Pre-B cell receptor signaling in acute lymphoblastic leukemia (NIH R01CA137060)
Pre-B cell receptor signaling in acute lymphoblastic leukemia. B lymphocytes are not only the cells that produce antibodies as part of the human immune system, they are also the cell of origin in most cases of acute lymphoblastic leukemia (ALL). ALL represents by far the most frequent type of cancer in children and is also a common disease in adults. For many years, patients with ALL are treated with chemotherapy and current treatment protocols lead to cure rates of 80 percent for children and 55 percent for adult patients with ALL. Our goal is to better understand the biology of human ALL, namely as a catastrophic aberration of normal B lymphocyte development. During normal B lymphocyte development, the pre-B cell receptor represents a critical signaling unit that guides early B lymphocyte precursors on their path of maturation. If signaling from the pre-B cell receptor is compromised, as for instance in patients with innate immune defects, the B lymphocyte precursors are arrested in their development at a primitive stage -as in ALL cells. Therefore, we propose to investigate the function of the pre-B cell receptor signaling unit (1) as a potential target to disrupt aberrant cell signaling that promotes leukemic growth and (2) to restore normal pre-B cell receptor signaling in the leukemia cells.
AID-mediated genetic instability in B cell lineage leukemia (NIH R01CA139032)
Recent work by our group demonstrated that the oncogenic BCR-ABL1 kinase in Ph+ ALL and blast crisis, but not chronic phase CML, induces expression of a mutator enzyme, termed AID (Activation-induced Cytidine Deaminase). The mutations that confer drug-resistance in Ph+ ALL and blast crisis CML can indeed be explained by activity of the AID enzyme. Additional experiments showed that engineered expression of AID in AID-negative chronic phase CML cells introduces the same mutations that cause drug-resistance in patients with AID-positive Ph+ ALL and CML blast crisis. Based on these observations, we propose three series of experiments to address the following questions: (1) Is the AID enzyme required for drug-resistance in AID-positive Ph+ ALL? To test this hypothesis, we will investigate whether BCR-ABL1-induced leukemia cells from mice carrying a deletion of the AID-gene fail to develop drug-resistance. (2) Does the AID enzyme play a critical role in the progression of chronic phase CML into blast crisis? Chronic phase CML can be treated very successfully for many years, whereas blast crisis represents a final and often fatal stage of the disease. (3) Which are the factors that induce aberrant expression of the AID enzyme in Ph+ ALL and blast crisis CML? The identification of such factors will likely help to understand how expression of this deleterious mutator enzyme can be prevented.
Infectious origins of childhood leukemia (NIH R01CA157664)
Childhood leukemia typically arises from a prenatally established pre-leukemic clone (Greaves and Wiemels, 2003). Genetic abnormalities that were acquired in utero are detected in cord blood samples, Guthrie cards and concordant twins who developed leukemia after a variable postnatal latency of up to 14 years. This led to a scenario, in which the initial prenatal lesion represents a first, albeit insufficient hit, which is followed by a series of additional transforming events, which ultimately cause leukemia (Greaves, 2009). Strikingly, some of these prenatal lesions define childhood leukemia subtypes yet are commonly detected in cord blood and on Guthrie cards from healthy individuals who will never develop disease. For instance, the TEL-AML1 gene rearrangement was detected in 6 of 567 (~1%) cord blood samples but leads to overt leukemia in less than 1% of these cases (with a cumulative risk of TEL- AML1 ALL at 1:14,000; Mori et al., 2004). These findings strongly support the notion that covert pre-leukemic clones are frequent but only a small minority of these pre-leukemic clones develop into frank leukemia after they acquired critical secondary genetic lesions. However, the postnatal mechanism(s) that drive the evolution of the fetal pre-leukemic clone towards childhood leukemia remain elusive and are at the center of this proposal. Since only a small minority of fetal pre-leukemic clones give rise to leukemia after birth, it appears critical to understand which mechanism(s) influence the clonal evolution of these pre-leukemic cells. This information will be highly significant, because control over these mechanisms could make childhood ALL a largely preventable disease.
Role of BCL6 in leukemia stem cell survival (NIH R21CA152497)
Current cytotoxic therapy approaches target proliferating bulk leukemia cells rather than quiescent leukemia stem cells. We now discovered that BCL6, a factor known to play a central role in B cell lymphomas, also plays a key role in the maintenance of leukemia stem cells. Since leukemia stem cells represent the origin of relapse and drug-resistance in leukemia in many cases, the identification of BCL6 as a target for leukemia stem cell eradication holds great promise. BCL6 is a master regulatory factor that controls the production of many different important genes. BCL6 was not previously known to be involved in leukemias. In preliminary studies for this proposal, we have discovered aberrant expression of BCL6 as a central component of a fundamentally novel pathway of leukemia stem cell self-renewal and drug-resistance in a wide array of human leukemias, some of which are still difficult to treat. In these leukemias, drug-treatment results in aberrant production of BCL6 by the leukemia cells, which appears to allow leukemia stem cell to self-renew and become resistance against drug-treatment. Recently a drug has been developed that can attach to BCL6 and block its cancer-causing activities. We found that this BCL6 inhibitor, which is called RI-BPI, has strong synergistic activity when combined with conventional drug- treatment, which opens up a powerful new therapeutic strategy for leukemia stem cell eradication through targeted inhibition of BCL6. Based on the discovery of BCL6 as a key component of a novel pathway of drug-resistance and stem cell self-renewal in a wide array of leukemias, we propose three Aims to develop these findings towards application in patient care: (1) To test the hypothesis that aberrant expression of BCL6 in human leukemia cells promotes leukemia stem cell survival, (2) To determine the frequency and phenotype of BCL6-dependent leukemia stem cells in human B cell ALL and (3) To validate a the role of the BCL6 inhibitor RI-BPI as a therapy for targeted eradication of leukemia stem cells.
