U.S. patent application number 10/103859 was filed with the patent office on 2002-10-24 for method of treating sepsis and ards using chemokine beta-9.
This patent application is currently assigned to Smithkline Beecham Corporation. Invention is credited to Pelus, Louis M., White, John R..
Application Number | 20020155094 10/103859 |
Document ID | / |
Family ID | 21785003 |
Filed Date | 2002-10-24 |
United States Patent
Application |
20020155094 |
Kind Code |
A1 |
White, John R. ; et
al. |
October 24, 2002 |
Method of treating sepsis and ARDS using chemokine beta-9
Abstract
The invention relates to the method of preventing and treating
sepsis and ARDS using chemokine or biologically active fragment
thereof, alone or in conjunction with an anti-infective agent.
Inventors: |
White, John R.;
(Coatesville, PA) ; Pelus, Louis M.; (Richboro,
PA) |
Correspondence
Address: |
STERNE, KESSLER, GOLDSTEIN & FOX P.L.L.C.
1100 NEW YORK AVENUE, N.W.
SUITE 600
WASHINGTON
DC
20005-3934
US
|
Assignee: |
Smithkline Beecham
Corporation
|
Family ID: |
21785003 |
Appl. No.: |
10/103859 |
Filed: |
March 25, 2002 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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10103859 |
Mar 25, 2002 |
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09496273 |
Feb 1, 2000 |
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09496273 |
Feb 1, 2000 |
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08852212 |
May 6, 1997 |
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6290948 |
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60017871 |
May 14, 1996 |
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Current U.S.
Class: |
424/85.1 ;
514/200 |
Current CPC
Class: |
A61K 31/175 20130101;
A61P 31/00 20180101; Y10S 514/921 20130101; A61K 38/195 20130101;
A61P 11/00 20180101; Y10S 514/885 20130101; A61K 31/546 20130101;
A61K 38/195 20130101; A61P 31/04 20180101; A61K 31/175 20130101;
A61P 43/00 20180101; A61K 31/7036 20130101; A61K 31/546 20130101;
A61K 31/7036 20130101; A61K 2300/00 20130101; A61K 2300/00
20130101; A61K 2300/00 20130101; A61K 2300/00 20130101 |
Class at
Publication: |
424/85.1 ;
514/200 |
International
Class: |
A61K 038/19; A61K
031/545 |
Claims
What is claimed is:
1. A method of treating sepsis and ARDS comprising administering to
an animal in need thereof an effective amount of chemokine selected
from the group consisting of: (a) Macrophage Inflammatory
Protein-Gamma; (b) Macrophage Inflammatory Protein-3 and -4; (c)
Macrophage Migration Inhibitory Factor-3; (d) Human Chemokine
Beta-9; (e) Human Chemokine Polypeptides; (f) Human Chemokine
Beta-11 and Human Chemokine Alpha-1; (g) Human Chemokine Beta-13;
(h) Human Chemokine Beta-12; (i) Chemokine Alpha-2; (j) Chemokine
Alpha-3; (k) Novel Chemokine for Mobilizing Stem Cells; (l) Short
Form Chemokine Beta-8; or biologically active fragments
thereof.
2. A method according to claim 1 wherein said effective amount is
from about 1 to about 100 mg/kg/dose.
3. The method according to claim 1 wherein chemokine is
administered orally.
4. The method according to claim 1 wherein chemokine is
administered subcutaneously.
5. A method of treating sepsis and ARDS comprising administering to
an animal in need thereof an effective amount of chemokine or
biologically active fragments thereof in conjunction with an
effective amount of an anti-infective agent.
6. A method according to claim 5 wherein the anti-infective agent
is selected from the group consisting of gentamicin, augmentin or
ceftazidime.
7. A method for the prevention of sepsis and ARDS comprising
administering to an animal in need thereof an effective amount of
chemokine or biologically active fragment thereof.
8. A method according to claim 7 wherein the effective amount is
from about 1 to about 100 mg/kg/dose.
9. The method according to claim 7 wherein cheomkine or
biologically active fragment thereof is administered 1 to 2 days
prior to surgery.
