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The increasing importance of pathology in modern clinical trial conduct: OlympiA as a case in point

  • Lauren Kalinowski
    Affiliations
    The University of Queensland Centre for Clinical Research, University of Queensland, Brisbane, Qld, Australia

    Department of Histopathology, Sullivan Nicolaides Pathology, Bowen Hills, Brisbane, Qld, Australia
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  • Giuseppe Viale
    Affiliations
    European Institute of Oncology IRCCS, University of Milan, Milan, Italy
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  • Susan Domchek
    Affiliations
    Basser Center for BRCA, Abramson Cancer Center, University of Pennsylvania, Philadelphia, PA, USA
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  • Andrew Tutt
    Affiliations
    Breast Cancer Now Toby Robins Research Centre, The Institute of Cancer Research, London, UK

    Breast Cancer Now Unit, The School of Cancer Studies and Pharmaceutical Science, King's College London, London, UK
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  • Peter C. Lucas
    Affiliations
    University of Pittsburgh School of Medicine and UPMC Hillman Cancer Center, Pittsburgh, PA, USA
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  • Sunil R. Lakhani
    Correspondence
    Address for correspondence: Prof Sunil Lakhani, The University of Queensland Centre for Clinical Research, University of Queensland, Brisbane, Qld 4029, Australia.
    Affiliations
    The University of Queensland Centre for Clinical Research, University of Queensland, Brisbane, Qld, Australia

    Pathology Queensland, Royal Brisbane and Women's Hospital, Herston, Brisbane, Qld, Australia
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      Introduction

      Recently published results from the randomised phase III adjuvant therapy OlympiA trial are a significant development for patients with hereditary breast cancer in the era of precision medicine. OlympiA has shown adjuvant treatment with a poly (ADP-ribose) polymerase (PARP) inhibitor significantly improves outcomes in patients with germline BRCA-mutated early breast cancer.
      • Tutt A.N.J.
      • Garber J.E.
      • Kaufman B.
      • et al.
      Adjuvant olaparib for patients with BRCA1- or BRCA2-mutated breast cancer.
      ,
      • Tutt A.N.J.
      • Garber J.
      • Gelber R.D.
      • et al.
      Pre-specified event driven analysis of overall survival (OS) in the OlympiA phase III trial of adjuvant olaparib (OL) in germline BRCA1/2 mutation (gBRCAm) associated breast cancer. ESMO virtual plenary sessions.
      PARP inhibitors are already approved for use in Australia in patients with germline BRCA1/2 mutations that have either high stage, high grade ovarian carcinomas or metastatic HER2-negative breast cancer. The American Society of Clinical Onocology (ASCO), National Cancer Comprehensive Network (NCCN) and St Gallen Early Breast Cancer Consensus updated guidelines now also include treatment with olaparib to improve outcomes for patients with an inherited mutation in BRCA1 or BRCA2 with early stage, HER2-negative breast cancer who have a high recurrence risk based on the results of OlympiA.
      • Tutt A.N.J.
      • Garber J.E.
      • Kaufman B.
      • et al.
      Adjuvant olaparib for patients with BRCA1- or BRCA2-mutated breast cancer.
      ,
      • Tung N.M.
      • Zakalik D.
      • Somerfield M.R.
      Hereditary breast cancer guideline expert panel. Adjuvant PARP inhibitors in patients with high-risk early-stage HER2-negative breast cancer and germline BRCA mutations: ASCO hereditary breast cancer guideline rapid recommendation update.

      National Comprehensive Cancer Network. Breast cancer. Cited 11 Feb 2022. https://www.nccn.org/professionals/physician_gls/pdf/breast.pdf.

      • Burstein H.J.
      • Curigliano G.
      • Thurlimann B.
      • et al.
      Customizing local and systemic therapies for women with early breast cancer: the St. Gallen International Consensus Guidelines for treatment of early breast cancer 2021.
      The United States Food and Drug Administration (FDA) approved the PARP inhibitor olaparib in high recurrence risk early breast cancer patients with germline BRCA1/2 mutations in March 2022.

      US Food and Drug Administration. FDA approves olaparib for adjuvant treatment of high-risk early breast cancer. Updated 11 Mar 2022. https://www.fda.gov/drugs/resources-information-approved-drugs/fda-approves-olaparib-adjuvant-treatment-high-risk-early-breast-cancer.

      A large number of ongoing clinical trials are examining olaparib and other PARP inhibitors in early breast cancer. These recent advances are bringing PARP inhibitors into the clinic for early breast cancer patients. As more targeted therapies become available and new prognostic and predictive biomarkers emerge, the role that pathologists play in clinical trials and in changing clinical practice is becoming increasingly important.

