Biochemistry of cancer 101
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General Topic on Cancer Biochemistry, current reviews for MBBS, BDS and other UG classes
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Biochemistry of cancer 101
- 1. Biochemistry of Cancer Glimpse of chemistry, pathways & tumor markers
- 3. Introduction ▪ A disease caused by an uncontrolled division of abnormal cells in a part of the body. ▪ Cancer is characterized by ▪ Unrestrained cell growth ▪ Cell Immortality ▪ Local invasion & Distant metastasis ▪ Cancer is the second most important cause of death world-wide ▪ Can affect potentially any person with any demographic parameters (Age, gender, ethnicity, geographical location etc.). Cancer Chemistry 3
- 4. Causes & Characteristics ▪ Exact cause is unknown. ▪ Found to be associated various factors, both internal & external ▪ External factors are called Carcinogens ▪ Association & Causation of external factors ▪ Carcinogens can be chemical, physical, infectious agents (viral & bacterial). ▪ Internal factors so far identified are inherited genetic changes Cancer Chemistry 4
- 5. Two types of growth ▪ Benign: ▪ Well-defined mass of tissue ▪ demarcated from normal tissue ▪ slow-growing ▪ Mostly differentiated cells ▪ no invasion ▪ no distant metastasis. ▪ Malignant ▪ Ill-defined tissue growth, ▪ no demarcation from normal tissue, ▪ can be rapidly growing; ▪ poorly differentiated cells ▪ local invasion ▪ distant metastasis are usual. Cancer Chemistry 5
- 6. Two types of spread ▪ Local Invasion: ▪ Early stage of the disease ▪ Direct migration of cancer cells ▪ Penetration of basement membrane ▪ Distant metastasis: ▪ Later stage of the disease ▪ Direct migration of cancer cells or indirectly materials from cancer cells such as DNA ▪ Usually penetration and transport via lympho- vascular system Cancer Chemistry 6
- 7. External Factors (Carcinogens) ▪ Physical: Harmful Radiations (Ultraviolet, X and radiations) ▪ Chemical: from lifestyles (alcohol & cigarette smoking), diet (aflatoxins) etc. Class Compound Polycyclic aromatic hydrocarbon Benzo(a)pyrene, Dimethyl benzanthracene Aromatic amines Acetylaminofluorene, aminobenzene Nitrosamines Dimethyl and diethyl nitrosamines Drugs Cyclophosphamide Naturally occurring compounds Aflatoxin B1, Dactinomycin Inorganic compounds Arsenic, Asbestose, beryllium, cadmium, chromium Cancer Chemistry 7
- 8. External Factors (Carcinogens) ▪ Biological factors: Oncogenic viruses Virus Nucleic Acid Associated cancer Epstein Barr virus (EBV) dsDNA Burkitt lymphoma, Nasopharyngeal carcinoma Hu papilloma virus (HPV) dsDNA Uterine, Cervical carcinoma Hepatitis B virus (HBV) dsDNA - RT Hepatoma & Hepatocellular Carcinoma Cancer Chemistry 8
- 9. Biochemical Mechanisms What are the different cellular and molecular mechanisms involved in Cancer?
- 10. Mechanisms for tumor cell formation ▪ What happens when a normal cell becomes tumor cell? ▪ External or tumor microenvironment changes. ▪ Internal acquired/inherited cellular growth mechanisms Cancer Chemistry 10
- 11. External factors or Tumor- Microenvironment or Extracellular- Matrix (ECM) Cell 2011 144, 646-674DOI: (10.1016/j.cell.2011.02.013) Cancer Chemistry 11
- 12. Growth factors ▪ Abnormal amount of growth factors to sustain proliferative signaling or their increased susceptibility for certain types of cells. E.g. Insulin Like Growth Factor, Epidermal Growth Factor, Platelet Derived Growth Factor etc. Estrogen and Bread adenoma. Estrogen receptor hypersensitivity in Ca breast Cancer Chemistry 12
- 13. Mucopolysaccharides ▪ Stromal Contact inhibitors: Molecules which affect contact inhibition of cell proliferation such as Sialic acid and Hyaluronic acid residues. Due to higher content of these negatively charged long residues there is a loss of orientation of cells and repel apart. Cancer Chemistry 13
- 14. Cell::Cell interaction ▪ Abnormal cell adherence junction (tight junction) due to altered protein interaction due to structural modification ▪ Loss Apico-basal polarity due to abnormal regulation of distribution of protein ▪ Abnormal anchoring proteins to ECM e.g. integrins, fibronectin, vimentin etc. due to protein structure abnormality. Cancer Chemistry 14 ERM NF2 / Merlin Apical Lateral Dynamic cell:cell adhesion Stable cell:cell adhesion Brit. J. Cancer 2008; BBA 2007
- 15. Acetate and SCFA produced by microbiota of gut cells regulate abnormal growth of colonic cells Cancer Chemistry 15
- 16. External factors Altered immunity: 1. Abnormal recruitment Regulatory T-cells (T- regs) which controls the specific local infiltration of T- lymphocytes to weed out Cancer cells is checked. 2. Regulated Immuno “rheostat” or immunostat or Check- point inhibitors such as PD-1/PD-L1 Cancer Immunity 2013 39(1), 1-10 DOI: (10.1016/j.immuni.2013.07.012) Cancer Chemistry 16
- 17. CONFLUENCE OF EXTERNAL MECHANISMS IS THE RULE FOR MALIGNANT PROGRESSION Cancer Chemistry 17
- 18. Internal Factors ▪Acquired changes. ▪Hereditary changes. Cancer Chemistry 18
- 19. Broad Cellular Mechanisms Cell 2011 144, 646-674DOI: (10.1016/j.cell.2011.02.013) Cancer Chemistry 19
- 20. Internal Changes: Intracellular signaling cell viability circuits Cell 2011 144, 646-674DOI: (10.1016/j.cell.2011.02.013) Cancer Chemistry 20
- 21. Cell 131, December 14, 2007 p1204.e1–1204.e2 DOI 10.1016/j.cell.2007.11.036 Cancer Chemistry 22
- 22. Cell 131(5), December 30, p1018.e1–1018.e2 DOI 10.1016/j.cell.2007.11.013 Cancer Chemistry 23
- 23. Cell 133 (7) June 21, 2008 p1292–1292.e1 Cancer Chemistry 24
- 24. Cell 152 (3) June 21, 2008 p656–656.e1 Cancer Chemistry 25
- 25. Cell 133, May 2, 2008 DOI 10.1016/j.cell.2008.04.023 Cancer Chemistry 26
- 26. Snapshot: Tumor Angiogenesis Cell 149(6) p1408–1408.e1 8 June 2012 DOI 10.1016/j.cell.2012.05.025 Cancer Chemistry 27
- 27. Cancer Energetics Where does cancer cells get energy from?
