CANCER GENOMICS

CANCER GENOMICS A scale of the cancer problem Chances to develop invasive cancer Cancer type Birth-39 40-59 Birth – death All cancers, 1/69 1/52 1/12 1/24 1/11 1/2 1/8 1/3 Male/Female Male/Female Male/Female Breast Uterine corpus 1/229 1/567 Uterine cervix 1/2097 Prostate <1/10000 Lymphoma 1/591 1/1311 Lung 1/3060 1/3099 Colorectal 1/1508 1/1719 Melanoma 1/769 1/508 1/288 1/138 1/48 1/208 1/89 1/115 1/199 1/317 1/116 1/145 1/261 1/48 1/13 1/17 1/58 1/117 1/37 1/6 1/57 1/17 1/18 1/82 AGE-ADJUSTED CANCER DEATH RATES (Males) Last 70 years statistics GASTRIC CANCER down 4 times REASON? – better nutrition? Probably… LUNG CANCER – UP!!! 10 times (due to smoking and air pollution) AGE-ADJUSTED CANCER DEATH RATES (Females) Last 70 years statistics UTERINE CANCER (including cervix) – down 5 times due to early diagnostics GASTRIC CANCER down 4 times … Same as in males…. LUNG CANCER – UP!!! 10 times (due to smoking and air pollution) Does science help patients? 5-year survival rates YES! Cancer type Leukemia Prostate Breast Years 74-76 35% 68% 75% Years 83-85 42% 76% 79% Year 92-97 46% 97% 87% NO! Cancer type Years 74-76 Years 83-85 Year 92-97 Pancreas Liver Uterine corpus 3% 4% 89% 3% 6% 85% 4% 6% 86% HOW SCIENCE CAN HELP CANCER PATIENTS? Few things contribute Early diagnosis Specific treatment Early check for relapse NOT possible without understanding of the state of genes in every given tumor Tumors are very polymorphic entities There is no such a thing like tumor in general; they all differ in their way to survive and kill Multiple systems of tumor classification exist ORGAN – based (trivial) Breast cancer Brain cancer Liver cancer TISSUE - based (histological) WISDOM – based (clinicians’ experience) Carcinoma Sarcoma Lymphoma 1 2 Grading/staging (metastase/invasion) Pace of progression (slow of fast) (best working) Colon cancer Mucinous cystadenocarcinoma Carcinomas with spindle and/or giant cells ORGAN – based classification Breast cancer as an example Tumors can be derived from ductal cells or lobular cells ? Ductal carcinoma in situ (DCIS) malignant High chances to become Lobular carcinoma in situ (LCIS) benign Low chances to become Invasive carcinomas (ductal or lobular or even mixed) Very malignant inflammatory breast cancer, medullary carcinoma, mucinous carcinoma, Paget's disease of the nipple, Phyllodes tumor, and tubular carcinoma. TISSUE-based classification Carcinoma malignant neoplasm of epithelial origin, including internal epithelium (bladder lining, pancreatic duct lining etc…. ) Adenocarcinoma From tissue/cells producing secret – milk or mucous or enzymes Squamous cell carcinoma (SCC) From pure lining cells or from skin epithelium Other small variants, like basal cell carcinoma etc. Sarcoma Sarcoma refers to cancer that originates in supportive and connective tissues such as bones, tendons, cartilage, muscle, and fat. •Osteosarcoma or osteogenic sarcoma (bone) •Chondrosarcoma (cartilage) •Leiomyosarcoma (smooth muscle) •Rhabdomyosarcoma (skeletal muscle) •Mesothelial sarcoma or mesothelioma (membranous lining of body cavities) •Fibrosarcoma (fibrous tissue) •Angiosarcoma or hemangioendothelioma (blood vessels) •Liposarcoma (adipose tissue) •Glioma or astrocytoma (neurogenic connective tissue found in the brain) •Myxosarcoma (primitive embryonic connective tissue) Leukemia cancers of the bone marrow (liquid phase) overproduction of immature lymphocytes, which do not perform as well as they should, therefore the patient is often prone to infection Myelocytic/myelogenous lymphocytic erythroblastic Mixed (polycytemia vera) Lymphoma tumors from the lymphocytes from lymph nodes (solid tumors) Can be present in extranodal (non-lymph nodes) sites – stomach lymphoma, skin lymphoma. Lymphoma goes through the same stages of generalization (metastase) as any solid tumor Hodgkin disease Non-Hodgkin