JPH07508977A - Treatment of cell hyperproliferation by inhibition of interleukin-1 - Google Patents

Abstract

(57) [Summary] This bulletin contains application data before electronic filing, so abstract data is not recorded.

Description

[Detailed description of the invention] Reciprocal application for treatment of cell hyperproliferation by inhibition of interleukin-1 reference This application is filed under U.S. Patent Application No. 07/707.89, filed March 31, 1991. This is a continuation-in-part application of No. 7, and the disclosure of that U.S. application is incorporated herein by reference. It has been inserted.

Background of the invention The present invention relates to a method of treating cell hyperproliferation. These methods are useful for cells, especially an excess of epithelial cells and, more preferably, keratinocytes (keratinocytes) It is useful in the treatment of various pathological conditions characterized by proliferation.

Conditions characterized by hyperproliferation of cells, especially keratinocytes, are difficult to treat That had been proven. Traditional treatments for some of these conditions, such as dry drunkenness, The location was insufficient. Therefore, by reducing or preventing cell hyperproliferation, A need exists for treatments that can treat these conditions.

Cytokine levels have been studied in certain conditions characterized by cell hyperproliferation. It's been coming. In particular, interleukin-1α in normal skin and psoriatic skin. Attempts have been made to compare levels of interleukin-1β. For dry skin It has been reported that interleukin-1 activity in skin is decreased compared to normal skin. It was done. However, compared to the level of interleukin-1β in normal dermatome, In comparison, the levels of high molecular weight interleukin-1β in dry skin are more The invention relates to interleukin-1 (IL-1) activity, or intracellular interleukin activity. -1 receptor antagonist activity by inhibiting or reducing the activity of the cell, especially the A method of preventing or reducing hyperproliferation of skin cells is provided. Therefore, the present invention A method comprising contacting the cells with an excessive growth-inhibiting amount of a tarleukin-1 inhibitory compound. Regarding the law. Interleukin-1 inhibitory compounds include interleukin-1 or Contains compounds that reduce intracellular interleukin-1 receptor antagonist activity. be caught. According to one method, interleukin-1 activity is inhibited by inhibiting its synthesis or is inhibited by down-regulating it. others According to the method, interleukin-1 activity inhibits the conversion from the precursor form to the active form. It is inhibited by Inhibition of this conversion to the active form converts the precursor form into the active form. including, but not limited to, preventing or reducing the expression of converting enzymes that convert , may be achieved by one of several methods, or by converting the converting enzyme itself. This may also be achieved by inhibiting two or more. Appropriate interleukin-1 inhibition The compound includes interleukino-1, interleukin regulatory factor or interleukin A group of oligomers that regulate the expression of Kin-1 converting enzyme (antisense oligomers, three-stranded oligomers or triple-helical oligomer pairs). Interloiki Interleukin-1 modulators include compounds that modulate interleukin-1 activity and e.g. , intracellular interleukin-1 receptor antagonists.

Ike's interleukin-1 inhibitory compounds were targeted (i.e., inhibiting hyperproliferation). interleukin-1 or intracellular interleukin Select from various classes of compounds that reduce the activity level of -1 receptor antagonists You may do so.

According to a preferred method, the invention provides oligomer(s) in an amount that inhibits hyperproliferation. , specifically for hyperproliferation of skin or epithelial cells by contacting them with skin or epithelial cells. It relates to a method of treating a symptomatic pathological condition. This oligomer(s) It may be a chain oligomer, a third strand oligomer or a triple helix oligomer. Like this Antisense oligomers are RNs transcribed from target genes present in cells. It has a sequence complementary to the A sequence. The third strand oligomer is a marker present in the cell. It has a sequence complementary to the selected double-stranded nucleic acid sequence of the target gene. triple helix The oligomer pair is complementary to a single-stranded nucleic acid sequence of the target gene or its transcript. . The target gene is a cytokine that regulates intracellular proliferation or its regulatory factor (receptor). selected from genes encoding enzymes that convert It will be done.

According to another preferred method, the present invention provides antisense oligomers, third strand oligomers, One of the oligomers, which includes gomer or triple helical oligomer pairs, was allowed to over-proliferate. excessive proliferation of skin or epithelial cells by contacting the cells with an amount that inhibits Relating to methods for reducing or protecting against. , the oligomer is present within the cell or transcript It has a sequence complementary to the nucleic acid sequence from the target gene. The target gene is , cytokines that mediate cell hyperproliferation, or against such cytokines. modulators such as receptor antagonists, or converting enzymes, or important for its function. from genes encoding enzymes involved in the translation or post-translational modification of cytokines. selected.

Preferred target genes include interleukin-1β (IL-1β), Ikin-1α (IL-1α), intracellular interleukin-1 receptor antagonist (iclL-1ra or intracellular IL-1ra) or interleukin-1 conversion Contains genes that encode enzymes.

definition As used herein, unless otherwise specified, the following terms have the following meanings:

The term "purine" or "purine base" refers to the naturally occurring adenine and guanine Not only bases substituted at the 8-position or groups modified at the 6-position. modifications of those bases, such as anino analogs or adenine analogs, 2-aminopurine. It also includes decorations.

The term “nucleo7d” includes and can be used interchangeably with the nucleo7d unit. used to refer to a subunit of a nucleic acid that contains a pentose and a nitrogenous base.

This term applies not only to units having A, G, C, T and U as bases, but also to It further includes analogues and modified forms of its bases (such as 8-substituted purine groups). .

In RNA, the pentose is a lipoose, and in DNA, it is a 2-carbon sugar. - Deoxy/ribose. The term also refers to 2°-O-alkyl ribose, including analogs of such subunits, such as modified sugars.

"The term phosphonate is a compound of the formula (in which R represents a group represented by (represents an alkyl or aryl group). suitable alkyl or alkyl The lyl group contains a group that does not sterically interfere with the phosphonate bond or interact with each other. Contains many groups. The phosphonate group may be present in either the rRJ or "S" configuration. It's okay. The phosphonate group is located within the nucleononor that binds the nucleonyl unit. It can be used as a bond (or link) for a phosphorus group.

The term "phosphonoester" refers to a group of the formula: The diester group is the bond of the phosphorus group within the nucleonodyl unit that is bonded to the nucleonodyl unit. (or link).

“Non-nucleonodo monomer unit” is a basic skeleton of base, sugar and/or phosphorus. , represents a monomer unit substituted with another chemical moiety.

"Nucleoside/non-nucleoside polymer" refers to nucleoside and non-nucleoside polymers. Leonod Represents a polymer containing monomer units.

The term "oliconucleoside" or "oligomer" typically refers to about 6 to about 1 00 nucleosides in length, but even more than about 100 nucleosides in length. Represents a chain of nucleotides connected by a good, intranucleo7 bond. It is It is usually synthesized from nucleoside monomers, but may also be obtained by enzymatic methods.

