# US 10,946,098 B2 · Activatable two-component photosensitizers

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- Number: US 10,946,098 B2
- Document ID: US10946098B2
- Status: Granted
- Kind: B2
- Application number: 16/568,886
- Filed: 2019-09-12
- Published: 2020-03-12
- Granted: 2021-03-16
- Earliest claimed: 2014-11-17
- Assignees: Carnegie Mellon University
- Applicants: Carnegie Mellon University
- Inventors: Marcel P. Bruchez; Jianjun He; Yi Wang
- Web page: https://patentgrep.com/patent/US10946098B2

## CPC

- A61K 41/0057: {Photodynamic therapy with a photosensitizer, i.e. agent able to produce reactive oxygen species upon exposure to light or radiation, e.g. UV or visible light; photocleavage of nucleic acids with an agent}
- A61K 47/64: Drug-peptide, drug-protein or drug-polyamino acid conjugates, i.e. the modifying agent being a peptide, protein or polyamino acid which is covalently bonded or complexed to a therapeutically active agent (peptidic linkers A61K47/65)
- A61N 5/062: {Photodynamic therapy, i.e. excitation of an agent}
- A61P 35/00: Antineoplastic agents
- C07C 211/43: having amino groups bound to carbon atoms of six-membered aromatic rings of the carbon skeleton
- C07C 233/36: having the carbon atom of the carboxamide group bound to a hydrogen atom or to a carbon atom of an acyclic saturated carbon skeleton
- C07C 233/40: having the carbon atom of the carboxamide group bound to an acyclic carbon atom of a carbon skeleton containing six-membered aromatic rings
- C07K 14/001: {by chemical synthesis}
- C07K 14/71: for growth factors; for growth regulators
- C09B 11/10: Amino derivatives of triarylmethanes
- C09B 11/22: containing OH groups bound to an aryl nucleus {and their ethers and esters}
- A61N 2005/0659: {infrared}
- C07K 2317/622: Single chain antibody (scFv)
- C07K 2319/035: containing a signal for targeting to the external surface of a cell, e.g. to the outer membrane of Gram negative bacteria, GPI- anchored eukaryote proteins
- C07K 2319/30: Non-immunoglobulin-derived peptide or protein having an immunoglobulin constant or Fc region, or a fragment thereof, attached thereto
- 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)
- A61N: ELECTROTHERAPY; MAGNETOTHERAPY; RADIATION THERAPY; ULTRASOUND THERAPY (measurement of bioelectric currents A61B; surgical instruments, devices or methods for transferring non-mechanical forms of energy to or from the body A61B18/00; anaesthetic apparatus in general A61M; incandescent lamps H01K; infrared radiators for heating H05B)
- A61P: SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- C07C: ACYCLIC OR CARBOCYCLIC COMPOUNDS
- 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)
- C09B: ORGANIC DYES OR CLOSELY-RELATED COMPOUNDS FOR PRODUCING DYES {, e.g. PIGMENTS}; MORDANTS; LAKES (fermentation or enzyme using processes to synthesise a desired chemical compound C12P)

## Abstract

Provided herein is a two-component photosensitizer, which demonstrated robust and selective killing effects for transfected HEK cells and affibody targeted A431 cancer cells when exposed to near infrared light excitation. Free MG2I is a pure and stable fluorogen; it is easy to synthesize and modify, and has no toxicity to cells. Unlike conventional photosensitizers, the dye and FAP itself has no photosensitizing effect until they are bound. Also unlike other activation methods, the activation step is achieved by adding the fluorogen, not the presence of the targeted molecule, requiring an ‘active’ activation instead of a ‘passive’ activation. This method offers the ability to locally switch-on and selective generation of singlet oxygen at the target site and can be used for a wide variety of molecular targets.

