Use of polypeptides or nucleic acids encoding these of the gene family NM23 for the diagnosis or treatment of skin or intestinal disorders, and their use for the identification of pharmacologically active substances

Method of using of polypeptides or nucleic acids encoding these, of the gene family NM23, for the analysis and/or diagnosis and/or prevention and/or treatment of disorders of skin and/or intestinal disorders and/or wound healing and/or disorders of wound healing and/or for the identification of pharmacologically active substances.

SEQ ID No. 1 to SEQ ID No. 10 show the poly-peptides used according to the invention from human or mouse. SEQ ID No. 11 to SEQ ID No. 14 and SEQ ID No. 17 to SEQ ID No. 26 show DNA sequences of oligonucleotides which were used for the experiments of the present invention. SEQ ID No. 15 to SEQ ID No. 16 show DNA sequences of NM23 which were used for the preparation of probes for the RNase protection assay and in situ hybridization. 
 EXAMPLES 
 Example 1 
 Enrichment of Wound-Relevant cDNA by Means of Subtractive Hybridization and Identification of NM23-M1 as Wound-Relevant Gene Total RNA was isolated from intact skin and from wound tissue (wounding on the back 1 day before tissue sampling by scissors cut) of BALB/c mice by standard methods (Chomczynski and Sacchi, 1987, Anal. Biochem. 162: 156-159, Chomczynski and Mackey, 1995, Anal. Biochem. 225: 163-164). In order to obtain tissue of mice with poorly healing wounds, BALB/c mice were treated before wounding with dexamethasone (injection of 0.5 mg of dexamethasone in isotonic saline solution per kg of body weight twice per day for 5 days). The RNAs were then transcribed into cDNA with the aid of a reverse transcriptase. The cDNA synthesis was carried out using the “SMART PCR cDNA synthesis kit” from Clontech Laboratories GmbH, Heidelberg, according to the directions of the corresponding manual. In order to identify those cDNAs which occurred with differing frequency in the cDNA pools, a subtractive hybridization (Diatchenko et al., 1996, Proc. Natl. Acad. Sci. USA 93: 6025-30) was carried out. This was effected using the “PCR select cDNA subtraction kit” from Clontech Laboratories GmbH, Heidelberg, according to the directions of the corresponding manual, the removal of excess oligonucleotides after the cDNA synthesis being carried out by means of agarose gel electrophoresis. Four cDNA pools were set up, which were enriched for wound-relevant genes, where one pool was enriched for cDNA fragments which are expressed more strongly in the wound tissue in comparison with intact skin (“wound-specific cDNA pool”), one pool was enriched in cDNA fragments which are more strongly expressed in intact skin in comparison with wound tissue (“skin-specific cDNA pool”), one pool was enriched in cDNA fragments which are more strongly expressed in normally healing wounds in comparison with poorly healing wounds (“normally healing cDNA pool”) and one pool was enriched in cDNA fragments which are more strongly expressed in badly healing wounds in comparison with normally healing wounds (“badly healing cDNA pool”). In order to identify those genes which were contained in the cDNA pools relevant to wound healing, the presence of the corresponding cDNAs in the pools was analyzed by “reverse Northern blot”. Here, the cDNA fragments are immobilized on membranes in the form of arrays of many different cDNAs, and hybridized with a complex mixture of radio-labelled cDNA (Sambrook et al., 1989, Molecular Cloning: A Laboratory Manual, Cold Spring Harbor, Cold Spring Harbor Laboratory Press, New York, Chapter 9 page 9.47 to 9.58 and Chapter 10 page 10.38 to 10.50; Anderson and Young: Quantitative filter hybridization; in: Nucleic Acids Hybridization, A Practical Approach, 1985, Eds. Hames and Higgins, IRL Press Ltd.; Oxford, Chapter 4, page 73 to 112). For the preparation of suitable hybridization probes, the subtracted cDNA pools were treated with the restriction endonuclease RsaI and purified by means of agarose gel electrophoresis (Sambrook et al., supra, Chapter 6, page 6.1 to 6.35) in order to separate the cDNA synthesis and amplification primer (see manual “PCR-Select cDNA Subtraction Kit”, Clontech). The cDNAs were then radio-labelled using the “random hexamer priming” method (Feinberg and Vogelstein, 1983, Anal. Biochem. 