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Leukemogenesis. Ahmad Silmi Advanced Hematology Medi6304 Second Semester / Mar,3th,2017. Malignant Transformation.
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Leukemogenesis Ahmad Silmi Advanced Hematology Medi6304 Second Semester / Mar,3th,2017
Malignant Transformation • Is a group of malignant (neoplastic) disorders that are characterized by the clonal expansion and accumulation of one or more blood cell line(s) with eventual involvement of all hematopoietic organs and other organs.
Malignant Transformation • A clone of cells with characteristics of a malignant cell: • • Immortality (resistance to apoptosis) • • Growth factor independent growth • • Insensitivity to growth-inhibitory signals (increased proliferation rate) • • Ability to invade and metastasize • • Ability to stimulate angiogenesis • • Blockage of intracellular differentiation
Leukemia: Double hit theory • Leukemia is a result of two or more transforming somatic (due to genetic and/or epigenetic aberration) mutations within the genome of HSC or multipotential progenitors or precursors. • Leukemia: Double hit theory • Malignant transformation due to: • • Activation of specific proto oncogenes • • De-activation of tumor suppressor genes
Leukemia Double hit theory
Malignancy: Multistage development Three stages of malignancy development: 1)Initiation 2)Promotion and transformation 3)Progression
Leukemia: Initiation A result of genetic and/or epigenetic irreversible damage of DNA that ultimately resulted in either structural or functional alteration of DNA. •Mediated by exposure to carcinogen which include radiation, chemicals and Viral infections. •Initiating factors induce damage to one or more of cell cycle checkpoints that are required to ensure controlled replication of normal cell.
Leukemia: Promotion and transformation Tumor promotion comprises the selective clonal expansion of initiated cells to produces a larger population of cells that are at higher risk of further genetic changes and malignant conversion •Promoters reduce the latency period for tumor formation upon exposure to a tumor initiator and contribute to the process of carcinogenesis by the expansion of a population of initiated cells that will then be at risk for malignant conversion •Malignant conversion is the transformation of a pre-neoplastic cell into one that expresses the malignant phenotype. •Transformation requires further genetic changes that may result from infidelity of DNA synthesis.
Leukemia: Progression Evolution of progressively more malignant cells as a result of propensity for genomic instability and uncontrolled growth as well as development of metastatic colonies mediated by the secretion of proteases that allow invasion beyond the primary tumor location. •Influenced by: 1.Hormone responsiveness 2.Growth rate 3.Invasiveness 4.Additional mutations 5.Drug resistance 6.Host defense response
Self renewal capacity: Prolonged survival DNA polymerase is unable to replicate the ends (telomeres) of chromosomes. •Results in an “internal clock” because the number of telomeres is decreased with each cell cycle. •Normal stem cells and some malignant cells are capable of expressing telomerase, an enzyme that replenishes telomeres. •Normally, telomerase activity progressively decreases with differentiation.
Self renewal capacity: Prolonged survival • Telomerase activity can be detected in cells that are in the cell cycle but not in cells that are in G0. •Telomerase activity progressively decreases with differentiation •Lymphoid cells capable of cell division and clonal expansion have telomerase activity that increases when they proliferate.
Self renewal capacity: Prolonged survival • Telomere and telomerase status in normal and malignant hematopoietic cells: • • LT-HSCs have relatively long telomeres, but low telomerase activity. • • ST-HSCs have long telomeres and up-regulate telomerase, enabling them to actively amplify. • • LSCs have high telomerase activity but short telomeres that essentially require maintenance by telomerase
Telomerase activity in leukemia At least 80% of patients with acute leukemia tend to have high telomerase activity. expression of hTERT (the catalytic subunit of telomerase) in cultured human primary cells reconstitutes telomerase activity and allows immortal growth. TERT-mediated telomerase activation is able to cooperate with oncogenes in transforming cultured primary cells into neoplastic cells. Patients with CML have low levels of telomerase activity that increases during the blastic phase. Patients with MDS have low levels that slowly increase during the course of the disease. Telomerase inhibition enhances apoptosis in human acute leukemia cells; it has been shown that telomere shortening by telomestatin induces apoptosis in some primary blast cells of acute leukemia.
Resistance to Apoptosis Several mechanisms have been proposed that include: •Inactivation of the pro-apoptotic protein BAD • Activation of the NF-κB survival pathway •Induction of caspase inhibitors •Inhibition of ASK-1
Resistance to Apoptosis: P53 p53 is inhibited by MDM2, a ubiquitin ligase that targets p53 for destruction by the proteasome. MDM2 is inactivated by binding to ARF. Cellular stress, including that induced by chemotherapy or irradiation, activates p53 either directly, by inhibition of MDM2, or indirectly by activation of ARF. ARF can also be induced by proliferative oncogenes such as RAS. Active p53 trans-activates pro-apoptotic genes including BAX, NOXA, CD95 and TRAIL-R1 to promote apoptosis. TRAIL-R1, tumor-necrosis-factor-related apoptosis-inducing ligand receptor 1.
Growth factors independent growth Oncogenic mutations of growth factor receptor proteins. For example, Receptor tyrosine kinases (PDGFRβ, c-kit, Flt3, FGFR and others) Mutation of cytoplasmic “2nd messengers” that transduce signals to the nucleus. • SOS (son of sevenless): an exchange factor that converts GDP to GTP • •Ras – encodes small GP proteins that serve as molecular switches and activate raf. • •Raf – inhibits apoptosis and phosphorylates MEK • •MEK – MApk/ERK/ kinase • •MApk – mitogen activated protein kinase • •ERK – extracellular signal regulated kinase • •Fos – enters nucleus – a transcription factor • Mutation of transcription factors that affect genetic responses to growth factors (c-myb or c-myc)
Growth factors independent growth Tumor cells generate many of their own growth factors (autocrine stimulation) Cancer cells may also switch the types of extracellular matrix receptors (integrins), favoring ones that transmit pro-growth signals. Some tumor cells acquire the ability to stimulate their neighboring cells to release increased amounts of growth stimulating signals.
