Cytotoxic Chemotherapy (1)

define chemotherapy

• it is the use of cytotoxic drugs to kill cancer cells or inhibit their proliferation

• they target rapidly dividing cells by interfering with

state the 3 major approaches to treating cancer

what is used depends on tumour type and cancer stage, can also be used in combination

• surgical incision

• chemotherapy

• irradiation

describe the clinical contexts that chemotherapy can be used in

• induction therapy- initial intensive treatment used to induce remission; used for AML

• Adjuvant- done following primary mode of treatment e.g. surgery to eradicate any microscopic residual disease to reduce risk of recurrence e.g. in localised breast cancer

• Curative-intent therapy- chemotherapy used with the intent to eradicate the disease completely; e.g. R-CHOP for diffuse large B-cell lymphoma

• Palliative in advanced/metastatic disease- to relieve symptoms, slow progression and improve quality of life

define the cell cycle and describe the phases

the process during which a somatic cell duplicates its content to divide

• G1- cell grows and prepares for DNA synthesis

• S phase- DNA synthesis replication

• G2 phase- cell rapidly grows and prepares for mitosis

• M phase- mitosis

describe how the cells regulate proliferation and how this becomes dysregulated in tumours

• Cells pass through control points that regulate proliferation and these occur:

  • G1/S checkpoint- occurs before the S phase to commit the DNA replication

  • G2/M checkpoint- occurs after the S phase to allow entry of correctly replicated DNA into mitosis

  • Mitotic spindle checkpoint

• In tumour cells there is loss of normal checkpoint control leading to disrupted cell cycle regulation and consequently dysregulated proliferation, which is the primary characteristic of cancer cells

• imbalance between proliferation and cell loss

describe the non-cell cycle specific chemotherapy agents and give examples

• can damage cells in any phase of the cell cycle but cytotoxicity is usually greater in proliferating cells that resting G₀ cells

• effect correlates with total dose intensity (D-R relationship)

• examples

  • platinum compounds, alkylating agents e.g. nitrogen mustards, anthracycline antibiotics e.g. mitomycin

describe the cell cycle specific chemotherapy agents and give examples for each phase

• targets cells at specific phases of the cell cycle

• effect depends on the duration of exposure, the timing of the cycle phase and the fraction of cells entering the relevant phase

• examples

  • M (mitosis) phase- vinca alkaloids, taxanes

  • G2 phase- antitumour antibiotics e.g. bleomycin

  • S phase- antimetabolites

  • G1 phase- antitumour antibiotics e.g. dactinomycin

describe the MOA of cisplatin cytotoxicity

• enter the cell via transport-mediated uptake (CTR1) or passive diffusion due to small size and neutral charge

• aquation where Cl- ions are replaced with water as i/c Cl- is lower than e/c Cl-

• generates reactive positively charged Pt species that binds to negatively charged DNA specifically at N7 position of guanine bases

• forms predominantly intrastrand DNA crosslinks

• DNA damage triggers apoptosis either via p53 dependent or independent pathways

explain the difference between cisplatin MOA of cytotoxicity compared to other Pt groups

• core mechanism the same but leaving group (Cl-) in cisplatin is different in other drugs

• other drugs undergo hydrolysis instead of aquation

state the reasons why tumours are resistant to cisplatin

either intrinsic quality or acquired during therapy

• failure to achieve a sufficient amount of Pt reaching the DNA

• failure to achieve cell death after Pt-DNA adduct formation

explain why the cells fails to achieve a sufficient amount of Pt reaching the DNA

• Decreased uptake- reduced activity/expression of membrane transport systems needed for Pt entry

• Increased i/c detoxification- elevated levels of sulphur-rich molecules e.g. glutathione bind to reactive Pt species and form inactive complexes

• Enhanced efflux or sequestration mechanisms e.g. upregulation of copper exporting ATPases lower the effective i/c Pt concentration

explain why some cells fail to achieve cell death after Pt-DNA adduct formation

• Increased repair or Pt-induced DNA lesions, notably by upregulation of nucleotide excision repair (e.g. ERCC1)- increased removal of intrastrand adducts and reduced cytotoxicity

• Increased tolerance to DNA damage e.g. through defects in mismatch repair or specialised DNA polymerases that bypass Pt adducts

