Therapeutic Potential and Challenges of Targeting Receptor Tyrosine Kinase ROR1 with Monoclonal Antibodies in B-Cell Malignancies

Yang, Jiahui; Baskar, Sivasubramanian; Kwong, Ka Yin; Kennedy, Michael G.; Wiestner, Adrian; Rader, Christoph

doi:10.1371/journal.pone.0021018

Cited by 85 publications

(96 citation statements)

References 45 publications

(76 reference statements)

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“…The identified functional epitopes of ROR1 in CLL will be further used in the development of human antibodies against ROR1 for clinical trials. The efficacy of our MAbs are in contrast to those produced by Yang et al 51 indicating that their MAbs might be more suitable as conjugates or chimeric antigen receptors. Their antibodies alone did not induce apoptosis, or mediated CDC and a low ADCC activity.…”

Section: Cd5contrasting

confidence: 60%

Monoclonal antibodies against ROR1 induce apoptosis of chronic lymphocytic leukemia (CLL) cells

Hojjat‐Farsangi

Khan

et al. 2012

Leukemia

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ROR1 is a receptor tyrosine kinase (RTK) recently identified to be overexpressed at the gene and protein levels in chronic lymphocytic leukemia (CLL). Monoclonal antibodies (MAbs) against RTKs have been successfully applied for therapy of solid tumors. We generated five MAbs against the Ig (n ¼ 1), cysteine-rich (CRD) (n ¼ 2) and kringle (KNG) (n ¼ 2) domains, respectively, of the extracellular part of ROR1. All CLL patients (n ¼ 20) expressed ROR1 on the surface of the leukemic cells. A significantly higher frequency of ROR1 expression was found in patients with progressive versus non-progressive disease, and in those with unmutated versus mutated IgVH genes. All five MAbs alone induced apoptosis in the absence of complement or added effector cells (Annexin-V and MTT, as well as cleavage of poly-(ADP ribose)-polymerase, caspase-8 and caspase-9) of CLL cells but not of normal B cells. Most effective were MAbs against CRD and KNG, significantly superior to rituximab (Po0.005). Cross-linking of anti-ROR1 MAbs using the F(ab 0 ) 2 fragments of anti-Fc antibodies significantly augmented apoptosis. Two of the MAbs induced complement-dependent cytotoxicity (CDC) similar to that of rituximab and one anti-ROR1 MAb (KNG) (IgG1) showed killing activity by antibody-dependent cellular cytotoxicity. The identified ROR1 epitopes may provide a basis for generating human ROR1 MAbs for therapy.

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Section: Cd5contrasting

confidence: 60%

Monoclonal antibodies against ROR1 induce apoptosis of chronic lymphocytic leukemia (CLL) cells

Hojjat‐Farsangi

Khan

et al. 2012

Leukemia

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“…In a similar series of screens, we humanized a mouse (2A2) 28 and a rabbit (R11) 37 mAb specific for the cancer cell-associated surface marker ROR1. 30,37,38 Here, we used sets of humanized variable regions designed by grafting of CDRs into framework regions derived from a database of somatically hypermutated human sequences, and generated transposable libraries as described before.…”

Section: Transpo-mab Screening Of Humanized Anti-ror1 Antibody Librariesmentioning

confidence: 99%

“…30,37,38 Here, we used sets of humanized variable regions designed by grafting of CDRs into framework regions derived from a database of somatically hypermutated human sequences, and generated transposable libraries as described before. The humanized libraries consisted of 64 VH £ 49 VL and 101 VH £ 82 VL for 2A2 and R11, respectively ( Fig.…”

Section: Transpo-mab Screening Of Humanized Anti-ror1 Antibody Librariesmentioning

confidence: 99%

“…Parental anti-ROR1 mouse mAb 2A2 28 and rabbit mAb R11, 37 as well as anti-mesothelin mouse mAb MN, 60 were produced as chimeric, full-length IgG1 antibodies with human constant regions as follows: Variable region coding regions were produced by total gene synthesis (GenScript) using MNFGLRLIFLVLTLKGVQC as leader sequence, and were assembled with human IgH-g 1 and IgL-k constant regions in the expression vector pCB14b. This vector, a derivative of the episomal mammalian expression vector pCEP4 (Invitrogen), carries the Epstein-Barr virus (EBV) replication origin, encodes the EBV nuclear antigen (EBNA-1) to permit extrachromosomal replication, and contains a puromycin selection marker in place of the original hygromycin B resistance gene.…”

