Effect of fixation time on breast biomarker expression: a controlled study using cell line-derived xenografted (CDX) tumours

Kao, Kenneth R.; Hasan, Tahrim; Baptista, Armando; Truong, Thu H.; Gai, Luis; Smith, Adam C.; Li, S; Gonzales, Patrick R.; Voisey, Kim; Eriwvo, P; Power, J; Denic, Nash

doi:10.1136/jclinpath-2017-204381

Cited by 6 publications

(4 citation statements)

References 17 publications

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“…Over-fixation, on the other hand, is less likely to significantly alter the HER2 status. Immunohistochemical studies examining HER2 expression in breast carcinoma specimens and cell line-derived xenografts showed no difference in protein expression with fixation past 72 hr (54,55).…”

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confidence: 93%

Does Specimen Type Have an Impact on HER2 Status in Endometrial Serous Carcinoma? Discordant HER2 Status of Paired Endometrial Biopsy and Hysterectomy Specimens in the Presence of Frequent Intratumoral Heterogeneity

Rottmann

Assem

Matsumoto

et al. 2020

International Journal of Gynecological Pathology

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A recent clinical trial showed prolonged progression-free survival in human epidermal growth factor receptor 2 (HER2)-positive advanced stage and recurrent endometrial serous carcinomas when trastuzumab was added to traditional chemotherapy. Approximately one third of these tumors are HER2-positive and have been described to show unique characteristics of HER2 protein expression and gene amplification, including significant intratumoral heterogeneity, in recent studies. However, currently, there are no standard protocols for the selection of optimal specimen type or algorithm for HER2 testing in endometrial serous carcinomas. The current study aimed to evaluate the concordance of HER2 status between endometrial biopsy/curettage and subsequent hysterectomy specimens in endometrial serous carcinoma. A total of 57 patients with endometrial serous carcinoma with available HER2 status were identified during the study period, 14 of which (14/57, 25%) were HER2-positive by immunohistochemistry and/or fluorescent in situ hybridization (FISH). The final study cohort consisted of 40 paired endometrial biopsies/curettings and hysterectomies to include all 14 HER2-positive tumors and 26 selected HER2-negative tumors to represent an equal distribution of HER2 immunohistochemical scores. HER2 FISH was performed on all tumors with an immunohistochemical score of 2+. HER2 immunohistochemical scores, heterogeneity of HER2 expression, FISH results, and the overall HER2 status were compared between the 2 specimen types. HER2 status was successfully assigned in both specimen types in 37 cases, as three specimens showed inadequate FISH signals. Concordant HER2 status was observed in 84% of cases (31/37), with identical HER2 immunohistochemical scores in 65% (26/40) of tumors. Among the 6 tumors with a discordant HER2 status, 2 were HER2 negative in the biopsy and positive in the hysterectomy, and 4 were HER2-positive in the biopsy and negative in the hysterectomy. The false-negative rate would be 15.4% and 26.7% if only the biopsy or only the hysterectomy would be the basis for the result, respectively. Intratumoral heterogeneity of HER2 protein expression was present in 22 tumors (55%), including all cases with a discordant HER2 status. The concordance rate of HER2 status between paired endometrial biopsies/curettings and hysterectomies of endometrial serous carcinoma is lower than the reported rates of breast cancer, and comparable to those of gastric carcinomas. Frequent heterogeneity of HER2 protein expression combined with the possibility of a spatially more heterogenous sampling of endometrial cavity in biopsies and curettings, and the potential differences in specimen handling/fixation between the 2 specimen types may explain our findings. HER2 testing of multiple specimens may help identify a greater proportion of patients eligible for targeted trastuzumab therapy and should be taken into account in future efforts of developing endometrial cancer-specific HER2 testing algorithm.

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confidence: 93%

Does Specimen Type Have an Impact on HER2 Status in Endometrial Serous Carcinoma? Discordant HER2 Status of Paired Endometrial Biopsy and Hysterectomy Specimens in the Presence of Frequent Intratumoral Heterogeneity

Rottmann

Assem

Matsumoto

et al. 2020

International Journal of Gynecological Pathology

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“…It is important to realize that there are differences in the nature and quantity of DNA DSBs and the size of the RAD51 foci between the endogenous (RAD51-FFPE) and radiation-induced (RECAP) DNA damage. In addition, it cannot be ruled out that time-to-fixation might play a role as has previously been described for HER2 assessment [ 44 , 45 ]. While the fixation period is exactly two hours for all RECAP samples, it varies considerably for the RAD51-FFPE samples due to their diagnostic nature.…”

