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Abstract Background The direct membrane feeding assay (DMFA), whereby gametocyte-infected blood is collected from human donors and from which mosquitoes feed through a membrane, is proving essential for assessing parameters influencing Plasmodium transmission potential in endemic countries. The success of DMFAs is closely tied to gametocyte density in the blood, with relatively high gametocytaemia ensuring optimal infection levels in mosquitoes. As transmission intensity declines with control efforts, the occurrence of asymptomatic individuals with low gametocyte densities, who can significantly contribute to the infectious reservoir, is increasing. This poses a limitation to studies relying on the experimental infection of large numbers of mosquitoes with natural isolates of Plasmodium. A simple, field-applicable method is presented for improving parasite infectivity by concentrating Plasmodium falciparum gametocytes. Methods Anopheles gambiae received one of the following 5 blood treatments through DMFA: (i) whole blood (WB) samples from naturally-infected donors; (ii) donor blood whose plasma was replaced with the same volume of Plasmodium-naive AB + serum (1:1 control); (iii) plasma replaced with a volume of malaria-naïve AB + serum equivalent to half (1:1/2), or to a quarter (1:1/4), of the initial plasma volume; and (v) donor blood whose plasma was fully removed (RBC). The experiment was repeated 4 times using 4 distinct wild parasite isolates. Seven days post-infection, a total of 1,095 midguts were examined for oocyst presence. Results Substituting plasma with reduced amounts (1:1/2 and 1:1/4) of Plasmodium-naive AB + serum led to a 31% and 17% increase of the mosquito infection rate and to a 85% and 308% increase in infection intensity compared to the 1:1 control, respectively. The full removal of plasma (RBC) reduced the infection rate by 58% and the intensity by 64% compared to the 1:1 control. Reducing serum volumes (1:1/2; 1:1/4 and RBC) had no impact on mosquito feeding rate and survival when compared to the 1:1 control. Conclusions Concentrating gametocytic blood by replacing natural plasma by lower amount of naive serum can enhance the success of mosquito infection. In an area with low gametocyte density, this simple and practical method of parasite concentration can facilitate studies on human-to-mosquito transmission such as the evaluation of transmission-blocking interventions.

Abstract Anopheles gambiae and Anopheles coluzzii mosquitoes, two major malaria vectors in sub‐Saharan Africa, exhibit selectivity among plant species as potential food sources. However, it remains unclear if their preference aligns with optimal nutrient intake and survival. Following an extensive screening of the effects of 31 plant species on An. coluzzii in Burkina Faso, we selected three species for their contrasting effects on mosquito survival, namely Ixora coccinea, Caesalpinia pulcherrima, and Combretum indicum. We assessed the sugar content of these plants and their impact on mosquito fructose positivity, survival, and insemination rate, using Anopheles coluzzii and Anopheles gambiae, with glucose 5% and water as controls. Plants displayed varying sugar content and differentially affected the survival, sugar intake, and insemination rate of mosquitoes. All three plants were more attractive to mosquitoes than controls, with An. gambiae being more responsive than An. coluzzii. Notably, C. indicum was the most attractive but had the lowest sugar content and offered the lowest survival, insemination rate, and fructose positivity. Our findings unveil a performance–preference mismatch in An. coluzzii and An. gambiae regarding plant food sources. Several possible reasons for this negative correlation between performance and preference are discussed.