lymphoma Teratoma • Mixed type of tumor, often it is derived from embryonic or stem cells The most common elements are components of stratified squamous epithelium medstat.med.utah.edu/ Common types of cancer progression Adenoma Adenocarcinoma Papilloma Fibroma Squamous cell carcimoma Fibrosarcoma Lipoma Glioma Liposarcoma Anaplastic glioma Glioblastoma multiforme Chronic leukemia Remission with latent residual disease Acute leukemic crisis (blast crisis) WISDOM – based classification of tumors BENIGN MALIGNANT Borders No invasion No clear borders Invasion Normal blood vessels many leaky blood vessels newscenter.cancer.gov/sciencebehind/ cancer/cancer34.htm Histological picture of tumor malignisation newscenter.cancer.gov/sciencebehind/ cancer/cancer34.htm TUMOR GRADES are based on degree of cell differentiation tumor cells obtained from biopsy are evaluated by histopathologist Grade I (Cells are slightly abnormal and well differentiated) Grade II (Cells are more abnormal and moderately differentiated) Grade III (Cells are very abnormal and poorly differentiated) Grade IV (Cells are immature and undifferentiated) Tumor histology • Metaplasia – Replacement of normal differentiated tissue by another (differentiated) type like epithelial to mesenchime transition in breast carcinoma progression • Anaplasia – Loss of differentiated phenotype • Dysplasia – Loss of tissue organisation • Hyperplasia – Increase in cell division TUMOR STAGING Staging is the classification of the extent of the disease in the body. Most common systems are TNM system and Numerical system The tumor, nodes, metastases (TNM) system classifies cancer : -- by Tumor size, T N M -- the degree of regional spread or Node involvement -- and presence/absence of distant Metastasis. Tumor (T) T0 Tis (No evidence of tumor) (Carcinoma in situ limited to surface cells) T1–4 (Increasing tumor size and involvement) Node (N) N0 No lymph node involvement N1–4 Increasing degrees of lymph node involvement Nx Lymph node involvement cannot be assessed Metastases (M) M0 No evidence of distant metastases M1 Evidence of distant metastases Pancreatic carc. met. in liver www.ikp.unibe.ch/lab2/cytostatics/ sld014.htm Numerical system Stage 0 Stage 1 Stage 2 Stage 3 Stage 4 Cancer in situ (limited to surface cells) Cancer limited to the tissue of origin with evidence of tumor growth Limited local spread of cancerous cells Extensive local and regional spread Distant metastasis Grading / Staging and prognosis High-grade tumors progress faster Late-stage tumors prognosis is worse newscenter.cancer.gov/sciencebehind/ cancer/cancer34.htm CANCER TREATMENT Radical IDEAL GOAL: Completely eradicate tumor Pain relief ATTAINABLE GOAL: Remove primary tumor (surgically) or achieve tumor shrinkage (chemotherapy) Infection combat (especially important for leukemias and lymphomas) Chemo-related problems combat Palliative GOALS: RADICAL STRATEGIES of the TUMOR TREATMENT Radiotherapy Surgery Chemotherapy Hormonal therapy IMMUNOTHERAPY SURGICAL REMOVAL Primary care for most of the tumors Ablastic principle (all cuts are done within normal tissue; can be PCR controlled) On the advanced stages surgery always used in combination with chemotherapy MINUSES: -Not able to catch and remove micrometastases; -Not able to deal with ascytic (liquid) tumors CHEMOTHERAPY CHEMOTHERAPY is the treatment of cancer with drugs that can destroy cancer cells. Ideal anticancer drug should be able to kill tumor cell and be harmless for any normal cell Problem: No clear differences between normal and tumor cells FIRST PROBLEM WITH CHEMO = side effects Only the rate of cell division makes tumor cell more prone to poisonous effect of (Cytoreduction) chemotherapy Normal cells with fast pace of divisions are also very susceptible to chemo Side effects for BONE MARROW, COLON, HAIR FOLLICLES etc…. SIDE EFFECTS