132: 6-13) in order to prepare hybridization probes. The membrane was preincubated in 25 ml of hybridization solution for 30 min at 65° C. (25 mM sodium phosphate, pH&equals;7.5, 125 mM NaCl, 7% SDS). The hybridization probe was denatured at 100° C. for 10 min, then cooled on ice, about 100 CPM per ml were added to the hybridization solution and the hybridization was carried out in a hybridization oven for 16 hours at 65° C. The membrane was then washed twice with the hybridization solution without probe at 65° C. for 10 min. The membrane was then washed at 65° C. several times for 10 min with wash solution (2.5 mM sodium phosphate, pH&equals;7.5, 12.5 mM NaCl, 0.7% SDS) until it was no longer possible to detect any activity in the wash solution poured off. The radioactive signals were analyzed using a phosphoimager (BioRad, Quantity One®) ( FIG. 1 ). Those cDNAs were then selected which produced different signal intensities with the various probes. This resulted at the position of NM23-M2 on the membrane, in a slightly stronger signal intensity with the hybridization probe of the wound specific cDNA pool in comparison to the skin specific cDNA pool and a clearly stronger signal intensity with the hybridization probe of the poorly healing cDNA pool in comparison to the normally healing cDNA pool. 
 Example 2 
 Confirmation of the Expression Pattern of NM23-M2 by Means of “Real-Time Quantitative RT-PCR” A confirmation of the differential expression of the nucleic acids used according to the invention was carried out by real-time RT-PCR in the ABI Prism 7700 sequence detection system (PE Applied Biosystems). The apparatus was equipped with the ABI Prism 7200/7700 SDS software version 1.6.3 (1998). The detection of PCR products was carried out during the amplification of the cDNA with the aid of the stain SYBR Green 1, whose fluorescence is greatly increased by binding to double-stranded DNA (Karlsen et al. 1995, J. Virol. Methods. 55: 153-6; Wittwer et al., 1997, BioTechniques 22: 130-8, Morrison et al., 1998, BioTechniques 24: 954-62). The basis for the quantification is the PCR cycle (threshold cycle, C T -value) which is reached when the fluorescence signal exceeds a defined threshold. The analysis is carried out by means of the &Dgr;-C T method (User Bulletin &num;2, Relative Quantification of Gene Expression, PE Applied Biosystems, 1997). The abundance of the cDNAs were determined relative to an endogenous reference (GAPDH). The results are shown in FIG. 2 . Total RNA pools from skin and wound tissue from 16 animals each was obtained as described above and 1 &mgr;g of total RNA was subjected to reverse transcription in a thermocycler (GeneAmp PCR system 9700, PE) using the TaqMan reverse transcription reagent kit (PE) according to the recommendations of the manufacturer (SYBR Green PCR and RT-PCR Reagents Protocol, PE Applied Biosystems, 1998). The primers for the amplification of the NM23-M2 cDNA (NM23-Primer 1: TTCAAAACCAGGCACCATCC (SEQ ID No. 11), NM23-Primer 2: ACTCTCCACTGAATCACTGCCA (SEQ ID No. 12) and the reference (GAPDH primer 1: ATCAACGGGAAGCCCATCA (SEQ ID No. 13), GAPDH primer 2: GACATACTCAGCACCGGCCT (SEQ ID No. 14)) were selected with the aid of the Primer Express software for Macintosh PC Version 1.0 (PE Applied Biosystems, P/N 402089, 1998) based on the nucleic acid used according to the invention and the known sequence of GAPDH. For the PCR, the SYBR Green PCR Core Reagents Kit (4304886, PE Applied Biosystems) was used. The concentration of the primers in the PCR was initially optimized in the range from 50 nM to 600 nM and the specificity of the PCR was verified by analysis of the length of the amplified products by agarose gel electrophoresis. The efficiency of the PCR system was then determined by means of a dilution series (User Bulletin &num;2, Relative Quantification of Gene Expression, PE Applied Biosystems, 1997). It became apparent that for both cDNAs the efficiency of the amplification was 100%, i.e. at each 1:2 dilution of the cDNA one more cycle was needed in order to exceed the fluorescence threshold value. For the quantification, each batch of cDNA was amplified from 10 ng of reverse-transcribed total RNA in a total volume of 25 &mgr;l. The running conditions for the PCR corresponded to the details of the manufacturer (PE Applied Biosystems, SYBR Green® PCR and RT-PCR Reagents Protocol, 1998). The C T -values were analyzed and the abundance of NM23-M2 relative to GAPDH was calculated. It was possible to confirm the slight induction of NM23-M2 in normally healing wounds and the strong induction in poorly healing wounds of dexamethasone-treated animals ( FIG. 2 ). 