- 28. Warburg Effect ▪ Otto Heinrich Warburg (1883-1970) ▪ Identified that most cancer cells predominantly produce energy by high rate of glycolysis followed by lactic acid fermentation in cytosol even when plenty of oxygen is available. ▪ Nobel Prize in 1931 Cancer Chemistry 29
- 29. Mechanism of Warburg Effect Cancer Chemistry 30
- 30. Uses of Warburg effect Diagnostic ▪ 2-fluoro-2-deoxy glucose (FdG) (18F replaced C2 of glucose. ▪ Cancer cells uptake 10x more, trapped in cancer cells as 6-phosoho FdG ▪ Decay of 18F gives positron which are detected by Positron Emission Tomography Cancer Chemistry 31 Therapeutic ▪ Gleevec (Imatinib) inhibits Tyrosine Kinase enzyme preventing hexokinase activation is a clinically approved drug. ▪ Oxythiamine which inhibit transketolase enzyme in preclinical trials.
- 31. Changes in DNA What are the biochemical changes in the informational molecules? Cancer Chemistry 32
- 32. Categories of DNA modifications Types of Genes involved in cancer Oncogenes Tumor Suppressor Genes Stability Genes Cancer Chemistry 33
- 33. Oncogenes ▪ Oncogenes are mutant forms of the genes for proteins that regulate cell cycle. ▪ Are originally discovered from viruses, which was later found to be DNAs incorporated from animal hosts of their precursor viruses, called proto- oncogenes. Cancer Chemistry 34
- 34. Oncogenes ▪ Several mechanisms of oncogene formation ▪ Viral incorporation Replication errors in virus Reincorporation into human cells failed regulation of cell division ▪ Selective dominance: Mutations in certain genes give characteristic dominance in specific proteins which trigger for cell proliferation. E.g. nuclear transcription factors which control cell division (Jun, Fos, etc.). ▪ Activating mutations: Oncogene mutations lead to spontaneously activating mutations in certain growth factor cell membrane receptor. E.g. Mutations of oncoprotein ErbB lead to spontaneous activation of the EGF receptor, without binding of ligand. Cancer Chemistry 35
- 35. Tumor suppressor gene ▪ Tumor suppressor genes encode proteins that normally restrain cell division. ▪ Mutations of one/ more of these genes can lead to tumor formation ▪ Usually genetically recessive. (unlike oncogenes). ▪ Second hit: If one copy of the gene is mutated congenitally, and if a second copy is mutated in any of the 1012 cells of the body, tumor could potentially start forming from that cell. Cancer Chemistry 36
- 36. Tumor suppressor genes ▪ Retinoblastoma gene (Rb): Second hit usually associated retinoblastoma and many types of tumors such as cancers of lung, prostate, breast etc. later in life. ▪ Von Hippel Lindau (VHL) gene: Congenital absence of p.VHL protein lead to uninhibited vasculogenesis in cerebellar hemisphere (called hemangioblastoma), also associated with multiple cysts in kidneys, liver, pancreas etc. Cancer Chemistry 37
- 37. Stability genes ▪ Also known as Caretaker genes: e.g. ATM, BRCA1 gene family, XP gene family, TP53 etc. ▪ The proteins encoded by these genes function in the repair of major genetic defects resulted from aberrant DNA replication, ionizing radiation, or carcinogens. ▪ TP53 : called “Guardian of human genome”. This gene ▪ Activate DNA repair ▪ Arrest growth by holding cell cycle at G1/S phase ▪ Can initiate apoptosis (or programmed cell death) ▪ Senescence response to short telomeres ▪ Congenital defects of p53 protein lead to a rare disorder called Li- frumeni syndrome (associated with cancers in multiple organs). Cancer Chemistry 38
- 38. Overall Cancer generally results from accumulated genetic changes or mutations to oncogenes, tumor suppressor genes and stability genes over a period of time. Cancer Chemistry 39
- 39. What are the chemical changes in DNA? ▪ Point mutations: Substitutions (missense/nonsense), Deletion/duplications one or two nucleotide bases in the DNA ▪ Deletions/duplications of short stretch of DNA: called indels, micro- indels. ▪ Gene Rearrangements: of large regions of chromosomes giving some peculiar proteins called fusion proteins which gives some cell proliferative properties. For e.g. BCR-ABL fusion of in Chronic Myeloid Leukemia (CML). Entire ABL gene in chromosome 9 is juxtaposed/fused onto entire BCR gene of chromosome 22 producing a hybrid protein which gives uncontrolled tyrosine kinase activity for cell growth in certain cells of bone-marrow which led to CML. Cancer Chemistry 40
- 40. Tumor markers Proteins that can portend!
