CA2120337A1

CA2120337A1 - Composition and treatment with biologically active peptides and antibiotic

Info

Publication number: CA2120337A1
Application number: CA002120337A
Authority: CA
Inventors: Michael A. Zasloff; Barry Berkowitz
Original assignee: Individual
Current assignee: Childrens Hospital of Philadelphia CHOP
Priority date: 1991-10-16
Filing date: 1992-10-16
Publication date: 1993-04-17
Also published as: AU2901692A; WO1993007892A1; JPH07500342A

Abstract

A composition comprising at least one biologically active amphiphilic peptide or protein, said peptide or protein being an ion channel-forming peptide or protein, and an antibiotic selected from the class consisting of non-peptide antibiotics and non-ion channel-forming peptide antibiotics, and derivatives or analogues thereof. The biologically active amphiphilic peptide or protein and the antibioic may be administered in a combined amount effective to inhibit growth of a target cell. The biologically active amphiphilic peptide or protein and antibiotic may protentiate each other.

Description

WO 93~07892 2 1 % ~ 3 ~ 7 PC~/IJS92/088~3 Compo~ition and Treatm~nt ~ith Biologically Act~ve Peptid~s and l~ ibiotic This application is a ce~ntinuation-in-part of Applica'ciDn 5erial No. 402,642, filed Septem~er S, 1989, which is a continuation-in-part of Application 5erial No. 339"292, filed Apri 1 17, 1989 .
This invention relates to biologically activ~ pept~d~s and prc~teins, and more partic:ularly to compositlona; and use~
involving }:~iologically active pept~des or proteins and a~
antibicstic; in particular an An~ib~o~lc whic~l i2; ~ot an ior~
channel-forming peptide or protein, or derlvativ~2s or analogues thereof .
I t has been d~ sclo~ed t~dat agent~ suc~ a~ polymyxin norl~peptide, which di~rupt the outer membrane o~ Grnm-~egative bacteria, increase the poterlcy of certain antil:~iotics,- J3ucl~ ~
~u~idic al~id, novobiocirl, and erythromycin, again~t th~ organi~ms Vil~anen, et al, "5u cept{billty of Bram~N~gatîv~ Bacteria to Polymyxin B Nonapeptide, " =~
Vol. 2~, No. 6, Pgs. 701-70~ (June 1984~. Vil~neal, et al, in ~5uscepti3bility of Gram~ Negative Bacteria to the Synerglst~c Bactericidal Action of Serum and PolylTyxin B Nonapeptlde, n _ n.
J. Microbiol.Vol . ~2 pgs. 66-69 ( $986), disclo~e a ~ynergi~tic efect of polymyxin B n~napeptide and serum in ~actericidal ' action against E. coli strain~, strains of 5almonella ~yphimurium, Xlebisiella species, Enterobacter cloacae, P eudomona~

influenzae. Vaara, et al., have disclosed a ~ynergistic effect of polymyxin B nonapeptide and novobiocin, fusidic acid, ery~hromycin, clindamycin, nafcillin, and cloxacillin against smooth encapsulated E.coli and smooth Salmonella tY~himurium.
Antimicrobial Aaents and Chemotherapy, Vol. 24, No. 1, pg~.
107-113. (July 1983).
In accordance with an aspect of the pre~ent invention, there is provided a compos~tion which includes at least one biologically active peptide or protein, said peptid~ or protein being an ion-channel form~ng peptide or protein; and an antibiotic.
In accordance with anoth~r a~pect of the pre~ent invention, there is provided a process wherein there is administered to a host at least one biologically active peptide or protein, said peptide or protein being an ion channel-forming peptide or protein, and ~n antibiotic.
The antibiotic in which i~ combined is different from such peptide or protein; i.e., ~uch antibiotic iQ other than an ion channel-forming peptide or protein.
Although the invention i~ not to be l~mited to~nny theoretical reasoning, it is believed that the peptides or proteins employed in the present invention interact with the membranes of bacterial cells and such interaction enhances the ability o the above-mentioned antibiotic~ to cro~s the me~brane.
It i8 to be u~derstood~ however~ that ~e scope of the invention i~ not limited to such antibiotics.
Example~ of antibiotics which may be employed include, but are not limited to, a tetracycline(s); a p~eudomonic acld;
viomycin; a cephalo~porin; benzoyl peroxlde; ~thambutol;
isoniazid; ethionamide; a 3ulfonamide; trlmethoprim;
nitroimidazole~; an antimalariral agent; cyclo~erine; a penem antibiotic; an aminoglyco~ide; a hydrophobic antibiotic; a pen~cillin; a monobactam; a 50-S ribo~ome inhibitor; and an antibiotic having a large lipid-like lactone ring.

W093/07892 2 ~ 2 o 3 3 7 PCT/~S92/088~3 Tetracyclines which may be employed include, but are not limited to, tetracycline, doxycycline, oxytetracycline, as well as those isolated from new species of MicromonosPora and Actimonadura, such as, for example, Sch-36969, Sch-332~6, and Sch-34164 (Schering Plough).
In one embodiment, thee pseudomonic acid iE mupirocin (pseudomonic acid A~.
Cephalosporins which may be employed include, but are not limited to cefotaxime; cephalothin; cephalexin; cefazolin;
cephradine; cephapir~n; cefotetan; cefamandole; ceftazidime;
cefoxitin; cefuroxime; cefoperazone; ceftriaxone; ceftizoxime;
ceforanide; cefonicid; cefaclor; 3' quaternary ammonium cephalosporins; broad-spectrum cephalo~porins such as MI-4646, MI-4648, and MI-4659 (Mochida); YM-13115 (Yamanouchi); DQ-2522 and DQ-25~6 (Daiichi); B0-1232 and 80 ~236 (Banyu~; CL-118523 (American Cyanamid), and moxalactam. In o~e embodiment, the cephalosporin is cefotaxime.
Sulfonamides which may be employed include, but are not limit~d to, sulfanilamide; sulfadiazine; sulfabenzamide;
sulfamethoxazole; sulfisoxazole; sulfamethoxazole sulfathalidine;
sulfacetamide; sulfacytine; sulfadoxime; sulfamerazine;
sulfamethazine; sulsulfamethizole; sulfapyridine; sulfasalazi~e;
sulfathiazole; and sulfapyrazone.
Anti-malarial agents which may be employed include, but are not limited to, pyrimethamine; primaguine; chloroguine; quinine, and quinidine.
Penem antibiotics include, but are not limite~ to, imipenem, car~apenems, and 2-(n-azolyl) alkyl-substituted penem~.
Aminoglycoside antibiotics include, but are not limited to, tobramycin, kanamycin, amikacin, the gentamicins (e.g., gentamicin C1, gentamicin C2, gentamicin C1a), netilmicin, kanamycin, neomycin, streptomycin, and deri~atives and analo~ues thereof. The preferred aminoglycosides are tobramycin and the W093/07892 PCT/US~/08823 2l2a3~7 gentamicins. The aminoglycosides, and the bacitracins hereinafter described, tend to be hydrophilic and water-soluble.
Penicillins which may be employed in accordance with the present invention include, but are not limited to benzyl penicillin, ampicillin, methi~illin ~dimethoxyphenyl penicillin), ticaricillin, penicillin V (phenoxymethyl penicillin), oxacillin, cloxacillin, dicloxacillin, flucloxacilli~, amoxicillin, and amidinocillin. Preferred penicillins which may be employed are benzyl penicillin and ampicillin. A preferred monobactam which may be employed is aztreonam.
As representative examples of hydrophobic antibiotics which may be used in the present invention, there may be mentioned macrolides such as erythromycin, roxythromycin, clarithromycin, etc.; 9-N-alkyl derivatives of erythromycin; mid~camycin acetate;
azithromycin; flurithromycin; rifabutin rokitamycin; a 6-0-methyl ery~hromycin A known as TE-031 (Taisho); rifapêntine;
benzypipera~inyl rifamycins such as CGP-7040, C~P-5909, CGP-279353 5Ciba-Geigy); an erythromy~in A derivative with a cyclic carbamate fu~ed to the Cl1/C12 po~ition of a macrolide ring known as A-62~14 (Abbott); AC-7230 ~Toyo Jozo);
benzoxazinorifamycin; difficidin; dirithromycin; a 3-N-piperdinomethylzaino methyl rifamy in ~V known as FCE-222~50 (Farmitalia); M-119-a (Kirin Brewery); a 6-0-met~yl-1-4"-0-carbamoyl erythromycin known as A-6307~
(Abbott); 3-formylrifamycin S~J-hydrazones with diazabicycloalkyl side chains such as CGP-27S~7 and CGP-2986 ~Ciba-Geigy~; and 16-membered macrolides having a 3~0-alpha-~-cladi~o~yl moiety, ~uch as 3-0-alpha-L-cladinosyldeepoxy rosaramicin; tylosins and acyl demycinosyl tylosins.
In addition to the macrolides hereinabove described, rifamycin, rifampin, carbenicillin, and nafcillin may be employed as well.
Other antibiotics which may be used (whether or not hydrophobic) are antibiotics which are S0-S ribosome inhibitors WO 93/07892 2 ~ 2 0 3 3 7 PCT/US92/08823 such as lincomycin; clindamycin; and chloramphenirol; etc.;
antibiotics which have a large lipid like lactone ring, such as mystatin; pimaricin, etc.
The preferred hydrophobic antibiotics are the macrolides and in particular erythromycin and derivatives and analogues thereof.
Peptide antibiotics which are not ion channel-forming peptides or proteins which may be employed include, but are not limited to, a bacitracin; gramacidin S; polymyxin; vancomycin;
teichoplanin; and capreomycin; and derivatives and analogues thereof.
The biologically active amphiphilic peptideq employed in the present invention are generally water soluble to a concentration of at least 20 mg/ml at neutral pH in water. In addition, the structure of such peptide provides for flexibility of the peptide molecule. When the peptide i~ placed in wa~er, it does not assume an amphiphilic structure. When the p~ptide encoun~ers an oily surface or membrane, the peptide chain fold~ upon itself into a rod-like or alpha-helical structure.
In general, such peptides ha~e at lea~t 11 amino acids, and preferably at least 20 amino acid~. In most ca~es, such peptides do not have in excess of 50 amino acids.
In general, the biolo~ically active peptides or proteins~
employed in the present in~ention are ion cha~nel-forming peptides or proteins. An ion channel-orming peptide or protein or ionophore is a peptide or protein which increa~es the permeability for ions across a natural or synthetic lipid membrane. B. Christensen et al. PNAS ~ol. 85 P. 5072-76 (July, 1988~ describes methodology which indicates whether or not a pep~ide or; protein has ion channel~forming prop~rties and is therefore an ionophore. As used herein an ion channel-forming peptide or protein is a peptide or protein which has ion channel-forming properties as determined by the method of Christensen et al.

WO 93/0?892 PCT/US92/08823 212~3~7 An amphiphilic peptide or protein is a peptide which includes both hydrophobic and hydrophilic peptide regions.
Although the biologically active amphiphilic (amphipathic) ion channel-forming peptides or proteins are capable of forming ion channels, the ability of such peptides or proteins and the above-mentioned a~,tibiotics to potentiate each other is not necessarily dependent upon the antibiotic crossing a membrane through such channels. Thus, although the ability to form ion channels may be a characteristic of a type of peptide or protein used in the invention, the invention is not limited to the formation and/or use of such channels as part of the mechani~m for the peptide or protein potentiating the antibiotic or vice versa. Similarly, although Applicant believes that such peptides or proteins interact with the membrane of bacterial cells and such interaction is the mechanism by which the antib.~.otic potentiates the peptide or protein and vice versa, the pre~sent invention is not limited to such a mechanism.
The term "potentiate", as used herein, means either that the biologically active amphiphilic peptide or protcin is e~fective in increasing the biological activity of the above-mentioned antibiotics against a target cell so thereby the antibiotic may be employed in an amount lower than that which would be requi~ed for preven~ing, destroying or inhibiting growth of a tar~et cell, and/or that the peptide or protein may be employed in~an amount lower than that which would be required for preventing, destroying, or inhibiting growth of a target c~llo The administration of the biologically active amphiphilic peptides or proteins and antibiotic to a target cell may be direct administration to the cell or ~ystemic or topical administration to a host which includes the target cell, in order to prevent, destroy, or inhibit the growth of a target cell.
Target cells whose growth may be prevented, inhibited, or destroyed by the administration of the biolog~cally active W093/07892 2 ~ ~ 0 3 3 7 PCT/US92/OW23 amphiphilic peptide or protein and antibiotic include Gram-positive and Gram-negative bacteria.
For example, erythromycin, when employed without the above-mentioned peptides, is effective only against Gram-positive organisms. Applicants have found unexpec~edly that erythromycin, when employed in combination with the above-mentioned peptides or proteins, is potentiated such that it becomes biolcgically effective against Gram-negative bacteria. Moreover, the erythromycin may be employed against Gram-positive bacteria in amounts lower than tho~e normally used. Furthermore, such a res~lt can be achieved by using peptide or protein amounts lower tha~ those normally used.
The peptides or proteins employed in the present invention are capable of interacting selectively with membranes of bacteria.
In general, the peptide or protein is employed to provide peptide dosages of from 1 mg to 500 mg per kilogram of host weight, when administered systemically. When administered topically, the peptide or protein is used in a concentration of from 0.1% to 10%.
The antibiotic, such as those hereinabove described, or derivatives or analogues thexeof, when used topically, is generally employed in a concentration of about 0.1% to about 10%.
When used systemically, the anti~iotic or derivative or analogue thereof is generally employed in an amount of from O.lmg to about 45mg per kg of host weight per day.
The use of a combination of peptide or protein and an antibiotic such as those hereina~ove described, or derivatives or analogues thereof in a~cordance with the present invention is effective as an antibiotic, and may be employed to inhibit, prevent or destroy the growth or proliferation of microbes, such as bacteria.
The compositions have a broad range of poterlt antibiotic activity against a plurality of microorganisms, including WO ~3/07892 PCT/US92/08823 212~33~ ~

Gram-positive and Gram-negative bacteria. Such compositions may be employed for treating or controlling microbial infection caused by organisms which are sensitive to such composition. The treatment may comprise administering to a host organism or tissues acceptable to or affiliated with a microbial infection an anti-microbial amount of such peptide or protein and an antibiotic.
The compositions may also be used as preservatives or sterilants for materials susceptible to microbial contamination.
In accordance with a preferred embodiment, the peptide used in conjunction with an antibiotic such as those hereinabo~e described, or derivatives or analo~es t~ercof i8 a basic (positively charged) polypeptide having at least sixteen amino acids wherein the polypeptide includes at least eight hydrophobic amino acids a~d at least eight hydrophilic amino acids. Still more particularly, the hydrophobic amino acids are in groups o two adjacent amino acids, and each group of two hydrophobic amino acids i8 spaced from another group of two hydrophobic amino acids by at least one amino acid other than a hydrophobic amino acid (preferably at least two amino acids) and generally by no greater than four amino acids, and the amino acids ~etwe~n pairs of hydrophobic amino acids may or may not be hydrophilic.
The hydrophilic amino acids are generally also in groups of two adja~ent amino acids in which at lea~t one of ~he~two amino acids is a basic hydrophilic amino acid, with ~uch groups of two hydrophilic amino acids being ~paced from each other ~y at least one amino acid other than a hydrophilic amino acid (preferably at least two amino acids) and ~enerally no greater than four amino acids, and the amino acids between pairs of hydrophilic amino ~cids may or may not be hydropho~ic.
In accordance with a particularly preferred embodiment, the polypeptide comprises a chain of at least four group3 of amino acids, with each group consisting of four amino acids. Two of the four amino acids in each group are hydrophobic amino acids, WO ~3/07892 2 1 2 0 3 3 7 PCT/US92/08823 and two of the four amino acids in each group are hydrophilic, with at least one of the hydrophilic amino acids in each group being a basic hydrophilic amino acid and the other beinq a basic or neutral hydrophilic amino acid.
The hydrophobic amino acids may be selected from the class consisting of Ala, Cys, Phe, Gly, Ile, Leu, Met, Val, Trp, Tyr, norleucine (Nle), norvaline (Nva), and cyclohexylalanine (Cha).
The neutral hydrophilic amino acids may be selected from the class consisting of Asn, Gln, Ser, snd Thr. The ba~ic hydrophilic amino acids may be selected from the class con~isting of Lys, Arg, and His, Orn, homoarginine (Har), 2, 4-diaminobutyric acid (Dbu), and p-aminophenylalanine.
Each of the groups of four amino acids may be of the seguence ABCD, BCDA, CDA~, or DABC, wherein A and 8 are each hydrophobic amino acids and may be the same or different, one of C or D is a basic hydrophilic amino acid, and the other of C or D
is a basic or neutral hydrophilic amino acid and may be the same or different. In a preferred embodiment, the polypeptide chain may comprise 5 or 6 groups of one or more of the~e ~equences. In each group, each of A, B, C and D may be the Jame in ~ome or all of the groups or may be different in some cr all of the groups.
The polypeptide chain preferably has at least 16 amino acids, and ~o greater than 50 amino acids. It is to be understood, however, that the polypeptide does not have to consist entirely of the groups described abo~e. The polypeptide may have amino acids extendirlg rom either or both end~ of the noted groups fo~ming the polypeptide chain and/or there may be amino acids between one or more of ~he at le~st four groups and still remain within the scope of the invention.
The groups of amino acids may be repe~t1 ng groups of amino acids, or the amino acids in the various groups may vary provided that in each group of the at least four groups of amino acids there are two hydrophobic and two hydrophilic amino acids as hereinabove noted.

W093/07892 PCT/U~92/~8X23 212~3~7 ~

-10- , ,, Thus, in a preferred embodiment, the biologically active polypeptide comprises a chain including at least four groups of ,~
amino acids, each containing four amino acid~. Two of the four amino acids in each group are hydrophobic, at lea~t one amino acid is basic hydrophilic, and the remaining one i8 basic or neutral hydrophilic, with the polypeptide chain preferably having at least 20 amino acids but no greater than 50 amino acids.
In one em~odiment, each of the at least four groups of àmino acids which are in the peptide chain is of the seguence A-B-C-D, B-C-D-A, C-D-A-B or D-A-8-C wherein A and ~ are hydrophobic ~:
amino acids, one of C or D is basic hydrophilic amino acid, and the other of C or D is basic or neutral hydrophilic amino acid. :~
The resulting polypeptide chain, therefore, may have one of the following sequences:
(X1)a(A~B~C~D)n(Yl)b (X2 ) a(B~C~D~A)n(Y2 )~
(X3)a~C-D-A-B)n(Y3)b (x4)a(D-A-~-c)n(y4)b wherein X1 is D; C-D- or B-C-D-, Y1 is -A or :~
-A-B or -A-B-C -:

X2 is A-, D-A- or C-D-A-Y2 is -B, -B-C or B-C-D
X3is B-, A-B-, D-A-B-Y3 is -C, C-D, -C-D-A .
X~is C-, B-C-, A-B-C-Y4 is -D, -D-A, -~-A-~
a i5 0 or 1; b is 0 or 1 and n is at least 4.
It is to be understood that the peptide chain may include amino acids between the hereinabo~e noted groups of four amino acids provided that the spacing between such groups and the charge on the amino acids does not change the characteristics of WO93J07892 2 1 2 Q 3 3 7 PCT/US~2/08823 the peptide chain which provide amphiphilicity and a positive charge and do not adversely affect the folding characteristics of the chain to that which is significantly different from one in which the hereinabove noted group of four amino acids are not spaced from each other~
As representative examples of peptides in accordance with the present invention, there may be mentioned.
I Ala-Phe-Ser-Lys-Ala-Phe-Ser-Lys-Ala-Phe-Ser-Lys-Ala-Phe-Ser-Lys-Ala-Phe-Ser-Lys (SEQ ID
NO~
II Ala-Phe-Ser-Lys-Ala-Phe-Ser-Lys-Ala-Phe-Ser-Lys-Ala-Phe-Ser-Lys-Ala-Phe-Ser-Lys-Ala-Phe-Ser-Lys. (SEQ ID NO:2) III Phe-Ser-Lys-Al~-Phe-Ser-Lys-Ala-Phe-Ser-Lys-Ala-Phe-Ser-Lys-Ala- (SEQ ID NO:3).
IV Ser-Lys-Ala-Phe-Ser-Lys-Ala- -Phe-Ser-Lys-Ala-Phe-Ser-Lys-Ala-~he-Ser-Lys-Ala-Phe- (SEQ ID NO:4~ -V Lys-Ala-Phe-Ser-Ly~-Ala-Phe-Ser-~ys-Ala-Phe-Ser-Lys-Ala-Phe-Ser (SEQ ID NO:5) The peptide may have amino acids extending from either en~d of the chain. For example, the chains may have a Ser-Lys seguence before the "Ala" end, and/or an Ala-Phe seguence after the "Lys" end. Other amino acid seguences may al~o be attached to the "Ala" and/or the "Lys" end.
Simil~rly, in any polypeptide chain having at least four ~`
groups of amino acids of the se~ence as described above, the chain may have, for example, a C-D seguence before the first A-B-C-D group. Also other amino acid sequences may be attached to the "A" and/or the "Dl' end of one of these polypeptide chains Also there may be amino acids in the chain which ~pace one or more groups of the hereinabove noted four amino acids from each other.

