Patent Document (Category 0):

one aspect of the present invention contemplates a method for modulating hormone signalling in an animal , said method comprising up - regulating or down - regulating expression of a genetic sequence encoding a socs protein or its derivative or homologue or increasing or decreasing the activity of a socs protein or its derivative or homologue in said animal . reference herein to “ socs ” encompasses any or all members of the socs family . specific socs molecules may be defined numerically such as , for example , socs - 1 , socs - 2 and socs - 3 . the species from which the socs has been obtained may be indicated by a preface of single letter abbreviation where “ h ” is human , “ m ” is murine and “ r ” is rat . accordingly , “ msocs - 2 ”, for example , is a specific socs from a murine animal . reference herein to “ socs ” is not to imply that the protein solely suppresses cytokine - mediated signal transduction , as the molecule may modulate other effector - mediated signal transductions such as by hormones or other endogenous or exogenous molecules , antigen , microbes and microbial products , viruses or components thereof , ions , hormones and parasites . the term “ modulates ” encompass up - regulation , down - regulation as well as maintenance of particular levels . reference herein to a “ hormone ” includes protein hormones and cytokines as well as non - proteinaceous hormones . one particularly useful hormone is growth hormone . another useful hormones are insulin - like growth factor i ( igf - i ) and prolactin . an “ animal ” is preferably a mammal such as but not limited to a human , primate , livestock animal ( e . g . sheep , cow , pig , horse , donkey ), laboratory test animal ( e . g . rabbit , mouse , rat , guinea pig ), companion animal ( e . g . cat , dog ) or captive wild animal . the animal may be in the form of an animal model . useful animals for this purpose are laboratory test animals . genetically modifying livestock animals is useful in assisting in food production . the preferred animal is a human , primate animal or laboratory test animal . the most preferred animal is a human . reference herein to “ socs ” includes a protein comprising a socs box in its c - terminal region comprising the amino acid sequence : x 1 x 2 x 3 x 4 x 5 x 6 x 7 x 8 x 9 x 10 x 11 x 12 x 13 x 14 x 15 x 16 [ x i ] n x 17 x 18 x 19 x 20 x 21 x 22 x 23 [ x j ] n x 24 x 25 x 26 x 27 x 28 x 1 is l , i , v , m , a or p ; x 2 is any amino acid residue ; x 3 is p , t or s ; x 4 is l , i , v , m , a or p ; x 5 is any amino acid ; x 6 is any amino acid ; x 7 is l , i , v , m , a , f , y or w ; x 8 is c , t or s ; x 9 is r , k or h ; x 10 is any amino acid ; x 11 is any amino acid ; x 12 is l , i , v , m , a or p ; x 13 is any amino acid ; x 14 is any amino acid ; x 15 is any amino acid ; x 16 is l , i , v , m , a , p , g , c , t or s ; [ x i ] n is a sequence of n amino acids wherein n is from 1 to 50 amino acids and wherein the sequence x i may comprise the same or different amino acids selected from any amino acid residue ; x 17 is l , i , v , m , a or p ; x 18 is any amino acid ; x 19 is any amino acid ; x 20 is l , i , v , m , a or p ; x 21 is p ; x 22 is l , i , v , m , a , p or g ; x 23 is p or n ; [ x j ] n is a sequence of n amino acids wherein n is from 0 to 50 amino acids and wherein the x j may comprise the same or different amino acids selected from any amino acid residue ; x 24 is l , i , v , m , a or p ; x 25 is any amino acid ; x 26 is any amino acid ; x 27 is y or f ; x 28 is l , i , v , m , a or p . the socs protein also comprises a protein : molecule interacting region such as but not limited to one or more of an sh2 domain , wd40 repeats and / or ankyrin repeats , n - terminal of the socs box . modulating hormone signalling includes modulating growth control mechanisms . in addition , modulating growth hormone signalling has applications in increasing strength in elderly people , obesity control treatment of catabolic diseases , treatment of chronic inflammatory disease , treatment and / or prophylaxis of osteoporosis , treatment of cardiomyopathy and in the treatment of complicated fracture . another aspect of the present invention provides a method of modulating hormone signalling in an animal and in particular a human , said method comprising up - regulating or down - regulating expression of a genetic sequence encoding a socs protein or increasing or decreasing the activity of a socs protein in said animal and wherein said socs protein comprises a protein : molecule interacting region such as but not limited to an sh2 domain , wd40 repeats and / or ankyrin repeats , n terminal of a socs box , wherein said socs box comprises the amino acid sequence : x 1 x 2 x 3 x 4 x 5 x 6 x 7 x 8 x 9 x 10 x 11 x 12 x 13 x 14 x 15 x 16 [ x i ] n x 17 x 18 x 19 x 20 x 21 x 22 x 23 [ x j ] n x 24 x 25 x 26 x 27 x 28 x 1 is l , i , v , m , a or p ; x 2 is any amino acid residue ; x 3 is p , t or s ; x 4 is l , i , v , m , a or p ; x 5 is any amino acid ; x 6 is any amino acid ; x 7 is l , i , v , m , a , f , y or w ; x 8 is c , t or s ; x 9 is r , k or h ; x 10 is any amino acid ; x 11 is any amino acid ; x 12 is l , i , v , m , a or p ; x 13 is any amino acid ; x 14 is any amino acid ; x 15 is any amino acid ; x 16 is l , i , v , m , a , p , g , c , t or s ; [ x i ] n is a sequence of n amino acids wherein n is from 1 to 50 amino acids and wherein