- 41. What are tumor markers? ▪ Factors released by tumor cells, detected in blood and other body fluids and potentially indicate the presence of tumor in the body. ▪ Uses ▪ Diagnostic: Mere presence could be a sign of tumor, but cautious of non-malignant conditions ▪ Prognostic: Serum levels may roughly indicate the tumor load, which can predict the effect of treatment ▪ Localization: Certain tumor markers are suggestive of tumors in certain specific organs or part of organs. ▪ Therapeutic: Once the treatment is started, progressive determination of serum levels could mirror tumor remission. Cancer Chemistry 42
- 42. Classical tumor markers Cancer Chemistry 43
- 43. Clinically important Tumor markers 1. Alpha-fetoprotein (AFP) 2. Carcinoembryonic antigen (CEA) 3. Beta Human Chronic Gonadotrophin (-HCG) 4. Cancer Antigen 125 (CA-125) 5. Tissue Polypeptide Antigen (TPA) 6. Prostate Specific Antigen (PSA) Cancer Chemistry 44
- 44. Alpha fetoprotein (AFP) ▪ Fetal albumin like protein, about 70 kDa molecular weight. ▪ In adult males and non-pregnant females, it can be upto 15 ng/ml ▪ A value of upto 300 ng/ml can occur nonmalignant liver disease ▪ More than 300 ng/ml is usually associated with cancer ▪ Associated with mostly hepatocellular carcinoma. Cancer Chemistry 45
- 45. Carcinoembryonic antigen (CEA) ▪ Set of highly related glycoprotein closely belong to immunoglobulin superfamily by 29 genes. ▪ Normally produced by fetal gastrointestinal system, stops production by birth ▪ Molecular weight: as large as 185 kDa. ▪ Mostly elevated in adenocarcinomas of colon, lung, breast, stomach & pancreas (above approximately 2.5 µg/L) ▪ Left sided colon cancers have higher levels ▪ Higher levels are associated with lymph node spread. Cancer Chemistry 46
- 46. Cancer Antigen 125 (CA-125) ▪ A glycoprotein of more than 1 MDa molecular weight. ▪ Normal level in the blood is < 35 U/ml ▪ Elevated levels are associated with 75% of ovarian cancers. ▪ Elevated levels can also be found in 20% of pancreatic and GI cancers. Cancer Chemistry 47
- 47. -HCG ▪ Synthesized by syncytio-trophoblasts of placental villi ▪ Molecular weight of HCG is ~ 45 kDa. ▪ HCG has an -subunit shares common structure with FSH, LH & TSH. ▪ -subunit is unique to HCG. ▪ Increased in hydatidiform mole, choriocarcinoma & germ cell tumor ▪ Also associated with 60% of testicular cancers. ▪ Normal value is 20 IU/L (> 10,000 IU/L is trophoblastic tumor) Cancer Chemistry 48
- 48. Bence-Jones Proteins (BJP) ▪ Abnormal production of immunoglobulin when plasma cells proliferate. ▪ Plasmacytoma/Multiple myeloma ▪ Characterized by lytic bone lesions, anemia, para-proteinemia, proteinuria. ▪ BJP is seen in 20% cases of multiple myeloma Alpha-I Beta Gamma Albumin Cancer Chemistry 49
- 49. Newer tumor markers ▪ Estrogen Receptor (ER) ▪ Progesterone Receptor (PR) ▪ Beta2-macroglobulin ▪ Bladder tumor Antigen (BTA) ▪ Calcitonin ▪ Her2/neu ▪ Neuron Specific Enolase (NSE) ▪ Thyroglobulin (Tg) ▪ Thyroid Transcription Factor- 1 (TTF-1) ▪ Cytokeratin-7 (CK-7) ▪ Synaptophysin Cancer Chemistry 50
- 50. Thank you Dr. Prasanth. A. S.
Editor's Notes
- The Cells of the Tumor Microenvironment (Upper) An assemblage of distinct cell types constitutes most solid tumors. Both the parenchyma and stroma of tumors contain distinct cell types and subtypes that collectively enable tumor growth and progression. Notably, the immune inflammatory cells present in tumors can include both tumor-promoting as well as tumor-killing subclasses. (Lower) The distinctive microenvironments of tumors. The multiple stromal cell types create a succession of tumor microenvironments that change as tumors invade normal tissue and thereafter seed and colonize distant tissues. The abundance, histologic organization, and phenotypic characteristics of the stromal cell types, as well as of the extracellular matrix (hatched background), evolve during progression, thereby enabling primary, invasive, and then metastatic growth. The surrounding normal cells of the primary and metastatic sites, shown only schematically, likely also affect the character of the various neoplastic microenvironments. (Not shown are the premalignant stages in tumorigenesis, which also have distinctive microenvironments that are created by the abundance and characteristics of the assembled cells.)