Therefore, the term "oligomer" refers to a nucleotide that joins nucleoside monomers. Represents a chain of oligonucleotides with intraleonodo linkages and therefore Otides, nonionic oligonucleotides, alkyl- and aryl-phosphonates Title analogs, alkyl- and aryl-phosphonothioates, oligonucleotides phosphorothioate or phosphoronthioate analogue of oligonucleotide Neutral phosphate esters, such as phosphoramidate analogs, phosphotriesters Oligonucleoside analogs and other oligonucleoside analogs and modified oligonucleotides Contains gonucleosides, as well as nucleo/de/non-nucleoside polymers. Contains. This term means that one or more of the phosphorus linkages between the monomer units , formacecool bonds, sulfamate bonds or carbamate bonds. Polymers of nucleotides/nucleotides substituted by Contains. It also includes morpholino base analogs or polyamide base analogs. Similarly, nucleotides/non-nucleons in which both the tang and phosphorus moieties are substituted or modified. Also includes polymers. It also includes non-nucleotide bases, sugars and phosphorus. The basic backbone of the acid is replaced by a non-nucleoside moiety or Onodo moieties are inserted into nucleonodo/non-nucleonodo polymers Also includes nucleoside/non-nucleoside polymers. Non-nucleosides of the above The moiety is capable of interacting with the target sequence or is capable of being taken up into the target cell. may optionally be used to conjugate other small molecules that can alter the

The term "alkyl- or aryl-phosphonate olicomer" also has one alkyl- or aryl-phosphonate nucleonone bond represents the oligomer that is present. Suitable alkyl- or aryl-phosphonate groups include Alkyl- or aryl that does not sterically hinder or interact with the sulfonate bond Contains a - group. Preferred alkyl groups have from about 1 to about 6 carbon atoms. Contains lower alkyl groups. Suitable aryl groups have a conjugated pl electron system. at least one ring containing carbocyclic aryl and heterocyclic aryl groups. which are optionally substituted and are highly preferred having about 10 carbon atoms. stomach.

"Methylphosphonate oligomer" (or rMP-oligomer) The term refers to oligonucleotides having at least one methylphosphonate intranucleosin linkage. Represents gomer.

The term "neutral oligomer" refers to the nonionic link between the nucleonid monomers. Intracleosidic bonds (i.e., bonds that have no net positive or negative ionic charge) for example, alkyl- or aryl-phosphonates. bond, alkyl- or aryl-phosphonothioate, phosphotriester linkage Neutral phosphate ester bonds, especially neutral ethyltriester bonds, such as Non-linkages such as sulfomate, morpholino, formacecool and carbamate linkages with intranucleoside bonds, such as intranucleoside bonds containing phosphorus. represents the oligomer that is present. Neutral oligomers optionally include oligonucleotides or nucleotides. Cleonode/Non-nucleo/de between the polymer and the second molecule containing the conjugated partner may include a conjugate. Such a conjugate partner is an intake agent. ), alkylating reagents, binding substances for cell surface receptors, lipophilic reagents, sola Nucleic acid modifying groups including photo-crosslinking-linking reagents such as Ren and cleave target sites on nucleic acids. Capable groups and the like may be included. Such a conjugate partner is also an oligomeric It may promote uptake or modify the interaction of the oligomer with the target sequence. Alternatively, the pharmacodynamic distribution of the oligomer may be altered. What is effectively required is Oligonucleotides or nucleosides/non-nucleosides contained in oligomer conjugates The main thing is that the polymer must be neutral.

The term "neutral alkyl- or aryl-phosphonate oligomer" refers to neutral nuclei containing at least one alkyl- or aryl-phosphonate linkage; Represents a neutral oligomer with an intra-onodo bond.

The term "neutral methylphosphonate oligomer" refers to Represents a neutral oligomer with an intranucleonid linkage containing a tylphosphonate linkage. vinegar.

The term "tandem oligonucleotide" or "tandem oligomer" , complementary to a sequence located on either the 5′- or 3′-side of the target nucleic acid sequence. the hybridizing oligomer with a second oligomer complementary to the target sequence. represents a oligonucleotide or oligomer. Tandem (series) oligomers target nuclei The target sequence can be transformed into such oligomers, such as by reducing the secondary structure of the acid sequence. help these oligomers get closer to their targets. It is possible to improve Izee/Yono. Additionally, a pair of tandem oligomers one member of the above second oligomer (or vice versa) by promoting a helical structure at either of the 3-termini. It is possible to improve the hybrid stability of hybrid oligomers to target nucleic acid sequences. Wear.

The term "short chain aliphatic alcohol" has about 2 to about 20 carbon atoms; The fatty (alkyl) chain represents an alcohol that can be straight or branched, and can be primary, Includes secondary and tertiary alcohol groups, glycols and polyols.

The term ``flux enhancer'' refers to the transdermal flux (transdermal fluidity) of a compound. Represents a substance used to increase Franks accelerator is a compound that applied to the skin or mucous membranes in combination with compounds to increase lux. is typical. Accelerators act by disrupting the skin or mucus membrane barrier. , or by altering the distribution behavior of the drug in the skin or mucus. it is conceivable that.

-The term “a pair of triple helix orikoma” refers to a pair of RNA or DN, A-like The first target is complementary to the segment of the chain target cough acid and is capable of hydrogen bonding. and the second 11-comer, therefore, they and the single-stranded nucleic acid together represent the A triple hexagonal structure can be formed.

7 The term tertiary oligomer f refers to DNA double helix, RNA double helix or ;: DNA-RNA double helix-like double zero-stranded nucleic acid segment and hybridization It is possible to form a three-fold helical structure with these. Represents oligomer vs.

The term ``first and second'' refers to a pair of triple helix orikoma (or a pair of first and second orikoma). = and -) or in the case of the third strand oricomer: Well, the combination with the base sequence 'Wl+ of the nucleic acid Forms a triple helix structure through hydrogen bonding (and base pairing or hybridization) represents an oligomer having the base sequence.

Detailed description of the invention According to the invention, a condition characterized by hyperproliferation in cells, in particular epithelial cells, By using an ic-leukin-1 inhibitory compound in an amount that inhibits hyperproliferation. and treat it. Interleukin-1 inhibitory compounds include interleukin-1 or is a compound that reduces intracellular interleukin-1 receptor antagonist activity. be. Suitable interleukin-1 inhibitory compounds include interleukin-1, Interleukin-1 receptor antagonist-like Prevents expression of nodal factor or enzyme (converting enzyme) that converts IL-1 precursor into active form or compounds that reduce or reduce the peptides, thus competitive or Non-competitive inhibitors, small molecule inhibitors, antibodies, that bind to the active site of a protein. oligomers that modify the function of - or triple helical oligomer pairs. These ori The appropriate nucleotide sequence for gomer can be determined from the sequence of the target gene. Can be done. Preferred sequences of target regions are described later in this specification. Other inns Tarleukin-1 inhibitory compounds inhibit the converting enzyme, thereby inhibiting IL-1 Included are compounds that prevent conversion of the precursor to the active form. Other interleukins 1 inhibitory compounds inhibit the activity of intracellular interleukin-1 receptor antagonists. Contains compounds that reduce

(Margin below) A Preferred oligomer The oligomers selected can be charged or uncharged backbones (bank bones). It can be of any of a number of types.

