CD47 agonist peptides induce programmed cell death in refractory chronic lymphocytic leukemia B cells via PLCγ1 activation: evidence from mice and humans.
BACKGROUND
Chronic lymphocytic leukemia (CLL), the most common adulthood leukemia, is characterized by the accumulation of abnormal CD5+ B lymphocytes, which results in a progressive failure of the immune system. Despite intense research efforts, drug resistance remains a major cause of treatment failure in CLL, particularly in patients with dysfunctional TP53. The objective of our work was to identify potential approaches that might overcome CLL drug refractoriness by examining the pro-apoptotic potential of targeting the cell surface receptor CD47 with serum-stable agonist peptides.


RESULTS
In peripheral blood samples collected from 80 patients with CLL with positive and adverse prognostic features, we performed in vitro genetic and molecular analyses that demonstrate that the targeting of CD47 with peptides derived from the C-terminal domain of thrombospondin-1 efficiently kills the malignant CLL B cells, including those from high-risk individuals with a dysfunctional TP53 gene, while sparing the normal T and B lymphocytes from the CLL patients. Further studies reveal that the differential response of normal B lymphocytes, collected from 20 healthy donors, and leukemic B cells to CD47 peptide targeting results from the sustained activation in CLL B cells of phospholipase C gamma-1 (PLCγ1), a protein that is significantly over-expressed in CLL. Once phosphorylated at tyrosine 783, PLCγ1 enables a Ca2+-mediated, caspase-independent programmed cell death (PCD) pathway that is not down-modulated by the lymphocyte microenvironment. Accordingly, down-regulation of PLCγ1 or pharmacological inhibition of PLCγ1 phosphorylation abolishes CD47-mediated killing. Additionally, in a CLL-xenograft model developed in NOD/scid gamma mice, we demonstrate that the injection of CD47 agonist peptides reduces tumor burden without inducing anemia or toxicity in blood, liver, or kidney. The limitations of our study are mainly linked to the affinity of the peptides targeting CD47, which might be improved to reach the standard requirements in drug development, and the lack of a CLL animal model that fully mimics the human disease.


CONCLUSIONS
Our work provides substantial progress in (i) the development of serum-stable CD47 agonist peptides that are highly effective at inducing PCD in CLL, (ii) the understanding of the molecular events regulating a novel PCD pathway that overcomes CLL apoptotic avoidance, (iii) the identification of PLCγ1 as an over-expressed protein in CLL B cells, and (iv) the description of a novel peptide-based strategy against CLL.
Cell Death and Drug Resistance in Lymphoproliferative Disorders Team, INSERM UMRS1138, Centre de Recherche des Cordeliers, Paris, France; Sorbonne Universités, Université Pierre et Marie Curie Paris 6, UMRS1138, Centre de Recherche des Cordeliers, Paris, France; Université Paris Descartes, Sorbonne Paris Cité, UMRS1138, Centre de Recherche des Cordeliers, Paris, France.
Ana-Carolina
AC
Martinez-Torres
Cell Death and Drug Resistance in Lymphoproliferative Disorders Team, INSERM UMRS1138, Centre de Recherche des Cordeliers, Paris, France; Sorbonne Universités, Université Pierre et Marie Curie Paris 6, UMRS1138, Centre de Recherche des Cordeliers, Paris, France; Université Paris Descartes, Sorbonne Paris Cité, UMRS1138, Centre de Recherche des Cordeliers, Paris, France.
Claire
C
Quiney
INSERM U1149, Paris, France; Faculté de Médecine, Site Xavier Bichat, Université Paris Diderot, Sorbonne Paris Cité, Paris, France.
Tarik
T
Attout
Laboratoire des Biomolécules, UMR 7203 and FR 2769, Sorbonne Universités, Université Pierre et Marie Curie, Paris, France; Centre National de la Recherche Scientifique, UMR 7203, Paris, France; Département de Chimie, École Normale Supérieure, Paris, France.
Heloïse
H
Boullet
Cell Death and Drug Resistance in Lymphoproliferative Disorders Team, INSERM UMRS1138, Centre de Recherche des Cordeliers, Paris, France; Sorbonne Universités, Université Pierre et Marie Curie Paris 6, UMRS1138, Centre de Recherche des Cordeliers, Paris, France; Université Paris Descartes, Sorbonne Paris Cité, UMRS1138, Centre de Recherche des Cordeliers, Paris, France.
Linda
L
Herbi
Cell Death and Drug Resistance in Lymphoproliferative Disorders Team, INSERM UMRS1138, Centre de Recherche des Cordeliers, Paris, France; Sorbonne Universités, Université Pierre et Marie Curie Paris 6, UMRS1138, Centre de Recherche des Cordeliers, Paris, France; Université Paris Descartes, Sorbonne Paris Cité, UMRS1138, Centre de Recherche des Cordeliers, Paris, France.
Laura
L
Vela
Cell Death and Drug Resistance in Lymphoproliferative Disorders Team, INSERM UMRS1138, Centre de Recherche des Cordeliers, Paris, France; Sorbonne Universités, Université Pierre et Marie Curie Paris 6, UMRS1138, Centre de Recherche des Cordeliers, Paris, France; Université Paris Descartes, Sorbonne Paris Cité, UMRS1138, Centre de Recherche des Cordeliers, Paris, France.