Abstract Success in reducing malaria transmission through vector control is threatened by insecticide resistance in mosquitoes. Although the proximal molecular mechanisms and genetic determinants involved are well documented, little is known about the influence of the environment on mosquito resistance to insecticides. The aim of this study was to assess the effect of plant sugar feeding on the response of Anopheles gambiae sensu lato to insecticides. Adults were fed with one of four treatments, namely a 5% glucose control solution, nectariferous flowers of Barleria lupulina, of Cascabela thevetia and a combination of both B. lupulina + C. thevetia. WHO tube tests were performed with 0.05% and 0.5% deltamethrin, and knockdown rate (KD) and the 24 h mosquito mortality were measured. Plant diet significantly influenced mosquito KD rate at both concentrations of deltamethrin. Following exposure to 0.05% deltamethrin, the B. lupulina diet induced a 2.5 fold-increase in mosquito mortality compared to 5% glucose. Species molecular identification confirmed the predominance of An. gambiae (60% of the samples) over An. coluzzii and An. arabiensis in our study area. The kdr mutation L1014F displayed an allelic frequency of 0.75 and was positively associated with increased phenotypic resistance to deltamethrin. Plant diet, particularly B. lupulina, increased the susceptibility of mosquitoes to insecticides. The finding that B. lupulina-fed control individuals (i.e. not exposed to deltamethrin) also displayed increased 24 h mortality suggests that plant-mediated effects may be driven by a direct effect of plant diet on mosquito survival rather than indirect effects through interference with insecticide-resistance mechanisms. Thus, some plant species may weaken mosquitoes, making them less vigorous and more vulnerable to the insecticide. There is a need for further investigation, using a wider range of plant species and insecticides, in combination with other relevant environmental factors, to better understand the expression and evolution of insecticide resistance.

Abstract Background Besides feeding on blood, females of the malaria vector Anopheles gambiae sensu lato readily feed on natural sources of plant sugars. The impact of toxic secondary phytochemicals contained in plant-derived sugars on mosquito physiology and the development of Plasmodium parasites remains elusive. The focus of this study was to explore the influence of the alkaloid ricinine, found in the nectar of the castor bean Ricinus communis, on the ability of mosquitoes to transmit Plasmodium falciparum. Methods Females of Anopheles gambiae and its sibling species Anopheles coluzzii were exposed to ricinine through sugar feeding assays to assess the effect of this phytochemical on mosquito survival, level of P. falciparum infection and growth rate of the parasite. Results Ricinine induced a significant reduction in the longevity of both Anopheles species. Ricinine caused acceleration in the parasite growth rate with an earlier invasion of the salivary glands in both species. At a concentration of 0.04 g l−1 in An. coluzzii, ricinine had no effect on mosquito infection, while 0.08 g l−1 ricinine-5% glucose solution induced a 14% increase in An. gambiae infection rate. Conclusions Overall, our findings reveal that consumption of certain nectar phytochemicals can have unexpected and contrasting effects on key phenotypic traits that govern the intensity of malaria transmission. Further studies will be required before concluding on the putative role of ricinine as a novel control agent, including the development of ricinine-based toxic and transmission-blocking sugar baits. Testing other secondary phytochemicals in plant nectar will provide a broader understanding of the impact which plants can have on the transmission of vector-borne diseases. Graphical abstract

The spread of insecticide resistance in Anopheles mosquitoes and drug resistance in Plasmodium parasites is contributing to a global resurgence of malaria, making the generation of control tools that can overcome these roadblocks an urgent public health priority. We recently showed that the transmission of Plasmodium falciparum parasites can be efficiently blocked when exposing Anopheles gambiae females to antimalarials deposited on a treated surface, with no negative consequences on major components of mosquito fitness. Here, we demonstrate this approach can overcome the hurdles of insecticide resistance in mosquitoes and drug resistant in parasites. We show that the transmission-blocking efficacy of mosquito-targeted antimalarials is maintained when field-derived, insecticide resistant Anopheles are exposed to the potent cytochrome b inhibitor atovaquone, demonstrating that this drug escapes insecticide resistance mechanisms that could potentially interfere with its function. Moreover, this approach prevents transmission of field-derived, artemisinin resistant P. falciparum parasites (Kelch13 C580Y mutant), proving that this strategy could be used to prevent the spread of parasite mutations that induce resistance to front-line antimalarials. Atovaquone is also highly effective at limiting parasite development when ingested by mosquitoes in sugar solutions, including in ongoing infections. These data support the use of mosquito-targeted antimalarials as a promising tool to complement and extend the efficacy of current malaria control interventions.

Abstract Despite their recognised role in HER2-positive (HER2+) breast cancer (BC), the composition, localisation and functional orientation of immune cells within tumour microenvironment, as well as its dynamics during anti-HER2 treatment, is largely unknown. We here investigate changes in tumour-immune contexture, as assessed by stromal tumour-infiltrating lymphocytes (sTILs) and by multiplexed spatial cellular phenotyping, during treatment with lapatinib-trastuzumab in HER2+ BC patients (PAMELA trial). Moreover, we evaluate the relationship of tumour-immune contexture with hormone receptor status, intrinsic subtype and immune-related gene expression. sTIL levels increase after 2 weeks of HER2 blockade in HR-negative disease and HER2-enriched subtype. This is linked to a concomitant increase in cell density of all four immune subpopulations (CD3+, CD4+, CD8+, Foxp3+). Moreover, immune contexture analysis showed that immune cells spatially interacting with tumour cells have the strongest association with response to anti-HER2 treatment. Subsequently, sTILs consistently decrease at the surgery in patients achieving pathologic complete response, whereas most residual tumours at surgery remain inflamed, possibly reflecting a progressive loss of function of T cells. Understanding the features of the resulting tumour immunosuppressive microenvironment has crucial implications for the design of new strategies to de-escalate or escalate systemic therapy in early-stage HER2+ BC.

Background: Identification of biomarkers to optimize treatment strategies for early-stage triple-negative breast cancer (TNBC) is crucial. This study presents the development and validation of TNBC-DX, a novel test aimed at predicting both short- and long-term outcomes in early-stage TNBC. The objective of this study was to evaluate the association between TNBC-DX and efficacy outcomes [pathologic complete response (pCR), distant disease-free survival (DDFS) or event-free survival (EFS), and overall survival (OS)] in the validation cohorts., Methods: Information from 1259 patients with early-stage TNBC (SCAN-B, CALGB-40603, and BrighTNess) was used to establish the TNBC-DX scores. Independent validation of TNBC-DX was carried out in three studies: (i) WSG-ADAPT-TN; (ii) MMJ-CAR-2014-01; and (iii) NeoPACT, including 527 patients with stage I-III TNBC undergoing neoadjuvant chemotherapy. In WSG-ADAPT-TN, patients were randomized to receive nab-paclitaxel plus gemcitabine or carboplatin. In MMJ-CAR-2014-01, patients received carboplatin plus docetaxel. In NeoPACT, patients received carboplatin plus docetaxel and pembrolizumab., Results: TNBC-DX test was created incorporating the 10-gene Core Immune Gene module, the 4-gene tumor cell proliferation signature, tumor size, and nodal staging. In the two independent validation cohorts without pembrolizumab, the TNBC-DX pCR score was significantly associated with pCR after adjustment for clinicopathological variables and treatment regimen [odds ratio per 10-unit increment 1.34, 95% confidence interval (CI) 1.20-1.52, P < 0.001]. pCR rates for the TNBC-DX pCR-high, pCR-medium, and pCR-low categories were 56.3%, 53.6%, and 22.5% respectively (odds ratio for pCR-high versus pCR-low 3.48, 95% CI 1.72-7.15, P < 0.001). In addition, the TNBC-DX risk score was significantly associated with DDFS [hazard ratio (HR) high-risk versus low-risk 0.24, 95% CI 0.15-0.41, P < 0.001] and OS (HR 0.19, 95% CI 0.11-0.35, P < 0.001). In the validation cohort with pembrolizumab, the TNBC-DX scores were significantly associated with pCR, EFS, and OS., Conclusions: TNBC-DX predicts pCR to neoadjuvant taxane-carboplatin in stage I-III TNBC and helps to forecast the patient's long-term survival in the absence of neoadjuvant anthracycline-cyclophosphamide, and independent of pembrolizumab use., (Copyright © 2024 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Purpose: The purpose of this study was to assess the predictive capability of HER2DX assay following (neo)adjuvant trastuzumab-pertuzumab (HP)-based therapy in HER2-positive (HER2+) early breast cancer., Experimental Design: HER2DX was analyzed in baseline pretreatment tumors from the PHERGain trial. Patients with stage I-IIIA HER2+ early breast cancer were randomized to group A [docetaxel, carboplatin, and HP (TCHP)] and group B (HP ± endocrine therapy). PET response was evaluated after two cycles. Group A received TCHP for six cycles regardless of PET response. Group B continued with HP ± endocrine therapy for six cycles (PET responders) or with TCHP for six cycles (PET nonresponders). The primary objective of this retrospective study was to associate the HER2DX pathologic complete response (pCR) score with pCR. The secondary objective was the association of the HER2DX risk score with 3-year invasive disease-free survival (iDFS)., Results: HER2DX was performed on 292 (82.0%) tumors. The overall pCR rate was 38.0%, with pCR rates of 56.4% in group A and 33.8% in group B. In multivariable analysis including treatment and clinicopathologic factors, the HER2DX pCR score (continuous variable) significantly correlated with pCR [OR, 1.29; 95% confidence interval (CI), 1.10-1.54; P < 0.001]. HER2DX-defined pCR-high, -med, and -low groups exhibited pCR rates of 50.4%, 35.8%, and 23.2%, respectively (pCR-high vs. pCR-low OR, 3.27; 95% CI, 1.54-7.09; P < 0.001). In patients with residual disease, the HER2DX high-risk group demonstrated numerically worse 3-year iDFS than the low-risk group (89.8% vs. 100%; HR, 2.70; 95% CI, 0.60-12.18; P = 0.197)., Conclusions: HER2DX predicts pCR in the context of neoadjuvant HP-based therapy, regardless of chemotherapy addition, and might identify patients at higher risk of recurrence among patients with residual disease., (©2024 The Authors; Published by the American Association for Cancer Research.)