Insensitivity to growth inhibition • Antigrowth signals can block proliferation by two mechanisms. • •Forcing the cell out of the cell cycle and into G0. • •Inducing the cell to differentiate into a postmitotic state. • Mutation of tumor suppressor genes such as Rb, p53, ink4– many of which normally act by suppressing progression through the cell cycle. For example: • •Rb:E2F complexes regulate the G1 into S transition. • •Ink4 proteins inhibit CDK4 and CDK6. • • P53 is considered the guardian of the genome and halts the cell cycle upon DNA damage.
Proto-oncogens Proto-oncogenes are normal genes that usually encode proteins that help to regulate cell growth and differentiation. Proto-oncogenes are often involved in signal transduction and execution of mitogenic signals A proto-oncogene is a normal gene that can become an oncogene (a result of mutations or increased expression) to produce oncoprotein(s).
Proto-oncogens: Levels of activation • Levels of activations: • (1)Over expression following acquisitiin of a novel transcriptional promoter • (2)Over expression due to amplification (i.e. increased number of gene copies) • (3)Influence on the level of transcription (i.e. enhanced gene products) • (4)Juxtaposition of the encogene following to a chromosomal translocation with an immunoglobulin domain • (1)Alteration in the structure of the oncoprotein (enhanced function and or activity). • Translocation can disturb the regulation of an oncogene through: • A.Providing a new promoter region or some other control elements that would activate the oncogene • B.Altering the coding sequence of the gene and accordingly changing its protein product from benign to a malignant form
Proto-oncogens: Mechanisms of activation • Mechanisms of activation: • (1)point mutations are found in highly specific regions of the gene • (2)Amplification of chromosomal segments containing these genes. • (3)Chromosomal translocation
is the result of a reciprocal translocation between chromosome 9 and 22. The result is that a fusion gene is created by juxtapositioning the Abl1 gene (encodes tyrosine kinase) on chromosome 9 (region q34) to a part of the BCR ("breakpoint cluster region") gene on chromosome 22 Philadelphia chromosome t(9;22)(q34;q11): Since ABL activates a number of cell cycle regulatory proteins, the result of the BCR-Abl fusion enhanced cell divisions and proliferation causing further genomic instability as well as has the ability to inhibits DNA repair mechanisms
Philadelphia chromosome t(9;22)(q34;q11): • Three clinically important variants: • • p190 is generally associated with ALL • • p210 is generally associated with CML and occasionally in cases of ALL • • p230 is usually associated with CNL.
Tumor suppression genes • Tumor-suppressor genes have down regulate cell cycle and/or promote apoptosis. • Functions: • 1.Repression of genes that are essential for cell cycle. • 2.Providing a link of the cell cycle to DNA damage and mediating DNA repair or promoting apoptosis if repair can’t be achieved. • 3.metastasis suppressors: involved in cell adhesion and block loss of contact inhibition • 4.DNA repair: for example: HNPCC, MEN1 and BRCA.
Tumor suppression genes Loss of function of tumor-suppressor genes usually occurs in a bimodal fashion, and most frequently involves point mutations in one allele and loss of the second allele by a deletion, recombinational event, or chromosomal nondisjunction.
Tumor suppressor genes: P53 • p53 is encoded by the TP53 gene located on 17p13.1 • Function: P53 is involved in many cellular activities predominantly including: • 1. DNA repair • 2. cell cycle arrest at the G21/S check point upon DNA damage recognition • 3. Initiation of apoptosis.
EPIGENETIC CHANGES IN LEUKEMOGENESIS • Epigenetics is generally used to refer to mitotically and meiotically heritable changes in gene expression that occur without alteration of the DNA coding sequence. Epigenetic changes that underlie leukemogenesis have been described as falling into one of two major categories: • Changes in the DNA methylation (Box 1) state • Alterations in the histone modification (Box 2) pattern.
EPIGENETIC CHANGES IN LEUKEMOGENESIS Compared with normal cells, cancer cells exhibit global DNA hypomethylation accompanied by aberrant methylation of CpG islands within gene promoters or coding regions. In the context of leukemogenesis, aberrant CpG island methylation in promoter regions of tumor suppressor genes (ex. cyclin-dependent kinase inhibitor 2B (CDKN2B) is associated with transcriptional silencing, which also involves recruitment of methyl-binding proteins and HDACs to regions around the transcriptional initiation sites
EPIGENETIC CHANGES IN LEUKEMOGENESIS • Oncogeneic fusion proteins such as AML1-ETO, CBFB-MYH11, and PML-RARA recruit transcriptional co-repressor complexes (including NCOR1 and SMRT) which result in the loss of histone acetylation and the acquisition of repressive histone modification marks such as histone H3 lysine 9 (H3K9) methylation and H3K27 trimethylation, as well as DNA methylation, and thereby a closed chromatin structure.
EPIGENETIC CHANGES IN LEUKEMOGENESIS • MICRORNAS (miRNA) • miRNAs are critical regulators of many physiological processes such as development, cell apoptosis, differentiation, and proliferation. • miRNAs function in complex regulatory networks to regulate hematopoietic differentiation and contribute to leukemogenesis