- Abnormal cell adherence junction (tight junction): lead to loss of contact dependent restriction of proliferation of normal cells. This would lead to proliferation, migration and invasion of cells. Loss of E-cadherin, Curto M et al., Brit. J. Cancer 2008; Fivet et al., BBA 2007
- Production of acetate by the commensal microbiota and the subsequent uptake by colonic epithelial cells: Resistant starches and other indigestible polysaccharides are broken down on the surface of bacterial cells by carbohydrate-degrading enzymes (top panel). These hexoses enter the cell and are converted, through glycolysis, into pyruvate. Pyruvate in turn is converted into acetyl-CoA and CO2. Phosphotransacetylases exchange a phosphate for CoA to form acetyl-phosphate (acetyl-P). Acetate kinase catalyses the transfer of the phosphate from acetyl-P to ADP, which generates ATP and acetate. Alternatively, the CO2 released from pyruvate decarboxylation can enter the Wood–Ljungdahl pathway, which uses reductive acetogenesis to form acetyl-CoA from CO and 5-methyl-tetrahydrofolate (5-MTHF). Once formed, acetate can be exported from the bacterial cell into the lumen of the gut, where it is absorbed by one of two methods by the colonic epithelium (bottom panel). The first mechanism is passive diffusion of the protonated form of acetate, acetic acid, which is facilitated by the extremely high concentrations of acetate and the relatively low pH in the proximal colon and caecum. The second route of entry is through the active transport of acetate by monocarboxylate transporters (MCTs), in which acetate is co-transported with Na+ or H+ or exchanged for HCO3−. Dashed lines represent multistep reactions. 10-formyl-THF, 10-formyl-tetrahydrofolate; GAP, glyceraldehyde-3-phosphate; glucose-6-P, glucose-6-phosphate; fructose-6-P, fructose-6-P; SMCT, sodium-coupled MCT; THF, tetrahydrofolate doi:10.1038/nrc.2016.87 Nature Reviews Cancer 16, 615 (2016)
- Stimulatory and Inhibitory Factors in the Cancer-Immunity Cycle Each step of the Cancer-Immunity Cycle requires the coordination of numerous factors, both stimulatory and inhibitory in nature. Stimulatory factors shown in green promote immunity, whereas inhibitors shown in red help keep the process in check and reduce immune activity and/or prevent autoimmunity. Immune checkpoint proteins, such as CTLA4, can inhibit the development of an active immune response by acting primarily at the level of T cell development and proliferation (step 3). We distinguish these from immune rheostat (“immunostat”) factors, such as PD-L1, can have an inhibitory function that primarily acts to modulate active immune responses in the tumor bed (step 7). Examples of such factors and the primary steps at which they can act are shown. Abbreviations are as follows: IL, interleukin; TNF, tumor necrosis factor; IFN, interferon; CDN, cyclic dinucleotide; ATP, adenosine triphosphate; HMGB1, high-mobility group protein B1; TLR, Toll-like receptor; HVEM, herpes virus entry mediator; GITR, glucocorticoid-induced TNFR family-related gene; CTLA4, cytotoxic T-lympocyte antigen-4; PD-L1, programmed death-ligand 1; CXCL/CCL, chemokine motif ligands; LFA1, lymphocyte function-associated antigen-1; ICAM1, intracellular adhesion molecule 1; VEGF, vascular endothelial growth factor; IDO, indoleamine 2,3-dioxygenase; TGF, transforming growth factor; BTLA, B- and T-lymphocyte attenuator; VISTA, V-domain Ig suppressor of T cell activation; LAG-3, lymphocyte-activation gene 3 protein; MIC, MHC class I polypeptide-related sequence protein; TIM-3, T cell immunoglobulin domain and mucin domain-3. Although not illustrated, it is important to note that intratumoral T regulatory cells, macrophages, and myeloid-derived suppressor cells are key sources of many of these inhibitory factors. See text and Table 1 for details. Oncology Meets Immunology: The Cancer-Immunity Cycle Volume 39, Issue 1, 2013, 1–10 http://dx.doi.org/10.1016/j.immuni.2013.07.012
- Signaling Interactions in the Tumor Microenvironment during Malignant Progression (Upper) The assembly and collective contributions of the assorted cell types constituting the tumor microenvironment are orchestrated and maintained by reciprocal heterotypic signaling interactions, of which only a few are illustrated. (Lower) The intracellular signaling depicted in the upper panel within the tumor microenvironment is not static but instead changes during tumor progression as a result of reciprocal signaling interactions between cancer cells of the parenchyma and stromal cells that convey the increasingly aggressive phenotypes that underlie growth, invasion, and metastatic dissemination. Importantly, the predisposition to spawn metastatic lesions can begin early, being influenced by the differentiation program of the normal cell-of-origin or by initiating oncogenic lesions. Certain organ sites (sometimes referred to as “fertile soil” or “metastatic niches”) can be especially permissive for metastatic seeding and colonization by certain types of cancer cells, as a consequence of local properties that are either intrinsic to the normal tissue or induced at a distance by systemic actions of primary tumors. Cancer stem cells may be variably involved in some or all of the different stages of primary tumorigenesis and metastasis.