      The BRCA journey

      The observation that breast cancer ran in some families and that some young women were particularly susceptible to the development of breast cancer led to the identification of BRCA1 and BRCA2 pathogenic variants in the mid-1990s.
      • Miki Y.
      • Swensen J.
      • Shattuck-Eidens D.
      • et al.
      A strong candidate for the breast and ovarian cancer susceptibility gene BRCA1.
      ,
      • Wooster R.
      • Bignell G.
      • Lancaster J.
      • et al.
      Identification of the breast cancer susceptibility gene BRCA2.
      Pathogenic germline loss of function mutations in these genes leads to a significantly higher risk of both breast and ovarian cancer. These variants are the most common cause of hereditary breast cancer and are identified in approximately 3% of breast cancer patients.
      • Hu C.
      • Hart S.N.
      • Gnanaolivu R.
      • et al.
      A population-based study of genes previously implicated in breast cancer.
      ,
      • Dorling L.
      • Carvalho S.
      • et al.
      Breast Cancer Association Consortium
      Breast cancer risk genes - association analysis in more than 113,000 women.
      BRCA1 and BRCA2 are tumour suppressor genes that are critically involved in cell cycle checkpoint activation, gene transcription regulation and DNA damage repair. Their discovery provided the rationale for familial cancer screening, the early diagnosis of disease and the implementation of preventative strategies for reducing an individual's risk of developing cancer.
      Pathologists have been instrumental in elucidating the morphological and phenotypic characteristics of BRCA1/2 tumours. BRCA1 tumours are typically high grade with pushing margins, highly proliferative, associated with a prominent lymphocytic infiltrate and are often triple-negative
      • Lakhani S.R.
      • Van De Vijver M.J.
      • Jacquemier J.
      • et al.
      The pathology of familial breast cancer: predictive value of immunohistochemical markers estrogen receptor, progesterone receptor, HER-2, and p53 in patients with mutations in BRCA1 and BRCA2.
      • Mavaddat N.
      • Barrowdale D.
      • Andrulis I.L.
      • et al.
      Pathology of breast and ovarian cancers among BRCA1 and BRCA2 mutation carriers: results from the Consortium of Investigators of Modifiers of BRCA1/2 (CIMBA).
      • Southey M.C.
      • Ramus S.J.
      • Dowty J.G.
      • et al.
      Morphological predictors of BRCA1 germline mutations in young women with breast cancer.
      • Spurdle A.B.
      • Couch F.J.
      • Parsons M.T.
      • et al.
      Refined histopathological predictors of BRCA1 and BRCA2 mutation status: a large-scale analysis of breast cancer characteristics from the BCAC, CIMBA, and ENIGMA consortia.
      (Fig. 1A,B). Similarly, BRCA2 tumours are typically high grade with pushing margins but are more likely to have a luminal oestrogen receptor-positive phenotype than BRCA1 mutated tumours.
      • Bane A.L.
      • Beck J.C.
      • Bleiweiss I.
      • et al.
      BRCA2 mutation-associated breast cancers exhibit a distinguishing phenotype based on morphology and molecular profiles from tissue microarrays.
      Fig. 1
      Fig. 1Triple-negative breast cancer from patient with BRCA1, highlighting pushing edge of the tumour, dense lymphoid infiltrate and high tumour grade (H&E; A, low power; B, higher power).
      Proteins encoded by BRCA1 and BRCA2 are responsible for repairing DNA damage via the homologous recombination DNA repair pathway. The germline inheritance of a pathogenic mutation in one of these genes, together with somatic inactivation of the wild type allele (e.g., via loss or methylation) leads to the accumulation of DNA damage, in particular DNA double strand breaks. This damage is unable to be repaired correctly, leading to genetic instability and predisposing to cancer development. Resulting tumours are said to be homologous recombination deficient (HRD) and importantly sensitive to PARP inhibitors.
      • Bryant H.E.
      • Schultz N.
      • Thomas H.D.
      • et al.
      Specific killing of BRCA2-deficient tumours with inhibitors of poly(ADP-ribose) polymerase.
      • Farmer H.
      • McCabe N.
      • Lord C.J.
      • et al.
      Targeting the DNA repair defect in BRCA mutant cells as a therapeutic strategy.
      • Fong P.C.
      • Boss D.S.
      • Yap T.A.
      • et al.
      Inhibition of poly(ADP-ribose) polymerase in tumors from BRCA mutation carriers.
      • Tutt A.N.
      • Lord C.J.
      • McCabe N.
      • et al.
      Exploiting the DNA repair defect in BRCA mutant cells in the design of new therapeutic strategies for cancer.
      Whilst BRCA1/2 testing has historically been considered in the context of germline mutations, testing has now expanded to include tumour samples to investigate somatic indicators of BRCA1/2 deficiency. Examples include testing for somatic BRCA1/2 mutations, examining for genome wide measures of DNA copy number alterations, known as the HRD score
      • Telli M.L.
      • Hellyer J.
      • Audeh W.
      • et al.
      Homologous recombination deficiency (HRD) status predicts response to standard neoadjuvant chemotherapy in patients with triple-negative or BRCA1/2 mutation-associated breast cancer.
      and using a combination of somatic mutational signatures from whole genome sequencing, known as the HRDetect score.
      • Davies H.
      • Glodzik D.
      • Morganella S.
      • et al.
      HRDetect is a predictor of BRCA1 and BRCA2 deficiency based on mutational signatures.
      In one study examining the utility of the mutational signatures and the HRDetect score, up to 22% of the cohort of 560 breast cancers had patterns of somatic mutations reflecting a BRCA1/2 deficiency, with a proportion of these having no identifiable pathogenic germline or somatic alterations to BRCA1/2 or other members of the homologous recombination DNA repair pathway.
      • Davies H.
      • Glodzik D.
      • Morganella S.
      • et al.
      HRDetect is a predictor of BRCA1 and BRCA2 deficiency based on mutational signatures.
      Like tumours with germline BRCA mutations, those tumours with somatic BRCA alterations may also respond to platinum-based therapies
      • Zhao E.Y.
      • Shen Y.
      • Pleasance E.
      • et al.
      Homologous recombination deficiency and platinum-based therapy outcomes in advanced breast cancer.
      and are similarly susceptible to treatment with PARP inhibitors.
      • Tung N.M.
      • Robson M.E.
      • Ventz S.
      • et al.
      TBCRC 048: phase II study of olaparib for metastatic breast cancer and mutations in homologous recombination-related genes.
      The importance of recognising BRCA1/2 mutations in breast cancer is now heightened as it not only identifies patients who can potentially benefit from genetic counselling and familial disease prevention strategies if the mutations are proven to be germline in origin, but it may also inform possible treatment modalities for those with either germline or somatic alterations.