Preferred oligomers are alkyl- and aryl-phosphonate oligomers. Methylphosphonate oligomers are particularly preferred. Other preferred oligos Examples include phosphorothioate oligomers, morpholino analogs, formacetic Examples include cool cervical body and peptide nucleic acid (pNA) sequence body. Also, a small Oligomers having at least about 8 nucleotide units are preferred, and from about 8 to 40 More preferred are nucleonod units. nucleonode/non-nucleonode Also preferred are oligomers which are polymers. As a preferred oligomer, hybrid RNA.

Chimeric oligonucleotides that are DNA triads are also included [Inouni ( Inoue et al., rFEBsLett, j2115:327 (1987). light]. For example, methylphosphonate oligos with attached cleavage or cross-linking moieties. Oligomers with neutral compounds such as sesame are advantageous under certain conditions. I can prove something. Such oligomers allow the cleavage or cross-linking moiety to Its neutral structure can promote the inactivation of polynucleotide sequences while its neutral structure It has a relatively long in vitro half-life because it reduces the rate of enzyme digestion.

In one embodiment of the invention, these antisense oligomers are selected from It has a complementary sequence to a part of the RNA transcribed from the target gene. Accuracy of inhibition Although the molecular mechanism behind this is not conclusively elucidated, the first antisense due to the formation of a double helix between the RNA transcribed from the target gene and the RNA transcribed from the target gene. It has been suggested that. The double helix formed in this way is responsible for the translation of the mRNA sequence. translation, inhibiting prosenonog or transport or digestion with the enzyme RNase I It may be possible to induce

In another embodiment of the invention, the double-stranded or single-stranded nucleic acid is complementary to the target sequence. a third strand oligomer with a sequence selected to form a triple helisox complex By triple helinox formation using lycomer or triple helical oligo markers, Tau control/control of cell proliferation is achieved, thereby allowing target nucleic acid sequences to can be prevented or prevented from occurring. Formation of triple helix complex to prevent or interfere with the expression of double-stranded or single-stranded target sequences based on Regarding the use of oligomers (including third strand oligomers and triple helical oligomer pairs) Further information is provided in co-pending U.S. patent application Ser. No. 07/388,027, No. 07/751813 and No. 07/772081, which are cited is incorporated herein by.

As a general matter, the oligomer used has a sequence complementary to the target nucleic acid sequence. will do. However, absolute complementarity may not be necessary. Generally, the target nucleic acid column and a stable 2ffi helix (or, in some cases, a triple helix complex) Any oligomer with sufficient complementarity to form is considered suitable. Ru.

Stable double helix formation depends on the sequence and length of the hybridizing oligomer and the relatively long, as it depends on the degree of complementarity between the antisense oligomer and the target sequence. When oligomers are used, the fidelity (complementarity) may be low. This is triple This also applies to the oligomers forming the linox complex. However, the book Particularly suitable for use in the method of the invention are those having a melting temperature of about 1 degree under physiological conditions. 40'C and sufficient complementarity to form a double or triple helithox structure. It is an oligomer of about 8 to 40 nucleonod units in length.

The concentration of oligomer used depends on the type of hyperproliferative condition being treated, the tissue (i.e., whether administered locally or systemically), depends on a number of factors, including the type and specificity of the individual antisense oligomers used. There may be a variety of ways to do this. In the studies described herein, at concentrations in the 50 IIM range, Significant inhibition was observed in the test system used, but once the oligomer Other conditions higher or lower may also be preferred.

When oligomers are administered in cinnamon, neutral oligomers are preferred.

For parenteral (e.g., injection) administration, neutral oligomers or ionically charged bags may be used. Oligomers with 1-7 (i.e., with charged internal nucleonid units) ) may be used.

In one preferred embodiment, these oligomers are polynucleotides or or combinations of nucleonod/non-nucleonod polymers and association partners. It is in the font. Suitable association partners include intercalating agents such as acridine, alkyl binding agents for cell surface receptors, philophilic agents, psoral en)t; any photocrosslinking agent, other crosslinking agent, prochelate, or hydrolyzing agent; Nucleic acid markers such as nuclear degrading agents (such as copper 0-phenanthroline or EDTA ions) Examples include nucleic acid modifying agents that cleave the target moiety, and all of these may be taken into account.

The association partner is an enzyme-mediated modified nuclease or nucleotide analog. or by chemical modification of oligomers (e.g., non-nucleotide They can also be incorporated into oligomers (such as by using a licker).

- When used to prevent the function or expression of terminal or double-stranded nucleic acid sequences, these The majority of the oligomers are derivatized or modified to incorporate nucleic acid modifying groups so that the nucleic acid reaction occurs, irreversibly modifying the structure of the nucleic acid, thereby changing the function of the nucleic acid. can be eliminated.

U.S. Patent Application No. 565,299 describes psoralen-derivatized oligomers. (which is incorporated herein by reference).

As mentioned above, a wide variety of nucleic acid modifying groups can be used as association partners to oricomers can be induced. As nucleic acid modification groups, derivatized oligomers After forming a complex of single-stranded or double-stranded nucleic acid segments, the nucleic acid segments Also: = crosslinking, alkylation, cleavage, decomposition, or other modification of the target sequence E 7: cause activation or destruction, thereby altering the functionality of the nucleic acid segment. Examples include groups that irreversibly inhibit the ability and/or expression of the expression.

The position of the nucleic acid modifying group in the 1j commer varies and depends on the particular nucleic acid modifying group used. Varies depending on the decorating group and the target nucleic acid segment. Therefore, the nucleic acid modifying group ``can be present at the - terminus or placed intermediate between the two termini. Wear. Multiple nucleic acid modifying groups may be included.

In one preferred embodiment, the elegant nucleic acid modifying group is photoreactive (e.g., photosensitive). (activated by a certain wavelength or range of wavelengths), it reacts with light to produce an optical A reaction, or crosslinking, occurs between the oligomer and the nucleic acid target.

A specific example of a nucleic acid modifying group that can cause crosslinking is aminomethyltrimethylsol. These are psoralens such as ren group (AMT). Because AMT is photoreactive Conveniently, therefore, irradiation with light of a specific wavelength before cross-linking is achieved must be activated by Other crosslinks, photoreactive or not These oligomers may be derivatized with groups.

In another embodiment, a nucleic acid modifying group covalently binds to a nucleic acid segment to render it inactive. It may also be an alkylating agent. Suitable alkylating agent groups are well known in the chemical industry. is known, and includes groups derived from alkyl halides, haloacetamides, etc. It will be done. Polynucleotides capable of causing cleavage of polynucleotide segments Examples of demodifying groups include moieties that generate radicals, as well as moieties that can be modified by nuclear attack. This includes parts that promote decomposition. Ethylenenoaminetetraacetate (EDTA) also Transition metal chelate complexes, such as their neutral derivatives, can be used to generate radicals. I can be there. Other groups used to perform radical-mediated cleavage include: Examples include funinanthroline and porphyrin. When using EDTA, turn it off. To assist in the generation of radicals, it is preferred to link iron to oligomers. Iron- Although EDTA is the preferred polynucleotide cleavage group, other nitrogen-containing substances ( or other transition metal chelate complexes may also be useful. Ru.