of Chemotherapy Bone marrow toxicity – infection and hemorrhage (bleeding) Gut mucosa toxicity – diarrhea Oral mucosa toxicity – mucositis (oral dryness) Hair follicle toxicity – hair loss Genomics can help! Better understanding of differences between normal and tumor cells can help to invent new drugs with increased tumor-specific action SECOND PROBLEM WITH CHEMO = tumor resistance After rounds of chemotherapy and successful shrinkage of tumor and/or remission tumor cells become resistant to treatment Side effects prevail over benefits Clinician has to stop treatment and tumor start to grow again Genomics can help again! Understanding of genomic changes during development of resistance can help prevent such changes or help invent new drugs not producing common types of resistance Traditional chemotherapeutic drugs • I. Alkylating agents (e.g. cisplatin) • II. Antimetabolites (e.g. Folate, purine or pyrimidine analogs) • III. Plant derivates (vinca alcaloids, taxanes, etoposide) • IV. Anti-tumoral Antibiotics Tumor could become resistant to any of this compounds or their combinations I. Alkylating agents: Nitrogen mustards, Nitrosoureas, Platinum agents (Cisplatin), Cyclophosphamides The alkylating agents react with everything; impair cell function by forming covalent bonds with the amino, carboxyl, sulfhydryl, and phosphate groups in biologically important molecules Alkylating agents generally interact with DNA non-specifically: the more effective ones tend to crosslink DNA. The electron-rich nitrogen at the N7 position of guanine in DNA is particularly susceptible to alkylation. Monoalkylation Alkylating agents interact with DNA non-specifically Active even for the resting cells (not cell cycle/phase-specific) Crosslinking (Between strands) chlorambucil and melphalan Crosslinking (Same strand) Cis-platinum (damage cell now, kill it during next cell division) Therefore Alkylating Agents can be used for the treatment of slow-growing tumors; extensive damage of DNA will lead to cell death Nitrogen mustard – first chemotherapeutic substance (interstrand-linking agent) Nitrogen mustard drugs were developed from mustard gas, a war agent first used during the First World War at Ypres. Could penetrate all protective materials except urethan One of the toxic-effects of exposure is the destruction of the bone marrow and white blood cells. Mustard gas Cure for cancer of the lymph glands – Hodgkin's Disease. Nitrogen mustard Cisplatin (Same strand-linking agent) BETTER than INTERstrand-linking agent!! because of two reasons Cisplatin (Same strand-linking agent) 1. Same strand cross-links formed are “harder" to repair than cross-links between strands. 2. Cisplatin is able to replace zinc(II) ion in zinc-finger containing transcription factors, directly destroying transcription regulation Cisplatin Therapy Type of Cancer Complete cure Sensitive Responsive Resistant Testicular cancer Ovarian Bladder, Head & neck Cervix, Prostate, Esophegeal Activity shown Various, eg.: Non-small cell lung, Ostegenic sarcoma, Hodgkins lymphoma Synergy shown in combination with 5-fluorouracil, cytarabine and bleomycin, That allows for greater flexibility in the design of drug regimens. Tumor could develop resistance to alkylating agents Genomic changes in AA treated tumor cells may lead to: -- Increase in ability of treated cells to repair DNA defects; -- Decrease in cellular permeability to the Alkylating drugs; -- Increase in glutathione synthesis conjugation with glutathione leads to chemical inactivation of alkylating agents (catalyzed by glutathione S-transferase) II. Antimetabolite agents: 1. Antimetabolites are structural analogs of the naturally occurring metabolites involved in DNA and RNA synthesis. 