the sequence x i may comprise the same or different amino acids selected from any amino acid residue ; x 17 is l , i , v , m , a or p ; x 18 is any amino acid ; x 19 is any amino acid ; x 20 is l , i , v , m , a or p ; x 21 is p ; x 22 is l , i , v , m , a , p or g ; x 23 is p or n ; [ x j ] n is a sequence of n amino acids wherein n is from 0 to 50 amino acids and wherein the x j may comprise the same or different amino acids selected from any amino acid residue ; x 24 is l , i , v , m , a or p ; x 25 is any amino acid ; x 26 is any amino acid ; x 27 is y or f ; x 28 is l , i , v , m , a or p . the present invention extends to any socs molecule such as those disclosed in international patent application no . pct / au99 / 00729 [ wo 98 / 20023 ] which is herein incorporated by reference . however , in a particularly preferred embodiment , the present invention is directed to manipulating levels of socs - 2 , which murine form comprises the nucleotide and corresponding amino acid sequence as set forth in seq id no : 1 and seq id no : 2 , respectively . the present invention is hereinafter described with reference to murine socs - 2 ( msocs - 2 ), however , this is done with the understanding that the present invention encompasses the manipulation of levels of any socs molecule , such as but not limited to human socs - 2 ( hsocs - 2 ). the nucleotide sequence of hsocs - 2 and its corresponding amino acid sequence can be found at gen bank accession number af037989 ( see also reference 19 ). reference herein to a “ socs ” molecule such as socs - 2 includes any mutants thereof such as functional mutants . an example of a mutant is a single or multiple amino acid substitution , addition and / or deletion or truncation to the socs molecule or its corresponding dna or rna . another useful socs molecule is socs - 3 and manipulating levels of socs - 3 is encompassed by the present invention . the present invention is predicated in part on the determination that knock out mice for a socs gene exhibit altered hormone signalling . in particular , mice which substantially do not produce socs - 2 exhibit disrupted growth hormone signalling and , as a result , have a growth advantage over their littermates . this provides the basis for developing therapeutic protocols for subjects which either require more or require less growth hormone signalling . it also has important implications in the veterinary industry for manipulating animals to provide same with a growth advantage . this may be important for food production . alternatively , where a reduction in the size of an animal is desired , facilitating expression of a socs gene or facilitating socs protein activity will result in greater growth hormone signalling . although the present invention is specifically exemplified in relation to growth hormone , the instant invention extends to any hormone signalling such as facilitated by protein hormones ( i . e . cytokines ) and non - protenaceous hormones . accordingly , another aspect of the present invention contemplates a method for controlling hormone signalling such as growth hormone signalling in an animal such as a human or livestock animal , said method comprising modulating expression of a genetic sequence encoding a socs protein comprising a socs box and a protein : molecule interacting region n - terminal of said socs box wherein said socs box comprises the amino acid sequence : x 1 x 2 x 3 x 4 x 5 x 6 x 7 x 8 x 9 x 10 x 11 x 12 x 13 x 14 x 15 x 16 [ x i ] n x 17 x 18 x 19 x 20 x 21 x 22 x 23 [ x j ] n x 24 x 25 x 26 x 27 x 28 x 1 is l , i , v , m , a or p ; x 2 is any amino acid residue ; x 3 is p , t or s ; x 4 is l , i , v , m , a or p ; x 5 is any amino acid ; x 6 is any amino acid ; x 7 is l , i , v , m , a , f , y or w ; x 8 is c , t or s ; x 9 is r , k or h ; x 10 is any amino acid ; x 11 is any amino acid ; x 12 is l , i , v , m , a or p ; x 13 is any amino acid ; x 14 is any amino acid ; x 15 is any amino acid ; x 16 is l , i , v , m , a , p , g , c , t or s ; [ x i ] n is a sequence of n amino acids wherein n is from 1 to 50 amino acids and wherein the sequence x i may comprise the same or different amino acids selected from any amino acid residue ; x 17 is l , i , v , m , a or p ; x 18 is any amino acid ; x 19 is any amino acid ; x 20 is l , i , v , m , a or p ; x 21 is p ; x 22 is l , i , v , m , a , p or g ; x 23 is p or n ; [ x j ] n is a sequence of n amino acids wherein n is from 0 to 50 amino acids and wherein the x j may comprise the same or different amino acids selected from any amino acid residue ; x 24 is l , i , v , m , a or p ; x 25 is any amino acid ; x 26 is any amino acid ; x 27 is y or f ; x 28 is l , i , v , m , a or p . preferably , the socs protein - encoding genetic sequence comprises a nucleotide sequence substantially as set forth in seq id no : 1 or a nucleotide sequence having at least 60 % similarity thereto or a nucleotide sequence capable of hybridizing to seq id no : 1 or its complementary form under low stringency conditions at 42 ° c . even more preferably , the socs protein in a human homologue of the nucleotide sequence set forth in seq id no : 1 . the term “ similarity ” as used herein includes exact identity between compared sequences at the nucleotide or amino acid level . where there is non - identity at the nucleotide level , “ similarity ” includes differences between sequences which result in different amino acids that are nevertheless