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![ The extrinsic apoptotic pathway plays a major role in apoptosis.
Six known death receptors (DRs) including tumor necrosis factor(TNF),
Fas(APO-1 or CD95), and DR4/DR5 (receptors for TNF-related apoptosis
induce ligand [TRAIL]).
Contain a cytosolic domain called the death domain, which recruits
adaptor proteins such as Fadd/Mort-1 to the receptor complex after
binding to ligand.
The recruiter adaptor protein has a death domain end and a death
effector domain (DED).](https://image.slidesharecdn.com/cllpathogenesisandtargetedtherapy-180829185640/85/Cll-pathogenesis-and-targeted-therapy-21-320.jpg)
![ NOTCH1 mutations
Restricted to the C-terminal PEST [proline (P), glutamate (E), serine (S),
and threonine (T)] domain
normally limits the intensity and duration of NOTCH1 signaling.
Removal of the PEST domain impairs the degradation of NOTCH1
allowing accumulation of the active form of NOTCH1.
One recurrent mutation (c.7544_7545delCT) accounts for ∼77% of all
NOTCH1 mutations in CLL-
Can be rapidly detected by a simple polymerase chain reaction–based
strategy- providing a potential approach for a first-level
screening of NOTCH1 alterations.](https://image.slidesharecdn.com/cllpathogenesisandtargetedtherapy-180829185640/85/Cll-pathogenesis-and-targeted-therapy-46-320.jpg)
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Similar to Cll pathogenesis and targeted therapy
Cll pathogenesis and targeted therapy
- 1. PATHOGENESIS OF CLLAND ITS BASIS FOR TARGETED THERAPY Presenter- Dr Abha Singh Moderator- Dr Mrinalini Kotru
- 2. EPIDEMIOLOGY CLL hasaverage incidence of 5.17 persons per 100,000 in United States representing 20% of all mature B cell neoplasms. Accounts for 0.8 % of all cancers Incidence is twice in males as compared to females Generally, the neoplastic lymphocytes are of B cell origin In less than 2 % cases, however, the neoplastic cells are of T cell origin and are included in the category of T cell prolymphocytic leukemia.
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- 5. ETIOLOGY Mostly idiopathic Familial cases Cytogenetics- clonal chromosomal abnormalities are detected in approximately 50% of CLL patients Most common clonal abnormality is trisomy 12 Structural abnormalities of chromosome 13, 17 and 11 Patients with abnormal karyotypes have worse prognosis
- 6. CLINICAL FEATURES Patientsare more prone to viral or bacterial infections secondary to impaired T-cell immunity or hypogammaglobinemia respectively. Night sweats and fever (B symptoms) are uncommon and requires prompt evaluation for complicating infectious disease. 80% of CLL patients have non tender LAP involving the cervical, supraclavicular or axillary lymph nodes. Lymphedema is rare.
- 7. LAB EVALUATION Sustainedmonoclonal lymphocytosis greater than 5000/micro litre. In B cell, clonality is confirmed by expression of kappa or lambda light chain on surface membrane. 10-25% patients with CLL develop autoimmune haemolytic anemia, with a positive direct Coombs test. The marrow aspirates shows greater than 30% of the nucleated cells as being lymphoid.
- 8. Continue lab evaluation… Essential Physical exam : attention to node bearing areas including waldayers ring, and to the size of spleen B symptoms CBC, differential count, platelets Comprehensive metabolic panel
- 9. OTHER INVESTIGATIONS Quantitativeimmunoglobulins Reticulocyte count, haptoglobin and DCT Beta 2 microglobulin level LDH IGVH mutation status Zap 70 molecular investigation Radiological investigation before chemo-therapy (PET scan usually not required)
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- 11. Points to bediscussed Origin of CLL Possible drivers Pathogenesis Treatment Targeted therapy Approach to treatment
- 13. ORIGIN OF CLL B cells express CD5,CD19, CD23 and low levels of surface Ig Recent gene expression profiling studies have confirmed that CLL is probably derived from CD5 B cells similar to those found in healthy individuals.
