WO2004111080A1 - Epitopes of the cytomegalovirus pp65 protein - Google Patents
Epitopes of the cytomegalovirus pp65 protein Download PDFInfo
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Classifications
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
- G01N33/569—Immunoassay; Biospecific binding assay; Materials therefor for microorganisms, e.g. protozoa, bacteria, viruses
- G01N33/56983—Viruses
- G01N33/56994—Herpetoviridae, e.g. cytomegalovirus, Epstein-Barr virus
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- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K38/00—Medicinal preparations containing peptides
- A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- A61K38/162—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from virus
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K39/00—Medicinal preparations containing antigens or antibodies
- A61K39/46—Cellular immunotherapy
- A61K39/461—Cellular immunotherapy characterised by the cell type used
- A61K39/4611—T-cells, e.g. tumor infiltrating lymphocytes [TIL], lymphokine-activated killer cells [LAK] or regulatory T cells [Treg]
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
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- A61K39/00—Medicinal preparations containing antigens or antibodies
- A61K39/46—Cellular immunotherapy
- A61K39/461—Cellular immunotherapy characterised by the cell type used
- A61K39/4612—B-cells
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- A—HUMAN NECESSITIES
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- A61K39/46—Cellular immunotherapy
- A61K39/461—Cellular immunotherapy characterised by the cell type used
- A61K39/4614—Monocytes; Macrophages
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- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K39/00—Medicinal preparations containing antigens or antibodies
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- A61K39/461—Cellular immunotherapy characterised by the cell type used
- A61K39/4615—Dendritic cells
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K39/00—Medicinal preparations containing antigens or antibodies
- A61K39/46—Cellular immunotherapy
- A61K39/462—Cellular immunotherapy characterized by the effect or the function of the cells
- A61K39/4622—Antigen presenting cells
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- A—HUMAN NECESSITIES
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- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K39/00—Medicinal preparations containing antigens or antibodies
- A61K39/46—Cellular immunotherapy
- A61K39/464—Cellular immunotherapy characterised by the antigen targeted or presented
- A61K39/464838—Viral antigens
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- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- C07K14/005—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from viruses
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- A—HUMAN NECESSITIES
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- A61K39/00—Medicinal preparations containing antigens or antibodies
- A61K2039/51—Medicinal preparations containing antigens or antibodies comprising whole cells, viruses or DNA/RNA
- A61K2039/53—DNA (RNA) vaccination
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N2710/00—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA dsDNA viruses
- C12N2710/00011—Details
- C12N2710/16011—Herpesviridae
- C12N2710/16111—Cytomegalovirus, e.g. human herpesvirus 5
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Abstract
Novel antigenic epitopes from Cytomegalovirus (CMV) pp65 protein, pharmaceutical compositions containing the same and the use thereof in the preventive and therapeutic treatment of CMV infections, are disclosed.
Description
EPITOPES OF THE CYTOMEGALOVIRUS ΌΌ65 PROTEIN
The present invention relates to the identification of antigenic epitopes recognized by Th cells (T helper CD4) within the human Cytomegalovirus (CMV) pp65 protein. More specifically, the invention provides pp65 peptides able to induce a CMV-specific and Th-mediated immune response, pharmaceutical and diagnostic compositions containing them and the use thereof in the prophylaxis, therapy and diagnosis of CMV infections. BACKGROUND OF THE INVENTION
Cytomegalovirus is an opportunistic pathogenic agent that may cause infections or reactivate, when already present in the host organism, especially in immunocompromised patients, such as individuals transplanted with solid organs or with stem cells, or AIDS patients (1-4). The clinical experience shows that defects of cellular immunity (mediated by lymphocytes CD4/Th and CD8/CTL) increase the risk of CMV reactivation (5-7).
Experiments in animals have shown that latent CMV is controlled by the combined actions of specific CTL and Th cells (8,9). Studies in mice and humans showed the importance of adoptive cellular immunoreconstitution in the prophylaxis and therapy of CMV complications (10-14). The interest for this treatment is witnessed by several studies aimed at defining easy procedures to generate CTL lines specific for CMV (15-22). These studies led to the identification of CTL epitopes in the immunodominant pp65 tegument protein and in other viral antigens. Most of the pp65 epitopes so far identified are of the CTL-type, i.e. they are recognized by CTL cells in a MHC-class I molecule context. US patents No. 6251399, 6207161, 6156317, 6120989 and
6074645 describe pp65 peptide fragments able to induce the lysis of human cells infected by HCMV through the activation of cytotoxic T lymphocytes.