 Example 3 
 Verification of the Expression Pattern of NM23-H1 by Means of “RNase Protection Assays” The expression of NM23-H1 was verified with the aid of the “RNase protection assay”. The test was carried out as described in the literature (Sambrook et al., supra Chapter 7, page 7.71 to 7.78; Werner et al., 1992; Growth Factors and Receptors: A Practical Approach 175-197, Werner, 1998, Proc. Natl. Acad. Sci. USA 89: 6896-6900). Reverse-strand RNA which was transcribed in vitro and radio-labelled was used as a hybridization probe. A NM23-H1 fragment of 266 bp length (SEQ. ID No. 15) was cloned via blunt ends into the EcoRV-restriction site of the vector pBluescript II KS (Stratagene). The plasmid was linearized with XbaI before transcription (length of the transcript without vector sequence: 299 basepairs, sequence of the probe SEQ ID No. 15). The transcriptions were carried out with T3 polymerase (Roche Diagnostics, Mannheim) in the presence of 32 P-UTP (35 &mgr;Ci/batch) (Amersham, Brunswick) according to the details of the manufacturer. The probe was purified by gel electrophoresis and elution (Sambrook et al., supra, Kapitel 6, Seite 6.36 bis 6.48). For the hybridization reaction, about 10 5 CPM each of the labelled transcripts were employed. For this, 10 &mgr;g of total RNA which was isolated from either skin or intestine biopsies using standard methods (Chomczynski and Sacchi, 1987, Anal. Biochem. 162: 156-159, Chomczynski and Mackey, 1995, Anal. Biochem. 225: 163-164) was precipitated together with the transcript, taken up in 10 &mgr;l of hybridization buffer (80% deionized formamide, 400 mM NaCl, 40 mM Pipes pH 4.6, 1 mM EDTA) and hybridized overnight at 42° C. An RNase A/T1 digestion (Boehringer, RNase A: 0.8 &mgr;g/batch, RNase T1: 20 U/batch) was then carried out. After inactivation of the RNase by proteinase K digestion (Boehringer, 30 &mgr;g/batch) and phenol extraction, the samples were precipitated with ethanol according to standard methods (Sambrook et al., supra). The samples were then separated by gel electrophoresis on a denaturing 5% acrylamide gel (7_M urea) . The gel was dried and the radioactive signals were analyzed by means of autoradiography ( FIGS. 3 and 4 ). In the RNase protection assay with RNA isolated from biopsies of 3 different psoriasis and 4 different control patients, an increased expression of NM23-H1 could be observed in the skin samples from all psoriasis patients in comparison to the control skin samples ( FIG. 3 ). The RNase protection assay with RNA from tissue samples derived from 5 persons with Crohn's disease showed an increased expression of NM23-H1 in inflammatory slices of the intestine relative to control subjects ( FIG. 4 ). 