- 14. Cellular origin ofCLL SMZL DLBCL HCL,PLL CLL( mutated) MALT CLL(Unmutated) FL, DLBCL, BURKITT’S PLASMA CELL DYSCRASIA MANTLE CELL HL Naïve B cell Marginal zone GC MZ
- 15. Cellular origin ofCLL : MUTATED VS UNMUTATED CLL( mutated) CLL(Unmutated) Naïve B cell Marginal zone GC MZ ANTIGEN MEMORY B CELL
- 16. CD5+ B cells Gene expression studies on CLL cells revealed that leukemia cells that express mutated or unmutated IGHVs share common gene expression profile-suggesting common origin On basis of several studies: CLL cells with unmutated IGHVs are derived from mature CD5+ CD27- B cells CLL cells with mutated IGHVs are derived from distinct, previously unrecognised, subset of CD5+ CD27+ Post germinal centre B cells with mutated IGHVs
- 17. Development of chroniclymphocytic leukemia (CLL).
- 18. DRIVER EVENTS INCLL PATHOGENESIS CLL Mutations TP53, SF3B1, NOTCH1, ATM, MYD88 Loss of 13q, 11q,17p trisomy 12 Auto- activation BCR Signaling Apoptosis microenvironment Unknown antigen
- 19. Mutations- atany stage of B cell development HSCs bearing oncogenetic mutations may give rise to B cells with modest growth/survival advantages M-CLL- B cells with incurred immunoglobulin (Ig) somatic mutation express mutated Ig heavy-chain variable-region genes (IGHVs) U-CLL- B cells that have not undergone Ig somatic mutations express germ-line IGHVs. The expansion of a CLL (or MBL) clone is associated with de novo accumulation of additional genetic lesions, interactions between the leukemic cells, accessory cells and antigens in the leukemia microenvironment of lymphoid tissues .
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- 21. The extrinsicapoptotic pathway plays a major role in apoptosis. Six known death receptors (DRs) including tumor necrosis factor(TNF), Fas(APO-1 or CD95), and DR4/DR5 (receptors for TNF-related apoptosis induce ligand [TRAIL]). Contain a cytosolic domain called the death domain, which recruits adaptor proteins such as Fadd/Mort-1 to the receptor complex after binding to ligand. The recruiter adaptor protein has a death domain end and a death effector domain (DED).
- 22. Once boundto the TNF receptor, the DED binds to caspases 8 and 10, which then become activated by autoactivation . One of these inhibitors is FLICE inhibitory protein (FLIP), which is a homolog of procaspase 8 and contains two DED domains but lacks proteolytic activity. It been reported that CLL cells have high levels of FLIP expression, rendering the cells resistant to DR-induced apoptosis.
- 23. Modulators of theApoptotic Pathway in CLL The bcl-2 family consists of ∼20 members that can either promote or inhibit apoptosis. Located in the cell membrane, nuclear membrane, and mitochondrial membrane Function by binding to other proteins or influencing cell permeability and the release of cytochrome c from the mitochondria. bax, bcl-xS, bak, and bad-promote apoptosis bcl-2, bcl-xL, and mcl-1 - inhibit apoptosis Another group (e.g., bag-1) can influence the activities of the other family members. CLL cells have high bcl-2, bax, and bak levels but have low levels of bcl-xL and bad
- 24. p53 -induce apoptosis, and this occurs preferentially in tumor cells( a feature that may explain the relative tumor specificity of anticancer agents) Mechanisms p53-induced apoptosis – up-regulation in the expressions of the TRAIL DRs, DR4 and DR5, increased expression of the proapoptotic Bcl-2 family members, Bax, Noxa, and Puma p53 mutations are typically associated with deletions of the second allele (deletion 17p13) Mutations or p53 gene deletions are observed in 10 to 15% of CLL patients These abnormalities are associated with high lymphocyte counts, drug resistance to anticancer agents in vitro and in vivo, and poor patient survival
- 25. The ATMgene is located on chromosome 11q22–q23 Responsible for phosphorylation and activation of p53 after DNA damage Approximately 30% of CLL patients have a mutation of ATM These patients have a defect in cellular response to irradiation similar to that observed in patients with a p53 mutation Explains the drug resistance and poor clinical outcome in patients with an ATM mutation ATM mutations are seen in patients with an unmutated IgV gene, and these patients are known to have a poor prognosis
- 26. Abnormalities in CellDivision Both cyclins D2 and D3 are overexpressed in chronic lymphocytic leukemia cells, but the retinoblastoma (Rb) protein is not phosphorylated, perhaps related to overexpression of p27Kip1. CDK, cyclin dependent kinase.
