Molecular Pathology Seminar 8: Molecular Genetic Testing In Breast Cancer With Case Study
Molecular genetic testing in breast cancer involves analyzing tumor biomarkers such as HER2 amplification, estrogen/progesterone receptor status, and gene expression profiles (e.g., Oncotype DX, MammaPrint) to guide prognosis, predict therapeutic response, and personalize treatment decisions.
Molecular Genetic Testing in Breast Cancer with Case Study
This lecture by Prof US Khoo is divided into two major parts and two case studies. It addresses the why, when, and how of molecular testing in breast cancer — specifically HER2 testing (Part I) and Gene Expression Profiling (Part II). The overarching concept is that breast cancer is not one disease: its molecular characteristics dictate prognosis and treatment. The lecture connects pathology to clinical decision-making, showing how a single test result (HER2 status, Oncotype Dx score) can fundamentally change whether a patient receives targeted therapy or chemotherapy.
Learning Objectives (directly from the lecture) [1]:
- When should HER2 test be performed?
- What clinical material is used for the test?
- Who will benefit from this test?
- What do the HER2 IHC and FISH test results convey?
- What are the pitfalls and limitations of these tests?
- Understand the rationale for gene expression profiling and the major commercially available assays.
- Apply knowledge to clinical case studies.
How this fits into exams: HER2 testing is a perennial favourite for MCQ, SAQ, and mini-case questions. Past papers repeatedly test: IHC scoring, when to reflex to FISH, interpretation of FISH ratios, clinical implications of HER2 positivity, Herceptin side effects, and the concept of gene expression profiling guiding chemotherapy decisions. [3][4][5]
Part I: HER2 Testing in Breast Cancer
The treatment pathway begins with pre-operative diagnosis by core biopsy, which determines whether a patient proceeds to primary surgical resection, neo-adjuvant chemotherapy, or primary endocrine therapy. [1]
The pathological features of the resected specimen then guide adjuvant therapy. This is why accurate molecular testing at diagnosis is critical — it shapes the entire treatment trajectory.
Traditional pathologic prognostic features [1]:
- Tumor size
- Lymph node status (strongest single prognostic factor [2])
- Histologic features: type, grade, lymphovascular invasion, margins
- ER/PR status
- HER2 status
High Yield – Prognostic vs Predictive
ER/PR status is both prognostic (better outcomes) and predictive (response to hormonal therapy). HER2 status is prognostic (worse outcomes if positive, untreated) and predictive (response to anti-HER2 therapy and certain chemotherapy regimens). [1]
HER2 is the common name for the human epidermal growth factor receptor 2 gene (ERBB2). It encodes a transmembrane tyrosine kinase receptor protein found on the surface of cells. [1]
Why does HER2 matter?
Normal cells have ~2 copies of the HER2 gene and express low levels of HER2 protein. In approximately 20-25% of breast cancers, the HER2 gene is amplified (extra copies of the gene), leading to protein overexpression on the cell surface. When epidermal growth factor (or other ligands) binds to these overabundant receptors, there is excessive activation of downstream signaling cascades (RAS-MAPK, PI3K-AKT) → uncontrolled cell proliferation, survival, and invasion. [1]
HER2 gene amplification is the PRIMARY mechanism for HER2 protein overexpression. [1]
This is the fundamental molecular principle: gene amplification → mRNA overproduction → protein overexpression → oncogenic signaling.
HER2 protein overexpression is associated with [1]:
- Higher risk of recurrence and mortality
- Relative resistance to endocrine treatment
- Less benefit from some forms of chemotherapy
High Yield – Clinical Significance of HER2
Without anti-HER2 therapy, HER2-positive breast cancer has one of the worst prognoses. With targeted therapy (Herceptin), outcomes are dramatically improved. This is why accurate testing is non-negotiable.
| Agent | Mechanism | Clinical Benefit | Key Side Effects |
|---|---|---|---|
| Trastuzumab (Herceptin) | Monoclonal antibody targeting extracellular domain of HER2; interferes with signal transduction cascade | Reduces risk of recurrence by ~50% and mortality by ~1/3 in early-stage HER2+ breast cancer | Cardiac toxicity (cardiomyopathy); very expensive (~US$70-110K/year) |
| Lapatinib | Small molecule dual inhibitor of HER2 and EGFR tyrosine kinase activity | Improved outcomes in metastatic HER2+ breast cancer | Diarrhea, hepatotoxicity, rash |
Herceptin is effective ONLY for patients who are HER2 positive. [1]
This is precisely why testing accuracy is so critical. An inaccurate test has dire consequences in both directions.