Section: Antibody Productionmentioning

confidence: 99%

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Transpo-mAb display: Transposition-mediated B cell display and functional screening of full-length IgG antibody libraries

Waldmeier

Hellmann

Gutknecht

et al. 2016

mAbs

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In vitro antibody display and screening technologies geared toward the discovery and engineering of clinically applicable antibodies have evolved from screening artificial antibody formats, powered by microbial display technologies, to screening of natural, full-IgG molecules expressed in mammalian cells to readily yield lead antibodies with favorable properties in production and clinical applications. Here, we report the development and characterization of a novel, next-generation mammalian cell-based antibody display and screening platform called Transpo-mAb Display, offering straightforward and efficient generation of cellular libraries by using non-viral transposition technology to obtain stable antibody expression. Because Transpo-mAb Display uses DNA-transposable vectors with substantial cargo capacity, genomic antibody heavy chain expression constructs can be utilized that undergo the natural switch from membrane bound to secreted antibody expression in B cells by way of alternative splicing of Ig-heavy chain transcripts from the same genomic expression cassette. We demonstrate that stably transposed cells co-express transmembrane and secreted antibodies at levels comparable to those provided by dedicated constructs for secreted and membrane-associated IgGs. This unique feature expedites the screening and antibody characterization process by obviating the need for intermediate sequencing and re-cloning of individual antibody clones into separate expression vectors for functional screening purposes. In a series of proof-of-concept experiments, we demonstrate the seamless integration of antibody discovery with functional screening for various antibody properties, including binding affinity and suitability for preparation of antibody-drug conjugates.

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“…Nonetheless, all drugs currently on the market or in clinical trials are general immunosuppressants that fail to distinguish the pathogenic CLL cells from healthy B-cells. Ideally, drug targets would be identified that allow selective suppression or killing of the malignant CLL cells without interfering with the normal function of the humoral immune system (5).…”

Section: Chronic Lymphocytic Leukemia (Cll)mentioning

confidence: 99%

Targeting Stereotyped B Cell Receptors from Chronic Lymphocytic Leukemia Patients with Synthetic Antigen Surrogates

Sarkar

Liu

et al. 2016

Journal of Biological Chemistry

Self Cite

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Chronic lymphocytic leukemia (CLL) is a disease in which a single B-cell clone proliferates relentlessly in peripheral lymphoid organs, bone marrow, and blood. DNA sequencing experiments have shown that about 30% of CLL patients have stereotyped antigen-specific B-cell receptors (BCRs) with a high level of sequence homology in the variable domains of the heavy and light chains. These include many of the most aggressive cases that have IGHV-unmutated BCRs whose sequences have not diverged significantly from the germ line. This suggests a personalized therapy strategy in which a toxin or immune effector function is delivered selectively to the pathogenic B-cells but not to healthy B-cells. To execute this strategy, serum-stable, druglike compounds able to target the antigen-binding sites of most or all patients in a stereotyped subset are required. We demonstrate here the feasibility of this approach with the discovery of selective, high affinity ligands for CLL BCRs of the aggressive, stereotyped subset 7P that cross-react with the BCRs of several CLL patients in subset 7p, but not with BCRs from patients outside this subset.

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Therapeutic Potential and Challenges of Targeting Receptor Tyrosine Kinase ROR1 with Monoclonal Antibodies in B-Cell Malignancies

Cited by 85 publications

References 45 publications

Monoclonal antibodies against ROR1 induce apoptosis of chronic lymphocytic leukemia (CLL) cells

Monoclonal antibodies against ROR1 induce apoptosis of chronic lymphocytic leukemia (CLL) cells

Transpo-mAb display: Transposition-mediated B cell display and functional screening of full-length IgG antibody libraries

Targeting Stereotyped B Cell Receptors from Chronic Lymphocytic Leukemia Patients with Synthetic Antigen Surrogates

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