- 27. BCR SIGNALING BCR SIGNALING CHRONICACTIVE BCR SIGNALING TONIC BCR SIGNALING MAPK mTOR NFAT NFKB PI3K
- 29. Ligation ofB cell receptor (BCR) by antigen recruits kinases such as spleen tyrosine kinase (SYK) and the SRC kinase LYN that phosphorylate immunoreceptor tyrosine–based activation motifs (ITAMs) on the cytoplasmic domains of the immunoglobulin (Ig) coreceptors CD79a and CD79b. Such phosphorylation recruits and activates Bruton’s tyrosine kinase (BTK) and phosphatidylinositol 3-kinase (PI3K), subsequently activating many downstream targets, including AKT/mTOR (mammalian target of rapamycin), nuclear factor κB (NF-κB), and extracellular signal–regulated kinase (ERK). B cell signaling in chronic lymphocytic leukemia
- 30. B cell signalingin chronic lymphocytic leukemia This signaling can be enhanced by ζ-associated protein of 70 kD (ZAP- 70). CD38, CD49d, CD44, and matrix metalloproteinase (MMP)-9 may form a supramolecular complex with ZAP-70. This complex can also recruit ZAP-70 to the plasma membrane, where it can enhance BCR signaling.
- 31. BCR SIGNALING CLL vsNORMAL B CELLS CHARACTERISTIC CLL CELLS sIg expression Low (higher in U-CLL) CD79 expression Low Response to antigen stimulation Variable (higher in U-CLL) Calcium flux Variable/generally low Syn/Lyn/Btk expression elevated PI3K p110 delta expression normal PI3K Kinase activity elevated
- 32. Cell Survival Factorsin CLL CLL cells interact with the microenvironment through activation of their cell-surface receptors- main signal that promotes survival in CLL cells. CLL cells also produce cytokines-stimulate their own growth in an autocrine or paracrine fashion while inhibiting survival of normal lymphoid and marrow cells. This latter effect can lead to the immunosuppression and myelosuppression that typifies this disease.
- 33. The variouscytokines and their receptors that contribute to cell survival are B-Cell Receptor (BCR) Interleukins Tumor Necrosis Factor-α (TNF-α) Transforming Growth Factor-β (TGF-β) Adhesion Molecule Receptors BMCA, TAC1, and BAFF-R Vascular Endothelial Growth Factor (VEGF) Receptor CD40
- 34. Stromal-Derived GrowthFactor-1 (SDF-1) Receptor Basic Fibroblast Growth-Factor (bFGF) Receptor Albumin Lysophosphatidic Acid (LPA)
- 35. CLL microenvironment Chemokinereceptors and adhesion molecules expressed by CLL cells are critical for homing and retention of CLL cells within the tissue compartments. CLL cells receive prosurvival signals via contact with accessory stromal cells in the leukemia microenvironment. Nurselike cells express the chemokines CXCL12 and CXCL13, whereas marrow stromal cells express predominantly CXCL12. Nurselike cells and marrow stromal cells attract CLL cells via the G protein–coupled chemokine receptors CXCR4 and CXCR5, which are expressed at high levels on CLL cells.
- 36. CLL microenvironment Nurselikecells also express the TNF family member BAFF and a proliferation-inducing ligand, providing survival signals to CLL cells via the corresponding receptors (BCMA, TACI, BAFF receptor). Integrins, particularly very late antigen 4 integrins (CD49d), cooperate with chemokine receptors in establishing cell– cell adhesion through respective ligands on the stromal cells (vascular cell adhesion molecule 1 and fibronectin). Another important molecule involved in the interactions between leukemia cells and their microenvironment is CD44
- 37. CD38, CD49d,CD44, and matrix metalloproteinase (MMP)-9 may form a supramolecular complex with ZAP-70. This complex can also recruit ZAP-70 to the plasma membrane, where it can enhance BCR signaling Following binding to any or all of these receptors by ligands released by accessory cells in the leukemia microenvironment, AKT and ERK undergo enhanced activation. CLL microenvironment
- 38. CLL microenvironment CXCR4can also directly interact with CXCL12 to induce calcium mobilization, activation of PI3K/AKT, ERK, and serine phosphorylation of signal transducer and activator of transcription 3 (STAT3). Activation of Toll-like receptor (TLR) can also enhance or induce activation of NF-κB.
