Computational pathology in cancer diagnosis, prognosis, and prediction - present day and prospects.

Computational pathology refers to applying deep learning techniques and algorithms to analyse and interpret histopathology images. Advances in artificial intelligence (AI) have led to an explosion in innovation in computational pathology, ranging from the prospect of automation of routine diagnostic tasks to the discovery of new prognostic and predictive biomarkers from tissue morphology. Despite the promising potential of computational pathology, its integration in clinical settings has been limited by a range of obstacles including operational, technical, regulatory, ethical, financial, and cultural challenges. Here, we focus on the pathologists' perspective of computational pathology: we map its current translational research landscape, evaluate its clinical utility, and address the more common challenges slowing clinical adoption and implementation. We conclude by describing contemporary approaches to drive forward these techniques. © 2023 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.

Cell type-restricted activity of hnRNPM promotes breast cancer metastasis via regulating alternative splicing.

Tumor metastasis remains the major cause of cancer-related death, but its molecular basis is still not well understood. Here we uncovered a splicing-mediated pathway that is essential for breast cancer metastasis. We show that the RNA-binding protein heterogeneous nuclear ribonucleoprotein M (hnRNPM) promotes breast cancer metastasis by activating the switch of alternative splicing that occurs during epithelial-mesenchymal transition (EMT). Genome-wide deep sequencing analysis suggests that hnRNPM potentiates TGFß signaling and identifies CD44 as a key downstream target of hnRNPM. hnRNPM ablation prevents TGFß-induced EMT and inhibits breast cancer metastasis in mice, whereas enforced expression of the specific CD44 standard (CD44s) splice isoform overrides the loss of hnRNPM and permits EMT and metastasis. Mechanistically, we demonstrate that the ubiquitously expressed hnRNPM acts in a mesenchymal-specific manner to precisely control CD44 splice isoform switching during EMT. This restricted cell-type activity of hnRNPM is achieved by competition with ESRP1, an epithelial splicing regulator that binds to the same cis-regulatory RNA elements as hnRNPM and is repressed during EMT. Importantly, hnRNPM is associated with aggressive breast cancer and correlates with increased CD44s in patient specimens. These findings demonstrate a novel molecular mechanism through which tumor metastasis is endowed by the hnRNPM-mediated splicing program.

Phyllodes tumours of the breast: a consensus review.

Phyllodes tumours constitute an uncommon but complex group of mammary fibroepithelial lesions. Accurate and reproducible grading of these tumours has long been challenging, owing to the need to assess multiple stratified histological parameters, which may be weighted differently by individual pathologists. Distinction of benign phyllodes tumours from cellular fibroadenomas is fraught with difficulty, due to overlapping microscopic features. Similarly, separation of the malignant phyllodes tumour from spindle cell metaplastic carcinoma and primary breast sarcoma can be problematic. Phyllodes tumours are treated by surgical excision. However, there is no consensus on the definition of an appropriate surgical margin to ensure completeness of excision and reduction of recurrence risk. Interpretive subjectivity, overlapping histological diagnostic criteria, suboptimal correlation between histological classification and clinical behaviour and the lack of robust molecular predictors of outcome make further investigation of the pathogenesis of these fascinating tumours a matter of active research. This review consolidates the current understanding of their pathobiology and clinical behaviour, and includes proposals for a rational approach to the classification and management of phyllodes tumours.

A population-level digital histologic biomarker for enhanced prognosis of invasive breast cancer.

Breast cancer is a heterogeneous disease with variable survival outcomes. Pathologists grade the microscopic appearance of breast tissue using the Nottingham criteria, which are qualitative and do not account for noncancerous elements within the tumor microenvironment. Here we present the Histomic Prognostic Signature (HiPS), a comprehensive, interpretable scoring of the survival risk incurred by breast tumor microenvironment morphology. HiPS uses deep learning to accurately map cellular and tissue structures to measure epithelial, stromal, immune, and spatial interaction features. It was developed using a population-level cohort from the Cancer Prevention Study-II and validated using data from three independent cohorts, including the Prostate, Lung, Colorectal, and Ovarian Cancer trial, Cancer Prevention Study-3, and The Cancer Genome Atlas. HiPS consistently outperformed pathologists in predicting survival outcomes, independent of tumor-node-metastasis stage and pertinent variables. This was largely driven by stromal and immune features. In conclusion, HiPS is a robustly validated biomarker to support pathologists and improve patient prognosis.

