TITLE

BROAD SPECTRUM ERBB LlGAND BINDING MOLECULES AND METHODS FOR

PREPARING AND USING THEM

BACKGROUND

[0001] Receptor tyrosine kinases are involved in stimulating the growth of many cancers. In general, receptor tyrosine kinases are glycoproteins which consist of (1 ) an extracellular domain that is able to bind with a specific ligand, (2) a transmembrane region, (3) a juxtamembrane domain which may regulate the receptor activity by, for instance, protein phosphorylation, (4) a tyrosine kinase domain that is the enzymatic component of the receptor, and (5) a carboxyterminal tail. The ErbB family of type 1 receptor tyrosine kinases constitute one important class of receptors because of their importance in mediating cell growth, differentiation and survival in many solid tumors. Members of this receptor family include ErbBl (also known as HERJ), ErbB2 (HER2/neu), ErbB3 (HER3), and ErbB4 (HER4). More than a dozen ligands interact with the ErbB-family receptors. For example, EGF, Transforming Growth Factor α (TGFα), and amphiregulin all bind to ErbB l . Isoforms of ncuregulin, also known as Hcregulin and Neu Differentiation Factor (NDF) have specific affinity for ErbB3 and ErbB4. Ligands such as betacellulin, heparin-binding EGF and epiregulin bind to both ErbB l and ErbB4.

[0002] It is becoming clear that over expression of ErbB activating ligands can cause uncontrolled cellular proliferation similar to that of a deregulated receptor. In such cases, interference with the binding of the activating ligand to its receptor may provide an effective therapeutic strategy or that could accentuate current receptor based or other therapies. Therapeutics that interfere with ligand binding to ErbB3 may be particularly effective. ErbB3 differs from the other receptors in the EGFR family because its tyrosine kinase domain is functionally inactive; however, ErbB2/ErbB3 heterodimers transmit the most potent mitogenic signals of any homo- or heterodimer combination of the ErbB family. Therefore, ErbB3 is an important target, yet one that cannot be inhibited through small molecules that target the kinase region. Since ErbB3 requires an activating ligand, such as heregulin or NDF, before activated heterodimers can form, molecules that can interfere with the binding of ErbB3 receptor ligands could be used to block or interfere with the formation of ErbB dimcrs and heterodimers. One example of such a molecule would be a molecule that has ligand binding affinity and can therefore "trap" ligands and effectively reduce their concentration so that they cannot activate the ErbB3 receptor. In addition to ErbB3 ligands, the other known ErbB receptor ligands, have similar effects to varying degrees. Thus, binding molecules that can trap and sequester the full spectrum of ErbB ligands may be of even more use in the treatment of cancer.

[0003] Several therapeutics exist that have attempted this trapping or "decoy" strategy. For example, Enbrel™ (etanercept - Amgen) is a soluble, modified version of the TNFR receptor that binds and traps the pro-inflammatory ligand TNFoc. Jn addition, a soluble fusion protein of the VEGFRl and VEGFR2 receptors, called a VEGF Trap, is currently in clinical trials for the treatment of both macular degeneration and several forms of cancer (Regeneron Pharmaceuticals). An ErbB3 trap has also shown potency in vitro at enhancing the effects of a dual EGFR/ErbB2 inhibitor and reversed GW2974 (a small molecule inhibitor of ErbB 1 and ErbB2) resistance in cells treated with NDF.

[0004] AU currently approved ErbB inhibitors target either EGFR, ErbB2, ErbB3, ErbB4 or combinations of all 4 receptors. However, no therapeutic is known that interferes with the binding of ligands to multiple ErbB receptors simultaneously. Clearly, new binding molecules are needed that can be used to sequester receptor ligands, such as ErbB ligands, and thereby block ligand binding to multiple ErbB receptors and subsequent receptor activation. Binding molecules capable of binding all known ErbB ligands would be particularly useful.

[0005] A number of binding studies have been carried out to determine regions of ErbB3 that are important to the binding of its ligand, heregulin. Singer, et. al. (2001), J. Biol. Chem. 276, 44266-44274. Other studies using chimeric receptors have identified the relative contributions of the extracellular domains of ErbB 1 and ErbB4 to ligand-specific signaling. Kim, et. al. (2002), Eur. J. Biochem. 269, 2323-2329. These studies reveal that neuregulin binding to ErbB4 depends much more on domain 1 than on domain III and that domain 111 of ErbB l is primarily important for EGF binding. However, these studies were conducted on full length receptors which span the entire length of the receptors including the transmembrane and cytoplasmic domains. These large molecules present manufacturing and administration problems potentially leading to lower therapeutic efficacy. SUMMARY

[0006] Chimeric ErbB ligand binding molecules having detectable binding activity for more ErbB ligands than any one of native ErbB l , ErbB3 or ErbB4 are disclosed. Preferably, the binding molecules bind a broad spectrum and, more preferably, the full spectrum of ErbB ligands. The chimeric ErbB ligand binding molecule generally has a subdomain LI derived from one of EιbB3 or 4 and a subdomain LII derived from another distinct ErbB receptor type, such as ErbBl , which are linked. The sub-domain, Sl, can be used to join LI and LII. Subdomain SI can be from either one of the receptor types or can have portions from both.

[0007] In one embodiment the chimeric ErbB ligand binding molecule can include at least a portion of LI from ErbB4 linked to at least a portion of LII derived from ErbB l . LlI can be linked to module 1 of SIl from ErbB l . An SI subdomain can be used to link LI and LII subdomains and can be derived from either the ErbB l or ErbB4 receptor sequences or can be a mixture of both. Optionally, the ErbB chimera can be fused to an aggregant such as IgG2Fc.

[0008] In one embodiment the chimeric ErbB ligand binding molecule can include at least a portion of LI from ErbB4; an Sl region having a portion derived from ErbB4 and a portion derived from ErbB l , wherein the ErbB4 portion switches to the ErbB l portion in a region having homology between the two sequences; at least a portion of LII can be derived from ErbB l and module 1 of SIl of ErbB l .

[0009] In one embodiment the chimeric ErbB ligand binding molecule can include at least a portion of a subunit LI from ErbB3 linked to at least a portion of an LlI derived from ErbB l and module 1 of SIl from ErbB l . The linking region can be an SI domain from ErbB I , ErbB3 or their mixtures.

[0010] In one embodiment the chimeric ErbB ligand binding molecule can include at least a portion of a subunit Ll from ErbB3, an SI region having a portion is derived from ErbB3 and a portion is derived from ErbB l , at least a portion of LII derived from ErbB l and module I of SII derived from ErbB l .

[001 1 ] In one embodiment the chimeric ErbB ligand binding molecule can include at least a portion of a subunit Ll from ErbB4, an SI region having a portion derived from ErbB4 and a portion is derived from ErbB l , at least a portion of LIl from ErbB l and module 1 of SlI from ErbB 1.

[0012] In one embodiment the chimeric ErbB ligand binding molecule can include at least a portion of a subunit Ll from ErbB3; an SI region having a portion derived from ErbB3 and a portion derived from ErbBl, wherein the ErbB3 portion switches to the ErbBl portion in a region having homology between the two sequences; at least a portion of LIl derived from ErbBl and module 1 of SIl derived from ErbBl .

[0013] DNA sequences that encode the disclosed chimeric ErbB ligand binding molecules are also contemplated along with sequences that facilitate expression and host cells for the maintenance and expression of such DNA sequences.

[0014] Pharmaceutical compositions that contain the chimeric ErbB ligand binding molecules and a pharmaceutically acceptable cxcipient arc also contemplated.

[0015] The chimeric ErbB ligand binding molecule can be used by immobilizing it on a solid support which can in turn be used for binding ErbB ligands such that the ligands can be removed from biological fluids, particularly from patients suffering from diseases associated with over-expression of such ligands. The ErbB ligand depleted biological fluids can then be replaced in those patients.

[0016] Methods for treating patients having diseases that are associated with overexpression of ErbB ligands are also contemplated that involve administering a pharmaceutical composition that contain therapeutically effective amounts of the chimeric ErbB ligand binding molecule in a pharmaceutically acceptable excipient.

FIGURES

[0017] Figure 1 illustrates several potential subunit structures of a chimeric ErbB4- ErbB l chimera. The highlighted sequence shows one specific embodiment of a chimeric ErbB ligand binding molecule starting with the ErbB4 sequence.

[0018] Figure 2 illustrates several potential subunit structures of a chimeric ErbB3- ErbBl chimera. [0019] Figure 3 provides an alignment of ErbB l (bottom sequence) and ErbB4 (top sequence). The highlighted amino acid sequence shows one specific embodiment of a chimeric ErbB ligand binding molecule starting with the ErbB sequence.

DETAILED DESCRIPTION

[0020] The complete nucleotide sequences of the ErbBl , ErbB2, ErbB3 and ErbB4 arc known and can be found in Gcnbank as accession number NM 005228 for ErbBl, accession number NM 004448 for ErbB2, accession number M29366 or NM 001982 for ErbB3, and accession number NM 005235 for ErbB4. The full length ectodomain, the extracellular domain, for ErbB receptors contains four sub-domains, sometimes referred to as L l , CR l , L2 and CR2, where L and CR stand for large and cysteine rich, respectively. The sub-domains are also sometimes known as domains I-IV or alternatively as Ll, Sl, LlI and SlI as they extend from the amino terminus. Amino acid sequences of the receptors have been analyzed and the sequences appear to be homologous and have been aligned. Alignments of the ectodomains of ErbB l , ErbB2, ErbB3 and ErbB4 are provided in US Patent Publication No. 2006/0234343 in Figures I A and IB.

[0021 ] The sub-domains are composed of smaller domains known as subregions. For example, the Sl or CRl sub-domain contains 8 disulfide bonded subregions sometimes known as modules which are numbered 1-8 as they extend from the amino to carboxy terminal direction. SlI contains seven (7) modules numbered 1 -7. A large ordered loop has been identified in module 5 that is thought to project directly away from the ligand binding site. The amino acid sequence in this loop is highly conserved.