Exam Favourite – Herceptin Side Effects
Up to 18% of patients could receive Herceptin unnecessarily — leading to unnecessary expense, cardiac toxicity, and delay of appropriate treatment. [1]
Up to 10% of patients may NOT receive appropriate HER2-targeted therapy — denying them a life-saving treatment. [1]
This is why HER2 testing must follow strict quality-assured protocols. It is not simply a special stain — a single observation leads to a major difference in treatment. [1]
ALL patients with invasive breast carcinoma should be tested for HER2. [1]
HER2 analysis MUST be performed on the INVASIVE component of the breast cancer. [1]
Why the invasive component specifically? Because HER2 overexpression/amplification is frequently present in DCIS (in-situ cancer), but treatment decisions (especially regarding Herceptin) are based on the invasive component. Testing DCIS alone could yield a misleading result that does not reflect the biology of the invasive disease. [1]
Material: Formalin-fixed, paraffin-embedded (FFPE) tissue sections from core biopsy or surgical resection specimen. [1]
There are two main categories of assays, targeting different levels of the central dogma:
| Level | Assay | What It Detects | Notes |
|---|---|---|---|
| Protein | Immunohistochemistry (IHC) | HER2 protein overexpression on cell membrane | Cheaper, easier, first-line test |
| DNA | In Situ Hybridization (ISH) | HER2 gene copy number (amplification) | Reflex test for equivocal IHC |
ISH subtypes [1]:
- FISH — Fluorescence in situ hybridization (gold standard)
- CISH — Chromogenic ISH
- SISH — Silver-enhanced ISH
- DISH — Dual ISH (combined approach)
The central dogma reminder (DNA → mRNA → Protein) explains why we can test at different levels:
- IHC detects the end product (protein)
- ISH detects the root cause (gene amplification)
In-situ hybridization (ISH) is a method of localizing and detecting specific DNA or mRNA sequences in morphologically preserved tissue sections or cell preparations. [1]
How it works:
- Denaturation: Target DNA in the tissue section is heated to separate double-stranded DNA into single strands
- Hybridization: A complementary labeled probe binds (hybridizes) to the target sequence
- Visualization: The probe label is detected — by fluorescence (FISH), chromogenic reaction (CISH), or metallographic method (SISH)
Probe visualization methods [1]:
- Radioactive labels (historical)
- Non-radioactive labels: Chromogenic, Fluorescence dyes, Metallographic
This is absolutely critical for exams. The IHC score determines whether further testing (ISH) is needed.
| IHC Score | Staining Pattern | Interpretation |
|---|---|---|
| 0 | No staining | Negative |
| 1+ | Incomplete membrane, weak staining within > 10% of invasive cells | Negative |
| 2+ | Circumferential membrane, incomplete and/or weak/moderate staining in ≥10% of cells | Equivocal → requires reflex ISH testing |
| 3+ | Circumferential membrane, complete, intense staining in > 10% of invasive cells | Positive |
High Yield – IHC Scoring Discriminators
The key discriminators between scores:
- 0 vs 1+: Both negative, but 1+ has detectable (though weak/incomplete) staining
- 1+ vs 2+: 1+ is incomplete + weak; 2+ has circumferential (wrapping around cell) but still incomplete/weak-moderate staining
- 2+ vs 3+: 2+ is incomplete or weak-moderate circumferential; 3+ is COMPLETE + INTENSE circumferential