TBSsiiiivolvement of Th cells, which are essential to the survival and
clonal expansion of specific CTLs (23), is particularly important for inducing an effective immune response against CMV.
DESCRIPTION OF THE INVENTION
In order to identify novel Th epitopes and the respective restriction HLA-class II alleles, 138 peptides corresponding to contiguous and partially overlapping regions of the CMV ρp65 protein were synthesized. The immunogenic peptides were selected for their ability to induce secretion of gamma interferon (IFNγ) by peripheral lymphocytes (PBMC) obtained from CMV seropositive subjects, and to activate subpopulations of selected Th CD4+ cells. The screening of CMV-seropositive subjects and the subsequent in vitro functional assays allowed the identification of the following CMV pp65 Th peptides, which are the subject of the present invention (the corresponding locations within the pp65 sequence SwissProt P06725 are indicated in the right-hand column): PLKMLNIPSINVHHY (SEQ ID N. 1) aa 117-131
TRQQNQWKEPDVYYT (SEQ ID N. 2) aa 169-183
EPDVYYTSAFVFPTK (SEQ ID N. 3 ) aa 177- 191
KVYLESFCEDVPSGK (SEQ ID N. 4) aa 225-239
TLGSDVEEDLTMTRN (SEQ ID N. 5) aa 245-259 QPFMRPHERNGFTVL (SEQ ID N. 6) aa 261-275
YRIQGKLEYRHTWDR (SEQ ID N. 7) aa 373-379
TERKTPRVTGGGAMA (SEQ ID N. 8) aa 413-427
ASTSAGRKRKSASSA (SEQ ID N. 9) aa 429-443
ACTSGVMTRGRLKAE (SEQ ID N. 10) aa 445-459 AGILARNLVPMVATV (SEQ ID N. 11) aa 489-503
All the invention peptides proved able to activate T cells CD4+, to induce their in vitro proliferation and expansion, thereby inducing and sustaining a specific immune response against CMV (24). In particular,
mixtures of Th peptides proved able to induce CD4 T lymphocyte recognition of 1) immunogenic peptides from the same individual, 2) the entire pp65 protein and 3) the pp65 protein from a viral lysate.
Accordingly, the invention peptides can be applied to I. in vitro diagnosis of CD4 cell-mediated immunity,
2. ex vivo generation of CD4 lymphocyte lines for immunoadoptive therapy,
3. in vivo stimulation of the immune response against CMV with vaccines containing synthetic or recombinant peptides, or DNA. According to one aspect, the invention provides pharmaceutical compositions containing one or more peptides herein disclosed and the use thereof for the preventive or therapeutic treatment of CMV infections.
The invention peptides can be prepared in different ways. For example, they can be synthesised in solution or on a solid phase, according to known procedures, or with an authomatic synthesizer. The principles and methods for peptide synthesis are known to those skilled in the art, and can be found, for example, in Stewart e Young, (1984) Solid Phase Peptide Synthesis, 2nd and., Pierce Chemical Co.; Tam et al., J. Am. Chem. Soc, (1983) 105:6442; Merrifield, The Peptides, Gross and Meinehofer eds. Academic Press, (1979) New York, pp. 1-284.
The pharmaceutical compositions are preferably administered through the parenteral and particularly the subcutaneous or intramuscular routes, and contain an effective amount of peptide, i.e. an amount able to induce a cell- mediated immune response against CMV virus in either active or latent form. According to a preferred embodiment, the peptides are administered in the form of a vaccine. The pharmaceutical compositions may contain at least one, preferably at least four, more preferably at least seven peptides according to the invention, most preferably all the peptides SEQ ID N. 1 - SEQ ID N. 11.