Duy C, Hurtz C, Shojaee S, Cerchietti L, Geng H, Klemm L, Kim YM, Jumaa H, Koeffler HP, Yu JJ, Heisterkamp N, Graeber TG, Wu H, Ye BH, Melnick A & Müschen M. BCL6 enables Ph+ acute lymphoblastic leukemia cells to survive BCR-ABL1 kinase inhibition. Nature 473: 384-388 (2011) Download Paper
Nahar R, Ramezani-Rad P, Mossner M, Duy C, Cerchietti L, Geng H, Dovat S, Jumaa H, Ye BH, Melnick A & Müschen M. Pre-B cell receptor-mediated activation of BCL6 induces pre-B cell quiescence through transcriptional repression of MYC. Blood. In press (2011).
Hurtz C, Hatzi K, Cerchietti L, Braig M, Park E, Kim YM, Herzog S, Ramezani-Rad P, Jumaa H, Müller MC, Hofmann WK, Hochhaus A, Ye BH, Agarwal A, Druker BJ, Shah NP, Melnick AM & Müschen M. BCL6-mediated repression of p53 is critical for leukemia stem cell survival in chronic myeloid leukemia. J Exp Med. In press (2011)
Park E, Gang EJ, Hsieh YT, Schaefer P, Chae S, Klemm L, Huantes S, Loh M, Conway EM, Kang ES, Koo HH, Hofmann WK, Heisterkamp N, Pelus L, Keerthivasan G, Crispino J, Kahn M, Müschen M & Kim YM. Targeting survivin overcomes drug resistance in acute lymphoblastic leukemia. Blood. In press (2011)
Rubbi L, Titz B, Brown L, Galvan E, Komisopoulou E, Chen SS, Low T, Tahmasian M, Skaggs B, Müschen M, Pellegrini M & Graeber TG. Global phosphoproteomics reveals crosstalk between Bcr-Abl and negative feedback mechanisms controlling Src signaling. Science Signaling 4: ra18 (2011)
Müschen M. Genetic relicts from the origin of ALL. Blood 115: 3424-3425 (2010)
Duy C, Yu J, Polo JM, Klemm L, Cerchietti L, Melnick A, Ye BH & Müschen M. BCL6 is critical for the development of a diverse primary B cell repertoire. J Exp Med, 207: 1209-1221 (2010)
Gruber TA, Chang MS, Sposto R & Müschen M. Activation-induced cytidine deaminase accelerates clonal evolution in BCR-ABL1-driven acute lymphoblastic leukemia. Cancer Res. 70: 7411-7420 (2010)
Trageser D, Duy C, Klemm L, Gruber TA, Kim YM, Hofmann WK, Groffen J, Jäck HM, Jumaa H, Heisterkamp N & Müschen M. Pre-B cell receptor-mediated cell cycle arrest in Ph+ acute lymphoblastic leukemia requires IKAROS function. J Exp Med, 206: 1739-1753 (2009)
Klemm L, Duy C, Iacobucci I, Li Z, Feldhahn N, Henke N, Park E, Hoffmann TK, Kim YM, Hofmann WK, Jumaa H, Groffen J, Lieber MR, Casellas R & Müschen M. The B cell mutator AID promotes drug-resistance and B lymphoid blast crisis in chronic myeloid leukemia. Cancer Cell, 16: 232-245 (2009)
Tsai A, Lu H, Raghavan SC, Müschen M, Hsieh CL & Lieber MR. Human Chromosomal translocations at CpG sites and a theoretical basis for their lineage- and stage-specificity. Cell, 135:1130-42 (2008)
Feldhahn N, Henke N, Melchior K, Duy C, Soh BN, Klein F, von Levetzow G, Giebel B, Li A, Hofmann WK, Jumaa H & Müschen M. Activation-induced cytidine deaminase acts as a mutator in BCR-ABL1-transformed acute lymphoblastic leukemia cells. J Exp Med 204:1157-1166 (2007)
Meixlsperger S, Kohler F, Wossning T, Reppel M, Müschen M, Jumaa H. Conventional light chains inhibit the autonomous signaling capacity of the B cell receptor. Immunity 26:323-333 (2007)
Feldhahn N, Rio P, Soh BN, Liedtke S, Sprangers M, Klein F, Wernet P, Jumaa H, Hofmann WK, Hanenberg H, Rowley JD & Müschen M. Deficiency of Bruton's tyrosine kinase in B cell precursor leukemia cells. Proc Natl Acad Sci USA 102:13266-13271, 2005.
Feldhahn N, Klein F, Mooster JL, Hadweh P, Sprangers M, Wartenberg M, Bekhite MM, Hofmann WK, Herzog S, Jumaa H, Rowley JD & Müschen M. Mimicry of a constitutively active pre-B cell receptor in acute lymphoblastic leukemia cells. J Exp Med 201:1837-1852 (2005).
Klein F, Feldhahn N, Harder L, Wang H, Wartenberg M, Hofmann WK, Wernet P, Siebert R & Müschen M. The BCR-ABL1 kinase bypasses selection for the expression of a pre-B cell receptor in pre-B acute lymphoblastic leukemia cells. J Exp Med 199:673-685 (2004).
Klein F, Feldhahn N, Lee S, Wang H, Ciuffi F, von Elstermann M, Toribio ML, Sauer H, Wartenberg M, Barath VS, Kronke M, Wernet P, Rowley JD & Müschen M. T lymphoid differentiation in human bone marrow. Proc Natl Acad Sci USA 100:6747-6752 (2003).