- The threshold is > 10% of invasive cells for all positive scores
- Only 3+ is definitively HER2 positive by IHC alone
PathVysion HER2 DNA Probe Kit (FDA-approved) [1]:
- Dual-probe system (built-in control):
- HER2 DNA probe: SpectrumOrange labeled, 190 kb, targets HER2 gene locus at 17q11.2-q12
- CEP17 probe: SpectrumGreen labeled, 5.4 kb, targets centromeric alpha satellite DNA of chromosome 17 (17p11.1-q11.1) — serves as internal control for chromosome 17 copy number
FISH procedure [1]:
- Specimen DNA denatured to single-stranded form
- Allowed to hybridize with PathVysion probes
- Unbound probe washed away
- Nuclei counterstained with DAPI (blue fluorescent nuclear stain)
Signal enumeration [1]:
- Use DAPI/Orange/Green triple filter under fluorescence microscope
- Count HER2 (orange) and CEP17 (green) signals in 20 nuclei
- Calculate ratio = total HER2 signals / total CEP17 signals
- Calculate average HER2 signal per cell
Worked example from lecture [1]:
| Parameter | Value |
|---|---|
| Total HER2 signals (20 nuclei) | 176 |
| Total CEP17 signals (20 nuclei) | 51 |
| Average HER2/nucleus | 8.8 |
| Average CEP17/nucleus | 2.55 |
| HER2/CEP17 ratio | 3.45 |
This result = amplified (ratio ≥ 2.0)
INFORM HER2 Dual ISH DNA Probe Cocktail assay (Ventana Medical Systems) [1]:
- Processed automatically
- HER2 gene probe: dinitrophenol-labeled → detected via peroxidase/enzyme metallography (SISH) → appears black
- CEP17 probe: digoxigenin-labeled → detected via anti-digoxigenin antibody/CISH → appears red
- Same interpretation criteria as FISH
This is the dual-probe ISH algorithm from the 2018 ASCO/CAP guidelines [1]:
| Dual-Probe ISH Result | HER2/CEP17 Ratio | Average HER2 Copy Number/Cell | Interpretation | Concurrent IHC Needed? |
|---|---|---|---|---|
| Group 1 | ≥ 2.0 | ≥ 4.0 | ISH positive → HER2 positive | No (but IHC recommended) |
| Group 2 | ≥ 2.0 | < 4.0 | Requires concurrent IHC | Need IHC 3+ to be HER2+ |
| Group 3 | < 2.0 | ≥ 6.0 | Requires concurrent IHC | Need IHC 2+ or 3+ to be HER2+ |
| Group 4 | < 2.0 | ≥ 4.0 and < 6.0 | Requires concurrent IHC | Need IHC 3+ to be HER2+ |
| Group 5 | < 2.0 | < 4.0 | ISH negative → HER2 negative | No |
Assess IHC using sections from the SAME tissue sample used for ISH [1]
High Yield – 2018 ISH Algorithm Key Points
- The simple rule: HER2/CEP17 ratio ≥ 2.0 OR average HER2 copy number ≥ 6.0 = amplified (but with caveats in the 2018 guidelines requiring concurrent IHC in certain scenarios)
- Groups 2, 3, and 4 are the "grey zones" requiring concurrent IHC to determine final HER2 status
- This reflects the move toward requiring concordance between protein expression and gene amplification for treatment decisions
Critical pre-analytical factors [1]:
- ≤ 1 hour cold ischaemic time (time from removal of tissue to fixation)
- Record time tissue obtained and time fixed (for quality assurance)
- 10% Neutral Buffered Formalin fixation
- Minimum 6 hours fixation
- Maximum 72 hours fixation (over-fixation can mask antigens and degrade nucleic acids)
Why this matters: Poor fixation leads to false-negative IHC results (protein degradation) or failed FISH (DNA degradation). This is a source of pre-analytical error that directly impacts patient care.
Concurrently run control slides [1]:
- Normal control (non-amplified)
- Amplified or weakly amplified control
This ensures signal enumeration accuracy and that the assay is performing correctly.
