Lesson 15

Question 1

Speak about cancer and chemotherapy

Answer 1

INTRODUCTION TO CHEMOTHERAPY

Chemotherapy is commonly associated with cancer treatment; however, this term also encompasses antibacterial and antibiotic chemotherapy. In this context, we will focus on traditional cancer chemotherapy and not cover contemporary methods.

Neoplasia, a term preferred over cancer, includes both solid tumors and hematological malignancies such as leukemia. Neoplasia is characterized by the unwarranted, uncontrolled, and non-purposeful proliferation of malignant cells throughout the body. These are autologous cells that have lost the ability to regulate growth and migration.

Malignant cells exhibit numerous anomalies at genomic, chromosomal, epigenetic, biochemical, and structural levels. Their growth is deregulated, and they typically exhibit a lack of contact inhibition, though not universally. Malignant cells possess the capability to invade adjacent tissues—referred to as regional spread. For example, breast cancer in women commonly metastasizes to the thoracic region and lymph nodes. Systemic spread occurs when cells are transported via blood or lymphatic systems, leading to distant metastases, such as hepatic metastases originating from colon cancer, which despite its location, is still identified as colon cancer.

Cancer ranks as the second leading cause of death from disease, following cardiovascular diseases. Of the total population diagnosed with cancer, less than 25% can undergo surgical or radiotherapeutic intervention. For the majority, the aim is to improve quality of life and prolong survival rather than prevent cancer mortality. Notably, only about 10% of patients achieve a cure, with recent advances showing promising results in lymphoma treatments. Treatment efficacy varies significantly depending on the cancer type; some are amenable to chemotherapy, while others are not.

CHEMOTHERAPY MECHANISMS

Chemotherapy induces a cytotoxic event primarily targeting DNA, but can also exert metabolic toxicity. The ultimate goal is to achieve long-term survival or a cure.

Treatment and cure represent distinct concepts: treatment aims to extend life, whereas a cure implies complete eradication of the disease. Certain leukemias and small solid tumors are typically curable.

Rapidly dividing cells, such as those in skin, bone marrow, and gastrointestinal mucosa, are more susceptible to traditional chemotherapy’s effects, which can result in side effects like hair loss and mucosal inflammation. While some lymphomas are aggressive and more amenable to cure, indolent lymphomas are less responsive to treatment due to their slow cell replication rate.

Chemotherapy is often administered preoperatively to shrink tumors and facilitate surgical removal, particularly in solid tumors. Neoadjuvant chemotherapy refers to treatment before surgery, while adjuvant chemotherapy is administered postoperatively to target residual cancer cells. Maintenance chemotherapy can be employed during remission to prevent recurrence.

Question 2

Speak about Antimetabolites

Answer 2

CHEMOTHERAPEUTIC AGENTS: ANTIMETABOLITES

Various agents act at different stages of cellular replication and metabolism. Among the most significant are:

• Antimetabolites: These agents disrupt DNA synthesis at the nucleotide level by interfering with essential enzymes. For instance, fluorouracil leads to faulty DNA production by blocking thymidine synthesis, affecting cancers like those of the colon, stomach, and intestines.
• Intercalating agensts, Anthracyclines: Doxorubicin, a drug in this class, intercalates into DNA, obstructing topoisomerase II and generating reactive oxygen species (ROS) that damage nucleic acids. It is effective against various cancers but has cardiotoxic side effects.
• Adduct Forming Agents: Cisplatin creates cross-links between DNA bases, particularly guanines, interfering with DNA replication and transcription. This mechanism is similar to anthracyclines but involves covalent bonding.
• Alkylating Agents: Cyclophosphamide, a nitrogen mustard, requires bioactivation in the liver and forms covalent bonds with DNA, disrupting cellular replication. It is used to treat breast cancer and lymphomas.
• Mitotic Spindle Poisons: Substances like vinca alkaloids and taxanes interfere with microtubule function. Vinca alkaloids, derived from the periwinkle plant, inhibit microtubule assembly, leading to mitotic arrest. Taxanes stabilize microtubules, preventing chromosome separation during cell division.

Antimetabolites are a class of drugs that impede DNA synthesis by interfering with the enzymes necessary for nucleotide synthesis. These drugs mimic the structure of normal metabolites, thereby inhibiting key enzymatic reactions within the DNA replication process.