Other cleavage agents promote the addition of water to phosphorus-internal nucleotide bonds. These include nucleating agents and hydrolyzing agents. Such agents include amines , a substituted guaninonium group, an imidazole group, and the like.

1 Preferred endogenous oligomer preparation [Preferred starting oligomers include neutral alkyl- and aryl-phosphonates. oligomers and neutral oligos with morpholino or phosphorimidate linkages Mar is mentioned. Particularly preferred are neutral methylphosphonate oligomers. tape Skin that has been peeled off (the stratum corneum has been removed and is reported as a model of mucous membranes) - Methylphosphonate in terms of its ability to adhere to any skin Neutral methylphosphonate oligomers with only internal nucleo/binary bonds are unique. preferred.

A method for synthesizing methylphosphonate oligomers is described in Example 1 of this specification and Li-(Le rBiochemistry J27:3197-3203 (1988) and Miller, P.S. et al. mistryJ25 + 5092-5097 (1986) (These documents (herein incorporated by reference).

In another embodiment of the invention, the oligomer can be neutral until it enters the cell. Once inside, it is converted into a charged species through chemical or biological processes. Preferred are oligomers. Such charged oligonucleotides are nuclease Other moieties may be included to stabilize the oligonucleotide against degradation. 2゛ - Substituents such as methylribose group, various base modifications, and phosphorothioene Analogs of the phosphorus backbone, such as be able to. Additionally, methylphosphonates or other neutrals in oligomers The presence of internal nucleotide bonds confers exonuclease resistance.

Neutral oligomers with about 6 to about 40 nucleotides are preferred, with about 12 to about 2 nucleotides being preferred. More preferred is 0 nucleosides. If you want complementarity to a longer sequence, use , neutral oligomers with 20 or more nucleosides may be used, but relatively By using short neutral tandem oligomers, targets such as mRNA can be Solubility and penetration into the skin or mucous membranes while competing with the development of secondary structure of the target nucleic acid. It is desirable to maximize transparency. Alternatively, each oligomer may contain the same gene or are complementary to distinct target sequences that are part of different genes. The use of one or more neutral oligomers is advantageous.

If the neutral oligomer is an alkyl- or aryl-phosphonate oligomer, Incorporating a nucleonodo monomer unit with a modified ribosyl moiety is advantageous. In these neutral oligomers, 2'-0-alkyl- and and in particular the use of nucleotide units with a 2°-methyl-liponol moiety. Therefore, the hybridization of the oligomer to its complementary target sequence is advantageously modified. It will be good.

Suitable formulations include from about 0.0001 to about 2% by weight of neutral oligomer.

In one preferred embodiment, from about 2 to about 100% short chain aliphatic alcohols. A neutral oligomer formulation is provided. Suitable alcohols include ethanol, iso Examples include propyl alcohol, propylene glycol and glycerol. Ru. In some studies, formulations of neutral oligomers containing ethanol have been shown to be suitable It showed transdermal flux.

In particularly preferred embodiments, these neutral oligomer formulations further include flux. A speed accelerator may be included. Suitable flux promoters include those known in the art. and de/methyl sulfoquinide, trimethyl sulfoxide and PCT applications Listed in number PCT/US86102583 (publication number W○87103473) Examples include cyclic ketones, lactones, aldehydes, and esters. .

Some flux enhancers increase the retention of oligomers and therefore increase their retention within the skin. Some have the effect of increasing the concentration of oligomers.

Therefore, in oligomeric formulations for direct (topical) administration, such as topical application to the skin, This not only maximizes transdermal flux, but also improves the retention of oligomers in the skin. It is preferred to use a flux promoter that increases the flux. Additionally, certain cyclic ketones and lactone promoters have been reported to increase local retention and therefore may include facilitators for topical administration of the preferred class of oligomeric formulations.

For oligomeric formulations for systemic treatment, the increase in local retention of the oligomer is minimal. Therefore, it is preferable to use a flux accelerator that maximizes flux. .

2 Preferred target genes and target sequences In a preferred embodiment, the present invention Expression of cytokines that influence proliferation, from precursors of such cytokines agents that interfere with their conversion to active forms or the expression of their intracellular receptor antagonists. The present invention relates to a method of preventing or reducing cell proliferation using oligomers.

Suitable oligomers include antisense oligomers, third strand oligomers and Examples include triple helix oligomer pairs.

In one embodiment of the invention, IL-1β, IL-1α, 1clL-1ra Alternatively, the target sequence on the DNA encoding IL-1β converting enzyme or the target sequence converted therefrom. By administering an oligomer that is complementary to the mapped mRN, A , interleukin-1β (IL-1β), interleukin-1α (IL-1 α) or intracellular splice variants of interleukin-1 receptor antagonists body (“intracellular protein/tarleukin-1 receptor antagonist” or 1clL-1 preventing or preventing the expression of IL-1 converting enzymes such as ra) or IL-1β converting enzyme Methods are provided for reducing cell hyperproliferation by interfering with it.

Therefore, the present invention provides growth-inhibiting amounts of oligomers (antisense oligomers, 3-stranded oligomers or triple-helical oligomer pairs) and how to reduce hyperproliferation of keratinocytes or other epithelial cells. It is something that Antisense oligomers contain RNA sequences transcribed from target genes. Complementary to column. The third strand oligomer is formed by hydrogen bonding with the double-stranded nucleic acid sequence. Ki, Maf: 3 Base sequences selected to form an ffi helinox complex Has columns.

The first and second oligomers of the triple helical oligomer pair are attached to the target-heavy chain nucleic acid sequence. is complementary to and capable of forming hydrogen bonds with the sequence and together with the heavyweight nucleic acid. The sequence was selected so that it could form a triple υnox complex.

Target genes are cytokines that mediate cell proliferation or their receptor antagonists. Selected from a group consisting of genes encoding esters or their converting enzymes One IL-1α and IL-1β are listed as cytokines that suitably target r=tg. IL-1β is synthesized as a Dalton precursor in 31, and is an extracellular active resident. In order to obtain Therefore, the gene encoding IL-1β converting enzyme is another preferred target gene. It is a child. Other preferred target genes include IL-1α, IL-1β or rL-1 family 3, which is critical for IL-1ra function (myristylation, etc.) Genetic forces that coat other enzymes that are also involved in translational or post-translational modification of no. Can be mentioned. A region in the transcript of a selected gene is oligomerized versus dimorphic. is a preferred target for

When hybridizing to a target site, the site should be adjacent to the above site or A suitable length of oligomer (from about 8 to about 40 nucleotides are preferred, and about 12 to about 20 nucleotides are more preferred. (new). Such sites in pre-mRNA include the splice Acceptors, splice donors and splice branch points and polyA addition regions There are several areas. The preferred site in mRNA is the start codon or mRN Such as the 5° end (cap site) of A. The sequence of the oligomer corresponds to the location of the target region. It would be a reverse complementary notation of the column.

As a specific example, in the case of IL-1β, the human IL-1β gene (61bank access Referring to the nucleotide positions in Accession No. M15840), these sites are As shown below.