Validation Study of the Newly Proposed Refined Diagnostic Criteria for Malignant Phyllodes Tumor With 136 Borderline and Malignant Phyllodes Tumor Cases.

The World Health Organization (WHO) diagnostic criteria for malignant phyllodes tumor (MPT) may miss a significant number of MPTs with metastatic potential. New refined diagnostic criteria (Refined Criteria) for MPT were recently proposed. The aim of this study is to validate the Refined Criteria. This validation study included 136 borderline (borderline phyllodes tumor [BoPT]) and MPT cases that were not included in the initial study. We evaluated tumor classifications based on both the Refined Criteria and the WHO criteria. The Refined Criteria defines MPT when these criteria are met (1) stromal overgrowth with ≥ 1 feature(s) of marked stromal cellularity, marked stromal cytologic atypia, or ≥10 mitoses per 10 high-power fields (10 mitoses/10 HPFs) or (2) marked stromal cellularity with ≥1 feature(s) of marked stromal cytologic atypia, ≥10 mitoses/10 HPFs or permeative border. The WHO criteria require all 5 morphologic features (stromal overgrowth, permeative border, marked stromal cellularity, marked stromal cytologic atypia, and ≥10 mitoses/10 HPFs) for an MPT diagnosis. Using the Refined Criteria, none of the 61 BoPTs developed metastasis and 40.0% of the 75 MPTs developed metastases; local recurrence was seen in 11.5% BoPTs and 25.3% MPTs. Using the WHO criteria, 9.6% of the 94 BoPTs developed metastases and 50.0% of the 42 MPTs developed metastases; 14.9% of the BoPTs had local recurrence and 28.6% of the MPTs had local recurrence. Nine (30.0%) of the 30 tumors that developed distant metastases were diagnosed as BoPTs by the WHO criteria. When we combined the 75 MPTs from this validation cohort with the 65 MPT cases from the published data using the Refined Criteria, 50 (35.7%) of the 140 MPTs developed metastases, whereas 8 cases with metastases were <5 cm. In the univariate analysis with log-rank test, stromal overgrowth, marked stromal cellularity, marked stromal cytologic atypia, ≥10 mitoses/10 HPFs, presence of heterologous components other than liposarcomatous component, and presence of stromal necrosis were significantly associated with the risk of metastasis (all with P < 0.05). In multivariate analysis with Cox proportional hazard regression, stromal overgrowth and marked stromal cellularity were significantly associated with metastasis (both with P < 0.001). The Refined Criteria are superior to the WHO criteria in predicting the clinical outcomes of BoPTs and MPTs. Using the Refined Criteria, 35.7% of 140 patients with MPT developed metastases, whereas none (0%) of the patients with BoPT developed metastases. Patients with MPT have a high metastatic rate; these patients may benefit from systemic chemotherapy or targeted therapies. In contrast, patients with BoPT may be managed with complete local excision alone without chemotherapy.

Preliminary results of centralized HER2 testing in ductal carcinoma in situ (DCIS): NSABP B-43.

NSABP B-43 is the first prospective, randomized phase III multi-institution clinical trial targeting high-risk, HER2-positive DCIS. It compares whole breast irradiation alone with WBI given concurrently with trastuzumab in women with HER2-positive DCIS treated by lumpectomy. The primary aim is to determine if trastuzumab plus radiation will reduce in-breast tumor recurrence. HER2-positive DCIS was previously estimated at >50 %, occurring primarily in ER-negative, comedo-type DCIS of high nuclear grade. There has been no documented centralized multi-institutional HER2 analysis of DCIS. NSABP B-43 provides a unique opportunity to evaluate this in a large cohort of DCIS patients. Patients undergoing lumpectomy for DCIS without evidence of an invasive component are eligible. A central review of each patient's pure DCIS lesion is carried out by immunohistochemistry analysis. If the lesion is 2+, FISH analysis is performed. Patients whose tumors are HER2 3+ or FISH-positive are randomly assigned to receive two doses of trastuzumab during WBI or WBI alone. NSABP B-43 opened 11/9/08. As of 7/31/2013, 5,861 patients have had specimens received centrally, and 5,645 of those had analyzable blocks; 1,969 (34.9 %) were HER2 positive. A total of 1,428 patients have been accrued, 1,137 (79.6 %) of whom have follow-up information. The average follow-up time for the 1,137 patients is 23.3 months. No grade 4 or 5 toxicity has been observed. In NSABP B-43 the HER2-positive rate for pure DCIS among patients undergoing breast-preserving surgery is 34.9 %, lower than the previously reported rate. No trastuzumab-related safety signals have been observed. Interest in this trial has been robust.

Updates in the pathology of Pregnancy Associated Breast Cancer (PABC).

Pregnancy associated breast cancer (PABC) is defined as a breast cancer diagnosed during gestation, lactation or within 5 years postpartum. While the development of malignancy during pregnancy is rare, the incidence is increasing. Breast cancer is one of the most common cancers diagnosed during pregnancy, affecting up to 1 in 3000 deliveries. New understanding of the pathophysiology of PABC recently resulted in updated definitions distinguishing breast cancer diagnosed during pregnancy (PrBC) from cancer diagnosed during the postpartum period (PPBC) due to distinct biology and prognosis. Pregnancy has a dual effect on breast cancer development- both protective against cancer and promoting tumor growth. While several hypotheses have been proposed over the years to explain these effects, the most likely hypothesis for the development of PABC is the involution hypothesis, proposing that remodeling programs activated in the immediate postpartum period are similar to wound healing and inflammation that may be associated with tumor development and progression. Although PABCs reflect all subtypes of breast carcinomas, they are most commonly invasive ductal carcinomas of high tumor grade and large tumor size, with more advanced stage at presentation and higher rates of lymph node involvement. Most PABCs are hormone negative tumors (triple negative or HER2 amplified tumors) with high Ki-67 proliferation rates. Several studies have shown that PABCs have different genomic signatures than non-PABC tumors, showing increased expression of immune response mediators. Better understanding of the molecular pathways of tumor initiation and progression, along with prompt diagnosis and novel treatment protocols in the care of PrBC and PPBC are needed to improve outcomes for these young, high-risk breast cancer patients.

The role of PRAME and NY-ESO-1 as potential therapeutic and prognostic biomarkers in triple-negative breast carcinomas.

PRAME and NY-ESO-1 are cancer-testis antigens (CTAs) reported to be highly enriched in triple-negative breast cancers (TNBCs), against which vaccines and immunotherapies are currently being developed. This study aims to analyze PRAME and NY-ESO-1 expression in TNBCs and their correlation with clinical outcomes. This is a retrospective cohort study of TNBC patients who have undergone neoadjuvant chemotherapy. PRAME and NY-ESO-1 expression were assessed on pre-therapy biopsies as H-scores (percentage x intensity) with final H scores of 2-3 considered as positive. Association between expression and pathologic complete response (pCR), metastasis, and residual cancer burden (RCB) were assessed via logistic regression. Cox proportional hazards models were used to assess the association with progression-free survival. P-values < 0.05 were considered statistically significant. Sixty-three percent of 76 patients were positive for PRAME. In contrast, only 5 % were positive for NY-ESO-1. PRAME positivity was significantly associated with a lower likelihood of early metastatic disease (OR = 0.24, 95 % CI 0.08-0.62; P = 0.005). However, it was not significantly associated with pCR, RCB category, or progression-free survival. NY-ESO1 score was not significantly associated with early metastatic disease, pCR, RCB category, or progression-free survival. Our results suggest that PRAME positivity may be associated with a lower risk of early metastasis in TNBCs, but not with response to neoadjuvant chemotherapy or progression-free survival. The high expression of PRAME in TNBCs makes it a potential therapeutic target, while NY-ESO1 appears to be a less useful marker. However, further larger studies are needed to ascertain the utility of these markers.

Computational ranking-assisted identification of Plexin-B2 in homotypic and heterotypic clustering of circulating tumor cells in breast cancer metastasis.

Metastasis is the cause of over 90% of all deaths associated with breast cancer, yet the strategies to predict cancer spreading based on primary tumor profiles and therefore prevent metastasis are egregiously limited. As rare precursor cells to metastasis, circulating tumor cells (CTCs) in multicellular clusters in the blood are 20-50 times more likely to produce viable metastasis than single CTCs. However, the molecular mechanisms underlying various CTC clusters, such as homotypic tumor cell clusters and heterotypic tumor-immune cell clusters, are yet to be fully elucidated. Combining machine learning-assisted computational ranking with experimental demonstration to assess cell adhesion candidates, we identified a transmembrane protein Plexin- B2 (PB2) as a new therapeutic target that drives the formation of both homotypic and heterotypic CTC clusters. High PB2 expression in human primary tumors predicts an unfavorable distant metastasis-free survival and is enriched in CTC clusters compared to single CTCs in advanced breast cancers. Loss of PB2 reduces formation of homotypic tumor cell clusters as well as heterotypic tumor-myeloid cell clusters in triple-negative breast cancer. Interactions between PB2 and its ligand Sema4C on tumor cells promote homotypic cluster formation, and PB2 binding with Sema4A on myeloid cells (monocytes) drives heterotypic CTC cluster formation, suggesting that metastasizing tumor cells hijack the PB2/Sema family axis to promote lung metastasis in breast cancer. Additionally, using a global proteomic analysis, we identified novel downstream effectors of the PB2 pathway associated with cancer stemness, cell cycling, and tumor cell clustering in breast cancer. Thus, PB2 is a novel therapeutic target for preventing new metastasis.

Cytokeratin 7: a re-evaluation of the 'tried and true' in triple-negative breast cancers.

AIMS: Triple-negative breast cancers (TNBCs) are often poorly differentiated tumours that can present clinically as metastases of an unknown primary. Immunohistochemical panels are frequently used to determine the likelihood of a breast primary, but in this tumour subset cytokeratin (CK)7 may be the only positive finding. In this study we aimed to evaluate a commonly employed immunohistochemical panel using a large group of TNBCs (both basal-like and unclassified), and to analyse the CK7 staining patterns. METHODS AND RESULTS: Tissue microarrays containing 138 TNBCs were stained with antibodies against CK7, CK20, gross cystic disease fluid protein 15 (GCDFP-15), and mammaglobin. CK5/6 staining was used to identify basal-like tumours. CK7 staining was notably heterogeneous, with 14.5% of all cases demonstrating ≤20% tumour cell staining. A greater proportion of basal-like TNBCs than of unclassified TNBCs showed focal staining. GCDFP-15 and mammaglobin were not expressed in the majority of TNBCs, and were less frequently positive in basal-like than in unclassified TNBCs. CONCLUSION: TNBCs are commonly negative for most immunomarkers indicative of breast origin, with the exception of CK7. As about one in five TNBCs showed only focal CK7 positivity, use of this marker must be interpreted with caution, especially in small samples, so that the possibility of a breast primary is not overlooked.

Pathologic evaluation of specimens after neoadjuvant chemotherapy in breast cancer: Current recommendations and challenges.