related to each other at the structural , functional , biochemical and / or conformational levels . where there is non - identity at the amino acid level , “ similarity ” includes amino acids that are nevertheless related to each other at the structural , functional , biochemical and / or conformational levels . in a particularly preferred embodiment , nucleotide and sequence comparisons are made at the level of identity rather than similarity . terms used to describe sequence relationships between two or more polynucleotides or polypeptides include “ reference sequence ”, “ comparison window ”, “ sequence similarity ”, “ sequence identity ”, “ percentage of sequence similarity ”, “ percentage of sequence identity ”, “ substantially similar ” and “ substantial identity ”. a “ reference sequence ” is at least 12 but frequently 15 to 18 and often at least 25 or above , such as 30 monomer units , inclusive of nucleotides and amino acid residues , in length . because two polynucleotides may each comprise ( 1 ) a sequence ( i . e . only a portion of the complete polynucleotide sequence ) that is similar between the two polynucleotides , and ( 2 ) a sequence that is divergent between the two polynucleotides , sequence comparisons between two ( or more ) polynucleotides are typically performed by comparing sequences of the two polynucleotides over a “ comparison window ” to identify and compare local regions of sequence similarity . a “ comparison window ” refers to a conceptual segment of typically 12 contiguous residues that is compared to a reference sequence . the comparison window may comprise additions or deletions ( i . e . gaps ) of about 20 % or less as compared to the reference sequence ( which does not comprise additions or deletions ) for optimal alignment of the two sequences . optimal alignment of sequences for aligning a comparison window may be conducted by computerised implementations of algorithms ( gap , bestfit , fasta , and tfasta in the wisconsin genetics software package release 7 . 0 , genetics computer group , 575 science drive madison , wis ., usa ) or by inspection and the best alignment ( i . e . resulting in the highest percentage homology over the comparison window ) generated by any of the various methods selected . reference also may be made to the blast family of programs as , for example , disclosed by altschul et al . ( 16 ). a detailed discussion of sequence analysis can be found in unit 19 . 3 of ausubel et al . ( 17 ). the terms “ sequence similarity ” and “ sequence identity ” as used herein refers to the extent that sequences are identical or functionally or structurally similar on a nucleotide - by - nucleotide basis or an amino acid - by - amino acid basis over a window of comparison . thus , a “ percentage of sequence identity ”, for example , is calculated by comparing two optimally aligned sequences over the window of comparison , determining the number of positions at which the identical nucleic acid base ( e . g . a , t , c , g , i ) or the identical amino acid residue ( e . g . ala , pro , ser , thr , gly , val , leu , ile , phe , tyr , trp , lys , arg , his , asp , glu , asn , gln , cys and met ) occurs in both sequences to yield the number of matched positions , dividing the number of matched positions by the total number of positions in the window of comparison ( i . e ., the window size ), and multiplying the result by 100 to yield the percentage of sequence identity . for the purposes of the present invention , “ sequence identity ” will be understood to mean the “ match percentage ” calculated by the dnasis computer program ( version 2 . 5 for windows ; available from hitachi software engineering co ., ltd ., south san francisco , calif ., usa ) using standard defaults as used in the reference manual accompanying the software . similar comments apply in relation to sequence similarity . reference herein to a low stringency includes and encompasses from at least about 0 to at least about 15 % v / v formamide and from at least about 1 m to at least about 2 m salt for hybridization , and at least about 1 m to at least about 2 m salt for washing conditions . generally , low stringency is at from about 25 - 30 ° c . to about 42 ° c . the temperature may be altered and higher temperatures used to replace formamide and / or to give alternative stringency conditions . alternative stringency conditions may be applied where necessary , such as medium stringency , which includes and encompasses from at least about 16 % v / v to at least about 30 % v / v formamide and from at least about 0 . 5 m to at least about 0 . 9 m salt for hybridization , and at least about 0 . 5 m to at least about 0 . 9 m salt for washing conditions , or high stringency , which includes and encompasses from at least about 31 % v / v to at least about 50 % v / v formamide and from at least about 0 . 01 m to at least about 0 . 15 m salt for hybridization , and at least about 0 . 01 m to at least about 0 . 15 m salt for washing conditions . in general , washing is carried out t m = 69 . 3 + 0 . 41 ( g + c )% ( 18 ). however , the t m of a duplex dna decreases by 1 ° c . with every increase of 1 % in the number of mismatch base pairs ( 19 ). formamide is optional in these hybridization conditions . accordingly , particularly preferred levels of stringency are defined as follows : low stringency is 6 × ssc buffer , 0 . 1 % w / v sds at 25 - 42 ° c . ; a moderate stringency is 2 × ssc buffer , 0 . 