[0022] Chimeric ErbB ligand binding molecules are disclosed wherein sub-domains Ll, SI, LIl and module I of SIl from at least two different ErbB receptors are combined in a single binding molecule. For purposes of this disclosure, the phrase "chimeric" with respect to the ErbB ligand binding molecule is intended to mean a single ErbB ligand binding molecule made from and containing portions of more than one ErbB receptor ectodomain. The phrases "ErbB chimera" and "chimeric ErbB ligand binding molecule" are used interchangeably in this application and are intended to be synonymous and refer to the protein sequence of a monomer. As can be appreciated in some embodiments the chimera molecules may dimerize through formation of disulfide bonds.

[0023] Surprisingly, it has been found that portions of the amino acid sequence of ErbB3 or ErbB4 can be combined with portions of ErbB l through an SI linking domain to create a chimeric binding molecule that binds ligands to both of the combined receptors. Thus, the combination of ErbB l and ErbB4 domains can be used to produce a chimeric binding molecule that has affinity for both heregυlin (ErbB4 specific ligand) and TGFα (ErbB l specific ligand).

[0024] The switch in the amino acid sequence from one receptor type to another can be at any suitable location that provides for broad spectrum and high affinity binding of ErbB ligands. In some embodiments the switch can occur in the SI sub-domain which links Ll and LIl sub-domains. In some embodiments the switch will occur in regions where amino acid sequences are homologous or identical between the receptors being combined, such as in module 5 of the SI sub-domain.

[0025] For purposes of this disclosure the term "homology" is intended to mean a region of amino acid sequence having identical or conservative amino acid substitutions as that term is generally understood in the art. For example with respect to an ErbB4/ErbBl chimera, as shown in Figure 3, a switch from the ErbB4 to the ErbBl sequence can take place in module 5 of the Sl sub-domain such that the sequence ending in VYNP from the ErbB4 sequence is followed by TTYQ from the ErbB l sequence, as shown by the highlighted sequence in Figure 3.

[0026] The sequence can extend as far into the ErbB l LlI domain as is required for ErbB l ligand binding. The entire ErbB l LIl domain can be included and a portion or all of the subsequent SIl region from ErbB l can also be included. Figure 3 shows an ectodomain amino acid alignment of the Ll, Sl, LII and module 1 of the SII domain of ErbB4 and ErbB l , ending at amino acid 501 of ErbB 1.

[0027] With reference to Figure 3, one chimeric ErbB embodiment is shown in which subdomain LI is derived from ErbB4; the SI region is derived from a portion of the Sl region of ErbB4 and switches to a portion of the SI region of the ErbB l in module 5. This sequence continues on to include the ErbB l LIl subdomain and module 1 of the ErbB l SII domain. This can be designated as follows: LI (ErbB4) - Sl (ErbB4/ErbB l ) - LII (ErbB l ) - SIIm I (ErbB l ). More specifically: LI (ErbB4 amino acids 1-245) - SI (ErbB l starting with amino acid 249) - Ll I (ErbB l ) - SIImI (ErbB l ending with amino acid 501 according to the numbering of native ErbB l ).

[0028] All of the amino acid numbering in this application is intended to be exclusive of the native signal peptide. [0029] In certain embodiments the Sl domain can be composed of portions of each of the two sub-domains, as indicated previously. Further, it is also possible to introduce substitutions into the amino acid sequence for a variety of purposes. For example, the DNA sequence for the chimeric binding molecule can be changed to remove cysteines so that the formation of aggregates through cysteine-cysteine bonds can be avoided. Substitutions of amino acids in one subdomain can be used to modify ligand binding affinities. For example, an amino acid from ErbBl can be substituted into the ErbB4 Ll subdomain to make that domain's sequence more like that of ErbBl in order to modify the affinity of the molecule to ErbB ligands. Similarly, amino acid substitutions from ErbB3 or 4 LIl subdomains can be included into the ErbBl subdomain. Such substitutions can also be made in the SI and SIl subdomains. Although any number of such substitutions can be considered substitutions of glutaminc from ErbB l for serine in the ErbB4 portion at position 13, tyrosine for serine at position 42, argininc for tyrosine at position 123 are representative examples. Other examples can be identified by one of skill in the art simply by comparing sequences. Substitutions that are not homologous can also be considered. For example, asparagine could be substituted for serine at position 13 rather than the glutamine found in ErbB l or a residue that has intermediate characteristics of the residues found in both receptors may be used.

[0030] Any of these chimeric molecules could also be fused to other molecules or portions thereof including: other chimeric receptors (of any growth factor receptor family) or to sequences that facilitate purification of the product. The DNA sequences can be obtained from commercial sources and placed in any suitable expression vector and expressed from suitable hosts of which many are known.

[0031 ] In one embodiment the ErbB chimera can be fused with components that cause aggregative conjugate formation or extend protein half-life. For example, the ErbB chimera can be fused to the constant region of immunoglobulin molecule such as the Fc region of IgG. For purposes of this disclosure one suitable Fc region is known as lgG2Fc, although others are also known in the art and can be used.

[0032] For purposes of this application suitable binding affinities are affinities that are high enough to bind ErbB ligands in a physiological matrix. Preferably, dissociation constants will be no higher than about 10-fold to about 100-fold above the dissociation constants of the native receptors. More preferably, the dissociation constants for the ErbB chimera will be within 10-fold of their native receptor counterparts and more preferably within the same order of magnitude. Most preferably the binding affinities of the chimeric molecules will not be distinguishable from the native counterparts. Any affinity that is sufficient to bind and sequester ErbB ligands to thereby prevent or interfere with ligand binding and activating ErbB receptors is suitable for use and can find use in the disclosed methods. Binding affinity, which is a proxy for inhibitor potency, of the binding molecules can be measured using biosensor technology or by classic binding assays such as ELISA which are well known in the art.

[0033] DNA that encodes the chimeric ErbB ligand binding molecule sequences is also contemplated. One of skill can appreciate that the genetic code can be used to prepare suitable DNA sequences and codon preferences for specific expression hosts can also be incoφoratcd into such sequences. Also contemplated for use with these sequences are additional DNA sequences that can be used for the expression of these DNA sequences. A variety of these are known. As is well known in the art such sequences can also be introduced into host cells for the maintenance of the DNA and for its expression and such hosts that include these DNA sequences are also contemplated.

[0034] Pharmaceutical compositions comprising a disclosed chimeric ErbB ligand binding molecule are also contemplated. Such compositions comprise a therapeutically effective amount of a chimeric ErbB ligand binding molecule, and a pharmaceutically acceptable carrier. The term "pharmaceutically acceptable" means approved by a regulatory agency of lhc Federal or a state government or listed in the U.S. Pharmacopeia or other generally recognized pharmacopeia for use in animals, and more particularly, in humans. The term "carrier" refers to a diluent, adjuvant, excipient, or vehicle in which the therapeutic is administered. Such pharmaceutical carriers can be sterile liquids, such as water and oils, including those of petroleum, animal, vegetable or synthetic origin, such as peanut oil, soybean oil, mineral oil, sesame oil and the like. Suitable pharmaceutical excipients include starch, glucose, lactose, sucrose, gelatin, malt, rice, flour, chalk, silica gel, sodium stearate, glycerol monostcarate, talc, sodium chloride, dried skim milk, glycerol, propylene, glycol, water, ethanol and the like in which the chimeric ErbB ligand binding molecule is soluble and is chemically stable. The composition can also contain wetting or emulsifying agents, or pH buffering agents. These compositions can take the form of solutions, suspensions, emulsion, tablets, pills, capsules, powders, sustained-release formulations and the like. Pharmaceutically acceptable carriers include other ingredients for use in formulations such as DPPC, DOPE, DSPC and DOPC. Natural or synthetic surfactants may be used. PEG may be used (even apart from its use in derivatizing the protein or analog). Dcxtrans, such as cyclodcxtran, may be used. Cellulose and cellulose derivatives may be used. Amino acids may be used, such as use in a buffered formulation. Pharmaceutically acceptable diluents include buffers having various contents (e.g., Tris-HCl, acetate, phosphate), pH and ionic strength; additives such as detergents and solubilizing agents (e.g., Polysorbate 80), antioxidants (e.g., ascorbic acid, sodium metabisulfitc), preservatives (e.g., benzyl alcohol) and bulking substances (e.g., lactose, mannitol); incorporation of the material into particulate preparations of polymeric compounds such as polylactic acid, polyglycolic acid, etc. or into liposomes. Hyaluronic acid may also be used, and this may have the effect of promoting sustained duration in the circulation. Such compositions may influence the physical state, stability, rate of in vivo release, and rate of in vivo clearance of the present proteins and derivatives. See, e.g., Remington's Pharmaceutical Sciences, 18th Ed. ( 1990, Mack Publishing Co., Easton, PA 18042) pages 1435-1712 which are herein incorporated by reference. The compositions may be prepared in liquid form, or may be in dried powder, such as lyophilized form. Implantable sustained release formulations are also contemplated, as arc transdermal formulations. Liposome, microcapsule or microsphere, inclusion complexes, or other types of carriers are also contemplated.

[0035] The amount of the active chimeric binding molecule that will be effective for its intended therapeutic use can be determined by standard clinical techniques. In addition, in vitro assays may optionally be employed to help identify optimal dosage ranges. Generally, the daily regimen should be in the range of 0.1 -1000 micrograms of the active agent (API) kilogτam of body weight, preferably 0.1 -150 micrograms per kilogram. Effective doses may be extrapolated from dose-response curves derived from in vitro or suitable animal model test systems. Dosage amount and interval may be adjusted individually to provide plasma levels of the compounds that are sufficient to maintain therapeutic effect. In cases of local administration or selective uptake, the effective local concentration of the compounds may not be related to plasma concentration. The dosage regimen involved in a method for treatment can be determined by the attending physician, considering various factors which modify the action of drugs, e.g. the age, condition, body weight, sex and diet of the patient, the severity of disease, time of administration and other clinical factors.

[0036] The amount of compound administered will, of course, be dependent on the subject being treated, on the subject's weight, the severity of the affliction, the manner of administration, and the judgment of the prescribing physician. The therapy may be repeated intermittently while symptoms arc detectable or even when they are not detectable. The therapy may be provided alone or in combination with other drugs.