Neoadjuvant chemotherapy is increasingly used to optimize breast conservation surgery and is becoming a standard of care in a subset of breast cancer patients. An accurate pathologic assessment is crucial in guiding clinical decisions and subsequent management and prognosis. This review aims to summarize the most current literature, recommendations, and challenges in the pathologic evaluation of breast cancer after neoadjuvant chemotherapy. Included are the most current definitions of the different types of tumor response, the underlying factors that can affect tumor response, how to assess lymph nodes, margins, and tumor markers post-neoadjuvant chemotherapy, as well as the different classification systems a pathologist can use to assess residual disease. In this era of de-escalation of surgical treatment, studies on imaging techniques to assess residual disease and avoid surgery after neoadjuvant chemotherapy have also been done. However, at least for now, surgical treatment remains the preferred practice. As such, pathologists play an increasingly critical role in standardizing assessment of residual disease post-neoadjuvant chemotherapy, and in optimizing the knowledge gained by this approach to breast cancer therapy.

Determining PD-L1 Status in Patients With Triple-Negative Breast Cancer: Lessons Learned From IMpassion130.

Triple-negative breast cancer (TNBC) accounts for approximately 12% to 17% of all breast cancers and has an aggressive clinical behavior. Increased tumor-infiltrating lymphocyte counts are prognostic for survival in TNBC, making this disease a potential target for cancer immunotherapy. Research on immunophenotyping of tumor-infiltrating lymphocytes is revealing molecular and structural organization in the tumor microenvironment that may predict patient prognosis. The anti-programmed death-ligand 1 (PD-L1) antibody atezolizumab plus nab-paclitaxel was the first cancer immunotherapy combination to demonstrate progression-free survival benefit and clinically meaningful overall survival benefit in the first-line treatment of metastatic TNBC (mTNBC) in patients with PD-L1-expressing tumor-infiltrating immune cells in 1% or more of the tumor area. This led to its United States and European Union approval for mTNBC and US approval of the VENTANA PD-L1 (SP142) assay as a companion diagnostic immunohistochemistry assay. Subsequently, the anti-programmed death-1 (PD-1 ) antibody pembrolizumab plus chemotherapy was approved by the US Food and Drug Administration for mTNBC based on progression-free survival benefit in patients with a combined positive score of at least 10 by its concurrently approved 22C3 companion diagnostic assay. Treatment guidelines now recommend PD-L1 testing for patients with mTNBC, and the testing landscape will likely become increasingly complex as new anti-PD-L1 and anti-PD-1 agents and diagnostics are approved for TNBC. Integrating PD-L1 testing into current diagnostic workflows for mTNBC may provide more treatment options for these patients. Therefore, it is critical for medical oncologists and pathologists to understand the available assays and their relevance to therapeutic options to develop an appropriate workflow for immunohistochemistry testing.

Correlation of manual semi-quantitative and automated quantitative Ki-67 proliferative index with OncotypeDXTM recurrence score in invasive breast carcinoma.

BACKGROUND: Ki-67 immunohistochemistry (IHC) staining is a widely used cancer proliferation assay; however, its limitations could be improved with automated scoring. The OncotypeDXTM Recurrence Score (ORS), which primarily evaluates cancer proliferation genes, is a prognostic indicator for breast cancer chemotherapy response; however, it is more expensive and slower than Ki-67. OBJECTIVE: To compare manual Ki-67 (mKi-67) with automated Ki-67 (aKi-67) algorithm results based on manually selected Ki-67 "hot spots" in breast cancer, and correlate both with ORS. METHODS: 105 invasive breast carcinoma cases from 100 patients at our institution (2011-2013) with available ORS were evaluated. Concordance was assessed via Cohen's Kappa (κ). RESULTS: 57/105 cases showed agreement between mKi-67 and aKi-67 (κ 0.31, 95% CI 0.18-0.45), with 41 cases overestimated by aKi-67. Concordance was higher when estimated on the same image (κ 0.53, 95% CI 0.37-0.69). Concordance between mKi-67 score and ORS was fair (κ 0.27, 95% CI 0.11-0.42), and concordance between aKi-67 and ORS was poor (κ 0.10, 95% CI -0.03-0.23). CONCLUSIONS: These results highlight the limits of Ki-67 algorithms that use manual "hot spot" selection. Due to suboptimal concordance, Ki-67 is likely most useful as a complement to, rather than a surrogate for ORS, regardless of scoring method.

