Chemistry: molecular biology and microbiology – Animal cell – per se ; composition thereof; process of... – Primate cell – per se
Reexamination Certificate
1999-11-18
2002-05-07
Yucel, Remy (Department: 1636)
Chemistry: molecular biology and microbiology
Animal cell, per se ; composition thereof; process of...
Primate cell, per se
C435S320100, C435S069100, C424S093210
Reexamination Certificate
active
06383807
ABSTRACT:
The invention relates to cells designed as traps for antigens causing dysfunctions of the immune system, more particularly those which are implicated in naturally occurring (auto-immune diseases) or induced (grafts) dysfunctions of the immune system.
The invention relates more particularly to the provision of medicines for the prevention of diseases of the “Graft versus Host Diseases” (GVHT) (diseases due to the reaction of a transplant against the host) or for the treatment of auto-immune diseases, more particularly in association with the said cells designed as traps.
In what follows the expression “antigen” relates to all substances or all products against which tolerance on the part of the host is desired, althou; they would naturally tend to induce a defensive reaction or one of rejection on the part of the immune system of the host.
The objective of the invention is to remedy these deficiencies, more particularly to provide a different therapeutic approach based on the presence, ensured beforehand in the host, of lymphoid cells, in particular T lymphocytes trapped against an activation by the antigens with respect which tolerance is sought.
The invention thus relates more particularly to a population of hematopoietic cells, designed as traps to prevent an activation by antigens normally foreign to the host but with respect to which tolerance is desired, characterized in that these cells contain a genetic sequence, the expression product of which is able, when its production is enhanced to give a sufficient quantity of it within the cell traps, to react in situ with a pharmaceutically active substance to induce the destruction of these cells, and means specifically inducible by an activation signal supplied to these cells by the antigen to trigger the above-mentioned enhanced production.
The administration of such a pharmaceutically active substance to a host carrying. such a population of cell traps, can provide for the effic ient protection of the host against an activation by antigens of the appropriate specific lymphoid cells of the immune system, in particular of its T lymphocytes, as a result of the destruction of these lymphoid cells even before they have begun to divide. As a result of the destruction of the cell traps carried out at the command of the antigen itself, their division will be blocked.
Generally speaking the procedures used up to now to confront the diseases resulting from an intolerance on the part of the immune system tend either to cause a generalized immunodepression in thehost or modify some classes of the cells affected by this type of antigen, in a manner such as to confer on them the capacity to produce constitutively certain substances which are supposed to inhibit the development of such antigens.
The invention relates more particularly to a population of cells of the bone marrow, in particular imnature cells which can be used for bone marrow grafts, these cells being “designed as traps” under the conditions indicated above.
The invention thus also relates to the use for the production of medicines active against diseases of the GVHT type of a pharmaceutically active molecule which is able, when it is associated in vivo with a population of cells, in particular hematopoietic cells, designed as traps in the sense described above and reintroduced into the organism of the host, in particular subsequent to a bone marrow graft, to interact with the expression product of the genetic sequence contained in these cell traps to cause their destruction as soon as they are stimulated by the antigens against which tolerance is desired.
In a preferred embodiment of the invention, the population of cell traps is characterized in that the As providing for the specific triggering of the enhanced production of the expression product and the genetic i@sequence mentioned above are contained in a recombinant DNA sequence contained in these cells. Advantageously, the agents specifically inducible by a signal resulting from the activation of these cells by these antigens consist of regulatory sequences normally controlling the expression of a cytokine, in particular a lymphokine, for example interleukin 2, or the expression of receptors corresponding to this cytokine, in particular the interleukin 2 receptor, and the above-mentioned genetic sequence is then placed under the control of this promoter and codes for an expression product which is able, when its production is enhanced as a result of the activation of the above-mentioned regulatory sequences , to react with a pharmaceutically active substance to form a reaction product capable of inducing directly or indirectly the destruction of the cells in the process of being activated.
Advantageously, the regulatory sequence used is constituted by a promoter normally associated with interleukin 2 or preferably with an interleukin 2 receptor. In particular, recourse will be bad to the promoter for the alpha chain of the interleukin 2 receptor (sequence described in Cell, April 1987). When activated, this promoter leads to an amplified expression of the sequence placed under its control (in particular in JURKAT cells) which is of the order of 30 times that which is measured when it is not activated.
Advantageously, the pharmaceutical substance used induces directly or indirectly the interruption in situ of the replication of DNAs within the cells in question. In a manner also particularly preferred, the invention also makes use of the procedure called “obliteration by thymidine kinase” already described by Borelli et al. (1988) in cells already expressing it constitutively and in a different field of application. The population of cell traps conforming to a preferred embodiment of the invention is then characterized in that the expression product encoded in the said genetic sequence placed under the control of the said promoter is a molecule such as the thymidine kinase of the Herpes simplex virus 1 (HSV1-TK) which is able, when it is present at a sufficient concentration in the cells in question, to phosphorylate nucleoside analogues, such as acyclovir (9-/(2-hydroxyethoxy)methyl/guanine or gancyclovir (9-[1,3-10 hydroxy-2-propoxymethyl] guanine), to give the monophosphate derivatives, which can themselves be converted by cellular enzymes into nucleoside triphosphates which can be incorporated into nucleic acids in the course of chain elongation brought about by polymerases within the said cells, resulting in the interruption of the elongation of the chains and cell death which follows.
It will be apparent immediately that the “active pharmaceutical substance” utilisable in a therapeutic protocol which makes use of one of the preferred different populations of cell traps such as defined above must, in each case, exhibit the properties which will allow it to react with the corresponding expression product under the conditions which have been defined.
The invention in fact takes advantage of two distinct phenomena:
1) the expression of a toxic gene placed under the control of a promoter for a cytokine or a cytokine receptor in cell traps conforming to the invention is at the mst very low as long as they are not activated by the antigen in question;
2) the induction of the expression of the toxic genetic sequence may be made sufficiently rapid in those of the cell traps which are activated to trigger their destruction even before the first cell divisions of these cells occur. Consequently, the actlivation of the immune system of the host is blocked in its initial phase, when it is stimulated by an antigen towards which tolerance is in fact desired.
In order to construct for use in man—or, optionally, in animals—the cell traps conforming to the invention, recourse may be had to any adequate procedure, in particular by in vitro infection of the corresponding cells by a pseudo-viral particle of the amphotropic Moloney type, which thus also infects human cells. These viral particles are produced by a socalled “packaging” cell line which will have been constructed beforehand. A packaging line is capable o
Brown Rudnick Berlack & Israels LLP
Hofer Mark A.
Katcheves Konstantina
Universite Pierre et Marie Curie
Yucel Remy
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