- 39. IMMUNOGLOBULIN REPERTOIRE Bcells are crucial to adaptive immune responses. The probability that two independent B cell clones carry exactly the same BCR is extremely low (e.g., less than 10−12). CLL cells isolated from different patients often express similar, if not identical, BCRs with common stereotypic features and/or structural similarities. IGHVs, such as IGHV1–69, IGHV4–34, and IGHV3–7, are used by CLL cells at higher frequencies than those observed in normal B cells Incidence of somatic hypermutation is not uniform among IGHVs in CLL cells. The marked restriction in the Ig gene repertoire of CLL cells highlights the role played by one or more common self-or environmental antigens in leukemic B cell selection
- 40. ANTIGENS THAT MAYPLAY A ROLE IN LEUKEMIA B CELL SELECTION Some of the Ig expressed in CLL can react with antigen expressed by cells undergoing apoptosis, including cytoskeletal proteins Some Ig react with non muscle myosin heavy chain IIA, which is expressed on some apoptotic cells, namely myosin-exposed apoptotic cells (MEACs). Binding to MEACs is more commonly observed on CLL cells expressing unmutated IGHVs than on CLL cells expressing mutated IGHVs. In addition to self-antigen, several other microbial or virus-associated antigens may contribute to the selection of the Ig expressed in CLL.
- 41. GENETIC ALTERATIONS INCHRONIC LYMPHOCYTIC LEUKEMIA CLL cells commonly harbor deletions at 13q14, 11q22–q23, or 17p13 or may have an extra copy of chromosome 12 (trisomy 12) The advent of next-generation sequencing technologies, coupled with gene copy-number analyses, have identified additional genetic lesions in CLL, such as mutations in NOTCH1, SF3B1, and BIRC3 Such mutations could be used as potential therapeutic targets or as biomarkers that can distinguish among patients who may have disparate clinical outcomes
- 42. EVIDENCE OF BCRSIGNALING AS CLL DRIVER Strong association of somatic hypermutation status with overall survival IgM rarely lost in CLL sIgM levels have clinical implications Specific and limited V gene use Steriotyped BCRs Evidence for autonomous BCR activity Universal response to BCR pathway inhibition Resistance associated with pathway mutation
- 43. Pattern of ClonalEvolution and Antigen Dependence in CLL GERMINAL CENTER B CELL ANTIGEN MUTANT SUB CLONE WITH DYSREGULATED DRIVE BUT STILL RESPONSIVE TO ANTIGEN MUTATIONS CONVERTING BCR TO AUTONOMOUS ACTIVITY
- 45. NOTCH1 Encodes aligand-activated transcription factor Regulates several downstream pathways that induce the differentiation of hematopoietic progenitors immature T cells and of mature B cells antibody-secreting cells Activating mutations in NOTCH1 occur in ∼60% of T-lineage acute lymphoblastic leukemias. In CLL, activating NOTCH1 mutations have been detected in ∼10% of newly diagnosed cases. NOTCH1 mutations are also more frequent in CLL cell populations that express unmutated IGHVs and that have trisomy 12.
- 46. NOTCH1 mutations Restricted to the C-terminal PEST [proline (P), glutamate (E), serine (S), and threonine (T)] domain normally limits the intensity and duration of NOTCH1 signaling. Removal of the PEST domain impairs the degradation of NOTCH1 allowing accumulation of the active form of NOTCH1. One recurrent mutation (c.7544_7545delCT) accounts for ∼77% of all NOTCH1 mutations in CLL- Can be rapidly detected by a simple polymerase chain reaction–based strategy- providing a potential approach for a first-level screening of NOTCH1 alterations.
- 47. SF3B1 Encodes thesplicing factor 3B sub-unit 1 (SF3B1) Mutations in SF3B1 were observed in ∼10% of newly diagnosed CLL cases and in ∼17% of cases with progressive, late-stage disease requiring therapy. SF3B1 mutations- acquired during clonal evolution, the proportionate representation of sub-clones harboring SF3B1 mutations can increase over time, independently of cytoreductive therapy. SF3B1 regulates the alternative splicing program of genes controlling cell cycle progression and apoptosis Mutations in SF3B1 may enhance CLL cell proliferation and/or survival
- 48. BIRC3 BIRC3 encodesthe baculoviral inhibitor of apoptosis (IAP) repeat Contains 3 protein (BIRC3)- can inhibit apoptosis by binding to tumor necrosis factor (TNF) receptor– associated factors 1 and 2 (TRAF1 and TRAF2) possibly by interfering with activation of ICE-like proteases (caspases). BIRC3 – - act as an E3 ubiquitin–protein ligase that regulates nuclear factor κB (NF-κB) signaling- -acting to promote canonical NF-κB signaling while suppressing constitutive activation of noncanonical NF-κB signaling. Activation of canonical NFκB signaling can promote the growth and survival of CLL cells in vitro and in vivo.