- Perhaps these are the hallmarks of the Cancer as well.
- Intracellular Signaling Networks Regulate the Operations of the Cancer Cell
- Cell 131, December 14, 2007 ©2007 Elsevier Inc. DOI 10.1016/j.cell.2007.11.036 Molecules labeled in green and blue play positive and negative roles in Wnt/β-catenin signaling, respectively. Molecules that are labeled in both colors have dual roles. (Top left) Wnt biogenesis. A lipid modification is added to Wnt ligands by Porc in the endoplasmic reticulum (ER). Wnt ligands are glycosylated in the ER and Golgi and require Wls (also known as Evi) to traffic through the Golgi to the plasma membrane. The retromer complex is also important for Wnt secretion, particularly for long range signaling. Mature Wnt ligands may interact with lipoprotein particles. Proteoglycans Dally and Kny/Glypican may also facilitate Wnt distribution. (Bottom right) Wnt receptor biogenesis. In cells that respond to Wnt the ER protein MESD is required for folding and trafficking of the Wnt receptor LRP5/6 to the plasma membrane, and the ER protein Shisa prevents folding and trafficking of the Fz protein to the plasma membrane. (Right) Wnt/β-catenin signaling OFF. sFRP and WIF1 directly bind Wnt ligands and prevent Wnts from binding to receptors. SOST/WISE, CTGF/Cyr61 bind to LRP6 (CTGF may also bind to Fz) to prevent the formation of the Wnt-Fz-LRP5/6 receptor complex. DKK binds to and inhibits LRP5/6 in cooperation with the KRM receptor. In the absence of Wnt signaling, the scaffolding protein Axin and tumor suppressor APC form a β-catenin destruction complex that binds cytosolic β-catenin and facilitates sequential phosphorylation of β-catenin by CK1 (at S45) and GSK3 (at S33/S37/T41). The tumor suppressor WTX may also reside in this complex. Phosphorylated β-catenin is recognized by β-Trcp and ubiquitinated for degradation by the proteasome. In the nucleus, TCF assembles a transcriptional repressor complex to silence Wnt target genes via recruiting Gro, CtBP, and HDACs. Residual β-catenin is exported from the nucleus by RanBP3 and APC or bound by CBY or ICAT that prevents β-catenin association with TCF/LEF. (Left) Wnt/β-catenin signaling ON. The Wnt ligand binds to Fz and LRP5/6 receptors to form a Fz-LRP5/6 complex. Dally and Kny can also bind Wnt and enrich Wnt concentration locally or help Wnt gradient distribution. Rspo proteins and Norrin are secreted agonists that bind to LRP5/6 and/or Fz to activate Wnt/β-catenin signaling. Formation of the Fz-LRP6 complex via Dvl promotes LRP6 phosphorylation by GSK3 and CK1γ and other CK1. Fz binds Dvl and phosphorylated LRP6 recruits Axin to the plasma membrane, resulting in inhibition of β-catenin phosphorylation/degradation by an as yet unknown mechanism. Fz-LRP6-Dvl aggregation may be involved. LRP6 association with caveolin may promote its endocytosis and signaling. Translocation of Axin is facilitated by MACF1. Trimeric G proteins may act between Fz and Dvl, and β-arrestin may associate with Dvl and Axin. Stabilized β-catenin is translocated to the nucleus where it binds TCF/LEF and recruits coactivators such as the Lgs/Pygo complex, CBP/p300, Brahma, MED12/Mediator, and the PAF1/Hyrax complex to initiate RNA transcription and elongation. TCF/β-catenin controls the expression of many genes that affect cell proliferation (i.e., c-myc, cyclin D1) and differentiation and also the expression of Wnt signaling inhibitory proteins (i.e., Dkk1, Axin2) that act as a negative feedback loop. In addition to TCF/LEF, β-catenin also binds to other transcription factors such as Prop1 and Pitx2 to coactivate non-TCF/LEF target genes. Other regulatory molecules (1) Stimulatory roles: Dvl phosphorylation by CK1ε and Par1 isoforms; GSK3 bindng by GBP/FRAT; Axin dephosphorylation by PP1, which prevents Axin-GSK3 binding. (2) Inhibitory roles: Dvl binding by NKD and IDAX; Dvl degradation by Inv and by the KLHL12-CUL3 complex. The primary cilia may suppress Wnt/β-catenin signaling via Inv-mediated Dvl degradation and other mechanisms. PP2A may have both stimulatory and inhibitory roles at different steps of the Wnt/β-catenin pathway depending on the regulatory subunits (PR55, PR61, PR72, and PR130). Frodo/DPR binds to Dsh/Dvl and may have stimulatory and inhibitory roles. Other signaling pathways that affect β-catenin signaling PKA can phosphorylate β-catenin at S675 and inhibit β-catenin degradation; Akt/PKB can phosphorylate β-catenin at S552 and promote β-catenin nuclear localization. At the adherens junction, β-catenin binds to cadherin to coordinate cell adhesion with actin cytoskeleton through α-catenin. Plakoglobin/γ-catenin (not shown) is closely related to β-catenin although its primary function appears to be at the adherens junction. RTK signaling phosphorylates cadherin-bound β-catenin at Y142 and Y654, via RTK and/or c-Src/Fyn/Fer, to promote dissociation of β-catenin from cadherin, resulting in an increase in cytosolic β-catenin and TCF/β-catenin signaling. RTKs (such as the PDGF receptor) can directly regulate the Axin complex via c-Abl-mediated phosphorylation of p68 (an RNA helicase), which displaces β-catenin from Axin and promotes β-catenin nuclear accumulation/signaling. TCF may be phosphorylated by NLK, which is activated by TAK1, to prevent TCF/β-catenin from binding to DNA.