1 % w / v sds at a temperature in the range 20 ° c . to 65 ° c . ; high stringency is 0 . 1 × ssc buffer , 0 . 1 % w / v sds at a temperature of at least 65 ° c . in an alternative embodiment , the modulation of growth hormone signalling is accomplished at the protein level using either a socs antagonist or agonist or using a composition comprising socs such as socs - 2 or its derivative , homologue or analogue . accordingly , another aspect of the present invention contemplates a method for controlling growth hormone signalling in an animal such as human or livestock animal , said method comprising administering to said animal a control - effective amount of a socs protein or functional part or homologue or analogue thereof or an antagonist or agonist of a socs protein for a time and under conditions sufficient to modulate growth hormone signalling . preferably , the socs protein is socs - 2 . where a socs protein is administered , it may be an isolated form of the naturally occurring socs protein or it may be a derivative , homologue or analogue thereof . a further aspect contemplates genetically modified animals such as livestock animals . such animals , having altered hormone signalling , are useful inter alia for food production . a “ derivative ” includes a part , portion or fragment thereof such as a molecule comprising a single or multiple amino acid substitution , deletion and / or addition . a “ homologue ” includes a functionally similar molecule from either the same species or another species . analogues contemplated herein include , but are not limited to , modification to side chains , incorporating of unnatural amino acids and / or their derivatives during peptide , polypeptide or protein synthesis and the use of crosslinkers and other methods which impose conformational constraints on the proteinaceous molecule or their analogues . examples of side chain modifications contemplated by the present invention include modifications of amino groups such as by reductive alkylation by reaction with an aldehyde followed by reduction with nabh 4 ; amidination with methylacetimidate ; acylation with acetic anhydride ; carbamoylation of amino groups with cyanate ; trinitrobenzylation of amino groups with 2 , 4 , 6 - trinitrobenzene sulphonic acid ( tnbs ); acylation of amino groups with succinic anhydride and tetrahydrophthalic anhydride ; and pyridoxylation of lysine with pyridoxal - 5 - phosphate followed by reduction with nabh 4 . the guanidine group of arginine residues may be modified by the formation of heterocyclic condensation products with reagents such as 2 , 3 - butanedione , phenylglyoxal and glyoxal . the carboxyl group may be modified by carbodiimide activation via o - acylisourea formation followed by subsequent derivitization , for example , to a corresponding amide . sulphydryl groups may be modified by methods such as carboxymethylation with iodoacetic acid or iodoacetamide ; performic acid oxidation to cysteic acid ; formation of a mixed disulphides with other thiol compounds ; reaction with maleimide , maleic anhydride or other substituted maleimide ; formation of mercurial derivatives using 4 - chloromercuribenzoate , 4 - chloromercuriphenylsulphonic acid , phenylmercury chloride , 2 - chloromercuri - 4 - nitrophenol and other mercurials ; carbamoylation with cyanate at alkaline ph . tryptophan residues may be modified by , for example , oxidation with n - bromosuccinimide or alkylation of the indole ring with 2 - hydroxy - 5 - nitrobenzyl bromide or sulphenyl halides . tyrosine residues on the other hand , may be altered by nitration with tetranitromethane to form a 3 - nitrotyrosine derivative . modification of the imidazole ring of a histidine residue may be accomplished by alkylation with iodoacetic acid derivatives or n - carbethoxylation with diethylpyrocarbonate . examples of incorporating unnatural amino acids and derivatives during peptide synthesis include , but are not limited to , use of norleucine , 4 - amino butyric acid , 4 - amino - 3 - hydroxy - 5 - phenylpentanoic acid , 6 - aminohexanoic acid , t - butylglycine , norvaline , phenylglycine , ornithine , sarcosine , 4 - amino - 3 - hydroxy - 6 - methylheptanoic acid , 2 - thienyl alanine and / or d - isomers of amino acids . a list of unnatural amino acid , contemplated herein is shown in table 1 . crosslinkers can be used , for example , to stabilize 3d conformations , using homo - bifunctional crosslinkers such as the bifunctional imido esters having ( ch 2 ) n spacer groups with n = 1 to n = 6 , glutaraldehyde , n - hydroxysuccinimide esters and hetero - bifunctional reagents which usually contain an amino - reactive moiety such as n - hydroxysuccinimide and another group specific - reactive moiety such as maleimido or dithio moiety ( sh ) or carbodiimide ( cooh ). in addition , peptides can be conformationally constrained by , for example , incorporation of c α and n α - methylamino acids , introduction of double bonds between c α and c β atoms of amino acids and the formation of cyclic peptides or analogues by introducing covalent bonds such as forming an amide bond between the n and c termini , between two side chains or between a side chain and the n or c terminus . the present invention further contemplates chemical analogues of socs molecules capable of acting as antagonists or agonists of a socs or which can act as functional analogues of socs . chemical analogues may not necessarily be derived from socs but may share certain conformational similarities . alternatively , chemical analogues may be specifically designed to mimic certain physiochemical properties of socs . chemical analogues may be chemically synthesized or may be detected following , for example , natural product screening . other derivatives contemplated by the present invention include a range of glycosylation variants from a completely unglycosylated molecule to a modified glycosylated molecule . altered glycosylation patterns may result from expression of recombinant molecules in different host cells . the present invention extends to compositions such as pharmaceutical compositions comprising one or more of a socs protein , a derivative , homologue or analogue thereof and / or an antagonist or agonist of a socs protein and one or more pharmaceutically acceptable carriers and / or diluents . such compositions are useful for modulating growth hormone signalling in an animal such as a human or livestock animal . preferably , the composition comprises socs - 2 or a derivative , homologue or analogue thereof . in order to facilitate passage of the socs protein through various membranes , the molecule may be fused to or co - expressed with tat , ( from hiv ) or penetratin . alternatively , the socs protein may be administered following laser treatment . the preparation of pharmaceutical compositions is well known in the art and is described , for example , in remington &# 39 ; s pharmaceutical sciences , eaton publishing , pennsylvania usa or in international patent publication no . pct / au99 / 00729 [ wo 98 / 20023 ]. the present invention also provides for the genetic control of socs levels in animals . for example , compositions comprising antisense rna or dna , ribozymes or sense molecules ( for co - suppression ) may be administered either locally or systemically to manipulate expression of socs genes or translation of socs mrna . another aspect of the present invention provides a genetically modified animal exhibiting altered hormone and in particular growth hormone signalling . more particularly , the present invention is directed to a genetically modified animal exhibiting increased or decreased levels of a socs protein or a derivative or homologue thereof wherein the socs protein , in a generally unaltered animal , comprises a protein : molecule interacting portion , n - terminal of a socs box , which socs box comprises the amino acid sequence : x 1 x 2 x 3 x 4 x 5 x 6 x 7 x 8 x 9 x 10 x 11 x 12 x 13 x 14 x 15 x 16 [ x i ] n x 17 x 18 x 19 x 20 x 21 x 22 x 23 [ x j ] n x 24 x 25 x 26 x 27 x 28 x 1 is l , i , v , m , a or p ; x 2 is any amino acid residue ; x 3 is p , t or s ; x 4 is l , i , v , m , a or p ; x 5 is any amino acid ; x 6 is any amino acid ; x 7 is l , i , v , m , a , f , y or w ; x 8 is c , t or s ; x 9 is r , k or h ; x 10 is any amino acid ; x 11 is any amino acid ; x 12 is l , i , v , m , a or p ; x 13 is any amino acid ; x 14 is any amino acid ; x 15 is any amino acid ; x 16 is l , i , v , m , a , p , g , c , t or s ; [ x i ] n is a sequence of n amino acids wherein n is from 1 to 50 amino acids and wherein the sequence x i may comprise the same or different amino acids selected from any amino acid residue ; x 17 is l , i , v , m , a or p ; x 18 is any amino acid ; x 19 is any amino acid ; x 20 is l , i , v , m , a or p ; x 21 is p ; x 22 is l , i , v , m , a , p or g ; x 23 is p or n ; [ x j ] n is a sequence of n amino acids wherein n is from 0 to 50 amino acids and wherein the x j may comprise the same or different amino acids selected from any amino acid residue ; x 24 is l , i , v , m , a or p ; x 25 is any amino acid ; x 26 is any amino acid ; x 27 is y or f ; x 28 is l , i , v , m , a or p . preferably , the socs protein is socs - 2 or its derivative or homologue and comprises the amino acid sequence substantially as set forth in seq id no : 2 or an amino acid sequence having at least about 60 % similarity thereto . in a particularly preferred embodiment , the present invention provides a genetically modified animal wherein the animal , before genetic modification , comprises a genetic sequence which comprises a sequence of nucleotides substantially corresponding to the nucleotide sequence set forth in seq id no : 1 or its complementary form or is able to hybridize under low stringency conditions at 42 ° c . to seq id no : 1 or its complementary form and wherein said genetic modification either results in a deletion of one or more nucleotides from said genetic sequence from said animal or substantially prevents , reduces or down - regulates expression of said genetic sequence . the term “ expression ” includes transcription and / or translation of a genetic sequence . in a particularly preferred embodiment , the present invention provides a genetically modified animal comprising a substantial deletion in a genetic sequence comprising a nucleotide sequence substantially corresponding to seq id no : 1 or capable of hybridizing to seq id no : 1 under low stringency conditions at 42 ° c . and wherein said animal exhibits altered hormone regulation such as growth hormone regulation . genetically modified animals contemplated by the present invention include mice , rats , guinea pigs , rabbits and livestock animals ( e . g . pigs , cows , sheep , donkeys , horses ). genetically modified livestock animals are particularly useful in food production . the genetically modified animals of the present invention are also useful animal models for screening for therapeutic agents capable of modulating growth hormone signalling . the present invention further extends to agents identified using the animal model of this aspect of the present invention . the present invention is further described by the following non - limiting examples . a genomic socs - 2 fragment extending approximately 2 . 