 Example 4 
 Analysis of the Expression of NM23-H1 in Tissue Cuts of Mouse Wounds The expression of NM23-M1 in wounds was analyzed by means of in situ hybridization. The test was performed as described in the literature (Werner et al., 1992; Growth Factors and Receptors: A Practical Approach 175-197, Werner, 1998, Proc. Natl. Acad. Sci. USA 89: 6896-6900). A NM23-M1 fragment of 256 bp length (SEQ. ID No. 16) was cloned via blunt ends into the EcoRV-restriction site of the vector pBluescript II KS (Stratagene). Using this vector, a radiolabelled reverse-strand RNA as hybridization probe was produced and used as described (Werner, 1998, supra). This probe was used to perform hybridizations on frozen tissue slices of 5 day-wounds of mice. It became apparent that the NM23-M1 gene was increasingly expressed in the hyperproliferative epithelium at the woundedge. In contrast, in healthy parts of the skin, a faint to nondetectable staining was observed. 
 Example 5 
 Analysis of Murine Patterns of Expression of NM23-M1 and NM23-M2 mRNAs During Wound Healing by Means of “TaqMan Analysis” The kinetics of regulation of expression of NM23 mRNA's NM23-M1 and NM23-M2 during normal wound healing of the adult mouse was analyzed using “TaqMan Analysis” in GeneAmp5700 of Applied Biosystems. Normally healing wounds and intact skin were taken from 10 week old BALB/c mice using scissors cut as described above. In order to isolate RNA, the biopsies were homogenized in the presence of RNAclean buffer (AGS, Heidelberg) that was supplemented by 1/100 volume fraction of 2-mercapto-ethanol using a disperser. Subsequently the RNA was extracted by a twice repeated phenolization using water saturated acidic phenol in the presence of 1-bromo-3-chloro-propane. Then the RNA was precipitated using isopropanol and ethanol precipitation and the RNA was washed using 75% ethanol. Afterwards the RNA was treated with DNaseI. 20 &mgr;g of RNA (ad 50 &mgr;l DEPC-treated water) was supplemented with 5.7 &mgr;l transcription buffer (Roche), 1 &mgr;l RNase-inhibitor (Roche); 40 U/&mgr;l) and 1 &mgr;l DNaseI (Roche); 10 U/&mgr;l) and incubated for 20 minutes at 37° C. Then another 1 &mgr;l DNaseI was added and the sample was incubated for another 20 minutes at 37° C. Subsequently the RNA was phenolized, ethanol precipitated and washed. All above mentioned steps were carried out using DEPC(diethylpyrocarbonate)-treated solutions and liquids unless they contained reactive aminogroups. Subsequently cDNA was synthesized from the extracted RNA. 20 &mgr;l RNA (50 ng/&mgr;l) were supplemented with 1× TaqMan RT-buffer (Perkin Elmer), 5.5 mM MgCl 2 (Perkin Elmer), 500 &mgr;M dNTPs each (Perkin Elmer), 2.5 &mgr;M random hexameres (Perkin Elmer), 1.25 &mgr;l Multiscribe Reverse Transcriptase (50 U/&mgr;l Perkin Elmer), 0,4 &mgr;l RNase-inhibitor (20 U/&mgr;l, Perkin Elmer) and DEPC-treated water (ad 100 &mgr;l volume). Upon addition of RNA and thorough mixing the solutions were divided into two 0.2 ml tubes (50 &mgr;l each) and the reverse transcription reaction was carried out in a thermocycler (10 min at 25° C.; 30 min at 48° C. and 5 min at 95° C.). The following quantification of cDNA was done by means of quantitative PCR using the SYBR Green PCR Master Mixes (Perkin Elmer), wherein for each NM23 cDNA species to be quantified, a triple determination (each time with target primers and GAPDH primers) was carried out. The stock solution for each triplet contained at a total volume of 57 &mgr;l, 37.5 &mgr;l 2× SYBR Master Mix, 0.75 &mgr;l Amp. Erase UNG(1 U/&mgr;l) and 18.75 &mgr;l DEPC-treated water. For each triple determination the 57 &mgr;l stock solution were supplemented with 1.5 &mgr;l forward- and reverse primer each in a concentration ratio that had been optimized before. Each 60 &mgr;l stock solution/primer mix were mixed with 15 &mgr;l cDNA solution (2 ng/&mgr;l) and divided to 3 wells. In parallel a stock solution with primers for the determination of GAPDH (SEQ ID No. 13 and SEQ ID No. 