Genetically modified tumor-targeted bacteria with reduced virulence

Abstract

The present invention is directed to mutant Salmonella sp. having a genetically modified msbB gene in which the mutant Salmonella is capable of targeting solid tumors. The invention is also directed to Salmonella sp. containing a genetically modified msbB gene as well as an genetic modification in a biosynthetic pathway gene such as the purl gene. The present invention further relates to the therapeutic use of the mutant Salmonella for growth inhibition and / or reduction in volume of solid tumors.

Description

[0001] This application is a continuation-in-part of application Ser. No. 08 / 926,636, filed Sep. 10, 1997, the entire disclosure of which is incorporated by reference herein in its entirety.1. FIELD OF THE INVENTION[0002] The present invention is concerned with the isolation of a gene of Salmonella which, when genetically disrupted, reduces both virulence and septic shock caused by this organism and increases sensitivity to agents which promote eradication of the bacteria, e.g., chelating agents. The nucleotide sequence of this gene and the means for its genetic disruption are provided, and examples of the use of tumor-targeted bacteria which possess a disruption in this gene to inhibit growth of cancers, including, but not limited to, melanoma, colon cancer, and other solid tumors are described. The present invention also provides for the genetic disruption of this gene in combination with disruption of an auxotrophic gene.2. BACKGROUND OF THE INVENTION[0003] Citation or identifica...

AI Technical Summary

Benefits of technology

[0025] The present invention provides a means to enhance the safety of tumor-targeted bacteria, for example, by genetic modification of the lipid A molecule. The modified tumor-targeted bacteria of the present invention induce TNF.alpha. less than the wild type bacteria and have reduced ability to directly kill normal mammalian cells or cause systemic disease compared to the wild type strain. The modified tumor-targeted bacteria of the present invention have increased therapeutic efficacy, i.e., more effective dosages of bacteria can be used and for extended time periods due to the lower toxicity in the form of less induced TNF.alpha. and systemic disease.

[0028] The present invention also provides a means for enhanced sensitivity for use in terminating therapy and for post therapy elimination. According to one embodiment of the present invention, the tumor-targeted bacteria having a genetically modified lipid A also have enhanced susceptibility to certain agents, e.g., chelating agents. It is a further advantage to modify tumor-targeted bacteria in this way because it increases the ability to eliminate the bacteria with agents which have an antibiotic-like effect, such as chelating agents including, but not limited to, Ethylenediaminetetraacetic Acid (EDTA), Ethylene Glycol-bis(.beta.-aminoethyl Ether) N,N,N',N',-Tetraacetic Acid (EGTA), and sodium citrate. Modification to enhance the ability to eliminate the bacteria via exogenous means, such as the administration of an agent to which the genetically modified bacteria are more sensitive than their wild type counterparts, is therefore useful.

Problems solved by technology

A major problem in the chemotherapy of solid tumor cancers is delivery of therapeutic agents, such as drugs, in sufficient concentrations to eradicate tumor cells while at the same time minimizing damage to normal cells.

However, Lee et al. and Jones et al. did not suggest the use of such mutants as therapeutic vectors, nor did they suggest the isolation of tumor-specific bacteria by selecting for mutants that show infection preference for melanoma or other cancers over normal cells of the body.

Without tumor-specificity or other forms of attenuation, such hyperinvasive Salmonella typhimurium as described by Lee et al. and Jones et al. would likely be pan-invasive, causing wide-spread infection in the cancer patient.

Furthermore, Pawelek et al. did not suggest that modifications to the lipid content of bacteria would alter their sensitivity to certain agents, such as chelating agents.

The problems associated with the use of bacteria as gene delivery vectors center on the general ability of bacteria to directly kill normal mammalian cells as well as their ability to overstimulate the immune system via TNF.alpha. which can have toxic consequences for the host (Bone, 1992 JAMA 268: 3452-3455; Dinarello et al., 1993 JAMA 269: 1829-1835).

In addition to these factors, resistance to antibiotics can severely complicate coping with the presence of bacteria within the human body (Tschape, 1996, D T W Dtsch Tierarztl Wochenschr 1996 103:273-7; Ramos et al., 1996, Enferm Infec. Microbiol. Clin. 14: 345-51).

Further, although Hone and Powell propose the therapeutic use of non-pyrogenic Salmonella with a mutation in the msbB gene, there is no enabling description of how to accomplish such use.

A preferred live bacterial vaccine must be immunogenic so that it elicits protective immunity; however, the vaccine must not be capable of excessive growth in vivo which might result in adverse reactions.

Thus, vaccine vectors which replicate minimally at normal body temperatures, would not be suitable for use as tumor-targeted vectors.

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