- The epidermal growth factor receptor (EGFR/ErbB) pathway plays pivotal roles in cell-cell communication in both vertebrate and invertebrates. In Drosophila and C. elegans the EGFR pathway participates in the determination of numerous cell fates, including the development of the compound eye and the vulva. The four EGFR orthologs in vertebrates form a layered signaling network that participates in specification of cell fate and coordinates cell proliferation. Mutations in components of the pathway are commonly involved in human cancer. Ligand processing: Different mechanisms are employed in vertebrates and invertebrates for processing the ligand in the signal-producing cell to generate the secreted, active form. In Drosophila, the ligand precursor for Spi, Grk, or Krn is retained in the ER. It associates with the chaperone Star and is trafficked to a late compartment where the intramembrane protease Rhomboid resides, cleaving the ligand to generate the secreted form. Rhomboid also cleaves the chaperone Star, thus attenuating the level of ligand that is trafficked. In vertebrates, the ligand precursors are trafficked to the plasma membrane, where they are cleaved by ADAM metalloproteases. (2) Receptor maturation: The nascent forms of EGFR and its sibling, HER2, associate in the ER with a complex comprising the HSP90 chaperone and the kinase-dedicated adaptor, CDC37. Upon glycosylation and delivery to the plasma membrane, only the mature form of HER2, as well as some naturally occurring EGFR mutants, remain associated with the chaperone. In polarized tissues the PDZ domain proteins LIN-2, -7, and -10 associate with and stabilize the receptor in the basolateral surface. (3) Receptor dimerization: In the absence of a ligand, EGFR exists in a conformation that suppresses kinase activity and restrains formation of receptor dimers. Ligand binding initiates a conformational alteration that unmasks a “dimerization loop,” triggering receptor dimerization. These transitions are relayed across the plasma membrane to activate the bilobular kinase domain: the mostly beta-strand N-terminal lobe of one receptor is juxtaposed next to the C-lobe of the dimer partner, thereby forming a catalytically active asymmetric dimer. Variations on this activation scheme are found in the ErbB family. ErbB-3 is kinase dead but is able to transactivate dimer partners, whereas HER2/ErbB-2 is a ligand-less oncogenic receptor “locked” in the active conformation. Drugs in current clinical use include two EGFR tyrosine kinase inhibitors, as well as a dual EGFR and HER2 inhibitor. Also approved for clinical applications are a humanized monoclonal anti-HER2 antibody and two anti- EGFR antibodies. (4) Downstream signaling: Receptor homo- or heterodimers undergo transphosphorylation on multiple tyrosine residues. This leads to the recruitment of a plethora of enzymes and adaptor proteins. For example, the SHC and GRB2 phosphotyrosine-binding adaptors link phosphorylated receptors, through a guanine nucleotide exchange protein (SOS) and a small GTP-binding protein (RAS), to a linear cascade culminating in ERK1 and ERK2, which translocate to the nucleus to stimulate various transcription factors. Nuclear translocation of active receptors, such as HER2 and ErbB-4, has also been described. (5) Switch off: Delayed activation of a variety of suppressive mechanisms attenuates ligand-stimulated signaling or switches cells back to the resting state. These processes include receptor ubiquitinylation and dephosphorylation, kinase inactivation, ligand depletion, removal of active receptors from the cell surface, or proteasomal degradation. In addition, an inducible set of transcriptional repressors and RNA-binding proteins ensure signal desensitization. (6) Endosomal sorting: Rapid clearance of active receptors and sorting for degradation in lysosomes involves receptor clustering over clathrin- or caveolin-coated regions and CBL-mediated conjugation of ubiquitin or NEDD8. Large ESCRT protein complexes sort ubiquitinylated receptors at the MVB. Independently, inflammatory cytokines and oxidative stress transregulate EGFR by phosphorylating and arresting the receptor in perinuclear vesicles. (7) Adhesion signaling: In some cell types, cell migration is regulated by EGF-induced activation of NCK and PLC-gamma and subsequent activation of RHO family GTPases necessary for formation of filopodia and lamellipodia.