0 kb from the protein initiation atg was generated by pcr . this fragment was fused to the atg of β - galactosidase via the bamhi site in the plasmid vector pβgalpaloxneo ( 24 ). the 3 ′ arm , an ecori fragment extending 3 . 7 kb downstream from the termination codon was blunted and ligated into the xhoi ( blunted ) site of pβgalpaloxneo that already contained the 5 ′ arm . this targetting vector was linearized with noti and electroporated into c57bl / 6 embryonic stem cells . transfected cells were selected in g418 and resistant clones picked and expanded . clones in which the targeting vector had recombined with the endogenous socs - 2 gene were identified by hybridizing ecorv - digested genomic dna with a 0 . 8 kb bamhi - nhei fragment situated in 5 ′ socs - 2 genomic sequence just outside the targeting vector ( fig1 ). this probe distinguished between the endogenous ( 16 kb ) and targeted ( 9 kb ) socs - 2 alleles . a targeted es cell clone was injected into balb / c blastocysts to generate chimaeric mice . male chimaeras were mated with c57bl / 6 females to yield socs - 2 heterozygotes which were interbred to produce wild - type ( socs - 2 +/+ ), heterozygous ( socs - 2 +/− ) and mutant ( socs - 2 −/− ) mice on a pure c57bl / 6 genetic background . the genotypes of offspring were determined by southern blot analysis of genomic dna extracted from tail biopsies as described above . the deletion of socs - 2 coding sequence and subsequent inability to produce socs - 2 mrna in mutant mice was confirmed in nucleic acid blots which were performed as previously described ( 25 ). northern blots were probed with a full - length socs - 2 coding region probe and then with a 1 . 2 kb psti chicken glyceraldehyde - 3 - phosphate dehydrogenase ( gapdh ) fragment . peripheral blood white cell and platelet counts were determined manually using haemocytometers . single cell suspensions from femoral bone marrow , spleen and liver were prepared and differential counts of peripheral blood , bone marrow and spleen were performed from stained smears and cytocentrifuge preparations . clonal cultures of 2 . 5 × 10 4 adult bone marrow cells were performed in 0 . 3 % agar as previously described ( 26 ). cultures were stimulated with recombinant purified gm - csf , g - csf , m - csf , il - 3 ( each at 10 ng / ml ), scf ( 100 ng / ml ), il - 6 ( 100 ng / ml ) or flk - ligand ( 500 ng / ml ) plus lif ( 10 3 u / ml ). agar cultures were fixed , sequentially stained for acetylcholinesterase , luxol fast blue and hematoxylin , and the cellular composition of each colony determined microscopically . tissue sections were prepared by standard techniques , stained with haematoxylin and eosin and examined by light microscopy . to analyze mup levels , 6 - 7 week old mice were made to urinate before samples were collected 3 and 5 . 5 hours later . samples were pooled and centrifuged ( 13 , 000 rpm × 3 min ) before 0 . 5 μl of supernatant was electorphoresed in 12 % sds - polyacrylamide gels and stained with coomassie blue . cohorts of mice were weighed at weekly intervals for 12 weeks from birth . after sacrifice , animals were pinned down through the oral cavity and lightly stretched by the tail for nose - anus ( body length ) and anus - tail ( tail length ) measurements . to measure skeletal dimensions , limbs were subsequently removed and oriented in a consistent manner for x - ray photography and bone length measurement . serum igf - i levels in serum from orbital bleeds were determined using an eia kit ( rat igf dsl - 10 - 2900 diagnostic systems laboratories , webster , tx ) according to the manufacturer &# 39 ; s instructions . igf - i rna expression was determined in rnase protections assays as previously described ( 23 ) using β - actin as an internal standard . to construct the socs - 2 targeting vector , a 5 ′ arm extending approximately 2 . 0 kb from the protein initiation atg was generated by pcr using specific socs - 2 oligonucleotides and genomic clone pgmsocs - 2 57 - 60 - 1 - 45 as template . this fragment was fused to the atg of β - galactosidase via the bamhi site in the plasmid vector pβgalpaloxneo . the 3 ′ arm , a 3 . 7 kb ecori fragment from pgmsocs - 2 57 - 60 - 145 was blunted and ligated into the xhoi ( blunted ) site of pβgalphaloxneo that already contained the 5 ′ arm . this targeting vector was linearized with noti and electroporated into c57bl / 6 embryonic stem cells . transfected cells were selected in g418 and resistant clones picked and expanded . clones in which the targeting vector has recombined with the endogenous socs - 2 gene were identified by hybridising ecorv - digested genomic dna with a 1 . 8 kb ecori - ecorv fragment from pgmsocs - 2 57 - 60 - 1 - 45 . this probe ( probe a , fig1 ), which is located 3 ′ to the socs - 2 sequences in the targeting vector , distinguished between the endogenous ( greater than 14 kb ) and targeted ( 7 . 