5-20% of children worldwide (Williams, H. et al. . J Allergy Clin. Immunol. 1999;103(1 Pt 1):125-138), and is also prevalent in many adults. The prevalence of atopic dermatitis is increasing, with some form affecting 11% of the U.S. population (Shaw, T.E. et al. . J Invest Dermatol. 2011;131(1):67-73), or about 35 million people, causing direct losses of $5 billion in the US alone. The main feature of the disease is dry, scaly, itchy skin. Despite the worldwide prevalence of atopic dermatitis and its high disease burden, few targeted and effectiv...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K35/74A61K38/17A61K9/00A61P17/00
CPCA61K9/0014A61K38/00A61K35/00C07K14/4713C07K2319/02C07K2319/10C12N1/20C12N15/74A61K38/1709A61P17/00A61K35/74A61K38/179C07K14/71
Inventor T.M.惠特菲尔
Owner AZITRA INC
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