14) as a reference as mixed with additional 15 &mgr;l of the same cDNA solution and distributed onto 3 wells. In order to obtain a standard graph for the GAPDH-PCR, a dilution series of different cDNA solutions was made (4 ng/&mgr;l; 2 ng/&mgr;l; 1 ng/&mgr;l; 0.5 ng/&mgr;l and 0.25 ng/&mgr;l). For the determination of GAPDH,15 &mgr;l each of the CDNA solutions of the dilution series was mixed with 60 &mgr;l stock solution/primer mix and distributed onto 3 wells. A standard graph for each of the PCRs of the NM23 homologues to be analyzed was obtained; wherein the same dilutions used for the GAPDH standard graph were used. As a control a PCR without cDNA was used. The stock solution/primer mix of each the target and GAPDH were supplemented with 15 &mgr;l DEPC-water, mixed and distributed onto 3 wells. The amplification was performed using Gene Amp. 5700 (2 min at 50° C.; 10 min at 95° C., followed by 3 cycles with 15 s at 96° C. and 2 min at 60° C.; followed by 37 cycles with 15 s at 95° C. and 1 min at 60° C.) . The analysis was done by determining the abundance for each NM23 gene relative to the GAPDH-reference. The standard curve was determined first by plotting the C T -values of the dilution series against the logarithm of the amount of cDNA and the PCR (ng of transcribed RNA) and the slope of the graph was determined. The efficiency (E) of the PCR can be calculated as follows: E&equals;10 −1/s −1. The relative abundance (X) of the NM23 cDNA species (Y) under investigation with respect to GAPDH is: X&equals;(1&plus;E GAPDH ) T C (GAPDH) /(1&plus;E Y ) T C (Y) . Subsequently the values were standardized by equatizing the amount of cDNA in intact skin of adult 10 weeks old BALB/c control animals with 1. The relative changes of expression of NM23-M1 and NM23-M2 respectively at different points of time after wounding of adult mice are depicted in FIG. 7 . Using appropriate primers to detect NM23-M1 (NM23-primer 3: TCC TGG CAC AGT CAG ACA ACA (SEQ ID No. 17); NM23-primer 4: TTC ACA ACC TCA CAC ATC CTC C (SEQ ID No. 18)) and NM23-M2 (NM23-primer 1: (SEQ ID No. 11) and NM23-primer 2 (SEQ ID No. 12)), it could be shown that the expression of both homologues was reduced during wound healing in adult mice ( FIG. 7 ). In the course of the wound healing NM23-M1 expression was overall constantly reduced to about 50% of the amount observed in intact skin. A similar pattern was detected in NM23-M2 expression, where during wound healing between 1 h and 7 d after wounding the expression was reduced considerably relative to the intact skin. Only after d 14 after wounding the expression level rebounded to approximately the original level. Taken together both homologues showed a comparably reduced level of expression over a long time of the wound healing. This result seems to contradict the examples 1 and 2, where a weak increase in the amount of NM23 expression in normal healing wounds could be observed. However, in example 4 it was demonstrated, that there is no overall increase in NM23 expression in wound tissue slices. An increased staining was observed in the hyperproliferative epithelium, but no significant expression was detected in other layers of the tissue. This result implies that wound healing-dependent changes in the amount of mRNA during complex changes of the spatial pattern of expression can only be analyzed in an insufficient way since the sensitivity to detect mRNA is lower in the case of in situ hybridization than in the case of “real time RT-PCR”: due to the complex changes in the spatial patterns of NM23 expression, subtle variabilities during the taking of the biopsies may lead to different results concerning wound healing dependent changes of the amount of NM23 mRNA determined by means of “real time RT-PCR” (example 2) and subtractive hybridization (example 1) as opposed to a determination by means of “TaqMan analysis” (example 5). It was surprising that genes of the NM23 gene family could be identified despite of the difficult conditions. 