5 kb ) socs - 2 loci ( fig1 ). several targeted es cells clones were identified , one of which was injected into balb / c blastocysts to generate chimeric mice . germline chimeras were mated with c57 / bl / 6 mice to produce socs - 2 +/− mice which were interbred to yield socs - 2 - deficient animals . southern blot analysis at weaning revealed that offspring of heterozygous parents included mice of each of the three expected genotypes in approximately mendelian proportions ( 22 : 51 : 22 for socs - 2 +/+ : socs - 2 +/− : socs - 2 −/− ). northern blots of rna extracted from a range of organs confirmed that socs - 2 transcripts were absent in homozygous mutant mice . this result is consistent with the findings that the socs - 2 gene had been functionally deleted . socs - 2 −/− mice appeared outwardly normal , however , male mice and perhaps to a lesser extend female mice developed to a significantly larger body weight than their normal littermates ( fig2 ; males at 3 months of age : socs - 2 −/− 41 . 2 ± 4 . 8 , wild - type 29 . 3 ± 1 . 8 ; females : socs - 2 −/− 25 . 3 ± 2 . 2 , wild - type 21 . 9 ± 2 . 2 grams ). in male mice the increased body weight was parallelled by increased size of several organs , in particular the pancreas , liver , lungs and heart ( fig3 ). upon removal of all organs the resulting means carcass weight also highlighted the increased size of socs - 2 −/− mice ( 21 . 5 ± 1 . 5 g ), compared to wild - type controls ( 13 . 1 ± 1 . 9 g ). an extensive analysis has suggested that mice socs - 2 −/− organs were histologically normal , with the exception of a marked thickening of the skin which develops as a result of increase collagen deposition . both male and female socs - 2 −/− have been found to be fertile . adult socs - 2 +/− males exhibited an intermediate body weight (−/−: 36 . 6 ± 1 . 0 g ; +/−: 30 . 8 ± 1 . 3 g ; +/+: 27 . 1 ± 1 . 6 g , at 12 weeks of age , n = 5 - 20 mice per group ). increased growth was also significant but less dramatic in female socs - 2 −/− mice . adult socs - 2 −/− females typically attained weights of wild - type male mice , but heterozygous socs - 2 females were not significantly heavier that sex - matched wild - type littermates (−/−: 26 . 2 ± 1 . 5 g ; +/−: 21 . 8 ± 0 . 7 g ; +/+: 20 . 5 ± 1 . 4 g , at 12 weeks of age , n = 7 - 20 mice per group ). consistent with this interpretation , the femur , tibia , radius and humerus in socs - 2 −/− mice were all significantly longer than in wild - type controls ( table 2 ). body length in male socs - 2 - deficient mice was also greater , although tail length was normal ( table 2 ). the mean frequency of hepatic nuclei per 10 high power fields in socs - 2 −/− liver sections was no greater than that in wild type mice ( 27 . 3 ± 4 . 4 , n = 4 versus 27 . 7 ± 2 . 6 , n = 4 ) and striated muscle cell width was normal in the thighs of socs - 2 - deficient animals . thus , increased organ weights in these mice appear to have resulted from elevated cell numbers rather than increased cell size . a similar , but less pronounced trend was also observed when organ and carcass weights and body , tail and bone lengths were assessed in female socs - 2 −/− mice ( table 2 ). a thorough hematological survey has not identified any hematological abnormalities in socs - 2 −/− mice ( table 3 ). the increased growth of mice is considered to be due to disruption in the pulsatile nature of growth hormone signalling in animals and in particular male animals . to further analyze this , the phenomenon is studied and compared in animals homozygous and heterozygous for animals homozygous and heterozygous for the socs - 2 gene . in these animals , levels of growth hormone and growth hormone controlled factors such as igf - 1 is determined . the activation status of stat5a and in particular stat5b , which are major mediators of growth hormone and prolactin signalling , is determined in the heterozygous and homozygous mice . in addition , growth hormone , pulse - regulated , sexually dimorphic gene expression of genes such as mup and cyp is also determined . furthermore , the sensitivity of knock out mice to growth hormone signalling is further analyzed . to more directly examine the effects of socs - 2 on the growth hormone system , the inventors initiated crosses of the socs - 2 −/− mice with stat5b −/− and little mice . stat5b is required for the sexually dimorphic effects of growth hormone . consequently , male mice lacking socs - 5b grow no larger than female mice , which are themselves normal size . a range of outcomes to this experiment is envisaged . if socs - 2 is required to regulate growth hormone ( gh ) signalling , mice lacking both socs - 2 and stat5b are expected to reproduce a stat - 5b - deficient phenotype as gh signalling would not properly operate in the absence of this key signalling molecule making the presence or absence of socs - 2 irrelevant . if socs - 2 acts independently of gh signalling , an intermediate phenotype might ensue . conversely , given that little mice are not entirely gh - deficient , if socs - 2 acts to regulate gh signalling , a little mouse that is also deficient in socs - 2 might exhibit amelioration of dwarfism . if socs - 2 acts on the gh or igf - i signalling pathways , a strong prediction would be that the socs - 2 - deficient cells or the socs - 2 −/− mice themselves would be significantly more sensitive than their wild - type counterparts to the effects of these cytokines . the deregulating of gh signalling might result in heightened