  • In a normal cell, the mismatched repair can repair to a defect and respond to it

  • But a defect in mismatched repair protein, means that the cell cannot respond to it

• Altered apoptosis signalling- loss of p53 function or increased anti-apoptotic proteins (e.g. BCL-2)

• what regimen is oxaliplatin used in

• describe the side effects of oxaliplatin

• how is oxaliplatin administered

• used as part of FOLFOX regimen (FOLinic acid + Fluorouracil (5-FU) + Oxaliplatin)

• Side effects: tingling, numbness, nausea. Neurotoxicity is the most frequent dose-limiting toxicity

• Administered via i.v. infusion, primarily for colorectal cancer

describe the structural differences between cisplatin and oxaliplatin and how this forms the basis of oxaliplatin overcoming tumour resistance

• differences- Cl- ions replaced with oxalate leaving group and amine group replaced with bulky DACH (diaminocyclohexane) moiety  

• Oxalate leaving group forms a stable ring-like structure around Pt which slows down hydrolysis in the cells and leads to slower activation rate

• Oxaliplatin keeps its bulky DACH moiety upon DNA binding protruding from the DNA helix and this creates a DNA conformational distortion

• The bulky DACH ligand results in DNA adducts that are less efficiently recognised and removed by nucleotide excision repair

• Oxaliplatin-induced DNA adducts are processed differently by DNA repair pathways

describe the cellular response to oxaliplatin

• Under normal conditions, p53 levels are kept low because it is ubiquitinated by MDMA and tagged for proteasomal degradation

• But oxaliplatin-induced DNA damage activates ATM/ATR kinases, which phosphorylates p53, and prevents MDM2-mediated ubiquitination and that allows p53 accumulation

• High p53 levels leads to either cell cycle arrest at G2/M checkpoint to repair the DNA or if the DNA damage is too severe, it will trigger apoptosis

describe the acute neurotoxicity associated with oxaliplatin

• Occurs in majority of patients during infusion or within hours

• Symptoms are induced by exposure to cold

• Sensory symptoms- distal paraesthesia and/or dysesthesias (tingling or numbness in extremities), perioral (mouth) numbness

• 1-2% of patients report cold-induced pharyngolaryngeal dysesthesia- intense discomfort in their throat, feels like choking

• Motor symptoms- muscle cramps or tetanic spasms, transient neuromuscular hyperexcitability

• May reflect hyperexcitability of peripheral neurons rather than axonal degradation

• Symptoms are transient and reversible between cycles

describe the chronic sensory neuropathy associated with oxaliplatin

• Seen in 10-15% of patients after repeated exposure (dose-dependent)

• Primarily non cold-related dysesthesias and paraesthesia of the extremities

• Clinical features- impaired sensation, sensory ataxia, distal dysesthesias and paraesthesia's, deficit in fine sensory motor coordination

• Symptoms generally persist between cycles and worsen with cumulative doses

• At higher grades, symptoms interfere with daily activity

• Often partially reversible- most with grade 3 neurotoxicity improve to grade 1 or less within 6-12 months after discontinuing treatment

name the 3 mechanisms of oxaliplatin induced ACUTE neuropathy and describe where it occurs and why

occurs in the DRG sensory neurons which are vulnerable to damage as they lack BNB- permeable so drug can accumulate here

• ion channel dysfunction

• TRPA1 sensitisation → lowers cold activation threshold

• copper transporter (CTRs)

describe how oxaliplatin causes ion channel dysfunction

• Altered function of voltage gated Na+, K+ and Ca2+ channels

• This disrupted ion flux causes neuronal hyperexcitability and abnormal sensory signal transmission

• This neuronal hyperexcitability happens because of two reasons:

  1. The oxalate byproduct chelates Ca2+ and Mg2+ which disrupts ion channel gating

  2. Oxaliplatin also directly alters voltage gated Na+ and K+ ion channel function

• These changes are largely functional and reversible

describe how oxaliplatin causes TRPA1 sensitisation

• TRPA1 is a cold-sensitive ion channels

• Oxaliplatin enhances TRPA1 activity through mechanisms that include oxidative stress and altered Ca2+ homeostasis