WO 93/07892 P~T/US92/08823 2120~7 ::

The peptides may ~e produced by known techni~ues and obtained in substantially pure form. For example, the peptides may be synthesized on an automatic chemical peptide synthesizer.
Journal of the American Chemical Society, Vol. 85 Pages 2149-54(1963). It is also possi~le to produce ~uch peptides by recombinant ~NA technology using genetic engineeri~g techni~ues.
Methods in Enz~molo~Y, Vol. 68: "Recombinant DNA" (Wu, et al.) Academic Press, New York (1979); and Methods in Enz~oloaY~ Vol.
18~: "Gene Expression Technology" (~.V. Goeddel, et al.) Academic Press, New York (l990).
In accordance with another preferred embodiment, the peptide employed in conjunction with an antibiotic such as those hereinabove described, or derivatives or analogues thereof may be a magainin peptide.
A magainin peptide is either a magaini~ such as Magai~in I, II or III or an analogue or derivative thereof. The magainin peptides may inc'ude the following basic peptide structure X12 Rll R11 R12 ~13 Rll R14 R12 Rll R14 R12 ~ Rll R14a- (R15 ~n-R14a-R14 ~~
wherein R11 is a hydrophobic amino acid, R12 is a basic hydrcphilic amino acid; Rl3 is a hydrophobi~, neutral hydrophilic, or basic hydrophilic amino acid; Rl4 and R14a are~ -hydropho~ic or basic hydrophilic amino acids, Rl5 ~ 5 ~l~tamic acid or aspartic acid, or a hydrophohic or basic hydrophilic amino acid, and n is 0 or 1. In a preferred ~mbodiment, ~13 is a hydrophobic or net~tral hydrophilic amino acid, Rl4a is a hydropho~ic amino acid, and R15 is glutamic acid or aspartic acid.
Thus, for example, a magaini~ peptide may inrlude the following structure:
Y12 X12' where X12 is the here-~nabove described basic peptide structure and Y12 is (i) R12; .

2120~37 (ii) R14 - R12;
(iii) Rll-R14 -R12; or (iv) ~14 Rll R14a R12 Rll, R12, R14, and R14a are as previously defined.
A magainin peptide may al80 have the following structure:
-X12-Z12, wherein X12 is as previously defined and Z12 i5:
(i) R16 where R16 is a basic hydrophilic amino a~id or asparagine or glutamine; or ~:
(ii) R16-R17 where R17 is a neutral hydrophilic amino acid, a hydrophobic amino acid, or a basic hydrophilic amino acid. Preferably, R17 is a neutral hydrophilic amino acid.
A magainin peptide may al50 have the following structure (Y12)a X12 (Z12)b where Xl7, Y1 , and Z12 are as previously defined, and a is or 1 and b is 0 or 1. -:~
The magainin peptides may also include the f~llowing basic -~
peptide structure X13:
14 ~11 R14a R12 Rll~R~ 12~R13~~ R14~
12 R11 Rll ~12-' Rll' R12' R13, R14, and R14a are amino acids as here~na~ove describ~d.
The magainin peptide may also include the fo~lowing ~:
structure X13-Z~3; wherein X13 i8 the hereinaboYe described basic peptide structure and Z13 is (Rll)n-(Rll)n~ l)n-(Rl4a)n (Rls)n ( 14a)n (R14) ~~R16)n-(R17~n-~ whe~ein R11~ R14~ R14a' R15' 16~
R17 are amino acids as hereinabo~e described, and n is O or 1, and each n may be the same or different.
The magainin peptides generally include at least fourteen amino acids and may include up to forty amino acids. A magainin peptide preferably has 22 or 23 amino acids. A~cordingly, the hereinabove described basic peptide structures of a magainin peptide may incl~de additional amino acids at the amino end or at the carboxyl end, or at ~oth ends. ..
As representative examples of such magainin peptides, there may be mentioned peptides having the following primary sequences as given in the accompanying sequence listing, as well as appropriate analogues and derivatives thereof:
(a) (NH2) (SEQ ID NO:6) (OH) or (NH2) (Magainin I) (b) (NH2) (SEQ ID NO:7) (O~) or (NH2) :~
(Magainin II) (c) (NH2) ~SEQ ID NO:8) (OH) or (NH2) (Magainin III) The following are examples of peptide derivatives or analo~s .
t~f the basi~ structure:
(d) ~NH~) (SEQ ID NO:9) (OH) or (NH2, .-~e) (NH23 (SEQ ID NO:lO) (9H) or (NH23 (f) (NH2) ~SEQ ID NO:ll) (OH) or (NH2) .-Magainin peptides are described in Proc. Natl. Acad Sc~. ~
Vol. 84, pp. 5449-53 (Aug. 1987). The term "magainin peptides" ~i as used herein refers to the basic magainin stru~ture as well as derivatives and analogs thereof, including but not limited to the representative derivatives or analogs.
In ac~ordance with a further embodiment, the peptide employed in conjunctio~ with an antibioti~ such as bac~tracin, tobramycin or gentamicin or derivatives or analogues thereof may be a pGr~a peptide or an XPF peptide.
A PGLa peptide is either PGLa or an analogue or derivative thereof. The PGLa peptides preferably include the following basic peptide ~tru~ture X14:
~ 17 ~12 ~ 14 ~
Rll R14 ~12 Rll Rll R12 11 Rll Rll ~12 where R~ 12~ Rl4, and R17 are as previously defined.

wo g3/~,892 2 1 2 o 3 3 7 Pcr/US92/08823 The PGLa peptides generally include at least seventeen amino acids and may include as many as forty amino acids. Accordingly, the hereinabove described basic peptide stru~ture for a P~La peptide may include additional amino acids at the amino end or at the carboxyl end or at both the amino and carboxyl end.
Thus, for example, a PGLa peptide may have the following struct~re:

where X14 is as previously defined and Y14 iS
(i) Rll;
( ii ) R14 Rll where Rll is as pre~iously defined.
For example, a PGLa like peptide may also have the following structure:
14 14' where X14 is as previously defined; and Z14 is:
~i) Rll; or ~ ii ) 11 11 " ' where Rll is as previously defined.
A PGLa peptide may als~ have the following structure: !'`
(Y14)a X14 (Z14)~
where X14; Y14 and Z14 are as previously deined,~ a is O or 1 and b is O or 1.
An XPF peptide is either XPF or an analogue or derivative thereof. The XPF peptides preferably include the following ~asic peptide structur* X16:
Rll R17 R12~ 14-R18-R17-Rl1 R14-R12-Rl1-;Rl1-R12-Rll-Rll- Rll-R12-~15-Rll--~
Rl1, R12, R14, R15 and ~17 are as previously defined ~nd R18 is glutamine or asparagine.

The XPF peptides generally include at least nineteen amino acids and may include up to forty amino acids. Accordingly, the hereinabove described basic peptide structure of XPF may include ~:
additional amino acids at the amino end, or at the carboxyl end or at both the amino and carboxyl ends. :
Thus, for example, an XPF peptide may include the following -~-structure:
Y16 X16 ' where X16 is as previously defined and Y16 is (i) Rll or (ii) 14 where Rll and R14 are is as previously defined. ~
An XPF peptide may include the following structure: ~:
-X16-Z16, where X16 is as previously defined and Z16 is ~i) R11; or (ii) R11 R18; or (iii) 11 Rlg-Proline; or ( iv ) Rll-R18-Prline R12 An XPF peptide may also have the following struc~ure: :
(Y16~a 16( 1~)b -~.
where X16, Y16 and ~16 are as previously defined: a is 0 or 1 and b is 0 or 1.~
Preferred are XPF or PGLa peptides, which are characteri~ed by the following primary amino acid sequence as given in the accompanying sequenc~ listing:
PGLa : (SEQ ID N0:12) (NH2) XPF : (SEQ ID N0:13) A review of XPF and PGLa can be found in Hoffman et al, EMB0 J. 2:711-714; 19~3; Andreu et al, J B ochem. 149:531-535, 1985;
Gibson et al J. Biol. Chem. 261:5341 5349, 1986; and Giovannini et al, Biochem J. 243:113-120, 1987. ~

W093/07X92 2 1 2 ~ 3 3 7 PCT/US92/08823 In accordance with yet another embodiment, the peptide employed in conjunction with an antibiotic such as those hereinabove described, or derivatives or analogues thereof may be a CPF peptide or appropriate analogue or derviative thereof.
A basic CPF peptide structure as well as analogues and derivatives thereof are herein sometimes referred to collectively as CPF peptides.
The CPF peptide is preferably one which includes the following peptide structure X30: `
21 ~21 R22~R22~~21~R21~R23~R2~
-R21-R21-~'23-~?'21-~21-R24-R25-R21-wherein R21 is a hydrophobic amino acid;
R22 is a hydrophobic amino acid or a basic hydrophilic amino acid;
R23 is a basic hydrophilic amino acid; and R24 is a hydrophobic or neutral hydrophilic amino acId; and ~:
R25 is a basic or neutral hydrophilic amino a~id.
The hereinabove basic structure ic hereinafter symbolically indicated as X30.
The hydrophobic amino acids may be Ala, Cys, Phe, Gly, Ile, Leu, Met, Val, Trp, and Tyr.
The neutral hydrophilic amino acids may be Asn, Gln, Ser, and Thr.
The basic hydrophilic amino acids may be Ly~, Arg, and ~is, Orn, homoarginine (Har), 2, 4-diaminobutyri~ acid lDbu), and p-aminophenylalanine.
The CPF peptide may include only the hereinabove noted amino acids or may include additional amino acids at the amino end or car~oxyl end or both the amino and car~oxyl ~nd. In g~neral, the peptide does not include more than 40 amino acid3.
The CPF peptides including the above ba~ic peptide structure may have from 1 to 4 additional amino acids at the amino end.
Accordingly, such preferred peptides may be represented by the structural formula:

~V~ 93/07X92 PCT/US92/08823 wherein X30 is the hereinabove described basic peptide -~
structure and Y30 is (i) R25-' or (ii) R22 R25; or (iii) R2l-R22-R25 or (iv) 22 R2l R22~R25; preferably GlyCine -3~21-R22 R25 wherein R2~, R22, and R25 are as previously defined.
The carboxyl end of the basic peptide structure may also --have additional amino acids which may r~nge from l to 13 additional amino acids. ~-~
In a preferred em~odiment, the basic structure may have from 1 to 7 additional amino acids at the carboxyl end, which may be represented as follows~
-X30-Z30 wherein X30 is the hereinabove defined basic peptide structure and Z30 is ~:
(i) R21 ' ~:
( ii ) R21 R21 Iiii) R2l R21 ~24;
(iv) R2l R2l R2~ R~4;
(~) R2l i~21 ~24 ~24 R26;
(vi) R2l R2l R24~R24~~26-Gln; or ~vii ) R21 R21 i~24-~24-R26-G~ Gln~
wherein R21 and R24 are as previously defined, and R26 is proline or a hydrophobic amino acid.
Preferred peptides may be represented by the following structural ormula:
(Y30)a 30 (Z30)b wherein X30, Y30 and Z30 are as previously defined and a is 0 or l a~d b is 0 or l.

WO ~3/0789Z 2 1 2 o 3 3 7 PCT/USg2/-88 - 19- `~ .
~ .
Representative examples of CPF peptides which are useful in the present invention have been described in the literature and comprise the following sequences as given in the accompanying sequence listing:
(SEQ ID N0:14) (SEQ ID N0:15) (SEQ ID N0:16) `~
(SEQ ID N0:17) (SEQ ID N0:18) (SEQ ID N0:19) `
(SEQ ID N0:20) (SEQ ID N0:21) (SEQ ID N0:22) (SEQ ID N0:23) (SEQ ID N~:24) (SEQ ID N0:25) (SEQ ID NO:26) A review of the CPF peptides can b- found in Richter, K., Egger, R., and Kreil (1986) J. Biol. Chem. 261, 3676-3680;
Wakabayashi, T. Kato, H., and Tachibaba, S. (198~) Nucleic Acids '`5'~' Research 13, 1817-1828; Gibson, B.W., Poulter, L., Williams, D.H., and Maggio, J.E. (1986) J. Biol. Chem. 261, 5341-5349.
CPF peptides which may be employed in the present invention are represented by the following (single letter amino acid ~ode):
G12S3LG4ALKA5LKIG678LGG9(10)QQ
Where: -~
1 = F, L
- 2 = G, A
3 = F, L -4 = K, L
= A, G, T
6 = A, T
7 = H, N
8 = A, M, F, L

W093/07892 PCT/~S92/08823 212~337 ~

9 = A, S, T
= P, L
The numbered amino acids may be employed as described in any ~;~
combination to provide either a basic CPF peptide ~tructure or an analogue or derivative~ The term CPF peptide includes the ba~ic peptide structure as well as analogues or derivatives thereof. ~
In accordance with yet another embodiment, the peptide may ~ :
include one of the following basic structures X31 through X37 wherein: :
X31 is lR3l-R32-~32-R33-R3l-~32~R32l~n; -X32 is [R32 R32 ~33 R31~R32~R32~R313~n;
X33 is lR32 R33 R31 ~32 ~32 ~31 ~32] n;
X34 is lR33-~31-R32-R32-R31-R32-R32~-n;
X35 is -1R31-R32-R32-R31 R32 R32 33 n X36 is IR3~-~32-R3l-R32-R32-R33-R3l~- ; and 37 lR32 R31 R32-R32-~33-~31-R321-n;
wherein R31 is a basic hydrophilic amino ac~d, R32 i~ a hydropho~ic amino acid, R33 is a neutral hydrophilic or hydrophobic amino acid, and n is from 2 to S.
The basic hydrophilic amino acids may be selected from the class consisting of Lys, Arg, ~is, Orn, homoarginine (Har), 2,4-diamino-butyric acid (Dbu), and p-aminophenylalanine.
The hydrophobi~ amino acids may be selected ~rom the class consisting of Ala, Cys, Phe, Gly, Ile, Leu, Net, Val, Trp and Tyr,norleucine ~Nle), norvaline (Nval), and cyclohexylalanine (Cha).
The neutral hydrophilic smino acids may be selected from the class consisting of Asn, Gln, Ser and Thr.
In accordance with one embodiment, when ~he peptide includes the structure X31, the peptide may include the following structure:
Y31-X31, wherein X31 is as hereina~ove described, and Y
is:
(i) ~32; `

WO 93/07892 2 1 2 ~ ~ 3 7 PCT/US92/08823 ~

-21- ~:

(ii) R32-R3Z;
(iii) R31-R32 R32;
(iV) R33-R31-R32 R32;
(v) R32-R33-R3l R32 32 ( ) R32 ~32 R33 R3l-R32-R32~ wherein R31, R32, and R33 are as hereinabove described. -:
In accordance with another em~odiment, when the peptide ~::
includes the structure X31, the peptide may include the following structure:
X31-Z3l, wherein X3l is as hereinabove described, and Z31 is:
(i ~ R31;
(ii) R31 R32;
(iii) R31-~32 R32;
(iv) R31-R32 R32 33 (v) R31 R32-R32-R33-R31; or ~ vi) R3l-R3~-R32-R33 ~31 3Z
In accordance with yet another embodiment, the peptide may ~:
include the following structure: -(Y3l)a-x3l-(z3l)b~ wherein Y31 and Z31 are as pre~iously defined, a is 0 or 1, and b is 0 or l.
When the peptide includes the structure X32, the peptideCmay include the following structure: ;
Y32 ~ X32, wherein X32 is as hereinabo~e described, and Y32 i s :
(i) ~31; :~
(ii~ R32-R31;
(iii) R32-~32 ~31 (iv) R3l~3~-R32 ~31' R33 R31 R32 R32 31;
R32-R33-~31 R32 R32 R31-In another embodiment, when the peptide includes the structure X32, the peptide may include the following structure: :

2l 20337 -22~

X32 ~ Z32~ wherein X32 is as hereinabove described, and Z32 i s :
( ) R32;
(ii) R32-R32;
(iii) R32-R32 R33;
(iv) R32-R32-R33 ~31;
(v) R3~-R32-~33-R31-R32; or (vi) R32-R32-~33 R31 ~32 R32-In accordance with yet another embodiment, the peptide may :-include the following structure:
(Y32)a X32 (Z3Z)b' wherein Y3~ and Z32 are as previously defined, a is 0 or 1, and b is 0 or 1.
In accordance with another embodiment, when the peptide includes the structure X33, the peptide may include the following structure:
Y33 - X33 wherein ~33 is as hereinabove described, and Y33 is:
(i) R32;
(ii3 R3i R32;
(iii) R32-~31 R32;
(iv) R32-~32-}?31 R32;
(v) ~31 ~32 ~32 R31 R32; or t ) 33 ~31 R32 R32 R31 R32~ wherein R31, R32, and R33 are as hereinabove d~scribed.
In accordance with another embodiment, when ~he peptide includes the stru~ture X33, the peptide may include the following structu~:
X33 - Z33 wherein X33 is as hereinabove described, ~nd Z33 is: .
(i) ~32;
( i) R32 R33;
(iii) R32-R33 ~31;
(iv) ~32-R33-R31 R32;
(v) R32-~33-~31-R32-R32; or ~ ~ ~ r ~ ~ ~ PCT/US92/08823 -~3-(vi) R32-R33-R3l-R32 R32 31 In accordance with yet another embodiment, the peptide may include the following structure: `
(Y33)a X33 (Z33)b~ wherein Y33 and Z33 are as previously defined, a is 0 or l, and b is 0 or 1.
In accordance with yet another embodiment, when the peptide includes the structure X34, the peptide may include the following structure: , Y34 - X34, wherein X34 is as hereinabove described, and Y34 i s :
(i) R32;
(ii) R32 ~32;
(iii) R3l-~32 R32;
(iv) R3 Y2-R3l-R32 R32;
( ) R32 R32 R31 ~32 ~32; or -~
( ) R31 R32 ~32 ~31 R32-~32~ wherein R31, R32 and R33 are ~;
as hereinabove described.
In accordance with another embodiment, when the peptide -~
includes the structure X34, the peptide~may include the followin~
structure:
- X34-Z34, wherein X34 is as hereinabove described, and Z3 is:
(i) R33;
(ii) ~33-~31;
(iii) R33-R3l ~32;
(iv) R33-R3l-R32 R32;
( ) R33 ~31 R32 ~32-R31; or (Vi ) R33-~.31; R32-R32R31-R32 '' ' In accordance with yet another embodiment, the peptide may include the followiny structure:
(Y34)a X34 tZ34)b~ wherein X34 and Z34 are as previously defined, a is 0 or l, and b is 0 or l.