[0037] A method for treating a patient in need of treatment is disclosed that includes obtaining a chimeric ErbB ligand binding molecule that binds ErbB ligands and interferes with the interaction and effect of ligands on the ErbB receptor system of cancer cells, and administering a therapeutically effective amount of the molecule to a patient. Administration can be by parenteral routes such as i.v. administration, direct injection into a solid tumor such as through a syringe or catheter or by i.p. injection.

[0038] In one method of treatment the chimeric ErbB ligand binding molecules can be immobilized to a solid support such as an apheresis or biocore support by standard methods. When the binding molecule is immobilized to a solid support the serum, blood or other biologically relevant fluid of a patient can be placed in contact with the solid support in the apheresis column to remove ErbB ligands from the fluid. The serum, blood or fluid can then be reintroduced into the patient.

[0039] The binding molecules can also be used in combination therapies. Thus, the chimeric ErbB ligand binding molecule may be administered in combination with one or more additional compounds or therapies, including chemotherapeutic agents, surgery, catheter devices, and radiation. Combination therapy includes administration of a single pharmaceutical dosage formulation which contains a chimeric ErbB ligand binding molecule and one or more additional agents. The chimeric ErbB ligand binding molecule and one or more additional agent(s) can be administered in their own separate pharmaceutical dosage formulations or together in the same formulation. For example, a chimeric ErbB ligand binding molecule and a cytotoxic agent, a chemotherapeutic agent or a growth inhibitory agent can be administered to the patient together in a single dosage composition or each agent can be administered in a separate dosage formulation. More specifically, the chimeric ErbB ligand binding molecule can be used in combination therapies that include therapeutic agents such as Lapatinib®, Herceptin®, Erbitux® and the like. Where separate dosage formulations are used, the chimeric ErbB ligand binding molecules and one or more additional agents can be administered concurrently, or at separately staggered times, i.e., sequentially. One of skill in the art can appreciate that the combination must be such that the chimeric ErbB ligand binding molecule does not interfere, but rather, accentuates the second therapeutic in the combination. [0040] The following examples arc given by way of illustration only and in no way should be construed as limiting the subject matter of the present application. In the following examples, nucleotide sequences that encode the disclosed amino acid sequences are also contemplated. Many of the following examples disclose the sequence for IgG2Fc fused to the ErbB chimera. It should be appreciated that use of IgGFc is optional. In addition, the conservative replacement of an amino acid with another similar amino acid that does not substantially (about 10-fold) interfere with ligand binding activity is specifically contemplated. Conventional binding studies of the purified products can be used to determine whether substantial differences in binding affinities exist. Many of the structures described below can be better understood by reference to the diagrams of those structures in Figures 1 and 2. All sequences include a signal peptide derived from a mouse antibody heavy chain gene. All numbering is exclusive of the signal peptide, and the first amino acid of each ErbB sequence is underlined.

EXAMPLE 1

[0041] The present example specifically discloses a chimera containing a signal peptide (M E W S W V F L F F LS VTTG V H S) joined to the LI subdomain of the ErbB3 receptor and the Sl and LII subdomains of the ErbBl receptor terminating after residue 501 in module 1 of the ErbBl SlI sub-domain. The ErbB chimera has the following amino acid sequence.

SEQ. ID. NO.1

MEWSWVFLFFLSVTTGVHSSEVGNSQAVCPGTLNGLSVTGD

AENQYQTLYKLYERCEVVMGNLEJVLTGHNADLSFLQWIRE

VTGYVLVAMNEFSTLPLPNLRVVRGTQVYDGKFAIFVMLNY

NTNSSHALRQLRLTQLTEILSGGVYIEKNDKLCHMDTIDWR

DL VRDR DA EI V V KDNG RSCPPCDPSCPNGSCWG AG EENCQK

LTK 11 CAQQCSG RCRG KS PSDCC H N QCAAG CTG P RES DC L VC

RKFRDEATCKDTCPPLMLYNPTTYQMDVNPEGKYSFGATCV

KKCPRNYVVTDHGSCVRACGADSYEMEEDGVRKCKKCEGP

CRKVCNGIGIGEFKDSLSINATNIKHFKNCTSISGDLHILPVA

FRG DSFTH TPPLDPQELDI LKT VKEITGFLLIQA WPEN RTD LH

AFENLEIIRGRTKQHGQFSLAVVSLNITSLGLRSLKEISDGDV

I ISGN KN LCYANTIN WK KLFGTSGQKTKI ISN RG ENSCKATG

QVCHALCSPEGCWGPEPRDCVSVECPPCPAPPVAGPSVFLFP

PKPKDTLMISRTPEVTCVVVDVSHEDPEVQFNWYVDGMEVH

NAKTKPREEQFNSTFRVVSVLTVVHQDWLNGKEYKCKVSN

KGLPAPIEKTISKTKGQPREPQVYTLPPSREEMTKNQVSLTC

LV KG FY PS DLAV E WESNGQPEN N Y KTTPPM LDSDG SFFLY S

KLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK

Stop

Il EXAMPLE 2

[0042] The present example specifically discloses a chimera containing a signal peptide (M E W S W V F L F F L S VTTG V H S) joined to LI and SI subdomains of ErbB3 receptor and the LII subdomain of the ErbBl receptor terminating after residue 501 in module I of the ErbBl SlI sub-domain. The ErbB chimera has the following amino acid sequence:

SEQ. ID. NO.2

MEWSWVFLFFLSVTTGVHSSEVGNSQAVCPGTLNGLSVTGD

AENQYQTLY KLY ERCEV VMGN LEl V LTGH N ADLSFLQWI RE

VTGYVLVAMNEFSTLPLPNLRVVRGTQVYDGKFAIFVMLNY

NTNSSHALRQLRLTQLTEILSGGVYIEKNDKLCHMDTIDWR

DIVRDRDAEIVVKDNGRSCPPCHEVCKGRCWGPGSEDCQTL

TKTICAPQCNGHCFGPNPNQCCHDECAGGCSGPQDTDCFAC

RHFNDSGACVPRCPQPLVYN KLTFQLEPNPHTKYQYGGVCV

ASCPHNFVVDQTSCVRACPPDKMEVDKNGLKMCEPCGGLC

PKVCNG IG IG EF KDSLS IN ATN IKH FKNCTS I SG DLH I LPV A F

RGDSFTHTPPLDPQELDILKTVKEITGFLLIQAWPENRTDLH

AF EN LEl I RG RTKQH GQFS LAV VS LN ITSLG LRSLK El SDG DV l lSGNKN LCYANTINWKKLFGTSGQKTKlISNRGENSCKATG

QVCHALCSPEGCWGPEPRDCVSVECPPCPAPPVAGPSVFLFP

PKPKDTLM ISRTPEVTCVVVDVSHEDPEVQFNWYVDGMEVH

NAKTKPREEQFNSTFRVVSVLTVVHQDWLNGKEYKCKVSN

KG LPA Pl E KTl SKTKGQP R EPQV Y TLP PSRE EMTKN QV S LTC

LVKGFYPSDIAVEWESNGQPENNYKTTPPMLDSDGSFFLYS

KLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK

Stop

EXAMPLE 3

[0043] The present example discloses a chimera containing a signal peptide (M E W S WVF L FFLS VTTG V H S) joined to the LI subdomain and modules 1-2 of the SI subdomain of the ErbB3 receptor and modules 3-8 of the SI subdomain, the LIl subdomain and module I of SlI subdomain of the ErbBl receptor fused to lgG2Fc. The ErbB chimera has the following amino acid sequence:

SEQ. ID. NO.3

MEWSWVFLFFLSVTTGVHSSEVGNSQAVCPGTLNGLSVTGD

AENQYQTLYKLYERCEVVMGNLEIVLTGHNADLSFLQWI RE

VTG Y VLVAMNEFSTLPLPN LRVVRGTQVY DGKFAlFVM LN Y

NTNSSHALRQLRLTQLTEILSGGVYIEKNDKLCHMDTI DWR

DIVRDRDAEI VVKDNGRSCPPCHEVCKGRCWGPGSEDCQTL

TKTICAPQCNGHCFGPNPNQCCHNQCAAGCTGPRESDCLVC

RKFRDEATCKDTCPPLMLYNPTTYQMDVNPEGKYSFGATCV

KKCPRNYVVTDHGSCVRACGADSYEM EEDGVRKCKKCEGP CRK VCNG IG IG EFKDSLSl NATN I K HFKNCTS ISGDLH I LPV A

FRGDSFTHTPPLDPQELDILKTVKEITGFLLIQAWPENRTDLH

A FEN LE I I RG RTKQHGQFSLAVVS LN ITS LGLRSLK ElSDG DV

HSGN KN LCY ANTlN WKKLFGTSGQKTKI ISNRG ENSCKATG

QVCHALCSPEGCWGPEPRDCVSVECPPCPAPPVAGPSVFLFP

PKPKDTLMISRTPEVTCVVVDVSHEDPEVQFNWYVDGMEVH

NAKTKPREEQFNSTFRVVSVLTVVHQDWLNGKEYKCKVSN

KGLPAPIEKTISKTKGQPREPQVYTLPPSREEMTKNQVSLTC

LVKGFYPSDIAVEWESNGQPENNYKTTPPMLDSDGSFFLYS

KLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK

Stop

EXAMPLE 4

[0044] The present example discloses a chimera containing a signal peptide (M E W S W V F L FF LS VTTG V H S) joined to the LI subdomain and modules 1-3 of the SI subdomain of ErbB3 receptor and modules 4-8 of the SI subdomain, the LII subdomain and module I of SlI subdomain of the ErbBl receptor fused to IgG2Fc. The ErbB chimera has the following amino acid sequence:

