HER2-Low Expression is the Tip of the Iceberg in Breast Cancer Classification

The advent of antibody-drug conjugates has placed a renewed focus on the lesser thought of HER2 classifications of IHC 1+ and 0.

Jorge S. Reis-Filho, MD, PhD, FRCPath

Jorge S. Reis-Filho, MD, PhD, FRCPath

For the past several decades, pathologists have worked to develop optimal methods to distinguish HER2 expressors via immunohistochemistry (IHC) assay on the higher end of the spectrum (ie, 2+ and 3+). However, the advent of antibody-drug conjugates (ADCs) has placed a renewed focus on the lesser thought of HER2 classifications of IHC 1+ and 0, according to Jorge S. Reis-Filho, MD, PhD, FRCPath.1

“We are facing real challenges in terms of [HER2] identification in the clinic, and I would contend that we are in a state of flux in terms of the identification,” said Reis-Filho, chief of experimental pathology and director of the Experimental Pathology Fellowship Program at Memorial Sloan Kettering Cancer Center in New York, New York, during a presentation at the 40th Annual Miami Breast Cancer Conference®.

HER2 amplification is present in 11% to 15% of all invasive breast cancers and can result in massive overexpression of HER2 cell membranes and/or adapt to HER2 signaling, which promotes cell proliferation via the RAS/MAPK pathway.1,2

“The reason [investigators] sought to identify the 2 positive [classifications (2+ and 3+)] was because several clinical trials illustrated the efficacy of the humanized monoclonal antibody trastuzumab [Herceptin] for patients who have HER2-positive disease,” Reis-Filho explained in a retrospective look at HER2 identification in breast cancer. With consistent improvements in overall survival reported over the course of the past decade, Reis-Filho noted, “not surprisingly, we have seen a proliferation of therapies targeting HER2, including additional monoclonal antibodies, [such as] pertuzumab [Perjeta] and margetuximab-cmkb [Margenza], and bispecific antibodies…. But I think the agents that have continually elicited the most excitement over the past few years is the development of ADCs.”

Using agents such as ado-trastuzumab deruxtecan (Kadcyla; T-DM1) and fam-trastuzumab deruxtecan-nxki (Enhertu) work to deliver the cytotoxic payloads leveraging the construct of the antibody with the linker mechanism. “The concept here is different. We are not trying to silence the pathways activated by HER2. We’re using HER2 as a homing device, so the antibody should find the cancer cells so that they internalize the toxic payload.”

That has led to new requirements in divergent testing, Reis-Filho said. Rather than finding the HER2-amplified cancers, “we have to identify all cancers that express HER2 because just the HER2 expression would be sufficient enough, in theory, to be able to get the ADC to the cancer cells.”

With this new level of distinction, Reis-Filho said that this group of tumors now comprise up to 60% of breast cancers. The pivotal phase 3 trial spotlighting the role of ADCs in HER2-low expressors was DESTINY-Breast04 (NCT03734029).3 Among patients with hormone receptor–positive, HER2-low breast cancer—defined as IHC 1+ or 2+ with a negative in situ hybridization (ISH) test—trastuzumab deruxtecan elicited a median PFS of 10.1 months (95% CI, 9.5-11.5) compared with 5.4 months (95% CI, 4.4-7.1) for investigator’s choice of chemotherapy (HR, 0.51; 95% CI, 0.40-0.64; P < .001). The median overall survival (OS) was 23.9 months (95% CI, 20.8-24.8) vs 17.5 months (95% CI, 15.2-22.4), respectively (HR, 0.64; 95% CI, 0.40-0.86; P = .003).3

The FDA approved trastuzumab deruxtecan for patients with HER2-low breast cancer in August 2022.4

Reis-Filho noted that the trial used the American Society of Clinical Oncology (ASCO)/College of American Pathologists (CAP) guidelines for HER2 threshhold.5 In terms of HER2 mRNA expression, Reis-Filho noted that using the guideline definition, there was a substantial overlap in expression levels according to an analysis using IHC for those with IHC 0 and 1+.6

Despite most HER2-low breast cancers being estrogen receptor (ER)–positive and luminal A or B subtype, there were still instances of triple-negative disease as well as basal-like cancers identified in studies.6 “That speaks to our imprecise definition of HER2 0 and HER2 1+,” Reis-Filho said. “From a molecular standpoint, [HER2-low] does not seem to constitute a discrete entity at all. Actually, there is a great deal of heterogeneity.”

In an analysis of HER2-low breast cancer investigators found that at least 4 different molecular subtypes may exist independent of PAM50 subtypes in these tumors—group A (lymphocyte activated), group B (unique HER2 gain), group C (tumor stroma remodeling), and group D (actionable PIK3CA).7 However,from a genomics perspective, several studies have shown that there are negligible differences in the incidence of oncogenic mutations after adjusting for ER.8

“Consistent with this notion that from a pathologic and molecular perspective HER2-low breast cancers may not be the discrete entity, clinical studies have shown that there is no consistency in terms of the impact of HER2-low status on outcomes for patients with breast cancer,” Reis-Filho said. “In a comparison with HER2 0 breast cancers, HER2-low breast cancers do not seem to have a different overall survival even when stratifies according to hormone status.”9

Ongoing HER2 Classification Challenges

An “incredible” challenge exists from a pathologist standpoint, Reis-Filho noted, because HER2 status may not be stable throughout the course of the disease. Studies comparing tissue samples from primary and recurrent disease have shown a discordance of HER2 direction that is bidirectional. Approximately 14% of patients with HER2-low disease changed to HER2 0 status and up to 15% changed from HER2 0 status to HER2 low in samples from the same patients. Similar trends were observed among patients pre and post neoadjuvant therapy.10

“It’s very important [to note] that the stability of the HER2-low phenotype can be very modest and that these discordances can be bidirectional, and can affect up to 50% of patients,” Reis-Filho said. “We have a real challenge…because we now have to divide breast cancers into 3 groups: the HER2-positive group that we treat with the current available [options] for HER2-positive disease; the HER2-low patients who receive trastuzumab deruxtecan; and [the group with] the HER2 0 breast cancers who receive therapies other than those.”

