# US 12,006,352 B2 · Techniques for predicting, detecting and reducing aspecific protein interference in assays involving immunoglobulin single variable domains

> Agents: this data is available as JSON from the patentgrep API and MCP server — see https://patentgrep.com/llms.txt

- Number: US 12,006,352 B2
- Document ID: US12006352B2
- Status: Granted
- Kind: B2
- Application number: 16/823,412
- Filed: 2020-03-19
- Published: 2020-10-15
- Granted: 2024-06-11
- Earliest claimed: 2011-06-23
- Assignees: Ablynx N.V.; Sanofi
- Applicants: Ablynx N.V.; Sanofi
- Inventors: Judith Baumeister; Marie-Paule Lucienne Armanda Bouche; Carlo Boutton; Marie-Ange Buyse; Veerle Snoeck; Stephanie Staelens; Bruno Dombrecht; Peter Schotte; Cedric Jozef Neotere Ververken; Gerald Beste; Guy Hermans; Soren Steffensen; Alexander Szyroki; Tinneke Denayer
- Web page: https://patentgrep.com/patent/US12006352B2

## CPC

- C07K 16/00: Immunoglobulins [IG], e.g. monoclonal or polyclonal antibodies
- A61P 37/04: Immunostimulants
- C07K 16/18: against material from animals or humans
- A61K 39/39533: {against materials from animals}
- A61K 2039/505: {comprising antibodies}
- C07K 16/28: against receptors, cell surface antigens or cell surface determinants
- C07K 16/2875: {against the NGF/TNF superfamily, e.g. CD70, CD95L, CD153, CD154 (against NGF C07K16/22, against TNF C07K16/241)}
- C07K 16/42: against immunoglobulins
- C07K 16/4283: {against an allotypic or isotypic determinant on Ig}
- C07K 2317/22: from camelids, e.g. camel, llama or dromedary
- C07K 2317/31: multispecific
- C07K 2317/34: Identification of a linear epitope shorter than 20 amino acid residues or of a conformational epitope defined by amino acid residues
- C07K 2317/35: Valency
- C07K 2317/567: Framework region [FR]
- C07K 2317/569: Single domain, e.g. dAb, sdAb, VHH, VNAR or nanobody®
- C07K 2317/92: Affinity (KD), association rate (Ka), dissociation rate (Kd) or EC50 value
- C07K 2317/94: Stability, e.g. half-life, pH, temperature or enzyme-resistance
- C07K 2319/30: Non-immunoglobulin-derived peptide or protein having an immunoglobulin constant or Fc region, or a fragment thereof, attached thereto
- G01N 33/5306: {Improving reaction conditions, e.g. reduction of non-specific binding, promotion of specific binding}
- G01N 33/54393: {Improving reaction conditions or stability, e.g. by coating or irradiation of surface, by reduction of non-specific binding, by promotion of specific binding}
- G01N 33/6854: {Immunoglobulins}
- G01N 33/6857: {Antibody fragments}
- 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)
- A61P: SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- 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)
- 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)

## Abstract

This invention provides, and in certain specific but non-limiting aspects relates to: assays that can be used to predict whether a given ISV will be subject to protein interference as described herein and/or give rise to an (aspecific) signal in such an assay (such as for example in an ADA immunoassay). Such predictive assays could for example be used to test whether a given ISV could have a tendency to give rise to such protein interference and/or such a signal; to select ISV's that are not or less prone to such protein interference or to giving such a signal; as an assay or test that can be used to test whether certain modification(s) to an ISV will (fully or partially) reduce its tendency to give rise to such interference or such a signal; and/or as an assay or test that can be used to guide modification or improvement of an ISV so as to reduce its tendency to give rise to such protein interference or signal;—methods for modifying and/or improving ISV's to as to remove or reduce their tendency to give rise to such protein interference or such a signal;—modifications that can be introduced into an ISV that remove or reduce its tendency to give rise to such protein interference or such a signal; ISV's that have been specifically selected (for example, using the assay(s) described herein) to have no or low(er)/reduced tendency to give rise to such protein interference or such a signal; modified and/or improved ISV's that have no or a low(er)/reduced tendency to give rise to such protein interference or such a signal.

## Claims

### Claim 1

A polypeptide comprising a serum albumin binding immunoglobulin single variable domain (VH or VHH) at its C-terminal end, wherein the immunoglobulin single variable domain ends with the sequence VTVSS(X) n (SEQ ID NO: 34), in which n is 1 to 5, with each X being independently chosen from any amino acid, with the proviso that X is not cysteine (C); and wherein the immunoglobulin single variable domain:
(i) is not Alb-1; and
(ii) does not end with the sequence VTVSSA (SEQ ID NO: 90).

### Claim 2 (depends on 1)

The polypeptide of claim 1, in which:
(a) n is 2 or 3 in which each X is alanine (A) or glycine (G); or
(b) n is 2 or 3 in which each X is alanine (A); or
(c) n is 1, 2 or 3 in which each X is G glycine (G); or
(d) n is 2 or 3 in which at least one X is alanine (A) or glycine (G), with any remaining amino acid residue X being independently chosen from any naturally-occurring amino acid; or
(e) n is 2 or 3 in which all but one X is alanine (A) or glycine (G), with any remaining amino acid residue X being independently chosen from any naturally- occurring amino acid.

### Claim 3 (depends on 2)

The polypeptide of claim 2, in which in (d) or (e) the remaining amino acid residue X is independently chosen from the group consisting of alanine (A), glycine (G), valine (V), leucine (L) and isoleucine (I).

### Claim 4 (depends on 1)

The polypeptide of claim 1, wherein n is 1 or 2.

### Claim 5 (depends on 1)

The polypeptide of claim 1, wherein each X is a naturally-occurring amino acid.

### Claim 6 (depends on 1)

The polypeptide of claim 1, wherein each X is independently chosen from the group consisting of alanine (A), glycine (G), valine (V), leucine (L) and isoleucine (I).