SEQ. ID. NO.4

M EWSWV FLFFLSVTTGVHSSEVGNSQAVCPGTLNGLSVTGD

A ENQ YQTL YKL Y ERC E V VMGN LEI V LTGHN ADLSFLQWI RE

VTGYVLVAMNEFSTLPLPNLRVVRGTQVYDGKFAIFVMLN Y

NTN SSH ALRQLRLTQLTE1 LSGG V YI EKN DKLCH MDTlDWR

DIVRDRDAEI VVKDNGRSCPPCHEVCKGRCWGPGSEDCQTL

TKTICAPQCNGHCFGPNPNQCCHDECAGGCSGPQDTDCFVC

RKFRDEATCKDTCPPLM LYNPTTYQMDVNPEGKYSFGATCV

KKCPRNYVVTDHGSCVRACGADSYEMEEDGVRKCKKCEGP

CRKVCNGIGIGEFKDSLSINATNIKHFKNCTSISGDLHILPVA

FRGDSFTHTPPLDPQELDI LKTVKEITGFLLIQAWPEN RTDLH

A FEN LEI I RG RTKQH G QF S LAW S LN ITS LG LRS LKEI S DG D V

IISGN KN LCYANTINWKKLFGTSGQKTKIISNRGENSCKATG

QVCHALCSPEGCWGPEPRDCVSVECPPCPAPPVAGPSVFLFP

PKPKDTLM ISRTPEVTCVVVDVSHEDPEVQFN WYVDGMEVH

NAKTKPREEQFNSTFRVVSVLTVVHQDWLNGKEYKCKVSN

KG LPA Pl EKTl S KTKGQPREPQV YTLPPSREEMTKNQVS LTC

LVKGFYPSDIAVEWESNGQPENNYKTTPPMLDSDGSFFLYS

KLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK

Stop

EXAMPLE 5

[0045] The present example specifically discloses a chimera containing a signal peptide (MEWSWVFLFFLSVTTGVH S) joined to the LI subdomain and modules 1-4 of the SI subdomain of ErbB3 receptor with modules 5-8 of the Sl subdomain, the LII subdomain and module 1 of SlI subdomain of the ErbBl receptor fused to IgG2Fc. The ErbB chimera has the following amino acid sequence:

SEQ. ID. NO.5

MEWSWVFLFFLSVTTGVHSSEVGNSQAVCPGTLNGLSVTGD

AENQYQTLYKLYERCEVVMGNLEJVLTGHNADLSFLQWIRE

VTGYVLVAMNEFSTLPLPNLRVVRGTQVYDGKFAIFVMLNY

NTNSSHALRQLRLTQLTEILSGGVYIEKNDKLCHMDTIDWR

DI VRDRDAEL V VKDNGRSCPPCH EVCKGRCWGPGSEDCQTL

TKTICAPQCNGHCFGPNPNQCCHDECAGGCSGPQDTDCFAC

RHFNDSGACVPTCPPLMLYNPTTYQMDVNPEGKYSFGATCV

KKCPRNYVVTDHGSCVRACGADSYEMEEDGVRKCKKCEGP

CRKVCNGIGIGEFKDSLSINATNIKHFKNCTSISGDLHILPVA

FRG DSFTHTPP L D PQ ELDI LKTVK E ITGFLL1QAWP EN RTDLH

AFENLEIIRGRTKQHGQFSLAVVSLNITSLGLRSLKEISDGDV

l lSGN KNLCYANTINWKKLFGTSGQKTKllSNRGENSCKATG

QVCHALCSPEGCWGPEPRDCVSVECPPCPAPPVAGPSVFLFP

PKPKDTLMISRTPEVTCVVVDVSHEDPEVQFNWYVDGMEVH

NAKTKPREEQFNSTFRVVSVLTVVHQDWLNGKEYKCKVSN

KGLPAPIEKTISKTKGQPREPQVYTLPPSREEMTKNQVSLTC

LVKGFYPSDl A VE WESNGQPENN YKTTPPMLDSDGSFFLYS

KLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK

Stop

EXAMPLE 6

[0046] The present example specifically discloses a chimera containing a signal peptide (M EWSWVFLFFLSVTTGVHS) joined to the LI subdomain and modules 1-5 of the SI subdomain of ErbB3 receptor with the modules 6-8 of the Sl subdomain, the LII subdomain and module I of the SII subdomain of the ErbBl receptor fused to IgG2Fc. The ErbB chimera has the following amino acid sequence:

SEQ. ID. NO.6

MEWSWVFLFFLSVTTGVHSSEVGNSQAVCPGTLNGLSVTGD AENQYQTLYKLYERCEVVMGNLEl VLTGHN ADLSFLQWl RE VTGY VLVAMN EFSTLPLPNLRVVRGTQVYDGKFAIFVMLNY NTNSSHALRQLRLTQLTEI LSGGVYI EKNDKLCHMDTI DWR DI VRDRDAEl V VKDNGRSCPPCHEVCKGRC WGPGSEDCQTL TKTICAPQCNGHCFGPNPNQCCHDECAGGCSGPQDTDCFAC RHFNDSGACVPRCPQPLVYNKLTFQLEPNPHTKYQYGGVCV AKCPRNYVVTDHGSCVRACGADSYEMEEDGVRKCKKCEGP CRK VCNG IG IG EFKDS LS INATN I KHFKNCTSISG DLH I LP V A FRG D SFTHTP P LDPQELDl LKTV KEITG F LLI QA W PEN RTD L H A FEN LE II RG RTKQHGQFSLAVVS LN ITS LG LRSLKE1SDG DV 11 SG N KN LCY AN TIN W KKLFGTSGQKTKI I SN RG EN SCKATG QVCHALCSPEGCWGPEPRDCVSVECPPCPAPPVAGPSVFLFP PKPKDTLMISRTPEVTCVVVDVSHEDPEVQFNWYVDGMEVH NAKTKPREEQFNSTFRVVSVLTVVHQDWLNGKEYKCKVSN KG LPA Pl EKTl S KTKGQPR EPQV YTLPPS RE EMTKNQV S LTC LV K G FY PS D IA V EW ES NGQ P EN N Y KTTPPM LD S DG SF F LY S KLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK

Stop

EXAMPLE 7

[0047] The present example describes a chimera containing a signal peptide (M E W S W V F LFFLSVTTG V H S) joined to the Ll subdomain and modules 1-6 of the Sl subdomain of the ErbB3 receptor with the modules 7-8 of the Sl subdomain, the LIl subdomain and module 1 of the SIl subdomain of the ErbBl receptor fused to lgG2Fc. The ErbB chimera has the following amino acid sequence:

SEQ. ID. NO.7

MEWSWVFLFFLSVTTGVHSSEVGNSQAVCPGTLNGLSVTGD

AENQYQTLYKLYERCEVVMGNLEIVLTGHNADLSFLQWIRE

VTGY VLVAMN EFSTLPLPN LRV VRGTQVY DGKFA IFVM LN Y

NTN SS H ALRQLR LTQLTE I LSGG VY I EKN DKLC HM DTl D WR

DIVRDRDAEIVVKDNGRSCPPCHEVCKGRCWGPGSEDCQTL

TKTICAPQCNGHCFGPNPNQCCHDECAGGCSGPQDTDCFAC

RHFN DSGACVPRCPQPLVYNKLTFQLEPNPHTKYQYGGVCV

ASCPH NFVVDQTSCVRACGADSYEMEEDGVRKCKKCEGPC

RKVCNGIGIGEFKDSLSINATNIKHFKNCTSISGDLHILPVAF

RG DS FTHTPPLDPQELD1 LKTVKElTG FLL IQAWPEN RTDL H

A FEN L El I RG RTKQH GQFSLA V VS LN ITSLG LRS LKEI S DG DV

IlSGNKNLCYANTINWKKLFGTSGQKTKllSNRGENSCKATG

QVCHA LCSPEGCWGPEPRDCVSVECPPCPAPPVAGPSVFLFP

PKPKDTLMISRTPEVTCVVVDVSHEDPEVQFN WYVDGM EVH

NAKTKPREEQFNSTFRVVSVLTVVHQDWLNGKEY KCKVSN

KG LPAPI EKTl SKTKGQPREPQVYTLPPSREEMTKNQVS LTC

LV KG FY PSD IAV EWES NGQPEN NY KTTPPMLDSDGSFF LY S

KLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK

Stop

EXAMPLE 8

[0048] The present example discloses a chimera containing a signal peptide (M E W SW VF LFF LSVTTG VH S) joined to the Ll subdomain and modules 1-7 of the SI subdomain of the ErbB3 receptor and module 8 of the Sl subdomain, the LJI subdomain and module 1 of the SII subdomain of the ErbBl receptor fused to IgG2Fc. The ErbB chimera has the following amino acid sequence:

SEQ. ID. NO.8

M EWSWVFLFFLSVTTGVHSSEVGNSQAVCPGTLNGLSVTGD

AENQYQTLYKLYERCEVVMGNLEIVLTGHNADLSFLQWIRE VTG Y V L V A M N E FS T L P L P N L RV V RG TQ V Y DG K FA I F V M L N Y

NTNSSH A LRQLR LTQLTEl LSGGVY IEKN DKLCH M DTI DWR

DI VRDR DA El V VKDNG RSCPPC H EVCKGRCWG PGSEDCQTL

TKTICAPQCNGHCFGPNPNQCCHDECAGGCSGPQDTDCFAC

RH FNDSGACVPRCPQPLVYNKLTFQLEPNPHTKYQYGGVCV

ASCPHNFVVDQTSCVRACPPDKMEVDKNGLKMCEPCEGPCR

KVCNG IG I G EF K DS LS IN ATN I KH FK NCTS I SGDLH ILPV AFR

GDSFTHTPPLDPQELDILKTVKEITGFLLIQAWPENRTDLHA

FENLEIIRGRTKQHGQFSLAVVSLNITSLGLRSLKEISDGDVI

ISG N KN LCY ANTlN WKKLFGTSGQKTKIISNRG ENSCKATGQ

VCHALCSPEGCWGPEPRDCVSVECPPCPAPPVAGPSVFLFPP

KPKDTLMISRTPEVTCVVVDVSHEDPEVQFNWYVDGMEVH

NAKTKPREEQFNSTFRVVSVLTVVHQDWLNGKEYKCKVSN

KGLPAPI EKTISKTKGQPREPQVYTLPPSREEMTKNQVSLTC

LV KG FY PSD IAV EW ESNGQPEN N YKTTPPMLDSDGSFFLYS

KLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK

Stop

EXAMPLE 9

[0049] The present example discloses a chimera containing a signal peptide (M E W S W V F L F F L S VTTG V H S) joined to the Ll subdomain and module 1 of the SI subdomain of the ErbB3 receptor and modules 2-8 of the SI subdomain, the LII subdomain and module 1 of the SIl subdomain of the ErbBI receptor fused to lgG2Fc. The ErbB chimera has the following amino acid sequence:

SEQ. ID. NO.9

MEWSWVFLFFLSVTTGVHSSEVGNSQAVCPGTLNGLSVTGD

AENQYQTLYKLYERCEVVMGNLEIVLTGHNADLSFLQWIRE

VTGY V LVAM N EFSTLPLPN LRV VRGTQV YDG KFAl FVM LN Y

NTNSSHALRQLRLTQLTEILSGGVYIEKNDKLCHMDTIDWR

DIVRDRDAEIVVKDNGRSCPPCHEVCKGRCWGPGSEDCQTL

TKI ICAQQCSGRCRGKSPSDCCHNQCAAGCTGPRESDCLVC

RK FR DEATC KDTCPP LM LYN PTTYQMDVN P EG KY SFG ATC V

KKCPRNYVVTDHGSCVRACGADSYEMEEDGVRKCKKCEGP

CRK VCNGIGIGEFKDSLS INATNIKH FKNCTSISGDLH ILPVA

FRG DSFTHTPPLDPQE LDlLKTV K EITG F LLlQA WPEN RTDLH

AFEN LEI IRGRTKQHGQFSLAVVSLNITSLGLRSLKEISDGDV

IISGNKN LCYANTINWKKLFGTSGQKTKIISNRGENSCKATG

QVCHALCSPEGCWGPEPRDCVSVECPPCPAPPVAGPSVFLFP

PKPKDTLMISRTPEVTCVVVDVSHEDPEVQFNWYVDGMEVH

NAKTKPREEQFNSTFRVVSVLTVVHQDWLNGKEYKCKVSN

KG LPAPI EKTISKTKGQPREPQVYTLPPSREEMTKNQVSLTC

LVKGFY PSDIAVEWESNGQPENNYKTTPPM LDSDGSFFLYS

KLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK

Stop EXAMPLE 10

[0050] The present example discloses a chimera containing a signal peptide (M E W SWVFLFFLSVTTGVH S) joined to the LI subdomain of the ErbB4 receptor and the Sl subdomain, the LIl subdomain and module I of the SlI subdomain of the ErbBl receptor fused to lgG2Fc. The ErbB chimera has the following amino acid sequence:

SEQ. ID. NO.10

M EWSWVFLFFLSVTTGVHSQSVCAGTENKLSSLSDLEQQYR

ALRK Y Y ENCEV VMG N LEITSl EHN RDLSFLRS VRE VTG Y V L

VALNQFRYLPLENLRI I RGTKLYEDRYALAIFLNYRKDGNFG

LQELG LKN LTEl LNGG VYVDQN KFLCYADT1HWQDI VRN PW

PSNLTLVSTNGSSGCGRCDPSCPNGSCWGAGEENCQKLTKII

CAQQCSGRCRGKSPSDCCHNQCAAGCTGPRESDCLVCRKFR

DEATCKDTCPPLM LYN PTTYQMDVNPEGKYSFGATCVKKCP

RNYVVTDHGSCVRACGADSYEMEEDGVRKCKKCEGPCRKV

CN G IG IG E FKDSLSIN ATN IKH FKNCTSISGDLH I LPVAFRG D

SFTHTPPLDPQELDI LKTVKEITGFLLIQAWPENRTDLHAFEN

LEl I RGRTKQHGQFSLAV V S LN ITS LG LRSLKEIS DGDV II SG

N KN LC Y AN Tl N W KK LFGTSG Q KTK I I SN RG EN SC KATGQVC

HALCSPEGCWGPEPRDCVSVECPPCPAPPVAGPSVFLFPPKP

KDTLM ISRTPEVTCVVVDVSHEDPEVQFNWYVDGM EVHNA

KTKPREEQFNSTFRVVSV LTVVHQDWLNGKEYKCKVSN KG

LPAPI EKTISKTKGQPR EPQVYTLPPSREEMTKNQVSLTCLV

KGFYPSDIAVEWESNGQPENNYKTTPPMLDSDGSFFLYSKLT

VDKSRWQQGNVFSCSVMH EALHNHYTQKSLSLSPGK Stop

EXAMPLE 11

[0051] The present example discloses a chimera containing a signal peptide (M E W SWVFLFFLSVTTGVH S) joined to the LI subdomain and the Sl subdomain of the ErbB4 receptor and LIl subdomain and module 1 of the SlI subdomain of the ErbBl receptor fused to lgG2Fc. The ErbB chimera has the following amino acid sequence:

SEQ. ID. NO.11

MEWSWVFLFFLSVTTGVHSQSVCAGTENKLSSLSDLEQQYR A LRK Y Y ENCEV V MGNLEITSI EH N RDLSFLRSVREVTG Y V L VALNQFRYLPLENLRII RGTKLY EDRYALAIFLNYRKDGNFG LQELGLKNLTEILNGGVYVDQNKFLCYADTIHWQDIVRNPW PSN LTLVSTNGSSGCG RCHKSCTG RCWGPTEN HCQTLTRTV CAEQCDGRCYGPYVSDCCHRECAGGCSGPKDTDCFACMNF NDSGACVTQCPQTFVYNPTTFQLEHNFNAKYTYGAFCVKKC PH N FV V DSSSCV RACPSSKM EV EE NG I KMCKPCTDIC PK VC NG I G IG E FKDS LS INATN I KH FK NCTS I SG DLH ILPVA FRG DS FTHTPPLDPQELDILKTVKEITGFLLIQAWPENRTDLHAFENL EIIRGRTKQHGQFSLAVVSLNITSLGLRSLKEISDGDV I ISGN KN LCYA NTlN W KK LFGTSGQKTKI I SN RG EN SCKATGQVC H ALCSPEGCWGPEPRDCVSVECPPCPAPPVAGPSVFLFPPKPK DTLM I S RTP EVTCVVV DVSH EDPEVQFN WY VDGM EV H NAK TKPREEQFNSTFRVVSVLTVVHQDWLNGKEYKCKVSNKGLP APIEKTISKTKGQPREPQVYTLPPSREEMTKNQVSLTCLVKG FY PS Dl A V EW ESNGQPEN N YKTTPPM LDSDG SFFLY SK LTV D KSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK Stop

EXAMPLE 12

[0052] The present example refers to a chimera containing a signal peptide (M E W S WVFLFFLSVTTGVH S) joined to the LI subdomain and the modules 1-2 of the SI subdomain of the ErbB4 receptor and the modules 3-8 of the Sl subdomain, the LII subdomain and module 1 of the SlI subdomain of the ErbBl receptor fused to lgG2Fc. The ErbB chimera has the following amino acid sequence:

SEQ. ID. NO.12

MEWSWVFLFFLSVTTGVHSQSVCAGTENKLSSLSDLEQQYR ALRKY YENCEV VMGN LEITSl EHN RDLSFLRSVREVTG YV L VA LNQFRY LPLEN LRl IRGTKLY EDRYALAIFLN YRKDGN FG LQELG LKN LTE I LNG G VY VDQN KFLC YADTl H WQDl VRN P W PSNLTLVSTNGSSGCGRCHKSCTGRCWGPTENHCQTLTRTV CAEQCDGRCYGPYVSDCCHNQCAAGCTGPRESDCLVCRKFR DEATCKDTCPPLMLYNPTTYQMDVNPEGKYSFGATCVKKCP RN YVVTDHGSCVRACGADSYEMEEDGVRKCKKCEGPCRK V CNG IG IG EFKDS LSI NATN IKH FKNCTSl SGDLH ILPVA FRGD S FTH TP P L DPQ E LDI LKTVK E ITG FL Ll QA WPEN RTD LH A F EN LEl I RG RTKQHGQ FS LAW S LN ITSLG LRSLKE ISDG DV I ISG NKN LCYANTINWK KLFGTSGQKTKIISN RGENSCKATGQVC HALCSPEGCWGPEPRDCVSVECPPCPAPPVAGPSVFLFPPKP KDTLMISRTPEVTCVVVDVSH EDPEVQFNWYVDGM EVHNA KTKPREEQFNSTFRVVSVLTVVHQDWLNGKEYKCKVSNKG L PA Pl EKTI SKT K GQ P REPQV YTL P PS RE EMT K N QV S LTC L V KGFYPSDIAVEW ESNGQPENNYKTTPPMLDSDGSFFLYSKLT VDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK Stop

EXAMPLE 13

[0053] The present example refers to a chimera containing a signal peptide (M E W S WVFLFF LSVTTG VH S) joined to the Ll subdomain and the modules 1-3 of the SI subdomain of the ErbB4 receptor and the modules 4-8 of the Sl subdomain, the LII subdomain and module 1 of the SIl subdomain of the ErbBl receptor fused to lgG2Fc. The ErbB chimera has the following amino acid sequence:

SEQ. ID. NO.13

MEWSWVFLFFLSVTTGVHSQSVCAGTEN KLSSLSDLEQQYR ALRKYY ENCEVVMGNLEITSIEHNRDLSFLRSVREVTGYVL VALNQFRYLPLENLRLLRGTKLYEDRYALAIFLNYRKDGNFG

