Identification of HER-2

The evaluation of HER-2 is recommended in all cases of breast cancer at the time of initial diagnosis and / or at the time of recurrence to guide therapy
A number of methods are used for HER-2 testing including:
- Immunohistochemistry (IHC)
- Florescence in situ hybridization (FISH)
- Enzyme-linked immunosorbent assay (ELISA) analysis of serum and tumor
- Western blot
- Southern blot
- Chromogenic in situ hybridization (CISH)
- Silver in situ hybridization (SISH)
- Polymerase chain reaction (PCR)
IHC and FISH:
- Are currently the main testing modalities for HER-2-positive breast cancer
HER-2 expression in breast cancer:
- Is primarily assessed semiquantitatively by IHC
The 2013 American Society of Clinical Oncology (ASCO) and the College of American Pathologists (CAP) guidelines:
- Outline an IHC scoring method based on four classes:
  - 0, 1+, 2+, and 3+
- A score of 0 is negative:
  - Indicating no observed staining in invasive tumor cells
- A score of 1+ is negative:
  - Indicates:
    - Weak, incomplete membrane staining in any proportion of invasive tumor cells or
    - Weak, complete membrane staining in less than 10% of invasive tumor cells
- A score of 2+ is equivocal:
  - Indicating:
    - Circumferential membrane staining that is incomplete and / or
    - Weak / moderate and in more than 10% of invasive tumor cells or
    - Complete and circumferential membrane staining that is intense and in 10% or less of invasive tumor cells
  - All 2+ equivocal cases undergo subsequent testing by FISH
- A score of 3+ is positive:
  - Indicates circumferential membrane staining that is complete and intense in a homogeneous and contiguous population, present in more than 10% of invasive tumor cells, and readily appreciated using a low-power objective
FISH:
- Is a sensitive and accurate method of scoring invasive breast tumor tissue for HER-2 expression
- Initial gene amplification studies by FISH assessment:
  - Used chromosome 17 centromere (CEP17) or another gene on the same chromosome as an internal control:
    - With a ratio of 2.0 or greater:
      - Considered evidence of HER-2 amplification
- These criteria were used as the cutoff for enrollment in trials evaluating HER-2 targeted therapies
In 2007, the ASCO / CAP guidelines were changed to define HER-2 amplified:
- As a ratio of 2.2 or greater
More recent guidelines:
- Have changed the ratio cutoff back to a ratio of 2.0 or greater with the inclusion of criteria to account for HER-2 copy number per tumor cell
- Based on the recent guidelines, HER-2 is amplified in cases where the:
  - HER-2 / CEP17 ratio is 2.0 or greater with an average HER-2 copy number of more than 4.0 signals/cell or
  - HER-2 / CEP17 ratio is less than 2.0 with an average HER-2 copy number of 6.0 or more signals / cell using a dual probe or a HER-2 copy number of 6.0 or more copies/cell using a single probe
- FISH testing is negative for HER-2 amplification with a:
  - HER-2 / CEP17 ratio of less than 2.0 with an average HER-2 copy number of less than 4.0 signals/cell or an average HER-2 copy number of less than 4.0 signals / cell using a single probe