      The rise of the PARP inhibitor

      The implications of BRCA1/2 mutations for DNA repair pathways led to the hypothesis that cancer cells harbouring these mutations may be sensitive to PARP inhibition.
      • Bryant H.E.
      • Schultz N.
      • Thomas H.D.
      • et al.
      Specific killing of BRCA2-deficient tumours with inhibitors of poly(ADP-ribose) polymerase.
      ,
      • Farmer H.
      • McCabe N.
      • Lord C.J.
      • et al.
      Targeting the DNA repair defect in BRCA mutant cells as a therapeutic strategy.
      ,
      • Tutt A.N.
      • Lord C.J.
      • McCabe N.
      • et al.
      Exploiting the DNA repair defect in BRCA mutant cells in the design of new therapeutic strategies for cancer.
      ,
      • Lord C.J.
      • Tutt A.N.
      • Ashworth A.
      Synthetic lethality and cancer therapy: lessons learned from the development of PARP inhibitors.
      PARPs are a family of enzymes that are activated to repair single stranded breaks in DNA, maintain aspects of the DNA replication process and protect replication fork stability when forks are obstructed. Inhibiting these enzymes in the context of biallelic inactivation of BRCA1 or BRCA2 and homologous recombination deficiency leads to tumour cell death via synthetic lethality. As both mechanisms of DNA repair are crippled in tumour cells, DNA lesions can progress to double stranded breaks, eventually leading to genome collapse and cell death.
      Normal cells survive this treatment, since they carry a wild type copy of BRCA1 and BRCA2 and hence functional homologous recombination DNA repair and replication fork stabilisation.
      The initial investigation of the utility of PARP inhibitors was in ovarian cancers with up to 20% of high grade serous carcinomas harbouring BRCA1/2 mutations.
      Cancer Genome Atlas Research Network
      Integrated genomic analyses of ovarian carcinoma.
      Many patients with high grade ovarian carcinomas initially treated with platinum-based chemotherapy that had longer survival rates were associated with BRCA1/2 mutations,
      • Fong P.C.
      • Yap T.A.
      • Boss D.S.
      • et al.
      Poly(ADP)-ribose polymerase inhibition: frequent durable responses in BRCA carrier ovarian cancer correlating with platinum-free interval.
      thought to be due to the underlying HRD, providing the initial impetus for the investigation of olaparib. Subsequent studies supported proof of concept for olaparib as a maintenance treatment for patients with germline BRCA1/2 mutations in the setting of platinum-sensitive disease that had relapsed.
      • Ledermann J.
      • Harter P.
      • Gourley C.
      • et al.
      Olaparib maintenance therapy in patients with platinum-sensitive relapsed serous ovarian cancer: a preplanned retrospective analysis of outcomes by BRCA status in a randomised phase 2 trial.
      ,
      • Pujade-Lauraine E.
      • Ledermann J.A.
      • Selle F.
      • et al.
      Olaparib tablets as maintenance therapy in patients with platinum-sensitive, relapsed ovarian cancer and a BRCA1/2 mutation (SOLO2/ENGOT-Ov21): a double-blind, randomised, placebo-controlled, phase 3 trial.
      The SOLO-1 phase III trial demonstrated maintenance treatment with olaparib significantly improved progression-free survival in patients with advanced ovarian cancer who had been treated with first-line platinum based chemotherapy and had germline BRCA mutations.
      • Moore K.
      • Colombo N.
      • Scambia G.
      • et al.
      Maintenance olaparib in patients with newly diagnosed advanced ovarian cancer.
      SOLO-2, although not statistically significant, demonstrated an improvement in overall survival, supporting the use of maintenance treatment with olaparib.
      • Poveda A.
      • Floquet A.
      • Ledermann J.A.
      • et al.
      Olaparib tablets as maintenance therapy in patients with platinum-sensitive relapsed ovarian cancer and a BRCA1/2 mutation (SOLO2/ENGOT-Ov21): a final analysis of a double-blind, randomised, placebo-controlled, phase 3 trial.
      The efficacy of PARP inhibitors as frontline treatment in advanced ovarian cancer has been demonstrated in key phase III clinical trials
      • Moore K.
      • Colombo N.
      • Scambia G.
      • et al.
      Maintenance olaparib in patients with newly diagnosed advanced ovarian cancer.
      • Poveda A.
      • Floquet A.
      • Ledermann J.A.
      • et al.
      Olaparib tablets as maintenance therapy in patients with platinum-sensitive relapsed ovarian cancer and a BRCA1/2 mutation (SOLO2/ENGOT-Ov21): a final analysis of a double-blind, randomised, placebo-controlled, phase 3 trial.