LQELGLKNLTEILNGGVYVDQNKFLCYADTIHWQDIVRNPW

PSNLTLVSTNGSSGCGRCHKSCTGRCWGPTENHCQTLTRTV

CAEQCDGRCYGPYVSDCCHRECAGGCSGPKDTDCFVCRKFR

DEATCKDTCPPLMLYNPTTYQMDVNPEGKYSFGATCVKKCP

RNYVVTDHGSCVRACGADSYEMEEDGVRKCKKCEGPCRKV

CNGIGIGEFKDSLSINATNIKHFKNCTSISGDLHILPVAFRGD

SFTHTPPLDPQELDILKTVKEITGFLLIQAWPENRTDLHAFEN

L EI I RGRTKQ H GQFS LA VVSLN JTSLG LRS LKEISDGDVI I SG

NKNLCYANTINWKKLFGTSGQKTKIISNRGENSCKATGQVC

HALCSPEGCWGPEPRDCVSVECPPCPAPPVAGPSVFLFPPKP

KDTLMISRTPEVTCVVVDVSHEDPEVQFNWYVDGMEVHNA

KTKPREEQFNSTFRVVSVLTVVHQDWLNGKEYKCKVSNKG

LPAPIEKTISKTKGQPREPQVYTLPPSREEMTKNQVSLTCLV

KGFYPSDIAVEWESNGQPENNYKTTPPMLDSDGSFFLYSKLT

VDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK STOP

EXAMPLE 14

[0054] The present example discloses a chimera containing a signal peptide (M E W SWVFLFFLSVTTGVHS) joined to the Ll subdomain and the modules 1 -4 of the Sl subdomain of the ErbB4 receptor and the modules 5-8 of the SI subdomain, the LII subdomain and module 1 of the SIl subdomain of the ErbBl receptor fused to IgG2Fc. The ErbB chimera has the following amino acid sequence:

SEQ. ID. NO.14

MEWSWVFLFFLSVTTGVHSQSVCAGTENKLSSLSDLEQQYR

ALRKYY ENCEVVMGNLEITSIEHNRDLSFLRSVREVTGYVL

VALNQFRYLPLENLRIIRGTKLYEDRYALAIFLNYRKDGNFG

LQELGLKN LTEl LNGGVY VDQN KFLCY ADTIH WQDI VRN PW

PSNLTLVSTNGSSGCGRCHKSCTGRCWGPTENHCQTLTRTV

CAEQCDGRCYGPYVSDCCHRECAGGCSGPKDTDCFACMNF

NDSGACVTTCPPLMLYNPTTYQMDVNPEGKYSFGATCVKK

CPRNYVVTDHGSCVRACGADSYEMEEDGVRKCKKCEGPCR

KVCNGIG IGEFK DSLSl N ATN IKH FKNCTSISGDLH ILPVAFR

GDSFTHTPPLDPQELDILKTVKEITGFLLIQAWPENRTDLHA

F EN LE I IRG RT KQHGQFSLA V VSLN ITS LGLRS LK EI SDGD VI

I SG N KN LCYANTl N WKK LFGTSGQKTK I IS N RG ENSCKATGQ

VCHALCSPEGCWGPEPRDCVSVECPPCPAPPVAGPSVFLFPP

KPKDTLMISRTPEVTCVVVDVSHEDPEVQFNWYVDGM EVH

NAKTKPREEQFNSTFRVVSVLTVVHQDWLNGKE.YKCKVSN

KGLPAPIEKTI SKTKGQPREPQVYTLPPSREEMTKNQVSLTC

L V KG FY PSDl A V EW ESNGQPEN N YKTTPPM LDS DGSFFLY S

KLTV DKSRWQQGNVFSCSVM HEALHNHYTQKSLSLSPGK

Stop EXAMPLE 15

[0055] The present example discloses a chimera containing a signal peptide (M E W SWVFLFFLSVTTGVH S) joined to the LI subdomain and the modules 1-5 of the SI subdomain of the ErbB4 receptor and the modules 6-8 of the SI subdomain, the LIl subdomain and module I of the SIl subdomain of the ErbBl receptor fused to lgG2Fc. The ErbB chimera has the following amino acid sequence:

SEQ. ID. NO.15

MEWSWVFLFFLSVTTGVHSQSVCAGTENKLSSLSDLEQQYR

A L RK YYENCEV VMGN LEl TS I EHN RDLSF LRS VREVTG Y VL

VA LN QFRY LPL EN LRI IRGTKLY EDRYALAI FLN Y RKDG N FG

LQELG LKN LTEl LNGGVYVDQN KF LCYADTI HWQDl VRN PW

PSNLTLVSTNGSSGCGRCHKSCTGRCWGPTENHCQTLTRTV

CAEQCDGRCYGPYVSDCCHRECAGGCSGPKDTDCFACMNF

NDSGACVTQCPQTFVYNPTTFQLEHNFNAKYTYGAFCVKKC

PRNYVVTDHGSCVRACGADSYEMEEDGVRKCKKCEGPCRK

VCNG IGIGEFKDSLSINATNIKHFKNCTSISGDLHILPVAFRG

DS FTHTPPLDPQ ELDl LKTVKEITG FLLIQAWP EN RTDLH AF E

N LEI I RGRTKQHGQFSLAVVSLN ITSLGLRSLK EISDGDVI IS

GNKN LCYANTINWKKLFGTSGQKTKIISNRGENSCKATGQV

CHALCSPEGCWGPEPRDCVSVECPPCPAPPVAGPSVFLFPPK

PKDTLMISRTPEVTCVVVDVSHEDPEVQFNWYVDGM EVHN

AKTKPREEQFNSTFRVVSVLTVVHQDWLNGKEYKCKVSNK

GLPAPl EKTISKTKGQPREPQVYTLPPSREEMTKNQVSLTCL

VKGFY PSDl A VEWESNGQPENN YKTTPPMLDSDG SFFLYSK

LTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK Stop

EXAMPLE 16

[0056] The present example refers to a chimera containing a signal peptide (M E W S WVFLFFLSVTTGVH S) joined to the LI subdomain and the modules 1-6 of the Sl subdomain of the ErbB4 receptor and the modules 7-8 of the SI subdomain, the LIl subdomain and module 1 of the SIl subdomain of the ErbBl receptor fused to IgG2Fc. The ErbB chimera has the following amino acid sequence:

SEQ. ID. NO.16

MEWSWVFLFFLSVTTGVHSQSVCAGTENKLSSLSDLEQQYR

ALRKYYENCEVVMGNLEITSIEHNRDLSFLRSVREVTGYVL

VALNQFRYLPLENLRIIRGTKLYEDRYALAIFLNYRKDGNFG

LQELGLKNLTEILNGGVYVDQNKFLCYADTIHWQDIVRNPW

PSNLTLVSTNGSSGCGRCHKSCTGRCWGPTENHCQTLTRTV

CAEQCDGRCYGPYVSDCCHRECAGGCSGPKDTDCFACMNF

NDSGACVTQCPQTFVYNPTTFQLEHNFNAKYTYGAFCVKKC

PHNFVVDSSSCVRACGADSYEMEEDGVRKCKKCEGPCRKV

CNGIGIGEFKDSLSINATNIKHFKNCTSISGDLHILPVAFRGD SFTHTPP LDPQELDl LKTVK EITG FLLIQAWPEN RTDLHA FEN LEI IRGRTKQHGOFSLAVVSLN lTSLGLRSLKElSDGDVIiSG NKNLCYANTINWKKLFGTSGQKTKIISNRGENSCKATGQVC HALCSPEGCWGPEPRDCVSVECPPCPAPPVAGPSVFLFPPKP KDTLM iSRTPEVTCVVVDVSHEDPEVQFNWYVDGMEVHNA KTKPREEQFNSTFRVVSVLTVVHQDWLNGKEYKCKVSNKG LPA Pl EKTl SKT K GQ PR E PQV Y TLP PS RE E MTK N Q VS LTC LV KG FY PSD IAV EW ES NGQPEN N Y KTTPPM LDSDG SFFLY S K LT VDKSRWQQGN VFSCSVMH EALHNHYTQKSLSLSPGK Stop

EXAMPLE 17

[0057] The present example refers to a chimera containing a signal peptide (M E W S WVFLFFLSVTTGVH S) joined to the Ll subdomain and the modules 1 -7 of the SI subdomain of the ErbB4 receptor and the module 8 of the Sl subdomain, the LlI subdomain and module 1 of the SlI subdomain of the ErbBI receptor fused to lgG2Fc. The ErbB chimera has the following amino acid sequence:

SEQ. ID. NO.17

MEWSWVFLFFLSVTTGVHSQSVCAGTENKLSSLSDLEQQYR ALRKYYENCEV VMGNLEITSl E H N RDL S F L R S V R EV TG Y VL VA LNQFRY LPLEN LRl IRGTKLY EDRYA LA IFLN YRKDGNFG LQELGLKN LTEl LNGGV Y VDQN KFLCYADTl H WQDlV RN PW PSNLTLVSTNGSSGCGRCHKSCTGRCWGPTENHCQTLTRTV CAEQCDGRCYGPYVSDCCHRECAGGCSGPKDTDCFACMNF NDSGACVTQCPQTFVYN PTTFQLEHNFNAKYTYGAFCVKKC PHNFVVDSSSCVRACPSSKMEVEENGIKMCKPCEGPCRKVC NG IG IG EFKDS LS IN ATN IKH FKNCTSISG DLHl LPVAFRGDS FTHTPPLDPQELD I LKTVKEITGFLLIQAWPEN RTDLHAFEN L EIIRGRTKQHGQFSLAVVSLNITSLGLRSLKElSDGDViISGN KN LCYA NTI N W KKLFGTSGQKTK 11 SN RGEN SCKATGQVCH ALCSPEGCWGPEPRDCVSVECPPCPAPPVAGPSVFLFPPKPK DTLM ISRTPEVTCVVVDVSHEDPEVQFNWYVDGMEVHNAK TKPREEQFNSTFRVVSVLTVVHQDWLNGKEYKCKVSNKGLP APIEKTISKTKGQPREPQVYTLPPSREEMTKNQVSLTCLVKG F Y PSDl A V EWESNGQPENN YKTTPPM LDSDGSFFLYSKLT VD KSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK Stop

EXAMPLE 18

[0058] The present example discloses a chimera containing a signal peptide (M E W S W V F L F F LS V TTG V H S) joined to the LI subdomain and module I of the Sl subdomain of the ErbB4 receptor and modules 2-8 of the Sl subdomain, the LlI subdomain and module I of the SIl subdomain of the ErbBI receptor fused to lgG2Fc. The chimera has the following amino acid sequence: SEQ. ID. NO.18

