Contributions
Type: Publication Only
Background
AML is a heterogeneous disease at both the phenotypic and molecular level with a variety of distinct genetic alterations giving rise to the disease. This heterogeneity extends to the leukemic stem cell (LSC), with this dynamic compartment evolving to overcome various selection pressures imposed upon it during disease progression. Since LSC are thought to be resistant to current chemotherapeutic regimens and mediate disease relapse, their study may have profound clinical implications. Various markers have been described to characterize the LSC. CD81 antigen belongs to the tetraspanin family (33 members in mammals) which are cell surface transmembrane proteins and may be involved in the re-entry of hematopoietic stem cells into quiescence.
Aims
In this project, we plan to investigate the role of CD81 in the heterogeneity of AML, its potential to induce tumor dormancy and if this is characterized by resistance to chemotherapy.
Methods
In order, to study CD81 expression on primary leukemic blasts, we designed a MFC panel (CD36, CD81, CD33, CD90, CD123, CD34, CD38, CD45).Cell surface marker expression was measured in fresh or thawed bone marrow samples from adult patients with AML at diagnosis (n=122) or at relapse (n=10) using the above mentioned panel. This is a retrospective study to test the prognostic value of CD81, in comparison with other markers of LSC/HSC, for survival (overall survival, event free survival, relapse free survival). AML was diagnosed between 2010 and 2013 at the CHRU of Lille. To further study the function of CD81, we will use AML xenografts. Blasts obtained from newly diagnosed AML, were FACS sorted based upon high CD45 intensity and were then directly injected into NSG mice. After engraftment, the mice were sacrificed, and the bone marrow and spleen were analyzed by flow cytometry. Serial engraftment is performed by injecting one part of the blast cells into a subsequent NSG mouse.
Results
We performed the analysis on 122 diagnosis bone marrow samples and we found that 35% of the samples were positive for CD81 expression. Classification by CD81 expression predicts OS and EFS in AML (EFS: p=0.012; OS: p=0.0066). 23 primary AMLs have been injected and 8 of them produced serial engraftments. Yet, we did not find any difference in phenotype between the AML xenografts and the AML diagnosis. However, the phenotype of the blasts differ between bone marrow and spleen (p= 0.027).
Summary
Our preliminary results show that CD81 may have an important role in AML as their expression on blast cells predicts worse clinical outcome (OS and EFS) in this pathology. Furthermore, we have put in place and phenotypically validated AML xenografts, and we will be able to study the functional role of CD81 in AML in the near future. Nevertheless, the impact of xenografts in general and in particular with regard to CD81 on prognosis will be studied once we have sufficient follow-up time and numbers of patients analyzed and injected into NSG mice.
Keyword(s): Acute myeloid leukemia, Leukemic stem cell, Survival prediction, Tetraspanin
Session topic: Publication Only
Type: Publication Only
Background
AML is a heterogeneous disease at both the phenotypic and molecular level with a variety of distinct genetic alterations giving rise to the disease. This heterogeneity extends to the leukemic stem cell (LSC), with this dynamic compartment evolving to overcome various selection pressures imposed upon it during disease progression. Since LSC are thought to be resistant to current chemotherapeutic regimens and mediate disease relapse, their study may have profound clinical implications. Various markers have been described to characterize the LSC. CD81 antigen belongs to the tetraspanin family (33 members in mammals) which are cell surface transmembrane proteins and may be involved in the re-entry of hematopoietic stem cells into quiescence.
Aims
In this project, we plan to investigate the role of CD81 in the heterogeneity of AML, its potential to induce tumor dormancy and if this is characterized by resistance to chemotherapy.
Methods
In order, to study CD81 expression on primary leukemic blasts, we designed a MFC panel (CD36, CD81, CD33, CD90, CD123, CD34, CD38, CD45).Cell surface marker expression was measured in fresh or thawed bone marrow samples from adult patients with AML at diagnosis (n=122) or at relapse (n=10) using the above mentioned panel. This is a retrospective study to test the prognostic value of CD81, in comparison with other markers of LSC/HSC, for survival (overall survival, event free survival, relapse free survival). AML was diagnosed between 2010 and 2013 at the CHRU of Lille. To further study the function of CD81, we will use AML xenografts. Blasts obtained from newly diagnosed AML, were FACS sorted based upon high CD45 intensity and were then directly injected into NSG mice. After engraftment, the mice were sacrificed, and the bone marrow and spleen were analyzed by flow cytometry. Serial engraftment is performed by injecting one part of the blast cells into a subsequent NSG mouse.
Results
We performed the analysis on 122 diagnosis bone marrow samples and we found that 35% of the samples were positive for CD81 expression. Classification by CD81 expression predicts OS and EFS in AML (EFS: p=0.012; OS: p=0.0066). 23 primary AMLs have been injected and 8 of them produced serial engraftments. Yet, we did not find any difference in phenotype between the AML xenografts and the AML diagnosis. However, the phenotype of the blasts differ between bone marrow and spleen (p= 0.027).
Summary
Our preliminary results show that CD81 may have an important role in AML as their expression on blast cells predicts worse clinical outcome (OS and EFS) in this pathology. Furthermore, we have put in place and phenotypically validated AML xenografts, and we will be able to study the functional role of CD81 in AML in the near future. Nevertheless, the impact of xenografts in general and in particular with regard to CD81 on prognosis will be studied once we have sufficient follow-up time and numbers of patients analyzed and injected into NSG mice.
Keyword(s): Acute myeloid leukemia, Leukemic stem cell, Survival prediction, Tetraspanin
Session topic: Publication Only