External Quality Assessment 2.0: The Importance of a Standardized Implementation of TILs for Daily and Trial Practices.

Increasing data suggests that an intact immune system is required for improvedoutcomes in patients with Human Epidermal Growth Factor Receptor 2 (HER2+) and Triple Negative Breast Cancer (TNBC) [...].

Clinicopathologic features of unexpectedly HER2 positive breast carcinomas: An institutional experience.

Human epidermal growth factor receptor 2 (HER2) overexpression occurs in 15-20 % of all breast carcinomas. These tumors are usually high-grade which often correlates with reduced overall survival and increased rates of recurrence. In a retrospective review, we identified 19 cases of unexpectedly HER2 positive (by immunohistochemistry and/or fluorescence in-situ hybridization) invasive breast carcinomas on core needle biopsies from a registry at Northwestern Memorial Hospital. These cases included low-grade tumors, invasive lobular carcinomas, classic type, and invasive carcinomas with special subtype features. Twelve of the tumors were histologic grade 1 and 7 were histologic grade 2. One of the grade 1 tumors had tubular features (8 %), 1 had cribriform features (8 %), 2 had mucinous features (17 %), 2 were invasive lobular carcinomas, classic type (17 %), and the rest were invasive carcinoma, no special type (50 %). The histologic grade 2 tumors included 5 invasive lobular carcinomas, classic type (71 %) and 2 invasive ductal carcinomas with mucinous features (29 %). By immunohistochemistry, 13 (65 %) were HER2 score 3+, 7 were score 2+ (35 %), and reflex fluorescence in-situ hybridization (FISH) testing showed amplification in 6 cases, with 1 equivocal case amplified on excision. Despite the HER2 positive status in the selected cases, no unique morphologic features that would indicate aggressive behavior were identified. In clinical follow up, two patients were found to have recurrences, five had lymph node metastasis, and one had distant metastasis. None of the patients with recurrent disease were treated with trastuzumab, despite their positive HER2 results. These findings support that our population of HER2 positive carcinomas showed a similar rate of lymph node metastases and recurrence as poorly-differentiated tumors, supporting HER2 positivity as a poor prognostic indicator, irrespective of morphologic features. We recommend continuing to test all breast cancers, regardless of grade or special subtype features, to provide the most comprehensive treatment and prognostic information for both clinicians and patients.

The Use of Serial Circulating Tumor DNA to Detect Resistance Alterations in Progressive Metastatic Breast Cancer.

PURPOSE: Circulating tumor DNA (ctDNA) is a promising tool for noninvasive longitudinal monitoring of genomic alterations. We analyzed serial ctDNA to characterize genomic evolution in progressive metastatic breast cancer. EXPERIMENTAL DESIGN: This was a retrospective cohort between 2015 and 2019 obtained under an Institutional Review Board-approved protocol at Northwestern University (Chicago, IL). ctDNA samples were analyzed with Guardant360 next-generation sequencing (NGS) assay. A total of 86 patients had at least two serial ctDNA collections with the second drawn at first post-NGS progression (PN1) by imaging and clinical assessment. A total of 27 participants had ctDNA drawn at second post-NGS clinical progression (PN2). We analyzed alterations, mutant allele frequency (MAF), number of alterations (NOA), and sites of disease on imaging in close proximity to ctDNA evaluation. Matched pairs' variations in MAF, NOA, and alterations at progression were tested through Wilcoxon test. We identified an independent control cohort at Massachusetts General Hospital (Boston, MA) of 63 patients with serial ctDNA sampling and no evidence of progression. RESULTS: We identified 44 hormone receptor-positive, 20 HER2+, and 22 triple-negative breast cancer cases. The significant alterations observed between baseline and PN1 were TP53 (P < 0.0075), PIK3CA (P < 0.0126), AR (P < 0.0126), FGFR1 (P < 0.0455), and ESR1 (P < 0.0143). Paired analyses revealed increased MAF and NOA from baseline to PN1 (P = 0.0026, and P < 0.0001, respectively). When compared with controls without progression, patients with ctDNA collection at times of progression were associated with increased MAF and NOA (P = 0.0042 and P < 0.0001, respectively). CONCLUSIONS: Serial ctDNA testing identified resistance alterations and increased NOA and MAF were associated with disease progression. Prospective longitudinal ctDNA evaluation could potentially monitor tumor genomic evolution.

