U.S. patent application number 14/476559 was filed with the patent office on 2017-03-02 for process for preparation of secretory iga and secretory igm.
The applicant listed for this patent is Stephen C. Brown, Michael R. Simon. Invention is credited to Stephen C. Brown, Michael R. Simon.
Application Number | 20170058018 14/476559 |
Document ID | / |
Family ID | 47846003 |
Filed Date | 2017-03-02 |
United States Patent
Application |
20170058018 |
Kind Code |
A2 |
Brown; Stephen C. ; et
al. |
March 2, 2017 |
PROCESS FOR PREPARATION OF SECRETORY IgA AND SECRETORY IgM
Abstract
A process for synthesizing and separating secretory IgA from a
mixture of IgA monmer and IgA dimer is provided. The process
includes covalently binding affinity tagged or epitope tagged
recombinant secretory component to the IgA dimer in the mixture and
binding the affinity tagged or an epitope tagged secretory IgA to
immobilized moieties on the solid phase support resin to which the
affinity tag or epitope tag binds and then eluting the affinity
tagged or an epitope tagged secretory IgA with release buffer. A
process for synthesizing and separating secretory IgM from a
mixture of IgM and other plasma proteins is also provided. The
process includes covalently binding affinity tagged or an epitope
tagged recombinant secretory component to the IgM in the mixture
and binding the affinity tagged or an epitope tagged secretory IgM
to immobilized moieties on the solid phase support resin and then
eluting the peptide tagged secretory IgM with a release buffer.
Inventors: |
Brown; Stephen C.; (Ann
Arbor, MI) ; Simon; Michael R.; (Ann Arbor,
MI) |
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Applicant: |
Name |
City |
State |
Country |
Type |
Brown; Stephen C.
Simon; Michael R. |
Ann Arbor
Ann Arbor |
MI
MI |
US
US |
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|
Prior
Publication: |
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Document Identifier |
Publication Date |
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US 20140371431 A1 |
December 18, 2014 |
|
|
Family ID: |
47846003 |
Appl. No.: |
14/476559 |
Filed: |
September 3, 2014 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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13935417 |
Jul 3, 2013 |
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14476559 |
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PCT/EP2013/054697 |
Mar 8, 2013 |
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13935417 |
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Current U.S.
Class: |
1/1 |
Current CPC
Class: |
C07K 2319/32 20130101;
A61K 2039/505 20130101; C07K 2317/21 20130101; B01D 15/3804
20130101; A61P 29/00 20180101; A61P 31/00 20180101; A61P 17/02
20180101; C07K 2319/036 20130101; C07K 1/14 20130101; C07K 14/70535
20130101; C07K 16/18 20130101; A61P 31/04 20180101; C07K 16/065
20130101; C07K 16/00 20130101; C07K 2319/10 20130101 |
International
Class: |
C07K 16/00 20060101
C07K016/00 |
Claims
1. A process for synthesizing and separating an immunoglobulin of
secretory IgA from an a monomer and dimer mixture of either IgA
mixed with other proteins comprising: adding secretory component
tagged with an affinity tag to the protein mixture containing IgA
dimers, facilitating the covalent binding of the affinity tagged
secretory component to the IgA dimer; recovering the secretory IgA
by adhesion of the affinity tagged secretory IgA to immobilized
moieties on the solid phase support resin to which the affinity tag
or epitope tag binds; and eluting the bound secretory IgA with a
release buffer.
2. The process of claim 1 wherein the secretory component is
human.
3. The process of claim 1 wherein the monomer and dimer mixture is
obtained from human plasma.
4. The process of claim 1 wherein the peptide tag is removed from
the secretory IgA.
5. The process of claim 1 wherein the peptide tag is histidine or
polyhistidine.
6. The process of claim 1 wherein the moiety on the solid phase
support resin to which the peptide binds is a divalent cation.
7. The process of claim 1 further comprising washing the unbound
IgA monomer through the resin prior to the eluting the secretory
IgA.
8. The process of claim 1 wherein the secretory component is
recombinant.
9. The process of claim 8 wherein the secretory component is
expressed with said histidine or polyhistidine.
10. The process of claim 1 further comprising coupling said
histidine or polyhistidine to the secretory component in vitro.
11. A composition comprising: a dimeric IgA; secretory component
bonded to said dimeric IgA; and a histidine or polyhistidine
covalently bonded to said secretory component.
12. A process for synthesizing and separating a secretory IgM
immunoglobulin from pentameric IgM mixed with other proteins
comprising: adding secretory component tagged with an epitope tag
to the protein mixture containing IgM pentamers, facilitating the
covalent binding of the epitope tagged secretory component to the
IgM pentamer; recovering the secretory IgM by adhesion of the
epitope tagged secretory IgM to immobilized moieties on the solid
phase support resin to which the affinity tag or epitope tag binds;
and eluting the bound secretory IgM with a release buffer.
13. The process of claim 12 wherein the secretory component is
human.
14. The process of claim 12 wherein the mixture is obtained from
human plasma.
15. The process of claim 12 wherein the epitope tag is removed from
secretory IgM.
16. The process of claim 12 wherein the moiety on the solid phase
support resin to which the epitope tag binds is a divalent
cation.
17. The process of claim 12 further comprising washing the unbound
IgM monomer through the resin prior to the eluting the secretory
IgM.
18. The process of claim 12 wherein the secretory component is
recombinant.
19. The process of claim 12 further comprising coupling said
histidine or polyhistidine to the secretory component in vitro.
20. A composition comprising: a pentameric IgM; secretory component
bonded to said pentameric IgM; and a histidine or polyhistidine
covalently bonded to said secretory component.