10. A method for the prevention of sepsis and ARDS comprising
administering to an animal in need thereof an effective amount of
chemokine or biologically active fragment thereof, in conjunction
with an effective amount of an anti-infective agent.
11. A method according to claim 10 wherein the anti-infective agent
is selected from the group consisting of gentamicin, augmentin or
ceftazidime.
12. A method for the treatment of sepsis and ARDS comprising
administering to an animal in need thereof an effective amount of
chemokine or biologically active fragment thereof.
13. A method according to claim 12 wherein the effective amount is
from about 1 to about 100 mg/kg/dose.
14. The method according to claim 12 wherein chemokine or
biologically active fragment thereof is administered 2 hours to 24
hours after surgery.
15. A method for the treatment of sepsis and ARDS comprising
administering to an animal in need the an effective amount of
chemokine or biologically active fragment thereof, in conjunction
with an effective amount of an anti-infective agent.
16. A method according to claim 15 wherein the anti-infective agent
is selected from the group consisting of gentamicin, augmentin or
ceftazidime.
Description
RELATED APPLICATION
[0001] This application claims the benefit of U.S. Provisional
Application No. 60/017,871 filed May 14, 1996.
FIELD OF INVENTION
[0002] This invention relates to the method of preventing and
treating sepsis and adult respiratory distress syndrome using
certain chemokines or biologically active fragments thereof alone
or in conjunction with an anti-infective agent or hematopoietic
maturing agent.
BACKGROUND OF INVENTION
[0003] Sepsis, as used herein, is broadly defined to mean
situations when the invasion of a host by a microbial agent is
associated with the clinical manifestations of infection including
but not limited to:
[0004] (1) temperature >38.degree. C. or <36.degree. C.; (2)
heart rate >90 beats per minute; (3) respiratory rate >20
breaths per minute or PaCO.sub.2 <32 mm Hg; (4) white blood cell
count >12,000/cu mm, <4,000/cu mm, or >10% immature (band)
forms; (5) organ dysfunction, hypoperfusion, or hypotension.
Hypoperfusion and perfusion abnormalities may include, but are not
limited to lactic acidosis, oliguria, or an acute alteration in
mental states. (Chest 1992; 101: 1644-1566)
[0005] Sepsis can occur in hospitalized patients having underlying
diseases or conditions that render them susceptible to bloodstream
invasion or in bum, trama or surgical patents. In many cases of
sepsis, the predominant pathogen is Escherichia coli, followed by
other Gram-negative bacteria such as the
Klebsiella-Enterobacter-Serratia group and then Pseudomonas.
Although comprising a somewhat smaller percentage of infection,
Gram-positive microbes such as Staphylococcus and systemic viral
and fungal infections are included by the term sepsis as used
herein. The genitourinary tract is the most common site of
infection, the gastrointestinal tract and respiratory tract being
the next most frequent sources of sepsis. Other common foci are
wound, burn, and pelvic infections and infected intravenous
catheters.
[0006] A serious consequence of bacterial sepsis often is septic
shock. Septic shock is characterized by inadequate tissue
perfusion, leading to insufficient oxygen supply to tissues,
hypotension and olgiuria.
[0007] Septic shock occurs because bacterial products react with
cells and components of the coagulation, complement, fibrinolytic
and bradykinin systems to release proteases which injure cells and
alter blood flow, especially in the capillaries.
[0008] Microorganisms frequently activate the classical complement
pathway, and endotoxin activates the alternative pathway.
Complement activation, leukotriene generation and the direct
effects of bacterial products on neutrophils lead to accumulation
of these inflammatory cells in the lungs, release of their
proteolytic enzymes and toxic oxygen radicals which damage the
pulmonary endothelium and initiate the adult respiratory distress
syndrome ("ARDS"). ARDS is a major cause of death in patients with
septic shock and is characterized by pulmonary congestion,
granulocyte aggregation, haemorrhage and capillary thrombi.
[0009] Septic shock is a major cause of death in intensive care
units. There are an estimated 200,000 cases per year of septic
shock in the United States, and despite advances in technology
(i.e., respiratory support) and antibiotic therapy, the mortality
rate for septic shock remains in excess of 40%. In fact, mortality
for established septic shock has decreased very little since the
comprehensive description by Waisbren (Arch. Intern. Med.