2~2~337 In accordance with a further embodiment, when the peptide includes the structure X35, the peptide may include the following structure:
Y35-X3~, wherein X35 is as hereinabove described, and Y
is:
(i) R33;
(ii) R32-~33;
(iii) R32~R32~R33;
(i~) R3l-R32 R32 R33;
(v) R32-R3l-R3~ R32 33;
( ) R32 R32 ~31 R32 R32-R33~ wherein R31, ~32~ and R33 are as hereinabove described.
In accordance with another embodiment, when the peptide includes the structure X35, the peptide may include the following structure:
X35 - Z35 wherein X35 is as hereinabove described, and Z
is:
(i ) R31;
(ii) R31'R32;
(iii) R31-~32 R32;
(iV) R31-~32-R32 R31;
(v~ 31 R32 R32~R31~R32; or (vi) R3l-~32-~32-R31 R32 32 In accordance with yet another embodîment, the peptide may include the followin~ structure:
~ Y35)a- X35 ~Z35)~, wherein X35 and Z35 are as previously defined, a is 0 or 1, ~nd b is 0 or l.
In accord~nce with a further embodiment, when the peptide includes the structure X36, the peptide may include the following structure:
Y3~ ~ X36 wherein X36 is as hereinabove described, and Y36 is:
(i) ~31;
(ii) R33-R31;

W093~07892 - 2 1 2 0 3 3 7 PCT/US92/08823 (iii) R32~R33 R31;
(iv) R32~~32~R33 R31;
(v) R31 R32-R32-R33-R31; or ( ) R32 R31 R32 ~32-R33-R31~ wherein R31, R32, and R33 are as hereinabove described.
In accordance with another ~m~odiment, when the pe~tide :~
includes the structure X36, the peptide may include the following structure:
X36-Z36, wherein X36 is as hereina~ove described, and Z
i s :
(i) R32; ~' (ii) R32-~32;
(iii ) R32-~32 R31;
(iv) R32-R32 R31 32 (v) R~2-R32-R3l-R32 R32;
(vi) R32-R32-R31 ~32 32 33 In accordance with yet another embodiment, the peptide may include the following structure:
(Y ) - X (Z 6)~ wherein Y36 and Z36 are as previ~uslY
defined, a is O or 1, and b is 0 or 1.
I~ accordance with one embodiment, when the peptide includes -~
the structure X37, the peptide may includes the structure . ~:
Y37-X37, wherein X37 is as hereinabove de3cribed, and Y37 is:
~i) R3~, (ii) R31-R32;
(iii) R33-~31 R32;
(iv) R32-~33-R31 R32;
(v) R -~ -~33-~31-R32; or ( ) 31 ~32 R32 R33 R31 R32, wherein R31, R32, and R33 are as hereinabove described.
In accordance with a further embodiment, when the peptide includes the structure X37, the peptide may include the follo,wing structure:

21203~7 `

X37 - Z37 wherein X37 is as hereinabove described, and Z37 -i s :
( ) 32;
(ii) R32-R31;
(iii~ R32-R31 R32;
(iv) R32-R31-R32 R32;
(v) R32 ~31 R32 32 33;
(vi) R32-R3l-R32 ~32 33 31 In accordance with yet another embodiment, the peptide may include the following structure:
(Y37)a X37 (Z37)b' wherein Y37 and Z37 are a~ previously defined, a is O or 1, and b is O or 1.
In a preferred embodiment, n is 3, and most preferably the peptide is of one of the following structures as given in the accompanying sequen~e listing:
(Lys Ile Ala Gly Lys Ile Ala)3 ~SEQ ID NO:27).
(Lys Ile Ala Lys Ile Ala Gly)3 (SEQ ID NO:28).
~Lys Ile Ala Gly Lys Ile Gly)3 (SEQ ID NO:29).
(~ys Leu Ala Gly Lys Leu Ala)3 (SEQ ID NO:30).
(Lys Phe Ala Gly Lys Phe Ala)3 (SEQ ID NO:31).
(Lys Ala Leu Ser Lys Ala ~eu~3 (SEQ ID NO:32). :~
(Lys Leu Leu Lyæ Ala Leu Gly)3 (SEQ ID NO:33). -~
(Lys Ala Ile Gly Ly~ Ala Ile)3 (5EQ ID NO:34~.
~Gly Ile Ala Lys Ile Ala Lys)3 (SEQ ID NO:35). -(Lys Ile Ala ~ys Ile Phe Gly)3 (SEQ ID NO:36).
(Gly Ile Ala Ar~ Ile Ala Lys)3 ~SEQ ID NO:373.
(Lys Phe Ala Arg Ile Ala Gly)3 (SEQ ID NO:38).
(Gly Phe Ala Lys Ile Ala Lys)3 (SEQ ID NO:39).
(Lys Ile Ala Gly Orn Ile Ala)3 (SEQ ID NO:40).
(Lys Ile Ala Arg Ile Ala Gly)3 (SEQ ID NO:41).
(Orn Ile Ala Gly Lys Ile Ala)3 (SEQ ID NO:42).
(G~y Ile Ala Arg Ile Phe Lys)3 (SEQ ID NO:43).
(Lys Nle Ala Gly Lys Nle Ala)3 (SEQ ID NO:44).
(Lys Nle Ala Gly Lys Ile Ala)3 (SEQ ID NO:4~

W093/07892 2 1 ~ ~ 3 3 7 PCT/US92/08823 tLYs Ile Ala Gly Lys Nle Ala)3 (SEQ ID NO:46).
(Lys Nva Ala Gly Lys Nva Ala)3 (SEQ ID NO:47).
(Lys Nva Ala Gly Lys Ile Ala)3 (SEQ ID NO:48).
(Lys Leu Leu Ser Lys Leu Gly)3 (SEQ ID NO:49~.
(Lys Leu Leu Ser Lys Phe Gly)3 (S~Q ID NO:50).
(Lys Ile Ala Gly Lys Nva Ala)3 (SEQ ID NO:51).
(His Ile Ala Gly His Ile Ala)3 (SEQ ID NO:52).
(Ala Gly Lys Ile Ala Lys Ile)3 (SEQ ID NO:53).
(Ile Ala Lys Ile Ala Gly Lys)3 ~SEQ ID NO:54). -~
(Lys Ile Ala Gly Arg Il* Ala)3 ~SEQ ID NO:55).
(Arg Ile Ala Gly Arg Ile Ala)3 (SEQ ID NO:56).~:
(Lys Val Ala Gly Lys Ile Ala)3 (SEQ ID NO:57). -~
~Lys Ile Ala Gly Lys Val Al~)3 (SEQ ID NO:58).
(Ala Lys Ile Ala Gly Lys Ile)3 (SEQ ID NO:59).
(Orn Ile Ala Gly Orn Ile Ala)3 (SEQ ID NO:60).
(Lys Phe Ala Gly Lys Ile Ala)3 (SEQ ID NO:61).
(Lys Ile Ala Gly Lys Phe Ala)3 (SEQ ID NO:62).
(Lys Ch~ Ala Gly Lys Ile Ala)3 (SEQ ID NO:63).
(Lys Nle Ala Lys Ile ,~la Gly)3 (SEQ ID NO:643. :
(Arg Ile Ala Gly Lys Ile Ala)3 (5EQ ID NO:65~. :
(Har Ile Ala Gly Har Ile Ala)3 (SEQ 'D NO:663.
(X~a Ile Ala Gly Lys Ile Ala)3 (SEQ ID NO:67~.
(Lys Ile Ala Gly Xaa Ile Ala)3 (SEQ ID NO:68~.
In (SEQ ID NO:67) and ~SEQ ID NO:68~, Xaa i~
p-aminophenylalanine.
In accordance with another embodiment, the biologically active amphiphilic peptide in~ludes the following basic structure R31-R32-R32-R33-R34-R32-R32-~?31-R32_R32_~.3~ R34-~.32_~,32, wherein R31 ~ R32, and R33 are a~ hereinabove described, a R34 is a basic hydrophilic or hydrophobic amino acid.
In accordance with one em~odiment, the peptide may includ the following structure:

W~g~/~7~9~3 7 PCT/US92/08823 Y40-X40, wherein X40 is as herei~abo~e described, and Y40 i s :
(i) R32;
(ii) R32-R32;
(iii) R34-~32-R32;
(iv) R33-R34-R32-~32;
(v) 32 33 R34 ~32 R32;
(v) R32 R32 R33 R34 R32-R32, or R ( ) R31 R32 R32 ~33-R34 ~32~R32,wherein R31, ~32~ R33 and 34 are as hereinabove de~cribed.
In accordance with another embodiment, the peptide may include the following structure:
X40-Z40, wherein X40 is as hereinabove described and Z40 is:
(i ) R31;
(ii) R31 ~32;
(iii ) R31-R32 R32;
(iv) R31-R3~ R32 33 (v) R31-~32 ~32 33 34 (vi) R31 ~32 ~32 ~33 R34 R32; or `~
( ) ~31 R32 ~32 ~33~~34~R32~~32, wherein R31, R32, ~33, and R34 are as hereinabove described.
In accordance with yet another embodim~nt the peptide may~ -:
include the following structure:
(Y40)a-x4o-(z4o)b~ wherein Y40 and ~40 are as previously defined, a is O or 1, and b is O or 1. In a preferred embodiment, ~he peptide has the following ~tructural formula 8S ~ :-given in the accompanying sequence listing:
~ SEQ ID NO:69) In another preferred em~odiment, the ~eptide has the following structural formula as given in the accompanying sPquence li sting:
(SEQ ID NO:70) , W093/07892 2 1 2 o ~ 3 7 PcT/us92/o8823 -29- ~:

In accordance with a further embodiment, the peptide has one ~.
of the one of the following structural formulae as given in the accompanying sequence listing:
(SEQ I~ N0:71) ~
(SEQ ID N0:72) .
(SEQ ID N0:73) ~:
(SEQ ID N0:74) (SEQ I~ N0:7S) ~
~SEQ ID N0:76) -~:
~SEQ ID N0:77) ~SEQ ID N0:78) .
(SEQ ID N0:79) (SEQ ID N0:80) (SEQ ID NO:B1) ~SEQ ,ID N0:82) ~SEQ ID NO:B3) --~SEQ ID NO:B4) ~SEQ ID N0:85 In acc~rdance with another embodiment, the peptide may ~`
include the following stru~tural ~ormula: .~:
- ~Lys Ile Ala Lys Lys Ile Ala)-n, wherein n is from 2 to 5. .' Preferably, n is 3, and the peptide has the following structu~al ~:`
formula:
~Lys Ile Ala Lys Lys Ile Ala) (SEQ ID NO:B6 In accordanee with another embodiment, the peptide may be selected from the group consisting of the followinq structural formulae as given in the accompanying sequence listing~
~SEQ ID N0:87) ~ E~
(SEQ ID N0:88) ~;
~SEQ ID N0:89) (SEQ IP N0:90) In accordance with another em~odiment, the peptide may includes the ollowing basic structure XS0: `

W093/07892 PCT/US92/0~823 R41 is a hydrophobic amino acid, and R42 is a basi.c hydrophilic or neutral hydrophilic ami~o acid.
In one embodiment, the peptide includes the basic structure Y50-X50 wherein X50 is as hereinabove described and Y50 is: -~ i) R41;
(ii) R42-R41; or (iii) R42-R42-R41, wherein R41 and R42 are as hereinabo~e described.
In one embodiment, R41 is leucine. In another embodiment, R42 is lysine. Representative examples of peptides in acordance with this aspect of the present invention include those having the following structures:
(SEQ ID N0: 91) (SEQ ID N0: g2) (SE~ ID N0: 93) ~SEQ ID N0: 94) In accordance with another em~odiment, the includes the ~ollowing basic structure X~2:
42 41 R42 R42 R41 R41 ~42-R42-R41-Rg2-R42, wherein R~l is a hydropho~ic amino acid, and R2 is a basic hydrophilic or neutral hydrophilic amino acid.
In o~e embodiment R41 is leucine. In another embodiment, R42 is lysine-In one embodiment, the peptide includes the basic structure Y52-X52, wherein X52 is as hereinabove described, and YS2 is: ~-(i) R42; ~;, (ii) R41 42;
(iii) R41 ~41 R42;
(iv~ R42-~41-~41 R42;
(v3 R42 ~42 ~41 R41 R42-In one embodiment, the peptide may have the following structure:

WO 93/07892 2 l 2 0 3 3 7 PCT/US92/08823 Lys Lys Leu Leu Lys Lys Leu Lys Lys Leu Leu Lys Lys Leu Arg Arg (SEQ ID NO.:9~) In another embodiment, the peptide includes the basic structure X~2 ~ Z~2' wherein X~2 is as hereinabove described, and :
Z~2 is:
(i) R41; ..
(ii) R41 R41;
liii) 41 R41 R42; :
(iv) R4l-R41-R42 ~42' (v) R41 R41 R42-~4~ 41;
In one embodiment, the peptide may have the following structure: ~.
Lys Leu Lys Lys Leu Leu Lys Lys Leu Lys Lys Leu Leu Lys Lys Leu ~ 10 15 :.
(SEQ ID NO: 96) In another embodiment, the peptide may include the structure: ~ `
52 a ~2 ~ ~23b~ wherein X52, Ys2 and Z52 are as hereinabove described, and a i5 0 or 1, and b is 0 or 1.
In one embodiment, the hereinabove described peptides may be ~:
acetylated with a CH3~O-group at the N-terminal.
In accordance with another embodiment, each of the amino acid residues may be a D-amino acid residue or glycine. ;~.
Although the scope of this particular embodimen~ is not to be limited to any theoretical reasoning, it is b~lieved that the peptides and proteins of the present invention, when consisting entirely of D-amino acid or glycine residue~, have in~reased resistance to proteolytic enzymes while retaining their biological activity. Such peptides may thus be administered WO 93/07892 P~T/USg2/08~3 2~203~7 orally. Also, in accordance with another embodiment, all of the amino acid residues may be D-amino acid or glycine residues, or L-amino acid or glycine residues.
In still another embodiment, the peptide employed in conjunction with an anti~iotic such as those hereinabo~e described, or derivatives or analogues thereof i8 a cecropin.
The term cecropins includes the basic structure as well as analogues and derivatives thereof. The cecropins and analogues and deri~atives thereof are described in Ann. Rev. Microbiol 1987, Vol. 41 pages 103-26, in particular p. 108 ~nd Christensen at al PNAS Vol. 8S p. 5072-76, which are hereby incorporated by reference.
The term cecropin includes the basic structure as well as analogues and derivati~es.
In yet another embodiment, the peptide employed in conjunction with an anti~iotic such as those hereinabove described, or derivatives or analogues thereof is a sarcotoxin.
The sarcotoxins and analogues and derivatives thereof are ~`
described in Molecular Entomoloav, pages 369-78, in particular p.
375 Alan R. Liss Inc. (1987), which is hereby incorporated by r~ference.
The term sarcotoxin includes the basic materials as well as analogues and derivati~es.
Ion channel-forming proteins or peptides which may be employed include defensins, also known a~ human neutrophil antimicrobial peptides (~NP), major basic protein (MBP) of eosinophils, bactericidal permeability-increasing protein (BPI), and a pore-forming cytotoxin called variously perforin, cytolysin, or pore-forming protein. Defensins are described in Selsted, et al., J. Clin. Invest., Vol. 76, pgs. 1436-1439 (198~). MBP proteins are described in Wa~moen, et al., J. Biol .
Chem., Vol. 263, pgs. 12559-12563 (1988). BPI proteins are described in Ooi , et al., J. Biol. Chem., Vol. 262, pgs.
14891-14894 (1981). Perforin is described in HenXart, et al. J.

W0 93/0~892 2 1 2 0 3 3 7 PCT/US92/08823 EXP. Med., 160:75 (1984), and in Podack, et al. J. EXP. Med., 160:695 (1984). The above articles are hereby incorporated by reference.
The term ion channel-forming proteins includes the basic structures of the ion channel-forming proteins as well as analogues or derivatives.
Pseudomonic acids are produced by Pseudomonas fluoressens.
A preferred pseudomonic acid which may be employed i5 mupirocin, or pseudomonic acid A, which has the following sturcture:

~'j ~h C; ~c_o~

JYO

The cephalosporins include a 7-aminocephalosporanic acid nucleus. A preferred cephalosporin is cefotaxime, which has the :
following structure:

~O-~c~ _ O _ ~/_ C,~

C00f~a Etham~utol, isoniazid, and ethionamide are especially useful in treating mycobacterial infections, and in particular infections of M. tuberculosis. Ethambutol is a synthetic, WOg3/0~892 . PCT/US92/08823 2120 ~7 - 3~ -water soluble, heat stable compound and is the D-isomer of the structure below: .~
CH20H C2H5 . :, H--C--NH--( CH2)2--Nt~ CH

Ethambutol may be dispensed as the dihydrocholoride salt. Ehtionamide, a close chemical relative of isoniazid, H
has the following structure: . ;
H5C ~ ~

.'.

S" `NH2 -~ ' Sulfonamides have the following nucleus:

to which various R radical in the amido`group ~ S02NHR) have been attached or in which various substitutions of the amino group ~NH23 are made. Representative exa~ples of the sulfonamides and their structure are given hereinbelow:
52 NH~
~J ;

SUBSTITUTE SHEET

W093/078s2 2 1 2 0 3 3 7 PCT/US92/~8823 - 3s - ~-sulfadiazine :
S 2 N H\~

sulfisoxazole S2 NH~

[~3 ~CH3 sulfamethoxazole SO2NH~

COOH

NHCO~
sulfathalidine Penem antibiotics which may be employed include~
but are not limited to, imipenem, carbapenems such as RS-533 (Sankyo), 2-(N-azolyl~ ~lkyl-substitu~ted penems, sucb as CGP-29~718 (Ciba-Geigy), and broad-spectrum penems such as Sch-34343 (Schering Plough). Imipenem, also known as a thienamycin antibiotic, has the following structure:
NH

COOH

SVB~T~ i ~ J ~ FT

W093/07892 PCT/U~92/08823 212~337 36 -Preferred aminoglycoside antibiotics which may be employed are tobramhcin and the gentamicins. Tobramycin has the following structure:
H~C--NH2 N~ NHz ~0~ ;.

H H~

Tbe gentamicins (Gentamicin C~, Gentamicin C2, and Gentamicin C1~), as well as netilmicin, have the following basic structure: R1 :
C~NHR2 NH2 NHR3 . ~:
~ '' N~2HO ~ OH ~
~ H3 :

For Gentamicin Cl, R1 and R2 are each CH3, the Cb-C5 bond is a single bond, and R3 is H. For Gentamcin C2, R1 is CH3, R2 and R3 are each H, and the C~-C5 bond is a single bo~d. For Gentamicin C1a, R1, R2 and R3 ea h are H, and the C~-C5 bon~ ic a single bond. For netilmicin, R1 and R2 each are H, R3 is C2~5, and the Cb-C5 bond is a double bond.
Other aminoglycosides which may al~o be employed within the scope of the present invention include, but ~re not limited to, kanamycin and amikacin, as well as metilmicin, neomycin, and streptomycin, which is alo effective in treatihg tuberculosis. Kanamycin ~nd amikacin both have the following basic structure:
H2C--N11,! N~NII-R
HO~ ~( ., ~LI CH20 H1:1 0U HO ~

S ~J 3S ~1 , . . _ ~ ~

W093~07892 2 1 2 ~ 3 3 ~ PCT/US92/08823 For kanamycin, R is H, and for amikacin, R i5:
o OH
Il /

Erythromycin, which is isolated from Streptomyces erythreus, is a member of a group of compounds known as macrolides. The basic structure of a group of compounds ~;
known as macrolides. The basic structure is a large lactone ring to which unusual sugars are attached. The term "macrolide" refers to a large ring formed from a chain of 14 to 20 carbon atoms by lactone condensation of a carboxyl and hydroxyl group. Other macrolides include oleandomycin, spiramycin, kitasamycin, and carbonmycin.
Erythromycin has the the following structure:

~ 3 0/~ ~
CH3 CIIJCII3~l 1 1 \/ o/\~\
/ ¦ N(Cl13)2 1~1 (I I oll H3 ~ ~ - Cll 3 Cll 2 0 0~ O y Cll 3 Cli3 Cl130 ~

~ IP..C~T'T~ ~ ~T`~

W093/07892 PCT/US92/0~23 212~337 _ 38 It is also to be understood that other macrolide antibiotics, such as roxythromycin, clarithromycin, and others hereinabove described, may be employed as well.

For purposes of illustration of exanples of other macrolide structures, the following examples, in addition to :
the above structure of erythromycin, are given below.

Rokitamycin is of the following structure:
CH3 C}i~CH2 ~CH~
HO~ O ~ H ¦ CH3 -^OH ~ 1 OCCH~CHJ
Cll~ ~ H\ N~C~/¦ ~ OdCN.CH,CN.
O D ~ H -CGP-7040, a benzapiperazinyl rifamycin, has the following structure:

CH 3 ~H 3 Cll 3 CH3CDO ~ ~

I CH3 ~ -J OH OH ~ ~ CH 3 ~CH3 1 ¦ I

¦ ~ ~ N ~ N ~ H

CH3 ~;
SUBSTITUTE SHEE~T ~

., .". .,. ~ ." .~:

W093/07892 2 1 2 0 3 3 7 PCT/US92/0~23 It is to be understood, however, that the scope of the present invention is not to be limited to the specific macrolides or macrolide structures hereinabove described.
Preferred penicillins which may be employed in accordance with the present invention are benzyl penicillin (penicillin G) and ampicillin (alpha-amino-benzyl penicillin). Benzyl penicillin is of the following structure:

~Cll 2 - C- NllF~ C~l 3 COOli Ampicillin has the following structure:
O Cll~
C}l-C-N1l ~ ~ ~ Cll~

COO}l A preferred monobactam which may be employed in accordance with the present invention is aztreonam, which has the following structure:

O
` 11 (;113 ~1 1 G--~--Nl~
~N N ~N
z I O ~S~
O - C ( C ~ 3 ) 2 C~QII

IR.C~T~T~

W093/07892 . PCTrUS92/0~23 2 12 ~ 3 3 l - 39/1-The bacitracins which may be employed in accordance with the present invention tend to be hydrophilic and water ~:
solubleO The preferred bacitracin is bacitracin A, which has the following structure:
,~S ~ G,H --CIICII z Cll 3 ~¦ Nl12 Cll~
N
.1 ' ':
C= O .
L-l{is- D-Asp-L-Asn L-Leu D-phe~ D-Glu L-Ile -D-Orn- L- a -Lys-L-lle Peptide antibiotics, which are not ion channel-forming peptides or proteins, which may be employed include, but are not limited to, gramacidin-S, polymyxin, vancomycin, :
teichoplanin, and capreomycin.