- 49. BIRC3 mutationsin CLL are predicted to disrupt the C-terminal RING domain, essential for proteasomal degradation of MAP3K14 by BIRC3 CLL cells harboring mutations in BIRC3 display constitutive NF-κB activation appear less responsive to conventional chemotherapy than CLL cells without such mutations Inhibitors of NF-κB may have clinical activity in CLL-particularly in cases harboring alterations in BIRC3.
- 50. MYD88 MYD88 encodesa critical adaptor molecule of the Toll-like receptor (TLR) Mutations in MYD88 are observed in 3% to 10% of CLL cases at diagnosis Also mutated in other B cell malignancies- lymphoplasmacytic lymphoma diffuse large B cell lymphoma (DLBCL) marginal zone B cell lymphoma. Mutations in other genes encoding proteins involved in the activation of TLR signaling and NF-κB have also been observed in DLBCL. In contrast to the mutations in NOTCH1 or SF3B1, mutations in MYD88 appear to be present in the vast majority (if not all) cells within the CLL clone; MYD88 appears to be an early driver mutation in at least a subset of CLL cases.
- 54. TREATMENT In general,it is estimated that one third of CLL patients never require therapy one third need treatment as soon as they are seen one third have disease progression over the years and require therapy at some point. The indications to treat in CLL have been reviewed and are as follows: 1. Rai stage 0–II disease in patients who are symptomatic (weight loss of >10% body weight in previous 6 months, extreme fatigue, night sweats or fevers of >100.5°F for >2 weeks without evidence of infection), have progressive anemia/thrombocytopenia or lymphocytosis (>50% increase over 2 months or doubling time <6 months) 2. Rai stages III/IV to improve the hemoglobin level and/or platelet counts, although asymptomatic patients can be monitored and treatment initiated when there is clear evidence of disease progression 3. Bulky or progressive lymphadenopathy (masses >10 cm in diameter) or splenomegaly (>6 cm below left costal margin) 4. AIHA/ITP
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- 57. TREATMENT COMPONENTS 2. COMBINATIONCHEMOTHERAPY 4. COMBINATIONS USING TARGETED THERAPY 1. SINGLE AGENTS a) Cytostatic agents b) Monoclonal antibodies c) Agents targeting B-cell receptor (ibrutinib) d) BCL-2 inhibitor e) Immunomodulatory drugs 3. CHEMOIMMUNOTHEREPY a) Rituximab b) Ofatumumab c) Alemtuzumab d) Obinutuzumab
- 59. TARGETING PATHOGENESIS ACTIVATINGTRANSLOCATIONS: if they affect a targetable gene BCR, cyclin D1, MYC ACTIVATING MUTATIONS: if they affect a targetable gene and are true disease drivers Kinases CONSTITUTIVELY OR ABERRANTLY ACTIVATED PATHWAYS THAT ARE NOT MUTATED- BCR signaling
- 61. TARGETED THERAPY Forpatients who have stopped responding to treatment new drug combination may be effective option Based on phase 3 clinical trials venetoclax and rituximab reduced the risk of cancer progression by more than 80% We are entering an era in which CLL treatment is not one size fits all but instead tailored to the biology of each person’s disease, depending on markers and genetic mutations. Clinicians now recommend that all patients with CLL to get some sort of genetic screening.
- 62. TARGETED THERAPY (Cont.) First class drug works by interfering with B cell receptors. These receptors are crucial for proper proliferation- if they go rogue- turn cancerous. New drugs take advantage of this by jamming this signalling system, creating agents that bind to receptors and block those that allow cancerous cells to multiply or by interfering with kinases.
- 63. IBRUTINIB First-in-class Brutontyrosine kinase (BTK) inhibitor Blocks B cell receptor (BCR) signaling, a key pathway for CLL cell survival and proliferation. Has potent activity even in high-risk groups such as previously treated CLL or CLL with TP53 aberrations Approved for all CLL patients based on improved progression free survival in treatment-naive disease and a favorable safety profile Increasingly used as monotherapy or tested in combination regimens.
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- 65. IMBRUVICA FDA approved Blocks Bruton’s tyrosine kinase (BTK), and several other kinases For patients with 17p genetic mutation this is first line treatment
- 66. ZYEDELIG( IDEALASIB) FDAapproved Blocks phosphoinositide 3 kinase (PI3K) Most affective when combine with Rituximab
- 67. VENCLEXTA FDA approved Targets BCL-2 protein that blocks apoptosis
- 68. Others GAZYVA( obinutuzumab)-CD 20 monoclonal antibody Approved as first line treatment in CLL in combination with the chemotherapy drug chlorambucil ARZERRA (ofatumumab)- has been approved in later lines of therapy
- 73.