Sandrine
S
Barbier
Sorbonne Universités, Université Pierre et Marie Curie Paris 6, UMRS1138, Centre de Recherche des Cordeliers, Paris, France; Université Paris Descartes, Sorbonne Paris Cité, UMRS1138, Centre de Recherche des Cordeliers, Paris, France; Intestine: Nutrition, Barrier, and Diseases Team, INSERM U1138, Centre de Recherche des Cordeliers, Paris, France.
Danielle
D
Chateau
Cell Death and Drug Resistance in Lymphoproliferative Disorders Team, INSERM UMRS1138, Centre de Recherche des Cordeliers, Paris, France; Sorbonne Universités, Université Pierre et Marie Curie Paris 6, UMRS1138, Centre de Recherche des Cordeliers, Paris, France; Service d'Hématologie Biologique, Groupe Hospitalier Pitié-Salpêtrière, Assistance Publique-Hôpitaux de Paris, Paris, France.
Elise
E
Chapiro
Cell Death and Drug Resistance in Lymphoproliferative Disorders Team, INSERM UMRS1138, Centre de Recherche des Cordeliers, Paris, France; Sorbonne Universités, Université Pierre et Marie Curie Paris 6, UMRS1138, Centre de Recherche des Cordeliers, Paris, France; Service d'Hématologie Biologique, Groupe Hospitalier Pitié-Salpêtrière, Assistance Publique-Hôpitaux de Paris, Paris, France.
Florence
F
Nguyen-Khac
Cell Death and Drug Resistance in Lymphoproliferative Disorders Team, INSERM UMRS1138, Centre de Recherche des Cordeliers, Paris, France; Sorbonne Universités, Université Pierre et Marie Curie Paris 6, UMRS1138, Centre de Recherche des Cordeliers, Paris, France; Service d'Hématologie Biologique, Groupe Hospitalier Pitié-Salpêtrière, Assistance Publique-Hôpitaux de Paris, Paris, France.
Frédéric
F
Davi
Cell Death and Drug Resistance in Lymphoproliferative Disorders Team, INSERM UMRS1138, Centre de Recherche des Cordeliers, Paris, France; Sorbonne Universités, Université Pierre et Marie Curie Paris 6, UMRS1138, Centre de Recherche des Cordeliers, Paris, France; Service d'Hématologie Biologique, Groupe Hospitalier Pitié-Salpêtrière, Assistance Publique-Hôpitaux de Paris, Paris, France.
Magali
M
Le Garff-Tavernier
Laboratoire des Biomolécules, UMR 7203 and FR 2769, Sorbonne Universités, Université Pierre et Marie Curie, Paris, France; Centre National de la Recherche Scientifique, UMR 7203, Paris, France; Département de Chimie, École Normale Supérieure, Paris, France.
Roba
R
Moumné
Immunoregulation Laboratory, Centre de Recherche du Centre Hospitalier de l'Université de Montréal, Montréal, Quebec, Canada.
Marika
M
Sarfati
Laboratoire des Biomolécules, UMR 7203 and FR 2769, Sorbonne Universités, Université Pierre et Marie Curie, Paris, France; Centre National de la Recherche Scientifique, UMR 7203, Paris, France; Département de Chimie, École Normale Supérieure, Paris, France.
Philippe
P
Karoyan
Cell Death and Drug Resistance in Lymphoproliferative Disorders Team, INSERM UMRS1138, Centre de Recherche des Cordeliers, Paris, France; Sorbonne Universités, Université Pierre et Marie Curie Paris 6, UMRS1138, Centre de Recherche des Cordeliers, Paris, France; Service d'Hématologie Biologique, Groupe Hospitalier Pitié-Salpêtrière, Assistance Publique-Hôpitaux de Paris, Paris, France.
Hélène
H
Merle-Béral
INSERM U1149, Paris, France; Faculté de Médecine, Site Xavier Bichat, Université Paris Diderot, Sorbonne Paris Cité, Paris, France.
Pierre
P
Launay
Cell Death and Drug Resistance in Lymphoproliferative Disorders Team, INSERM UMRS1138, Centre de Recherche des Cordeliers, Paris, France; Sorbonne Universités, Université Pierre et Marie Curie Paris 6, UMRS1138, Centre de Recherche des Cordeliers, Paris, France; Université Paris Descartes, Sorbonne Paris Cité, UMRS1138, Centre de Recherche des Cordeliers, Paris, France.
Santos A
SA
Susin
D000368:Aged; D000369:Aged, 80 and over; D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D001402:B-Lymphocytes / Q000378:metabolism*; D051928:CD47 Antigen / Q000378:metabolism*; D019008:Drug Resistance, Neoplasm; D005260:Female; D006801:Humans; D015451:Leukemia, Lymphocytic, Chronic, B-Cell / Q000097:blood / Q000188:drug therapy / Q000378:metabolism*; D008297:Male; D051379:Mice; D016688:Mice, Inbred NOD; D008875:Middle Aged; D010455:Peptides / Q000494:pharmacology* / Q000627:therapeutic use; D051966:Phospholipase C gamma / Q000378:metabolism*; D019700:Thrombospondin 1 / Q000627:therapeutic use
D051928:CD47 Antigen; D010455:Peptides; D019700:Thrombospondin 1; D051966:Phospholipase C gamma