~J~ ~T'.~JT~ ~H~FT

W093/07892 2 1 2 0 3 ~ 7 PCT/US92/08823 -4~-Vancomycin is a tricyclic glycopeptide antibiotic. Its chemical formula is C66H15C12H9024, and it hafi a molecular weight of 1,44~.

It is to be understood, however, that the ~cope of the invention is not to be limited to the specific antibiotics and antibiotic structures hereinabove described.
The present invention will be further described with respect to the following examples, however, the ~cope of the invention is not to be limited thereby.
ExamDle 1 `:
S. aure~s organisms are grown to mid log phase, and then diluted to 10 organisms/ml in 1/~ ~trength trypticase soy broth.
After the incubation of the organisms, the minimal inhibitory ~`-concentraion in ug~ml for Z-52 MG-2(amide-terminated), PGLa, CPF -~
Z-50, A-97, Magainin II, and Z-74 peptides, agai`nst S.aureus was measured when each peptide was added alone to the organisms.
"MG-2 (amide)" is amide-terminated Magainin II, and 1'Magainin~
is car~oxy-terminated Magainin II. A-C7 peptide is of the following structureo (SEQ ID N0:97) amide Z-74 peptide is of the following str~lcture:
~ 5EQ ID N0:98)-amide The PGLa peptide is of the followinq structure:
~ SEQ ID N0:12)-amide.
The minimal inhibitory concentration (MIC) for each peptide was then measured when 20% of the minimal inhi~itory concentration of bacitracin, to~ramycin or gentamicin, respecitvely, is added to the organisms along with the peptide.
The gentamicin employed is a mixture of Gentamicin Cl, Gentamicin Cla, and Gentamicin C2. For Examples 1-3, the MlC values for bacitracin are 2 ~g/ml against S. aureus, 64 ~g/ml against E. :
coli, and >2~6 ~g/ml against 8~e9~e~a~YD~o~kqh~8~- The MIC
values for gentamicin are 256 ~g/ml again~t S. aureus, 2 ~g/ml against E. coli, and >256 ~g/ml against P. aeru~inosa. The MIC
values for tobramycin are >256 ~g/ml against S. aureus, 2 ~g/ml against E. coli, and 128 ~g/ml again~t P. aeru~inosa. CP~ Z-50 peptide is (5EQ ID N0:21) of the CPF peptides hereinabove described. Z-52 peptide is of the structure (SEQ ID NO:~9)-NH2.
The minimal inhibitory concentrations are given below in Table I.
TABLE I
MIC With Peptide _dded Antibiotic Pe~tide Alone _Bacitracin~ Gentamicin Tobram~cin MG-2 amide256 32 ~ 32 PGLa 32 16 2 32 ~-Maqainin 2256 128 4 32 Z-50 16 16 2 8 -~
Z-74 1~ 16 4 8 -E.coli organisms were incu~ated according to the prcedure described in Example 1. After the incubation, the minimal inhibitory concentrations against E.coli of each of the peptides described in Example 1 alone, as well as of each peptide when employed in ~ombination with 20% of the MIC for bactracin, were then measured. The results are given in Table 2 below.

wo g3/07892 2 1 2 ~ 3 3 7 Pcr/US92~08823 Ta~le 2 PePtide MIC
PePtide Pe~t_de with Alone _ Bacitracin Z-52 ~ ~
MG-2 amide 16 4 PGLa 16 8 Magainin II32 8 (amide) Z-7~ 16 8 ExamPle 3 In this example, P.aeru~inosa organisms were incubated according to the procedure described in Example 1. After the incubation, the minimal inhibitory ~oncentration~ of the peptides -~.
hereinabQve described alone, as well a~ the peptides in combination with 20~ of the MIC of bacitracln, were then measured. The results are given below in Table Table 3 PePtide MIC .
Peptide Pe~tide with Alone Bacitracin Added Z-52 ~6 MG-2 amide128 8 PGLa 64 8 Z-~0 32 4 Magainin ~I256 1 EXAMPLE 4 :~

In this example, E. coli or P. aeruqinosa organisms were incubated according to the procedure described in Example 1.
After, the incubation, the minimal inhibitory concentrations of the peptides hereinabove described alone, as well a5 the peptides in combination with 20% of the minimal inhibitory concentrations (MIC) of benzyl penicillin or ampicillin, were then measured.
The MIC of benzyl peni illin against E. coli i~ 64 ~g/ml, and of ampicillin against E. coli is 4 ~g/ml. The results are given below in Table 4.

E. coli-MIC ..
Peptide Peptide and Peptide and :~
PePtide alone Ben~yl ~enicill_nAm~icillin ~.

Mg 2-amide 64 32 8 PGT.a 32 16 Magainin A-97 32 16 . 16 P. aeru~inosa-Pe~tide PGLa 128 32 32 Mg~2 amide 256 128 128 W O 93/07892 . 2 1 2 0 3 3 7 PC~r/US92/08823 E~U~MPLE S
E. coli and P. aeruq nosa organisms were incubated according to the procedure described in Example 1. After the incubation, the minimal inhibitory concentrations of the peptides hereina~ove described alone, as well as the peptides in combination with 20%
of the MIC of the monobactam antibiotic aztreonam, were then measured. The MIC of aztreonam against E. col$ is a 2 ~g/ml, and ~
against P. aeru~inosa i~ 8 ~g/ml. The results are given below in ~.:
Table 5.

:.:
':
- .

WO 93/0~8g2 PCT/US92/08823 2l2n337 :~

TABLE S
E. coli-MIC
Peptide Peptide with :~
PePtide alone Aztreonam Mg 2-amide 64 32 PCLa 32 4 Magainin Z-?4 4 8 P. aeru~inosa-Pe~tide Z-~2 32 32 Mg-2 amide 128 64 PGLa 128 64 Z-~0 64 3X

Magainin The above r~sùlts indicate that when one of the biologically active ion-channel forming peptides hereinabove described is added in combination with bacitracin, tobramycin, gen~amicin,, benzyl penicill~n, ampicillin, or aztreonam, in an amount of 20%
of the MIC of these antibiotics, against S.aureus, E.coli, or WO 93/07X92 2 1 2 ~ 3 3 7 PCT/US92/08823 4~

P.aeruqinosa, there is, in most cases, a resulting synergy between the peptide and the antibiotic. In most cases, less peptide and less antibiotic may be used against these organisms when peptide and antibiotic are employed in combination, than if peptide or antibiotic alone were employed.
Exam~le 6 In this example, the effect of a combination of erythromycin and biologically active amphiphilic ion channel-forming peptide will be measured against K.Pneumoniae, P.aeruqinosa, E.coli, and 5.aureus.
For each of the organisms listed in Table 6 below, 105 organisms were mid-log inoculated into 200 ~1 of one-half strength trypticase soy broth. The organisms were incubated for 15 hrs. at 37C. After the incubation of the organisms, the minimal inhi~itory concentration in ~g/ml for Magainin II
(amide-terminated), shown as MGN2, CPF Z-50 (amide-terminated) peptide, Z-52 (amide-terminated) peptide, against each species of organism was measured wherein 10 ~g~ml of erythromycin was added and wherein no erythromycin was added. The minimal inhibitory concentration of erythromycin alone against each species of organism was also measured. The minimal inhibitory concentrations in ~g/ml are given below in Table 6. For purpQses of explanation, the "~" and "-" signs below the "Erythromycin (10 ~g/ml)" column indicate the presence or a~sence, respectively, of erythromycin ~dministered in combination with one of the `-biologically active peptides. K.~neumQniae, P.aeruqinosa, and E.coli are Gram-negative bacteria.

W093/07892 PCT/US92fO8B23 ~33~ `

Table 6 Minimal Inhibitory Concentration (~g/ml) Erthro- MGN2 CPF Z-52 Erytho mycin Organism (10~/ml) amide z-50 amide mycin amide K.Pneumoniae - 5 ~0.5 1.5 >100 + <0.5 <o.~~0.5 P.ae~uquinosa - 50 100 ~500 5-15 ~100 + <5.0 <0.5~1.5 E.coli - 5 <5.0 l.S >100 + 0.5 <0.5<0.5 The above results show that when one of the ~-biologically active ion channel-forming peptides shown in Table 6 is added to 10 ~g/ml of erythromycin, such a combination of peptide and erythromycin is effective against the Gram-negative organisms shown, whereas grea~er than 100 ~g/ml of erythromycin alone is required for effective biological a~tivity against these Gram-negative organisms.
The addi*ion of erythromycin to the biologically active peptides also enables one to use less of the biologically active peptide against Gram-negatiYe organis~. Thus, there is provided a synergistic effect against Gram-negatiYe organisms when erythromycin and a biologically active amphiphilic ion channel-fo~ming peptide are administered to inhibit growth of Gram-negative organisms.
It was also found that a concentration from a~out 0.6 ~g/ml to about 1.25 ~g~ml of erythromycin alone was required for effectiYe biological activity agai.nst S._aureus, a Gram-positive organism, and that greater than 250 ~g/ml of amide-terminated organism, and that greater than 250 ~g/ml of amide-te~minated Magainin II alone was required for effective biological activity against S.aureus. A combination of lO
~g~ml of amide-terminated SUBS~TU ~ r ~HFFT

WO 93/0789~ 2 1 2 0 3 3 7 PCT/US92/08823 M~gainin II and 0.03 ug/ml of erythromycin, however, al80 showed effective biological activity against S.aureus. Thus, it has also been shown that an enhanced effect against Gram-positive organisms is also obtained when erythromycin and a biologically active amphiphilic ion channel-forming peptide are administered to inhi~it growth of Gram-positive organisms. The addition of erythromycin to the bioloyically active peptide enables one to use less of the biologically active peptide against a Gram-positive organism.

ExamPle 7 -:~
The procedure for t~is assay, which is a checkerboard assay, is performed as described in Antibiotics and Laboratorv Medicine, 2 nd ed., Victor Lorian, M.D., Editor, pgs. 540-546 (19~6).
The checkerboard assay is carried out in a microtiter plate having wells arranged in rows and columns, 100 ~1 of plain broth is added to every row of wells. 100 ~1 of peptide (SEQ ID
N0:100) at 512 ~g/ml is added to the top row of wells, and serially dil~ted (1:2) through the columns. 50~1 o vancomycin in 4X concentrations is added to each appropriate column of wells to give a range of vancomycin concentrations from 16 to 1,024 ~g/ml. 50~1 of P. aeru~lnosa strain ATCC 27853 is thenr added to each well. The plate is incubated at 35-37C for 18 to 24 hours, and the wells are then examined for the presence of visible growth, i.e., turbidity.
Peptide (SEQ ID N0:100~-N~2 was tested alone or in combination with vancomyein for activity against _ aeruainosa --strain ATCC 27853.
In this assay, the MIC of (SEQ ID N0:100)-NH2 wa~ 32 ~g/ml, and the MIC of vancomycin was greater than 1,024 ~g/ml; i.e., vancomycin at a concentration of 1,024 ~g/ml did not inhibit growth of P. aeru~inosa.

WO g3J07892 PCT/US92/08823 2~2D337 -4g-The following combinations of (SEQ tD N0:100)-N~2 and vancomycin were found to be inhibitory.
~ ~q/ml ~SEQ ID NO:100)-NH2 plus 16 ~g/ml ~ancomycin 8 ~g/ml (SEQ ID NO:100)-NH2 plus 32 ~g/ml vancomycin 8 ~g/ml (SEQ ID N0:100)-NH2 plus 64 ~g/ml vancomycin 8 ~g/ml (SEQ ID N0:100)-NH2 plus 128 ~g/ml vancomycin 8 ~g/ml (SEQ ID NO:100)-NH2 plus 256 ~g/ml vancomycin 8 ~g/ml (SEQ ID NO:100)-NH2 plus S12 ~g/ml vancomycin 8 ~g/ml (SEQ ID N0:100)-NH2 plus 1,024 ~g/ml vancomycin The above results thus indicate that an enhanced effect against P. aeruainosa is obtained when vancomycin and a biologically active ion channel-forming peptide are administered to inhibit growth of P. aeruqinosa.
The peptide or protein and antibiotic such as those hereinabove des~ribed, may be enployed for treating a wide variety of hosts. In accordance with a preferred embodiment, a host is an animal, and such animal may be a human or non-human animal~ It is also possible to administer the peptide or protein and antibiotic in separate forms. For example, the antibiotic may be administered systemically and the peptide or protein may be administered topically.
The peptide or pr~tein and/or antibiotic such as those hereina~ove described, may be employed in a wide variety of pharmaceutical compositions in combination with a non-toxic pharmaceutical carrier or vehicle such a~ a filler, nun-toxic buffer, or physiological saline solution. Such pharmaceutical compositions may be used topically or systemically and may be in any suitable form such as a liquid, solid, semi-solid, injectable soiution, tablet, ointment, lotion, paste, capsule, or the like.
The peptide or protein and/or antibioti~ such as those hereinabove described may also be used in combination with adjuvants, protease lnhibitors, or compatible drugs where such a combination is seen to be desirable or advantàgeous in controlling infection caused by harmful microorganisms.

WO 93/07892 2 1 2 0 3 3 7 P~T/US92/08X23 ~

When the peptide or protein is administered topically, it is administered in combination with a water-soluble vehicle, said water-soluble vehicle being in the form of an ointment, cream, lotion, paste, or the like. Examples of water-soluble vehicles which may be employed include, but are not limited to, glycols, such as polyethylene glycol, hydroxycellulose, and KY Jelly. The water-soluble vehicle is preferably free of an oily substance.
The combination of peptide or protein and antibiotic of the present invention may be administered to a host; in particular an animal, in an effective antibiotic amount. When used to inhibit growth of bacterial cells, the combination, whether administered as a mixture or separately, is employed in an effective antibacterial amount. When used to inhibit growth of fungi, such components are administered in an effective antifungal amount.
As representative examples of administering the peptide or ~-protein and antibiotic for topical or local administration, the ~-peptide or protein could be administered in an amount of from about O.l,o to about 10% weight to weight; and the antibiotic is delivered in an amount of from about 0.1~ to about 10% weight to weight. -~;
Numerous modifications and variations of the present invention are possible in light of the above teachings and, therefore, within the scope of the appended claims, the invention may be practiced otherwise than as partiGularly d~cribed.

SEQUENCE Ll STING

(1) GENERAL INFORMATION:
(i) APPLICANT: Berkowitz, Barry A.
Zasl~ff, Michael (ii) TITLE OF INVENTION: Composition and Treatment wi~h Biologically Active Peptides and Antibiotic.

(iii) NUMBER OF SEQUENCES: 100 (iv) CORRESPONDENCE ADDRESS:

(A) ADDRESSEE: Carella, Byrne, Bain, Gilfillan, Cecchi ~ Stewart (B) STREET: 6 Becker Farm Road (C) CITY: Roseland ~D) STATE: New Jer~ey (E) COUNTRY: USA
(F) ZIP: 07068 ~v) COMPUTER READABLE FORM: ~
(A) M~DIUM TYPE: 3.5 inch diskette ~ -(B) COMPUTER: IBM PS/2 (C) OPERATING SYSTEM: PC-DOS
~D) SOFTWARE: DW4.V2 (~i) CURRENT APPLICATION DATA
(A) APPLICATION`NUMBER:
(B) FILING DATE:
(C) CLASSIFICATION:

(vii) PRIOR APPLICATION DATA:

-5~-(A~ APPLICATION NUMBER:

~B) FILING DATE: 05-SEP-1989 (A) APPLICATION NUMBER: ~:
US073392g2 "' ~8) FILING DATE: 17-APR-1989 (viii) ATTORNEY/AGENT INFORMATION:
(A) NAME: Ol~tein, Elliot M.
(B) REGISTRATION NUMBER: 24,025 (C) REFERENCE/DOCKET NUMB~: 421250 (ix) TELECOMMUNICATION INFORMATION:
(A) TELEPXONE: 201-994-1700 ~B) TELEFAX: 201-994-1744 (2) I~FORMATION FOR S Q ID NO:1: `~
(i) SEQUENCE CHARACTERISTICS .~;
(A) LENGTH: 20 amino acids ~
(B) TYPE: amino acid ,l:
(C) STRANDEDNESS~
(D) TOPOLOGY: linear (ii) MOLECULE TYPE: peptide (x) PUBLICATION INFO~MATION:
(H) DOCUMENT NUM8ER: W089/11290 -~
(I) FILING DATE: 19-MAY-1989 ~:
(J, PUBLICATION DRTE: 30-NOV-1989 ~:

WOg3t0~892 PCT/US92/~8823 2120337 "`

..

(xi) SEQUENCE DESCRIPTION: SEQ ID NO:1:
Ala Phe Ser Lys Ala Phe Ser Lys Ala Phe Ser Lys Ala Phe Ser Lys Ala Phe Ser Lys (2) INFORMATION FOR SEQ ID NO:2 (i) SEQUENCE C~ARACTERISTICS:
(A) LENGT~: 24 amino acids (B) m E: amino acid (C~ STRANDEDNESS:
(D) TOPOLOGY: linear (ii) MOLECULE TYPE: peptide (x) PUBLICATION INFORMATION: ;
(H) DOCUMENT NUMBER: W089/11290 (I) FILI~G DATE: l9-MAY-19~9 (J) PUBLICATION DATE: 30-NOV-1989 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:2:
Ala Phe Ser Lys Ala Phe Ser Lys Ala Phe Ser Lys Ala Phe Ser ~ys Ala Phe Ser Lys 1~ 20 Ala Phe Ser Lys (2) IN~ORMATION FOR SEQ ID NO:3:
(i) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 16 amino acids (B) m E: amino acid (C) STRANDEDNESS:
(D~ TOPOLOGY: linear WO 93~078922 1 2 0 3 ~ ~ PCI/US92/0~823 -54- -:

(ii) MOLECULE TYPE: peptide (x) PUBLICATION INFORMATION:
(H) DOCUMENT NUM8ER: W089/11290 (I) FILING DATE: l9-MAY-l9~
(J) PUBLICATION DATE: 30-NOV-1989 (xi) SEQUENCE DESCRIPTION: SEQ ~D NO:3:
Phe Ser Lys Ala Phe Ser Lys Ala Phe Ser :~
Lys Ala Phe Ser Lys Ala (2) INFORMATION FOR SEQ ID NO:4~
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH~ 20 amino acids (B) TYP : amino acid ::
(C) STRANDEDNESS:
~D) TOPOLOGY: linear (ii) MOLECULE TYPE: peptide (x) PUBLICATION TNFORMATION:
(H) DOCUMENT NUM8ER: W089/11290 (I) FILING DATE: 19~MAY-1989 (J) PUBLICATION DATE: ~O-NOV~1~89 ~xi) SEQUENCE DESCRIPTION: SEQ ID NO:4: ~-Ser Lys Ala Phe Ser Lys Ala Phe Ser Lys Ala Phe Ser Lys Ala Phe 5er Lys Ala Phe (2~ INFORMATION FOR S~:~ ID NO:5:

W093/~7892 PCT/US92/08~23 212~3~7 (i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 16 amino acids ( B ) TYPE: ami ~o acid (C) STRANDEDNESS:
(D) TOPOLOGY: linear (ii) MOLECULE TYPE: peptide (x) PUBLICATION INFORMATION:
(~) DOCUMENT NUMBER: W089/11290 (I) FILING DATE: l9-MAY-1989 (J) PUBLICATION DATE: 30-NOV-1989 (xi~ SEQUENCE DE~CRIPTION: SEQ ID NO:5:
~ Lys Ala Phe Ser Lys Al~ Phe Ser Lys Ala Phe Ser Lys Ala Phe Ser (2) INFORMATION FOR SEQ ID NO:6:
~i) SEQUENCE CHARACTERISTICS :.
(A) LENGTH: ~23 ~mino acids (8) TYPE: amino acid (C) STRANDEDNESS:
(D) TOPOLOGY: linear .
MOLECULE TYPE: peptide (A) NAME/KEY: Magainin I peptide.