Description
RELATED APPLICATIONS
[0001] This application is a continuation of U.S. patent
application Ser. No. 13/935,417; filed Jul. 3, 2013; and a
continuation-in-part of PCT application Serial No.
PCT/EP2013/054697 filed Mar. 8, 2013 that in turn claims priority
benefit of EP 12158931.1 filed Mar. 9, 2012 and EP 12168343.7 filed
May 16, 2012; the contents of which are hereby incorporated by
reference.
FIELD OF THE INVENTION
[0002] This invention relates in general to a process for the
preparation of secretory IgA and secretory IgM from plasma proteins
containing IgA and/or IgM, and in particular to a process that is
scalable to allow the production of commercial quantities of
medicaments containing the same.
BACKGROUND OF THE INVENTION
[0003] Clostridium difficile (C. difficile) is a gram-positive
anaerobic bacillus. Disease is associated with C. difficule
infection.
[0004] Antibiotic associated pseudomembranous colitis results from
the use of broad-spectrum antibiotic agents such as clindamycin.
These antibiotics cause diarrhea in about 10% of treated patients
and pseudomembranous colitis in about 1%. C. difficile causes
antibiotic associated diarrhea and almost all cases of
pseudomembranous colitis.
[0005] Pseudomembranous colitis results from the production of C.
difficile toxin A (MW, 308,000) and toxin B (MW, 270,000) in the
colon (Barroso et al., Nucleic Acids Res., 18:4004; Dove et al.,
Infect. Immun., 58:480-488; Lyerly et al., Clin. Microbiol. Rev.,
1:1-18). Toxin A probably causes most of the gastrointestinal
symptoms because of its enterotoxic activity (Lyerly et al.,
Infect. Immun., 35:1147-1150; Lyerly et al., Infect. Immun.,
47:349-352). The toxins may act synergistically and the initial
pathology caused by toxin A allows toxin B to manifest its toxicity
(Lyerly et al., Infect. Immun., 47:349-352).
[0006] Most patients with C. difficile associated disease are
treated effectively with vancomycin or metronidazole. Other
treatment modalities include tolevemer, a toxin binding polymer (T.
J. Louie et al., Clin. Infect. Dis. 2006; 43:411), and an
antiparasitic medication, nitazoxanide (Med. Letter Drugs Ther.
2006; 48:89). Current treatments for C. difficile associated
disease use antibiotics such as metronidazole and vancomycin. These
drugs result in further disruption of the intestinal flora and are
associated with a 20-25% incidence of disease relapse. Therefore,
there is still a need for additional treatments of Clostridium
difficile associated disease in humans
[0007] Immunological treatment is valuable against C. difficile.
Vaccination against toxins A and B stimulates active immunity
against C. difficile disease in animals (Libby et al., Infect.
Immun., 36:822-829).
[0008] Passive immunization is another immunological treatment
against C. dfficile. Serum antibodies against C. difficile have
been shown to protect hamsters against C. difficile disease after
oral administration. Passive immunization with bovine antibodies
has been proposed as a treatment for other infectious diseases of
the gastrointestinal tract, such as diseases caused by rotavirus,
enteropathogenic and enterotoxigenic Escherichia coli, Vibrio
cholerae, and Cryptosporidium parvum. Preliminary studies indicate
that such passive immunization provides protection
(Boesman-Finkelstein et al., Infect. Immun., 57:1227-1234; Brussow
et al., J. Clin. Microbiol., 25:982-986; Fayer et al., Infect.
Immun., 58:2962-2965; Hilpert et al., J. Infect. Dis., 156:158-166;
Mietens et al., Eur. J. Pediatr., 132:239-252; Tacket et al., N.
Engl. J. Med., 318:1240-1243; Yoshiyama et al., Immunology,
61:543-547).
[0009] It has been reported that bovine immunoglobulin G (IgG)
concentrate from the colostrum of cows vaccinated with C. difficile
toxoid protects hamsters against antibiotic associated cecitis. The
hamsters were protected when treated before the onset of diarrhea
but not after diarrhea began (Lyerly et al., Infection and
Immunity, Vol. 59, No. 6, pages 2215-2218 (1991)). IgG directed
against toxins A and B of C. difficile are present in the general
population (Bacon and Fekety, Diagn. Microbiol. Infect. Dis., 1994;
18:205-209). Human intravenous immunoglobulin derived from plasma
donors has facilitated treatment in some patients, especially
patients who lack circulating antibodies to the C. difficile toxins
(Leung D. Y. et al., J. Pediatr. 1991 April; 118(4 (Pt 1)):633-7;
Salcedo J. et al., Gut 1997; 41:366-370; Wilcox M. H., J.
Antimicrob. Chemoth. 2004; 53:882-884; McPherson S. et al., Dis.
Colon Rectum 2006; 49:640-645; Cone L. A. et al., Infect. Dis.
Clin. Pract. 2006; 14:217-220).
[0010] In vitro experiments have demonstrated that polymeric IgA is
superior to monomeric IgA and IgG in preventing C. difficile toxin
damage to intestinal epithelial cell monolayers (Stubbe H. et al.,
J. Immunol. 2000; 164:1952-1960). Selective neutralization of C.
difficile toxin by serum IgA has also been demonstrated (Johnson S.
et al., Infect. Immun. 1995; 63:3166-3173).