88:467-488 (1951)). Although effective antibiotics are available,
and there is an increased awareness of the septic shock syndrome,
the incidence of septic shock over the last several decades has
actually increased. With the appreciation that antimicrobial agents
have failed to completely abrogate septic mortality, it is clear
that other agents must be developed to be used alone or in
conjunction with antimicrobials in order to rectify the
deficiencies of current established therapy.
BRIEF DESCRIPTION OF THE INVENTION
[0010] This invention relates to a method of preventing or treating
sepsis and ARDS comprising administering to an animal, including
humans, in need thereof an effective amount of chemokine protein or
biologically active fragments thereof.
[0011] This invention further relates to a method of preventing or
treating sepsis and ARDS comprising administering to an animal
(including humans) in need thereof an effective amount of chemokine
protein or biologically active fragments thereof, either before, in
conjunction with or after an anti-infective agent.
DETAILED DESCRIPTION OF THE INVENTION
[0012] It is the object of this invention to provide a new method
of treatment of sepsis and ARDS comprising administering to an
animal in need thereof, including humans, an effective amoudt of
chemokine protein or biologically active fragments thereof, alone
or in combination with other anti-infective agents. As used herein
the term "chemokine" means those polypeptides claimed in the patent
applications set forth in Table 1.
1TABLE 1 Chemokine Patent Applications Application Gene Name Date
Filed Number Macrophage Inflammatory Protein-Gamma 22-Dec-93
08/173,209 Macrophage Inflammatory Protein-3 and -4 8-Mar-84
WO95/17092 Macrophage Migration Inhibitory Factor-3 16-May-
WO95/31468 94 Human Chemokine Beta-9 6-Jun-95 WO96/06169 Human
Chemokine Polypeptides 23-Aug-94 WO96/05856 Human Chemokine Beta-11
and Human Chemokine Alpha-1 8-Feb-95 US95/01780 Human Chemokine
Beta-13 5-Jun-95 08/464,594 Human Chemokine Beta-12 6-Jun-95
08/468,541 Chemokine Alpha-2 19-Mar-96 60/013,653 Chemokine Alpha-3
18-Mar-96 US96/03686 Novel Chemokine for Mobilizing Stem Cells
29-Sep-95 60/006,051 Short Form Chemokine Beta-8 24-Oct-95
60/004,517
[0013] This invention further relates to a method of preventing
sepsis and ARDS comprising administering to an animal in need
thereof an effective amount of modified chemokine protein or
biologically active fragments thereof alone or in combination with
other anti-infective agents.
[0014] Known anti-infective agents include, without limitation,
anti-microbial agents routinely used for the treatment of sepsis
such as amino-glycosides (such as amikacin, tobramycin, netilmicin,
and gentamicin), cephalosporins such as ceftazidime, related
beta-lactam agents such as maxalactam, carbopenems such as
imipenem, monobactam agents such as aztreonam; ampicillin and
broad-spectrum penicillins, (e.g., penicillinase-resistant
penicillins, ureidopenicillins or antipseudomonal penicillin or
Augmentin) that are active against P. aeruginosa, Enterobacter
species, indole-positive Proteus species, and Serratia. Also
included within the definition of anti-infective agents are
antifungal agents, amphotericin and the like as well as anti-viral
agents such as famvir and acyclovir.
[0015] The compound is useful in the treatment and prevention of
sepsis and ARDS in humans and other animals such as dairy cattle,
horses, calves or poultry.
[0016] Chemokine protein or biologically active fragments thereof
have been described in the applications in Table 1, incorporated by
reference herein. The use of chemokine protein or biologically
active fragments thereof for the prevention and treatment of sepsis
has not been reported. It has now been discovered that chemokine
protein or biologically active fragments thereof significantly
increases the survival of animals challenged with lethal sepsis
causing organisms. Treatment with the compound of this invention,
alone or in combination with an anti-infective agent prior to
contemplated thoracic or abdominal surgery would be useful in
reducing the likelihood of post-operative sepsis. It may also be
used post-operatively for the treatment of sepsis and ARDS caused
by a variety of reasons as outlined previously.