Comparison of Radiation With or Without Concurrent Trastuzumab for HER2-Positive Ductal Carcinoma In Situ Resected by Lumpectomy: A Phase III Clinical Trial.

PURPOSE: Preclinical studies report that trastuzumab (T) can boost radiotherapy (RT) effectiveness. The primary aim of the B-43 trial was to assess the efficacy of RT alone vs concurrent RT plus T in preventing recurrence of ipsilateral breast cancer (IBTR) in women with ductal carcinoma in situ (DCIS). PATIENTS AND METHODS: Eligibility: Eastern Cooperative Oncology Group (ECOG) performance status 0 or 1, DCIS resected by lumpectomy, known estrogen receptor (ER) and/or progesterone receptor (PgR), and human epidermal growth factor receptor 2 (HER2) status by centralized testing. Whole-breast RT was given concurrently with T. Stratification was by menopausal status, adjuvant endocrine therapy plan, and nuclear grade. Definitive intent-to-treat primary analysis was to be conducted when either 163 IBTR events occurred or all accrued patients were on study ≥ 5 years. RESULTS: There were 2,014 participants who were randomly assigned. Median follow-up time as of December 31, 2019, was 79.2 months. At primary definitive analysis, 114 IBTR events occurred: RT arm, 63 and RT plus T arm, 51 (hazard ratio [HR], 0.81; 95% CI, 0.56 to 1.17; P value = .26). There were 34 who were invasive: RT arm, 18 and RT plus T arm, 20 (HR, 1.11; 95% CI, 0.59 to 2.10; P value = .71). Seventy-six were DCIS: RT arm, 45 and RT plus T arm, 31 (HR, 0.68; 95% CI, 0.43 to 1.08; P value = .11). Annual IBTR event rates were: RT arm, 0.99%/y and RT plus T arm, 0.79%/y. The study did not reach the 163 protocol-specified events, so the definitive analysis was triggered by all patients having been on study for ≥ 5 years. CONCLUSION: Addition of T to RT did not achieve the objective of 36% reduction in IBTR rate but did achieve a modest but statistically nonsignificant reduction of 19%. Nonetheless, this trial had negative results. Further exploration of RT plus T is needed in HER2-positive DCIS before its routine delivery in patients with DCIS resected by lumpectomy.

Interobserver Agreement of PD-L1/SP142 Immunohistochemistry and Tumor-Infiltrating Lymphocytes (TILs) in Distant Metastases of Triple-Negative Breast Cancer: A Proof-of-Concept Study. A Report on Behalf of the International Immuno-Oncology Biomarker Working Group.

