Use of one or more metal carriers to selectively kill mammalian cells

a technology of metal carriers and mammalian cells, which is applied in the direction of transferrins, peptides, powder delivery, etc., can solve the problems of inducing rapid death of mammalian cells, and achieve the effect of enhancing the accuracy of toxic metal delivery and high affinity

Inactive Publication Date: 2010-06-03
ZOLTAN LAB
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Benefits of technology

[0008]Embodiments of the invention include two-carrier systems that aid in specific transport of toxic metals into targeted cells resulting in cell death. As used in this application, “targeted” cells or tissues mean non-normal or non-healthy. Such cells or tissues include cancer cells. The first carrier is a peptide or protein that binds several toxic metals but does not enter the cells or enters them only slowly. Using one of the several available methods, the first carrier-metal complex is directly implanted into or targeted to the required tissue. The second carrier is generally represented by a group of chemically synthesized compounds that have higher affinity for the metals compared to the metal-carrying peptides or proteins. Therefore, the second carrier will attract toxic metal atoms from the first carrier, already in or in the vicinity of the target tissue, and carry these metals inside the nearest cells in the target tissue thereby inducing their rapid death. In some embodiments of the invention, the second carrier molecules can be modified by fusing with targeting molecules or moieties to further enhance the accuracy of delivery of toxic metals. In other embodiments, suitable metal-bound second carrier molecules, modified by a targeting moiety, can be used to kill tumor cells in vivo independent of the first carrier.
[0010]Some embodiments of the invention include methods, which allow preferential localization of the metal-loaded first carrier in a target tissue. These methods include implantation of peptide / protein into the target tissue by using specific devices, or fusing the peptide / protein with a targeting moiety that allows specific delivery of the complex to the target tissue. Once the accumulation of the first carrier / metal complex molecules in the target tissue reach an optimum level, subsequent application of the second carrier allows transfer of toxic metal atoms into the cells localized in the target tissue.

Problems solved by technology

Therefore, the second carrier will attract toxic metal atoms from the first carrier, already in or in the vicinity of the target tissue, and carry these metals inside the nearest cells in the target tissue thereby inducing their rapid death.

Method used

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  • Use of one or more metal carriers to selectively kill mammalian cells
  • Use of one or more metal carriers to selectively kill mammalian cells
  • Use of one or more metal carriers to selectively kill mammalian cells

Examples

Experimental program
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example 1

Synthesis of Non Protein Carrier 3-aminopropylpiperidine-1-carbodithioate.HCl Salt (CCK3)

[0074]20.43-g (0.24 mol) piperidine was dissolved in a mixture of 8.0-g (0.2 mol) of NaOH and 35-ml of water and the mixture was cooled to 0-2° C. Then 0.2 mol of CS2 was added drop-wise over one hour period with intensive stirring of the mixture. This was followed by the stirring of the mixture for four additional hours at room temperature (22° C.). The precipitated N-piperidinedithiocarbamic acid was dissolved in 200 ml of water, filtered, mixed with 300 ml of acetone, and kept overnight at 4° C. The white precipitate was filtered, washed twice with ice-cold acetone, and dried. Seventeen g of N-piperidinedithiocarbamic acid was dissolved in 20-ml of water and slowly mixed with 0.011 mol (in 10 ml water) of 3-chloropropylamine (HCL salt). After stirring the mixture for an additional 4 hours period, the mixture was kept overnight at 4° C. The precipitated CCK3 compound was filtered, washed wit...

example 2

Preparation of MTLP-25

[0075]The sequence of the MTLP-25 peptide is as follows:

(SEQ ID NO: 1)Ac-K(ε-NH2)-DCGCSGASSCNCGSGCSCSNCGSG-NH2.

This structure allows the use of terminal NH2 group to bind selected targeting units via amide bond. The peptide was synthesized on MBHA resin (0.56 mmol / g) in 0.1 mmol quantity with Boc-strategy by automatic peptide synthesizer (AB 410). The tri-functional amino acids were protected as follows: Boc-Cys(Mbzl), Boc-Ser(Bzl) and Boc-Lys(Fmoc). For coupling, the DCC / HOBt method was used. After the coupling was complete, the protected peptide, still on the resin, was removed from the synthesizer. The N-terminal amino group was acetylated with the acetyl-OPFP activated ester and the ε-amino group was liberated from the Fmoc group of the N-terminal lysine. In the next step, the side function groups and the peptide were removed from the resin by using HF and scavengers. The solution, which had the pH of 2.0, was divided into three parts. One part was lyophil...

example 3

Demonstration and Quantification of Cell Death

[0076]Cells which lost viability do not synthesize DNA. To quantify cell death, around the time when significant and characteristic morphological changes took place, cells were pulse-labeled for 30 min. with [3H]thymidine to measure DNA activity as described in[Tomono, M. Crilly, K. S. and Kiss, Z. (1995) [Synergistic potentiating effects of choline phosphate and ethanolamine on insulin-induced DNA synthesis in NIH 3T3 fibroblasts. Biochem. Biophys. Res. Commun. 213, 980-985]. Also, cells were collected by making cell suspensions with trypsin, followed by re-plating using fresh tissue culture medium. If no viable cells were obtained after 1 week in culture, it meant that the cells at the time of re-plating were practically 100% dead. Survival values obtained with untreated cells were considered 100%. For the determination of relative number of viable cells after treatments, the MTT assay was used. This colorimetric assay is based on the...

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Abstract

Compositions and methods for decreasing the viability of cells, particularly aberrant non-healthy cells, and most particularly cancer cells, are disclosed. The primary agent that causes cell death is a toxic metal atom or ion. Embodiments of the invention provide compositions and methods to ensure that the toxic metal is directed to the desired cell or tissue. In one embodiment, the metal is bound to a sulfur-rich peptide or protein carrier containing a targeting moiety. Such metal-protein complex is targeted to the selected cells or tissues to enrich the cell or tissue site with the metal-containing peptide or protein molecules followed by administering a dithiocarbonyl which carries the metal from the protein inside the cells to induce cell death.

Description

PRIORITY CLAIM [0001]This application is a continuation of U.S. application Ser. No. 11 / 472,763, filed Jun. 22, 2006, entitled “Use of one or More Metal Carriers to Selectively Kill Mammalian Cells” which claims the benefit of U.S. Provisional Application No. 60 / 697,181, filed Jul. 7, 2005, entitled “Use of one or More Metal Carriers to Selectively Kill Mammalian Cells” both of which are incorporated by reference in their entirety.FIELD OF THE INVENTION [0002]This invention relates to compositions designed to selectively decrease the viability of targeted diseased cells, such as cancer cells, by facilitating transport of potentially toxic metals such as zinc, cadmium, copper or others, across the cell membrane.BACKGROUND [0003]There are many pathological conditions, particularly cancer, when aberrant tissue growth poses serious health hazard that would require rapid, selective, and effective elimination of the constituent aberrant cells. There are three major procedures in the medic...

Claims

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Application Information

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Patent Type & Authority Applications(United States)
IPC IPC(8): C07K2/00C07D475/04C07K14/79C07D493/22C07K7/23C07H21/00C07F15/00C07K16/00A61K33/24A61K33/241A61K33/242A61K33/243
CPCA61K33/24A61K33/30A61K38/16A61K47/48076A61K47/48353A61K47/48884B82Y5/00A61K2300/00A61K47/547A61K47/665A61K47/6929A61K33/242A61K33/241A61K33/243
Inventor KISS, ZOLTAN
Owner ZOLTAN LAB
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