[0011] Administration of an immunoglobulin product containing
specific antibodies to C. difficile results in the elimination of
C. difficile toxins and also killing of the bacteria within the
colon as detailed in U.S. Pat. No. 5,773,000. Such passive
immunization therefore provides an effective approach for the
treatment of C. difficile associated diseases such as colitis,
pseudomembranous colitis and antibiotic associated diarrhea. This
is especially important for patients experiencing multiple
relapses.
[0012] Oral human immunoglobulin treatment has shown efficacy in
treating acute C. difficile infections. Monomeric IgA admixed with
IgG (2:1) was derived from plasma (IgAbulin, Immuno, Vienna) (100
mg/mL); four mL administered orally 3 times daily for 3 weeks to a
three and one-half year old child with antibiotic-associated
diarrhea and C. difficile toxin A in his stools proved effective in
the context of concurrent vancomycin administration. The child
improved on this treatment (Tjellstrom B. et al., Lancet 1993;
341:701702). This report demonstrates the efficacy of passive
immunization with IgA derived from the general population.
[0013] The isolation of secretory forms of immunoglobulins is more
difficult that securing forms of a respective immunoglobulin that
circulate in the blood and lack secretory component.
[0014] Thus, there exists a need for IgA and IgM therapeutics that
are resistant to gastrointestinal tract degradation. There also
exists a need to provide such a therapeutic in a dosing form well
suited for treating an infected subject. Human plasma derived IgA
and IgM has been successfully combined with recombinant secretory
component to produce secretory IgA and secretory IgM with
biological activity (Longet et al 2013).
SUMMARY OF THE INVENTION
[0015] A process for the preparation of secretory IgA and/or
secretory IgM and their separation from unwanted other substances
including proteins is provided. The process involves the
application of an IgA source, which may contain a mixture of
monomers and dimers, or a plasma protein solution containing IgM,
to secretory component that is modified to contain an affinity tag
or an epitope tag to form secretory IgA and/or secretory IgM
containing said affinity tag or epitope tag that is useful for
capture by a solid phase support resin. The protein solution which
now contains the affinity tagged or epitope -tagged secretory IgA
or affinity tagged or epitope -tagged secretory IgM is then applied
to a solid phase support resin. The adherence of the affinity
tagged or epitope tagged secretory IgA or secretory IgM to the
resin and the unwanted components flow through and are thus
removed. The desired product, secretory IgA or secretory IgM is
then eluted from the solid phase support resin using a release
agent.
[0016] For histidine and "FLAG" peptides, examples of the moieties
on the solid phase support resin to which the peptide binds are
immobilized divalent cations or anti polyhistidine antibodies and
immobilized anti-FLAG peptide antibodies. A new composition of
histidine tagged secretory IgA or secretory IgM is also
provided.
BRIEF DESCRIPTION OF THE DRAWINGS
[0017] FIG. 1. ELISA .DELTA. colostral secretory IgA (sIgA) and
.quadrature. synthesized sIgA. Plate was coated with mouse
anti-secretory component antibody.
[0018] FIG. 2. Tris acetate PAGE showing sIgA, prepared with
histidine tagged secretory component eluted from nickel resin (Cube
Biotech, Monheim, Germany). Lane 1: MW ladder; Lane 2: colostral
secretory IgA; Lane 3 blank; Lane 4 nickel resin flow-through; Lane
5: nickel resin imidazole eluate with semisynthetic sIgA.
DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS
[0019] The present invention has utility for the preparation of
secretory IgA or secretory IgM. In some inventive embodiments, the
IgA is derived from a mixture of monomeric and dimeric plasma IgA.
or in other inventive embodiments, the preparation of secretory IgM
is derived from a mixture of IgM with other plasma proteins. The
present invention is superior to monomeric IgA or pentameric IgM
administered orally because the presence of secretory component
protects the IgA or IgM from digestion in the gastrointestinal
tract. Without intending to be bound to particular theory, it is
believed that the increased efficacy of the present invention is
achieved for secretory IgA or secretory IgM owing to the propensity
of monomeric IgA and pentameric IgM to degrade in the
gastrointestinal tract. The resultant dosing requirements increase
treatment costs. While the present invention is further detailed
principally with respect to IgA, it is appreciated that the process
and medicaments that result are equally applicable to IgM and the
resulting secretory IgM, regardless of whether the tag is retained
or removed.
[0020] An affinity tag or an epitope tag that are efficaceous for
the present invention is one of: peptide tags:AviTag, a peptide
allowing biotinylation by the enzyme BirA so the protein can be
isolated by streptavidin; GLNDIFEAQKIEWHE (SEQ ID No. 1);
calmodulin-tag, a peptide bound by the protein calmodulin
(KRRWKKNFIAVSAANRFKKISSSGAL (SEQ ID No. 2); FLAG-tag, a peptide
recognized by an antibody DYKDDDDK (SEQ ID No. 3);
Hemaglutinin-tag, a peptide recognized by an antibody YPYDVPDYA
(SEQ ID No. 4); His-tag, 5-10 histidines bound by a nickel or
cobalt or other divalent cation chelate HHHHHH (SEQ ID No. 6);
Myc-tag, a short peptide recognized by an antibody EQKLISEEDL (SEQ
ID No. 7); S-tag KETAAAKFERQHMDS (SEQ ID No. 8); SBP-tag, a peptide
which binds to streptavidin MDEKTTGWRGGHVVEGLAGELEQLRARLEHHPQGQREP
(SEQ ID No. 9); Softag 1, for mammalian expression SLAELLNAGLGGS
(SEQ ID No. 10); Softag 3, for prokaryotic expression TQDPSRVG (SEQ
ID No. 11); V5 tag, a peptide recognized by an antibody
GKPIPNPLLGLDST (SEQ ID No. 12); Xpress tag DLYDDDDK (SEQ ID No.
