Anti-CLL1 specific single-chain chimeric antigen receptors (scCARs) for cancer immunotherapy

A chimeric antigen receptor, specific technology, applied in the direction of antibody mimics/scaffolds, DNA/RNA fragments, the use of vectors to introduce foreign genetic material, etc., can solve the problem of no therapeutic effect

Active Publication Date: 2017-11-28
CELLECTIS SA
View PDF52 Cites 9 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, no therapeutic effect could be attributed to the CCL-1 targeting peptide itself

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • Anti-CLL1 specific single-chain chimeric antigen receptors (scCARs) for cancer immunotherapy
  • Anti-CLL1 specific single-chain chimeric antigen receptors (scCARs) for cancer immunotherapy
  • Anti-CLL1 specific single-chain chimeric antigen receptors (scCARs) for cancer immunotherapy

Examples

Experimental program
Comparison scheme
Effect test

Embodiment approach

[0424] According to another embodiment, the CLL1-specific chimeric antigen receptor (anti-CLL1 CAR) comprises an extracellular binding domain, wherein at least two epitopes are inserted in the extracellular domain in such a way that the VH and VL chains are located between them, all of these components are optionally separated by at least one linker.

[0425] In a preferred embodiment, the CLL1-specific chimeric antigen receptor (anti-CLL1 CAR) has one of the polypeptide structures selected from V1, V3 or V5, such as figure 2 As shown, the structure at least comprises an anti-CLL1 extracellular ligand-binding domain, a CD8α transmembrane domain, a 4-1BB co-stimulatory domain, a CD3ζ signaling domain, and two of the epitopes are inserted in this way In the extracellular domain, such that the VH and VL chains are located between them, all these components are optionally separated by at least one linker.

[0426] In a more preferred embodiment, the CLL1-specific chimeric antige...

Embodiment 1

[0683] Example 1: Proliferation of TCRα-inactivated cells expressing CLL1-scCAR

[0684] designed and produced a heterodimeric TALE nuclease that targets two 17-bp long sequences (termed half targets) that are expressed in the T cell receptor alpha constant chain region (TRAC) gene The 15-bp spacer (spacer) inside separates. Each half-target is recognized by the repeat sequence of the half-TALE nucleases listed in Table 10.

[0685] Table 10: TAL nucleases targeting the TCRα gene

[0686]

[0687] Each TALE nuclease construct was subcloned by restriction enzyme digestion in a mammalian expression vector under the control of the T7 promoter. The mRNA encoding the TALE nuclease-cleaved TRAC genomic sequence was synthesized from a plasmid carrying the coding sequence downstream of the T7 promoter.

[0688] Purified T cells preactivated with anti-CD3 / CD28 coated beads during 72 hours were transfected with each of the 2 mRNAs encoding the two half TRAC_T01 TALE nucleases. ...

Embodiment 2

[0691] Example 2: Construction of CLL1 scCAR using various anti-CLL1 antibody fragments

[0692] Primary T cell culture

[0693] Using Ficoll gradient density medium (Ficoll PaquePLUS / GE Healthcare Life Sciences), by EFS (Etablissement du Sang, Paris, France) to purify T cells from buffy coat samples. The PBMC layer was recovered and T cells were purified using a commercial T cell enrichment kit (Stem Cell Technologies). In X-Vivo supplemented with 20 ng / mL human IL-2 (MiltenyiBiotech), 5% human serum (Sera Laboratories), and Dynabeads human T activator CD3 / CD28, beads: cell ratio 1:1 (Life Technologies), TM -15 medium (Lonza) to activate purified T cells. After activation, in X-Vivo supplemented with 20ng / mL human IL-2 (Miltenyi Biotec) and 5% human serum (Sera Laboratories) TM Cells were grown and maintained in -15 medium (Lonza).

[0694] scCAR mRNA transfection

[0695] Transfection was performed on day 4 or 11 after T cell purification and activation. 5 million ...

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to view more

PUM

No PUM Login to view more

Abstract

The present invention relates to Chimeric Antigen Receptors (CAR) that are recombinant chimeric proteins able to redirect immune cell specificity and reactivity toward CLL1 positive cells. The engineered immune cells endowed with such CARs are particularly suited for immunotherapy for treating cancer, in particular leukemia.

Description

technical field [0001] C-type lectin-like molecule-1 (CLL1) has been identified as frequently overexpressed in most acute myeloid leukemia (AML) compared with normal hematopoietic stem cells or other cell types on stem cells (LSCs). The present invention relates to a method for targeting CLL1-positive malignant cells using a Chimeric Antigen Receptor (anti-CLL1-CAR), wherein the above-mentioned Chimeric Antigen Receptor is a recombinant chimeric protein capable of redirecting to a selected Immune cell specificity and reactivity to the membrane antigen CLL1. These anti-CLL1 CARs more specifically comprise extracellular ligand binding comprising scFV derived from some specific anti-CLL1 monoclonal antibodies. Engineered immune cells endowed with such CARs provide adoptive immunity against CLL1-positive cells as part of various cell therapies for more efficient treatment of cancer, especially blood cancer. Background technique [0002] Adoptive immunotherapy, which involves...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to view more

Application Information

Patent Timeline
no application Login to view more
Patent Type & Authority Applications(China)
IPC IPC(8): C07K19/00C12N15/62C12N5/10A61K35/17A61P35/02
CPCC07K14/7051C07K16/2866C07K16/3061C07K16/2803A61K2039/5158A61K2039/5156C07K2317/24C07K2317/622C07K2319/03C12N2510/00C07K14/70596C12N5/0636A61K39/0011A61P35/00C07K2319/02C07K2319/33A61K35/17C07K2319/00C12N5/0093C12N5/0638A61K2039/6056A61K2039/64A61P35/02A61P43/00A61K39/001176C07K16/28C07K16/30A61K39/395C07K14/70517C07K14/70578C07K19/00C12N5/10C12N15/63A61K39/39558A61K2039/505C07K14/70535C07K2317/53C07K16/2887C07K2317/56G01N15/1031G01N15/14G01N2015/008G01N2015/1006G01N2015/1081G01N2015/149
Inventor 朱莉安娜·史密斯朱利安·瓦尔顿亚历山大·朱耶拉特菲利普·迪谢托巴尔布拉·约翰逊·萨苏阿尔温德·拉吉帕尔
Owner CELLECTIS SA
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Try Eureka
PatSnap group products

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap