Podcasts about calm af10

In this episode of the Epigenetics Podcast, we talked with Ani Deshpande from Sanford Burnham Prebys about his work on epigenetic regulation and developing small molecules through high throughput screens for AML. Throughout our discussion, we delve into Dr. Despande's journey into the field of biology and science, tracing his evolution from a literature enthusiast in Mumbai to a dedicated cancer researcher. He reflects on his formative experiences during his PhD at Ludwig Maximilian University in Munich, where she developed murine models for refractory acute myeloid leukemia (AML). We examine these models' contributions to therapeutic discovery and understanding the intricate mechanisms underscoring AML's complexities. Transitioning to his postdoctoral work at Scott Armstrong's lab in Boston, Dr. Despande shares his insights on the importance of epigenetic regulators, such as DOT1L, in leukemias, and how they can serve as strategic therapeutic targets. His ambitious pursuit of translational research is further highlighted through his efforts in developing a conditional knockout mouse model and his collaborative work utilizing CRISPR technology to refine our understanding of epigenetic regulation in cancer pathogenesis. Moreover, we engage in a conversation about the challenges and opportunities that arise when establishing his lab at Sanford Burnham Prebys. Dr. Despande candidly discusses the delicate balance between pursuing topics of genuine interest versus adhering to grant fundability, underlining the tension researchers face in the current scientific landscape. His emphasis on the critical need for innovation within lab settings serves as a motivational call for emerging scientists to venture beyond the established templates that often inhibit groundbreaking discoveries. We conclude our dialogue with an exploration of his recent projects, which involve targeting specific epigenetic modifiers and how his lab's findings can contribute to greater understanding and potential treatments for not only AML but also other pediatric cancers driven by gene fusions. Dr. Despande's insights into the integration of modern technologies, such as CRISPR libraries, exemplify his commitment to pushing the boundaries of cancer research. In addition to discussing his scientific contributions, we touch upon Dr. Despande's foray into podcasting (The Discovery Dialogues), shedding light on his motivation to bridge the communication gap between scientists and the broader public. He articulates his desire to demystify scientific discoveries and promote awareness about the intricate journey of research that lays the groundwork for medical advancements. This multidimensional discussion not only highlights his scientific achievements but also emphasizes the importance of effective science communication in fostering public understanding and appreciation of research.   References Deshpande AJ, Cusan M, Rawat VP, Reuter H, Krause A, Pott C, Quintanilla-Martinez L, Kakadia P, Kuchenbauer F, Ahmed F, Delabesse E, Hahn M, Lichter P, Kneba M, Hiddemann W, Macintyre E, Mecucci C, Ludwig WD, Humphries RK, Bohlander SK, Feuring-Buske M, Buske C. Acute myeloid leukemia is propagated by a leukemic stem cell with lymphoid characteristics in a mouse model of CALM/AF10-positive leukemia. Cancer Cell. 2006 Nov;10(5):363-74. doi: 10.1016/j.ccr.2006.08.023. PMID: 17097559. Deshpande AJ, Deshpande A, Sinha AU, Chen L, Chang J, Cihan A, Fazio M, Chen CW, Zhu N, Koche R, Dzhekieva L, Ibáñez G, Dias S, Banka D, Krivtsov A, Luo M, Roeder RG, Bradner JE, Bernt KM, Armstrong SA. AF10 regulates progressive H3K79 methylation and HOX gene expression in diverse AML subtypes. Cancer Cell. 2014 Dec 8;26(6):896-908. doi: 10.1016/j.ccell.2014.10.009. Epub 2014 Nov 20. PMID: 25464900; PMCID: PMC4291116. Sinha S, Barbosa K, Cheng K, Leiserson MDM, Jain P, Deshpande A, Wilson DM 3rd, Ryan BM, Luo J, Ronai ZA, Lee JS, Deshpande AJ, Ruppin E. A systematic genome-wide mapping of oncogenic mutation selection during CRISPR-Cas9 genome editing. Nat Commun. 2021 Nov 11;12(1):6512. doi: 10.1038/s41467-021-26788-6. Erratum in: Nat Commun. 2022 May 16;13(1):2828. doi: 10.1038/s41467-022-30475-5. PMID: 34764240; PMCID: PMC8586238.   Related Episodes Targeting COMPASS to Cure Childhood Leukemia (Ali Shilatifard) The Menin-MLL Complex and Small Molecule Inhibitors (Yadira Soto-Feliciano) MLL Proteins in Mixed-Lineage Leukemia (Yali Dou)   Contact Epigenetics Podcast on Mastodon Epigenetics Podcast on Bluesky Dr. Stefan Dillinger on LinkedIn Active Motif on LinkedIn Active Motif on Bluesky Email: podcast@activemotif.com

Mon, 7 Jul 2014 12:00:00 +0100 https://edoc.ub.uni-muenchen.de/17426/ https://edoc.ub.uni-muenchen.de/17426/1/Dutta_Sayantanee.pdf Dutta, Sayantanee ddc:610, ddc:600, Medizinische Fakultät 0

Chromosomal translocations are common in human leukemias. Detailed studies of chromosomal translocation have been useful in understanding the pathogenesis and identifying therapeutic targets in hematologic malignancies. Some translocations result in the formation of fusion genes. These fusion proteins play an important role in leukemogenesis. The t(10;11)(p12;q14) translocation is rare but recurring and results in the formation of the CALM/AF10 fusion protein. Patients with this translocation have a bad prognosis. To understand how CALM/AF10 leads to leukemia, various mouse models have been established. In a murine bone marrow retroviral transduction and transplantation model Deshpande et al. (2006) showed that mice expressing CALM/AF10 in their bone marrow cells developed an acute myeloid leukemia with a penetrance of 100% and a short latency period of 110 days. Using a transgenic mouse model, in which CALM/AF10 was under the control of Vav promoter, Peter Aplan and colleagues demonstrated that only 40% to 50% of mice developed leukemia after a long latency of 10 to 12 months. Two classical transgenic CALM/AF10 models were established in our group using the immunoglobulin heavy chain enhancer/promoter (IgH-CALM/AF10) and proximal murine LcK promoter (pLck-CALM/AF10) to drive CALM/AF10 expression. These transgenic mice did not show any leukemic phenotype even after an observation period of 15 months. Taken together these studies strongly suggest that additional collaborating factors are required for the CALM/AF10 fusion gene to induce leukemia. Meis1, a Hox cofactor, is known to collaborate with several Hox genes and Hox fusion genes such as HOXA9 and NUP98-HOXD13. In these studies, Meis1 played a critical role in accelerating the development of leukemia. It could also be shown that MEIS1 is highly expressed in CALM/AF10 positive human leukemia cells. Therefore, I sought to determine whether the homeobox gene Meis1 collaborates with CALM/AF10 in inducing leukemia. In order to achieve this goal, lethally irradiated non-transgenic mice were transplanted with IgH-CALM/AF10 transgenic bone marrow cells transduced with a Meis1 expressing retrovirus. The transplanted mice developed an acute leukemia with a penetrance of 100% and a median latency period of 187 days. The leukemia showed predominantly myeloid features such as the presence of myeloid marker positive cells. The myeloid blast cells infiltrated in multiple hematopoietic as well as non-hematopoietic organs. The leukemic cells were also positive for the B-cell marker B220. Cells that were positive for both lymphoid and myeloid markers, a characteristic feature of CALM/AF10-induced leukemia, were also detected in all the mice. The leukemic cells had clonal DJH rearrangements. Overall, these data suggest that the transformed cell might be an early progenitor cell capable of lymphoid as well as myeloid differentiation or that the leukemia was initiated by a B220+ IgH DJ rearranged cell with blocked lymphoid differentiation, which started a default myeloid differentiation program. By performing serial secondary and tertiary transplantations the leukemic nature of the disease could be confirmed. Colony forming cell assays showed that CALM/AF10 in collaboration with Meis1 failed to induce the transformation of hematopoietic progenitors in vitro. This could either be due to the lack of required growth factors and conditions necessary for the proliferation of the transformable cell or lack of additional events essential for progression towards leukemia development. In conclusion, I have demonstrated that Meis1 collaborates with CALM/AF10 in inducing acute myeloid leukemia. Additional, detailed analyses of the leukemia initiating cell in these models would help to better understand the pathogenesis of CALM/AF10-induced leukemia.

Mon, 27 Jul 2009 12:00:00 +0100 https://edoc.ub.uni-muenchen.de/10480/ https://edoc.ub.uni-muenchen.de/10480/1/Mulaw_Medhanie_Assmelash.pdf Mulaw, Medhanie Assmelash ddc:610, ddc:600, Medizinische Fakul

The Clathrin Assembly Lymphoid Myeloid leukemia gene (CALM) was first identified as the fusion partner of AF10 in the t(10;11)(p13;q14) translocation. The CALM/AF10 fusion protein plays a crucial role in t(10;11)(p13;q14) associated leukemogenesis. Using the N-terminal half of CALM as a bait in a yeast two-hybrid screen a novel protein named CATS (CALM interacting protein expressed in thymus and spleen) was identified as CALM interacting partner. Multiple tissue Northern blot analysis showed predominant expression of CATS in lymphoid tissues. CATS codes for two protein isoforms of 238 and 248 amino acids. The interaction between CALM and CATS was confirmed by co-immunoprecipitation and colocalization experiments. The CATS interaction domain of CALM was mapped to amino acids 221 to 294 of CALM. This domain is contained in the CALM/AF10 fusion protein. CATS localizes to the nucleus and shows a preference for nucleoli. Expression of CATS was able to markedly increase the nuclear localization of CALM and of the leukemogenic fusion protein CALM/AF10. This effect of CATS seems to be stronger on CALM/AF10 than on CALM. Several monoclonal antibodies against the C-terminus of human CATS were generated. These antibodies recognize both the human and the murine CATS protein. Western blot analyses showed that CATS is strongly expressed in different human leukemia, lymphoma and tumor cell lines but not in resting T-cells. High CATS expression in proliferating cells as well as its nucleolar localization suggest a role of CATS in the control of cell proliferation. In order to gain further insight into CATS function we used CATS as a bait in a yeast two-hybrid screen. Several CATS interacting proteins with apparently unrelated function were identified. Interestingly, on closer scrutiny these proteins could be associated with three key regulatory pathways: signaling, apoptosis and cell cycle control. We discuss in detail the biological relevance of the CATS interaction with the two apoptosis-associated proteins HAX1 and SIVA, the cell cycle regulator KIS and the CALM interacting ribonucleoprotein PCBP1. Our results indicate that the subcellular localization of CALM and CALM/AF10 could depend in part on the presence of CATS with a greater fraction of CALM or CALM/AF10 being present in the nucleus of cells with high CATS expression (e.g. lymphoid cells have high CATS expression). Moreover we provide evidences that CATS function might be tightly linked to cancer initiation and/or progression. The CALM-CATS interaction might thus play an important role in CALM/AF10 mediated leukemogenesis.