Exclusion criteria for HER2 FISH assay [1]:
Sample limitations:
- Samples with only limited invasive carcinoma (insufficient tumor cells)
- Tissue fixed in other than NBF or for greater than 72 hours
Test performance limitations:
- Controls with unexpected results (assay failure)
- FISH signals non-uniform, background obscures signal, or non-optimal enzymatic digestion
Histopathologic discordance = when HER2 status does not match what is expected based on histologic features [1]
Key histopathologic patterns [1]:
- Grade 3 invasive carcinoma is more likely to be HER2+
- Good-prognosis subtypes (tubular, mucinous, cribriform, adenoid cystic, ER/PR-positive invasive ductal/lobular carcinoma) are usually NOT HER2+
When discordance occurs (e.g., low-grade ER+ tumor testing HER2+, or grade 3 tumor testing HER2-), the ASCO guideline recommends reflex testing using an alternative assay. [1]
HER2 status (positive or negative) must be determined in ALL patients with invasive breast cancer (early stage or recurrence) on the basis of one or more HER2 test results. [1]
HER2-targeted therapy is recommended if HER2 positive: IHC score 3+ OR ISH amplified. [1]
If HER2 equivocal OR apparent histopathologic discordance → reflex testing using alternative assay. [1]
This is an emerging concept that changes the paradigm:
Novel anti-HER2 compound T-DXd (trastuzumab deruxtecan, Enhertu®) is an antibody-drug conjugate (ADC) that delivers chemotherapy payload directly to the tumor site. [1]
T-DXd is active against breast cancers that LACK protein overexpression or gene amplification — called HER2-low breast cancer. [1]
HER2-low definition [1]:
- IHC 1+ (incomplete membrane, weak staining in > 10% of invasive cells)
- IHC 2+ non-amplified (equivocal IHC with negative ISH)
High Yield – 2023 ASCO-CAP Recommendations for HER2-low
Previous recommendations for classic anti-HER2 therapies remain unchanged. However, for metastatic patients without HER2 overexpression or gene amplification, an IHC 1+ or 2+ result may make patients eligible for treatment with trastuzumab-deruxtecan (the only currently available agent for HER2-low). [1]
This means:
- IHC 0 = truly HER2-negative (no HER2-targeted therapy at all)
- IHC 1+ or IHC 2+ non-amplified = HER2-low (eligible for T-DXd in metastatic setting)
- IHC 3+ or ISH amplified = HER2-positive (eligible for classic anti-HER2 therapies)
Part II: Gene Expression Profiling for Clinical Management
Breast cancer is very heterogeneous. Patients with the same stage of disease can have markedly different treatment responses and overall outcomes. [1]
Traditional prognostic features (size, grade, LN status, ER/PR/HER2) have limitations:
- Tumors with similar features may have different clinical outcomes [1]
- Cannot reliably predict response to specific therapeutic options for individual patients
Gene Expression Profiling detects the activity of thousands of genes at once to create a global molecular picture. It can:
- Assess prognosis (risk of recurrence)
- Predict response to therapy (especially chemotherapy benefit) [1]
Two approaches [1]:
- Bottom-up (biology-driven): Start with clinical outcome data → identify candidate genes → build prognostic model
- Top-down (hypothesis-driven): Start with biologic hypothesis → select candidate genes → validate against outcomes
Gene expression profiling has revealed that breast cancer comprises distinct molecular subtypes with different prognoses and treatment responses. [1]
| Subtype | ER/PR | HER2 | Grade | Prognosis | Treatment | Frequency |
|---|---|---|---|---|---|---|
| Luminal A | + | - | Low | Best | Hormonal therapy | ~40% |
| Luminal B | + | ± | Higher | Worse than A | Hormonal ± chemo ± anti-HER2 | ~20% |
| HER2-enriched | - | + | High | Poor (without Tx) | Anti-HER2 + chemo | ~10-15% |
| Basal-like / Triple Negative | - | - | High | Poor | Chemo only (platinum, PARP inhibitors) | ~15-20% |
Connection to BRCA [6]:
- BRCA1 mutations → typically triple-negative/basal-like (ER/PR/HER2 negative, CK5/6+, high Ki67)
- BRCA2 mutations → typically luminal A (ER/PR positive, HER2 negative)
| Feature | MammaPrint | OncotypeDx | PAM50 (Prosigna) | EndoPredict |
|---|---|---|---|---|
| Number of genes | 70 | 21 | 50 | 12 |
| Tissue type | Fresh frozen (originally; now FFPE available) | FFPE | FFPE | FFPE |
| Target population | Early-stage (I/II), LN-negative, ER+ AND ER-, age < 61, tumor < 5cm | Early-stage (I/II), LN-negative, ER+ | Incorporates ER, nodal, grade, size; provides 10-year risk | ER+, HER2- |
| Output | Low vs High risk of recurrence | Recurrence Score: Low (< 18), Intermediate (18-31), High ( > 31) | Risk of recurrence score | EPclin Risk Score: < 3.33 (low) vs > 3.33 (high) |
| Key trial | MINDACT | TAILORx (NSABP B-20) | DGCG89D | — |
High Yield – OncotypeDx Recurrence Score and Chemotherapy Benefit
The landmark NSABP B-20 study showed [1]:
- RS ≥ 31 (High): 28% absolute benefit from adding chemotherapy to tamoxifen (P < 0.001) → SHOULD receive chemoendocrine therapy
- RS 18-30 (Intermediate): Small, non-significant benefit from chemotherapy (4.4% absolute benefit, P = 0.39) → Uncertain benefit
- RS < 18 (Low): Very low likelihood of distant recurrence; no significant benefit from adding chemotherapy (P = 0.61) → Endocrine therapy alone sufficient
This is the clinical utility: OncotypeDx identifies patients who can safely AVOID chemotherapy (low RS) and those who clearly benefit from it (high RS).
