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PA6.2-3 | Molecular Basis of Cancer & Carcinogenesis — SDL Guide (Part 2)

Apoptosis Regulators and DNA Repair Genes

A four-panel medical diagram explaining BCL2 apoptosis inhibition, mismatch repair failure, BRCA-related DNA repair defects, PARP inhibitor synthetic lethality, and epigenetic tumor suppressor silencing. — **Panel A:** BCL2, outer mitochondrial membrane, cytochrome-c release, apoptosis, t(14;18), immunoglobulin heavy-chain promoter, BCL2 overexpression, follicular lymphoma, lymphocyte accumulation. **Panel B:** DNA replication mismatch, mismatch repair, MLH1, MSH2, MSH6, PMS2, Lynch syndrome/HNPCC, microsatellite instability, colorectal cancer, endometrial cancer. **Panel C:** Double-strand DNA break, BRCA1, BRCA2, homologous recombination, chromosomal instability, error-prone repair, PARP inhibitor, synthetic lethality, BRCA-deficient tumor cell. **Panel D:** Global hypomethylation, transposable element activation, proto-oncogene activation, CpG island promoter hypermethylation, tumor suppressor gene silencing, epigenetic second hit.

Apoptosis regulators:
BCL2 (B-cell lymphoma 2) is the prototype anti-apoptotic protein. It resides on the outer mitochondrial membrane and prevents cytochrome-c release. In follicular lymphoma, the t(14;18) translocation places BCL2 under the immunoglobulin heavy-chain promoter → constitutive overexpression → lymphocytes cannot die → accumulate as low-grade lymphoma. BCL2 does NOT drive proliferation; it prevents death — an important distinction from oncogenes.

DNA repair genes — 'caretaker' genes:
When DNA repair fails, mutation rate rises and clonal evolution accelerates.

Mismatch repair (MMR) genes (MLH1, MSH2, MSH6, PMS2) — correct base-pair mismatches after replication. Germline loss → Lynch syndrome (hereditary non-polyposis colorectal cancer, HNPCC): early-onset colorectal and endometrial cancers with microsatellite instability (MSI) as the molecular signature. Somatic MMR loss also occurs in sporadic colon, gastric, and endometrial cancers.

BRCA1/2 — as noted above, enable homologous recombination. Loss → error-prone repair, chromosomal instability, double-strand break accumulation. BRCA-deficient tumours are exquisitely sensitive to PARP inhibitors (synthetic lethality).

Epigenetic alterations:
Cancer genomes are globally hypomethylated (activating transposable elements and proto-oncogenes) yet show focal hypermethylation of TSG promoter CpG islands, silencing them without mutation — the so-called 'epigenetic second hit'.

MicroRNAs (miRNAs):
miRNAs are small non-coding RNAs that post-transcriptionally repress target mRNAs. OncomiRs (e.g., miR-21) suppress TSGs; tumour-suppressor miRNAs (e.g., miR-15/16) suppress BCL2. miRNA dysregulation is now a recognised mechanism of carcinogenesis and a potential biomarker.

CLINICAL PEARL

Practical yield of MMR/MSI testing: Universal MMR immunohistochemistry (or MSI-PCR) is now standard on all newly diagnosed colorectal cancers. MSI-high status predicts: (1) Lynch syndrome workup needed if patient is <70 years, (2) poor response to standard 5-FU adjuvant chemotherapy, but (3) excellent response to immune checkpoint inhibitors (pembrolizumab). Understanding the biology directly changes management.

Multistep Carcinogenesis and Clonal Evolution

No single mutation transforms a normal cell into a fully malignant cancer. Multistep carcinogenesis is the sequential acquisition of mutations over years to decades, each conferring a selective growth advantage — Darwinian evolution within a tissue.

The colon provides the best-characterised model (Fearon–Vogelstein sequence):

APC loss → aberrant crypt foci → early adenoma (KRAS mutation) → intermediate adenoma (SMAD4/DPC4 loss, TGF-β pathway) → late adenoma (TP53 loss) → invasive adenocarcinoma

This sequence typically takes 10–15 years, providing a window for screening colonoscopy to interrupt progression.

