Lecture 3 Pharmacology of Anti-Cancer Drugs Wong

THE MAMMALIAN CELL CYCLE:

1. G1 (GAP one) phase

• Manufacturing of protein

• Surveillance of DNA integrity

2. S (Synthesis) phase

• Synthetic phase of genetic material

3. G2 (GAP two) phase

• Late manufacturing of material for cell division

• Surveillance of integrity of newly replicated DNA

4. M (Mitosis) phase

• Actual cell division occurs, producing two identical daughter cells

• Sub-divided into 4 phases: prophase, metaphase, anaphase and telophase

• The 2 daughter cells separate their cytoplasmic connection at cytokinesis

5. Go (GAP zero) phase

• Resting phase, cells are not replicating

• Reversible, not part of the cell cycle

WHAT CONTROLS THE CYCLE?

• Progression of the cell cycle is controlled by proteins known as

cyclins, and their functions are regulated by cyclin-dependent kinase

o Oncogenes = promote cell cycle progression

o Tumor suppression genes = arrest cell cycle progression

DNA REPLICATION AND CELL DIVISION:

• Why target DNA replication?

o DNA replication is essential step before cell division can take place

o DNA replication only happens before cell division, representing a

unique marker for proliferating cells

• Steps in DNA replication and cell division:

o Synthesis of nucleotides

o DNA replication & topology (↑ sensitivity to DNA damaging agents)

o Equal segregation of chromosomes into 2 nuclei

o The division of a single cell into two daughter cells

NUCLEOSIDE SYNTHESIS INHIBITION:

DEOXYNUCLEOTIDES: BUILDING BLOCKS OF DNA

• DNA are polymers made up of 4 deoxynucleotide building blocks

• Each deoxynucleotide contains a sugar backbone (deoxyribose)

and a different base (total of 4)

o 2 purines: A (adenine), G (guanine)

o 2 pyrimidines: C (cytosine), T (thymine)

• A always pairs with T, C pairs with G

PYRMIDINE SYNTHESIS:

NUCLEOSIDE SYNTHESIS INHIBITION:

• Inhibition of Folate Acid Pathway: methotrexate

• Inhibition of thymine synthesis: 5-fluorouracil

FOLATE ANALOGS:

RALTITREXED:

MOA • Quinazoline (fused benzene & pyramidine rings) = analog of folic acid; potency increases by cellular poly-glutamation

• Specifically inhibits thymidylate synthase in the presence of Dump

• Competitively inhibits binding of MTHF to TS

• Stops purine and thymidine synthesis

• Cells are arrested at S phase of cell cycle

Route • IV infusion

Toxicities • Myelosuppression

• GI disturbances (moderate-low emetogenic)

Resistance Mechanisms

• Impaired cellular uptake

• Increased cellular levels of TS

• Reduced polyglutamation

METHOTREXATE:

MOA • Folate analog inhibits enzyme DHFR

• Stop purines and thymidine synthesis

• Cells are arrested at S phase of cell cycle

• Cancer cells more susceptible than normal cells for the apoptotic-inducing effects of MTX

Route • Administered PO, IV and IT

Toxicities • Myelosuppression; GI disturbances

• High emetogenic potential = dose-dependent

Folinic Acid Rescue

• Folinic acid (Leucovorin) rescue allowed lethal dose of MTX be used in aggressive chemotherapy regimens

• Administer exogenous tetrahydrofolate (folinic acid) 24-36h after MTX, as damage with MTX already done to the rapidly dividing cancer cells, but can be reversed in the slower-dividing normal cells

Resistance Mechanisms

• Impaired cellular uptake

• DHFR mutations or gene amplification

• Increased cellular export by drug efflux pump MRP

OTHER FOLATE PATHWAY INHIBITORS: pemetrexed

Lecture 3 Pharmacology of Anti-Cancer Drugs Wong

NUCLEOSIDE SYNTHESIS INHIBITION:

NUCLEIC ACIDS ANALOGS:

PYRMIDINE ANALOGS:

5-fluorouracil Capecitabine

MOA • Halogenated pyrmidine analog

• Prodrug, metabolically converted to FdUMP (5-fluoro-2’-deoxyuridylate)