 Example 6 
 Differential Expression of NM23-H1 and NM23-H2 in Human Wounds The differential regulation of NM23-H1 in psoriasis and Crohn's disease was analyzed in example 3. Using normally healing wounds it was investigated whether the differential regulation of expression of NM23-M1 and NM23-M2 shown in example 5 can also be observed in humans. Skin samples were taken from untreated intact skin, day-1 wounds or day-5 wounds of healthy subjects by means of isolating 4 mm or 6 mm punch skin samples respectively. For each group (intact skin, day-1 wound, day-5 wound) biopsies of 14 subjects each were pooled. The biopsies were desintegrated in a swing mill and the RNA was isolated as described in example 5, then DNAseI digested and reverse transcribed into cDNA. A quantification of wound healing relevant cDNA was performed as described in example 5. The results of the experiments are depicted in FIG. 8 . For the analysis of NM23-homologues primers for the amplification (hGAPDH-Primer 1: CATGGGTGTGAACCATGAGAAG (SEQ ID Nr. 25); hGAPDH-Primer 2: GCTAAGCAGTTGGTGGTGCA (SEQ ID Nr. 26), NM23-H1 Primer 1: GAAATTCATGCAAGCTTCCGA (SEQ ID Nr. 19), NM23-H1 Primer 2: CAGGTCAACGTAGTGTTCCTTGAG (SEQ ID Nr. 20); NM23-H2 Primer 1: CTGGTTGACTACAAGTCTTGTGCTC (SEQ ID Nr. 21); NM23-H2 Primer 2: TCCACCTCTTATTCATAGACCCAGT (SEQ ID Nr. 22) were selected based on known sequences of human GAPDH (GenBank:M17851) and human NM23-H1 and NM23-H2 (EMBL:X17620 and EMBL:X58965) . cDNA resulting from reverse transcription of 10 ng total RNA was amplified in a total volume of 25 &mgr;l for quantification. PCR was performed according to the instructions of the manufacturer (PE Applied Biosystems, SYBR Green PCR and RT-PCR reagents protocol, 1998). C T -values were determined and the abundance of NM23 mRNA relative to GAPDH-mRNA was calculated. The results of the experiment are depicted in FIG. 8 . It was observed that wound tissue showed a slight decrease of expression. In contrast to the analysis of murine biopsies of example 5 two different points of time were selected. Using human tissue a significant coincidence of the results regarding the kinetics of wound healing of mice and of humans was observed (example 5): compared to the initial value, the NM23-M1 and NM23-H1 expression level in biopsies at day 1 after wounding is reduced by 70% and both mRNA levels show a slight increase to 66% or 70% of the initial value respectively 5 days after wounding. The weaker level of expression of NM23-M2 and NM23-H2 in day-5 wounds is also comparable (48% vs. 60% of the initial value). Thus it could be confirmed that both genes play an essential role in the regulation of wound healing and that the modulation of the amount of at least one homologue, preferentially both homologues can be used for the prevention and/or diagnosis and/or treatment of disorders of skin cells. 