Antimetabolites exert their effects by acting as false substrates or inhibitors for enzymes involved in the synthesis of nucleotides, the building blocks of DNA. For instance 5-Fluorouracil targets the enzyme thymidylate synthase. During normal DNA synthesis, deoxyuridine monophosphate (dUMP) is methylated to form deoxythymidine monophosphate (dTMP). 5-Fluorouracil is structurally similar to uracil and competes with dUMP. When incorporated into the DNA synthesis pathway, it interferes with the methylation process, preventing the conversion of dUMP to dTMP, and ultimately inhibiting DNA synthesis.

Question 3

Speak about intercalating agents

Answer 3

INTERCALATING AGENTS: ANTHRACYCLINES

Anthracyclines represent a crucial class of anticancer drugs, initially discovered during the search for new antibiotics in Italy by an Italian scientist. Among them, doxorubicin, also known as “Adriamycin,” is a prominent chemotherapeutic agent utilized in the treatment of various cancers such as breast, ovarian, lung, stomach, and sarcomas.

Doxorubicin operates through multiple mechanisms:

1. DNA Intercalation: The drug intercalates between DNA base pairs, obstructing the enzymatic action of topoisomerase II, which is essential for DNA unwinding during replication and transcription processes.Doxorubicin’s structure allows it to form hydrogen and covalent bonds with DNA: It establishes two hydrogen bonds—one with a nitrogen base and one with an oxygen atom in the DNA. It intercalates between two guanine bases on opposite strands, forming a covalent bond.
2. Free Radical Formation: Doxorubicin facilitates the generation of reactive oxygen species (ROS), specifically hydroxyl radicals (OH*), which can induce double-strand breaks in DNA.The metabolism of doxorubicin involves the Fenton reaction, which generates highly reactive hydroxyl radicals: Doxorubicin is reduced, transferring an electron to a quinone group within its structure, leading to the formation of a semiquinone radical with the help of NADPH. This reaction involves iron, which cycles between Fe²⁺ and Fe³⁺ states, facilitating the conversion of hydrogen peroxide (H₂O₂) into hydroxyl radicals (OH*) and other ROS.
3. Inhibition of Nucleic Acid Synthesis: By intercalating into DNA, anthracyclines prevent the synthesis of DNA and RNA and can lead to strand scission.The hydroxyl radicals attack DNA by abstracting a hydrogen atom from the deoxyribose sugar, leading to the cleavage of the sugar-phosphate backbone and subsequent DNA strand breaks. This damage is not limited to nuclear DNA but also extends to mitochondrial DNA, illustrating the drug’s wide-ranging effects within the cell.

Initially, doxorubicin treatments were associated with significant cardiotoxicity, as the generated hydroxyl radicals could damage not only cancer cells but also the cells of the heart. This led to the development of new drugs aimed at mitigating the formation of OH* radicals by sequestering iron—these are known as chelating agents.

Question 4

Speak about Cisplatin

Answer 4

Cisplatin, a platinum-based chemotherapy drug, is an important treatment option for various types of cancer, including ovarian, cervical, breast, bladder, brain, and lung cancers. Its molecular structure is relatively simple, consisting of a central platinum (Pt) atom surrounded by two amine groups and two chloride ions.

Cisplatin exerts its anticancer effects by forming cross-links with purine bases in DNA, predominantly with guanine. This cross-linking interferes with the DNA’s normal functions, such as replication and transcription, ultimately leading to cell death.

Cisplatin becomes activated in the intracellular environment, where its chloride ions are displaced in a reaction facilitated by the nucleophilic character of water molecules due to the relatively acidic environment (presence of H+ ions). This displacement leads to the formation of a positively charged platinum complex that can readily form coordinate bonds with the nitrogen atoms of the purine bases in DNA.

A coordinate bond, also known as a dative covalent bond, occurs when one atom donates a pair of electrons to another atom; in the case of cisplatin, this involves the platinum atom and the nitrogen atoms of guanine (and sometimes adenine). The amine groups stabilize the complex by neutralizing the positive charges on the platinum.