Splice acceptor, nucleotide position 445/446:908/90 9; Donano Yank/Yono 970/971; 1535/1536; 1587/ 1588:3575/3576;3777/3778;4322/4323:4 487/4488:572215723;585315854;3569/6 570 poly A addition nognal nucleotide position 7367-7372 start codon Nucleotide position 924-926 mRNA cap site Nucleotide position 374 As a specific example of a preferred site, in the case of IL-1α, human TL-1α gene If you refer to the sequence number of the gene (Genbank accession number X03833), this Their sites and nucleotide positions are as follows.

Splice acceptor, nucleotide position 1488/1489:2152 /2153: Donor/Yank/Yo/2207/2208:3165/3166. 3214/3215:4102/4103+4325/4326:6261/6 2626432/6433; 7814/7815; 7939/7940:102 89/10290 Poly A addition nognal nucleotide position 11618-116 23 start codon nucleotide position ffi 2161-2163 mRNA cap Site Nucleotide position 1438 The site shown below is the preferred site of 1clL-1ra. is an example of a new site, and the site and nucleotide position are those of the human IL-1ra gene. Intracellular splice variants of the gene (Genbank accession number M55646) It is related to

Start codon nucleotide position 123-125 More preferred target site is IL -1ra gene mRNA cap site or splice/yanokunoyono Ru.

As a specific example of a preferred site, in the case of IL-1β converting enzyme, human IL-1β gene The site of the child converting enzyme (Genbank accession number M 87 D O7) and Nucleotide position 1 is as follows.

Start codon nucleotide position 18-20 poly A nognal nucleotide position f fi 1316-1321 or 1335-1340 Furthermore, Ike's preferred site is the mRNA cap site of human IL-1β converting enzyme. or include splices.

Thus, in a preferred embodiment of the present invention, IL-1β, IL-1α, 1c The above nucleotide positions described for IL-1ra or IL-1β converting enzyme Will it rapidly hybridize to sites adjacent to the target site, as determined by the or an arrangement that hybridizes to cover part or all of the site. Oligomers of suitable length (about 8 to 40 nucleotides are preferred) to have a sequence of (with about 12 to 20 nucleometes being more preferred).

When using an antisense oligomer, the sequence of the oligomer is similar to that of the target region. This is a reverse Aiura arrangement.

When using third-strand oligomers, a sequence-specific hydrogen bonding phase is established between the double-stranded nucleic acid target and the double-stranded nucleic acid target. Oligomers are selected to form interactions.

When using a 3m helical oligomer pair, - sequence-specific hydrogen bonding between the heavy chain nucleic acid The first and and a second oligomer.

(Margin below) 3. Preferred treatment instructions Preferred treatment instructions include (a) cutaneous, benign overgrowth: (b) cutaneous, malignant (C) epithelial, benign hyperproliferation; (d) epithelial, malignant hyperproliferation; and ( e) Concerning conditions that can be classified as non-epithelial hyperproliferation.

Skin conditions where excessive proliferation of the epidermis leads to symptoms include dry skin, ichthyosis, hives, rub ra pilaris (PRP), chronic dermatitis, psoriatic dermatitis, atopic Dermatitis, viral eaves cell tumors (warts), other benign growths, chronic lichen pruritus, This includes, but is not limited to, fungiform fungus/5ezary syndrome. do not have.

Malignant skin hyperproliferation of epidermal keratinocytes includes lepidoma, basal cell tumor, ultraviolet They include linear keratoses, keratocytoma, and epithelial neoplasms of the skin of the pond.

Non-malignant hyperproliferative conditions of epithelium include oral mucosa, vagina, cervix, esophagus, lung and gastric II hyperplasia. These include growth and dysplasia, pharyngeal papillomas and bladder cysts.

Epithelial, non-epidermal epithelial cells in malignant deafness include squamous cells and dendritic structures in the head and neck, lungs , including epithelioma of the intestines, breasts, bladder, cervix, uterus, and vagina.

Non-epithelial hyperproliferation may also respond. These include rheumatoid arthritis, polycystic kidney disease, disease, restenosis, and fibrosis of various organs. Non-epithelial cancer is also a target. Dew.

In addition, the method of the present invention can also be used to prevent the prophylactic expression of the IL-1 family. This drug secondarily inhibits the release of cytokines from the skin, and as a result, symptoms are improved. It can also be used to treat certain conditions. Such conditions include rheumatoid arthritis, multiple Forces that include sexual sclerosis, inflammatory bowel disease, grape pancreatitis, or Inuda's disease (these 1 The fundamental points that are essential to this limitation are that Inuda has an immunological abnormality in which the force λ is unregulated. The second possibility is that it is caused by the proliferation of narlatinocytes. I L-11: Shows activity in a wide range of narrow capsule types, and has no pathological effects seen in dry snoring. vinegar. In addition, IL-1: A node failure is dry! It has been reported by 11 organizations, and the :: There can be consequences. E coo/(-ra(Cooper, ni, D, et at,) J , Invest, Dermatol, 95.245-265 (1990) ]. IL-1α levels in psoriasis tissues are ~1/10 lower than in normal tissues. ), IL-1β levels have been reported to be increased (~2x) [Cooper Cooper.

K, D, et al,) J, Immunol, 144.4593-460 3 (1990)]. In addition, IL-1 receptor antagonists showed no inhibition in Inita's skin. It is involved in overwhelming IL-1 inhibitory activity [Kin et al. l, )”Psoriatic 5kin Reveals the in vivo Presence@ofthe Epidermal IL-I Inhibitor+ ^rch, DerII Iatol, Res, 1991 (in Press) Intracellular forms of L-1 receptor antagonists are expressed in normal epidermis and in Inuda epidermis. It was reported that functional suppression was carried out [Hammerbarb et al. berg, C.

et al, )”Interleukin-I Receptor Ant agonist in Normal and Psori≠Kaname@Epid ermis''J, C11n, Invest, 1992 (in Pre ss) Ko. In the epidermis, all members of IL-1 are released, with few exceptions. (Exists within cells and is not secreted. Cytokines produce both precursors and products. can exist as However, IL-1β in normal or xerotic tissues is When measured on the basis of cell activation, it is converted into an inactive form. this Their results showed that I in the control of extracellular or intracellular proliferation of keratocytes Suggests a potential autothalinic role for L-1 and/or IL-1ra.

Furthermore, the intracellular concentrations of IL-1α, IL-1β, and IL-1ra are When cells enter the proliferation cycle, they increase in size, and conversely, an abnormal keratinocyte population appears on the dry skin. The presence of S is associated with a rapid decrease in IL-1α, TL-1β and IL-1ra Phase inhibition occurs [HamIIerberg et al. et al.) Abstract, J, 1. D,, ^pri1. 1992 Ko .

IL-1α, IL-1β and 1clL-1ra in keratinocyte proliferation Few studies examine the functional significance of gene products to assess their role within cells. One alternative method is provided by antisense oligomers. Gene expression Due to genetic limitation, genes that are not only related to each other, but also highly related to each other, can also be specifically distinguished. Two different keratinocyte cultures were was used in research. Isolate the epidermis from normal human skin and culture keratinocytes. A normal keratinocyte culture was obtained by culturing. [Birdsgaard et al. (B aadsgard, 0. et al,) J, Invest, Dermat ol, 95:275-282 (1990). Other cultures■ is a keratinocyte cell line (cell 1i) that retains the properties of gelatinocytes. ne), which shows terminal differentiation (terIIin) when transplanted into nude mice. al differentiation), forming onychokeratosis [boo]. Boukamp et al. J, Ce11. Biol.