Purpose: This study aimed to investigate the current therapeutic management of patients with early-stage HER2-positive (HER2+) breast cancer in Spain, while also exploring the perceptions surrounding HER2DX in terms of its credibility, clinical relevance, and impact on therapeutic decision-making. Understanding these aspects is crucial for optimizing treatment strategies and enhancing patient outcomes in the context of HER2+ breast cancer., Methods: An online questionnaire was conducted by an independent third-party between April and May 2022 across 70 medical oncologists highly specialized in breast cancer management in Spain. The survey included 37 questions regarding treatment decision making in HER2+ early breast cancer., Results: The management of patients with HER2+ early breast cancer exhibited a high degree of heterogeneity. Among the interviewed oncologists, 53% would recommend upfront surgery for node negative tumors measuring 1 cm or less. Interestingly, 69% and 56% of interviewers were open to deescalate the duration of adjuvant trastuzumab in pT1a and pT1b N0 tumors, respectively. Certain clinicopathological characteristics, such as high grade, high Ki-67, and young age, influenced the decision to prescribe neoadjuvant treatment for patients with clinical stage 1 disease. In cases where neoadjuvant treatment was prescribed for cT1-2 N0 tumors, there was a wide variation in the choice of chemotherapeutic and anti-HER2 regimens. Regarding the use of adjuvant trastuzumab emtansine (T-DM1) in patients with residual disease after neoadjuvant therapy, there was diversity in practice, and a common concern emerged that T-DM1 might be overtreating some patients. HER2DX, as a diagnostic tool, was deemed trustworthy, and the reported scores were considered clinically useful. However, 86% of interviewees believed that a prospective trial was necessary before fully integrating the test into routine clinical practice., Conclusion: In the context of early-stage HER2+ breast cancer in Spain, a notable diversity in therapeutic approaches was observed. The majority of interviewed medical oncologists acknowledged HER2DX as a clinically valuable test for specific patients, in line with the 2022 SEOM-GEICAM-SOLTI clinical guidelines for early-stage breast cancer. To facilitate the full integration of HER2DX into clinical guidelines, conducting prospective studies to further validate its efficacy and utility was recommended., (© 2024. The Author(s).)

Arbuscular mycorrhizal (AM) fungi are difficult to manipulate and observe due to their permanent association with plant roots and propagation in the rhizosphere. Typically, AM fungi are cultured under in vivo conditions in pot culture with an autotrophic host or under in vitro conditions with Ri Transfer-DNA transformed roots (heterotrophic host) in a Petri dish. Additionally, the cultivation of AM fungi in pot culture occurs in an opaque and non-sterile environment. In contrast, in vitro culture involves the propagation of AM fungi in a sterile, transparent environment. The superabsorbent polymer-based autotrophic system (SAP-AS) has recently been developed and shown to combine the advantages of both methods while avoiding their respective limitations (opacity and heterotrophic host, sterility). Here, we present a detailed protocol for easy preparation, single spore inoculation, and observation of AM fungi in SAP-AS. By modifying the Petri dishes, high-resolution photographic and video observations were possible on living specimens, which would have been difficult or impossible with current in vivo and in vitro techniques.