The t(10;11)(p13;q14) is a recurring translocation resulting in the fusion of the CALM and AF10 genes. The leukemogenic CALM/AF10 fusion genes codes for a 1595 amino acids protein. This translocation was first identified in a patient with hystiocytic lymphoma and was subsequently found in patients with AML, T-ALL and malignant lymphoma. This translocation is found in younger patients and is associated with a poor prognosis. The CALM/AF10-associated leukemias can exhibit myeloid, lymphoid or mixed lymphoid-meyloid features, indicating a stem cell or an early commited progenitor as the target cell of leukemic transformation. At the present time the target cells in CALM/AF10-associated leukemogenesis are unknown. It is also not known which target genes are up or downregulated by the presence of the CALM/AF10 fusion protein. To answer these questions, the following experiments were performed: 1) Five transgenic mouse lines, two expressing CALM/AF10 under the control of the immunoglobulin heavy chain enhancer promoter and three under the control of the murine proximal Lck promoter were generated. Although the CALM/AF10 expression was confirmed to be present and specific to the cells targeted by the promoters used (B- and T-cell progenitors for IgH and Lck promoters, respectively), the transgenic animals did not show a phenotype that could be detected after meticulous clinical, haematological, immunological, flow cytometrical and immunohistopatological analysis . 2) We performed molecular characterization of several CALM/AF10 patient samples: A group of 13 patients with different types of leukemia: case 1 (AML M2), case 2 (Acute Biphetnotypic leukemia), case 3 (Pre T-ALL), case 4 (Acute Undifferentiated Leukemia), case 5 (PreT-ALL), cases 6 and 7 (ProT-ALL), case 8 (T-ALL), case 9 (AML), case 14 (T-ALL), case 15, 16 and 17 (AML) with a t(10;11) translocation detected by cytogenetic analysis suggesting a CALM/AF10-rearrangement. The samples were analyzed for the presence of the CALM/AF10 and AF10/CALM fusion transcripts by RT-PCR and sequence analysis. All these patients were found to be positive for the CALM/AF10 fusion. In addition, we analyzed a series of twenty-nine patients with T-ALL with T-cell receptor ≥¥ rearrangement. Among these patients, four (case 10 to 13) were positive for the CALM/AF10 fusion transcript, indicating a high incidence of CALM/AF10 fusions in this group of leukemia. Three different breakpoints in CALM at nucleotide 1926, 2091 and a new exon, with 106 bases inserted after nt 2064 of CALM in patient 4 were found. In AF10 four breakpoints were identified: at nucleotide position 424, 589, 883 and 979. In patient 16 we found an extra exon before nt 424 of AF10. In seven patients it was also possible to amplify the reciprocal AF10/CALM fusion transcript (case 1, 3, 4, 8, 9, 10 and 14). There was no correlation between disease phenotype and breakpoint location. Ten CALM/AF10 positive patients were analyzed using oligonucleotide microarrays representing 33,000 different genes (U133 set, Affymetrix). Analysis of microarray gene expression signatures of these patients revealed high expression levels of the polycomb group gene BMI1, the homeobox gene MEIS1 and the HOXA cluster genes HOXA1, HOXA4, HOXA5, HOXA7, HOXA9, and HOXA10. The overexpression of HOX genes seen in these CALM/AF10 positive leukemias is reminiscent to the pattern seen in leukemias with rearrangements of the MLL gene, normal karyotypes and complex aberrant karyotypes suggesting a common effector pathway (i.e. HOX gene deregulation) for these diverse leukemias. In addition, the general pattern of gene expression of CALM/AF10 patients when compared to other leukemia subtypes and to normal bone marrow was dominated by a global downregulation of genes some of them with function identified as related to important molecular mechanisms, such as membrane trafficking, cell growth regulation, proliferation, differentiation and tumor suppression. 3) We cloned CALM/AF10 fusion gene into a vector that allowed us to induce the expression of CALM/AF10 using doxycycline in transiently and stably-transfected NIH3T3 and HEK293 cells. This system will be an important tool to identify direct CALM/AF10 target genes and to answer the question whether continued CALM/AF10 expression is necessary to maintain the CALM/AF10-associated expression pattern.

We have demonstrated, that an acute leukemia with a predominantly myeloid phenotype can be propagated by a progenitor with lymphoid characteristics in a mouse model of the t(10;11) (p13;q14) translocation. Mice transplanted with bone marrow retrovirally engineered to express the leukemia specific CALM/AF10 fusion gene consistently developed an acute leukemia with a short latency. The leukemia showed characteristic myeloid features such as the presence of myeloid marker positive cells infiltrating multiple hematopoietic and non-hematopoietic organs, the positivity of blasts for myeloid specific histochemical stainings and the depletion of the lymphoid compartment in lymphoid organs. Apart from the major population of cells expressing myeloid but not lymphoid markers (M population), a smaller population of cells expressing myeloid markers as well as the lymphoid marker B220 ( B/M population) and a smaller population expressing only the B220 marker (B population) could be detected in all mice. We determined that the frequency of leukemia propagating cells was the highest in the B population and that this population could give rise to the other two populations of cells, namely the B/M and the M populations. This indicated that the leukemic stem cell candidate for the myeloid leukemia in this model of CALM/AF10 induced transformation is a B220 + cell. Further characterization of these candidate LSCs revealed the presence of D-JH rearrangements and the absence of Pax5 transcription. These cells were characterised as being CD43 + /AA4.1 + /HSA low-pos/CD19 -/FLT3R + /IL-7R low-neg c-kit low-neg and expressing the early B cell factor (EBF) transcripts as well as transcripts for the myeloperoxidase (MPO) gene,bearing a resemblance to Pax5 knockout preBI cells. These findings indicate that the leukemia-propagating cell in a subset of acute myeloid leukemias could be a cell with lymphoid characteristics. The fact that this progenitor cell expressed markers different from those expressed by the bulk leukemic population but could still propagate the leukemia raises the interesting possibility of selectively targeting these cells using novel therapeutic strategies that aim to eliminate these LSCs.