[0017] To effectively treat a human or other animal chemokine
protein or biologically active fragments thereof may be
administered by injection in the dose range of about 10 fg/kg to
about 100 mg/kg/dose, preferably between about 1 and 50 mg/kg/dose,
or orally in the dose range of about 10 fg/kg to about 100 mg/kg
body weight per dose, preferably between about 1 and 50 mg/kg body
weight; if administered by infusion or similar techniques, the dose
may be in the range of about 10 fg/kg to about 100 mg/kg/dose,
preferably between about 1 and 50 mg/kg/dose; if administered
subcutaneously the dose may be in the range of about 10 fg/kg to
about 100 mg/kg/dose, preferably between about 1 and 50
mg/kg/dose.
[0018] Depending on the patient's condition, the compounds of this
invention can be administered for prophylactic and/or therapeutic
treatments. In therapeutic application, the compound is
administered to a patient already suffering from a disease in an
amount sufficient to cure or at least partially arrest the disease
and its complications. It may be given at any time after surgery,
preferably prior to 24 hours after surgery. In prophylactic
applications, a composition containing chemokine protein or
biologically active fragments thereof, is administered to a patient
not already in a disease state to enhance the patient's resistance.
It may be given one day or one week prior to surgery, preferably
one to two days prior to surgery. It may be administered
parenterally or orally.
[0019] Single or multiple administrations of the compounds can be
carried out with dose levels and pattern being selected by the
treating physician. In any event, a quantity of the compounds of
the invention sufficient to effectively treat the patient should be
administered.
[0020] The compounds of this invention, may also be administered in
conjunction with a conventional anti-infective as disclosed herein
above, such a gentamicin, augmentin or ceftazidime. The particular
anti-infective chosen should be one to which the infective organism
is susceptible and is selected or modified during therapy as the
infecting microrganism is more particularly identified.
[0021] Additionally, various adjunctive agents in the treatment of
septic shock also may be useful in combination with the components
of this invention. They include sympathomimetic amines
(vasopressors) such as norepinephrine, epinephrine, isoproterenol,
dopamine, and dobutamine; anti-inflammatory agents such as
methylprednisolone anti-inflammatory agents such as indomethacin
and phenylbutazone; and corticosteroids such as betamethasone,
hydrocortisone, methylprednisolone, or dexamethasone;
anticoagulants such as heparin, anti-thrombin III or coumarin type
drugs for certain conditions and schedules; diuretics such as
furosemide or ethacrynic acid; and antagonist of opiates and
beta-endorphins such as naloxone; an antagonist of tumor necrosis
factor or of interleukin-1; phenothiazines; anti-histamines;
glucagon; .alpha.-adrenergic blocking agents, vasodilators; plasma
expanders; packed red blood cells; platelets; cryoprecipitates;
fresh frozen plasma; bacterial permeability protein; clindamycin;
and antibodies to (lipid A), the J5 mutant of E. coli or to
endotoxin core glycolipids. Methods for preparing such antibodies
are described widely in the literature.
[0022] One of the most important aspects in the treatment of the
clinical septic shock syndrome is its apparently intractable
resistance to the effects of a variety of highly potent
antimicrobial agents. Despite the development of newer
antimicrobial agents, the overall incidence of clinical sepsis has
increased, and mortality remains unacceptably high, often
approaching 60% of diagnosed patients. The discovery of the
increased survival with the treatment of chemokine protein or
biologically active fragments thereof both prophylactically and
after infection provides a new and usefull therapy of sepsis and
ARDS.
[0023] The compounds of this invention, may also be administered in
conjunction with hematopoietic maturation agents, such as G-CSF,
Flt3, M-CSF or GM-CSF. These compounds affect the mobilization of
the chemokines of the invention and are believed to enhance the
anti-sepsis and anti-ARDS efficacy of chemokines.
[0024] The biological activity of chemokine protein or biologically
active fragments thereof are demonstrated by the following
assays:
[0025] Rats. Male Fischer 344 rats obtained from Taconic farms
weighing 200 to 250 g. are utilized. The rats are housed 2 per cage
in standard plastic caging and fed lab chow and water ad
libitum.