13); Biotin Carboxyl Carrier Protein, a protein domain recognized
by streptavidin; Glutathione-S-transferase-tag, a protein which
binds to immobilized glutathione; Green fluorescent protein-tag, a
protein which is spontaneously fluorescent and can be bound by
nanobodies; Maltose binding protein-tag, a protein which binds to
amylose agarose; Nus-tag; Strep-tag, a peptide which binds to
streptavidin, or the modified streptavidin called streptactin
Strep-tag II: WSHPQFEK (SEQ ID No. 14); Thioredoxin-tag; TC tag; or
Ty tag.
[0021] Plasma IgA contains a mixture of monomer and dimer
(Delacroix et al. 1981; Delacroix et al. 1983; Longet et al. 2013).
In some embodiments of the present invention, plasma dimeric IgA in
the naturally occurring monomer-dimer mixture is covalently bound
to recombinant peptide tagged secretory component in vitro. In
other inventive embodiments native secretory component is
covalently bonded to one or more amino acid residues through
conventional synthetic techniques (Hermanson GT 1996). As an
example using a histidine tag, it is appreciated that a single
histidine residue or a poly histidine having typically between 2
and 20 histidine residues is added to secretory component,
regardless of whether produced by recombinant, synthetic addition,
or other technique. The secretory IgA is now histidine tagged by
virtue of the divalent bonding of the histidine tagged secretory
component to the IgA dimer, The novel method of obtaining purified
secretory IgA that is thus formed is to remove the secretory IgA
that is now tagged by affinity binding of one of the aforementioned
tags to a binding moiety immobilized on a resin a or simply by a
histidine residue(s) tagged to an immobilized nickel.sup.+2 resin.
Alternatively, other immobilized divalent metal ions such as
cobalt, zinc, copper or iron can be used. Alternatively, a FLAG
peptide is used in certain inventive embodiments and antibody to
the FLAG peptide is immobilized on the solid support resin. FLAG
tags have been detailed elsewhere as for example U.S. Pat. No.
4,703,004. The resultant secretory IgA has utility, for example, as
a treatment of C. difficile associated diseases such as Clostridium
difficile colitis, pseudomembranous colitis and antibiotic
associated diarrhea and in particular to secretory immunoglobulin A
(IgA) compositions administered in the form of pharmaceutical
compositions. The above process is equally applicable to IgM to
form purified secretory IgM.
[0022] A more detailed description of an exemplary isolation of an
IgA component as a byproduct from pooled human plasma or
hyperimmune pooled human plasma is as follows. Ethanol
fractionation of pooled human plasma is a well-known process to
prepare immunoglobulin G. Pooled human plasma is first obtained
from licensed plasmapheresis centers in the United States and
tested for various pathogens including the HIV virus. The first
manufacturing step of most commercial immunoglobulin G preparations
involves a modified cold ethanol fractionation according to Cohn to
produce Cohn fraction II. In the fractionation process, many
infectious viruses are eliminated from the pooled human plasma.
Following fractionation, the Cohn fraction II is subjected to
adsorption onto an ion exchange medium. This step may selectively
reduce the IgA concentration to less than 0.1%. Such a step is
important for producing immunoglobulin G for intravenous infusion
into humans. This is because some individuals undergo an
anaphylactic-like reaction if treated with intravenous IgG that
contains IgA as an impurity.
[0023] The modified cold ethanol fractionation process according to
Cohn is a series of fractionations using various levels of ethanol,
pH, and temperature to produce a fraction II which is further
treated to produce immunoglobulins as described above. In the
fractionation process, pooled human plasma is first treated to
produce a cryoprecipitate and cryo-supernatant. Alternatively, it
is appreciated that the source plasma may be autologous plasma or
hyperimmune human plasma, either pooled, or from a single
individual who has been immunized against a specific disease.
[0024] In another embodiment, the IgA component is be prepared by
hybridoma techniques to provide antigen-specific IgA. Hybridoma
techniques are described originally in Kohler and Milstein, Nature
1975; 256:495-497 with more recent advances summarized in Berzofsky
et al., Fundamental Immunology, Third Edition, 1993, pp 455-62.
[0025] Regardless of the source, the cryo-supernatant is subjected
to a first ethanol fractionation to yield a supernatant I.
Supernatant I is subjected to a second ethanol fractionation to
yield fraction II+III. Fraction II+III is subjected to a third
ethanol fractionation procedure to yield a supernatant III and
Fraction III precipitate.
[0026] The fraction III precipitate enriched in IgA is generally
discarded as an unwanted byproduct. According to the present
invention, this unwanted IgA following ion exchange adsorption
purification is further treated by incubation with immobilized
hydrolases to inactivate viruses and vasoactive substances. Such
treatment has been proven to eliminate many viruses tested
including HIV, Sindbis, and vaccinia. Other antiviral treatments,
as known to those skilled in the art, are used in combination and
consist of solvent detergent processes, nanofiltration and/or heat
inactivation. Usually three antiviral steps are implemented.
Following incubation to remove viruses, the concentration of the
active material is adjusted with sterile saline or buffered
solutions to ensure a constant amount of active material per
milliliter of reconstituted product. Finally, the solution with a
constant amount of reconstituted product is sterilized by
filtration before use.
[0027] The ethanol fractionation process according to Cohn is well
known in the art and is described in Cohn et al., J. Am. Chem. Soc.