Once bound to DNA, the platinum atom can form various types of adducts:

1. Intrastrand Cross-Links: These are the most common and involve the platinum atom binding to two guanine bases or to a guanine and an adenine base on the same DNA strand.
2. Interstrand Cross-Links: These are less common but are considered more cytotoxic because they link opposite strands of the DNA, effectively preventing the strands from separating, which is essential for replication and transcription.
3. Monoadducts: These involve the binding of the platinum atom to a single nitrogen base.

The binding of cisplatin to DNA creates a physical blockage for the DNA polymerase enzyme, which is unable to bypass the platinum-induced cross-links. This obstruction halts DNA replication and transcription, leading to apoptosis or programmed cell death.

The DNA cross-linking activity of cisplatin is the basis of its use in cancer chemotherapy. However, its effectiveness must be balanced against potential side effects, including nephrotoxicity, neurotoxicity, and ototoxicity. The development of resistance to cisplatin is also a significant concern, often necessitating combination therapy or the use of alternative platinum-based drugs.

Question 5

Speak about alkalinating agents

Answer 5

Alkylating agents represent a class of chemotherapeutic drugs that function by transferring alkyl groups, which are hydrocarbon chains, to biologically active molecules such as DNA. This results in the formation of DNA adducts, which can interfere with DNA replication and lead to cell death, especially in rapidly dividing cells such as cancer cells.

The CHOP regimen is a chemotherapy protocol that incorporates multiple drugs to treat lymphoma, with cyclophosphamide being a critical component.

Cyclophosphamide is a prominent member of the nitrogen mustard group of alkylating agents. It is a prodrug that requires metabolic activation, predominantly by the liver, to exert its antitumor effects. This compound has shown efficacy against a broad spectrum of cancers, particularly breast cancer and lymphomas, and can be administered either orally or intravenously. In its inactive form, cyclophosphamide does not exhibit cytotoxicity in vitro, such as in human lymphocyte cultures or neoplastic cell lines. However, once metabolized in the liver, primarily through the action of the cytochrome P450 enzyme CYP2B6, it is converted into its active form, 4-hydroxy-cyclophosphamide. This hydroxylation is a part of the phase I metabolic pathway. Cyclophosphamide contains a phosphate in place of a carbon in the amide group and two chlorine atoms, similar to cisplatin. These chlorine atoms are crucial for the drug’s activity. Following biotransformation, cyclophosphamide may yield an inactive metabolite excreted in urine or undergo non-enzymatic conversion to aldophosphamide. Aldophosphamide then spontaneously tautomerizes into two compounds: the toxic byproduct acrolein and the active form phosphamide mustard, with the latter being the actual cytotoxic agent.

4-Hydroxy-cyclophosphamide is in equilibrium with aldophosphamide, which can produce both the active mustard and acrolein. Acrolein is a potent pulmonary irritant and a corrosive agent for the bladder urothelium, capable of causing hemorrhagic cystitis.

To mitigate the risk of hemorrhagic cystitis, patients may be administered fluimucil—an acetylcysteine compound—which serves as a detoxifying agent. In phase II metabolism, glutathione transferase catalyzes the conjugation of glutathione to acrolein, leading to a less toxic, excretable metabolite. This conjugation is essential for blocking acrolein’s toxicity. Acrolein conjugated with cysteine is then eliminated via the biliary pathway. To ensure the liver’s detoxification system functions adequately, it is crucial to maintain a supply of fresh cysteine for glutathione synthesis, with glycine and glutamate being recycled components of the tripeptide.

Chlormethine, also known as mustine, is a simpler structure compared to cyclophosphamide and has historically been used as a chemical warfare agent. It shares a similar mechanism of action with cisplatin and cyclophosphamide, whereby it loses a chloride ion, and the remaining CH2 group becomes a positively charged carbocation. The nitrogen forms a coordinate covalent bond, resulting in a transient triangular structure that is highly unstable in the cell nucleus. This instability leads to the formation of cross-links between guanine bases, causing cytotoxicity in replicating cells.

Alkylating agents, including cyclophosphamide, are not cancer-specific and can therefore damage healthy cells such as those in the skin, blood, and mucosal lining of the gastrointestinal tract.