Key principles:
• The order of mutations matters — APC loss is the obligate early event; TP53 loss is typically late
• 'Driver mutations' provide selective advantage; 'passenger mutations' accumulate but are not necessary
• Clonal evolution: a single initiated cell with a growth advantage outcompetes neighbours, then subclones with further mutations arise within that clone — the basis of tumour heterogeneity
• Epigenetic changes and chromosomal instability (CIN) amplify the mutation rate at each step

Diagram showing the stepwise progression from normal colonic crypt epithelium to invasive adenocarcinoma with APC, KRAS, SMAD4/DPC4, and TP53 changes plus clonal evolution and screening relevance. — **Panel A:** Fearon-Vogelstein sequence: normal colonic crypt epithelium, APC loss, aberrant crypt focus, early adenoma with KRAS mutation, intermediate adenoma with SMAD4/DPC4 loss and TGF-beta pathway disruption, late adenoma with TP53 loss, invasive adenocarcinoma crossing the basement membrane, 10-15 year window, colonoscopy screening interruption.. **Panel B:** Clonal evolution in a colonic crypt: initiated APC-mutant cell, selective growth advantage, expanding clone, KRAS subclone, SMAD4/DPC4 subclone, TP53 subclone, tumour heterogeneity, branching phylogeny.. **Panel C:** Key principles: driver mutation, passenger mutation, chromosomal instability, epigenetic silencing, APC as obligate early event, TP53 as late event, mutation order matters..

SELF-CHECK

A 45-year-old man undergoes surveillance colonoscopy for familial adenomatous polyposis (FAP). A 1.2 cm tubular adenoma is removed. Sequencing reveals: APC germline mutation (inherited) + somatic KRAS Gly12Val mutation. According to the multistep model, what is the MOST likely next molecular event if this adenoma were to progress toward carcinoma?

A. Somatic APC second-hit mutation

B. KRAS amplification

C. Loss of SMAD4 or TP53 function

D. BCL2 translocation

Reveal Answer

Answer: C. Loss of SMAD4 or TP53 function

In the Fearon–Vogelstein sequence, APC loss and KRAS mutation represent early events. The intermediate-to-late adenoma transitions are driven by SMAD4 loss (disabling TGF-β growth arrest) and ultimately TP53 loss (abrogating the DNA-damage checkpoint), enabling invasive behaviour. BCL2 translocation is associated with follicular lymphoma, not colorectal carcinogenesis.

Chemical Carcinogenesis

Chemical carcinogenesis proceeds in two separable stages:

Initiation: an irreversible, heritable DNA mutation induced by a chemical carcinogen. The cell is 'initiated' but not yet transformed.

Promotion: repeated, prolonged exposure to a promoter (not itself mutagenic) causes clonal expansion of the initiated cell by stimulating proliferation. Promotion is reversible if the promoter is withdrawn early.

Direct-acting vs procarcinogens:
• Direct-acting carcinogens (e.g., alkylating agents, nitrogen mustard, cyclophosphamide) react with DNA without metabolic activation.
• Procarcinogens require metabolic activation by cytochrome P450 enzymes (CYP1A1, CYP2E1) in the liver or target organ to form the ultimate carcinogen — the reactive electrophile that attacks DNA.

Key examples:

Carcinogen	Source	Target organ	Mechanism/Note
Polycyclic aromatic hydrocarbons (PAHs) — benzo[a]pyrene	Tobacco smoke, charbroiled food	Lung, oral cavity	Procarcinogen; CYP-activated to diol-epoxide; G→T transversions in TP53/KRAS
β-Naphthylamine, benzidine	Rubber/dye industry	Urinary bladder	Procarcinogen; hepatic acetylation detoxifies, but urinary deconjugation reactivates it at bladder urothelium
Aflatoxin B1	Aspergillus flavus on peanuts/grain	Liver (HCC)	Procarcinogen; CYP3A4 → epoxide; G→T transversion at codon 249 of TP53 (hotspot mutation)
Nitrosamines	Cured/pickled foods; tobacco	Gastric, oesophageal	CYP-activated; prevalent in high-gastric-cancer regions
Asbestos	Mining, insulation	Mesothelioma, lung (co-carcinogen with tobacco)	Physical fibre mechanism + ROS; synergises with smoking for lung cancer (multiplicative risk)
Vinyl chloride	PVC industry	Angiosarcoma liver	Direct alkylation after CYP activation

Note the tissue specificity of carcinogens — explained by local metabolic activation, target cell vulnerability, or topographic concentration (e.g., urothelium concentrates urinary carcinogens).