MEWSWVFLFFLSVTTGVHSQSVCAGTENKLSSLSDLEQQYR

ALRKYYENCEVVMGNLEITSIEHNRDLSFLRSVREVTGYVL

VA LNQFRY LPLEN LRI IRGTKLY EDRYALAI FLN Y RKDGN FG

LQELG LKN LTEl LNGG V Y VDQN KFLC Y ADTIH WQDI VRN P W

PSNLTLVSTNGSSGCGRCHKSCTGRCWGPTENHCQTLTKIIC

AQQCSGRCRGKSPSDCCHNQCAAGCTGPRESDCLVCRKFRD

EATCKDTCPPLMLYNPTTYQMDVNPEGKYSFGATCVKKCPR

NYVVTDHGSCVRACGADSYEMEEDGVRKCKKCEGPCRKVC

NGIGIGEFKDSLSINATNIKHFKNCTSISGDLHILPVAFRGDS

FTHTPPLDPQELDILKTVKEITGFLLIQAWPENRTDLHAFEN L

El I RG RTKQHGQFS LA VVSLN ITSLG LRS LKElSDG DVl ISGN

KNLCYANTINWKKLFGTSGQKTKIISNRGENSCKATGQVCH

ALCSPEGCWGPEPRDCVSVECPPCPAPPVAGPSVFLFPPKPK

DTLM IS RTP EVTCVV V DVSH EDPEVQFN WYV DGM EV HN A K

TKPREEQFNSTFRVVSVLTVVHQDWLNGKEYKCKVSNKGLP

APIEKTISKTKGQPREPQVYTLPPSREEMTKNQVSLTCLVKG

FYPSDIAVEWESNGQPENNYKTTPPMLDSDGSFFLYSKLTVD

KSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK Stop

EXAMPLE 19

[0059] The present example discloses a chimera containing a signal peptide (M E W SWVFLFFLSVTTGVH S) joined to the Ll subdomain and modules 1-5 of the SI subdomain of the ErbB4 receptor (through amino acid residue 245 of the ErbB4 receptor in the alignment of Figure 3) and modules 5-8 of the Sl subdomain of the ErbBl receptor (starting at amino acid residue 249 of the ErbBl receptor in the alignment of Figure 3), the LIl subdomain and module 1 of the SII subdomain of the ErbBl receptor, fused to lgG2Fc after amino acid 501 in the SII subdomain. Amino acid 245 is based on numbering of the ErbB4 receptor while amino acids 249 and 501 are based on numbering of the ErbBl receptor. The chimera has the following amino acid sequence:

SEQ. ID. NO.19

MEWSWVFLFFLSVTTGVHSQSVCAGTENKLSSLSDLEQQYR

ALRKYYENCEVVMGNLEITSIEHNRDLSFLRSVREVTGYVL

VALNQFRYLPLENLRIIRGTKLYEDRYALAIFLNYRKDGNFG

LQ E LG L KN LTE I LN GG VY V DQN KF LCY A DTI H WQD IV RN PW

PSNLTLVSTNGSSGCGRCHKSCTGRCWGPTENHCQTLTRTV

CAEQCDGRCYGPYVSDCCHRECAGGCSGPKDTDCFACMNF

NDSGACVTQCPQTFVYNPTTYQMDVNPEGKYSFGATCVKK

CPRNYVVTDHGSCVRACGADSYEMEEDGVRKCKKCEGPCR

KVC NG IG IG EFKDS LSI NATN I KH FKNCTSL SGDLHI LPVA FR

GDSFTHTPPLDPQ ELDI LKTV KEITG FLL1QAWPEN RTDLH A

FENLEIIRGRTKQHGQFSLAVVSLNITSLGLRSLKEISDGDVI

LSGNKNLCYANTINWKKLFGTSGQKTKILSNRGENSCKATGQ

VCHALCSPEGCWGPEPRDCVSVECPPCPAPPVAGPSVFLFPP KPKDTLMISRTPEVTCVVVDVSHEDPEVQFNWYVDGMEVH NAKTKPREEQFNSTFRVVSVLTVVHQDWLNGKEYKCKVSN KG LPA Pl EKTI S KTKGQPR EPQV YTLPPSREEMTKNQVSLTC LVKG F Y PS D IAV E W ESNGQPEN N Y KTTPPMLD S DG SFFLY S KLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK

Stop

EXAMPLE 20

[0060] The present example discloses a chimera containing the LI subdomain and modules 1-5 of the SI subdomain of the ErbB4 receptor (through amino acid residue 245 of the ErbB4 receptor in the alignment of Figure 3) and modules 5-8 of the SI subdomain of the ErbBl receptor (starting at amino acid residue 249 of the ErbBl receptor in the alignment of Figure 3), the LIl subdomain and module 1 of the SlI subdomain of the ErbBl receptor, fused to lgG2Fc after amino acid 501 in the SII subdomain. Amino acid 245 is based on numbering of the ErbB4 receptor while amino acids 249 and 501 are based on numbering of the ErbBl receptor. Additionally, this sequence contains 2 cysteine (Cys) to serine (Ser) substitutions in the hinge region of lgG2Fc. These substitutions are C226S and C229S, based on numbering of the IgG2Fc protein and are underlined below. The chimera has the following amino acid sequence:

SEQ. ID. NO.20

MEWSWVFLFFLSVTTGVHSQSVCAGTEN KLSSLSDLEQQY R

ALRKYY ENCEV VMGN LElTSI EHNRDLSFLRSVREVTG Y VL

VA LNQFRY LPLEN LRI IRGTKLY EDRY ALAl FLN YR KDG N FG

LQELGLKNLTEI LNGGVYVDQNKFLCYADTIHWQDIVRNPW

PSNLTLVSTNGSSGCGRCHKSCTGRCWGPTENHCQTLTRTV

CAEQCDGRCYGPYVSDCCHRECAGGCSGPKDTDCFACMNF

NDSGACVTQCPQTFVYNPTTYQMDVNPEGKYSFGATCVKK

CPRNYVVTDHGSCVRACGADSYEMEEDGVRKCKKCEGPCR

KVC NG I G IG E FK D SLS IN ATN IKH FKNCTS I SG DLH ILP V A FR

GDSFTHTPPLDPQ ELDl LKTV KE ITG FLLl QA WPEN RTDLH A

FEN LE 11 RG RTKQHGQFS LA VVSLN ITS LG LRSLK El SDGDVl

ISGN KN LCY ANTINWKKLFGTSGQKTKI ISNRGENSCKATGQ

VCHALCSPEGCWGPEPRDCVSVESPPSPAPPVAGPSVFLFPP

KPKDTLM ISRTP EVTC VVV DVSH EDPEVQFN W Y VDG M EV H

NAKTKPREEQFNSTFRVVSVLTVVHQDWLNGKEYKCKVSN

KGLPAPI EKTISKTKGQPREPQVYTLPPSREEMTKNQVSLTC

LVKGFY PSDIAV EWESNGQPENNYKTTPPMLDSDGSFFLYS

KLTVDKSRWQQGNVFSCSVMHEALHN HYTQKSLSLSPGK

Stop EXAMPLE 21

[0061 ] This example demonstrates that ErbB chimeras having substantial binding affinity for ligands to both receptors can be created from the selective combination of two different ErbB receptors. Four constructs were tested for binding to HRG l β and TGFα including the construct from Example 19 (Seq ID No. 19), designated for purposes of this example as E l/E4-Fc, and the construct from Example 20 (Seq ID No. 20) which is designated E l/E4-MFc. The controls included amino acids 1 -501 of ErbB I joined to the lgG2Fc portion, designated E l and amino acids 1 -497 of ErbB4 joined to the lgG2Fc portion, designated E4. In some studies another E l control was used.

[0062] Trap DNA molecules were synthesized by starting with the desired amino acid sequence and optimizing the DNA sequence for mammalian system expression. Trap DNA sequences were cloned into a suitable mammalian expression vector (pCpGfree-vitroHmcs) that can be selected using hygromycin and contains MAR/SAR sequences (insulator and boundary regions) and promoters and enhancers for trap expression. Vectors containing trap sequences were transfected into CHO cells by standard transfection methods and the cells were selected for vector integration with hygromycin. Traps were purified from stably transfected cell lines by collecting cell culture medium and purifying by standard methods (protein A column binding). Traps were eluted from protein A by standard methods and quantitated using a custom derived IgG-Fc sandwich ELlSA assay.

[0063] Traps were purified using protein A and run on a polyacrylamide gel, under non-reducing (NR) and reducing (R) conditions. Disulfide linked dimers run at approximately 220-240 kDa, while reduced monomers run at 120-130 kDa. The chimeric trap monomer with mutated cysteines that prevent disulfide formation runs at approximately 120-130 kDa under both non-reducing and reducing conditions. The El -Fc, E4-Fc, E l/E4-Fc appeared to be in the 220-240 kDa range when run under nonreducing conditions and in the range of about 120- 130 when run under reducing conditions. The E l/E4-Fc construct appeared to have molecular weight of about 120- 130 under both nonreducing and reducing conditions.

[0064] Purified trap molecules were coated on 96 well plates and incubated with either TGFα or HRG I β. Then detection antibodies against TGFα or HRG lβ were used to measure the affinity (Kd) and amount of bound ligand (Bmax).

[0065] Traps were coated on 96 well plates and incubated with varying concentrations of either TGFα or HRG I β. Detection antibodies against TGFα or HRG l β were used to measure the affinity (Kd) and amount of bound ligand (Bmax). Data obtained in these studies is provided in Tables 1-4 below.

[0066] As set forth in Tables I and 2, the chimeric monomer (El/E4-Fc) bound TGFα with an affinity in the range of about 400- 620 nM and ErbBl -IgGFc (El-Fc) bound with an affinity (Kd) in the range of about 27 - 40 nM.

TABLE 1

TGFα Binding Study

TABLE 2

TGFα Binding Study

E1/E4- EI/E4M-

El-Fc E4-Fc El/E4-Fc MFc 50%

Bmax 781,267 280,121 834,302 695,464 779,310

Kd (nM) 40.33 10601 344.8 621.2 1953

Std Error

Bmax 45,287 2,180,000 13,865 137,557 338,644

Std Error

Kd (nM) 9.847 89561 116.3 250.5 1208

[0067] As shown in Tables 3 and 4, the chimeric monomer (El /E4-Fc) bound FTRG I β with an affinity in the range of about approximately 15 -30 nM which was essentially the same as ErbB4-lgGFc (E4-Fc).