 Example 7 
 Lokalization of NM23-H2 in Biopsies of Intact Skin, of Normally Healing Wounds, Ulcer, As Well As Non-Lesional and Lesional Psoriatic Skin by Means of in Situ Hybridization For the experiment biopsies of healthy skin as well as normally healing day-5 wounds were taken from a healthy subject as described in example 6. In addition biopsies of non-lesional and lesional skin of 10 psoriasis patients each and from intact skin and the wound of an ulcer patient (Ulcus cruris venosum) were taken as described above. The tissue slices were fixed in 4% paraformaldehyd, treated with proteinase K (1 &mgr;g/ml isotonic saline) for 10 min at 37° C., and subsequently treated with paraformaldehyde and then with acetanhydride (0.5 ml in 0.1 M triethanolamine, pH 8,0). The mRNA of human NM23-H2 was localized by radioactive in situ hybridization. Paraformaldehyde fixed slices were embedded in paraffin. The synthesis of the hybridization probe was based on in vitro transcription of a partial NM23-H2 cDNA fragment in the presence of &agr;- 35 S-UTP. In order to obtain the PCR product promoter sequences for the transcription in sense and anti-sense direction were added to the primers. (T3-NM23-H2-primer: AATTAACCCTCACTAAAGGGGGAGGGGCTGAACGTGGTGAAGAC (sense control primers with T3-promoter; SEQ ID No. 23), Sp6-NM23-primer: ATTTAGGTGACACTATAGAATACACGCCGTGCTGAAGGAGACTGC (antisense primer with Sp6-promoter; SEQ ID No. 24). For the in vitro transcription 60 &mgr;Ci 35 S-UTP and 5 mM ATP, GTP and CTP each, as well as either 25 U T3- or T7-RNA polymerase (Roche), 1 &mgr;g PCR-product, 10 mM Dithiothreitol, 40 U RNAse inhibitor (Roche) and 1× TB-buffer (Roche) were used. Human tissue slices (see above) were mounted onto slides, treated with proteinase K, fixed with para-formaldehyde and were subsequently acetylated. The slices were transferred into a humid chamber containing Whatman tissue paper soaked in 50% formamide/4× SSC. The slices were covered with 30 &mgr;l hybridization solution and incubated for 2.5 h at 60° C. Afterwards, the slices were incubated with 30 &mgr;l hybridization solution containing 0.7×10 6 CPM of radioactively labelled riboprobe for 16 h at 60° C. Then the slices were washed under stringent conditions incubated with RNAse A and dehydrated with ethanol. The slices were then covered with photo emulsion (Kodak IBO 1433) in the absence of light and oxygen for 2-6 weeks at 40° C. and subsequently developed using photographic developer and fixative (Kodak IBO 1433) according to the instructions of the manufacturer. No or very weak signals were observed in intact skin of the healthy subject and the ulcer-patient as well as in non-lesional skin of the psoriasis patients. In contrast, tissue slices of normal healing day-5 wounds showed signals in the basal cell layer of the hyperproliferative epithelium. This indicates that the induction of NM23-H2 expression is particularly essential in the cell layer that is responsible for the closure of the wound by means of proliferation and migration. This is consistent with the observation that no significant labelling was detected at wound edge and the wound ground of the non- or badly-healing Ulcer wound. Thus, the wound specific regulation of NM23-H2-expression is essential for the normal process in wound healing. In comparison to intact skin of the healthy subject or non-lesional skin of the psoriasis patients, the lesional psoriatic skin biopsies also showed significantly increased labelling intensity in the basal cell layers of the hyperproliferative epithelium. This is consistent with a result of the experiment of example 3 where it was shown that the amount of NM23-H1 is increased in psoriatic skin. This experiment elucidates that the regulation of NM23 expression is essential for intact, healthy skin as well as for the normal process of wound healing and that a dysfunctional regulation of the expression can lead to disorders of skin cells for example to a delayed wound healing or psoriasis and it shows that NM23, preferentially both homologues can be used for the prevention and/or diagnosis and/or treatment of disorders of skin cells. Badly healing wounds are associated with a reduced amount of NM23, whereas in psoriasis patients, whose keratinocytes are characterised by pathological proliferation, exhibit an increase in the level of NM23 expression. For the treatment of skin cells the expression or activity of NM23 should be modulated, preferentially by activating the expression or activity of NM23 in the case of disorders of wound healing. The activity or expression of NM23 should be preferentially inhibited in the case of hyperproliferative disorders of skin cells, especially in the case of psoriasis. It will be apparent to those skilled in the art that various modifications can be made to the compositions and processes of this invention. Thus, it is intended that the present invention cover such modifications and variations, provided they come within the scope of the appended claims and their equivalents. Priority application DE 10008330.7-41, filed Feb. 23, 2000 and priority application U.S. Ser. No. 60/199,312 filed Apr. 24, 2000. All publications cited herein are incorporated in their entireties by reference.