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Production of a biological factor and creation of an immunologically privileged environment using genetically altered sertoli cells

a technology of sertoli cells and biological factors, which is applied in the direction of genital tract cells, enzymes, biocides, etc., can solve the problems of transplants, inability to produce proteins or other biological products, and inability to use specific immunosuppression agents

Inactive Publication Date: 2008-06-19
SERTOLI TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Benefits of technology

[0007]The present invention provides compositions and methods for providing an individual with a biological factor or intermediate thereof which comprises introducing into the individual a ther...

Problems solved by technology

Certain chronic diseases destroy the functional cells in affected organs individuals with such diseases are often unable to produce proteins or other biological products necessary to maintain homeostasis and usually require numerous exogenous substances to survive.
Such transplants, however, are often rejected by the body due to an immune response initiated in response to the foreign tissue or cells.
The use of nonspecific immunosuppression agents however, is fraught with unwanted side effects such as increased susceptibility to infection, hypertension, renal failure and tumor growth.
Immune rejection of genetically altered cells during gene therapy also remains a problem.
Such cells however, may be difficult to obtain due to the diseased state of the patient and are often destroyed during harvesting.
Those cells that survive harvesting are often difficult to grow in vitro and require complicated culture conditions and purification methodologies.

Method used

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Examples

Experimental program
Comparison scheme
Effect test

example 1

[0063]Sertoli cells are isolated from Lewis rats and cultured for two to seven days using standard techniques (Selawry, H. P. and Cameron, D. F., 1993 Cell Transplantation 2:123-129; Korbutt, G. S., et. al., 1997 Diabetes 46:317-322). For example, testicles from 15 to 21-day-old Lewis rates are collected in Hanks' balanced salt solution (HBSS), chopped into 1-mm pieces with scissors, and digested for 10 minutes at 37 C with collagenase (2.5 mg / ml; Sigma Type V, St. Louis, Mo.) in HBSS. The digest is washed three times with calcium- and magnesium-free HBSS containing 1 mmol / EDTA and 0.5% bovine serum albumin (Sigma) (HBSS / EDTA), digested for 10 minutes at 37° C. with trypsin (25 μg; Boehringer) in HBSS / EDTA, and washed four times in HBSS; the final cell pellet is resuspended in HAM's F10 media supplemented with 10 mmol / l D-glucose, 2 mmol / l L-glutamine, 50 μmol / l isobutylmethylxanthine, 0.5% bovine serum albumin, 10 mmol / l nicotinamide, 100 U / ml penicillin, 100 ng / ml streptomycin, an...

example 2

[0069]The collected Sertoli cells from Example 1 are then pooled and transplanted in a normal rat under the kidney (renal) sub-capsular space. The transplantation protocol used to transplant the stably transfected cells is the same as that used for transplantation of islet cells and Sertoli cells described in Rajotte et al. (1997) Diabetes 46:317-322. For example, each recipient receives between about 5.5±0.3 to 11±0.4×106 transgenic Sertoli cells. Cells are aspirated into polyethylene tubing such as PE-50, pelleted by centrifugation, and gently placed under the left renal subcapsular space of anesthetized (e.g., halothane-anesthetized) animals. The number of transgenic Sertoli cells grafted in each animal is assessed by measuring the DNA content of triplicate representative aliquots of each cell preparation before transplantation, and calculations are based on the finding that freshly isolated testicular cells contain about 20 pg DNA per cell. Therefore, the total DNA content trans...

example 3

[0073]Lewis rat Sertoli cells are transfected with a retroviral vector constituting the alkaline phosphatase gene. The RetroExpress® system, available from Clontech is especially useful. After selection and expansion as described in Example 2, transfected Sertoli cells are transplanted into Wistar-Furth rats as described in Example 2.

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PUM

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Abstract

The present invention provides a method of providing an individual with a biological factor or intermediate thereof which comprises introducing into the individual Sertoli cells genetically altered to produce the biological factor or intermediate thereof. The genetically altered Sertoli cells are administered in an amount effective to produce the desired effect. Aside from producing the biological factor or intermediate thereof, the engineered Sertoli cells also create an immunologically privileged site. Vectors comprising a promoter which functions in Sertoli cells operably linked to coding sequence for a desired biological factor are also provided as are Sertoli cells comprising such vectors. A pharmaceutical composition comprising Sertoli cells genetically altered to produce a biological factor is also provided.

Description

FIELD OF THE INVENTION[0001]Transplants of healthy organs or cells into a patient suffering from a disease are often rejected by the body due to an immune response initiated in response to the foreign tissue or cells. Genetically altered cells administered during gene therapy are often met with a similar immune response. The present invention provides a method of cellular transplantation in which an immunologically privileged site is created, thus alleviating the rejection associated with conventional transplantation and gene therapies.[0002]Specifically, the present invention provides compositions and methods for providing an individual with a biological factor or intermediate thereof which comprises introducing into the individual a therapeutically effective amount of Sertoli cells genetically manipulated to produce the biological factor or intermediate thereof and wherein the Sertoli cells create an immunologically privileged site. A pharmaceutical composition comprising genetica...

Claims

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Application Information

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IPC IPC(8): A61K35/12C12N15/63C12N5/08A61P3/10A61K48/00C12N5/071C12N9/10C12N9/64
CPCA61K38/28A61K38/4846A61K48/00C12Y304/21022C12N5/0683C12N9/1051C12N9/644A61K2035/122A61P3/10
Inventor KIRKPATRICK, SHAUN A.GORES, PAULHALBERSTADT, CRAIG
Owner SERTOLI TECH
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