Section: Discussionmentioning

confidence: 99%

Performance of a RAD51-based functional HRD test on paraffin-embedded breast cancer tissue

van Wijk,

Vermeulen,

ter Haar

et al. 2023

Breast Cancer Res Treat

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Purpose BRCA-deficient breast cancers (BC) are highly sensitive to platinum-based chemotherapy and PARP inhibitors due to their deficiency in the homologous recombination (HR) pathway. However, HR deficiency (HRD) extends beyond BRCA-associated BC, highlighting the need for a sensitive method to enrich for HRD tumors in an alternative way. A promising approach is the use of functional HRD tests which evaluate the HR capability of tumor cells by measuring RAD51 protein accumulation at DNA damage sites. This study aims to evaluate the performance of a functional RAD51-based HRD test for the identification of HRD BC. Methods The functional HR status of 63 diagnostic formalin-fixed paraffin-embedded (FFPE) BC samples was determined by applying the RAD51-FFPE test. Samples were screened for the presence of (epi)genetic defects in HR and matching tumor samples were analyzed with the RECAP test, which requires ex vivo irradiated fresh tumor tissue on the premise that the HRD status as determined by the RECAP test faithfully represented the functional HR status. Results The RAD51-FFPE test identified 23 (37%) of the tumors as HRD, including three tumors with pathogenic variants in BRCA1/2. The RAD51-FFPE test showed a sensitivity of 88% and a specificity of 76% in determining the HR-class as defined by the RECAP test. Conclusion Given its high sensitivity and compatibility with FFPE samples, the RAD51-FFPE test holds great potential to enrich for HRD tumors, including those associated with BRCA-deficiency. This potential extends to situations where DNA-based testing may be challenging or not easily accessible in routine clinical practice. This is particularly important considering the potential implications for treatment decisions and patient stratification.

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“…Freshly sectioned slides (4 μm on positively charged slides) from formalin-fixed paraffin-embedded blocks were processed as previously described (17). The only modifications made to the previously published protocol were that pepsin was varied between 40 and 45 min depending on tissue requirements.…”

Section: Fluorescence In Situ Hybridizationmentioning

confidence: 99%

HER2-low and Overexpression in Mucinous Ovarian Cancer: Analysis of ASCO/CAP and ToGA Immunohistochemical Scoring

Han,

Madariaga,

Gonzalez-Ochoa

et al. 2024

International Journal of Gynecological Pathology

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Mucinous ovarian carcinoma is an uncommon malignancy characterized by resistance to chemotherapy and poor survival in the metastatic setting. HER2 amplification is a frequent late event in carcinogenesis, yet the incidence of HER2-low in mucinous ovarian carcinoma is unknown. Further, the optimal method for determining overexpression in these tumors is not established. We sought to assess the ASCO/CAP and ToGA trial scoring methods for HER2 IHC with correlation to FISH, p53, and mismatch repair protein status and to determine the incidence of HER2-low in mucinous ovarian carcinoma. A total of 29 tumors from 23 patients were included. Immunohistochemistry for HER2, p53, MLH1, PMS2, MSH2, and MSH6 was performed. Scoring was performed according to the ASCO/CAP and ToGA trial criteria. HER2 FISH was performed and scored according to the ASCO/CAP criteria. The proportion of HER2-low, defined as 1+ or 2+ staining with negative FISH, was determined. Using ASCO/CAP, 26% demonstrated 3+ while 35% demonstrated 2+ staining. Using ToGA, 30% demonstrated 3+ while 57% demonstrated 2+ staining. By FISH, 26% were positive for HER2 amplification. Both systems captured all FISH-positive cases; the use of ASCO/CAP resulted in fewer equivocal and false-positive cases. Among HER2-negative cases, 88% were HER2-low. Aberrant p53 expression was detected in 55% of cases; mismatch repair deficiency was not identified in any cases. ASCO/CAP guidelines are accurate and resource-effective in determining HER2 overexpression in mucinous ovarian carcinoma. HER2-low is common in these tumors; further studies to determine the role of HER2-targeted therapy including antibody-drug conjugates are indicated.

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Effect of fixation time on breast biomarker expression: a controlled study using cell line-derived xenografted (CDX) tumours

Abstract: Neither shortened nor prolonged fixation affects ER expression, consistent with previous findings. In equivocal Her2-expressing tumours, however, increasing fixation increased the sensitivity of Her2 IHC reporting while not affecting FISH.

Cited by 6 publications

References 17 publications

Does Specimen Type Have an Impact on HER2 Status in Endometrial Serous Carcinoma? Discordant HER2 Status of Paired Endometrial Biopsy and Hysterectomy Specimens in the Presence of Frequent Intratumoral Heterogeneity

Does Specimen Type Have an Impact on HER2 Status in Endometrial Serous Carcinoma? Discordant HER2 Status of Paired Endometrial Biopsy and Hysterectomy Specimens in the Presence of Frequent Intratumoral Heterogeneity

Performance of a RAD51-based functional HRD test on paraffin-embedded breast cancer tissue

HER2-low and Overexpression in Mucinous Ovarian Cancer: Analysis of ASCO/CAP and ToGA Immunohistochemical Scoring

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