[0026] Chemokine protein or biologically active fragments thereof,
is prepared in E. coli by the method given in Example 5. The
compound is dissolved in DPBS containing 0.5% heat inactivated
autologous normal rat serum. The animals are dosed
intraperitoneally with chemokine 24 hours and 2 hours before
infection. Control animals are dosed with dilution buffer on the
same schedule. Starting two hours after infection the rats are
treated twice daily with subcutaneous gentamicin. E. coli. A
clinical isolate of E. coli isolated from sputum is utilized. The
organisms are tested for antibiotic sensitivity by the disc-agar
diffusion technique and found to be sensitive to gentamicin,
ampicillin, cephalothin, chloramphenicol, kanamycin, tetracycline,
trimethoprin/sulfamethoxazole and resistant to penicillin G,
erythromycin, and vancomycin. The organism is animal passed in mice
and subsequently recovered and plated onto MacConkey's agar. The
reisolated organisms are grown overnight in brain-heart infusion
broth, and then stored frozen at 70.degree. C. The inoculate the
fibrin clot, organisms from thawed stocks are inoculated into
brainheart infusion broth and incubated overnight on a rotary
shaker (120 rpm) an 37.degree. C. The E. coli is harvested by
centrifugation, washed 3X and finally resuspended in normal saline.
The number or organisms is quantified by turbidimentry, and the
concentration adjusted with normal saline. All inoculum sizes are
based on viable counts determined by scoring colony forming units
on MacConkeys agar.
[0027] Fibrin Clot. The E. coli infected fibrin clots are made from
a 1% solution of bovine fibrinogen (Type 1-S, Sigma) in sterile
saline. The clot is formed by adding sequentially human thrombin
(Hanna Pharma.) bacteria, and fibrinogen solution to 24 well
plastic plates. Bacterial numbers of 2.0 to 3.0.times.10.sup.9 are
used in inoculate the fibrin clots. The resulting mixture is then
incubated at room temperature for 30 minutes before
implantation.
[0028] Animal Model. The rats are anethetized with
ketamine/xylazine (40 mg/kg/5 mg/kg) then the abdominal surfaced is
shaved and a midline laporatomy performed. Bacterial peritonitis is
induced by implanting a fibrin-thrombin clot containing E. coli
into the abdominal cavity. After implantation the muscle layers are
closed with 4-0 silk suture, and the wound closed with surgical
staples. The animals are closely observed, any animals obviously
moribound are euthanized.
[0029] Gentamicin. Rats are treated subcutaneously with gentamicin
sulfate (Elkins-Sinn, NJ) 5 mg/kg twice a day for five days.
[0030] Statistics. All continuously variable data are expressed as
the percent survival from several pooled studies. The Fisher's
Exact test is used to determine the statistical significance of the
differences between the survival rates at 14 days. The differences
between the groups are considered statistically significant at
p<0.05.
EXAMPLE 1
[0031] Prophylactically Administered Chemokine
[0032] Chemokine proteins may be prepared using known methods for
protein purification or as described in the patent applications
listed in Table 1.
[0033] The animals are dosed intraperitoneally with chemokine at
doses of 10, 100 and 1,000 fg/kg, and 10 and 100 mg/kg 24 hours and
2 hours before infection. Control animals are dosed with dilution
buffer on the same schedule. Starting two hours after infection the
rats are treated twice daily with subcutaneous gentamicin. On day 0
the rats are implanted with an E. coli containing fibrin-thrombin
clot. Starting two hours after infection the rats are treated with
gentamicin twice daily. The rats prophylactically treated with
chemokine at 33 or 100 fg/kg followed by gentamicin treatment
demonstrated significantly improved survival rates over the diluent
treated control rat receiving gentamicin therapy alone.
EXAMPLE 2
[0034] Theraputically Administered Chemokine
[0035] On day 0 the rats are implanted with an E. coli containing
fibrin-thrombin clot. The animals are dosed intraperitoneally with
chemokine at doses of 10, 100 and 1,000 fg/kg, and 10 and 100 mg/kg
as a single injection 2 hours after infection. Control animals are
dosed with dilution buffer on the same schedule. Starting two hours
after infection the rats are treated twice daily with subcutaneous
gentamicin. The rats theraputically treated with chemokine at 100
or 333 fg/kg followed by gentamicin treatment are assessed for
improved survival rates over the diluent treated control rat
receiving gentamicin therapy alone.
