THE HALLMARKS OF CANCER

THE HALLMARKS OF CANCER

ONCOGENES - Most of the oncogenes were first identified in retroviruses: EGFR (ErbB), Src, Ras, Myc, PI3K and others (slightly more than 30) - Mutated cellular genes incorporated in the viral genome (viruses as carriers of host genes)

Retroviruses can cause cancer by picking up mutated versions of normal cellular genes

ONCOGENES

COMMON PROTO-ONCOGENE ALTERATIONS

FUNCTIONAL CLASSES OF ONCOGENES 1. Growth factors: PDGF, EGF 2. Growth factor receptors: EGFR, FLT3 3. Signal transduction proteins: SRC, RAS, PI3K 4. Proteins that control cell cycle: cyclins, CDK 5. Proteins that control cell survival and/or apoptosis: Bcl-2 6. Transcription factors: myc, NF-kB

1. ONCOGENES: GROWTH FACTORS NGF (1952 Rita Levi Montalcini): secreted by murine cancer cells that promoted neurite outgrowth EGF (1962 Cohen): first proliferation factor for epithelial cells thta induced precocius eyelid opening and tooth eruption when injected in newborn mice 1986 Nobel prize to Cohen to Levi-Montalcini Others PDGF, Ins, IGF, FGF etc

2. ONCOGENES: RECEPTOR TYROSINE KINASES (RTK) Hubbard & Till 2001

EGFR FAMILY Extracellular domain, leucin rich - EGFR (HER1 or ErbB1) - HER2 (neu or ErbB2): no ligands, heterodimers with ErbB1 - HER3 (ErbB3): no intrinsic enzymatic activity - HER4 (ErbB4) Functional domains of interaction: SH2 (100 aa): binds tyrosine phosphorylated sequences SH3 (100 aa): binds proline-rich sequences PH: binds phospholipids (es. PIP3 in Akt, PDK1)

RECRUITMENT OF PROTEINS THROUGH THE SH2 DOMAINS

EGFR SIGNALLING Grb2-SOS: activates RAS/Raf/ERK pathway > Elk1 (c-fos) and c-myc PLC: hydrolysis of PIP2 in IP3 (Ca2+) and DAG (PKC) > NF-kB Vav1 and Nck: Rac1 GTPase (actin cytoskeleton reorganization) PAK1/MEEK/JNK > c-jun that together with c-fos forms AP-1 STAT1/STAT3 nuclear translocation transcriptional activation PI3K/Akt/mTOR: survival, metabolism and protein synthesis

PI3K

DEREGULATION OF EGFR IN HUMAN CANCERS 1980: Overexpression of EGFR in several epithelial tumors and autocrine stimulation Point Mutation and deletions resulted in increased catalytic activity Deletion of exons 2-7 (EGFRvIII): induced by rearrangement or alternative mrna splicing > constitutive active EGFR without an extracellular domain.

v-erbb: EGFR without an extracellular domain, constitutive activity v-erb: avian erythroblastosis virus

HER2 (1985) HER2 is the preferred heterodimerization partner for EGFR, HER3 and HER4, - Controls cell growth, proliferation, differentiation, apoptosis and remodelling of mammary gland Alteration of HER2 receptor in mammary and ovarian tumors (HER2 gene is amplified in 30% of invasive breast cancers) - Most common mutation Val- Gln664 substitution > tyrosine kinase (TK) with a constitutive activity

INCREASED EXPRESSION OF HER2 - Induces high survival and cell proliferation - Increases the metastatic potential of tumour cells > synthesis and secretion of matrix metalloproteinases (MMPs) - Overexpression of HER2 is associated with a poor prognosis in patients with mammary carcinoma - HER2 as a target of novel therapeutic approaches

Specific therapy for RTK

Second generation of EGFR mabs

NSCLC, non small cell lung cancer; mcrc, metastatic colorectal carcinoma HNSCC, head and neck squamous cell carcinoma