106:761-771 (1988)].

b. Inhibition of keratinocyte proliferation Inhibition of gene expression of IL-1β, IL-1a and IL-1ra Studies using oligomers complementary to parts of IL-1α and IL-1ram RNA It was done. Two different keratinocyte cell lines were used in these studies. normal case Latinocyte cell culture involves separating the epidermis and culturing keratinocyte cells. derived from normal human skin [Baadsgard et al. l,, supra. The second cell culture consists of keratinocytes that retain the properties of keratinocytes. It is a nocyto cell line, and when used as a nude mouse (HacaT cell line) cell type, the terminal Vine undergoes terminal differentiation and forms onychokeratosis (Barkamp, et al., (cited above).

The antisense oligomers used in these studies can be used to target individual gene target mRNAs. or towards the start codon of the splice jar/knoyon, making them Table 1 shows the pond sequences used to evaluate sequence specificity.

Todoroki-± 2 [14zz) Human IL-1β CAC-CTG-MT-MA-AAG X -1 lie x 11 ’ C-’! TG-AeT-TCT Ri&=+Nn4 [L48) l l: To XL-L G(:C-ATG-GGG-AGG-GCC open 1g copy Example 2 and By reviewing the results reported in Section 3 and 3, we determined that IL-1α or Expression of IL-1ra or IL-1ra in keratinocytes is essential for their proliferation. It seems like it's for use. Constitutive production of these cytokines by keratinocytes Due to their current and intracellular location, antisense oligomers are novel and sequential. It seems that synthesis is taking place.

Proliferation of keratinocytes in culture using sequence-specific antisense oligomers The fact that these oligomers can limit or reduce demonstrated to be useful in limiting keratinocyte proliferation in urticaria There is. In addition to IL-1α and IL-1β, keratinocytes are involved in the inflammatory process of dermatitis. leads to cells & moisture (also produces many other cytokines. Keratin in Iida) The site releases remaining cytokines that activate T-cells. [Chan( Chang, E, Y, )”T-cell Activation is Potentiated by Cytokines qe1eased by Regional Psoriatic, But Not NorlIa l, Epider++is, +personrch,, Dermatogeki A, Su bmitted, 1992)]. Inhibition of keratinocyte proliferation also affects the immune composition of the disease. The reason for this is that the IL-1 group is expected to have an effect on T-cells in Iida. as well as being one of several synergistic cytokines involved in the activation of , [, using antisense or tertiary antisense oligomers against -1β or IL-1α It also induces other cytokines by changing the intracellular cytokine environment. The reason is that it can inhibit the release of

To aid in understanding the invention, the following examples are set forth which present the results of a series of experiments. . Of course, the following examples relating to the invention are not to be considered as particularly limiting the invention. currently known or later developed that is within the conceivability of a person skilled in the art. Modifications are included within the scope of the invention as defined in the claims below.

Neutral methylphosphonate oligomers were prepared by chemical methods [Miller et al. 1illeret al,) Nucleic Ac1ds Res,, 1 1: 6225-6242 (1983); Jaguar (＾　Jage Chichij and Enfels (J, Engels), Tetrahedron Le tters 25: 1437-1440 (1984) and Thoman et al. A, Dorman et al,) Tetrahedron Lett ers 40°95-102 (19W 4): Synthesized using methylphosphonamidite monomer.

Solid-phase synthesis is performed using Milligen Model 8800 DNA Synthesis Synthesis Sound 2. I went. The programs used for the synthesizer are MTHL 06 (main) and CP L, AW 11 (coapling), these are provided by the vendor. was provided.

The sample mixture used is as follows.

] Activator Acetonitrile 90.45M Tetrazole 2, CapA Aceto 40% acetic anhydride 3 in nitrile, CapB 0625% in pyrinone/methylamino Pirituno 4, Deprotecting agent 2.5% dichloroacetic acid in dichloromethane 5 Oxidizing agent: Te 0.1M ft/2.6-lutidine in trahydrofuran Water (74,82/2510.18; v/v/v) 6 Cleaning liquid A・30 pI) f Acetonitrile 7 Washing liquid B containing water of fl or less: 301) Water of p11 or less Acetonitrile containing 8, monomers: All monomers are acetonitrile 00 Diluted to 8M 9. Support: Oligomers were derivatized with appropriate nucleosides. Synthesized using acrylate bead support The crude, protected methylphosphonate oligomer was purified by acetonitrile/ethano ammonium hydroxide (45/45/10; v/v/v) at room temperature. It was removed from the solid support by stirring for 30 minutes. Then, an equal volume of ethi Protected from base by adding diamine (high quality) and treating for 6 hours at room temperature. The protecting group was removed. The resulting solution was diluted 10 times with water and then neutralized with glacial acetic acid.

The solution containing the oligomer was prepared as per the manufacturer's instructions in 5ep-PakTll. C18 cartridge 01i11ipore/maters Bedford, MA). Wash the column with water and remove the oligomer with 50% acetonitrile in water. Elute.

The methylphosphonate oligomer can be further processed, e.g. Purify by tography: Beckman System Gold HPL C, What+man RACII 0DS-2 column (5 μ, 9 mm i, d, x 100 mm length). Buff yA=501! M-triethyl Ammonium acetate) (pH 7): Buffer B = 50% acetonitrile 1 50mM) ethyl ammonium acetate (pH 7). 10% acetonitrile / Load the sample dissolved in water onto the column using an external pump.

The column was then mounted on a [16d@an FIPLC system and 0-20% Buffer B gradient end for 5 minutes, then 20-60% Buffer B Flow Gradiend for 40 minutes at a flow rate of 3.0 ml/min. Collect fractions and complete length The fraction containing methylphosphonate oligomers was collected and distilled under reduced pressure to reduce the concentration to 50%. Resuspend in acetonitrile/water.

Literature on growth inhibition of normal keratinocytes using antisense oligomers [Barzga] Baadsgaard et at, J, Invest, Der matol, 95:275-282 (1990)]. Human keratinocytes were obtained, and keratinocyte growth medium (C1onetics, S an　Diego,　C^).

Oligomer stocks in 50% acetonitrile/water were added to the culture medium unless otherwise specified. It was diluted at least 100 times in water to 100 μM once. Acetonitrile in the medium The highest concentration of 100% was 0.05%.

Cells were placed in a 96-well cell culture plate with 3xlO” cells per well. plated. Cells were maintained at 37°C in a 002 incubator. proliferation After 4 days, add 3H-dT (1 μCi) and label at 6 p.m. unless otherwise specified. It was introduced for a while. Collect the acid-precipitable label and count it in a non-chloride/yellow counter. Ta.

Six wells (replicates) were set up for each treatment or control. 6 Excluding the highest and lowest counts among the four replicates, the remaining four replicates The mean value and error of the values were calculated.