Patients with advanced triple-negative breast cancer (TNBC) benefit from treatment with atezolizumab, provided that the tumor contains ≥1% of PD-L1/SP142-positive immune cells. Numbers of tumor-infiltrating lymphocytes (TILs) vary strongly according to the anatomic localization of TNBC metastases. We investigated inter-pathologist agreement in the assessment of PD-L1/SP142 immunohistochemistry and TILs. Ten pathologists evaluated PD-L1/SP142 expression in a proficiency test comprising 28 primary TNBCs, as well as PD-L1/SP142 expression and levels of TILs in 49 distant TNBC metastases with various localizations. Interobserver agreement for PD-L1 status (positive vs. negative) was high in the proficiency test: the corresponding scores as percentages showed good agreement with the consensus diagnosis. In TNBC metastases, there was substantial variability in PD-L1 status at the individual patient level. For one in five patients, the chance of treatment was essentially random, with half of the pathologists designating them as positive and half negative. Assessment of PD-L1/SP142 and TILs as percentages in TNBC metastases showed poor and moderate agreement, respectively. Additional training for metastatic TNBC is required to enhance interobserver agreement. Such training, focusing on metastatic specimens, seems worthwhile, since the same pathologists obtained high percentages of concordance (ranging from 93% to 100%) on the PD-L1 status of primary TNBCs.

Landscape of circulating tumour DNA in metastatic breast cancer.

BACKGROUND: We describe the genomic landscape of circulating tumour DNA (ctDNA) across pathological subtypes of metastatic breast cancer. METHODS: 255 clinically annotated patients with ctDNA testing by Guardant360 were stratified into HR+, HER2+, and TNBC cohorts. Frequency and heterogeneity of alterations were reported. Paired ctDNA and tissue sequencing were compared for a subset of patients. The association of ctDNA and metastatic sites of disease on imaging was also assessed. FINDINGS: 89% of patients had at least one ctDNA alteration detected. The most common single nucleotide variants (SNVs) for HR+ patients were PIK3CA, ESR1, and TP53. For HER2+, these were TP53, PIK3CA, and ERBB2 with ERBB2 as the most frequent copy number variant (CNV). For TNBC, the most common SNVs were TP53 and PIK3CA, and the most frequent CNVs were MYC, CCNE1, and PIK3CA. TNBC patients had a significantly higher mutant allele frequency (MAF) of the highest variant compared to HR+ or HER2+ patients (P<0.05). Overall, alterations in PIK3CA, ESR1, and ERBB2 were observed in 39.6%, 16.5%, and 21.6% of patients, respectively. Agreement between blood and tissue was 79-91%. MAF and number of alterations were significantly associated with number of metastatic sites on imaging (P<0.0001). INTERPRETATION: These data demonstrate the genetic heterogeneity of metastatic breast cancer in blood, the high prevalence of clinically actionable alterations, and the potential to utilise ctDNA as a surrogate for tumour burden on imaging. FUNDING: Lynn Sage Cancer Research Foundation, OncoSET Precision Medicine Program, and REDCap support was funded by the National Institutes of Health UL1TR001422.

The PD-1/PD-L1 Axis in HER2+ Ductal Carcinoma In Situ (DCIS) of the Breast.

OBJECTIVES: The aims were to evaluate the programmed death 1 (PD-1)/programmed death ligand 1 (PD-L1) axis in ductal carcinoma in situ (DCIS) of the breast. METHODS: We reviewed 85 pure DCIS cases treated with surgical excision at our institution, including 51 luminal A (estrogen receptor [ER] positive/human epidermal growth factor 2 [HER2] negative), 15 luminal B (ER+/HER2+), 13 HER2 (ER-/HER2+), and six basal-like (ER-/HER2-/CK5/6+). The extent and intensity of PD-1 and PD-L1 immunohistochemical staining in the tumor-infiltrating lymphocytes (TILs) and in the tumor cells were recorded. RESULTS: Our study found that moderate/severe inflammation around DCIS correlated with HER2 expression (20/28 HER2+ cases [71%] vs 21/57 HER2- cases [37%], P = .005). Of interest, over half of the TILs around the HER2 subtype expressed PD-L1 (7/13, 54%). In addition, about one-third of TILs around the HER2 subtype expressed PD-1 (4/13, 31%). CONCLUSIONS: These findings suggest that immune-based therapeutic strategies may be used as a potential therapy in DCIS cases with PD-L1 overexpression, especially those of the HER2 molecular subtype.