• The sensitised TRP channels lower the threshold for activation by cold stimuli

describe the role of the CTRs in oxaliplatin induced acute neuropathy

• CTR1 contributes to oxaliplatin uptake though it is less CTR1-dependent than cisplatin

name the 3 mechanisms of oxaliplatin induced CHRONIC neuropathy

• neuroinflammation

• DNA damage in DRG

  • oxidative stress

describe how neuroinflammation causes oxaliplatin induced chronic neuropathy

• Increased production of pro-inflammatory cytokines and chemokines

• The sustained activation of inflammatory contributes to progressive neuronal sensitisation

• There is abnormal communication between neurons and glial cells that sustains the neuropathic pain

• Neuroinflammation occurs early on the in treatment but the sustained neuroinflammation is characteristic of chronic neuropathy

describe how DNA damage in DRG causes oxaliplatin induced chronic neuropathy

• Neurons have limited DNA repair capacity because neurons are post mitotic (they don’t divide)

describe how oxidative stress in DRG causes oxaliplatin induced chronic neuropathy

• Develops with cumulative oxaliplatin doses

• Repeated treatment increased accumulation of ROS which leads to mitochondrial DNA damage

• Mito DNA are much less able to repair themselves than nuclear DNA which leads to dysfunction of respiratory complexes that reduces ATP production leading to ATP depletion

• Neurons are highly energy dependent so the ATP depletion contributes to energy failure which leads to axonal degeneration

describe the toxicity pattern of the Pt-based chemo agents

cisplatin

• Produces dose-limiting nephrotoxicity

• Causes moderate neurotoxicity and high ototoxicity

oxaliplatin

• Severe peripheral neuropathy and moderate myelosuppression

carboplatin

• High risk of myelosuppresion (bone marrow suppression)

describe the MOA of the cytotoxicity of alkylating agents

• attach alkyl groups to DNA, forming DNA adducts that may trigger apoptosis

• These adducts disrupt DNA replication and transcription, activate DNA damage pathways and can induce apoptosis

though cell specific, which cell cycle stage are cells more sensitive to alkylating agents and why

• These are cell cycle phase non-specific but cells in S phase are more vulnerable as replication forks stall at unrepaired adducts

describe the mechanism of DNA damage of alkylating agents

• generate reactive electrophilic intermediates that covalently bind to nucleophilic sites on DNA (most commonly N7 position of guanine)

• The alkylation results in

  • Abnormal base pairing e.g. guanine mispairing with thymine

  • Accumulation of secondary strand breaks following base excision repair attempts (DNA fragmentation)- occurs because alkylating agent overwhelms repair system

  • Intrastrand or interstrand crosslinks

• All of these interfere with DNA replication and transcription

• After DNA damage we get Bax/Bak activation which leads to pores forming in the OMM, leading to MOMP, CytC release and apoptosis/necrosis

describe the metabolism of nitrogen mustards

• administered as prodrugs that require metabolic activation in the liver by CYP450 enzymes (cyclophosphamide: CYP2B6 and ifosfamide: CYP3A4)

• These compounds can generate 2 metabolites:

  • Phosphoramide mustard- therapeutically active, cytotoxic species; gives us anti-tumour activity

  • Acrolein- highly reactive cytotoxic byproduct; responsible for urotoxicity

describe where nitrogen mustards are used

used in chemotherapy in range of cancers such as breast and ovarian

• cyclophosphamide is also used as immunosuppressant for many autoimmune conditions e.g. lupin

describe the toxicity profile of alkylating agents

myelosuppression

• most common dose limiting toxicity

• alkylation damages rapidly dividing marrow progenitor cells

• leukopenia, neutropenia, thrombocytopenia

urological toxicity

• Haemorrhagic cystitis is the most common- this is bladder mucosal damage

• characteristic of cyclophosphamide and ifosfamide due to the acrolein

  • Acrolein concentrated with the bladder and has direct contact with the urothelium- leads to OS and epithelial injury

describe how haemorrhagic cystitis caused by alkylating agents can be prevented/treated

• This can be prevented by aggressive hydration to flush away toxic metabolites, and the use of mesna

  • mesna coadministered with ifosfamide which has higher risk of HC

  • mesna detoxifies acrolein metabolite