~x~ PU~LICATION INFORMATION:
(A) AUTHOR: Zasloff, Michael (C) JOURNA~: Proceedin~s ~f ebe N~t_ooal Academ~
of Sciences (D) VOLUME: 84 ':~
:

W093~07~92 2 1 2 0 3 3 7 PCT/US92/08823 . -56-( F ) PAGES: 5449-5453 (G) DATE: AUG - 1987 ~H) DOCUMENT NUMBER: US 4810777 (I) FILING DATE: 04-MAR-19B7 (J) PUBLICATION DATE: 07-MAR-1989 '~
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:6 Gly Ile Gly Lys Phe Leu His Ser Ala Gly 5 10 ~
Lys Phe Gly Lys Ala Phe Val Gly Glu Ile :
Met Lys Ser (2) INFORMATION FOR SEQ ID NO:7: ;
(i) SEQUENCE CHARACTERISTICS
(A3 LENGTH: 23 amino acids (B) TYPE: amino acid ~C) ST~ANDE~N%SS:
(D~ TOPOLOGY: linear -, .
(ii) MOLECU~E TYPE: peptide (ix) FEAT~RE:~
(A) NAME/KEY: Magainin II peptide.

(x~ PUBLICATION INFO~MATION:
(A) AUTHOR: Zasloff, Michael (C) JOURNAL: Proceedinqs of the National AcademY
of Sciences (D) VOLUME: 84 (F) PAGES: 5449-5453 (G) DATE: AUG - 1987 ~H) DOCUMENT NUMB~R: US 4810777 (I) FI~ING DATE: 04-MA~-1987 ~J) PUBLICATION DhTE: 07-MAR-19B9 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:7 Gly Ile Gly Lys Phe Leu His Ser Ala Lys Lys Phe Gly Lys Ala Phe Val Gly Glu Ile Met Asn Ser (2) INFORMATION FOR SEQ ID NO:8: -:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 22 amino acids (B) TYPE: amino acid (C) STRANDEDNESS:
(D) TOPOLOGY: linear (ii) MOLECULE TYPE: peptide ~-(ix) FEATURE: -(A) NAME/KEY: Magainin III peptide.

(x) PUBLICATION INFORMATION:
(A) AUTHOR: Zasloff, Michael (C) JOURNAL: Proceedinqs of the National Aca~emv of Sciences (D) VOLUME: 84 (F) PAGES: 5449-54S3 (G) DATE: AUG - 1987 ~H) DOGUMENT NUMBER: US 4810777 (I) FILING DATE: 04-MAR^1987 (J) PUBLICATION DATE: 07-MAR-1989 W093/07892 2 1 2 ~ 3 3 7 PCT/US92/08823 -5fi-(xi) SEQUENCE DESC~IPTION: SEQ ID NO:8 Gly Ile Gly Lys Phe Leu His Ser Ala Ly~

Lys Phe Gly Lys Ala Phe Val Gly Glu Ile }~ 20 Met Asn :

(2) INFORMAT~ON FOR SEQ ID NO:9:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 22 amino acids (B) TYPE: amino acid (C) STRANDEDNESS:
(D) TOPOL~GY: linear (ii~ MOLECULE TYPE: peptide ~;

(ix) FEATURE:
(A) Nt~lE/KEY: magainin peptide.

( x ) PUBL I CAT I ON I NFORMAT I ON:
~A) AUTHOR: Zasloff, Michael (C) JOURNAL: Proceedinqs of the National Academv of Sciences ~;
( I) ) VOLUME: 84 (F) PAC;ES: 5449-5453 (G) DATE: AUG - 1987 (H) DOCUMENT NtJMBER: US 4810777 :-( I ) FILING DATE: 04-MAR-1987 (J) PUBLICATION DATE: 07-MAR-1989 WO 93/07892 PCT/USg~/08823 21~Q33~

-59- .

(xi3 SEQUENCE DESCRIPTION: SEQ ID NO:9:
Ile Gly Lys Phe Leu His Ser Ala Lys Lys Phe Gly Lys Ala Phe Val Gly Glu Ile Met Asn Ser (2) INFORMATION FOR SEQ ID NO:10:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amino acids (B) TYPE: amino acid ::
(C) STRANDEDNESS:
(D) TOPOLOGY: linear (ii) MOLECULE TYPE: peptide (ix) FEATURE:
(A) NAME/KEY: magainin peptide.

(x) PU8LICATION INFORMATION: ~ -:
~A) AUTHOR: Zasloff, ~ichael (C) JOURNAL: ~3~E~L~g__of the National Academy of Scien es (D) VOLUME: 84 (F3 PAC;ES: 5449-5453 (G) DAT~: AUG 1987 (H) DOCUMENT NUMBER: US 4810777 (I) FILING DATE: 04-MAR-19B7 ~J) PUBLICATION VAT : 07-MAR-1989 WO 93/07892 2 1 2 0 3 3 ~ PCT/US92/08~23 -6~- .

(xi) SEQUENCE DESCRIPTION: SEQ ID NO:10:
Gly Lys Phe Leu His Ser Ala Lys Lys Phe Gly Lys Ala Phe Val Gly Glu Ile Met Asn 1~ 20 Ser (2) INFORMATION FOR SEQ ID NO:ll:
(i~ SEQU~NCE CHARACTERISTICS
5A~ LENGTH: 20 amino acids (B~ m~ amino acid ~`
(C) STRANDEDNESS:
(D) TOPOLOGY: linear ~ -.'.
(ii) MOLECULE mE peptide (ix) FEATURE:
~A) NAME/KEY: magainin peptide. ~:

(x) PUBLICATIQN INFORMATION:
~A) AUIHOR: Za510ff, Michael (C) JOURNAL: Proceedinqs of the Nati nal AcademY
of Sciences (D) VOLUME: 84 (F) PAGES: 5449-5453 ~G) DATE: AUG - 1987 -:
~H) DOCUMENT NUMBER: US 4810777 ~I) FILING DATE: 04-MAR-1987 (J) PUBLICATIO~ DATE: 07-MAR-1989 W093/0~892 PCT/US92/08823 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:ll:
Lys Phe Leu His Ser Ala Lys Lys Phe Gly ~:
S 10 , Lys Ala Phe Val Gly Glu Ile Met Asn Ser - ..
15 ~ :

(2) INFORMATION FOR SEQ ID NO:12: :~
(i) SE~UENCE CHARACTERISTICS ~
(A) LENGTH: 21 amino acids ::
(B) m E: amino acid :~
(C) STRANDEDNESS:
(D) TOPOLOGY: linear ;
'~."
(ii) MQLECULE TYPE: peptide -:

(ix) FEATURE: ~
(A) NAME/KEY: PGLa peptide. .~.

(x) PUBLICATION INFORMATION:
(A) AUTHOR: Hoffman, et al.
(C) JOURNAL: EMBO J.
(D) VOLUME: 2 (F) PAGES: 711-714 ~G) DATE: l9B3 (A) AUTHOR: Andreu, et al.
(C) JOURNAL: Journal of Bio hemis~rY
(D3 VOLUME: 149 (F) PAGES: 531-535 :~:
(G) ~ATE: l9a5~
(A) AUT~OR: Gibson, et al.
(C~ JOURNAL: J. Biol. Chem.
(D) VOLUME: 261 (F) PAGES: 5341-5349 (G1 DATE: 1986 WO 93/07892 2 1 2 0 3 3 7 PCT/US92/0~23 . :

(A) AUIHOR: Giovannini, et al.
(C) JOURNAL: Biochem J.
(D) VOLUME: 243 (F) PAGES: 113-120 (G) DATE: 1987 -:~

(xij SEQUENCE DESCRIPTION: SEQ ID NO:12:
Gly Met Ala Ser Lys Ala Gly Ala Ile Ala .
.~,.
Gly Lys Ile Ala Lys Val Ala Leu Lys Ala lS 20 ~-Leu (2) INFORMATION FOR SEQ ID NO:13:
(i) S~QUE~CE CHARACTERISTICS
(A) LENGTH: 25 amino acids (B) TYP~: amino acid (C) ST~ANDEDNESS:
(D) TOPOLO5Y: linear :

(ii) MOLECU~E TYPE: peptide (ix) FEATURE.
(A) NAME/KEY: XPF peptide.

(x) PUBLICATION INFORMATION:
(A) AUTHOR: Hoffman, et al.1 (C) JOURNAL: MBO J.
~D) VOLUME: 2 (F) PAGES: 711-714 (~) DATF~: 1983 (A) AUTHOR: Andreu, et al.
~C) JOURNAL: Journal of Biochemistr~
tD) VOLUME: 149 WO 93/07892 P~T/US92/08823 (F) PAGES: 531-535 (G) DATE: 1985 (A) AUTHOR: Gibson, et al.
~C) JOURNAL: J. Biol. Chem.
(D) VOLUME: 261 (F) PAGES: 5341-5349 (G) DATE: 1986 (A) AUTHOR: Giovannini, et al.
(C) JOURNAL: Biochem J.
(D3 VOLUME: 243 (F) PAGES: 113-12 (G) DATE: 1987 (xi) CE~UENCE DESCRIPTION: SEQ ID NO:13:
Gly Trp Ala Ser Lys Ile Gly Gln Thr Leu 5 lO
Gly Lys Ile Ala Lys Val Gly Leu Lys Glu Leu I le Gln Pro Lys :

(2) INFORMATION FOR SEQ ID NO: 14:
( i ) SEQUENCE CHA~ TERISTICS
(A) LENGTH: 27 amino acids ~ B ) TYPE: amino acid ( C ) STRANDEDNESS:
(D) TOPOLOt;Y: linear ( ii ) MOLECULE TYPE: peptide ( i x ) FEATURE:
~A) NAME/KEY: C~F peptide. ~

( x ) PUBLICATION INFORMATION: -WO 93/07892 . 2 ~ 2 0 3 3 7 PCT/US92/0%823 -64- ~:

(A) AUTHOR: Richter, K.
Egger, R.
Kreil (C) JOURNAL: J. Biol. Chem.
(D) VOLUME: 261 (F) PAGES: 3676-3680 (G) DATE: 1986 (A) AU~HOR: Wakabayashi, T.
Kato, H.
Tachibaba, S.
(C) JOURNAL: Nucleic Acids Re~earch (D) VOLUME: 13 ~F) PAGES: 1817-1828 (G) DATE: 198$
(A1 AUTHOR: Gibson, ~.W.
Poulter, L.
Williams, D.~.
Maggio, J.E. ``
~C) JOURNAL: J Biol. Chem.
(D~ VOLUME: 261 (F) PAGES: 5341-5349 (G) DATE: 1986 ~H) DOCUMENT NUMBER: WOa0/04407 (I) FILING DATE: 16-OCT-1989 ~J) PUBLICATION DATE: 03-MAY-1990 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:14:
Gly Phe Gly Ser Phe Leu Gly 1eu Ala Leu Lys Ala Ala Leu Lys Ile Gly Ala Asn Ala Leu Gly Gly Ala Pro Gln Gln ~5 W093/~78g~ PCT/US92/08823 (23 INFORMATION FOR SEQ ID NO:lS:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 27 amino acids (B) TYPE: amino acid tc) STRANDEDNESS: ~
(D) TOPOLOGY: linear :

(ii) MOLECULE TYPE: peptide (ix) FEATURE:
(A) NAME/KEY: CPF peptide.

(x) PUBLICATION INFORMATION:
(A) AUTHOR: Rirht~r, K
Egger, R. :
Kreil ::.
(C) JOURNAL: J. Biol. Chem.
(D) VOLUME: 251 ~:
(F) PAGES: 3~76-3680 ~:
(G) DATE: 198S
(A) AUTHOR: Wakabayashi, I.
Kato, H.
Tachibaba, S.
(C) JOURNAL: ~Es~h~ a~s_Research (D~ VOLUME: 13 (~) PAGES: 1817-1828 (~) DATE: lg85 (A) AUTHO~: Gibson, B.W.
Poulter, L.
Williams, D.H.
Maggio, J.E.
(C) JOURNAL: J. Biol. Chem.
(D) ~OLUM~: 261 .
(F) PAGES: 5341-5349 - ~
' '.

2 1203~7 W093/07~92 PCT/US9~/08823 (G) DATE: 1986 (H) DOCUMENT NUMBER: W090/04407 (I) FILING DATE: 16-OCT-1989 (J) PUBLICATION DA~E: 03-MAY-1990 (xi) 5EQUENCE DESCRIPTION: SEQ ID NO:lS:
Gly Leu Ala Ser Phe Leu Gly Lys Ala Leu Lys Ala Gly Leu Lys Ile Gly Ala His Leu lS 20 Le~ Gly Gly Ala Pro Gln Gln :

2) INFORMATION FOR SEQ ID NO:16:
(i) SEQUENCE CHARACTERISTICS
~A) LENGTH: 27 amino acids ~B) TYPE: amino acid (C) STRANDEDNESS:
~D) TOPOLOGY: linear (ii) MOLECULE TYPE: peptide (ix) FEATURE:
(A) NAME/KEY: CPF peptide.

(x~ PUBLICATION IN~ORMATION:
(A~ AUTHOR: Richter, K.
Egger, R.
Kreil (C) JOU~NAL: J. Biol. Chem (D) VOLUME: ~61 (F~ PAGES: 3676-368 (G) DATE: 1986 (A) AUTHOR: Wakabayashi~ T.

WO 93/07892 . PCI`/US92/08823 Kato, H.
Ta ::hibaba, S .
(C) JOURNAL: Nucleic Acids Resear~h (D ) VOLVME: 13 (F) PAGES: 1817-1828 (G) DATE: 1985 ::
( A ) AUTHO~: Gibson, B . W .
Poulter, L.
Williams, D.H. -Maggi o, J . E .
(C) JOURNAL: J. Biol. Chem.
( D ) VOL~ME: 2 61 (F) PAGES: 5341-~34 ( G ) DATE: 1986 (H) DOCUMENT NUMBER: W090/04407 (I) FILING DATE: 16-OCT-1~8g ( J ) PIJBLICATION DATE: 03-MAY- 1990 (xi ) SEQUENCE DESCRIPTION: SEQ ID NO: 16: ::
Gly Leu Ala Ser Leu Leu Gly Lys Ala Le~

Lys Ala Gly Leu Lys Ile Gly Thr E~is Phe 2û
Leu Gly Gly Ala Pro Gln Gln (2 ) INFORMATION F5:)R SEQ ID NO: 17:
i ) SEQUENCE C~ARACTERI STICS
(A) LENGTH: 27 amino acids (B~ TYPE: amino acid ( C ) STRANDEDNESS:
( D ) TOPOLOC;Y: 1 ine ar ( ii ) MOLECULE TYPE: peptide WO 93/07892 ~ PCT/US92/08823 (ix) FEATURE:
(A) NAME/KEY: CPF peptide.

(x) PUBLICATION INFORMATION:
(A) AUTHOR: Richter, K.
Egger, R.
Kreil (C) JOURNAL: J. 8iol. Chem.
(D) VOLUME: 261 (F) PAGES: 3676-3680 ~G) DATE: 1986 :
(A) AUTHOR: Wakabayashi, T. :
Kato, H.
Tachibaba, S.
(C) ~OURNAL: Nucleic Acids Research (D) VOLUME: 13 (F) PAGES: lB17-1828 ~) DATE: 1985 (A) AUTHOR: Gibson, B.W.
Poulter, L.
Williams, D.H.
Maggio, J.E. ~ ~`
(C~ JOURNAL: J. Biol. Chem.
(D) VOLUME: 261 (F) PAGES: 5341-5349 (G) DATE: 1986 (H) DOCUMENT NUMBER: WO90/04407 (I) FILING DATE: 16-OCT-1989 ~J) PUBLICATION DATE: 03-MAY l99O

WO 93/07892 PCT/USg2/08823 2121)337 -6g- :

(xi) SEQUENCE DESCRIPTION: SEQ ID NO:17:
Gly Leu Ala Ser Leu Leu Gly Lys Ala Leu .

Lys Ala Thr Leu Lys Ile Gly Thr His Phe ~.
Leu Gly Gly Ala Pro Gln Gln -(2) INFORMATION FOR SEQ ID NO:18~
(i) SEQUE~CE CHARACTERISTICS
(A) LEN~TH: 27 amino acids ~B) TYPE: amino acid (C) STRANDEDNESS: .
(D) TOPOLOGY: linear ::

(ii) MOLECULE TYPE: pe~tide ~

(ix) FEATURE:
(A) NAME/KEY: C~F pepti~e.

(x) PUBLICATION INFORMATIO~:
~A) AUIHOR: Richter, K. , :-Egger, R.
Kreil (C) JOURNAL: J. Biol. Chem (D) VOLUME: 261 -(E) PAGES: 3676-3680 (G) DATE: 1986 (A) AUTHOR: Wakabayashi, T.
:
Kato, H.
Tachibaba, S.
(C) JOURNAL: Nucleic Acids_Research (D) VOLUME: 13 (F) PAGES: 1817-1828 -wo 93/0,8g2 2 1 2 0 3 3 7 PCT/US92/~8823 -70~.

(G) DATE: 1985 .~
(A) AUTHOR: Gibson, B.W. :
Poulter, ~.
Williams, D.H.
Maggio, J.E.
(C) JOURNAL: J. Biol. Chem (D) VOLUME: 261 (F) PAGES: 5341-5349 (G) DATE: 1986 (H) DOCUMENT NVM~ER: WO90/04407 ~I) FILING DATE: 15-OCT-1~89 (J) PUBLICATION DATE: 03-MAY-1990 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:18:
Gly Phe Ala Ser Phe Leu Gly Lys Ala Leu Lys Ala Ala Leu Lys Ile Gly Ala Asn Met ~0 Leu Gly Gly Thr Pro Gln Gln ~:.
.

~2) INFORMATION FOR SEQ ID NO:1:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 27 amino acids (B) TYPE: amino arid ~C) STRANDEDNESS:
~D) TOPOLOGY: linear ~ii) MOLECULE TYPE: peptide (ix) FEATURE:
(A) NAME~KEY: CPF peptide.

(x) PUBLICATION INFORMATION:

W093/07~2 PCT/US92/~823 (A) AUIHOR: Richter, K.
Egger, R.
Kreil (C) JOURNAL: J. ~iol. Chem. -~-(D) VOLUME: 261 (F) PAGES: 3676-3680 (Gj DATE: 1986 (A) AUTHOR: Wakabayashi, T.
Kato, H.
Tachibaba, S.
(C) JOURNAL: I~ucleic Acids_Researoh (D) VOLUME: 13 (F) PAGES: 1817-1828 (G) DATE: 1985 (A) AUIHOR: Gibson, B.W.
Poulter, L.
Williams, D.B.
Maggio, J.E.
(C) JOURNAL- J. Biol. Chem.
~D) VOLVME: 261 :~
(F) PAGES: 5341-5349 (G~ DATE: la86 ~H) DOCUMENT NUMBER: WO90/04407 ~I) FILING DATE: 16-OCT-1989 (J) PUBLICATION DATE: 03-MAY-1990 ~xi~ SEQUENCE DESC~IPTI~N: SEQ ID NO:19:
Gly Phe Gly Ser Phe Leu 51y Lys Ala Leu :~
Lys Ala Ala Leu Lys Ile Gly Ala Asn Ala Leu Gly Gly Ala Pro Gln Gln ::.

W093/07892 2 1 2 ~ 3 3 7 pCT~US92/08823 (2) INFORMATION FOR SEQ ID NO:20: -(i) SEQUENCE CHARACTERISTIC5 (A) LENGTH: 27 amino acids (B) TYPE: amino acid (C) STRANDEDNESS:
(D) TOPOLOGY: linear (ii) MOLECULE TYPE: peptide (ix) FEATURE: ~
(A) NAME/KEY: CPF peptide.