      • Ray-Coquard I.
      • Pautier P.
      • Pignata S.
      • et al.
      Olaparib plus bevacizumab as first-line maintenance in ovarian cancer.
      • Gonzalez-Martin A.
      • Pothuri B.
      • Vergote I.
      • et al.
      Niraparib in patients with newly diagnosed advanced ovarian cancer.
      and they are now considered standard of care first line maintenance therapy for patients with BRCA-related advanced ovarian cancers.
      • Moore K.N.
      • Pothuri B.
      Poly(ADP-Ribose) polymerase inhibitor inhibition in ovarian cancer: a comprehensive review.
      The first phase I trial considering the use of a PARP inhibitor in treating patients had an expansion cohort in those with BRCA1/2 mutated tumours, including breast cancers, and was published in 2009.
      • Fong P.C.
      • Boss D.S.
      • Yap T.A.
      • et al.
      Inhibition of poly(ADP-ribose) polymerase in tumors from BRCA mutation carriers.
      Promising initial results in the expansion cohort led to subsequent proof of concept phase II
      • Tutt A.
      • Robson M.
      • Garber J.E.
      • et al.
      Oral poly(ADP-ribose) polymerase inhibitor olaparib in patients with BRCA1 or BRCA2 mutations and advanced breast cancer: a proof-of-concept trial.
      and randomised phase III trials in the breast cancer metastatic setting for patients with germline BRCA1/2 mutations.
      • Robson M.
      • Im S.A.
      • Senkus E.
      • et al.
      Olaparib for metastatic breast cancer in patients with a germline BRCA mutation.
      ,
      • Litton J.K.
      • Rugo H.S.
      • Ettl J.
      • et al.
      Talazoparib in patients with advanced breast cancer and a germline BRCA mutation.
      Based on these trials, the PARP inhibitors olaparib and talazoparib are now approved for use in metastatic breast cancer patients with germline BRCA-mutated, HER2-negative breast cancer based on two key clinical trials. OlympiAD was a phase III randomised clinical trial of metastatic breast cancer patients that demonstrated olaparib improved progression-free survival compared to physican's choice standard chemotherapy with a simultaneous improvement of health-related quality of life.
      • Robson M.
      • Im S.A.
      • Senkus E.
      • et al.
      Olaparib for metastatic breast cancer in patients with a germline BRCA mutation.
      ,
      • Robson M.E.
      • Tung N.
      • Conte P.
      • et al.
      OlympiAD final overall survival and tolerability results: olaparib versus chemotherapy treatment of physician's choice in patients with a germline BRCA mutation and HER2-negative metastatic breast cancer.
      The EMBRACA phase III trial showed similar results for talazoparib with improved progression-free survival.
      • Litton J.K.
      • Rugo H.S.
      • Ettl J.
      • et al.
      Talazoparib in patients with advanced breast cancer and a germline BRCA mutation.
      ,
      • Litton J.K.
      • Hurvitz S.A.
      • Mina L.A.
      • et al.
      Talazoparib versus chemotherapy in patients with germline BRCA1/2-mutated HER2-negative advanced breast cancer: final overall survival results from the EMBRACA trial.
      There is also emerging evidence that somatic BRCA mutation status may be utilised as a predictive biomarker for response to PARP inhibition in both ovarian and breast cancers.
      • Tung N.M.
      • Robson M.E.
      • Ventz S.
      • et al.
      TBCRC 048: phase II study of olaparib for metastatic breast cancer and mutations in homologous recombination-related genes.
      ,
      • Dougherty B.A.
      • Lai Z.
      • Hodgson D.R.
      • et al.
      Biological and clinical evidence for somatic mutations in BRCA1 and BRCA2 as predictive markers for olaparib response in high-grade serous ovarian cancers in the maintenance setting.
      ,
      • Shimomura A.
      • Yonemori K.
      • Yoshida M.
      • et al.
      Gene alterations in triple-negative breast cancer patients in a Phase I/II study of eribulin and olaparib combination therapy.
      Studies examining the use of PARP inhibitors in breast cancer patients with somatic BRCA1/2 mutations or germline PALB2 mutations have shown some promising initial results, suggesting an even broader population of patients may benefit from PARP inhibition.
      • Tung N.M.
      • Robson M.E.
      • Ventz S.
      • et al.
      TBCRC 048: phase II study of olaparib for metastatic breast cancer and mutations in homologous recombination-related genes.
      ,
      • Grellety T.
      • Peyraud F.
      • Sevenet N.
      • et al.
      Dramatic response to PARP inhibition in a PALB2-mutated breast cancer: moving beyond BRCA.