- 50-gene subtype predictor using Nanostring platform [1]
- Incorporates standard clinical parameters (ER status, nodal status, grade, size)
- Provides 10-year probability of risk of recurrence
- DGCG89D trial: Patients with HER2-enriched breast cancer derived substantial benefit from anthracycline chemotherapy; anthracyclines are NOT essential for luminal subtypes [1]
Proliferation genes are the common driving force in ALL prognostic gene expression signatures. [1]
This makes intuitive sense: rapidly proliferating tumors are more aggressive, more likely to recur, and more likely to respond to chemotherapy (which targets dividing cells).
Clinical scenario: 35-year-old woman, left breast mass, grade 1 invasive carcinoma (no special type), 9mm, LVI negative, sentinel LN negative (0/1), margins clear. IHC: ER positive (Allred 6/8), PR positive (Allred 6/8), HER2 equivocal (2+), Ki67 10%. [1]
Q1: What additional test would the pathologist perform? → HER2 ISH (FISH or DISH) — because HER2 IHC is equivocal (2+), requiring reflex testing to determine gene amplification status [1]
Q2: What is the purpose? → To determine whether HER2 gene is amplified, which would indicate eligibility for anti-HER2 targeted therapy (Herceptin) [1]
Q3: On what material? → FFPE tissue of the INVASIVE component of the breast cancer (not the DCIS component) [1]
Q4: Which field is appropriate? → Field showing INVASIVE carcinoma (not in-situ component) [1]
Q5: FISH interpretation (green = CEP17, red = HER2): → Count HER2 (red) and CEP17 (green) signals in 20 nuclei → calculate ratio and average HER2 copy number → interpret per 2018 algorithm [1]
Clinical context note: This is a low-grade, small, ER-positive, LN-negative tumor with low Ki67. If HER2 turns out negative (which is expected given the favorable histology), this would be a luminal A-type cancer that likely only needs endocrine therapy. If unexpectedly HER2-positive, this would be a case of histopathologic discordance warranting repeat testing. [1]
Clinical scenario: 52-year-old Mrs. M, simple mastectomy + SLNB for right breast cancer. Grade III (3,3,2), 2.8cm, LVI negative, sentinel LN negative (0/7), margins clear. ER positive (Allred 7/8), PR positive (Allred 7/8), HER2 negative (1+), Ki67 20%. [1]
Q1: Rationale for Gene Expression Profiling? → To provide more accurate prognostic information and predict benefit from adjuvant chemotherapy beyond what traditional pathologic features can offer. This patient has conflicting features — high grade (suggesting aggressive biology) but ER/PR positive and node negative (suggesting favorable prognosis). Gene expression profiling helps resolve this uncertainty. [1]
Q2: Possible GEP tests? → OncotypeDx (21-gene, FFPE, for ER+, LN-negative early-stage cancer) — most appropriate [1] → MammaPrint (70-gene) — applicable to ER+ or ER-, but was originally based on fresh frozen tissue → PAM50 (Prosigna), EndoPredict — also options for ER+, HER2- cancers [1]
Q3: Material? → Formalin-fixed, paraffin-embedded tissue (for OncotypeDx, PAM50, EndoPredict) [1]
Q4: Oncotype Dx RS = 5 (Low). Clinical implication? → RS < 18 = Low recurrence score. The patient has a very low likelihood of distant recurrence. The benefit of adding chemotherapy to endocrine therapy is minimal and not statistically significant. Therefore, this patient can be managed with endocrine therapy alone (tamoxifen or aromatase inhibitor), sparing her the toxicity of chemotherapy. [1]
Clinical Pearl – Case Study 2
Despite having a grade III, 2.8cm tumor (traditionally considered high-risk), the OncotypeDx RS of 5 reclassifies her as LOW risk. This exemplifies the power of gene expression profiling to override traditional histopathologic features and avoid unnecessary chemotherapy.