(x) PUBLICATION INFORMATION:
(A) AUTHOR: Richter, K.
Egger, R.
Kreil (C) JOURNAL: J. Biol. Chem.
(D) VOLUME: 261 (F) PAGES: 3676-3680 (G3 DATE: l9B6 (A) AUTHOR: Wakabayashi, T.
Kato, H.
Taçhibaba, S.
(C) JOURNAL: Nucleic Acids Research (D) VOLUME: 13 (F~ PAGES: 1817-1828 (G) DATE: 1985 (A) AUTHOR: Gibson, B.W.
Poulter, L.
William , D.H.
Maggio, J.E.
(C) JOURNAL: J. Biol. Chem.
(D) VOLUME: 261 (F) PAGES: 5341-5349 WO 93/07892 PCI/USg2/08823 2 12~337 ( G ) DATE: 1986 (H) DOCUMENT ~UM8ER: W090/04407 ( I ~ FILING DATE: 16-OCT- 1989 (J) PUBLICATION DATE: 03-MAY-1990 (xi) SEQUENCE DE5CRIPTION: SEQ ID NO:20 Gly Phe Gly Ser Phe Leu Gly Lys Al a Leu S 10 . ,,~
Lys Ala Ala Leu Lys Ile Gly Ala Asn Ala 1~ 20 ~
I.eu Gly Gly Ser Pro Gln Gln .~.

~2) INFORMATION FOR SEQ ID NO:21: ~.
~i ) SEQUENCE CHARACTERISTICS
(A) LENGTH: 27 amino acids ~:
(8) TYPE: amino a~id ( C ) STRANDEDNESS:
(D ) TOPOLOGY: linear ~ii ) MOLECULE TYPE: peptide ( i x ) FE:ATURE:
~ A ) NAME/KEY: CPF pepti de .

( x ) PUBL I CATI QN INFORMAT I ON:
( A ) AUTHOR: Ri chter, K .
Egger, R.
Krei 1 (C) JOURNAL: J. Biol. Chem.
(D) VOLUME: 261 (F) PAGES: 3676-3680 (G) DATE: l9B6 (A) AUTHOR: Waka~ayashî, T.

212~337 WO 93/07892 PCT/USg2/0~823 Kato, H.
Tachibaba, S.
(C) JOURNAL: Nucleic Acids Research (D~ VOLUME: 13 (F) PAGES: 1817-1828 (G~ DATE: 198~
(A) AUTHOR: Gibson, B.W.
Poulter, L. :~
Williams, ~
Maggio, J.E.
(C) JOURNAL: J. Biol. Chem.
(D) VOLUME: 261 (F) PAGES: 5341-5349 (G) DATE: 1986 :
(H) DOCUMENT NUMBER: WO90/04407 (I) EILING DATE: 16-OCT-1989 ~J) PUBLICATION DATE: 03-MAY-1990 -~

(xi) SEQUENCE DESCRIPTION: SEQ ID NO:21:
Gly Phe Ala Ser Phe Leu Gly Ly~ Ala Leu Lys Ala Ala Leu Lys lle ~ly Ala Asn Leu 1~ 20 Leu Gly Gly Thr Pro ~ln Gln 2~

(2) INEORMATION FOR SEQ ID NO:22:
~i) SEQUENCE CRARACTERISTICS :`
(A) LENGTH: 27 amino acids (B) TYPE: amino acid (C) STRANDEDNESS:
~D) TOPQLOGY: linear (ii) MOLECVLE TYPE: peptide WO g3/078g2 . PCI`~US92/08823 212~337 .
( ix ) FEATURE:
(A) NAME/KE;Y: CPF peptide.

. .
( x ) PUBLIC~TION INFORMATION:
( A ) AUTHOR: Ri chter, K .
Egger, R.
Kre i l ~C) JOURNAL: J. Biol. Chem ~.
(D) VOLUME: 261 (F) PAGES: 3676-3680 ::.
( G ) DATE: 1986 ~;
~A) AUT~OR: Wakabayashi, T.
Kato, H.
Tachibaba, S.
(C) JOURNAL: Nucleic Acids Research (D) VOL~JME: 13 ~ F ) PAGES: 1817-1828 ( C; ) DATE: l9S5 ~ -( A ) AUT~OR: Gi}:~son, B . w .
Poult~r, L.
Williams, D.H.
Maggi o, J . E .
(C) JOIJRNAL J. Biol. Chem.
~ D ~ VOLUME 2 61 (F) PAGE5: 5341-5349 ~ G ) DATE: 1986 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:22:
Gly Phe Ala Ser Phe Leu Gly Lys Ala Leu Lys Ala Ala Leu Lys Ile Gly Ala Asn Ala Leu Gly ~;ly Ala Pro Gln Gln - :

wo 93/078g2 2 1 2 0 3 3 7 PCT/US92/08X23 -7~- :

(2) INFORMATION FOR SEQ ID NO:23~
~i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 27 amino acids (B) TYPE: amino a~id `
(C) STRANDEDNESS: -(D) TOPOL WY: linear (ii) MOLECULE TYPE: peptide (ix) FEATU~E:
(A) NAME/KEY: CPF peptide.

(x) PUBLICATION INFO~MATION:
(A) AUTHOR: Richter, ~. ~
Egger, R.-~.
Kreil (C) JOURNAL: J. Biol. Chem.
(D) VOLUME: 261 ~::
(~) PAGES: 3676-3680 (G) DATE: 1~86 (A) AUIHOR: Wakabayashi, T. ~-Kato, H.
Tachibaba, S.
(C) JOURNAL:
~D~ VOLUME: 13 (F~ PA~ES: 1817-18~8 (G) DATE: 1985 ~A~ AUT~OR: Gibson, B.W.
Poulter, L.
Williams, D.H.

Maggio, J.E.
(C) JOURNAL: J. Biol. Chem. ~:
~D) VOLUME: 261 (F) PAGES: 5341 5349 -:.
.-:

W~ ~3,~,0 ~ 7 PCT/US92/~8823 (G) DATE: 1986 .

(xi) SEQUENCE DESCRIPTION: SEQ ID NO:23:
Gly Phe Ala Ser Phe Leu Gly Lys Ala Leu Lys Ala Ala Leu Lys Ile Gly Ala Asn Met lS 20 Leu Gly Gly Ala Pro Gln Gln (2) IN~ORMAT~ON FOR SEQ ID NO:24:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 27 amino acids (B~ TYPE: amino acid (C) STRAN~EDNESS:
(D) TOPOLOGY: linear (ii) MOLECULE ~YPE: peptide (ix) FEATURE:
(A) NAME/KEY: CPF peptide.

(x) PUBLICATION I~FORMATION:
(A) AUT~OR: Richter~ K.
Egger, R.
Kreil (C) JOURNAL: J. ~iol~ Chem.
(~) VOLUME: 261 (F) PAGES: 3676-3680 (G) DATE: l9B6 (A) AUTHOR: Wakabayashi, T.
Kato, H.
Tachibaba, S.
(C) JOURNAL: Nucleic_Acids Research W093/07892 2 1 2 0 3 3 7 Pcr/US92/0~823 -78- ::

(D) VOLUME: 13 ~F) PAGES: 1817-1828 :~.
(G) DATE: 1985 ~A) AUTXOR: Gibson, 8.W.
Poulter, L.
Williams, D.H.
Maggio, J.E. :
(C) JOURNAL: J. Biol. Chem.
tD) VOLUME: 261 ~F) PAGES: 5341-5349 (G) DATE: 1986 ~xi) SEQUENCE DESCRIPTION: SEQ ID NO:24: ::~
Gly Phe Gly Ser Phe Leu Gly Lys Ala Leu 5 10 :
Ly5 Ala Ala Leu Lys Ile Gly Ala Asn Ala lS 20 Leu Gly Gly Ser Leu Gln Gln :

(2~ INFORMATION FOR SEQ ID NQ:2S:
(i) SEQUENCE CHARACTERISTICS
~A) LENGTH: 27 amino acids (B) TYPE: ami~o acid (C) STRANDEDNES5:
(D) TOPOLOGY: linear (ii) MOLECULE IYPE: peptide (ix) FEATURE:
(A) NAME/KEY: CPF peptide.

(x) PUBLICATION INFORMATION:
(A) AUTHOR: Richter, K.

W093/07892 PCT/US92~08823 2 1 ~3~7 Egger, R.
Kreil (C) JOURNAL: J. Biol. Chem.
(D) VOLVME: 261 ~F) PAGES: 3676-3680 (G) DATE: 1986 ~A) AUTHOR: Wakabayashi, T.
Kato, H.
Tachibaba, 5.
(C) JOURNA~: Nucleic Acids R~cear~h (D) VOLUME: 13 (F) PAGES: 1817-1828 (G) DATE: l9a5 (A) AUTHOR: G~bson, B.W.
Poulter, L.
Williams, D.H.
Maggio, J.E.
(C) JOURNAL: J Biol. Chem.
(D) VOLUME: 261 ~F) PAGE5: 5341-534 (G) DATE: 1986 . .
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:25:
Gly Phe Gly Ser Phe Leu Gly Lys Ala Leu 5 1~
Lys Ala ~ly Leu Lys Ile Gly Thr Asn Phe ~5 ~0 Leu Gly Gly Ala Pro Gln Gln (2~ INFO~MATION FOR S~Q ID NO:26:
(i) SEQUENCE CHARACTERISTICS
(A) ~ENGTH: 27 amino aeids W093/û7892 2l2n337 PCI`/US~2/08823 ( B ) TYPE: amino acid ( c ) STRANl:~EDNESS:
(D ) TOPOLOGY: linear ( ii ) MOLECULE TYPE: peptide ( i x ) FEATURE:
(A) NAME/KEY CPF peptide.

( x ) PUBLI CATI ON INFORMP.TION:
(A) AUTHOR: Richter, K
Egger, R.
Krei 1 ~:
(C~ JOURNAL: J. Biol. Chem.
(D~ VOLUME: 261 (F) PAGES: 3676-3680 ( G ) DATE: 1986 ( A ) AUTHOR: Wakabayashi, T .
Kato, H.
Tachi~aba, S.
(C) JOURNAL: Nucleic Acids Research (D) VOLUME: 13 ( F ) PAGES: 1817-1828 (G) DATE: l9B5 ( A ~ At~OR: Gibson, B . W .
Poulter, L.
Williams, D.H.
Maggio, J. E.
( C ) JOURNAL: J . _ Biol . Chem .
( D ) VOLUME: 2 61 (F) PAGES: 5341-53 (G) DATE: 1986 WO 93/07892 PCT/uS92/0X823 2l2n.~ 7 (xi) SEQUENCE DESCRIPTION: SEQ I~ NO:Z6:
Gly Leu Ala Ser Leu Leu Gly Lys Ala Leu Lys Ala Ala Leu Lys Ile Gly Ala Asn Ala lS 20 Leu Gly Gly Ser Pro Gln Gln (2~ INFO~MATION FOR SEQ ID NO:2~:
(i) SEQUENCE CHAR~CTERISTICS
~A) LENGTH: 2l amino acids (B) TYP~: amino acid (C~ STRANDEDNESS:
~) TOPOLOGY: linear ~ii) MOLECULE TYPE: peptide (xi) SEQUENCE DESCRIPTION: SEQ ID NO:27:
Lys Ile Ala Gly Lys Ile Ala Lys Ile A1B
Gly Lys Ile Ala Lys Ile Ala Gly Lys Ile lS 20 Ala (2) I~IFORMATION FOR SEQ ID NO:28:
(i) SEQUENCE CHA~ACTERISTICS:
~A) LEN~TH: 2l amino acids (B~ m E: amino acid (C) STRANDEDNESS:
[D) TOPOLOGY: linear (ii) MOLECVLE TYPE: peptide WO 93/07892 2 1 2 0 3 ~ 7 PCT/US92/08823 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:28:
Lys Ile Ala Lys Ile Ala Gly Lys Ile Ala .

Lys Ile Ala Gly Lys Ile Ala Lys Ile Ala Gly (2) INFORMATION FOR SEQ ID NO:29:
(i) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 21 amino acids (B) TYPE: amino acid ~C~ STRANDEDNESS: :
(D) TOPOLOGY: linear (ii) MOLECULE TYPE: peptide -(xi) SEQUENCE DESCRIPTION: SEQ ID NO:29:
Lys Ile Ala Gly Lys Ile Gly Lys Ile Ala Gly Lys Ile Gly Lys Ile Ala Gly Lys Ile 1~ 20 Gly (2) INFORMATION ~OR SEQ ID NO:30:
(i) SEQUENCE C~ARACTERISTICS
(A) LENGTH: 21 amino acids ~B) TYPE- amino acid (C) STRANDEDNESS:
(D) TOPOLOGY: linear (ii) MOLECULE TYPE: peptide WO 93/~7892 - PCT/US92/08823 21203~7 ` ~`

-83- :
.

(xi) SEQUENCE DESCRIPTION: SEQ ID NO:30:
Lys Leu Ala Gly Lys Leu Ala Lys Leu Ala Gly Lys Leu Ala Lys Leu Ala Gly Lys Leu Ala (2) INEORMATION ~OR SEQ ID NO:31:
(i) SEQUENCE ~HARACTERISTICS :~:
(A) LENGT~: 21 ami~o acids (B) TypF: amino acid (C~ STRANDED~ESS:
~D) TOPOLOGY: linear (ii) MOLECULE TYPE: peptide ~xi) SE~UENCE D~SCRIPTION: SEQ ID NO:31: 5'' Lys Phe Ala Gly Lys Phe Ala Lys Phe ~la Gly Lys Phe Ala Lys Phe Ala Gly Lys Phe Ala (2~ INFORMATION FOR SEQ ID NO:32:

(i3 SEQUENCE CHARACTERISTIC5 (A) LENGTH: 21 amino acids (B) TYPE: amino a~id ~C) STRANDEDNESS:
(D) TOPOLOGY: linear (ii) MOLECULE TYPE: peptide WO 93/O~X92 2 ~ 2 ~ 3 3 7 PcT/vs92/o88~3 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:32: -Lys Ala Leu Ser Lys Ala Leu Lys Ala Leu 5 10 :-~
Ser Lys Ala Leu Lys Ala Leu Ser Lys Ala ~-~
lS 2D ::
Leu :

(2) INFORMATION FOR SEQ ID NO:33:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amino acid~
~B) m E: amino acid (C) STRANDEDNESS:
(D) TOPOLOGY: linear (ii) MOLECULE TYPE: peptide (xi) SEQUENCE DESCRIPTION: SEQ ~D NO:33:
Lys Leu Leu Lys Ala Leu Gly Lys Leu Leu Ly5 Al a Leu Gly Lys Leu Leu Lys Ala Leu ~;ly (2) INFORMATION EOR SEQ ID NO:34:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amino a~ids (B) m E: amino acid (C) STRANDEDNESS;
~D) TOPOLOGY: linear (ii) MOLECULE TYPE: peptide .~. .

, . .

W093/~7892 PCT/US92/08823 212033~

(xi) SEQUENCE DESCRIPTION: SEQ ID NO:34:
Lys Ala Ile Gly Lys A~a Ile Lys Ala Ile Gly Lys Ala Ile Lys Ala Ile Gly Lys Ala -Ile (2) INFORMATION FOR SEQ ID NO:35:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amino acids (B) TYPE: amino a~id (C) STRANDEDNESS:
(D) TOPOLOGY: linear (ii) MOLECULE TYPE: peptide ~ .
(xi~ SEQUENCE DESCRIPTION: SEQ ID NO:35: ~-Gly Ile Ala Lys Ile Ala ~ys Gly Ile Ala 5 10 ~
Lys Ile Ala Lys Gly Ile Ala Lys Ile Ala :
Lys (2) INFORMATION FOR SEQ ID NO:36:
(i) SEQUENCE CHARACTERISTICS
(A~ LENGTH: 21 amino acids (B) TYPE: amino acid (C) STRANDEDNESS:
(D) TOPOLOGY: linear (ii) MOLECULE TYPE: peptide .,,~ .
,~ .

WO 93/07892 2 1 2 0 3 3 7 PCT~US92/0~8~3 ~

-8~- :

~xi) SEQUENCE DESCRIPTION: SEQ ID NO:36:
Lys Ile Ala Lys Ile Phe Gly Lys Ile Ala Lys Ile Phe Gly Lys Ile Ala Lys Ile Phe Gly ~2) INFORMATION FOR SEQ ID NO:37:
(i) SEQUENCE CHARACT~RISTICS
(A) LENGTH: 21 amino acid~
(B) TYPE: amino acid (C) STRANDEDNESS:
(D) TOPOLOGY: linear (ii) NOLECULE TYPE: peptide (xi) SEQUENCE DESCRIPTION: SEQ I~ NO:37:
_Gly ~le Ala Arg Ile Ala Lys Gly Ile Ala Arg Ile Ala Lys Gly Ile Ala Arg Ile Ala Lys (2) INFORMATION FOR SEQ ID NO:38:
(i~ SEQUENCE CRARACTERISTICS
~A) LENGTH: 21 amin~ acids (B) TYPE: amino acid (C) STRANDEDNESS:
(D) TOPOLOGY: linear (ii) MO~ECULE ~YPE: peptide (xi) SEQUENCE DESCRIPTION: SEQ ID NO:3B:
Lys Phe Ala Arg Ile Ala Gly Lys Phe Ala Arg Ile Ala Gly Lys Phe Ala Arg Ile Ala Gly ~2) IMFOXMATION FOR SEQ ID NO:39:
(i) SEQUENCE CHARACTERISTICS
- (A) LENGTH: 21 amino acids (B) TYPE: amino acid ~C) STRANDEDNESS:
(D) TOPOLOGY: linear (ii) MOLECULE TYPE: peptide (xi) SEQ~ENC~ DESCRIPTION: SEQ ID NO:39:
_Gly Phe Ala Lys Ile Ala Lys 51y Phe Ala Lys I le Ala Lys Gly Phe Ala Lys Ile Ala Lys `~

(2) INFORMATION FOR SE~ ID NO:40:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amino acids (B) TY}?E: amino acid ~ C ~ STRANDEDNESS:
( 1) ) TO~OLQGY: 1 i ne ar .
(ii) MOLECULE TYPE: peptide 212~3~7 .

(xi) SEQUENCE DESCRIPTION: SEQ ID NO:40:
. Lys Ile Ala Gly Orn Ile Ala Lys Ile Ala Gly Orn Ile Ala Lys Ile Ala Gly Orn Ile Ala (2) INFORMATION F0R SEQ ID N0:41:
(i) SEQUENCE C~AR~CTERISTICS
~ ~A) LENGTH: 21 amino acids ::
~B) TYPE: amino acid ( C ) STRANDEDNESS:
D ~ TOPOLOGY: linear :.
(ii~ MOLECULE TYPE;: peptide ::

(xi) SEQIJENCE DESCRIPTION: SEQ ID NO:41:
Lys Ile Ala Arg Ile Ala Gly Lys lle Ala Arg Ile Ala Gly Lys Ile ~la Arg Ile Ala ~0 Gly ~2) IN~0RMATI0N ~OR SEQ ID NO:42:
(i) SEQUENCE CHARACTERISTICS
~A) LENGTH: 21 amino acids (B) TYPE: amino acid C ) STRANDEDNE S S:
~ D ) TOPOI.OGY: l i ne a r (ii) MOLECULE TYPE: peptide WO 93/07892 PCT/US92/08~23 :

(xi) SEQUENCE DESCRIPTION: SEQ ID NO:42:
Orn Ile Ala Gly Lys Ile Ala Orn Ile Ala 5 10 .:
Gly Lys Ile Ala Orn Ile Ala Gly Lys Ile 15 20 :
Ala ~:~

J (2) INFORMATION FOR S~Q ~D NO:43:
(i) SEQUENCE CHARACTERISTICS ::
- (A) LENGTH: 21 amino acids (B) m E: amino acid (C) 5TRANDE~NESS: ~
~D) TOPOLOGY: linear :
.:
Iii) MOLECULE TYPE: peptide (xi) SEQUENCE DESCRIPTION: SEQ ID NO:43:
Gly Ile Ala Arg Ile Phe Lys Gly Ile Ala Arg Ile Phe Lys Gly Ile Ala Ar~ Ile Phe .~:
Lys :~
(2) INFO~;ATION FOR SEQ ID NO:44:
(i) SEQUENCE C~ARACTERISTICS
(A) LENGTH: 21 amino acids (B) m E: amino acid (C) STRANDEDNESS:
(D) TOPOLOGY: linear (ii) MOLECULE TYPE: peptide 21203~7 WO 93!07892 PCT/US92/08823 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:44:
Lys Nle Ala Gly Lys Nle Ala Lys Nle Ala 5 10 ;
Gly Lys Nle Ala Lys Nle Ala Gly Lys Nle Ala (2) INFORMATION FOR SEQ ID NO:45:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amino acids (B) TYPE: amino acid (C) STRANDEDN~SS:
(D) TOPOLOGY: linear (ii) MOLECUL~ TYPE: peptide (xi) SEQUENCE DESCRI~TIO~: SEQ ID NO:54:
Lys Nle Ala Gly Lys Ile Ala Lys Mle Ala Gly Lys Ile Ala ~ys Nle Ala Gly Ly~ Ile 1~ 20 Ala ~2) INFORMATION ~OR SEQ ID NO:46:
(i) SEQUENCE C~ARACTERISTICS
~A) LENGTH: 21 amino acids (~1 TYPE: amino acid (C) STRANDEDNESS:
(D) TOPOLOG~: linear (ii) MOLECULE mE peptide WO 93r07892 PCI~US92/08~23 :
212U3~7 :

-gl- ~.

