# US 10,371,705 B2 · Atomic description of immune complex that causes heparin-induced thrombocytopenia

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- Number: US 10,371,705 B2
- Document ID: US10371705B2
- Status: Granted
- Kind: B2
- Application number: 15/524,511
- Filed: 2015-11-05
- Published: 2018-01-25
- Granted: 2019-08-06
- Earliest claimed: 2014-11-06
- Assignees: The Trustees of the University of Pennsylvania
- Applicants: The Trustees of The University of Pennsylvania
- Inventors: Mark I. Greene; Douglas B. Cines; Zheng Cai; Zhiqiang Zhu
- Web page: https://patentgrep.com/patent/US10371705B2

## CPC

- G01N 33/6863: {Cytokines, i.e. immune system proteins modifying a biological response such as cell growth proliferation or differentiation, e.g. TNF, CNF, GM-CSF, lymphotoxin, MIF or their receptors}
- A61K 38/195: {Chemokines, e.g. RANTES}
- A61K 39/3955: {against proteinaceous materials, e.g. enzymes, hormones, lymphokines}
- C07K 14/522: {Alpha-chemokines, e.g. NAP-2, ENA-78, GRO-alpha/MGSA/NAP-3, GRO-beta/MIP-2alpha, GRO-gamma/MIP-2beta, IP-10, GCP-2, MIG, PBSF, PF-4, KC}
- C07K 16/24: against cytokines, lymphokines or interferons
- G01N 33/86: involving blood coagulating time {or factors, or their receptors}
- A61K 2039/505: {comprising antibodies}
- G01N 2333/522: Alpha-chemokines, e.g. NAP-2, ENA-78, GRO-alpha/MGSA/NAP-3, GRO-beta/MIP-2alpha, GRO-gamma/MIP-2beta, IP-10, GCP-2, MIG, PBSF, PF-4 or KC
- G01N 2400/40: Glycosaminoglycans, i.e. GAG or mucopolysaccharides, e.g. chondroitin sulfate, dermatan sulfate, hyaluronic acid, heparin, heparan sulfate, and related sulfated polysaccharides
- G01N 2800/222: Platelet disorders
- G01N 2800/50: Determining the risk of developing a disease
- G01N 2800/52: Predicting or monitoring the response to treatment, e.g. for selection of therapy based on assay results in personalised medicine; Prognosis
- G01N: INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES (measuring or testing apparatus or processes other than immunoassay, involving enzymes or microorganisms C12M, C12Q)
- A61K: PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES (devices or methods specially adapted for bringing pharmaceutical products into particular physical or administering forms A61J3/00; chemical aspects of, or use of materials for deodorisation of air, for disinfection or sterilisation, or for bandages, dressings, absorbent pads or surgical articles A61L; soap compositions C11D)
- C07K: PEPTIDES (peptides in foodstuffs A23; obtaining protein compositions for foodstuffs, working-up proteins for foodstuffs A23J; preparations for medicinal purposes A61K; peptides containing beta-lactam rings C07D; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, C07D; ergot alkaloids of the cyclic peptide type C07D519/02; macromolecular compounds having statistically distributed amino acid units in their molecules, i.e. when the preparation does not provide for a specific; but for a random sequence of the amino acid units, homopolyamides and block copolyamides derived from amino acids C08G69/00; macromolecular products derived from proteins C08H1/00; preparation of glue or gelatine C09H; single cell proteins, enzymes C12N; genetic engineering processes for obtaining peptides C12N15/00; compositions for measuring or testing processes involving enzymes C12Q; investigation or analysis of biological material G01N33/00)

## Abstract

The present invention provides a humanized antibody or antibody fragment comprising (a) a humanized light chain comprising 1) Complementarity Determining Region (CDR)-L1, the sequence of which is identical to the sequence of SEQ ID NO: 3; 2) CDR-L2, the sequence of which is identical to the sequence of SEQ ID NO: 4; and 3) CDR-L3, the sequence of which is identical to the sequence of SEQ ID NO: 5, and (b) a humanized heavy chain comprising 1) CDR-H1, the sequence of which is identical to the sequence of SEQ ID NO: 6; 2) CDR-H2, the sequence of which is identical to the sequence of SEQ ID NO: 7; and 3) CDR-H3, the sequence of which is identical to the sequence of SEQ ID NO: 8, as well as methods for treating, diagnosing, and monitoring the progression of HIT. The present invention also provides methods for assessing the antigenicity and ability to cause HIT of anionic anticoagulants. The present invention also provides a mutant protein which has the same amino acid sequence of a wild type PF4 monomer except that (i) at least one amino acid of the wild type PF4 monomer has been deleted, (ii) at least one amino acid of the wild type PF4 monomer has been replaced by another amino acid, or (iii) a combination of such changes has been made. The present invention also provides methods of treating or reducing the likelihood of HIT, treating angiogenesis, treating abnormal cell growth, or affecting coagulation pathologies that lead to thrombus formation, by administering such mutant proteins to a patient.

## Claims

### Claim 1

A mutant platelet factor 4 (PF4) monomer which has the same amino acid sequence of wild type PF4 monomer set forth in SEQ ID NO: 11 except that
(i) the amino acid at position 9 is serine, the amino acid at position 10 is cysteine, and the amino acid at position 11 is valine, or
(ii) the amino acid at position 9 is serine, the amino acid at position 10 is cysteine, the amino acid at position 11 is valine, and the amino acid at position 55 is arginine.

### Claim 2 (depends on 1)

The mutant platelet factor 4 (PF4) monomer of claim 1, wherein the amino acid sequence of the mutant PF4 monomer comprises SEQ ID NO:15.

### Claim 3 (depends on 1)

The mutant protein of claim 1, wherein the mutant platelet factor 4 (PF4) monomer interferes with wild type PF4 tetramer oligomerization.

### Claim 4

A mutant platelet factor 4 (PF4) monomer which has the same amino acid sequence of wild type PF4 monomer set forth in SEQ ID NO: 11 except that
(i) the amino acid at position 9 is serine, the amino acid at position 10 is cysteine, and the amino acid at position 11 is valine; or
(ii) the amino acid at position 9 is serine, the amino acid at position 10 is cysteine, the amino acid at position 11 is valine, and the amino acid at position 55 is arginine; and wherein the mutant PF4 monomer of part (i) or part (ii) further comprises up to 8 amino acids that are deleted or substituted from the N-terminal end of the PF4 monomer.

### Claim 5 (depends on 4)

The mutant platelet factor 4 (PF4) monomer of claim 4, wherein at least 5 amino acids are deleted or substituted from the N-terminal end of the PF4 monomer.