Freshly obtained normal keratinocytes cultured in Enox vipo (ex viv0) Methylphosphonate (MP) antisene against IL-1α or IL-1β Treatment with oligomers has an effect on proliferation when cytokines are targeted. In order to evaluate this, we varied the evaluation time. The results showed that IL-1α or ■ Normal keratinocytes in culture by targeting either L-1β. (Table 11) (measured by incorporation of 'H-thymine into DNA). The degree of inhibition is dependent on the length of treatment, and antisense-treated cells A reduction in the growth rate of the cells was demonstrated.

Table II 7 2, LSt ± 1,476 --- 7 662 ± 231 69% 7 861±21a 6ot 4L, L4:L amount 11) 57 7 552 * 811 70% l° ・Ratio to control on the same day To test the sequence specificity of growth inhibition, IL-1β oligomers (1252) and A methane that has the same base composition but the positions of the two nucleotides in the molecule are exchanged. phosphonate (“MP”) oligomers; We synthesized oligomers that were predicted to have low or no affinity for oligomer was incubated with normal keratinocyte cultures at 100 μM for 4 days. ■L -1β [oligomer 1 (1252)] The normal inhibition of keratinocyte proliferation was 54%, but mismatched oligomers [oligo MarCI (1480)] did not inhibit proliferation and within the standard deviation of this analysis. showed increased uptake (Table III). Normal keratino seen in oligomer 1 Inhibition of site proliferation is sequence-specific, with oligomer 01 targeting two nucleotides within the sequence. By exchanging the compound, any inhibitory activity of the added oligomers disappeared.

vs. PlI 4 days [1,02B 1 1,169 --- (oligomer - None) To obtain a continuous cell source, we used the human keratinocyte established cell line HaCaT. was also used. IE-conjugated systems retain some of the properties of keratinocytes and are of particular interest. When using immune-deficient mouse nude mouse (HacaT cell line) cell type, Boukamp et al. (Boukamp, P., e. t, al,) J, Ce1l.

Biol, 106: 761-771 (19g!()co.

The Ca T cell line was maintained in DSIEM containing 10% fetal serum. Ta.

Ori: de-Stonok was preserved and translated as described in Example 2. shake cells Incubate, label, and precipitate the label as described in Example 2. It was measured separately.

ILL-β oligomer (oligomer 1) The concentration dependence of HaCaT cell inhibition was determined using mismatched oligomers. It is shown in Table IV along with that of CI. Dose-dependent antisense ILL-β ori Although HaCaT cell inhibition by gomer (oligomer 1) was observed, two oligomers No decrease in proliferation was observed for oligomer 01 with reotide mismatch. It was. In the keratinocyte cell line, similar results were observed in normal human keratinocytes. , inhibition by antisense ILI-β [oligomer 1(1225)] was observed. 2 of the HaCaT keratinocyte cell line with MP-ligoma against LI-β. 5: LS7,513: 10.142 19% m2,5 150.205 : 14498 1 [11t6.2 1” J Yu, s73 ± 6,589 27 25 LE19. Eli 3 engineering 4,653 story 12.5 1fill, 14f amount 1 5.618 4lg, 2 i91,57 ± 6.5BB c Example 4 Oligomers and HaCaT cells were treated as described in Example 3.

IL-1α oligomerization of HaCaT cells after 4 days with 0-100 μM oligomer The concentration dependence of gomer [oligomer 3 (1251-3)] treatment was determined using two nucleotides. antisense IL-1α oligomer [oligomer C3 (15 88-1)].

The role of IL-1α in the inhibition of HaCaT cell line growth observed in the second experiment. The clarity is lower than that described in Example 2, which uses normal keratinocytes. won. Even when antisense IL-1α oligomer (oligomer 3) was added, H aCaT cell development was not significantly reduced, which was similar to the mismatched control oligomer C3 and the effect They were similar in terms of This finding is based on the HaCaT cell line and cultured human keratinocytes. This reflects the difference between the

Oligomers and cells were treated as described in Examples 2 and 3.

Intracellular IL-1 receptor antagonist (ic IL-1ra) is antisense Targeted to MP olicomer, its role in keratinocyte proliferation has been investigated. te990)). Intracellular LL-1ra is unique in that IL- It has a start codon region that is not unique to the soluble form of 1ra. child region is a unique target for distinguishing between intracellular and soluble forms of gene products give. antisense MP oligomer against 1clL-1ra in normal human When incubated with keratinocytes, growth was strongly inhibited by >90%. 1 cL-1ra and IL-1β are highly homologous in their amino acid sequences as protein derivatives. 1clL-1ra has similar intracellular functions as IL-1β. There is a possibility [Klarch, C.J., et al., Nature (Lon d,) 315+641 (1985)],] treatment c IL 1 Nitrogen ■ may not play a role as a receptor antagonist within the cell.

Control (no oligomer) 2 6.630±1.218 ---4 13.98 1 soil 2,722 --- 4 2 479±7892% [IL-1ra Intracellular splice variant 4293±1897% (1487-2)] 1) Comparison to same-day control Different degrees of oligomer 4 (1487) were used with HaCaT cells to The role of 1clL-1ra in nocyte cultures was investigated (see Table Vl).

The antisense oligomer (oligomer 4) against ic'IL-1ra had the highest The lower concentration inhibited proliferation by 60% after 4 days of incubation. However, ori Decreasing Gomer 4 by 1 degree showed no reduction, and 125 and 6.25 μM oligo A rapid change in its efficacy occurred during the Ma. The highest concentration of oligomer 4 The inhibition of HaCaT cells in this study is in line with that observed using normal keratinocyte cultures. The results were similar to those observed in the survey (Table 7). 4 converted from oligomer 4 (1487) Oligomer C4 (1607) of mismatched oligomers with nucleotides did not inhibit nocyte proliferation. Table V and Table Vl show that 1clL-1ra is keratin demonstrated that it can play a functional role in regulating cell proliferation. Tsuki d drunk Junjibi tanyo Domeiko Enshu Oeshu ball hit Inhibition of HaCaT cells by Control (no oligomer) 109.264±1634 100 49, 518±7 ．． 840 62% [I L-1r a 50 120.073±11.537 8% intracellular splice variant 25109.852±26.472 16% (+ 487-3) Ko 12.5 97.371 ± 35.155 25% 6.2 166.954±9.994 (+28%) C4100153,711±37 ．． 747 (418%) [I L-1r a 50 158.493±6.70 0 (+20%) Intracellular splice variant 25 170.470 Sat 17.7 Disagreement (1607-1) 1 12.5 139.9 for 67 (+31%) 90±30.983 (+8%) 6.2 186.836±26.101 ( 443%) Continuation of front page (51) Int, C1, 6 identification symbol Internal office reference number A61K 31/70 A.C.J. 48100 8314-4C C12N 15109 (72) Inventor: Hammerberg, Craig, Michigan, USA 48159 , 17241 English Road I, Manchester (72) Inventor: Maxwell, Cameron W. United States 920 67 California, Rancho Santa Hue, Avenida Calma 1603AM (72) Inventor Tseng, Peng Wai Cabston Do, San Diego, California 92130, United States Live number 1325

Claims (40)