To increase immunogenicity, the peptides can be cross-linked or coupled to immunogenic carriers or to suitable adjuvants. Furthermore, the peptides can be conjugated to lipids, glucides, or different peptides to improve biovailability or HLA molecule-affinity. DNA molecules containing a peptide-encoding sequence can be used in the vaccination against CMV infections. The techniques for DNA immunization are known to those skilled in the art (Donnelly JJ. et al., 1994, The immunologist 2:1) and include parenteral, mucosal or intramuscular administration of peptide-encoding DNA, or of a suitable genetic construct thereof, optionally inserted in a vector.
The peptides, peptide-encoding nucleic acid molecules or pharmaceutical compositions according to the invention can be advantageously used to treat CMV-infected or seropositive subjects, individuals carrying latent CMV, subjects at risk of exposure to the virus or to viral-reactivation, especially immunosuppressed patients after organ or cell transplantation, or previously-infected subjects.
Mixtures of different HLA class Il-restricted peptides are prefarbly used to increase the efficacy of the treatment and broaden the activity spectrum.
Furthermore, the pp65 epitopes herein disclosed can be combined with known Th epitopes, such as LLQTGIHVRVSQPSL, IIKPGKISHIMLDVA,
EHPTFTSQYRIQGKL, KYQEFFWDANDIYRI (25, 26, 27).
The invention further provides an in vitro method for the activation of antigen-specific T lymphocytes, which comprises exposing cultured lymphocytes to the invention peptides or to cells exposing on their surface said peptides linked to HLA molecules.
The invention is also directed to antigen presenting cells (APC) or dendritic cells bearing an invention peptide in association with class-II HLA molecules. The cells can be exposed in vitro to one or more peptides of the
invention and then utilized to induce or stimulate a CMV-specific immune response either in vitro or in vivo. Before their reintroduction into the patient, the APCs can be purified, for example by means of affinity column chromatography utilizing an antibody to a surface marker. The invention is also directed to diagnostic compositions or reagents that utilize the peptides herein disclosed. In a typical diagnostic assay, the response or activation of T cells from a CMV-seropositive subject or from a subject at risk of CMV-exposure are analyzed by contacting the cells with APCs previously pulsed with one or more invention peptides, in a suitable in vitro system.
EXPERIMENTAL Materials and Methods Antigens and media
CMV antigen (strain AD 169, Microbix, Toronto, Canada) and protein pp65 (His-tagged, affinity-purified recombinant protein expressed in E. CoIi, kindly gifted by G. Palύ, Padova) were used at 5 and 1 μg/ml, respectively.
Peptides from pp65 (SwissProt P06725) were 15mers overlapped by 11 residues. Peptides were used at 1 μg/ml in lymphoproliferation, ELISPOT or for the generation of CD4 lines. Cultures were performed in RPMI 1640 (BioWhittaker, Verviers, BE) (10 mM L-glutamine, 5% autologous plasma).
Human recombinant IL-2 (Proleukin, Chiron, Emeryville, US) was used at 30
U/ml final concentration.
Lymphoproliferation
IgG-CMV-seropositive donors (age 18-55) were HLA-typed serologically for class I antigens (BAG, Lich, Grmany) and molecularly typed at low resolution for class II (Dynal, Meresyside, UK). Heparinized blood was spun on Biocoll (Biochrom, Berlin, DE). PBMCs were resuspended in RPMI-
1640 (2 x 106/ml) and dispensed in flat-bottom microtiter plates (Costar,
Cambridge, MA) at 200 μl/well with or without antigens. Plates were pulsed after 4 days (0.018 MBq 3H-thymidine, Amersham, Aylesbury, UK) and harvested after 24 hours. The filters were counted with a scintillator (Matrix- 9600, Cmberra-Packard, Meriden, CT); results are shown as Kcpm (cpm x 103).
ELISPOT assay
PVDF plates (Millipore, Bedford, MA) were coated with anti-IFNγ mAb (ELISPOT-hIFNg, Mabtech, Nacka, SE) following the manufacturer's instructions. PBMC (2xlO5) or fractionated cells or T cells (2xlO3) from established lines were dispensed in 200μl. The plates were incubated for 20hrs, processed as indicated by the manufacturer and developed (alkaline- phosphate-conjugate kit, Bio-Rad, Hercules, CA). Spot-forming units (SFU) were enumerated under a stereomicroscope. Responses with >20 spots/well and positive:negative ratio >3 were scored as positive. Results are shown as SFU/106 cells.