From GC 156 slides [6]:
BRCA1-associated breast cancers are typically triple-negative (ER/PR/HER2 negative), high grade, high Ki67, basal-like (CK5/6 positive). They show distinct genomic expression, are found in 80-90% of BRCA1 mutation carriers, present in younger patients, frequently as "interval cancers" with rapid progression, and metastasize to brain and lung rather than lymph nodes.
BRCA2-associated breast cancers are typically luminal A type (ER/PR positive, HER2 negative), medium/high grade, and include tubular, cribriform, mucinous, or lobular histology.
This connects to the HER2 testing lecture because:
- BRCA1 cancers are usually HER2-negative → no benefit from Herceptin
- BRCA2 cancers are usually HER2-negative → endocrine therapy is the mainstay
- Gene expression profiling (e.g., OncotypeDx) is particularly useful for BRCA2-type (ER+, HER2-) cancers to determine chemotherapy benefit
Exam Intelligence
| Question Type | Pattern | Key Points |
|---|---|---|
| MCQ – IHC 3+ next step | "IHC score 3+, what is the next step?" | Start trastuzumab (NOT FISH — FISH is for equivocal 2+ only) [3][4] |
| MCQ – IHC 2+ next step | "IHC score 2+, what is the next step?" | Perform HER2 ISH (FISH/DISH) |
| MCQ – Herceptin side effect | "Most commonly known side effect of trastuzumab?" | Cardiomyopathy [5] |
| SAQ – Testing pathway | "Describe the pathway for HER2 testing" | IHC first → 0/1+ negative, 3+ positive, 2+ → reflex ISH |
| SAQ – FISH interpretation | "HER2/CEP17 ratio = 2.9, avg HER2 = 7.5" | Amplified (ratio ≥ 2.0 AND avg ≥ 4.0 = Group 1 positive) [3] |
| SAQ – OncotypeDx utility | "How does OncotypeDx help management?" | Stratifies recurrence risk; identifies patients who can avoid chemotherapy |
| Case study – Material | "What material should be used?" | FFPE tissue of the invasive component |
Common Traps
- Trap: After IHC 3+, choosing "Perform FISH" — Wrong. IHC 3+ is definitively HER2-positive. Start anti-HER2 therapy. FISH is only for equivocal (2+).
- Trap: Testing DCIS for HER2 to guide Herceptin — Wrong. Must test the INVASIVE component.
- Trap: Confusing prognostic vs predictive — ER+ is predictive of hormonal therapy response; HER2+ is predictive of anti-HER2 therapy response. Both are also prognostic.
- Trap: Thinking all HER2-negative patients get no anti-HER2 therapy — Wrong in 2023. HER2-low (IHC 1+ or 2+/non-amplified) may be eligible for T-DXd in metastatic setting.
- Trap: MammaPrint uses FFPE tissue — Originally based on fresh frozen tissue (though newer versions can use FFPE). OncotypeDx, PAM50, EndoPredict use FFPE.