FLT3 (FMS-like tyrosine kinase 3) FLT3 is a RTK regulating proliferation, differentiation and survival of staminal haemopoietic cells FLT3 Expressed by early myeloid and lymphoid progenitors FLT3L is produced by different haemopoietic cell types of spleen, thymus, bone marrow and peripheral blood. FLT3L in prostate, ovary, lung, gut, testis, kidney, placenta. NO BRAIN

STRUCTURE AND SIGNALLING RTK of 993 amino acids with an extracellular domain of five Ig-like domains, a transmembrane domain, and two intracellular tyrosine kinase domains (TKDs) linked by a kinase-insert domain FLT3 receptors reside as inactive monomers in the plasma membrane Ligand-mediated dimerization induces a conformational changes with activation of the TKD

FLT3 FUNCTIONS IN NORMAL CELLS Activation of FLT3 promotes the in vitro growth of early progenitor cells Promotes the monocytic differentiation of early haematopoietic progenitor cell without proliferation if stimulated in the absence of other factors FLT3 stimulation in combination with other growth factors (IL-3, GM- CSF ) produced a more vigorous proliferative response Stimulation of FLT3 promotes dendritic-cell development and expansion in the bone marrow, spleen, lymph nodes and peripheral blood Stimulation of FLT3 in combination with IL-7 and IL-11 is also important in T and B lymphoctye development.

FLT3 MUTATIONS - Identified and described for the first time in 1996 by Nakao et al. - Mutated in patients with acute lymphoblastic leukaemia (ALL, 1 3%), myelodysplasia (5 10%), acute myeloid leukaemia (AML, 15 35%) - Most frequent mutation: in tandem duplication (ITD) of exons 14 and 15. - Mutations within the tyrosine kinase domain (TKD): D835Y most frequent

ITD AND POINT MUTATIONS INDUCE THE CONSTITUTIVE ACTIVATION OF FLT3 - ITDs probably promote ligand-independent dimerization and activation of FLT3 through changing the conformation of the expressed receptor - Missense mutations in TKD might also promote ligand independent activation.

FLT3 SPECIFIC THERAPY CEP-701: inhibits autophosphorylation of mutated and wt Flt3, highly specific Sugen, MLN518, PKC412: also inhibit other RTK, such as PDGFR and VEGFR

3. ONCOGENES: NON-RECEPTOR TYROSINE KINASES (SRC)

C-SRC VS V-SRC Identified in 1970 within the genome of Rous sarcoma virus

REGULATION OF SRC ACTIVITY PTP1B CSK C-terminal Y is phosphorylated by Csk and binds to the SH2 domain of Src In this conformational status the SH3 domain interacts with the kinase domain thus causing the c-src molecule to assume a closed configuration that covers the kinase domain and reduces its potential for substrate interaction Dephosphorylation of C- terminal Y by phosphatases (i.e. PTPa or PTP1B) induces the activation of Src

CELLULAR TARGETS OFSRC

SRC AND CELL ADHESION - E-cadherin: A cytoplasmic complex consisting of α-catenin, β-catenin and p120 catenin (p120ctn) links E-cadherin homodimers to the actin cytoskeleton. c-src associates with this complex and, when activated, is able to promote the disruption of the adherens junction. - Focal adhesions, heterodimers of α- and β-integrin subunits bind the extracellular matrix through their extracellular domains. Their cytoplasmic domains bind to a complex consisting of a range of proteins, including paxilin, talin, vinculin, tensin and α-actinin, which connect integrins to the actin cytoskeleton. Several signalling molecules also associate with this complex, c-src, which can promote the turnover of the focal adhesion when activated, to promote cellular motility > disruption of focal adhesion

-c-src stimulates the ubiquitylation of E- cadherin, leading to its endocytosis. - c-src induces tyrosine phosphorylation and activation of FAK, which results in the loss of focal adhesions. - c-src phosphorylates RRAS and inhibits integrin function. - Motility and invasiveness - Angiogenesis

SRC and TUMORS Increased activity of SRC in colorectal, hepatocellular, pancreatic, gastric, mammary, ovarian and lung tumors Inhibitors of Src (Sugen, SKI-606, AP23464) in clinical trials > effective only in solid tumors with high level of Src activity