CD4 and CD 8 fractionation
Magnetic beads coated with anti-CD4 or anti-CD8 antibodies (M-450, Dynal, Oslo, NO) were used for negative fractionation of PBMC. Standard separations were performed on 5x106 PBMC. Generation of CD4 cell lines
PBMC were stimulated with autologous APCs and with IL-2 (28, 29, 30). PBMC were pulsed with protein or peptide antigens at 10xl06/ml and plated at 2xlO6/ml in 24-well plates. After 4 days IL2 was added. Cells were split to 0.2 -lxlO6/ml. After 3 weeks, 5xlO5 cells were stimulated with 106 autologous irradiated (30Gy) PBMC pulsed with antigens. Restimulations were performed every 3 weeks. These conditions select CD4 lymphocytes (>96% after 2 cycles). Specific proliferation was checked on 2x104 cells with 105 irradiated PBMC.
Results
Proliferative responses to CMV antigens
Proliferation to CMV and to pp65 was determined on 30 CMV- seropositive healthy donors. All the CMV responders (>2 ΔKcpm) responded also to pp65, with a correlation coefficient between the two proliferation levels of 0.65 (Fig. 1).
ELISPOT on PBMC with pρ65 peptides
The panel of 138 overlapped peptides of pp65 was used for the screening. Figure 2 shows representative ELISPOT profiles, with examples of broad responses (left panels) and focussed responses (right panel). Screening on 20 informative subjects allowed the identification of pp65 regions that were frequently recognized (hot spots, solid bars) or regions that were never recognized by these donors (cold spots, open bars) (Fig. 3). However, this assay did not discriminate between CD4 and CD8 responses, but indicated peptides deserving further analysis (Fig. 3).
ELISPOT on CD4 and CD8 cells
To define Th peptides, PBMC from 18 donors were fractionated with magnetic beads to obtain CD4 and CD8 cell subsets. Fig. 4 shows examples of
CD4 and CD8 subsets tested with peptides encompassing Th or CTL epitopes. Responses to pp65 were CD4 mediated in these donors, while in 5 donors (not shown) pp65 stimulated both CD4 and CD8, as reported by others (14, 22).
Peptides were recognized by CD4 (pep 62, 128, 57) or CDS cells (pep 123,
104), with mutually exclusive patterns in these donors. Peptides NV9 (HLA-
A2) and TMlO (HLA-B 7), within 15mers pep 123 and pep 104, confirmed recognition by CD8 cells.
Definition of CD4 peptides
Peptides defined in Fig. 3 were tested on CD4 and CD8 cells from 18 donors. 11 peptides CD4 were identified in this study for the first time. When
overlapped peptides were stimulators, only the most antigenic have been indicated in the Table. The Table shows the serial number of the peptide, the amino acid positions and sequences, the number of responders versus the number of subjects expressing the indicated HLA-DR allele as peptide restriction element. Four peptides, as shown, were originally reported by others (25, 26, 27) (Table).
Our main goal was the identification of Th peptides that could be used in combination for stimulating different donors independently from their own HLA-class II alleles. Seven peptides Th contained known CTL epitopes (underlined sequences in the Table).