      The OLYMPIA trial

      OlympiA
      • Tutt A.N.J.
      • Garber J.E.
      • Kaufman B.
      • et al.
      Adjuvant olaparib for patients with BRCA1- or BRCA2-mutated breast cancer.
      is a multi-centre trial spanning a large number of institutions, recruiting patients from around the world, and designed to assess the efficacy of adjuvant PARP inhibition in germline BRCA1/2 mutated breast cancer patients. Many of these patients present at an early age, at higher stages and with more aggressive biological recurrence risk features, hence the need to seek additional adjuvant therapies.
      The trial is a phase III, double-blind, randomised controlled trial that randomised 1836 high risk, early stage, HER2-negative breast cancer patients treated with adjuvant or neo-adjuvant chemotherapy, surgery and radiotherapy where indicated.
      • Tutt A.N.J.
      • Garber J.E.
      • Kaufman B.
      • et al.
      Adjuvant olaparib for patients with BRCA1- or BRCA2-mutated breast cancer.
      Eligible patients were then randomised to either one year of adjuvant olaparib or placebo.
      • Tutt A.N.J.
      • Garber J.E.
      • Kaufman B.
      • et al.
      Adjuvant olaparib for patients with BRCA1- or BRCA2-mutated breast cancer.
      At the time of the interim analysis with a median follow-up of 2.5 years, 85.9% of patients treated with adjuvant olaparib were alive and free of recurrent invasive cancer compared with 77.1% of placebo-treated patients.
      • Tutt A.N.J.
      • Garber J.E.
      • Kaufman B.
      • et al.
      Adjuvant olaparib for patients with BRCA1- or BRCA2-mutated breast cancer.
      In these initial results, when compared with the placebo, adjuvant olaparib reduced the risk of local recurrence, metastatic recurrence, other new cancers and death due to any cause by 42% (p<0.0001).
      • Tutt A.N.J.
      • Garber J.E.
      • Kaufman B.
      • et al.
      Adjuvant olaparib for patients with BRCA1- or BRCA2-mutated breast cancer.
      Overall survival data continue to mature, and have recently been reported at the ESMO Virtual Plenary presentation showing overall survival benefits had crossed the pre-specified significance with a 32% reduction in risk of death.
      • Tutt A.N.J.
      • Garber J.
      • Gelber R.D.
      • et al.
      Pre-specified event driven analysis of overall survival (OS) in the OlympiA phase III trial of adjuvant olaparib (OL) in germline BRCA1/2 mutation (gBRCAm) associated breast cancer. ESMO virtual plenary sessions.
      These results indicate that adjuvant olaparib is a valuable addition to the standard of care for patients with high recurrence risk, germline BRCA-mutated, early stage, HER2-negative breast cancer in patients who have received (neo)adjuvant chemotherapy.