Data from the phase 2 DAISY trial (NCT04132960) of trastuzumab deruxtecan in patients with HER2 0, showed that the ADC may have efficacy to support its use in this population. The overall response rate among 37 patients with IHC 0+ status was 29.7% (95% CI, 15.9%-47.0%).11

The advent of a new classification has emerged with the initiation of the phase 3 DESTINY-Breast06 trial (NCT04494425) for patients with ultra-low HER2 expression, defined as IHC greater than 0 but less than 1+.

“In the past 2 or 3 years, we had to identify HER2 positives and negatives. Then, after investigators presented DESTINY-Breast03, we had to identify the HER2 lows, the positives, and the negatives. Now, with DESTINY-Breast06, we need to identify the ultra lows, as well,” Reis-Filho said, citing a cycle of HER2 reclassification set out by Andre et al.12 “But I think where we are going to land is that there will be the HER2-positive breast cancers—the 2+ and 3+—and, for the others, we need to provide some form of granular quantification of HER2 to help us prioritize which ADC we are using for each patient population.”

Editor’s Note: Reis-Filho has the following disclosures: scientific advisory board with paid honoraria from Paige.AI, REPARE Therapeutics, Personalis, Roche Tissue Diagnostics, Daiichi Sankyo and Merck. He is a member of board of directors at Oncoclinicas and a consultant for Paige.AI, Goldman Sachs, Bain Capital, REPARE Therapeutics and SAGA Diagnostics.


  1. Reis-Filho JS. Real-world treatment patterns and effectiveness of palbociclib plus an aromatase inhibitor in patients with metastatic breast cancer aged 75 years or above. Presented at: 40th Annual Miami Breast Cancer Conference®; March 2-5, 2023; Miami Beach, FL.
  2. Hudis CA. Trastuzumab--mechanism of action and use in clinical practice. N Engl J Med. 2007;357(1):39-51. doi:10.1056/NEJMra043186
  3. Modi S, Jacot W, Yamashita T, et al; DESTINY-Breast04 Investigators. Trastuzumab deruxtecan in previously treated HER2-low advanced breast cancer. N Engl J Med. 2022;387(1):9-20. doi:10.1056/NEJMoa2203690
  4. FDA approves fam-trastuzumab deruxtecan-nxki for HER2-low breast cancer. FDA. August 5, 2023. Accessed March 3, 2023. bit.ly/3zGfOPt
  5. Wolff AC, Hammond MEH, Hicks DG, et al; American Society of Clinical Oncology; College of American Pathologists. Recommendations for human epidermal growth factor receptor 2 testing in breast cancer: American Society of Clinical Oncology/College of American Pathologists clinical practice guideline update. J Clin Oncol. 2013;31(31):3997-4013. doi:10.1200/JCO.2013.50.9984
  6. Schettini F, Chic N, Brasó-Maristany F, et al. Clinical, pathological, and PAM50 gene expression features of HER2-low breast cancer. NPJ Breast Cancer. 2021;7(1):1. doi:10.1038/s41523-020-00208-2
  7. Berrino E, Annaratone L, Bellomo SE, et al. Integrative genomic and transcriptomic analyses illuminate the ontology of HER2-low breast carcinomas. Genome Med. 2022;14(1):98. doi:10.1186/s13073-022-01104-z
  8. Tarantino P, Gupta HV, Hughes ME, et al. Integrative genomic and transcriptomic analyses illuminate the ontology of HER2-low breast carcinomas. Presented at: 2022 San Antonio Breast Cancer Symposium; December 6-10, 2022; San Antonio, TX.
  9. Peiffer D, Howard FM, Chen N, et al. Integrative genomic and transcriptomic analyses illuminate the ontology of HER2-low breast carcinomas. Presented at: 2022 San Antonio Breast Cancer Symposium; December 6-10, 2022; San Antonio, TX.
  10. Miglietta F, Griguolo G, Bottosso M, et al. Evolution of HER2-low expression from primary to recurrent breast cancer. NPJ Breast Cancer. 2021;7(1):137. doi:10.1038/s41523-021-00343-4
  11. Diéras V, Deluche E, Lusque A, et al. Trastuzumab deruxtecan (T-DXd) for advanced breast cancer patients (ABC), regardless HER2 status: a phase II study with biomarkers analysis (DAISY). Clin Cancer Res. 2022;82(suppl 4):PD8-02. doi:10.1158/1538-7445.SABCS21-PD8-02
  12. Andre F, Fernanda M, Deluche E, et al. Mechanism of action and resistance to trastuzumab deruxtecan in patients with metastatic breast cancer: the DAISY trial. Research Square. September 26, 2022. doi:10.21203/rs.3.rs-2083650/v1
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