"B cell Chronic Lymphocytic Leukemia (CLL) is the most common leukemia in western countries. Although substantial therapeutic improvements have been obtained, the disease is incurable and causes 4000 deaths/year in the sole USA.We plan to in vivo evaluate CLL B cells division by measuring Deuterium (2H) incorporation into newly synthesized DNA. 2H given to patients in the form of deuterated water (2H2O) is a very practical, safe, non invasive approach that has been already exploited in CLL patients.Preliminary data obtained in 15 CLL patients using the 2H labeling approach are showing in each CLL clone two sub-populations that have 10 times different proliferation rates and can be identified using two well known surface markers, CXCR4 and CD5. The data also indicate that isolated resting cells express higher mRNA for genes found in hematopoietic stem cells (HSC) such as ABCB1, ABCB4 and Notch1. On the other hand proliferating cells express higher levels of known activation markers some of which (CD38, CD52, CD23, others) are target of monoclonal antibodies already in use.We want to refine the description of proliferating and resting cells within each clone by running a more extended immunophenotype and gene expression profile. The detailed description of highly resting and highly proliferating cells will provide new insights of the biology of the disease and a powerful tool for developing drugs aimed at target these fractions with high specificity.Moreover, RAG2-/-yc-/- mice will be used to test if highly defined resting and proliferating cells differ in their capacity to recapitulate the tumor. This mouse background has been successfully used for reconstituting a human hematopoietic system by transferring CD34+ cells and was found to support with high efficiency expansion of the CLL cell line MEC-1. We expect to find CLL resting cells to have a higher potential to recapitulate the tumor in mice model providing the first proof of a Leukemic Stem Cell in CLL"