      Role of pathology in clinical trials and in Olympia

      Successful patient management and indeed clinical trial design requires the coordinated efforts of many individuals with diverse areas of expertise, including pathology, oncology, surgery, radiology, bioinformatics, statistics, nursing and administration. In the OlympiA trial, pathologists have been involved throughout all of the stages of the clinical trial process. Their broad ranging role encompasses initial conceptualisation and study design, pre-trial eligibility and recruitment, monitoring, collation of biospecimens and trial data, publication of the initial results as well as contributing to the ongoing research project (Table 1). Input from pathologists regarding the rationale for inclusion critieria and the feasibility, accuracy and reproducibility of eligibility criteria were key contributions to the initial study design.
      Table 1Contribution of pathology to clinical trials
      Contribution of pathology to clinical trials
      • Clinical trial design and planning
      • Eligibility criteria
        • Histological assessment & pathological staging
        • Biomarker assessment (e.g. ER, PR and HER2 status)
        • Molecular testing including genetic testing
      • • Central histology review
      • • Monitoring for response
        • Pathological response to treatment
      • Collation of data and reviewing results
      • • Supplying diagnostic or archival material for testing
      • • Quality control and assessment
      • • Tissue biobanking
      Pathology testing plays a critical role in the management of breast cancer patients and in directing the use of currently available therapeutics, with many of these findings also crucial in determining clinical trial eligibility. In breast cancer, pathological evaluation is performed for tumour classification, to provide prognostic information (e.g., staging, gene expression assays), to predict response to treatment (e.g., HER-2 status; trastuzumab) and to investigate for underlying germline mutations. As such, results of histopathology, immunohistochemistry, in situ hybridisation testing and next generation sequencing techniques may all provide information required for clinical trial enrolment.
      Molecular genetic testing and histopathology results were also key determinants of inclusion in the OlympiA trial. Patients had to have undergone genetic testing for germline BRCA1/2 pathogenic variants. Histopathologists played a role in clinical trial recruitment by accurately reporting breast cancer specimens, especially HER2 status, as only HER2-negative patients were eligible. In OlympiA, hormone receptor and HER2 status results underwent a central review.
      • Tutt A.N.J.
      • Garber J.E.
      • Kaufman B.
      • et al.
      Adjuvant olaparib for patients with BRCA1- or BRCA2-mutated breast cancer.
      This was necessary since interpretation of immunohistochemistry is not straightforward and is an evolving area with changes in criteria and cut-off values occurring over time. Further, it was important in trial that the misclassification rates were not significantly different between groups. Oestrogen receptor (ER) and progesterone receptor (PR) have a low diagnostic threshold with 1% nuclear staining considered a low positive result.
      • Burstein H.J.
      • Curigliano G.
      • Thurlimann B.
      • et al.
      Customizing local and systemic therapies for women with early breast cancer: the St. Gallen International Consensus Guidelines for treatment of early breast cancer 2021.
      HER2 is evaluated in the first instance using immunohistochemistry and if the result is equivocal or positive, further evaluation of the HER2 copy number via in situ hybridisation is performed.
      • Wolff A.C.
      • Hammond M.E.H.
      • Allison K.H.
      • et al.
      Human epidermal growth factor receptor 2 testing in breast cancer: American Society of Clinical Oncology/College of American Pathologists clinical practice guideline focused update.
      Adequacy and length of fixation for breast specimens can impact immunohistochemistry and in situ hybridisation testing, where interpretation relies heavily on the quality of both specimen fixation and staining. Subtle variations in the interpretation of diagnostic criteria may affect whether a tumour is considered hormone receptor-positive or triple-negative which can have implications not only for treatment but also for clinical trial inclusion. Clinical trial participation may act as a mechanism of additional quality assurance with central review to confirm eligibility criteria providing external validation of results at no additional cost to a laboratory.
      Other reported pathological findings such as tumour size and lymph node status were also important aspects of OlympiA inclusion criteria. For example, ER-positive patients who underwent adjuvant chemotherapy had to have at least four involved lymph nodes to be considered for the trial.
      • Tutt A.N.J.
      • Garber J.E.
      • Kaufman B.
      • et al.
      Adjuvant olaparib for patients with BRCA1- or BRCA2-mutated breast cancer.
      For triple-negative patients treated with adjuvant chemotherapy, their nodal status and size of invasive carcinoma (>2 cm) determined their trial eligibility.
      • Tutt A.N.J.
      • Garber J.E.
      • Kaufman B.
      • et al.
      Adjuvant olaparib for patients with BRCA1- or BRCA2-mutated breast cancer.
      Moreover, patients who had neoadjuvant treatment needed to have residual primary tumour or residual disease in lymph nodes to be considered eligible.
      • Tutt A.N.J.
      • Garber J.E.
      • Kaufman B.
      • et al.
      Adjuvant olaparib for patients with BRCA1- or BRCA2-mutated breast cancer.
      Hormone receptor-positive patients treated with neoadjuvant chemotherapy required an incomplete pathological response. These patients had to have a clinical and pathological (CPS) and oestrogen receptor and grade (EG) score of 3 or higher.
      • Tutt A.N.J.
      • Garber J.E.
      • Kaufman B.
      • et al.
      Adjuvant olaparib for patients with BRCA1- or BRCA2-mutated breast cancer.
      The CPS+EG score is on a scale of 0–6 and is used to estimate risk of relapse, with higher scores indicating a higher risk of relapse.
      • Mittendorf E.A.
      • Jeruss J.S.
      • Tucker S.L.
      • et al.
      Validation of a novel staging system for disease-specific survival in patients with breast cancer treated with neoadjuvant chemotherapy.
      The score is derived from a combined assessment of the clinical stage, the pathological stage, the ER status and the nuclear grade which are each allocated a score.
      • Mittendorf E.A.
      • Jeruss J.S.
      • Tucker S.L.
      • et al.
      Validation of a novel staging system for disease-specific survival in patients with breast cancer treated with neoadjuvant chemotherapy.
      Trial inclusion criteria require complete and thorough pathological assessment of the tumour to determine patient eligibility. The strict application of eligibility criteria is critical for the overall value and impact of the clinical trial. Including only patients likely to benefit from a therapy and excluding those who are ineligible that may dilute the possible benefit of a therapy is important not only for guaranteeing the validity of the clinical trial results but also ensuring that patients who will receive no benefit from the treatment and experience possible side effects are not unnecessarily exposed. Without standardisation of results and robust quality control, the results of a trial cannot be compared, reproduced, or implemented in clinical practice. Centralised review plays an important part of this process and is also important in developing and evaluating other pathological and molecular parameters (including features identified through digital analysis) that may separate responders from non-responders. Including a pathologist on the steering committee of a cancer clinical trial may aid not only in the interpretation of pathology results, but early recognition of possible challenges or pitfalls that may arise from the use of biological specimens, leading to improved reliability and reproducibility of clinical trial results.
      Pathology laboratories have long been the repositories of diagnostic material and the storage of tissue blocks and slides for many years leads to the accumulation of robust libraries of archival material. Many pathology laboratories also host tissue biobanks. The role that pathologists play as custodians of this biological material is incredibly important in the execution of both prospective and retrospective studies. In the context of cancer trials, a pathologist is often required to assess whether the material available is representative with the aim of selecting the best tissue block. Ensuring diagnostic material is available when required for clinical trials for central confirmation of eligibility criteria and/or facilitating translational research applications, the safe transit of this valuable resource which may require sending tissue blocks or slides interstate or overseas, together with meeting regulatory requirements that govern the appropriate transportation and preservation of diagnostic material, are all significant considerations.