EXAMPLE 3
[0036] Therapeutically Administered Chemokine in S. aureus
sepsis:
[0037] On day 0 the rats are implanted with a S. aureus containing
fibrin-thrombin clot. The animals are dosed intraperitoneally with
chemokine at doses of 10, 100 and 1,000 fg/kg, and 10 and 100 mg/kg
as a single injection 2 hours after infection. Control animals are
dosed with dilution buffer on the same schedule. Starting two hours
after infection the rats are treated twice daily with subcutaneous
gentamicin. The rats theraputically treated with chemokine followed
by gentamicin treatment are assessed for improved survival rates
over the diluent treated control rat receiving gentamicin therapy
alone.
Sequence CWU 1
1
2 1 405 DNA Homo sapiens CDS (1)..(405) 1 atg gct cag tca ctg gct
ctg agc ctc ctt atc ctg gtt ctg gcc ttt 48 Met Ala Gln Ser Leu Ala
Leu Ser Leu Leu Ile Leu Val Leu Ala Phe 1 5 10 15 ggc atc ccc agg
acc caa ggc agt gat gga ggg gct cag gac tgt tgc 96 Gly Ile Pro Arg
Thr Gln Gly Ser Asp Gly Gly Ala Gln Asp Cys Cys 20 25 30 ctc aag
tac agc caa agg aag att ccc gcc aag gtt gtc cgc agc tac 144 Leu Lys
Tyr Ser Gln Arg Lys Ile Pro Ala Lys Val Val Arg Ser Tyr 35 40 45
cgg aag cag gaa cca agc tta ggc tgc tcc atc cca gct atc ctg ttc 192
Arg Lys Gln Glu Pro Ser Leu Gly Cys Ser Ile Pro Ala Ile Leu Phe 50
55 60 ttg ccc cgc aag cgc tct cag gca gag cta tgt gca gac cca aag
gag 240 Leu Pro Arg Lys Arg Ser Gln Ala Glu Leu Cys Ala Asp Pro Lys
Glu 65 70 75 80 ctc tgg gtg cag cag ctg atg cag cat ctg gac aag aca
cca tcc cca 288 Leu Trp Val Gln Gln Leu Met Gln His Leu Asp Lys Thr
Pro Ser Pro 85 90 95 cag aaa cca gcc cag ggc tgc agg aag gac agg
ggg gcc tcc aag act 336 Gln Lys Pro Ala Gln Gly Cys Arg Lys Asp Arg
Gly Ala Ser Lys Thr 100 105 110 ggc aag aaa gga aag ggc tcc aaa ggc
tgc aag agg act gag cgg tca 384 Gly Lys Lys Gly Lys Gly Ser Lys Gly
Cys Lys Arg Thr Glu Arg Ser 115 120 125 cag acc cct aaa ggg cca tag
405 Gln Thr Pro Lys Gly Pro 130 2 134 PRT Homo sapiens 2 Met Ala
Gln Ser Leu Ala Leu Ser Leu Leu Ile Leu Val Leu Ala Phe 1 5 10 15
Gly Ile Pro Arg Thr Gln Gly Ser Asp Gly Gly Ala Gln Asp Cys Cys 20
25 30 Leu Lys Tyr Ser Gln Arg Lys Ile Pro Ala Lys Val Val Arg Ser
Tyr 35 40 45 Arg Lys Gln Glu Pro Ser Leu Gly Cys Ser Ile Pro Ala
Ile Leu Phe 50 55 60 Leu Pro Arg Lys Arg Ser Gln Ala Glu Leu Cys
Ala Asp Pro Lys Glu 65 70 75 80 Leu Trp Val Gln Gln Leu Met Gln His
Leu Asp Lys Thr Pro Ser Pro 85 90 95 Gln Lys Pro Ala Gln Gly Cys
Arg Lys Asp Arg Gly Ala Ser Lys Thr 100 105 110 Gly Lys Lys Gly Lys
Gly Ser Lys Gly Cys Lys Arg Thr Glu Arg Ser 115 120 125 Gln Thr Pro
Lys Gly Pro 130
* * * * *