2. Antimetabolites exert their cytotoxic activity either by competing with normal metabolites for the catalytic or regulatory site of a key enzyme or by direct substituting for a nucleotide that is normally incorporated into DNA and RNA. 3. Antimetabolites are most active in S phase cells and have little effect on cells in G0. Consequently, these drugs are most effective in rapidly dividing tumors Types of Antimetabolites: Folate analogs -- Methotrexate (MTX) (bind to catalytic site of dihydrofolate reductase DHFR) Purine analogs -- Mercaptopurine,Fludarabine (inhibits enzymes involved in purine metabolism) Pyrimidine analogs -- Flurouracil (5-FU), Ara-C (inhibits enzymes involved in pyrimidine metabolism) Methotrexate (MTX) www.vet.purdue.edu/ Looks very same to folic acid – necessary nutrient. Methotrexate DHFR like MTX more than folic acid No production of No THF-co-enzymes tetrahydrofolic acid for one-carbon transfer No de novo purines and pyrimidines Broad range Treat leukemia, lymphomas, and osteosarcoma. It is also used in the treatment of AIDS and rheumatoid arthritis No DNA synthesis Methotrexate (MTX) resistance as an example of antimetabolite resistance Genomic changes in MTX-treated tumor cells may lead to: -- decrease in drug transport into the cell (less MTX) -- alteration in gene DHFR encoding dihydrofolate reductase enzyme with lower affinity for methotrexate (MTX-durable DHFR) -- quantitative increase in DHFR concentration by DHFR gene amplification, or by increased DHFR mRNA production (more DHFR) III. Plant derivates (heterogenous group) • Vinca alcaloids (Vinblastine, Vincristine) from periwinkle plant Vinca rosea (native to Madagaskar); • Taxanes (Paclitaxel = Taxol and docetaxel = Taxotere) from needles of Pacific Yew plant Taxus brevifolia; • Podophyllotoxin (etoposide) from Podophyllum peltatum (India) • Camptothecins (topotecan, irinotecan) from Camptotheca acuminata (China) Vinblastine, Vincristine -- Mechanism of action = mitosis block: bind to tubulin and induce microtubule depolymerization (Disassembly) Cells are arrested at metaphase as no chromosome segregation possible http://www.img.cas.cz/dbc/gallery.htm Red – anti-tubulin; Green – anti-vimentin; Blue -- DNA Mouse 3T3 fibroblast (normal) 3T3 after vincristine Vinca alcaloids bind to tubulin dimers at a specific recognition site on the protein. Tubulin form paracrystaline aggregates Concentration of the free dimers is reduced Mitotic spindles can not be formed Equilibrium of growth/shrinkage of tubules shifted to the shrinkage www.staff.ncl.ac.uk/i.r.hardcastle/ antibiotics.html www.chem.wisc.edu/~bestchem/ taxol.gif Paclitaxel (Taxol) and docetaxel (Taxotere) -- Mechanism of action = mitosis block: promote microtubule assembly and super stability, action is opposite to vinca alcaloids, but effect is the same Bundling of accumulated, disorganised microtubule filaments. + Taxol Used for treatment of otherwise resistant ovarian cancers and recurrent breast tumors Podophyllotoxin (etoposide) = Topo II inhibitor Reconnection of DNA Etoposide stabilizes TOPO II – DNA complex in “non-sealable” form Both ends of DNA can freely rotate within the enzyme Etoposide inhibit topoisomerase II that catalyses transient breaking and re-joining of ds DNA strands (DNA becomes damaged Camptothecins (topotecan, irinotecan) = Topo I inhibitor topoisomerase I produces reversible single-strand breaks in DNA These single-strand breaks relieve torsional strains, and allow DNA replication to proceed. Topotecan binds to the topoisomerase I-DNA complex and prevents ligation of the DNA strand, resulting in DNA breakage during the elongation and cell death TOPO I is not required for the viability of cells (TOPO II can substitute). Therefore, TOPO I – negative cancer cells are viable and resistant to topotecan IV. Antitumoral antibiotics (produced by fungi) Anthracyclines: Doxorubicin (Adriamycin) and Daunorubicin Produced by fungus Streptomyces percetus var caesius. Doxorubicin Bleomycin