## Claims

### Claim 1

A kit comprising:
a. a first vessel containing a heavy atom-modified malachite green derivative having the structure:
where X and X′ are, independently, Br, I, As, Se, Ga, Ge, or Sb, R1, R2, R3, R4, R5, R6, R7, R8, R9, and R10 are, independently H or F, R11, R12, R13 and R14 are, independently, methyl, H, aziridine or azetidine, wherein when R11, R12, R13, and/or R14 are aziridine or azetidine, R11 and R12 form a single ring and/or R13 and R14 form a single ring, and where R is selected from —OH, —COO−, —SO3−, —PO4−, —NO2, —NH2, —N(CH3)(R15), —OR16, alkyl, ether, polyether, PEG1_30, —(C1-C4 alkyl)-R17, heterocyles containing N, S or O atoms, substituted acetylenic groups, cyano, and carbohydrate groups, wherein R15 and R16 are: straight- or branched-chain alkyl; straight or branched-chain C1-6 alkyl; straight-chain or branched poly(C1-C4 alkyl amide); straight-chain or branched poly(C1-C4 alkyl amide) having from 2 to 6 amide moieties; poly(C1-C4 alkylene glycol); poly(C1-C4 alkylene glycol) having from 2 to 30 or from 2-10 C1-C4 alkylene glycol moieties; straight-chain or branched poly(C1-C4 alkyl amide):poly(C1-C4 alkylene glycol) diblock copolymer; straight-chain or branched poly(C1-C4 alkyl amide):poly(C1-C4 alkylene glycol) diblock copolymer having from 2 to 6 amide moieties and from 2 to 10 C1-C4 alkylene glycol moieties; sulfonyl- or bis-sulfonyl-terminated straight-chain or branched poly(C1-C4 alkyl amide); bis-taurine-terminated branched poly(C1-C4 alkyl amide); ethyl butyrate; C1-6 alkyl C1-6 alkanoate; —(CH2)n-C(O)—O—(CH2)m-CH3, where n=1-4 and m=0-3, and wherein R17 is selected from H, —OH, —COO—, —SO3—, —PO4—, —NO2, or —NH2; and
b. a targeting activator in the first vessel or in a second vessel, comprising a targeting moiety that selectively binds a target compound of a cell and an activator moiety that selectively binds the heavy atom-modified malachite green derivative having an excitation wavelength so that the heavy atom-modified malachite green derivative produces singlet oxygen when bound by the targeting activator and exposed to light at the excitation wavelength.

### Claim 2 (depends on 1)

The kit of claim 1, wherein R1, R2, R3, R4, R5, R6, R7, R8, R9, and R10 are H.

### Claim 3 (depends on 1)

The kit of claim 1, wherein X and X′ are independently Br or I.

### Claim 4 (depends on 1)

The kit of claim 1, wherein X and X′ are I.

### Claim 5 (depends on 1)

The kit of claim 1, wherein R11, R12, R13 and R14 are, independently, methyl or H, or R11, R12, R13 and R14 are methyl.

### Claim 6 (depends on 1)

The kit of claim 1, wherein R is —OR16, and R16 is ethylbutyrate.

### Claim 7 (depends on 1)

The kit of claim 1, wherein the heavy atom-modified malachite green derivative has having the structure:

### Claim 8 (depends on 1)

The kit of claim 1, wherein the targeting activator is a fusion protein of an scFv activator moiety and an affibody targeting moiety.

### Claim 9 (depends on 8)

The kit of claim 8, wherein the scFv is an L5-MG scFv peptide.

### Claim 10 (depends on 8)

The kit of claim 8, wherein the scFv is one of SEQ ID NOS: 1-4.

### Claim 11 (depends on 1)

The kit of claim 1, wherein the targeting moiety is selective for an epidermal growth factor receptor.

### Claim 12 (depends on 11)

The kit of claim 11, wherein the epidermal growth factor receptor is HER1 (human epidermal growth factor receptor 1) or HER2 (human epidermal growth factor receptor 2).

### Claim 13 (depends on 1)

The kit of claim 1, wherein the targeting activator comprises a sequence selected from SEQ ID NOS: 1-4, 10-15, 17 and 18.

### Claim 14 (depends on 1)

The kit of claim 1, wherein R is —OR16, and R16 is sulfonyl- or bis-sulfonyl-terminated straight-chain or branched poly(C1-C4 alkyl amide) having from 2 to 6 amide moieties.