1946; 68:459-475, Oncley et al., J. Am. Chem. Soc. 1949;
71:541-550, and in most detail in pages 576-602, Kirk-Othmer
Encyclopedia of Chemical Technology, Vol. 3, second edition (1963).
Alternatively, ion exchange chromatography may be used to obtain
the dimeric and polymeric IgA byproduct during the manufacture of
intravenous immunoglobulin. From 4% to 22% of plasma IgA is dimeric
and polymeric IgA (Delacroix et al. 1981; Delacroix et al. 1983).
The resulting dimeric IgA-J chains are purified to form a
medicament.
[0028] The dimeric and polymeric IgA present in the plasma IgA
monomer-polymer mixture is further coupled to secretory component
that is recombinantly produced to include a histidine tag or
another of the aforementioned tags; or subsequently covalently
bonded to a peptide tag such as histidine or poly-histine
oligopeptide. In certain inventive embodiments, the coupling of IgA
to secretory component is accomplished by forming disulfide bonds
under mildly oxidizing conditions. (Jones R. M. L., Schweikart F.,
Frutiger S., Jaton J-C., Hughes G. J. Thiol-disulfide redox buffers
maintain a structure of immunoglobulin A that is essential for
optimal in vitro binding to secretory component. Biochimica et
Biophysica Acta 1998; 1429:265-274.) Dimeric and polymeric IgA
containing both J chain and secretory component is purified from
the mixture by immobilized metal ion affinity chromatography, as
performed by those of skill in the art of protein purification.
[0029] It has been previously found that it is possible to separate
recombinant proteins from cell supernates by producing such
proteins with histidine tails or other of the aforementioned tags
and then passing the cell supernates through nickel bound solid
support resins. The histidine or other tag adheres to the nickel or
other suitable tag specific binding moiety and is retained while
the unwanted proteins are washed through. The tagged secretory
immunoglobulin protein is then recovered by eluting it with an
imidazole buffer in the case of an amide-metal bond between target
protein and resin (Block H et al 2009).
[0030] The mixture of histidine tagged secretory IgA and residual
monomeric IgA is buffer exchanged into a binding buffer containing
low concentrations of imidazole (<40 mM). Another release agent
operative to exchange histidine tagged secretory IgA or secretory
IgM illustratively includes 1) ethylene diamine tetraacetic acid
(EDTA) at 10 mM and 2) an elution buffer of pH 5.5 or lower.
Typical binding buffer imidazole concentrations range from 0.1 to
100 millimolar (mM). It is appreciated that the initial binding
buffer pH is somewhat variable and readily discerned for a given
chemical structure of buffer and concentration through routine
experimentation. The chromatography medium operative herein is
selected to be stable in the presence of the binding buffer and
able to separate histidine tagged secretory IgA. Suitable media
illustratively include Exemplary of these media are nickel, cobalt
and zinc immobilized on sepharose. In a preferred embodiment, the
affinity medium is washed in a wash buffer containing from 0 to 100
mM imidazole to remove unbound monomeric IgA. The bound histidine
tagged secretory IgA is recovered using an elution buffer of a
higher imidazole concentration (e.g. 100 to 1000 mM). With
successive elutions separation of monomeric from histidine tagged
secretory component bound dimeric IgA is exacted. It is appreciated
that the inventive process is amenable to scaling to produce
quantities sufficient to treat numerous subjects. It is appreciated
that similar selective binding pairs is achieved between other
inventive tagged secretory component containing immunoglobulin
proteins and resins are conventional to the art for each of the
aforementioned tags.
[0031] By way of a specific example, the binding and wash buffers
are 50 mM NaH.sub.2PO.sub.4 300 mM NaCl and 20 mM imidazole, is
adjusted to pH 8. The mixture of IgA monomer and secretory IgA is
dissolved in that buffer. The elution buffer is identical to the
binding buffer with the exception that the imidazole is at a higher
concentration, e.g 100 to 1000 mM.
[0032] The remaining histidine tagged secretory IgA is then eluted
from the divalent immobilized metal resin with the elution buffer
according to conventional techniques and conditions that include an
exemplary basic pH of for example 8 to 10, see FIGS. 1 and 2.
[0033] Purified secretory IgA containing histidine tagged secretory
component is stabilized in some embodiments for example by the
addition of human serum albumin to a final concentration of 5%
total weight albumen.
[0034] In another embodiment, the tag is removed from the recovered
secretory IgA and native secretory IgA is available for usage as a
medicament. For a histidine tagged secretory IgA a procedure for
tag removal is known to the art (Kopera E et al 2012).
[0035] In summary, the inventive process is the addition of tagged
secretory component in either recombinatant or post expression
tagging to a mixture of plasma derived IgA monomers and dimers, in
which the tagged secretory component combines with the IgA dimer
forming secretory IgA and allows recovery of the newly formed
secretory IgA by adhesion to immobilized divalent metal ions or
other solid phase moiety, and subsequent elution therefrom.
[0036] Plasma IgM can be recovered from the byproducts of the
production of intravenous immunoglobulin. An example of such a
byproduct is Cohn fraction III precipitate. The IgM is most easily
solublized from Cohn fraction III precipitate by 20 mM sodium
acetate. Other plasma proteins are similarly solublized along with
the IgM. The plasma IgM in this protein mixture is covalently bound
to recombinant histidine tagged secretory component in vitro
forming secretory IgM within the protein mixture. The secretory IgM
is now tagged by virtue of the divalent bonding of the tagged
secretory component to the IgM. The novel method of obtaining
purified secretory IgM that is thus formed is to remove the
secretory IgM that is now tagged by affinity binding of the tag to
a immobilized nickel.sup.+2 or other divalent metal ion or other
suitable binding moiety conventional to the art that is part of a
resin column. The resultant semisynthetic secretory IgM has
utility, for example, as a treatment of Clostridium difficile
associated diseases such as Clostridium difficile colitis,
pseudomembranous colitis and antibiotic associated diarrhea and in
particular to secretory immunoglobulin M compositions administered
in the form of pharmaceutical compositions.