There are mnay different kinds of alkalinatying damages:

• Monoalkylation: The addition of an alkyl group to one DNA base.
• Interstrand Cross-Linking: Linking of two opposite DNA strands.
• Intrastrand Cross-Linking: Linking of bases on the same DNA strand, leading to distortion of the DNA helix and improper reading during replication and transcription.

These cross-links impede the progression of DNA polymerase and the DNA repair mechanisms, potentially leading to the amplification of damage. Alkylating agents are also mutagenic and have been associated with secondary cancers in patients treated with chemotherapy.

Question 6

Speak about mitotic spindle poisons

Answer 6

MITOTIC SPINDLE POISONS

Understanding the distinction between the centromere and kinetochore is crucial in the context of cell division and the action of certain chemotherapy agents:

• Centromere: The centromere is a constricted region of a chromosome where sister chromatids are most closely held together. It serves as an anchor point for the kinetochore and is essential for the proper segregation of chromosomes during cell division.
• Kinetochore: The kinetochore is a proteinaceous complex that assembles on the centromere. It is the attachment site for microtubule fibers of the mitotic spindle. The kinetochore plays a pivotal role in chromosome movement and segregation.

During metaphase, chromosomes align at the center of the spindle, and microtubules from opposite spindle poles attach to the kinetochores of sister chromatids. Not all microtubules attach to chromosomes; some serve other structural and mechanical functions. During anaphase, the kinetochore microtubules shorten as tubulin subunits are removed from their plus ends (at the kinetochores), pulling the chromatids toward the spindle poles.

Chemotherapeutic agents may target the microtubules, particularly those associated with the kinetochore. By destabilizing these structures, the drugs prevent the proper alignment and segregation of chromosomes, ultimately leading to cell death. Chemotherapeutic agents known as microtubule “poisons” are compounds that interfere with the dynamics of microtubules, which are critical for cell division. Two major classes of these agents have been derived from plant sources and have significantly impacted cancer treatment.

• Derived from the Vinca rosea plant, vinca alkaloids are a group of powerful chemotherapy drugs. Among this group, vinblastine (with a methyl group, R=CH3) and vincristine (with an aldehyde group, R=CHO) are structurally similar and widely used in the treatment of various cancers including lymphomas, lung cancer, bladder cancer, brain cancer, and others. Vinca alkaloids function by inhibiting the formation of microtubules. These compounds bind to alpha-beta tubulin dimers, the building blocks of microtubules, preventing their polymerization. This sequestration of tubulin dimers disrupts the normal dynamic equilibrium necessary for microtubule formation. Upon binding, vinca alkaloids stabilize the tubulin dimers, making them unavailable for microtubule assembly. This action is visually represented where vinca alkaloids (depicted in pink) bind to the alpha-beta tubulin dimers (white and blue) and prevent their polymerization. Additionally, vinca alkaloids can induce the formation of paracrystalline aggregates of tubulin, further inhibiting microtubule dynamics. Vinca alkaloids induce mitotic catastrophe, a type of cell death triggered by aberrations in mitosis, leading to:
  
  • Polyploid Cells: Cells with more than two sets of chromosomes.
  • Aneuploid Cells: Cells with an abnormal number of chromosomes.
  These aberrant cells often appear as enlarged multinucleated giant cells under microscopic examination, which ultimately undergo lysis and cell death.

• Another class of microtubule “poisons” are the taxanes, derived from the Taxus baccata plant. Docetaxel, a prominent member of this family, is a complex molecule used to treat a variety of cancers, including breast, prostate, ovarian, lung, kidney cancer, and melanoma. It is notably effective in prostate cancer therapy. Taxanes have an opposing effect to vinca alkaloids, they bind to beta-tubulin within microtubules and promote their stabilization, preventing disassembly. This action hinders the normal process of microtubule shortening necessary for chromosome segregation during cell division. These drugs bind to the ends of microtubules, known as “caps.” Microtubule caps with GTP-bound tubulin are stable, whereas those with GDP-bound tubulin are unstable and prone to depolymerization. Taxanes effectively “freeze” the microtubules in a polymerized state, blocking the movement of chromatids during mitosis. When cancer cells are in metaphase, taxanes prevent the shortening of microtubules, which is crucial for the separation and migration of chromatids to the spindle poles. The inability to complete this phase of cell division results in cell death.