Produced by Streptomyces verticillus as a mixture of small-molecular-weight copper-chelating glycopeptides Bleomycin Anthracyclines: Doxorubicin (Adriamycin) and Daunorubicin Mechanisms of action: 1) intercalation between DNA base pairs and inhibition of DNA topoisomerases I and II. 2) Altering membrane fluidity and ion transport Active in Hodgkin's disease, non-Hodgkin's lymphomas, sarcomas, acute leukemia, and breast, lung, and ovarian carcinomas Bleomycin Mechanisms of action - multiple: 1) binds to double- and single-stranded DNA produces site-specific and non-specific SS and DS breaks ratio single:double = 10:1; Cleave DNA at G-C and G-T sequences; DNA in open chromatin esp. sensitive (where genes are expressed) 2) Makes non-covalent interstrand links 3) Inhibits DNA reparation by suppression of DNA ligase Rapidly degraded by bleomycin hydrolase present in most tissues except skin and lung No bone marrow and intestinal toxicity!! (…Pulmonary fibrosis and skin effects are strong...) Multidrug resistance phenomenon Is a membrane associated phenomenon that represents a serious obstacle to chemotherapy of cancer Cell that are treated with (let’s say) etoposide and develop resistance to etoposide, simultaneously become resistant to Methotrexate MDR caused by induction of membrane proteins that effectively and (almost) non-specifically expel small molecules from the cell! P-glycoprotein (170 kDa) Belongs to traffic-ATPase superfamily of transport proteins Other transporters BCRP, ABC 1,2,3 … etc…. P-glycoprotein This is an ATP-dependent export pump (= efflux pump) of broad specificity This efflux pump expel hydrophobic drugs, natural products and peptides. The transport function of P-glycoprotein can be blocked by the action of another group of compounds known as MDR modulators, or chemosensitizers cyclosporin A, verapamil, tamoxifen RADICAL STRATEGIES of the TUMOR TREATMENT Radiotherapy Surgery Chemotherapy IMMUNOTHERAPY Hormonal therapy RADIOTHERAPY • General principle is the same – reduce a mass of rapidly dividing cells (cytoreduction principle, the same as for chemo) Side effects are the same as for chemo Difference – radiotherapy is regional. IR could be focused like a beam of light to a treated area (internal or external) HORMONAL THERAPY Good for hormone-dependent tumors Speaking generally, hormonal therapy can be discussed as trophic factor removal therapy Estrogens are trophic factors for breast epithelium and uterine epithelium Removal of estrogen from ER-positive tumors leads to stop of their growth Tamoxifen (Nolvadex) Tamoxifen mimics the action of estrogene and binds to estrogen receptor (ER). The tamoxifen-ER complex dimerises Than transported from the cytosol into the cell's nucleus The dimeric tamoxifen-ER complex binds to DNA DNA-tamox-ER complex is unable to function in the same way as the DNA-estrogene-ER complex In reality mechanism is not known completely !!!! In other tissues of the body Tamoxifen plays exactly like estrogene itself !!! IMMUNOTHERAPY Any intervention to enhance the body's natural ability to defend itself against malignant tumors. Cytokines: IFN-alpha, IL-2, Tumor necrosis factors (TNFs) Monoclonal antibodies: anti-CD20 Rituximab for lymphomas, anti-HER2 trastuzumab (herceptin) for breast cancer Cancer Vaccines: cancer-specific antigens + other components boosting immunity (e.g. dendritic cell vaccines) RADICAL TUMOR TREATMENT STRATEGIES Radiotherapy Surgery Chemotherapy Hormonal therapy IMMUNOTHERAPY MODERN science-oriented therapies Modern science for cancer treatment improvement SURGERY – PCR assisted surgery CHEMOTHERAPY – new target-specific drugs, tumor-specific delivery RADIOTHERAPY – normal (p53 positive) cell can be protected by p53 inhibitor pifithrin HORMONAL THERAPY – designed chemical inhibitors of hormone receptors IMMUNOTHERAPY – entirely modern area