Efforts to Improve HER2 Testing Continue ASCO Panelist Sees Progress

Antonio C. Wolff, MD

The movement to improve the accuracy and predictive value of tumor marker testing in patients with breast cancer has resulted in a dramatic upswing in the number of accredited laboratories, as well as a continuing effort to clarify definitions that guide therapy decisions.

Antonio C. Wolff, MD, who has helped lead such efforts, reviewed several of the key controversies that have surrounded guidelines research. He is an associate professor of Oncology and a research scientist in the Breast Cancer Program at the Sidney Kimmel Comprehensive Cancer Center at Johns Hopkins Hospital, Baltimore, Maryland.

Wolff co-chaired the cooperative panel of the American Society of Clinical Oncology (ASCO) and the College of American Pathologists (CAP) that issued guidelines in 2007 for testing for the human epidermal growth factor receptor 2 (HER2) in invasive breast cancer. The marker guides decisions for trastuzumab (Herceptin) and lapatinib (Tykerb), and possibly in other therapies.

“We wanted to standardize methodology, making sure that the same test done in the same tissue with the same patient anywhere would as much as possible yield the same information,” Wolff said. “The panel never set out to change the eligibility for trastuzumab. This has been a huge source of confusion.”

Table 1. Key Points in 2011 Clinical Notice

HER2

HER2 ER and PgR

Cold ischemic time

Changed from "short" to ≤1 hour

Interval ≤1 hour

Document time between tissue removal and start of fixation

Fixation time in neutral buffered formalin

6-48 hours in NBF; less fixation time permissible for needle biopsy specimens

6-72 hours in NBF for all specimens

Optimal sample for testing

Resection specimens recommended

Core needle biopsies recommended

Pathologist discretion in selecting sample for testing

ER indicates estrogen receptor; HER2, human epidermal growth factor receptor 2; NBF, neutral buffered formalin; PgR, progesterone receptor.

ASCO Guidelines Clinical Notice, American Society of Clinical Oncology Website. http://goo.gl/pVdGb. Updated January 13, 2011. Accessed August 22, 2011.

In 2005, researchers believed that up to approximately 20% of all HER2 testing worldwide yielded false positives. Wolff said the picture for false negatives was not as clear, with estimates ranging from 5% to 10%.

While researchers set out to compare results from local laboratories with central facilities, the issues turned on broader questions of testing protocols. “It’s not an issue of whether the test was done centrally or locally, but whether the test was done correctly,” Wolff said.

Today, there are indications that the frequency of HER2 false positives has dropped to less than 6% in the United States, Wolff said in an interview.

Meanwhile, the number of HER2- testing laboratories participating in the CAP accreditation program has risen markedly for both immunohistochemistry (IHC) and fluorescence in situ hybridization (FISH) methods. There were 1120 IHC and 316 FISH laboratories participating in the program this year, up from 125 IHC and 174 FISH facilities in 2004, according to a CAP survey. Likewise, more laboratories are becoming accredited for estrogen receptor (ER) testing, with the numbers rising from 97 in 2004 to 1200 this year.

There are about 1500 laboratories doing HER2 testing in the United States, Wolff said.

The joint guidelines panel recommended tissue-handling guidelines for HER2 testing in 2007, with a key factor being a gap between gathering of a specimen at a hospital and testing the sample in a laboratory (J Clin Oncol. 2007;25(1):118-145).

“

It’s not an issue of which is the right test to do—IHC or FISH— but whether you’re doing the test right. ”

—Antonio C. Wolff, MD

Although clinicians and administrators at each point were following protocols, more coordination was necessary, Wolff said. “We are all responsible,” he said. “The enemy ultimately was us.”

The stakes for an accurate determination of HER2 status have grown increasingly higher, Wolff noted. “We’re not talking about palliation any more,” he said. “We’re talking about literally life and death for women with curable disease in the adjuvant setting.”

Better Standards for Hormone Testing

In 2010, the joint panel revisited the HER2 guidelines and also reviewed procedures for ER and progesterone receptor (PgR) testing. In January, ASCO issued a Clinical Notice summarizing recommendations of both the 2007 and 2010 panels.

“Because the same tissue coming out of the same patient will be used both for ER and for HER2 testing, we felt that it was important to reconcile and to harmonize some of the original recommendations made for HER2,” Wolff said in an interview.