      The future of pathology and clinical trials

      In this era of precision medicine, ongoing rigorous clinical trials are required before new therapeutics are introduced into mainstream clinical practice. Pathology has a critical role in many clinical trials both in determining clinical trial eligibility and in the assessment of outcomes, data upon which all of the subsequent trial findings are based. Given the pivotal nature of this role, it is ideal that pathologists are involved in the trial process from initial discussions around trial participation to the site initiation visit. Where possible, it is optimal for a pathologist to be specifically designated as responsible for the conduct of the trial in order to streamline a point of contact and to ensure that there is dedicated oversight should questions or queries arise.
      The value of formal pathology participation in clinical trial design has recently been highlighted with the standard protocol items: recommendations for interventional trials (SPIRIT-Path extension).
      • Kendall T.J.
      • Robinson M.
      • Brierley D.J.
      • et al.
      Guidelines for cellular and molecular pathology content in clinical trial protocols: the SPIRIT-Path extension.
      These recently published consensus guidelines provide specific recommendations around clinical trial protocol development involving cellular and molecular pathology.
      • Kendall T.J.
      • Robinson M.
      • Brierley D.J.
      • et al.
      Guidelines for cellular and molecular pathology content in clinical trial protocols: the SPIRIT-Path extension.
      These guidelines recognise how important pathological assessment is in the clinical trial process and aim to formalise aspects of specimen handling and reporting, with the intention that the recommendations be specifically addressed in clinical trial protocols to facilitate standardisation and study reproducibility.
      • Kendall T.J.
      • Robinson M.
      • Brierley D.J.
      • et al.
      Guidelines for cellular and molecular pathology content in clinical trial protocols: the SPIRIT-Path extension.
      Some of the recommendations include documenting the laboratory and accreditation status, pathology reporting methods and requirements for central pathological review, outlining assessment of methods and timing of tissue sampling for pathology-specific outcomes and stating specific methods for specimen assessment by histochemical, immunohistochemical or molecular techniques.
      • Kendall T.J.
      • Robinson M.
      • Brierley D.J.
      • et al.
      Guidelines for cellular and molecular pathology content in clinical trial protocols: the SPIRIT-Path extension.
      Encouraging the adoption of these guidelines will also help to recognise the contribution of pathologists and scientists in clinical trial development and execution.
      • Kendall T.J.
      • Robinson M.
      • Brierley D.J.
      • et al.
      Guidelines for cellular and molecular pathology content in clinical trial protocols: the SPIRIT-Path extension.
      From an economic perspective, acknowledging the contribution of pathologists with adequate reimbursement for the service that they provide is also important. The financial agreement between trial sponsors and local sites should include adequate compensation for both the pathologist and their laboratory.
      As we move forward, the contribution of pathology in clinical trial medicine is shifting from simply providing diagnosis and basic tumour classification to incorporating nuanced analyses of complex molecular markers, setting up-to-date standards and benchmarks for biomarker evaluation, examination of treatment response in tissue (e.g., pathological complete response may be a primary endpoint in many oncology trials) and in the future possibly monitoring for disease recurrence (e.g., circulating tumour DNA; ctDNA). In the context of PARP inhibitors, it is plausible that in the future ctDNA may be used to monitor for mutations that confer resistance.
      • Dias M.P.
      • Moser S.C.
      • Ganesan S.
      • Jonkers J.
      Understanding and overcoming resistance to PARP inhibitors in cancer therapy.
      Patients treated with PARP inhibitors are known to develop resistance, with one of the key mechanisms being reversion mutations that can induce re-expression of BRCA1/2 wild-type proteins.
      • Dias M.P.
      • Moser S.C.
      • Ganesan S.
      • Jonkers J.
      Understanding and overcoming resistance to PARP inhibitors in cancer therapy.
      ,
      • Pettitt S.J.
      • Frankum J.R.
      • Punta M.
      • et al.
      Clinical BRCA1/2 reversion analysis identifies hotspot mutations and predicted neoantigens associated with therapy resistance.