(xi ) SEQUENCE DESCRIPTION: SEQ II) NO: 46:
Lys Ile Ala Gly Ly~i Nle Ala Lys Ile Ala Gly Lys Nle Ala Lys Ile Ala Cly Lys Nle -Ala ( 2 ) INFORMATION FOR SEQ ID NO: 47:
( i ) SEQUENCE CHARACTERISTICS :~
(A) LENGTEI: 21 amino acid~ ~' (B) TYPF~: amino acid ( C ) STRANDEDNE S S:
(D) TOPOLOGY: linear ( ii ) MOLECULE TYPE . peptide :

(xi ) SEQUENCE DESCRIPTION: SEO ID NO: 47:
- Lys Nva Ala Gly Lys Nva Ala Lys Nva Ala 5 10 .
Gly Lys Nva Ala Lys Nva Ala Gly Lys Nva Ala 2 ) INFORMATION FC)R SEQ ID NO: ~a (i ~ SEQUENCE C~ARACTERISTICS
(A) LENGTHo 21 smino acids ( ~ ~ TYPE o amino acid ~ C ) STRANDEDNESS:
(D~ TOPOLOGY: linear ( i i ) MOLECULE TYPE: pept i de WO 93/07892 2 ~ 2 D 3 3 7 Pcr/US92/0~823 (xi) SEQUENCE DESC~IPTION: SEQ ID NO-48:
Lys Nva Ala Gly Lys Ile Ala Lys Nva Ala lQ
Gly Lys Ile Ala Lys Nva Ala Gly Ly5 Nva Ala ( 2 ) INFORMATION FOR SEQ ID NO: 49:
( ~ ) SEQUENCE CHARACTERISTICS
- (A) I.ENGl~l: 21 amino acid~
(B) TYPE: amino acid ( C ) STRAN~EDNES S:
(D ) TOPOLOGY: linear ( i i ) MOLECULE ~YPE: peptide (xi) SE5~UENCE D5C~IPTION: SEQ ID NO:49:
_ Lys I.eu Leu Ser Lys Leu Gly Lys Leu Leu Ser Lys Leu Gly l.ys Leu Leu Ser Lys Leu Gly (2) INFORMATION FOR SEQ Il:) NO:50:
~i ) SEQIJENGE CH~RAC:TERISTIC5 (A) LENGTH: 21 amino acids ( B ) ~:PE: aminc~ acid C ) STRANDE:DNESS:
~D) TOPOLOGY: lYne~r ( ii ) MOLECULE TYPE: peptide WO93/078g2 PCT/uS92/08$23 ::~
2:12a337 ~

-93- -~

(xi) SEQUENCE DESCRIPTION: SEQ ID NO:50: ~
Lys Leu Leu Ser Lys Phe Gly Lys Leu Leu ~:
Ser Lys Phe Gly Lys Leu Leu Ser Lys Phe Gly (2) INFORMATION FOR SEQ ID NO:51:
(i) SEQUENCE CHARACTERISTICS
~A) LENGTH: 21 amino a~ids ~B) TYPE: amino acid ::
(C~ STRANDEDNESS:
(D) TOPOLOGY: linear (ii) MOLECULE TYPE: peptide :

(xi) SEQUENCE DESC~IPTION: SEQ ID NO:51:
~ Lys Ile Ala Gly Lys Nva Al~ Lys Ile Ala Gly Lys Nva Ala Lys Ile Ala Gly ~ys Nva Ala (2) INFO~MATION FOR S~Q ID NO:52:
EQUENCE CHARACTERISTICS
(A) ~ENGT~: 21 ami~o acids (B) m E: amino acid (C) STRANDEDNESS:
(D) TOPOLOGY: linear (ii) MO~ECULE TYPE: peptide wo 93~0~8g2 2 1 2 0 3 3 7 PCT/~S92/08823 .
(xi) S~QUENCE DESCRIPTION: SEQ ID NO:52:
His Ile Ala Gly His Ile Ala His Ile Ala Gly His Ile Ala His Ile Ala Gly His Ile Ala (2) INFORMATION FOR SEQ ID NO:53:
(i) SEQUENCE CHARACTERI5TICS
(A) LENGTH: 21 amin~ acids (B) TYPE: amino acid tC) STRANDEDNE55 (D) TOPOLOGY: linear (ii) MOLECULE TYPE: peptide (xi) SEQUENCE DESCRIPTION: SEQ ID NO:53:
~Ala Gly Lys Ile Ala Lys Ile Ala Gly Lys Ile Ala Lys Ile Ala Gly ~ys Ile Ala Lys `~
Ile (2) INFORMATION FOR SEQ ID NO:54:
(i) SEQUENCE CHARACTERISTICS
(A) LENGT~: 21 amino acids (B~ TYPE: amino acid SC) STRANDEDNESS
(D) TOPOLOGY: linear (ii) MOLECULE TYPE: peptide WO 93/07892 PCT~US92108823 2121)337 -95- :

(xi) SEQUENCE DESCRXPTION: SEQ ID NO:54:
Ile Ala Lys Ile Ala Gly Lys Ile Ala Lys Ile Ala Gly Lys Ile Ala Lys Ile Ala Gly 1~ 20 Lys .
(2) INFORMATION FOR SEQ ID NO:55:
(i) SEQUENCE CHARACT~RISTICS
(A) LENGTH: 21 amino acids `
(B) TYPE: amino acid (C) STRANDEDNESS:
(D) TOPOLOGY: linear (ii) MOLECULE TYPE- peptide (xi) SEQUENCE DESC~IPTION: SEQ ID NO:55: -.
Lys Ile Ala Gly Arg Ile Ala Lys Ile Ala f 5 10 ~ly Arg Ile Ala Lys Ile Ala Gly Arg Ile Ala (2) INFORMATION FOR SEQ ID NO:56:
(i) SEQUENCE ~HARACTERISTICS
(A) LENGTH: 21 amino acids ~B) TYPE: amino acid ~C) STRANDEDNESS: -:
(D) TOPOLOGY: linear (ii) MOLECULE mE peptide WO 93/07892 2 12 D 3 3 ~ PCr/lJS92/08X23 _9~_ (xi ) SEQUENCE DESCRIPTION: SEQ ID NO: S6:
Arg Ile Ala Gly Arg Ile Ala Arg Ile Ala C;ly Arg Ile Ala Arg Ile Ala Gly Arg Ile i~0 .
Ala (2) INFORMATION FOR SEQ ID NO:57:
( i ) SEQUENCE CHARACTERI STIC:S
(A) LENGS~I: 21 amis~o acids (~) mE: arnino acid ( C ) STRA~DEDNESS:
( D ) TOPOLOGY: 1 ine ar ( ii ) MOLE5ULE TYPE: peptide ( xi ) SEQUENCE DESCRIPTION: SEQ ID NO: 57: ~:~
Lys Val Ala Gly Lys Ile Ala Lys Val Ala ;~
Gly Lys Ile Ala Lys Val Ala Gly Lys Ile :~
1~ 20 :~:
Ala ~`
(2) INFORMATION FOR SEQ ID NO:58:
( i ) SEQUENt:E CHARACTERISTICS
(A) LENGTH: 21 aminc: acids (B~ mE: amino acid C ~ 5TRANDE:DNESS: -(D) TOPOLOGY: 1 inear (ii) MOLECULE TS!PE: peptide WO 93/07892 PCT/US92/08823 ~
2120337 ~

(xi) SEQUENCE DESCRIPTION: SEQ ID NO:58:
Lys Ile Ala Gly Lys Val Ala Lys Ile Ala S 10 .,:
Gly Lys Val Ala Lys Ile Ala Gly Lys Val Ala (2) INEORMATION FOR SEQ ID NO:59:
(i) SEQUENCE CHARACTERIST$CS -~
~ (A) LENGTH: 21 amino acid~ :
(B) TYPE: amino a~id ~:
~C) STRANDEDNESS:
(D) TOPOLOGY: linear ~ii) MO~ECULE m E: peptide (xi) SEQUENCE DESCRIPTION: SEQ ID NO:59:
~Ala Lys Ile Ala Gly Lys Ile Ala Lys Ile :-",."
Ala Gly Lys Ile Ala Lys Ile Ala Gly Lys .
15 20 -~
Ile (2) INFORMATION ~OR SEQ ID NO:60:
(i) SEQUENCE CHARACTERISTICS
(~ LENGTH: 21 amin~ acids (B) TYPE: amino acid ~C) STRANDEDNESS: :
(D) TOPOLOGY: 1inear MOLECULE TYPE: peptide W093/~7892 2 1 2 ~ 3 3 7 PCT/US92/08823 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:60:
Orn Ile Ala Gly Orn Ile Ala Orn Ile Ala Gly Orn Ile Ala Orn Ile Ala Gly Orn Ile Ala (2) INFORMATION FOR SEQ ID NO:61:
(i) SEQUENCE CHARACTERISTICS
~A) LENGTH: 21 amin~ acids -(B) TYPE: amino acid (C) STRANDEDNESS:
(D) TOPOLOGY: linear (ii) MOLECULE TYPE: peptide (xi) SEQUENCE DESCRIPTION: SEQ ID NO:61:
Lys Phe Ala Gly Lys Ile Ala Lys Phe Ala Gly Lys Ile Ala L~s Phe Ala ~ly Lys Ile ~
lS 20 t:~' Ala (2~ INFORMATION FOR SEQ ID NO:62:
(i) SEQUENCE ~HARACTERISTICS
~A) LENGTH: 21 amino acids (B3 ~YPE: amino a~id (C3 STRANDEDNESS:
~D) TOPOLOGY- line~r --~ii) MOLECULE TYPE: peptide WO 93/07892 PCT/US92/08~23 ~xi) SEQUENCE DESCRIPTION: SEQ ID NO:62:
Lys Ile Ala Gly Lys Phe Ala Lys Ile Ala Gly Lys Phe Ala Lys Ile Ala Gly Lys Phe .:
~:
Ala -(2) INFORMAT~ON FOR SEQ ID NO:63:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amin~ acids .:
(B) TYPE: amino acid :
(C) ST~ANDEDNESS:
(D) TOPOLOGY: linear (ii) MOLECULE TYPE: peptide (xi) SEQUENCE DESCRIPTION: SEQ ID NO:63: ::
Lys Cha Ala ~y Lys Ile Ala Lys ~ha Ala Gly Lys Ile Ala Lys Cha Ala Gly Lys Ile ~:
lS 2 Ala (2) INFORMATTON FOR SEQ TD NO:64:
~i) SEQUENCE CHARACTER~STICS
(A) LENGTH: 21 amino acids ~B) TYP~: amino acid (C) STRANDEDNESS:
~D) TOPOLOGY: linear ~ii) MOLECULE TYPE: peptide WO 93/07892 2 1 2 0 3 3 7 PCT/US9~/08823 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:64: ;
Lys Nle Ala Lys Ile Ala Gly Lys Nle Ala Lys Ile Ala Gly Lys Nle Ala Lys Ile Ala ::~
Gly (2) INFORMATION FOR SEQ ID NO:65:
(i) SEQUENCE CHARACTERISTICS:
~A) LENGTH: 21 AMINO ACI~S
(B) m E: amino acids j.
(C) STR~NDEDNE55:
(D) TOPOLOGY: linear (ii) MOLECULE TYPE: peptide (xi) SEQUENCE DESCRIPTION: SEQ ID NO:65:
Arg Ile Ala ~ly Lys Ile Ala Arg Ile Ala - 5 lO
Gly Lys Ile Ala Arg Ile Ala Gly Lys Ile :
1~ 20 Ala (2) INFORMATION FOR SEQ ID NO:66:
(i) SEQUENCE CHARACTERISTlCS
(A) LEN~TH: 21 amino acids (B) TYPE: amino acid (C) STRANDEDNESS:
(D) TOPOLOGY: linear (ii) MOLECULE TYPE: peptide ;~

W~93/07892 PCT/Us92/08823 2~ 20337 ~:

- 101 - :~

(xi) SEQUENCE DESCRIPTION: SEQ ID NO:66:
Har Ile Ala Gly Har Ile Ala Har Ile Ala :
Gly Har Ile Ala Har Ile Ala Gly Har Ile Ala (2)INFORMATION FOR SEQ ID NO:67:
(i) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 21 amino acids (B) m E: amino a~id (C) STRANDEDNESS:
(D) TOPOLOGY: linear (ii) MOLECULE TYPE: peptide (ix) FEATURE: Xaa is p-aminophenylalanine (xi) SEQUENCE DESCRIPTION: SEQ ID NO:67:
Xaa Ile Ala Gly Lys Ile Ala Xaa Ile Ala 5 1~ :
Gly Lys Ile Ala Xaa Ile Ala Gly Lys Ile 1~ ~0 Ala (2) INEORMATION FOR SEQ ID NO:6B:
(i) SEQUENCE CHARACTERISTICS
(A) ~ENGTH: Zl amino acids (B) TYP : amino acid (C) STRANDEDNESS:
(D) TOPOLOGY: linear (ii) MOLECULE TYPE: peptide WO 93/07892 2 ~ 2 ~ 3 3 7 PCT/US92/0~823 (ix~ FEATURE: Xaa is p-aminophenylalanine :~

(xi) SEQUENCE DESCRIPTION: SEQ ID NO:68:
Lys Ile Ala Gly Xaa Ile Ala Lys Ile Ala Gly Xaa Ile Ala Lys Ile Ala Gly Xaa Ile Ala (2) INFORMATION FOR SEQ ID NO:69:
SEQUENCE C~ARACTERISTICS
(A) ~NGTH: 21 amino acids .:
~B) TYPE: amino acid :~
(C) STRANDEDNESS:
(D) TOPOLOGY: linear (ii) MOLECULE TYPE: peptide ~xi) SEQUENCE DESCRIPTION: SEQ ID NO:69:
Lys Leu Ala Ser Lys Ala Gly Lys Ile Ala Gly :-Lys Ile Ala Lys Val Ala Leu Lys Alà Leu 15 20 ~:

(2) INFORMATION EOR SEQ ID NO:70:
(i) SEQUENCE C~ARACTERISTICS
(A~ LENGTH: 21 amino acids (B) TYPE: amino acid ~C) STRANDEDNESS: :~
(D) TOP~LOGY: linear (ii) MOLECULE TYPE: peptide WO 93/07892 PCI'/US92/08823 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:70:
Lys Ile Ala Gly Lys Ile Ala Lys Ile Ala Gly 5 lO j;
Orn Ile Ala Lys Ile Ala Gly Lys Ile Ala 15 2~ ~-(2) INFORMATION FOR SEQ ID NO:71:
(i) SEQUENCE CHARACTERISTICS
tA) LENGTH: 21 amino acids (B) TYPE: amino acid (C~ STRANDEDNESS:
(D) TOPOLOGY: linear ~ii) MOLECU~E TYPE: peptide ~ xi) SEQUENCE DESCRIPTION: SEQ ID NO:71:
! Lys Ile Ala Gly Lys Ile Ala Lys Ile Ala Gly Arg Ile Ala Lys Ile Ala Gly Lys Ile ~O
Ala (~) INFO~MATION ~OR SEQ ID NO:72:
(1) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amino acids (B) TYPE: amino acid (C~ STRANDEDNES5:
(D) TOPOLOGY: linear (ii) MOLECU~E TYP~: peptide WO 93/07892 2 1 7 ~ ~ 3 7 Pcr/US92/08823 ' ( xi ) SEQUENC~; DESCRIPTION: SEQ ID NO: 72:
Lys Ile Ala Gly l.ys Ile Ala Lys Ile Ala Gly Nle Ile Ala Lys Ile Ala Gly Lys lle Ala ( 2 ) INFORMATION FOR SEQ ID NO: 73:
(i) SEQUEMCE CHARACTERISTICS ;~
(A) LE:NGTH: 21 amino acids (B) TYPE: amino acid (C) STRANDEDNESS:
(D ) TOPOLOGY: linear (ii) MOLECU~E TYPE: peptide ~xi) SEQUENCE DESCRIPTI~N: SEQ ID NO:73:
Lys Ile Ala Gly Lys lle Ala Lys Ile Ala Gly Nva Ile Ala Lys Ile Ala Gly Lys Ile ~
15 20 .
Ala (2) INFORMATION FOR SEQ ID NO:74:
(i) SEQUENCE CHARACTER}STICS
~A) LE~GTH: 21 amino acids (B) TY~E: amino acid (C) STRAND D ~ESS:
(D) TOPOLOGY: linear (ii) MOLECULE TYPE: peptide (xi) SE~UENCE DESCRIPTION: SEQ ID NO:74:
Lys Phe Ala Gly Lys Phe Ala Lys Phe Ala Gly Orn Phe Ala Lys Phe Ala Gly Lys Phe Ala (2) INFORMATION FO~ SEQ ID NO:75:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amino acids (B) TYPE: amino acid (C) STRANDEDNESS:
(D) TOPOLOGY: linear (ii) MOLECULE TYPE: peptide (xi) SEQUENCE DESCRIPTION: SEQ }D NO.75:
Lys Tle Ala Gly Lys Phe Ala Lys Ile Ala Gly Orn Phe Ala Ly~ Ile Ala Gly Lys Phe Ala .

(2) INFORMATION FOR SEQ ID NO:76:
(i) SEQUENCE CHARACTERISTICS
~A) LENGT~: 21 amino acids (B) TYPE: amino acid ~C) STRANDEDNESS:
(D) TOPOLOGY: linear (ii) MOLECULE TYPE: peptide WO 93/07892 2 1 2 0 3 3 7 PCT/US92~0B823 - l o~ - , (xi) SEQUENCE DESCRIPTION: SEQ ID NO:76: :
Lys Ile Ala Gly Lys Nle Ala Lys Ile Ala Gly Orn Nle Ala Lys Ile Ala Gly Lys Nle ~:
lS 20 -;
Ala .:

~2) INFORMATION FO~ SEQ ID NO:77:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amino acid~ :
~B) TYPE: amino acid (C) STRAN~EDNESS: ;:
~D) TOPOLOGY: linear (ii) MOLECULE TYPE: peptide :~.

(xi) SEQUENCE D~SCRIP~ION: SEQ ID NO:77:
Lys Me~ Ala Ser Lys Ala Gly Lys Ile Ala :
5 1~ :~
Gly Lys Ile Ala Lys Vai Ala Leu Lys Al~
Leu (2) INFORMATION FOR SEQ ID NO:78:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amino acid3 (B3 TYPE: amino acid (C) STRANDEDNESS:
(D) TO~OLOGY: linear (ii) MOLECULE TYPE: peptide W093/n7892 PCT/US92/08~23 (xi) S~QUENCE DESCRIPTION: SEQ ID NO:78 Lys Ile Ala Ser Lys Ala Gly Lys Ile Ala Gly Lys Ile Ala Lys Val Ala Leu Lys Ala Leu 1~ 20 (2) INFORMATION FOR SEO ID NO:79:
~i) SEQUENCE CHARACTERISTICS
(A) ~ENGTH: 21 amino acid~
(B) TYPE: amino acid (C) STRANDEDNESS:
(D) TOPOLOGY: linear (ii).MOLECULE TYPE: peptide ~xi) SEQUENCE DESCRIPTION: SEQ ID NO:79:
Lys Ile Ala Ser Lys Ala Gly Lys Nle Ala Gly Lys Ile Ala Lys Val Ala Leu Lys Ala Leu ~2) INFORMATION FOR SEQ ID NO:BO:
(i) SEQUENCE CHA~ACTERI 5T I CS
tA) LEN~TH: 21 amino acids ~B) TYPE: amino acid (C) STRANDEDNES s ~D) TOPOLOGY: linear (ii) MOLECULE TYP~: peptide W093/07892 2 1 2 ~ 3 3 7 PCT/US92/08823 -108- :~

(xi) SEQUENCE D~SCRIPTION: SEQ ID NO:80:
Lys Leu Ala Ser Lys Ala Gly Lys Nle Ala Gly Lys Ile Ala Lys Val Ala Leu Lys Ala 15 20 ~:.
Leu ~.