- Ras is a monomeric membrane-associated GTP-binding protein that regulates cell proliferation and survival in response to extracellular stimuli, such as activation of epidermal growth factor receptor (EGFR) or T cell receptor (TCR). Originally identified as an oncogene in murine sarcoma viruses, activating mutations in Ras have been found in about 30% of human tumors. Dysregulation of the Ras signaling pathway plays a key role in the progression of cancer. When bound to GTP, Ras is active and stimulates several downstream targets by direct interactions. Ras has intrinsic GTPase activity, which can be activated by GTPase-activating proteins (GAPs) such as the NF1 tumor suppressor gene product and p120GAP. Activation of Ras occurs largely through guanine nucleotide exchange factors (GEFs) such as Sos and RasGRP1, which catalyze the exchange of Ras-bound GDP with free GTP. Multiple downstream effector pathways mediate Ras signaling, including the Raf/MEK/ERK kinase cascade, the PI 3-kinase/ Akt/mTor pathway, and the Ral GTPase pathway. Raf/MEK/ERK is the prototypical mitogen-activated protein kinase (MAPK) cascade, wherein each kinase phosphorylates and activates its downstream target, culminating in the activation of multiple targets including several transcription factors. Much of the recent work on the MAPK cascade has focused on negative feedback loops, such as the phosphorylation of Raf and Sos by ERK, as well as the transcription of negative regulators such as Sprouty and Spred. The second of the major downstream Ras targets, the PI 3-kinase/Akt/mTor pathway, contributes to cell growth, proliferation, and survival downstream of Ras. PI 3-kinase is a lipid kinase that phosphorylates phosphatidylinositol (4,5) bisphosphate (PIP2) to generate the second messenger phosphatidylinositol (3,4,5) trisphosphate (PIP3). PIP3 activates Akt (PKB) and also GEFs for Rac GTPases, which regulate the actin cytoskeleton. Finally, Ras is able to activate another GTPase, Ral, through stimulation of the RalGDS family of guanine nucleotide exchange factors. Activation of Ral results in increased endocytosis and activation of the transcription factors Jun and Fos. 1292.e1 Cell 133, June 27, 2008 ©2008 Elsevier Inc. DOI 10.1016/j.cell.2008.06.020
- p38 : A Stress-Activated Protein Kinase with Multiple Functions Mitogen-activated protein kinase (MAPK) pathways are important regulators of cellular responses to many extracellular stimuli. Typically, eukaryotic cells have several parallel MAPK pathways, which allow the integration of signals from different stimuli. One of these, the p38 MAPK pathway, has been conserved from yeast to mammals, in which there are four family members: p38a (MAPK14), p38b (MAPK11), p38d (MAPK13), and p38g (MAPK12). Most of what has been published on p38 MAPK signaling refers to p38a, which is ubiquitously expressed at high levels in most cell types. In contrast, p38b seems to be normally expressed at lower levels. The other two family members have more restricted tissue expression patterns. Activation of p38a is induced by most stress stimuli, including UV light, oxidative stress, and heat or osmotic shock, but also when cells are exposed to cytokines, chemokines, hormones, or growth factors. Taken together, it appears that p38a signaling helps cells to adequately respond to changing environmental conditions. The extracellular stimuli usually lead to the activation of MAPKs via a cascade of phosphorylation events that involves at least two other kinases acting sequentially. MAP2Ks directly phosphorylate the activation loop of p38a on Thr and Tyr residues, leading to a conformational change that results in kinase activation. The three MAP2Ks that are known to activate p38a are, in turn, activated by phosphorylation on two conserved residues catalyzed by ten MAP3Ks. Upstream in the pathway, there is more diversity, and the activation of different MAP3Ks involves mechanisms like phosphorylation, ubiquitination, or protein-protein interaction, which facilitate integration of a wide range of signals. There is also evidence for the activation of p38a in particular cases by a noncanonical mechanism based on autophosphorylation, independently of MAP2Ks. Once p38a becomes activated, it can phosphorylate many substrates on Ser or Thr residues. This schematic depicts upstream regulators leading to p38a activation and the myriad downstream targets of p38a. p38 Substrates as a Source of Functional Diversity A large number of publications have described the implications of the p38a-signaling pathway in multiple functions. However, comprehensive information about the p38a targets that are phosphorylated in response to different stimuli has not been compiled. We have found reports for 96 proteins that can be phosphorylated by p38a. A companion Snap- Shot that will be published in the February 14 issue will provide additional information about reported substrates, what residues are modified, and functional consequences. About 55% of the known p38a substrates are located in the nucleus. These are mainly DNA- or RNA-binding proteins that are involved in the regulation of gene expression. There is evidence that 31 transcription factors can be directly phosphorylated by p38a, which in most cases results in the activation of transcription. Recent work has also connected p38a with chromatin remodeling via phosphorylation of BAF60c and p18Hamlet, which are structural components of the SWI/SNF and SRCAP complexes, respectively. In addition, there are p38a substrates that can regulate mRNA processing (FBP2/3 and SPF45) or stability (HuR and KSRP). Another important group of p38a substrates comprises proteins that are involved in signal transduction. These include two membrane receptors with Tyr kinase activity, EGFR and FGFR, and ten Ser/Thr kinases, which in turn can phosphorylate additional proteins and diversify the signal. Thus, MSK1 and MSK2 can regulate gene expression by direct phosphorylation of the transcription factors CREB and ATF1 and the chromatin protein