TABLE 3

HRG l β Binding Study

TABLE 4

HRG l β Binding Study

[0068] The E l /E4-MFc 50% data point shows that when one half of the amount of the purified chimeric receptor is added to a well that binding goes down by half and therefore binding is proportional to the amount of added receptor.

[0069] This example demonstrates that ErbB chimeras can be created and purified that have substantial affinity for ligands to both of its receptor subcomponents. Further this example shows that mutations can be introduced into the chimeric receptor binding molecules to change disulfides to prevent the formation of dimers through disulfide bond formation.

EXAMPLE 22

[0070] The present example discloses a chimera containing a signal peptide (M E W S W V F L F F L S V T T G V H S) joined to amino acids 1 -245 of the ErbB4 receptor and amino acids 249 to 501 of the ErbB l receptor. The fusion also includes lgG2-Fc as in the sequences above. Also incorporated into this sequence is a substitution of glutamine for serine at position 1 3 and the two cysteine to serine modifications from example 20 that prevent disulfide based dimerization of the molecule. The glutamine at position 13 (underlined below) is found in the ErbB l sequence at that location and is incorporated in the sequence to increase the affinity of the chimera for TGFα. The ErbB chimera has the following amino acid sequence: SEQ. ID. NO.21

MEWSWVFLFFLSVTTGVHSQSVCAGTENKLSQLSDLEQQY R

ALRKYYENCEVVMGNLEITSIEHNRDLSFLRSVREVTGYVL

VA LNQFRY LP LEN LRI I RGTKLY EDRYA LA IFLN Y RKDG N FG

LQELGLKN LTEI LNGGVYVDQNKFLCYADTIHWQDIVRNPW

PSNLTLVSTNGSSGCGRCHKSCTGRCWGPTENHCQTLTRTV

CAEQCDGRCYGPYVSDCCHRECAGGCSGPKDTDCFACMNF

NDSGACVTQCPQTFVYNPTTYQMDVNPEGKYSFGATCVKK

CPRNYVVTDHGSCVRACGADSYEMEEDGVRKCKKCEGPCR

KVCNGlG IG EFKDSLSIN ATN I KHFKNCTSISGDLHILPVAFR

G DS FT HTP P LD PQ ELD ILKTVKE ITG F L LI QA WPEN RTDLH A

FEN LEI IRG RTKQHGQFS LAVVSLN ITSLG LRSLKEISDG DV I

ISGNKN LCY A NTIN WKKLFGTSGQKTKI ISNRGEN SCKATGQ

VCHALCSPEGCWGPEPRDCVSVESPPSPAPPVAGPSVFLFPP

K PKDTLM IS RTPEVTCV VV DVSH EDPEVQFN W Y VDG M EV H

NAKTKPREEQFNSTFRVVSVLTVVHQDWLNGKEYKCKVSN

KG LPAPl EKTl SKTKGQ PREPQV YTLPPS REEMTKNQV S LTC

LV KGFY PS D IA V EWESNGQPEN N Y KTTP PMLDSDGS FFLYS

KLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK

Stop

EXAMPLE 23

[0071] The present example discloses a chimera containing a signal peptide (M E W SW V FLFF LSVTTG V H S) joined to amino acids 1-245 of the ErbB4 receptor and amino acids 249 to 501 of the ErbBl receptor. The fusion also includes lgG2-Fc as in the sequences above. Also incorporated into this sequence is a substitution of tyrosine for serine at position 42 and the two cysteine to serine modifications from example 20 that prevent disulfide based dimerization of the molecule. The tyrosine (underlined below) at position 42 is found in the ErbBl sequence at that location and is incorporated in the sequence to increase the affinity of the chimera for TGFα. The ErbB chimera has the following amino acid sequence:

SEQ. ID. NO.22

MEWSWVFLFFLSVTTGVHSQSVCAGTENKLSSLSDLEQQYR ALRKYY ENC EV VMGNLEITYIEHN RDLSFLRSVREVTGY V L VALNQFRYLPLENLRM RGTKLYEDRYALAIFLNY RKDGN FG LQELGLKN LTEl LNGG V Y VDQNKFLCY ADTIH WQDI VRN PW PSNLTLVSTNGSSGCGRCHKSCTGRCWGPTENHCQTLTRTV CAEQCDGRCYGPYVSDCCH RECAGGCSGPKDTDCFACMN F NDSGACVTQCPQTFVYNPTTYQMDVNPEGKYSFGATCVKK CPRNYVVTDHGSCVRACGA DSYEM EEDGVRKCKKCEGPCR KVCNG IG IG EFK DS LS I N ATN I KH FKNCTS ISG DLH I LPVA FR G DSFTHTPPLDPQ ELD I LKTV KE ITG FLLIQAWPEN RTDLH A FEN LEI I RG RTKQHGQFSLAVVSLN ITS LG LRSLKEISDGDV 1 ISGNKN LCYANTINWKKLFGTSGQKTKI ISNRGENSCKATGQ VCHALCSPEGCWGPEPRDCVSVESPPSPAPPVAGPSVFLFPP KPKDTLMISRTPEVTCVVVDVSHEDPEVQFNWYVDGMEVH NAKTKPREEQFNSTFRVVSVLTVVHQDWLNGKEYKCKVSN KG LPAPL EKTI SKTKGQPREPQVYTLPPS REEMTKNQVSLTC LVKGFYPSDIAVEWESNGQPENNYKTTPPMLDSDGSFFLYS KLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK

Stop

EXAMPLE 24

[0072] The present example discloses a chimera containing a signal peptide (M E W SWVFLF FLSVTTG VH S) joined to amino acids 1-245 of the ErbB4 receptor and amino acids 249 to 501 of the ErbBl receptor. The fusion also includes IgG2-Fc as in the sequences above. Also incorporated into this sequence is a substitution of arginine for tyrosine at position 123 (underlined) and the two cysteine to serine modifications from example 20 that prevent disulfide based dimerization of the molecule. The arginine at position 123 is found in the ErbBl sequence at that location and is incorporated in the sequence to increase the affinity of the chimera for TGFα. The ErbB chimera has the following amino acid sequence:

SEQ. ID. NO.23

MEWSWVFLFFLSVTTGVHSQSVCAGTENKLSSLSDLEQQYR

ALRKYYENCEVVMGNLEITSI EHNRDLSFLRSVREVTGYVL

VA LNQFR Y LPLEN LRl IRGTKLY EDRY A LAlFLN YRKDGN FG

LQE LG LK N LTE ILNG G VRV DQN KFLCYA DTI H WQDI V RN P W

PSNLTLVSTNGSSGCGRCHKSCTGRCWGPTENHCQTLTRTV

CAEQCDGRCYGPYVSDCCHRECAGGCSGPKDTDCFACMNF

NDSGACVTQCPQTFVYNPTTYQMDVNPEGKYSFGATCVKK

CPRNYVVTDHGSCVRACGADSYEMEEDGVRKCKKCEGPCR

K VCNG IG IG EFK DSLSl NATNl KHFKNCTSISGDLH ILPVAFR

G DS FTH TPP L DPQ E LD ILKTV KEITG FLL IQA WPEN RTDLH A

FENLEIIRGRTKQHGQFSLAVVSLNITSLGLRSLKEISDGDVI

ISGN KNLCYANTI NWKKLFGTSGQKTKIISNRGENSCKATGQ

VCHALCSPEGCWGPEPRDCVSVESPPSPAPPVAGPSVFLFPP

KPKDTLMISRTPEVTCVVVDVSHEDPEVQFNWYVDGMEVH

NAKTKPREEQFNSTFRVVSVLTVVHQDWLNGKEYKCKVSN

KGLPAPIEKTISKTKGQPREPQVYTLPPSREEMTKNQVSLTC

LVKGFYPSDIAVEWESNGQPENNYKTTPPM LDSDGSFFLYS

KLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK

Stop

EXAMPLE 25

[0073] The present example discloses a chimera containing a signal peptide (M E W SWV FLFFLS VTTG V H S) joined to amino acids 1-183 of the ErbB4 receptor and amino acids 187-501 of the ErbBl receptor. The sequence is essentially the same as the sequence in Example 18, except that it contains two cysteine to serine modifications in the lgG2Fc region to prevent dimerization (as in example 20). This fusion contains more ErbBI sequence in order to increase the binding affinity of the chimera for TGFα. The ErbB chimera has the following amino acid sequence:

SEQ. ID. NO.24

MEWSWVFLFFLSVTTGVHSQSVCAGTENKLSSLSDLEQQYR

ALRKYY ENCEV VMGN LEITSIEHNRDLSFLRSVREVTGYVL

VALNQFRY LPLEN LRI IRGTKLY EDRYA LAl FLN Y RKDG N FG

LQELGLKN LTEILNGGVYVDQNKFLCYADTIHWQDIVRNPW

PSNLTLVSTNGSSGCGRCHKSCTGRCWGPTENHCQTLTKIIC

AQQCSGRCRGKSPSDCCHNQCAAGCTGPRESDCLVCRKFRD

EATCKDTCPPLM L.Y N PTTYQMDVN PEGKYSFGATCVKKCPR

NYVVTDHGSCVRACGADSYEMEEDGVRKCKKCEGPCRKVC

NGIGIGEFKDSLSINATNIKHFKNCTSISGDLHILPVAFRGDS

FTHTPPLDPQE LDl LKTVKElTG F LL1QAWPENRTDLH AFEN L

El IRGRTKQH GQFSLA VVS LNlTSLG LRSLKEISDGDV1 ISGN

KNLCYANTINWKKLFGTSGQKTKIISNRGENSCKATGQVCH

ALCSPEGCWGPEPRDCVSVESPPSPAPPVAGPSVFLFPPKPK

DTLMISRTPEVTCVVVDVSHEDPEVQFNWYVDGMEVHNAK

TKPREEQFNSTFRVVSVLTVVHQDWLNGKEYKCKVSNKGLP

API EKTI SKTKGQPREPQVYTLPPSREEMTKNQVS LTCLV KG

FYPSDIA VEWESNGQPENNYKTTPPMLDSDGSFFLYSKLTVD

KSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK Stop