• FdUMP, together with cofactor MTHF (methylene-tetrahydrofolate), formed a stable complex with thymidylate synthase

• Abolish dTTP production and DNA synthesis

Synergy • Folinic acid (Leucovorin) co-administration increases the concentration of MTHF for the 5FU-MTHF-TS ternary complex formation

Combo therapy

• Co-administered with leucovorin to increase the levels of MTHF

• Co-administered with methotrexate in breast cancer

• Co-administered with oxaliplatin or irinotecan in metastatic colorectal cancer

• Potent radiation sensitizer

Activated • Activated in target (tumor) cells

• Administered IP or IV (various forms)

• Orally available pro-drug of 5-FU

• Metabolized to 5’-deoxy-5-fluorocytidine (5’-DFCR) in liver and activated in target cells

Route • Administered IP or IV (various forms) • Orally administered, recommended to be taken with food

Toxicities • Myelosuppression

• GI disturbances

• Cardio-toxicities

• Palmar-plantar erythrodysesthesia

• Similar to 5-FU

• Hepatic disturbances

PG • Genetic deficiency of dihydro-pyrimidine dehydrogenase (DPD) are associated with severe drug toxicities

Resistance Mechanisms

• Changes in TS levels

• Decrease levels of MTHF Increase expression of anti-apoptosis proteins

CYTIDINE ANALOGS:

5-fluorouracil Capecitabine

MOA • Cytidine analogs converted into AradCTP (AraC) and dFdCTP (gemcitabine) in vivo

• Competes with dCTP for DNA polymerase

• Incorporation of activated drugs into the growing DNA molecule will lead to chain termination & ecll death

Activated • Activated intracellular in target cells • Activated in target cells

Route • IV, SC and IT administered • IV infusion

Toxicities • Cerebellar toxicity and myelosuppression

• Ocular toxicity

• High moderate emetogenic potential is dose-related

• Myelosuppression, hematouria

• Hepatic toxicities

• Pulmonary toxicities

Resistance Mechanisms

• Increase metabolism

• Decrease drug import

• Increase inactivation by metabolic enzymes

• Decrease drug import

AGENTS THAT WORK THROUGH SIMILAR MECHANISMS:

FLUDARABINE:

• Adenine analog

• Chain termination

6-THIOGUANINE AND 6-MERCAPTOPURINE

• Guanine analogs

• Elicit MMR activation and apoptosis

TOPOISOMERASE INHIBITORS:

DNA REPLICATION AND TOPOLOGY:

• DNA chain termination: cytosine arabinoside

• Topoisomerase I inhibitor: irinotecan

• Topoisomerase II inhibitor: etoposide

TOPOISOMERASES:

• Topoisomerases removed excess DNA supercoiling by breaking rotating and

religating DNA strands

• At least 5 members of topoisomerases in human cells:

o Topoisomerase I is found in transcription and replication complexes

o Topoisomerase II (2 subtypes, α and β) enzymes seem to function

after replication during separation of daughter DNA strands

o Topoisomerase III (2 subtypes, α and β) function in humans is

currently unknown

• Topo I and II inhibitors are potent anti-neoplastic agents)

TOPOISOMERASE INHIBITORS:

Topo I inhibitors Top II inhibitors

• Act in both cell cycle specific and non-specific manner

• Induce both single and double stranded DNA breaks

• Camptothecins and derivatives (irinotecan, topotecan)

• Cell cycle specific

• Induce double stranded DNA breaks

• Etoposides and other podophyllotoxin derivatives (teniposide)

Lecture 3 Pharmacology of Anti-Cancer Drugs Wong

TOPOISOMERASE INHIBITORS:

TOPOISOMERASE I INHIBITOR:

Camptothecin Irinotecan

MOA • Semisynthetic molecules o Chinese tree Camptotheca acuminate

• Specifically binds to and stabilizes Topoisomerase I-nicked DNA complex, creating single stranded DNA breaks During DNA replication, polymerase enzyme binds to the camptothecan-DNA-topo I complex, resulting in double strand breaks