histone H3, whereas MNK1 and MNK2 can regulate protein synthesis by phosphorylation of the initiation factor eIF4E. One of the first reported p38a substrates, MAPKAPK-2 (MK2), as well as the closely related MK3 can regulate mRNA stability by phosphorylation of ARE-binding proteins such as TTP or HuR. MK2 and MK3 also play important roles in actin filament remodeling by phosphorylation of Hsp27. Interestingly, some proteins can be potentially phosphorylated by both p38a and one of its downstream kinases, such as the MK2 substrates Cdc25B and HuR, or the MSK1/2 substrate histone H3. This double targeting of downstream substrates might function as a fail-safe mechanism to limit inappropriate effector activation. A number of cytoplasmic proteins that are involved in different aspects of cell regulation can also be phosphorylated by p38a. This group includes proteins that mediate p38a antiproliferative functions, such as stress-induced cell-cycle arrest (p57Kip2 and cyclin D1/3), and apoptosis (Bax and BimEL). However, p38a has also been reported to regulate cell survival through the phosphorylation of caspase-3 and caspase-8. Other cytoplasmic substrates of p38a may regulate proliferation and differentiation or specific processes such as cytoskeleton organization and intracellular membrane trafficking. Protein turnover can also be regulated by p38a at different levels either by phosphorylation-induced changes in the stability of the substrates or by phosphorylation of E3 ubiquitin ligases such as Siah2. In addition, p38a may inhibit proteasome activity by phosphorylation of the proteasome subunit Rpn2. In summary, p38a plays key roles in the stress responses but is also implicated in multiple cellular functions not related to stress. Although many more p38a substrates likely remain to be discovered, the variety of known targets supports the notion that this signaling pathway connects many different stimuli to a broad spectrum of cell responses. 656.e1 Cell 152, January 31, 2013 ©2013 Elsevier Inc. DOI http://dx.doi.org/10.1016/j.cell.2013.01.029
- The PTEN (phosphatase and tensin homolog deleted on chromosome 10) gene encodes a plasma membrane lipid phosphatase that is recurrently lost in various human cancers. Heterozygous mutation of PTEN is causative for familial diseases including Cowden syndrome, which is characterized by multiple hamartomas, developmental defects, and increased cancer susceptibility. The major tumor-suppressive activity of PTEN is attributed to its ability to hydrolyze the 3′ phosphate of phosphoinositides at the plasma membrane and thereby negatively regulate phosphoinositide 3-kinase (PI3K) signaling, a promoter of cell growth and survival. In mouse models, Pten is haploinsufficient for tumor suppression. Progressive reduction of Pten dose is also associated with more aggressive cancers in the mouse, leading to the notion that subtle variations in Pten levels could have critical consequences for tumor progression. Indeed, in addition to the loss of PTEN function through genetic alterations at the 10q23 locus, emerging lines of evidence demonstrate that PTEN is tightly regulated by transcriptional, posttranscriptional, and posttranslational mechanisms. Recent studies report that PTEN has tumor-suppressive functions from within the nucleus, including the regulation of chromosomal stability, the DNA repair response, and the cell cycle. Aberrant cytoplasmic localization of PTEN is associated with cancer progression, suggesting that the presence of PTEN in the nucleus is required for its tumorsuppressive activity. How PTEN gets in and out of the nucleus is still not completely understood. Nuclear pore-mediated import as well as the monoubiquitination of PTEN can modulate its subcellular compartmentalization, and germline mutations of residues that are ubiquitinated have been associated with the nuclear exclusion of PTEN in cancerous lesions. 550.e1 Cell 133, May 2, 2008 ©2008 Elsevier Inc. DOI 10.1016/j.cell.2008.04.023
- How Do Tumors Recruit Blood Vessels? Blood vessels are indispensible for tumor growth and metastasis. Hence, tumors exploit multiple avenues to recruit blood vessels. Angiogenesis—the sprouting of new blood vessels from the existing vasculature—is the most widely investigated mode of new vessel formation in tumors. There are five other mechanisms of new vessel recruitment (top panels; adapted from Carmeliet and Jain, 2011). However, their relevance in cancer is still being debated, and their molecular mechanisms are not well understood. Vasculogenesis involves vessel formation by endothelial progenitor cells (EPCs), which are recruited from the bone marrow and/or are resident in vascular walls. Intussusception is the splitting of pre-existing vessels to give rise to daughter vessels. Vessel co-option occurs when cancer cells grow around and co-opt the existing vasculature. Vascular mimicry is a process in which cancer cells get incorporated into the blood vessel wall. Tumor stem cell to EC differentiation occurs when cancer stem-like cells differentiate into endothelial cells (ECs). For historical reasons and, now, for convenience, the term “angiogenesis” is used to describe all of these methods of blood vessel recruitment by tumors. 1408.e1 Cell 149, June 8, 2012 ©2012 Elsevier Inc. DOI 10.1016/j.cell.2012.05.025
- The anaerobic metabolism of glucose in tumor cells yields far less ATP (2 per glucose) than the complete oxidation to CO2 that takes place in healthy cells under aerobic conditions (,30 ATP per glucose), so a tumor cell must consume much more glucose to produce the same amount of ATP. Glucose transporters and most of the glycolytic enzymes are overproduced in tumors. Compounds that inhibit hexokinase, glucose 6-phosphate dehydrogenase, or transketolase block ATP production by glycolysis, thus depriving the cancer cell of energy and killing it. 1. HIF-1 induce VEGF and glycolytic enzymes 2. Cells with mutant p53 has defective mitochondrial electron transport chain and are forced to rely more on glycolysis.