[0037] In another embodiment, the tag is removed from the recovered
secretory IgM and native secretory IgM is available for usage as a
medicament. For a histidine tagged secretory IgM a procedure for
tag removal is known to the art (Kopera E et al 2012).
[0038] Thus, an inventive process provides the addition of peptide
tagged secretory component to a mixture of plasma derived IgM and
other plasma proteins, in which the peptide tagged secretory
component combines with the IgM forming secretory IgM and allows
recovery of the newly formed secretory IgM by adhesion to moieties
on the solid phase support resin to which the peptide binds, and
subsequent elution therefrom using an elution buffer.
REFERENCES
[0039] Aoyama K, Chiba J. Separation of different molecular forms
of mouse IgA and IgM monoclonal antibodies by high-performance
liquid chromatography on spherical hydroxyapatite beads. J Immunol
Methods. 1993; 162(2):201-10.
[0040] Bacon A. E. 3rd, Fekety R. Immunoglobulin G directed against
toxins A and B of Clostridium difficile in the general population
and patients with antibiotic-associated diarrhea. Diagn. Microbiol.
Infect. Dis. 1994; 18:205-209.
[0041] Barroso L. A., Wang S. Z., Phelps C. J., Johnson J. L.,
Wilkins T. D. Nucleotide sequence of Clostridium difficile toxin B
gene. Nucleic Acids Res. 1990; 18:4004.
[0042] Berzofsky J. A., Berkower I. J., Epstein S. L., Monoclonal
Antibodies in Chapter 12, Antigen-Antibody Interactions and
Monoclonal Antibodies, pp. 455-465 in Fundamental Immunology, Third
Edition, W. E. Paul (ed), Raven Press, NY 1993. Berzofsky et al.,
Fundamental Immunology, Third Edition, 1993, pp 455-462.
[0043] Block H, Maertens B, Spriestersbach A, Brinker N, Kubicek J,
Fabis R, Labahn J, Schafer F. Immobilized-Metal Affinity
Chromatography (IMAC): A Review. CHAPTER 27 in Methods in
Enzymology, Volume 463, 2009.
[0044] Boesman-Finkelstein M., Walton N. E., Finkelstein R. A.
Bovine lactogenic immunity against cholera toxin-related
enterotoxins and Vibrio cholerae outer membranes. Infect. Immun.
1989; 57:1227-1234.
[0045] Brussow H., Hilpert H., Walther I., Sidoti J., Mietens C.,
Bachmann P. Bovine milk immunoglobulins for passive immunity to
infantile rotavirus gastroenteritis. J. Clin. Microbiol. 1987;
25:982-986.
[0046] Cohn E. J., Strong L. E., Hughes W. L., Jr., Mulford D. J.,
Ashworth J. N., Melin M., Taylor H. L., Preparation and Properties
of Serum and Plasma Proteins IV. A System for the Separation into
Fractions of the Protein and Lipoprotein Components of Biological
Tissues and Fluids, J. Am. Chem. Soc. 1946; 68; 459-475.
[0047] Cone L. A., Lopez C., Tarleton H. L., Jodoin D., Taylor M.,
Gade-Andavolu R., Dreisbach L. P. A durable response to relapsing
Clostridium difficile colitis may require combined therapy with
high-dose oral vancomycin and intravenous immune globulin. Infect.
Dis. Clin. Pract. 2006; 14:217-220.
[0048] Corthesy B., Recombinant Secretory IgA for Immune
Intervention Against Mucosal Pathogens, Biochem. Soc. Trans. 1997,
25; 471-475.
[0049] Corthier et al., Emergence in Gnotobiotic Mice of
Nontoxinogenic Clones of clostridium difficile from a Toxinogenic
One, Infection and Immunity, June 1988, pp. 1500-1504.
[0050] Corthier et al., Protection Against Experimental
Pseudomembranous Colitis in Gnotobiotic Mice by Use of Monoclonal
Antibodies Against clostridium difficile Toxin A, Infection and
Immunity, March 1991, pp. 1192-1195.
[0051] Crottet P., Cottet S., Corthesy B., Expression, Purification
and Biochemical Characterization of Recombinant Murine Secretory
Component, A Novel Tool in Mucosal Immunology, Biochem. J. 1999,
341; 299-306.
[0052] Delacroix D. L., Hodgson H. J., McPherson A., Dive C.,
Vaerman J. P. Selective transport of polymeric immunoglobulin A in
bile. Quantitative relationships of monomeric and polymeric
immunoglobulin A, immunoglobulin M, and other proteins in serum,
bile, and saliva. J. Clin. Invest. 1982 August; 70(2):230-41
[0053] Delacroix D. L., Elkom K. B., Geubel A. P., Hodgson H. F.,
Dive C., Vaerman J. P. Changes in size, subclass, and metabolic
properties of serum immunoglobulin A in liver diseases and in other
diseases with high serum immunoglobulin A. J. Clin. Invest. 1983
February; 71(2):358-67.
[0054] Dove C. H., Wang S. Z., Price S. B., Phelps C. J., Lyerly D.
M., Wilkins T. D. and Johnson J. L.; Lyerly et al. Molecular
characterization of the Clostridium difficile toxin A gene. Infect.