- Trap: Giving Herceptin concurrently with anthracyclines — Wrong. Both are cardiotoxic. Sequential use only.
| Paper | Question | Topic | Key Answer |
|---|---|---|---|
| AOS Pathology [3] | MCQ | "IHC 3+ in > 10% cells, next action?" | D. Start trastuzumab treatment |
| AOS Pathology [3] | SAQ | "Predictive marker, technique, material, equivocal, further test, interpretation" | HER2 IHC on FFPE → equivocal → FISH → ratio 2.9, copy 7.5 = amplified |
| M26 MCQ Q18 [4] | MCQ | "Grade 3 IDC, LN+, ER/PR negative, HER2 3+. Next step?" | D. Start Herceptin |
| M26 MCQ Q63 [5] | MCQ | "Side effect of trastuzumab?" | A. Cardiomyopathy |
| 2024 MCQ Q70 [7] | MCQ | "Triple negative stage III breast CA with brain mets" | D. WBRT + steroid (no hormonal/targeted therapy options) |
| M26 MCQ Q91 [5] | MCQ | "Advantage of neoadjuvant systemic treatment?" | B. Early control of subclinical micrometastatic disease |
Q1 (MCQ-style): A 58-year-old lady has grade 3 invasive ductal carcinoma with LN metastases. IHC: ER negative, PR negative, HER2 3+. What is the most appropriate next step? → Start trastuzumab (Herceptin) treatment — IHC 3+ is definitively HER2 positive; no need for FISH.
Q2 (SAQ-style): A patient has HER2 IHC score 2+. What test should be done next, on what material, and how is the result interpreted? → HER2 ISH (FISH or DISH) on FFPE invasive breast cancer tissue. Count HER2 and CEP17 signals in 20 nuclei. Amplified if HER2/CEP17 ratio ≥ 2.0 or average HER2 copy number ≥ 6.0/cell.
Q3 (SAQ-style): What is OncotypeDx and how does it guide management of ER+, node-negative breast cancer? → 21-gene assay on FFPE tissue. Generates Recurrence Score. RS < 18 = low risk, endocrine therapy alone sufficient. RS > 31 = high risk, benefit from adding chemotherapy to endocrine therapy.
Q4 (MCQ-style): What is the most commonly known side effect of trastuzumab? → Cardiomyopathy
Q5 (SAQ-style): Define HER2-low breast cancer and its clinical significance. → IHC 1+ or IHC 2+ non-amplified. In the metastatic setting, these patients may be eligible for trastuzumab deruxtecan (T-DXd), an antibody-drug conjugate active against cancers without HER2 overexpression/amplification.
High Yield Summary
- HER2 gene amplification is the primary mechanism for HER2 protein overexpression, found in 20-25% of breast cancers
- All invasive breast carcinomas must be tested for HER2; testing must be on the invasive component using FFPE tissue
- IHC is first-line: 0/1+ = negative; 2+ = equivocal (reflex to ISH); 3+ = positive
- ISH (FISH/DISH): HER2/CEP17 ratio ≥ 2.0 or average HER2 ≥ 6.0/cell = amplified (with concurrent IHC requirement in grey-zone groups per 2018 guidelines)
- Trastuzumab (Herceptin) reduces recurrence by 50% and mortality by 33% in HER2+ early-stage breast cancer; key side effect = cardiotoxicity
- HER2-low (IHC 1+ or 2+/non-amplified) is a new category; eligible for T-DXd in metastatic setting
- Gene expression profiling (OncotypeDx, MammaPrint, PAM50, EndoPredict) identifies patients who can avoid chemotherapy
- OncotypeDx RS < 18: endocrine therapy alone sufficient; RS > 31: add chemotherapy for 28% absolute benefit
- Proliferation genes are the common driving force in all prognostic signatures
- Tissue handling: ≤1hr cold ischaemia, NBF fixation, 6-72 hours fixation time
Active Recall - Lecture Notes
[1] Lecture slides: MBBS IV Molecuar genetic testing in breast cancer HER2.pdf (all pages/sections) [2] Senior notes: Maksim Surgery Notes.pdf (pp. 182-186, breast malignancy and adjuvant therapy) [3] AOS material: AOS - Pathology.pdf (pp. 32, 43 — HER2 testing MCQ and SAQ) [4] Past papers: M26 4th Summative MCQ.pdf (Q18 — HER2 3+ next step) [5] Past papers: M26 4th Summative MCQ.pdf (Q63 — trastuzumab side effect; Q91 — neoadjuvant advantage) [6] GC lecture slides: GC 156. Many of my family members have cancers Cancer genetics and cytogenetics (File 2).pdf (pp. 14, 18 — BRCA1/2 histologic subtypes and molecular types) [7] Past papers: 2024 Fourth Summative MCQ.pdf (Q70 — triple negative breast cancer management)