Administration • Prodrug that is activated by carboxyesterase to SN-38

• Administered by IV infusion

Toxicities • Unpredictable toxicities limited clinical use • Less toxic and more soluble than camptothecins

• Cholinergic syndrome

• High emetogenic potential

• May cause severe GI toxicity that can be life-threatening susceptibility predicted by UGT enzyme polymorphism GROUPS PREVALENCE RISK OF TOXICITY

All patients 10%

Homozygous mutant 10% 50%

Heterozygous 40% 12.5%

Homozygous WT 50% 0%

TOPOISOMERASE II INHIBITOR: ETOPOSIDE

MOA • Semi-synthetic derivatives of a natural compound (isolated from Podophyllum peltatum – Mayapple tree)

• Inhibit the re-ligation of double strand DNA breaks generated by Topo II

• Arrest cells in the S and G2 phase of the cell cycle

Route • Available orally or by IV (preferred)

Major toxicities • Myelosuppression

• Congestive heart failure; myocardial infarction

• Type I hypersensitivity reactions

Resistance Mechanisms • Over-expression of drug efflux pump (p-glycoprotein)

Use • Secondary leukemia in Acute Lymphoblastic Leukemia

CHROMOSOME SEGREGATION:

THE FORMATION OF MICROTUBULE:

• Microtubules are dynamic structures that undergo continual assembly and disassembly within the cell (dynamic instability)

• Microtubules function to determine cell shape and cell movements, involving in processes such as:

o Cell locomotion

o Intracellular transport of organelles

o Separation of chromosomes during mitosis

VINCA ALKALOIDS: agents that destabilize the microtubule formation

Vinblastine Vincristine

MOA • Natural products isolated from the Madagascar peri-winkle plant

• Binds to specific sites on β-tubulin tubulin dimers are unable to polymerize into microtubules

o Shift the equilibrium of towards disassembly o Block mitosis by inducing metaphase arrest

Route • IV infusion

• Lethal if injected via IT route

Toxicities • Myelosuppression

• Peripheral neuropathy

• Bronchospasm

• Peripheral neuropathy

• Neurotoxicities

• Myelosuppression less severe than vinblastine

Resistance mechanisms

• Increase expression of p-glycoprotein

• Mutations in beta-tubulin binding sites

TAXANES: agents that stabilize microtubule formation

Paclitaxel Docetaxel

Derived • First isolated compound

• Western Yew tree

• Semi-synthetic & more potent analog of paclitaxel

MOA • Binds to β-tubulin disrupt equilibrium between free tubulin & microtubule formation by stabilizing the formation of polymers

o Also induce formation of abnormal bundles of microtubules

• Arrest cells in mitotic phase of the cell cycle, thereby inducing apoptosis in both p53 dependent and independent manners

Route • IV infusion and IP • IV infusion

Toxicities • Myelosuppression

• Peripheral neuropathy

• Hypersensitivity

• Cardio-toxicities

• Myelosuppression

• Pulmonary toxicities

• Hypersensitivity

Resistance mechanisms

• Increase expression of p-glycoprotein

• Mutations in beta-tubulin binding sites

• Increase expression of anti-apoptotic proteins

Lecture 3 Pharmacology of Anti-Cancer Drugs Wong

SUMMARY: MAMMALIAN CELL CYCLE AND CANCER AGENTS TARGETING SPECIFIC PHASES OF THE CELL CYCLE

• The mammalian cell cycle can be divided into 4 major phases and each phase is controlled by specific signal transduction events

• Cell-cycle specific chemotherapeutic agents target various steps in the replication and segregation of DNA in cell division

• Anti-metabolites halt the growth of cancer cells by inhibiting DNA synthesis

• Topoisomerase inhibitors created single-and double-stranded DNA breaks at specific phases of the cell cycle

• Anti-mitotic agents such as the vinca alkaloids and taxanes bind to β-tubulin and interfere with spindle assembly

• Cells that are arrested at various cell cycle checkpoints will need to be repaired or to be eliminated via apoptotic pathways

• Increase repair capacity and reduction in apoptosis are common PD resistance mechanisms