Immun. 1990; 58:480-488.
[0055] Ehrich et al., Production of clostridium difficile
Antitoxin, Infection and Immunity, June 1980, pp. 1041-1043.
[0056] Fayer R., Guidry A., Blagburn B. L. Immunotherapeutic
efficacy of bovine colostral immunoglobulins from a hyperimmunized
cow against cryptosporidiosis in neonatal mice. Infect. Immun.,
1990; 58:2962-2965.
[0057] Gerding et al., clostridium difficile-Associated Diarrhea,
Archives of Internal Medicine, vol. 146, January 1986, pp.
95-100.
[0058] Hermanson G T. Bioconjugate Techniques. Academic Press, San
Diego 1996.
[0059] Hilpert H., Brussow H., Mietens C., Sidoti J., Lerner L.,
Werchau H. Use of bovine milk concentrate containing antibody to
rotavirus to treat rotavirus gastroenteritis in infants. J. Infect.
Dis. 1987; 156:158-166.
[0060] Johnson S. et al. Infect. Immun. 1995; 63:3166-3173.
[0061] Jones R. M. L., Schweikart F., Frutiger S., Jaton J-C.,
Hughes G. J. Thiol-disulfide redox buffers maintain a structure of
immunoglobulin A that is essential for optimal in vitro binding to
secretory component. Biochimica et Biophysica Acta 1998;
1429:265-274.
[0062] Kelly et al., clostridium difficile Colitis, New England
Journal of Medicine, vol. 330, January 1994, pp. 257-262.
[0063] Kelly et al., Human Colonic Aspirates Containing
Immunoglobulin A Antibody to clostridium difficile Toxin A Inhibit
Toxin A-Receptor Binding, Gastroenterology, vol. 102, No. 1, pp.
35-40.
[0064] Kohler G., Milstein C., Continuous Cultures of Fused Cells
Secreting Antibody of Predetermined Specificity, Nature 1975;
256;495-497.
[0065] Kopera E, Belczyk-Ciesielska A, Bal W. Application of
Ni(II)-assisted peptide bond hydrolysis to non-enzymatic affinity
tag removal. PLoS One. 2012; 7(5):e36350. doi:
10.1371/journal.pone.0036350. Epub 2012 May 4.
[0066] Leung D. Y., Kelly C. P., Boguniewicz M., Pothoulakis C.,
LaMont J. T., Flores A. Treatment with intravenously administered
gamma globulin of chronic relapsing colitis induced by Clostridium
difficile toxin. J. Pediatr. 1991 April; 118(4 (Pt 1)):633-637.
[0067] Libby J. M., Jortner B. S., Wilkins T. D. Effects of the two
toxins of Clostridium difficile in antibiotic-associated cecitis in
hamsters. Infect. Immun. 1982 May; 36(2):822-829.
[0068] Lima et al., Effects of clostridium difficile Toxins A and B
in Rabbit Small and Large Intestine In Vivo and on Cultured Cells
In Vitro, Infection and Immunity, March 1988, pp. 582-588.
[0069] Longet S, Miled S, Lotscher M, Miescher S M, Zuercher A W,
Corthesy B. Human plasma-derived polymeric IgA and IgM antibodies
associate with secretory component to yield biologically active
secretory-like antibodies. J Biol Chem. 2013 Feb. 8;
288:4085-94.
[0070] Louie T. J., Peppe J., Watt C. K., Johnson D., Mohammed R.,
Dow G., Weiss K., Simon S., John J. F. Jr., Garber G., Chasan-Taber
S., Davidson D. M.; Tolevamer Study Investigator Group. Tolevamer,
a novel nonantibiotic polymer, compared with vancomycin in the
treatment of mild to moderately severe Clostridium
difficile-associated diarrhea. Clin. Infect. Dis. 2006;
43:411-20.
[0071] Luellau, E., von Stockar, U., Vogt, S., Freitag, R.
Development of a downstream process for the isolation and
separation of monoclonal immunoglobulin A monomers, dimers and
polymers from cell culture supernatant, Journal of Chromatography
A, 1998, 796:165-175.
[0072] Lullau E., Heyse S., Vogel H., Marison I., von Stockar U.,
Kraehanbuhl J-P., Corthesy B., Antigen Binding Properties of
Purified Immunoglulin A Antibodies, J. Biol. Chem. 1996;
271:16300-16309.
[0073] Lyerly D. M., Krivan H. C., Wilkins T. D. Clostridium
difficile: its disease and toxins. Clin. Microbiol. Rev. 1988;
1:1-18.
[0074] Lyerly D. M., Phelps C. J., Toth J., Wilkins T. D.
Characterization of toxins A and B of Clostridium difficile with
monoclonal antibodies. Infect. Immun. 1986; 54:70-76.
[0075] Lyerly D. M., Bostwick E. F., Binion S. B., Wilkins T. D.
Passive immunization of hamsters against disease caused by
Clostridium difficile by use of bovine immunoglobulin G
concentrate. Infect. Immun. 1991; 59:2215-2218.
[0076] Lyerly D. M., Lockwood D. E., Richardson S. H., Wilkins T.
D. Biological activities of toxins A and B of Clostridium
difficile. Infect. Immun. 1982; 35:1147-1150.
[0077] Lyerly D. M., Saum K. E., MacDonald D. K., Wilkins T. D.
Effects of Clostridium difficile toxins given intragastrically to
animals. Infect. Immun. 1985; 47:349-352.
[0078] Mahe et al., Effect of Various Diets on Toxin Production by
Two Strains of clostridium difficile in Gnotobiotic Mice, Infection
and Immunity, August 1987, pp. 1801-1805.