(2) INFORMATION FOR SEQ ID NO:Bl:
(i) SEQUENCE CHARACTERISTICS
~ (A) LEN~TH: 21 amino acids (B) TYPE: amino acid (C) STRAND~DNESS:
(D) TOPOLOGY: linear (ii) MOLECULE TYPE: peptide (xi) SEQUENCE DESCRIPTION: SEQ ID NO:81: ~.
Lys Nle Ala Ser Lys Ala Gly Lys Nle Ala 5 10 ~:
~ly Lys Ile Ala ~ys Val Ala Leu Lys Ala Leu 1~ 20 `~

(2) INFORMATION FOR SEQ I~ NO:82:
(i) SEQUENCE CHARACTERISTICS:
(A~ LENGTH: 21 amino acids (B) TYPE: aminc acid (C) STRANDEDNESS:
(~) TOPOLOGY: linear (ii) MOLECULE TYPE: peptide (ix) FEATURE:
(D) OTHER INFORMATION: Xaa is p-aminophenylalanine.

~r'~ ,r~

W0 93/0789~ PCT/U592/U8823 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:~2:
Lys Ile Ala Gly Ly~ Ile Ala Lys Ile Ala Gly Xaa Ile Ala Lys Ile Ala Gly Lys Ile Ala (2) INFORMATION FOR SEQ ID NO:83:
(i) S~QUENCE CHARACTERI5TICS:
tA) LENGTH: 21 amino acids ~B) TYPE: amino acid (C) STRANDEDNESS:
(D) TOPOLOGY: linear (ii) MOLECULE TYPE: peptide ~
'`;
(xi) SEQUENCE DESCRIPTION: SEQ I~ NO:83:
~ Lys Ile Ala Gly Ala Ile Ala ~ys Ile Ala S 10 -~
Gly Lys Ile Ala Lys Ile Ala Gly Ly~ Ile Ala (2) INFORMATION FOR SEQ ID NO:84:
(i) SEQUEN OE CHARACTERISTICS: :
(A) LENGTH: 21 amino acids (B) TYPE: aminc) acid ( C ) ST~DEDNESS:
(D ~ TOPOLOGY: l inear (ii) MOLECULE TYPE: peptide WO 93/07892 2 1 ~ ~ 3 3 7 PCl`/US92/0~823 (xi ) SEQUENCE DESCRIPTION: SEQ ID NO: B4:
Lys Ile Ala Gly Lys Ile Ala Lys Ile Ala Gly Ala Ile Ala Lys Ile Ala Gly Lys Ile -Ala :

(2) INFORMATION FOR SEQ ID MO:85 ( i ) SEQUENCE C~IARACTER I ST I CS: ~
(A) LENGTH: 21 amino acids ~;
(B) TYPE: amino acid (C) STRANDEDNESS:
( D ) TOPOLOGY: l i ne ar ( ii ) MOLECULE TYPI:: peptide ~ xi ~ SEQUE;NC DE:SCRIPTION: SEO ID NO: 85:
~ Lys Ile Ala Gly Lys Ile Ala Lys Ile P,la 5 10 :
Gly Lys Ile Ala Lys Ile Ala Gly Ala Ile Ala ~2) INFORMATION EOR SEQ ID NO:86:
i ) 5EOIJENCE CHARACTERI STIC5 (A) LENGTH: 21 amino acids (B) mE: ~mino acid (C) STRANDEI)NESS:
~D) TOPOLOGY: linear (ii) MOLECULE TYPE: peptide 2 1 2 0 3 3 7 PCT/US92/~8823 ..
(xi) S~QUENCE DESCRIPTION: SEQ ID NO:96:
- Lys Ile Ala Lys Lys Ile Ala Lys Ile Ala 5 10 :~:
- Lys Lys Ile Ala Lys Ile Ala Lys Lys Ile 15 20 ~
Ala .

(2) INFORMATION FOR SEQ ID NO:87:
(i) SEQUENCE CHARACTERISTICS
(A) ~ENGTH: 21 amino acids (B) m E: amino acid (C) STRANDEDNESS:
~D) TOPOLOGY: linear (ii) MOLECULE TYPE: peptide (xi) SEQUENCE DESCRIPTION: SEQ ID NO:87:
Ala Ile Ala Gly Lys Il~ Ala ~yci Ile Ala - S 10 ~, Gly Lys Ile Ala Lys Ile Ala Gly Lys Ile :
1~ 20 ~
Ala ~-:

~2) INFORMATION FOR SEQ ID NO:8~:
(i) S~QUENCE CHARACTERISTICS
~A) LENGTH: 21 amin~ acids (B) mE amino acid (C) STRA~DEDNESSi (~) TOPO~OGY: ~inear :
(ii3 MOLECULE TYPE: peptide WO 93/07892 2 1 2 0 3 3 7 PcT~us92/o8x23 (xi) SEQUENCE DESCRIPTION: SEQ ID NO 88:
Lys Ile Ala Gly Lys Ile Ala Ala Ile Ala Gly Lys Ile Ala Lys Ile Ala Gly Lys Ile lS 20 Ala (2) INFORMATION FOR SEQ ID NO:B9:
(i) SEQUEN OE CHARACTERISTICS
- (A) LENGTH: 21 amino acids (B) TYPF.: amino acid ~C) STRANDEDNESS:
(D) TOPOLOGY: linear (ii) MOLECULE TYPE: peptide (xi) SEQUENCE DE5CRIPTION: SEQ ID NO:89:
Lys Ile Ala Gly Lys Ile Ala ~ys Ile Ala ~ 5 10 Gly Lys Ile Ala Ala Ile Ala Gly Lys Ile Ala ~2) INFORMAT1ON FOR SEQ ID NO:90:
(i~ SEQUENCE C~ARACTERISTICS
(A) LENGTH: 21 amino acids (B) TYPE: amino acid ~C1 STRANDEDNESS:
~D) TOPOLOGY. linear (ii) MOLECULE TYPE: peptide 2 ~ ~ ~ 3 ~ 7 PCT/~592/08~3 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:9~:
Gly Met Ala Ser Lys Ala Gly Lys Ile Ala Gly Lys Ile Ala Lys Val Ala Leu Lys Ala Leu (2) INFORMATION FOR SEQ ID NO:91:
~i) SE~UENCE CHARACTERISTICS
~ (A) LE~G~H: 11 amino acids ~B) TYPE: ami~o ~cid (C) STRANDEDNESS:
(D) TOPOLOGY: linear (ii) MOLECULE TYPE: peptide ~:.
(ix~ FEATURE~
(D) OTH~R INFORMATION: May be a C-terminal amide, and/or may be acetylated at N-terminus. :
.:
(xi~ SEQUENCE DESCRIPTION:SEQ ID NO~
Leu Lys Lys Leu Lys Lys Leu Leu Lys Leu 5 10 -~.
Leu (2) INFORMATION ~OR SEQ ID NO:92: ~-(i) SEQUENCE C~ARACTERISTICS
(A) ~ENGT~: 12 amino acids (B) TYPE: amino acid (C) STRANDEDN~SS:
(D) TOPOLOGY: linear (ii) MOLECULE TYPE: peptide WO 93/07892 2 1 2 ~ 3 3 7 PCT/US92/08823 (ix) FEATURE:
(D~ OTHER INFORMATION: May be a C-terminal amide, and/or may be acetylated at N-terminus.

(xi) SEQUENCE DESCRIPTION:SEQ ID NO:92:
Leu Leu Lys Lys Leu Lys Lys Leu Leu Lys Leu Leu a (2) INFORMATION FOR SEQ ID NO:a3:
(i) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 13 amino acids :.
(B) TYPE: amino acid (C) 5TRANDE9NESS:
(D) TOPOLOGY: linear (ii) MOLECULE TYPE: peptide (ix) FEATURE:
(D) OTHER INFORMATION: May be a C-terminal amide, and/or may be acetylated at N-terminus.

(xi) SEQUENCE DESCRIPTION:SEQ ID NO:93:

Lys Leu t~eU ~ys Lys Leu Lys Lys Leu Leu Lys Leu Leu (2) INFORMATION FOR SEQ ID NO:94:
(i) SE~UENCE CHARACTERISTICS:
(A) LENGTH: 14 amino acids (B) TYPE: amino a~id -llS-(C) STRANDED~ESS:
(D) TOPOLOGY: linear (ii) MOLECULE TYPE: peptide (ix) FEATURE:
(D) OTHER INFORMATION: May be a C-terminal amide, and/or may be acetylated at N-terminus.

(xi) SEQUENCE DESCRIPTION:SEQ ID NO:94:

Lys Lys Leu Leu Lys Lys L~u Lys Lys Leu Leu Lys Leu Leu ,,.

(2) INFORMATION FOR SEQ ID NO:95:
(i) SEQU~NCE CHARACTERISTICS:
(A) LENGTH: 16 amino acids (B) TYPE: amino acid .
(C) STRANDEDNESS:
(D) TOPOLOGY: linear -(ii) MOLECULE TYP : peptide (ix) F~ATURE:
(D) OTHER INFORMAT$0N: May be a C-terminal amide, and/or may be acetylated at N-term~nu~.
(ix) SEQUENCE DESCRIPTION: SEQ ID MO:95~
Lys Lys Leu L~u Lys Lys Leu Lys LYB Leu Leu Lys Ly~ Leu Arg Arg ~ 10 lS

(2) INFORMATION FOR SEQ ID NO:96:
(i) SEQUENCE CHARACTERISTICs:
(A) LENCTH: 16 amino acids W093/07892 2 1 2 0 3 3 ~ PCT/US~2/08823 -l16-.:
(B) TYPE: amino acid (C) STRANDEDNESS:
(D) TOPOLOGY: linear (ii) MOLECULE TYPE: peptide (ix) FEATURE:
~D) OT~ER INFORMATION: May be a C-t~rminal amide, andfor may be acetylated at N-terminu~.

~xi~ SEQUENCE DESCRIPTION:SEQ ID NO:96:

Lys Leu Lys Lys Leu Leu Lys Lys Leu Lys Lys Leu Leu 1ys Leu Leu (2) INFORMATION FOR SEQ ID NO:97:
(i) SEQUNCE CHARACTERISTIGS
(A) LENGTH: 22 amino acids (B) TYP~: amino acid (C) STRANDEDNESS:
(D) TOPOLOGY: linear (ii~ MOLECULE TYPE peptide (xi) SEQUENCE ~ESCRIPTION: SEQ ID NO:97:
Gly Ile Gly Lys Phe Leu His Ser Ala Gly Lys Phe Gly Lys Ala Phe Val Lys Ile Met Lys Ser W093/07892 PCT~S9~/08823 2~21)337 -117- . :~

(2) INFORMATION FOR SEQ ID NO:98:
(i) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 24 amino acids (B) TYPE: amino acid -.
(C) STRANDEDNESS:
(D) TOPOLOGY: linear (ii) MOLECULE m E: peptide ~xi) SEQUEN OE DESCRIPTION: SEQ ID NO:98:
Ala Gly Ile Gly Lys Phe Le~ His Ala Ala Lys Lys Phe Ala Lys Ala Phe Val Ala Glu 15 20 ~:
Ile Met Asn Ser (2) INFORMATlON FOR SEQ ID NO:99:
~- (i) SEQUE~C~ CHARACTERISTICS:
(A) LENGTH: 24 amino acids (B~ TYPE: amino acid ~C) STRANDED~SS: ~:
(D) TOPOLOGY: linear ~ MOLECULE TYPE: peptide ~`

(xi~ SEQUENCE DESCRIPTION: SEQ ID NO:99:
Ala Leu Ser Lys Ala Leu Ser Lys Ala Leu Ser Lys Ala Leu S~r Lys Ala Leu Ser Lys ~5 20 Ala Leu Ser Lys (2) INFORMATION FOR SEQ ID NO:100:
(i) SEQUENCE CHARACTERIST~CS:

WO 93/07892 2 1 2 0 3 3 7 p~/U~i92/08~23 ~ 118-(A) LENGTH: 16 am~no acids (B) TYPF~: amino acid ( C ) STRANDEDNESS:
(D) TOPOLOGY: linear ( ii ) MOLECULE ~FYPE: peptide (xi ) SEQUENCE DESCRIPTION: SEQ ID NO: lOO

Gly Ile Lys Lys Phe Leu Lys Lys Ala Gly Lys Phe Gly Lys Ala Phe ,.

Claims

WHAT IS CLAIMED IS

1. A process of inhibiting growth of a target cell in a host, comprising:
administering to a host at least one biologically active amphiphilic peptide or protein, said peptide or protein being an ion channel-forming peptide or protein; and an antibiotic which is not an ion channel-forming peptide or protein, said biologically active amphiphilic peptide or protein and said antibiotic being administered in a combined amount effective to inhibit growth of a target cell in a host.

2. The process of Claim 1 wherein said antibiotic is selected from the class consisting of a tetracycline; a pseudomonic acid; benzoyl peroxide; neomycin; viomycin; a cephalosporin; rifampin; ethambutol; streptomycin; isoniazid;
ethionamide; a sulfonaide; trimethoprin; cycloserine; a nitroimidazole; and a penem antibiotic.

3. The process of Claim 2 wherein said antibiotic is a cephalosporin.

4. The process of Claim 3 wherein said cephalosporin is selected from the group consisting of cephalothin, cephalexin, cefazolin, cephradine, cephapirin, cefamandole, cefoxitin, cefoperazone, cefotaxime, moxalactam, and 3' quaternary ammonium cephalosporins.

5. The process of Claim 4 wherein the cephalosporin is cefotaxime.

6. The process of Claim 2 wherein said antibiotic is a tetracycline.

7. The process of Claim 2 wherein said antibiotic is a sulfonamide.

8. The process of Claim 7 wherein said sulfonamide is selected from the class consisting of sulfanilamide, sulfadiazine, sulfamethoxazole, sulfisoxazole, sulfamethoxazole, and sulfathalide.

9. The process of Claim 1 wherein the peptide is a basic polypeptide having at least sixteen amino acids, wherein said basic polypeptide includes at least eight hydrophobic amino acids and at least eight hydrophilic amino acids.

10. The process of Claim 1 wherein the peptide is a magainin peptide.

11. The process of Claim 1 wherein the peptide is an XPF
peptide.

12. The process of Claim 1 wherein the peptide is a PGLa peptide.

13. The process of Claim 1 wherein the peptide is a CPF
peptide.

14. The process of Claim 1 wherein the peptide includes one of the following basic structures X31 through X37, wherein:
X31 is -[R31-R32-R32-R33-R31-R32-R32]n-;
X32 is -[R32-R32-R33-R31-R32-R32-R31]n-;
X33 is -[R32-R33-R31-R32-R32-R31-R32]n-;
X34 is -[R33-R31-R32-R32-R31-R32-R32]n-;
X35 is -[R31-R32-R32-R31-R32-R32-R33]n-;
X36 is -[R32-R32-R31-R32-R32-R33-R31]n-; and X37 is -[R32-R31-R32-R32-R33-R31-R32]n-, wherien R31 is a basic hydrophilic amino acid, R32 is a hydrophobic amino acid, R33 is a neutral hydrophilic or hydrophobic amino acid, and n is from 2 to 5.

15. The process of Claim 1 wherein the peptide includes the following basic structure X40:
R31-R32-R32-R33-R34-R32-R32-R31-R32-R32-R34-R32-R32, wherein R31 is a basic hydrophilic amino acid, R32 is a hydrophobic amino acid, and R33 is a neutral hydrophilic, hydrophobic or basic hydrophilic amino acid, and R34 is a basic hydrophilic or hydrophobic amino acid.

16. The process of Claim 1 wherein the peptide includes the following basic structure X50:
R41-R42-R42-R41-R42-R42-R41-R41-R42-R41-R41, wherein R41 is a hydrophobic amino acid, and R42 is a basic hydrophilic or neutral hydrophilic amino acid.

17. The process of Claim 1 wherein the peptide includes the following basic structure X52:
R42-R41-R42-R42-R41-R41-R42-R42-R41-R42-R42, wherein R41 is a hydrophobic amino acid, and R42 is a basic hydrophilic or neutral hydrophilic amino acid.

18. A composition comprising:
(a) at least one biologically active amphiphilic peptide or protein, said peptide or protein being an ion channel-forming peptide or protein and (b) an antibiotic which is not an ion channel-forming peptide or protein, and derivatives and analogues thereof.

19. The composition of Claim 18 wherein said components (a) and (b) are present in a combined amount effective to inhibit growth of a target cell.

20. The composition of Claim 19 wherein said antibiotic is selected from the class consisting of a tetracycline; a pseudomonic acid; benzoyl peroxide; neomycin; viomycin; a cephalosporin; rifampin; ethambutol; streptomycin; isoniazid;
ethionamide; a sulfomamide; trimethoprim; cycloserine; a nitroimidazole, and a penem antibiotic.

21. The composition of Claim 20 wherein the non-peptide antibiotic is a cephalosporin.

22. The composition of Claim 21 wherein said cephalosporin is selected from the group consisting of cephalothin, cephalexin, cefazolin, cephradine, cephapirin, cefamandole, cefoxitin, cefoperazone, cefotaxime, moxalactam, and 3' quaternary ammonium cephalosporins.

23. The composition of Claim 22 wherein the cephalosporin is cefotaxime.

24. The composition of Claim 20 wherein said antibiotic is a tetracycline.

25. The composition of Claim 20 wherein said antibiotic is a sulfonamide.

26. The composition of Claim 25 wherein said sulfonamide is selected from the class consisting of sulfanilamide, sulfadiazine, sulfamethoxazole, sulfisoxazole, and sulfathalidine.

27. The composition of Claim 18 wherein the peptide is a basic polypeptide having at least sixteen amino acids, wherein said basic polypeptide includes at least eight hydrophobic amino acids and at least eight hydrophilic amino acids.

28. The composition of Claim 18 wherein the peptide is a magainin peptide.

29. The composition of Claim 18 wherein the peptide is an XPF peptide.

30. The composition of Claim 18 wherein the peptide is a PGLa peptide.

31. The composition of Claim 18 wherein the peptide is a CPF peptide.

32. The composition of Claim 18 wherein the peptide includes one of the following basic structures X31 through X37, wherein:
X31 is -[R31-R32-R32-R33-R31-R32-R32]n-;
X32 is -[R32-R32-R33-R31-R32-R32-R31]n-;
X33 is -[R32-R33-R31-R32-R32-R31-R32]n-;
X34 is -[R33-R31-R32-R32-R31-R32-R32]n-;
X35 is -[R31-R32-R32-R31-R32-R32-R33]n-;
X36 is -[R32-R32-R31-R32-R32-R33-R31]n-; and X37 is -[R32-R31-R32-R32-R33-R31-R32]n-, wherein R3? is a basic hydrophilic amino acid, R32 is a hydrophobic amino acid, R33 is a neutral hydrophilic or hydrophobic amino acid, and n is from 2 to 5.

33. The composition of Claim 18 wherein the peptide includes the following basic structure X40:

R31-R32-R32-R33-R34-R32-R43-R31-R32-R32-R32-R34-R32-R32, wherein R31 is a basic hydrophilic amino acid, R32 is a hydrophobic amino acid, and R33 is a neutral hydrophilic, hydrophobic, or basic hydrophilic amino acid, and R34 is a basic hydrophilic or hydrophobic amino acid.

34. The composition of Claim 18 wherein the peptide includes the following basic structure X50:
R41-R42-R42-R41-R42-R42-R41-R41-R42-R41-R41, wherein R41 is a hydrophobic amino acid, and R42 is a basic hydrophilic or neutral hydrophilic amino acid.

35. The composition of Claim 18 wherein the peptide includes the following basic structure X52:
R42-R41-R42-R42-R41-R41-R42-R42-R41-R42-R42, wherein R41 is a hydrophilic amino acid.

36. The process of Claim 1 wherein each of said peptide and said antibiotic is administered in an amount ineffective in inhibiting growth of a target cell in a host if administered alone to a host.

37. The process of Claim 1 wherein the peptide includes the following structure formula:
-(Lys Ile Ala Lys Ile Ala)n-, wherein n is from 2 to 5.

38. The composition of Claim 18 wherein the peptide includes the following structural formula:
-(Lys Ile Ala Lys Lys Ile Ala)n-, wherein n is from 2 to 5.