Pep n residues sequence ϊsponders DR restriction references
11 41-55 LLQTGIHVRVSQPSL 2/4 15 21 (DQ6), 32 (DR15)
30 117-131 PLKMLNIPSINVHHY 3/7 1 tc 2/3 3
2/6 7
43 169-183 TRQQNQWKEPDVYYT 1/7 1
45 177-191 EPDVYYTSAFVFPTK 2/6 7
57 225-239 KVYLESFCEDVPSGK 2/4 15
62 245-259 TLGSDVEEDLTMTRN 1/3 - 1/6 3 - 7
66 261-275 QPFMRPHERNGFTVL 1/3 13
1/9 11
71 281-295 IIKPGKISHIMLDVA 2/2 4 32 (DR53)
1/7 - 1/6 1 - 7 32 (DR53)
92 365-379 EHPTFTSQYRIQGKL 4/9 11 21 (DRIl), 49 (DRIl)
94 373-387 YRIQGKLEYRHTWDR 1/7 - 1/3 1 - 3
104 413-427 TERKTPRVTGGGAMA 1/9 11
1/2 14
108 429-443 ASTSAGRKRKSASSA 2/9 11
112 445-459 ACTSGVMTRGRLKAE 5/7 1
123 489-503 AGILARNLVPMVATV 7/9 11
1/3 - 1/7 3 - 1
128 509-523 RIEFFWDANDIYKYQ 5/7 1
1/3 3 32 (DR52\ 49 fDR3(T
Legenda
Pep n: peptide serial number in the panel of 138; Peptide residues: peptide initial and end positions Sequences: amino acid sequence
For each peptide, the location within pp65 sequence (SwissProt P06725) and HLA/DR restriction profile are indicated. Some of the Th peptides identified (SEQ ID N. 1,2,6,8,11) contain CTL epitopes reported by others (underlined). Generation of CD4 T lines with peptide mixtures
CMV or pp65 can be used for the in vitro expansion of human CD4 and
CD8 lymphocytes (14, 22), but the antigens produced for diagnostic purposes are not always therapeutically useful (cell expansion for the purpose of in vivo reinfusion). On the contrary, peptides can be easily synthesized as GMP certified products and used accordingly.
We have shown that the identified peptides not only activate CD4 lymphocyte in terms of IFNγ secretion, but also promote their in vitro proliferation and expansion.
The peptides reported in the Table were pooled and used for stimulating PBMC. A representative example isv shown in Fig. 5, where PBMCs from a donor responded to peptides 30, 45, 62, 128, consistently with the donor's HLA DR13(17), DR7 alleles. Notably, the responses to PPD (tuberculin, prime-boost antigen) and to allogenic stimuli were evident in PBMC, as expected. As shown in the right-hand panel, cells stimulated with CD4 peptide pool proved specific for immunodominant peptides, with loss of the component specific for PPD and allostimulators after one in vitro stimulation- expansion cycle. CMV and pp65 were recognized, indicating that peptide stimulation had expanded proliferating CD4 cells able to recognize the relevant epitopes in the context of viral proteins, as expected in vivo. DESCRIPTION OF THE FIGURES
Fig. 1: proliferative responses to CMV and pp65.
Fig. 2: ELISPOT on PBMC with pρ65 peptides.
Fig. 3: Identification of stimulating pp65 peptides by ELISPOT. Results
of peptide screening are shown as the number of responding individuals out of 20 CMV-seropositive healthy donors.
Fig. 4: Discrimination between Th and CTL peptides. PBMC were depleted of CD4 and CD8 cells, giving selected CD8 and CD4 populations, respectively. Representative tests are shown: peptides 62 and 128 were recognized by CD4 cells from donor FI, peptide 57 by CD4 cells from donor CM. CD8 cells recognized peptides 123 and 104, containing the known CTL epitopes NV9 and TMlO, respectively.
Fig. 5: Use of a mixture of CD4 peptides for the generation of CD4 T cell lines. PBMC from donor FI were stimulated with peptides Th (Table). The cells cultured for three weeks were re-stimulated and tested against different antigens and alloantigens (right-hand panel), with respect to PBMC (left-hand panel). Proliferation in response to CMV and to pp65 was maintained.
Proliferation to peptides 30, 45, 62 and 128 was confirmed by expanded cells. PPD and alloreactive responses, clearly evident in PBMC, were absent in the peptide-expanded population.
NS pep: non-stimulating peptides, response to peptides which do not induce proliferation.
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Claims
1. Immunogenic peptides from Cytomegalovirus (CMV) pp65 protein, which are selected from the group consisting of: PLKMLNIPSINVHHY (SEQ ID N. 1)
TRQQNQWKEPDVYYT (SEQ ID N. 2)
EPDVYYTSAFVFPTK (SEQ ID N. 3)
KVYLESFCEDVPSGK (SEQ ID N. 4)
TLGSDVEEDLTMTRN (SEQ ID N. 5) QPFMRPHERNGFTVL (SEQ ID N. 6)
YRIQGKLEYRHTWDR (SEQ ID N. 7)
TERKTPRVTGGGAMA (SEQ ID N. 8)
ASTSAGRKRKSASSA (SEQ ID N. 9)
ACTSGVMTRGRLKAE (SEQ ID N. 10) AGILARNLVPMVATV (SEQ ID N. 11)
2. A nucleic acid molecule encoding a peptide according to claim 1.
3. A pharmaceutical composition containing a peptide according to claim 1 or a nucleic acid molecule according to claim 2, in association with pharmaceutically acceptable excipients.