Reports indicate that while HER2 testing has improved, there’s a long way to go in ER and PgR testing. As many as 20% of ER and PgR test results performed through IHC methods worldwide may yield either false positives or false negatives, chiefl y because of variations in preanalytic factors, thresholds for positivity, and interpretation criteria, the panel said (J Clin Oncol. 2010;28(16):2784-2795).

In the United States, there has been no standardized proficiency testing of IHC procedures for ER and PgR markers, although IHC became the predominant ER testng method in the 1990s.

Table 2. HER2 Testing Algorithms

FISH¹

HER2 gene amplification

IHC²

HER2 protein expression

Positive

FISH ratio >2.2

or HER2 gene copy >6.0

IHC 3 (uniform intense membrane staining in >30% of invasive tumor cells)

Negative

FISH ratio <1.8 or HER2 gene copy <4.0

IHC 0 or 1

*Equivocal

FISH ratio 1.8-2.2 or HER2 gene copy 4.0-6.0

IHC 2 (patients with uniform intense membrane staining in >10% and <30% of cells eligible for adjuvant trastuzumab trials)

Count additional cells for FISH, retest, or test with HER2 IHC

Test with assay for HER2 gene amplification

*Patients with HER2/CEP17 ratio ≥2.0 eligible for adjuvant trastuzumab trials.

FISH indicates fluorescence in situ hybridization; HER2, human epidermal growth factor receptor 2; IHC, immunohistochemistry.

1. Wolff AC, Hammond ME, Schwartz JN, et al. American Society of Clinical Oncology/College of American Pathologists guideline recommendations for human epidermal growth factor receptor 2 testing in breast cancer. J Clin Oncol. 2007;25(1):118-145.

2. Wolff AC, Hammond ME, Schwartz JN, Hayes DF. On behalf of the ASCO/CAP Panel on HER-2 Testing in Breast Cancer [correspondence]. J Clin Oncol. 2007;25(25):4021-4023. doi: 10.1200/JCO.2007.11.4363.

Studies in other countries, notably a quality assurance program in the United Kingdom, suggest significant variations and inaccurate results. An inquiry in the Canadian province of Newfoundland indicated that approximately one-third of 1023 ER tests conducted between 1997 and 2005 were scored falsely negative when compared with retested results from a central laboratory.

One key point to emerge from both panels is the optimal method for clinicians to use for collecting samples.

For HER2, the guidelines recommend that resection samples be preferentially used rather than core needle biopsies “because of the likelihood of artifacts that might confound interpretation.”

For ER and PgR testing, core needle biopsies are recommended “because of higher likelihood that the samples will be quickly fixed.”

Treatment Algorithms Clarified

Although the 2007 panel’s intention was to focus on improving testing accuracy, debate soon erupted over the appropriate algorithms for defining HER2- positive disease, and thus whether a patient would be a candidate for trastuzumab therapy.

Wolff said the panel was seeking simply to recommend that results be validated with IHC or FISH methodology if they fell into an equivocal category the first time the samples were tested.

In April, Wolff helped author a Clinical Notice letter that sought to clarify the issue. One figure already had been revised “to state that patients with a HER2-positive equivocal IHC test who had tumors with uniform intense membrane staining in more than 10% and less than 30% of cells were also eligible for the trastuzumab trials,” according to the notice. (published online ahead of print April 18, 2011. J Clin Oncol. doi/10.1200/JCO.2011.35.2245).

“The panel did not recommend withholding anti- HER2 treatment in those patients with an equivocal HER2 test (or tests) whose results fell within ranges that would have allowed them to be treated in the first generation of adjuvant HER2-targeted trials,” the document said.

In addition, Wolff is not recommending one test over the other. “It’s not an issue of which is the right test to do—IHC or FISH—but whether you’re doing the test right because both tests can be equally good in identifying patients who will benefit from therapy,” he said.

Discussions over HER2 guidelines are far from settled. Wolff said the panel will be re-established and that an updated guideline likely would be released next year.

He also noted that the focus on testing would be far-reaching.

“The attention to detail will go a long way toward improving the performance of these tests,” he said. “These issues do transcend beyond breast cancer. As we have more and more targeted therapies for various diseases, we need to have assays that are as good as the treatment that we give.”