[0079] Martinez et al., Purification and Characterization of
clostridium sordellii Hemorrhagic Toxin and Cross-Reactivity with
clostridium difficile Toxin A (Enterotoxin), Infection and
Immunity, May 1988, pp. 12-15-1221.
[0080] McFarland et al., Nosocomial Acquisition of clostridium
difficile Infection, The New England Journal of Medicine, January
1989, pp. 204-210.
[0081] McFarland et al., Review of clostridium difficile Associated
Diseases, American Journal of Infection Control, vol. 14, No. 3,
June 1986, pp. 99-104.
[0082] McPherson S., Rees C. J., Ellis R., Soo S. and Panter S. J.
Intravenous Immunoglobulin for the Treatment of Severe, Refractory,
and Recurrent Clostridium difficile Diarrhea. Diseases of the Colon
& Rectum. 2006; 49(5):640-645.
[0083] Med. Letter Drugs Ther. 2006; 48:89-90, 92.
[0084] Mietens C., Keinhorst H., Hilpert H., Gerber H., Amster H.,
Pahud J. J. Treatment of infantile E. coli gastroenteritis with
specific bovine anti-E. coli milk immunoglobulins. Eur. J. Pediatr.
1979; 132:239-252.
[0085] Mitchell et al., Effect of Toxin A and B of clostridium
difficile on Rabbit Ileum and Colon, Gut, 1986, vol. 27, pp.
78-85.
[0086] Morris et al., Role of Surgery in Antibiotic-Induced
Pseudomembranous Enterocolitis, The American Journal of Surgery,
vol. 160, November 1990, pp. 535-539.
[0087] Oncley J. L., Melin M., Richert D. A., Cameron J. W., Gross
P. M., Jr., The Separation of the Antibodies, Isoagglutinins,
Prothrombin, Plasminogen and .beta.1-Lipoprotein into Subfractions
of Human Plasma. J. Am. Chem. Soc. 1949; 71:541-550.
[0088] Pothoulakis C., LaMont J. T., Eglow R., Gao N., Rubins J.
B., Theoharides T. C., Dickey B. F. Characterization of rabbit
ileal receptors for Clostridium difficile toxin A. Evidence for a
receptor-coupled G protein. J. Clin. Invest. 1991; 88:119-25.
[0089] Rothman et al., Differential Cytotoxic Effects of Toxins A
and B Isolated from clostridium difficile, Infection and Immunity,
November 1984, pp. 324-331.
[0090] Salcedo J. et. al. Gut 1997; 41:366-370.
[0091] Strong L. E., Blood Fractionation, pp. 576-602 in vol. 3,
Kirk-Othmer Encyclopedia of Chemical Technology. Second Edition, H.
F. Mark, J. J. McKetta, D. F. Othmer (eds), Interscience
Publishers, NY 1963, pp. 576-602.
[0092] Stubbe H. et al. J. Immunol. 2000; 164:1952-1960.
[0093] Symersky J., Novak J., McPherson D. T., DeLucas L., Mestecky
J. Expression of the recombinant human immunoglobulin J chain in
Escherichia coli. Mol. Immunol. 2000; 37:133-140.
[0094] Tacket C. O., Losonsky G., Link H., Hoang Y., Guesry P.,
Hilpert H., Levine M. M. Protection by milk immunoglobulin
concentrate against oral challenge with enterotoxigenic Escherichia
coli. N. Engl. J. Med. 1988; 318:1240-3.
[0095] Tjellstrom B., Stenhammar L., Eriksson S., Magnusson K. E.
Oral immunoglobulin A supplement in treatment of Clostridium
difficile enteritis. Lancet 1993; 341(8846):701-702.
[0096] Triadafilopoulos et al., Differential Effects of clostridium
difficile Toxins A and B on Rabbit Ileum, Gastroenterology, 1987,
vol. 93, pp. 273-279.
[0097] Tucker et al., Toxin A of clostridium difficile Is a Potent
Cytotoxin, Journal of Clinical Microbiology, May 1990, pp.
869-871.
[0098] Weltzin R., Traina-Dorge V., Soike K., Zhang J. Y., Mack P.,
Soman G., Drabik G., Monath T. P., Intranasal Monoclonal IgA
Antibody against Respiratory Syncytial Virus Protects Rhesus
Monkeys against Upper and Lower Respiratory Tract Infection. J.
Infect. Dis. 1996; 174:256-261.
[0099] Weltzin R., Hsu S. A., Mittler E. S., Georgakopoulas K.,
Monath T. P., Intranasal Monoclonal Immunoglobulin A against
Respiratory Synctial Virus Protects against Upper and Lower
Respiratory Tract Infections in Mice. Antimicrob. Agents Chemother.
1994; 38:2785-2791.
[0100] Wilcox M. H.. J. Antimicrob. Chemoth. 2004; 53:882-884.
[0101] Yoshiyama Y., Brown W. R. Specific antibodies to cholera
toxin in rabbit milk are protective against Vibrio cholerae-induced
intestinal secretion. Immunology. 1987; 61:543-547.
[0102] Patent applications and publications mentioned in the
specification are indicative of the levels of those skilled in the
art to which the invention pertains. These applications and
publications are incorporated herein by reference to the same
extent as if each individual application or publication was
specifically and individually incorporated herein by reference.
[0103] The foregoing description is illustrative of particular
embodiments of the invention, but is not meant to be a limitation
upon the practice thereof. The following claims, including all
equivalents thereof, are intended to define the scope of the
invention.
* * * * *