4. A pharmaceutical composition according to claim 3, containing the peptides from SEQ ID N. 1 to SEQ ID N. 11.
5. A composition according to claims 3-4, which is in the form of vaccine.
6. Isolated Th lymphocytes able to recognize and bind a peptide according to claim 1.
7. Antigen presenting cells able to bind and present a peptide according to claim 1 in association with a HLA-class II molecule.
8. Antige presenting cells according to claim 7, which are selected from dendritic cells, monocytes-macrophages, B cells.
9. The use of a peptide according to claim 1, of a nucleic acid molecule according to claim 2, of Th lymphocytes according to claim 6 or of antigen presenting cells according to claim 7, for the preparation of a medicament or vaccine for preventing, controlling or curing CMV infections.
10. The use according to claim 9, for the treatment of CMV-infected subjects, subjects at risk of exposure to CMV, CMV-seropositive subjects carrying the virus in latent form.
11. The use according to claims 9 and 10, of a mixture of peptides SEQ ID No. 1-11.
12. The use of one or more peptides according to claim 1, for the preparation of a diagnostic composition.
13. A method for the activation of Th cells against CMV which comprises exposing said cells in culture to peptides according to claim 1.
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Cited By (5)
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| EP3049092A4 (en) * | 2013-09-24 | 2017-09-06 | Duke University | Compositions, methods and kits for eliciting an immune response |
| US10611800B2 (en) | 2016-03-11 | 2020-04-07 | Pfizer Inc. | Human cytomegalovirus gB polypeptide |
| JP2020534874A (en) * | 2017-09-29 | 2020-12-03 | ヴァクディアグン バイオテクノロジー カンパニー リミテッドＶａｃｄｉａｇｎ Ｂｉｏｔｅｃｈｎｏｌｏｇｙ Ｃｏ．， Ｌｔｄ | Fusion peptide of CD4 helper T cell epitope and its vaccine |
| US11629172B2 (en) | 2018-12-21 | 2023-04-18 | Pfizer Inc. | Human cytomegalovirus gB polypeptide |
| US11857622B2 (en) | 2020-06-21 | 2024-01-02 | Pfizer Inc. | Human cytomegalovirus GB polypeptide |
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Cited By (9)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| EP3049092A4 (en) * | 2013-09-24 | 2017-09-06 | Duke University | Compositions, methods and kits for eliciting an immune response |
| US9950056B2 (en) | 2013-09-24 | 2018-04-24 | Duke University | Compositions, methods and kits for eliciting an immune response |
| US10611800B2 (en) | 2016-03-11 | 2020-04-07 | Pfizer Inc. | Human cytomegalovirus gB polypeptide |
| JP2020534874A (en) * | 2017-09-29 | 2020-12-03 | ヴァクディアグン バイオテクノロジー カンパニー リミテッドＶａｃｄｉａｇｎ Ｂｉｏｔｅｃｈｎｏｌｏｇｙ Ｃｏ．， Ｌｔｄ | Fusion peptide of CD4 helper T cell epitope and its vaccine |
| EP3715359A4 (en) * | 2017-09-29 | 2021-06-23 | Vacdiagn Biotechnology Co., Ltd. | Cd4 helper t cell epitope fusion peptide and vaccine therewith |
| US11352411B2 (en) | 2017-09-29 | 2022-06-07 | Vacdiagn Biotechnology Co., Ltd | Fusion peptides of CD4 helper T cell epitopes and vaccines thereof |
| JP7146926B2 (en) | 2017-09-29 | 2022-10-04 | ヴァクディアグン バイオテクノロジー カンパニー リミテッド | Fusion peptide of CD4 helper T cell epitope and its vaccine |
| US11629172B2 (en) | 2018-12-21 | 2023-04-18 | Pfizer Inc. | Human cytomegalovirus gB polypeptide |
| US11857622B2 (en) | 2020-06-21 | 2024-01-02 | Pfizer Inc. | Human cytomegalovirus GB polypeptide |
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