activation of stat5b , the key stat involved in sexually dimorphic growth in mice . this is investigated in the livers of unmanipulated male socs - 2 −/− mice , together with those from unmanipulated wild - type mice and wild - type mice after injection with a maximally stimulating dose of gh . the levels of expression of downstream markers of pulse - regulated gh signalling , including mup , and cytochrome p450 - catalysed testosterone hydroxylase , are determined in northern blot analysis of liver rna and / or western blot analysis of protein extracts . differences in gh sensitivity is also investigated by comparing the cytokine concentration required to stimulate phosphorylation of stat5b in socs - 2 −/− and normal mice in dose response studies using preparations of primary hepatocytes and / or fibroblasts . the size parameters of socs - 2 / stat5b are shown in table 4 . the data in the table show that male socs - 2 −/− stat5b −/− mice have a growth phenotype intermediate to the large socs - 2 −/− stat5b +/+ and small socs - 2 +/+ stat5b −/− mice . in female mice , socs - 2 −/− stat5b −/− also appear to be intermediate in this regard compared with mice lacking socs - 2 or stat5b singly . this result implies that the excessive growth in socs - 2 −/− mice is dependent , at least in part , on stat5b . also , although not proof , this is further evidence consistent with abnormal growth hormone signalling in socs - 2 −/− mice . the possibility that socs - 2 acts directly in the igf - i signalling cascade is also being investigated . to achieve this , the phosphorylation status of the igf - i receptor and key downstream signalling molecules in cells or tissues of socs - 2 −/− mice is investigated . primary embryo fibroblast cultures are established from socs - 2 −/− and normal control mice . initially , the extent and duration of phosphorylation of the igf - i receptor , as well as the downstream effectors irs and shc is measured following stimulation of these cells , or in primary hepatocyte preparations , with a maximally - stimulating dose of igf - i . differences in igf - i sensitivity are also investigated by comparing the minimal cytokine concentration required to stimulate receptor and substrate phosphorylation in socs - 2 −/− and normal fibroblasts in dose response studies . transgenic mice are generated which constitutively express socs - 2 ubiquitously . the murine cdna encoding socs - 2 is linked to human ubiquitin c promoter , which drives high - level transgene expression in most mouse tissues . if the transgenic mice are small , and show evidence of resistance to gh or igf - i , this provides strong evidence consistent with the hypothesis that socs - 2 acts within the gh / igf - i axis . socs - 2 regulation of major urinary protein ( mup ) and insulin - like growth factor ( igf - 1 ) the inventors suspected that asspects of gh and / or igf - i signalling might be deregulated in socs - 2 −/− mice . to directly examine the gh / igf - i pathway , the inventors first investigated levels of major urinary protein ( mup ), a gh pulse - dependent product that is down - regulated in gh - overexpressing transgenic mice , where the usual pulsatile pattern of gh signalling is disrupted ( 21 ). to analyze mup levels , 6 - 7 week old mice were made to urinate before samples were collected 3 and 5 . 5 hours later . samples were pooled and centrifuged ( 13 , 000 rpm × 3 min ) before 0 . 5 μl of supernatant was electorphoresed in 12 % w / v sds - polyacrylamide gels and stained with coomassie blue . consistent with deregulated gh signalling , less mup was observed in samples of urine from each of 6 male and 5 female socs - 2 −/− mice compared with a similar number of sex - matched wild - type samples ( fig4 a ). production of igf - i is also stimulated by gh and , consistent with deregulated gh action , rnase protection assays revealed increased igf - i production in several organs including the heart , lungs and spleen of socs - 2 −/− mice ( fig4 b , c ). serum igf - i levels in serum from orbital bleeds were determined using an eia kit ( rat igf dsl - 10 - 2900 diagnostic systems laboratories , webster , tx ) according to the manufacturer &# 39 ; s instructions . igf - i rna expression was determined in rnase protections assays as previously described ( 23 ) using β - actin as an internal standard . however , excess production was not evident in the liver , bone , fat or muscle . no increase in serum igf - i concentration was observed in socs - 2 −/− mice ( male −/−: 278 ± 74 ng / ml ; +/+: 282 ± 45 ; female −/−: 310 ± 62 ; +/+: 298 ± 76 ; n = 6 mice per group ), consistent with normal production in the liver , which is the major source of circulating igf - i ( 22 , 23 ). those skilled in the art will appreciate that the invention described herein is susceptible to variations and modifications other than those specifically described . it is to be understood that the invention includes all such variations and modifications . the invention also includes all of the steps , features , compositions and compounds referred to or indicated in this specification , individually or collectively , and any and all combinations of any two or more of said steps or features . 1 . nicola , n . a . guidebook to cytokine and their receptors . oxford university press : oxford , 1994 . 3 . sprang et al , curr . opin . structural biol . 3 : 815 - 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