      There are already studies considering the applications of ctDNA in monitoring for reversion-related PARP inhibitor resistance.
      • Weigelt B.
      • Comino-Mendez I.
      • de Bruijn I.
      • et al.
      Diverse BRCA1 and BRCA2 reversion mutations in circulating cell-free DNA of therapy-resistant breast or ovarian cancer.
      ,
      • Quigley D.
      • Alumkal J.J.
      • Wyatt A.W.
      • et al.
      Analysis of circulating cell-free DNA identifies multiclonal heterogeneity of BRCA2 reversion mutations associated with resistance to PARP inhibitors.
      Currently pathologists are called upon to evaluate tumour cellularity, assess tumour viability and to determine whether tissue blocks or cytological specimens have sufficient tumour present to enable molecular evaluation. Tools such as HRDetect can be performed on formalin-fixed, paraffin-embedded tissue but need sufficient tumour cellularity to enable whole genome sequencing to be performed.
      • Davies H.
      • Glodzik D.
      • Morganella S.
      • et al.
      HRDetect is a predictor of BRCA1 and BRCA2 deficiency based on mutational signatures.
      Similarly, extracting DNA from frozen or formalin-fixed tissue to perform a HRD score requires pathological review to select areas of tumour that are enriched with the highest tumour cell density to facilitate the greatest tumour DNA yield.
      • Telli M.L.
      • Timms K.M.
      • Reid J.
      • et al.
      Homologous recombination deficiency (HRD) score predicts response to platinum-containing neoadjuvant chemotherapy in patients with triple-negative breast cancer.
      Pathologists may soon be required to determine whether there is sufficient tumour content to enable DNA extraction to perform assessment for HR deficiency or somatic BRCA1/2 mutations, molecular findings that in the future may be key to determining eligibility for drugs like PARP inhibitors.
      It is also important to highlight that once a new biomarker or drug has gone through the rigorous clinical trial process, it is only through pathology laboratories implementing these new companion diagnostic tests that they are incorporated into routine clinical practice. This requires constant adaptation as the role of pathology and pathology services continues to evolve. Effective and stringent quality measures are required so that clinical trial outcomes can be reproduced in clinical practice, and this often requires new validations, pathologist education, requirements for specimen handling and more. The diagnostic labs will also need to make a case to their managers for resources to deliver the service in a timely manner. The potential therapeutic opportunities of PARP inhibitors are just one example where pathology-based testing for BRCA and BRCA-like alterations will likely expand in the clinical setting to facilitate new treatment options for patients.

      Conclusion

      With pathology laboratories operating behind the scenes in the care of patients with cancer, the active role that pathology plays in large clinical trials and in the introduction of new therapies is easy to underestimate. It may not seem intuitive that pathologists and their teams have contributed to a new drug becoming available or the success of a clinical trial. Yet there are many individuals working at all levels within the discipline of pathology who help realise the goal of bringing new therapies into clinical practice. Pathologists, trainees, scientists and laboratory staff across all aspects of pathology must embrace their role in the clinical trial process as another way in which we are actively contributing to providing the best possible care for patients. With the increasing complexity of the pathological evaluation of disease, now more than ever it is necessary for pathologists, and pathology as a discipline, to step forward and proactively provide guidance, innovation, and leadership in clinical trial design and implementation.

      Acknowledgements

      We would like to acknowledge the work and support of the OlympiA Steering Committee, Pathology Group, and Translational Advisory Committee (TAC). We would also like to acknowledge the support of Breast Cancer Trials (BCT)—formerly the Australia and New Zealand Breast Cancer Trials Group (ANZBCTG)—who participated in the OlympiA trial and have encouraged and supported pilot and clinical fellowship funding in their clinical trials research.

      Conflicts of interest and sources of funding

      LK is part funded by a pathology clinical fellowship from the NHMRC CRE-TransBCR (Lideman/Lakhani). The authors state that there are no conflicts of interest to disclose.

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