[Claims] 1. (a) Complementary to the RNA sequence transcribed from the target gene present in the cell antisense oligomer having the sequence, (b) selection of target genes present in cells; a third strand oligomer having a sequence complementary to the selected double-stranded nucleic acid sequence, and (c) A triple helix oligomer complementary to a single-stranded nucleic acid sequence of a target gene or its transcript An excessive proliferation-inhibiting amount of an oligomer selected from hif or epithelium, characterized by reducing hyperproliferation of cells by exposure to A method of treating a pathological condition characterized by hyperproliferation of cells, the method comprising: Genes are linked to cytokines, cytokine regulators, and cytokines that mediate cell proliferation. The function of converting enzymes or cytokines that convert precursors of in to active cytokines. encodes an enzyme involved in translation or post-translational modification of the cytokine, which is important for A method in which genes are selected from among 2. The target gene is one that regulates cell proliferation through an intracellular mechanism. The method described in claim 1. 3. The target gene is 1L-1α, 1L-1β or icIL-1ra or IL 3. The method of claim 2, wherein the method encodes a -1β converting enzyme. 4. The method of claim 2, wherein the pathological condition includes benign hyperproliferation of keratinocytes. . 5. The method of claim 2, wherein the pathological condition includes malignant hyperproliferation of keratinocytes. . 6. 3. The method according to claim 2, wherein the pathological condition includes benign hyperproliferation of non-epidermal epithelial cells. Law. 7. 3. The method according to claim 2, wherein the pathological condition includes malignant hyperproliferation of non-epidermal epithelial cells. Law. 8. 3. The method according to claim 2, wherein the condition is psoriasis. 9. The pathological condition is regulated by 1L-1α, 1L-1β or ic1L-1ra production 3. The disease is a chronic inflammatory disease in which cytokine release is persistent. Method. 10. The chronic inflammatory disease may include rheumatoid arthritis, inflammatory bowel disease, psoriasis, and inflammation. The method according to claim 9, wherein the method is selected from among sexual eye diseases. 11. 2. The method of claim 1, wherein the oligomer is a neutral oligomer. 12. 2. The method of claim 1, wherein the oligomer is an antisense oligomer. 13. Claims that the oligomer is a third strand oligomer or a triple helix oligomer pair The method described in Section 1. 14. (a) Complementary to the RNA sequence transcribed from the target gene present in the cell (b) antisense oligomer with a sequence of the target gene present in the cell; a third strand oligomer having a sequence complementary to the selected double stranded nucleic acid sequence, and (c ) a triple helix oligomer complementary to the single-stranded nucleic acid sequence of the target gene or its transcript; - to human or epithelial cells, an amount of an oligomer selected from Preventing or reducing hyperproliferation of human or epithelial cells characterized by exposure to A method, wherein the target gene is a cytokine that mediates cell proliferation. regulator of cytokines, a converting enzyme that converts cytokine precursors into active cytokines , or translational or post-translational modifications of the cytokine that are important for the function of the cytokine. genes encoding enzymes involved in. 15. The cytokine or receptor antagonist acts on cells through intracellular mechanisms. 15. The method of claim 14, wherein the method mediates cell proliferation. 16. 15. The method of claim 14, wherein the oligomer is a neutral oligomer. 17. 15. The method of claim 14, wherein the oligomer is an antisense oligomer. 18. 15. The method of claim 14, wherein the oligomer is a third strand oligomer. 19. A method of preventing or reducing cell hyperproliferation, the method comprising: -1 or an agent that decreases intracellular interleukin-1 receptor antagonist activity. the cell is brought into contact with a growth-inhibiting amount of a noncoleukin-1 inhibitory compound. how to. 20. 20. The method of claim 19, wherein the cells are epithelial cells. 21. The interleukin-1 inhibitory compound prevents interleukin-1 expression compound that increases or decreases or converts a precursor of interleukin-1 to an active form A method selected from among compounds that inhibit or reduce. 22. The interleukin-1 inhibiting compound is a compound of interleukin-1 converted oxygen. or inhibit the activity of interleukin-1 converting enzyme. 22. The method according to claim 21. 23. The interleukin-1 inhibitory compound inhibits interleukin-1, interleukin-1, Preventing or reducing expression of leukin-1 regulatory factor or interleukin-1 converting enzyme 20. The method according to claim 19, wherein the oligomer is 24. Interleukin-1 inhibitory compounds inhibit intracellular interleukin-1 receptor activation. 20. The method of claim 19, wherein the method is selected from compounds that reduce antagonist activity. . 25. (a) Complementary to the RNA sequence transcribed from the target gene present in the cell (b) antisense oligomer with a sequence of the target gene present in the cell; a third strand oligomer having a sequence complementary to the selected double stranded nucleic acid sequence, and (c ) a triple helix oligomer complementary to the single-stranded nucleic acid sequence of the target gene or its transcript; - an excessive growth-inhibiting amount of the oligomer selected from A pharmaceutical composition comprising a carrier in which the target gene is a carrier that mediates cell proliferation. cytokines, cytokine regulators, and cytokine precursors to active cytokines. A converting enzyme that converts into a cytokine or a cytokine that is important for the function of the cytokine. selected from genes encoding enzymes involved in translation or post-translational modification of composition. 26. The target gene is 1L-1α, 1L-1β, ic1L-1ra or 1L-1 26. The composition of claim 25, encoding a β-converting enzyme. 27. 26. The composition of claim 25, wherein the oligomer is a neutral oligomer. 28. 28. The composition of claim 27, further comprising a flux promoter. 29. The target gene is 1L-1α, 1L-1β, ic1L-1ra or 1L-1 28. The composition of claim 27, encoding a β-converting enzyme. 30. 30. The composition of claim 29 further comprising a flux promoter. 31. Interleukin-1 or intracellular interleukin-1 receptor antagonist Excessive growth-inhibiting amount and production of interleukin-1 inhibitory compounds that reduce anti-steroid activity A pharmaceutical composition containing a pharmaceutically acceptable carrier. 32. The interleukin-1 inhibitory compound prevents the expression of interleukin-1. Compounds that inhibit or reduce interleukin-1 precursors or convert them to active forms 32. The composition of claim 31, wherein the composition is selected from compounds that inhibit or reduce . 33. The interleukin-1 inhibitory compound inhibits interleukin-1 converting enzyme. inhibit or reduce the synthesis or inhibit the activity of in-leukin-1 converting enzyme 32. The composition according to claim 31. 34. Interleukin-1 inhibitory compounds reduce intracellular 1L-1ra activity 32. The composition according to claim 31, selected from among the compounds. 35. Interleukin-1 inhibitory compound is interleukin-1, interleukin-1 Preventing or reducing the expression of Kin-1 regulatory factor or Interleukin-1 converting enzyme 32. The composition according to claim 31, which is an oligomer that can be used as an oligomer. 36. 36. The composition of claim 35, wherein the oligomer is a neutral oligomer. 37. 36. The composition of claim 35, further comprising a flux promoter. 38. 32. The interleukin-1 inhibitory compound is an oligomer. Composition of. 39. 39. The composition of claim 38, wherein the oligomer is a neutral oligomer. 40. 40. The composition of claim 39 further comprising a flux promoter.