Rhizophagus irregularis is the model species for arbuscular mycorrhizal fungi (AMF) research and the most widely propagated species for commercial plant biostimulants. Using asymbiotic and symbiotic cultivation systems initiated from single spores, advanced microscopy, Sanger sequencing of the glomalin gene, and PacBio sequencing of the partial 45S rRNA gene, we show that four strains of R. irregularis produce spores of two distinct morphotypes, one corresponding to the morphotype described in the R. irregularis protologue and the other having the phenotype of R. fasciculatus. The two spore morphs are easily distinguished by spore colour, thickness of the subtending hypha, thickness of the second wall layer, lamination of the innermost layer, and the dextrinoid reaction of the two outer spore wall layers to Melzer's reagent. The glomalin gene of the two spore morphs is identical and that of the PacBio sequences of the partial SSU-ITS-LSU region (2780 bp) obtained from single spores of the R. cf fasciculatus morphotype has a median pairwise similarity of 99.8% (SD = 0.005%) to the rDNA ribotypes of R. irregularis DAOM 197198. Based on these results, we conclude that the model AMF species R. irregularis is dimorphic, which has caused taxonomic confusion in culture collections and possibly in AMF research., (© 2023 His Majesty the King in Right of Canada and The Authors. New Phytologist published by John Wiley & Sons Ltd on behalf of New Phytologist Foundation. Reproduced with the permission of the Minister of Agriculture and Agri‐Food Canada.)

Chryseobacterium sp. MHB01, Rhodococcus qingshengii MHB02, and Agrobacterium tumefaciens MHB03 were isolated from superabsorbent polymer granules cultured with an arbuscular mycorrhizal fungus. Whole-genome sequencing of these three strains revealed genome sizes of 4.57 Mb, 7.13 Mb, and 5.49 Mb with G + C contents of 36.9%, 62.5%, and 58.2%, respectively., Competing Interests: The authors declare no conflict of interest.

Background: Early-stage triple-negative breast cancer (TNBC) displays clinical and biological diversity. From a biological standpoint, immune infiltration plays a crucial role in TNBC prognosis. Currently, there is a lack of genomic tools aiding in treatment decisions for TNBC. This study aims to assess the effectiveness of a B-cell/immunoglobulin signature (IGG) alone, or in combination with tumor burden, in predicting prognosis and treatment response in patients with TNBC., Methods: Genomic and clinical data were retrieved from 7 cohorts: SCAN-B (N = 874), BrighTNess (n = 482), CALGB-40603 (n = 389), METABRIC (n = 267), TCGA (n = 118), GSE58812 (n = 107), GSE21653 (n = 67). IGG and a risk score integrating IGG with tumor/nodal staging (IGG-Clin) were assessed for event-free survival (EFS) and overall survival (OS) in each cohort. Random effects model was used to derive pooled effect sizes. Association of IGG with pathological complete response (pCR) was assessed in CALGB-40603 and BrighTNess. Immune significance of IGG was estimated through CIBERSORTx and EcoTyper., Findings: IGG was associated with improved EFS (pooled HR = 0.77, [95% CI = 0.70-0.85], I 2  = 18%) and OS (pooled HR = 0.79, [0.73-0.85], I 2  = 0%) across cohorts, and was predictive of pCR in CALGB-40603 (OR 1.25, [1.10-1.50]) and BrighTNess (OR 1.57 [1.25-1.98]). IGG-Clin was predictive of recurrence (pooled HR = 2.11, [1.75-2.55], I 2  = 0%) and death (pooled HR = 1.99, 95% [0.84-4.73], I 2  = 79%) across cohorts. IGG was associated with adaptive immune response at CIBERSORTx and EcoTyper analysis., Interpretation: IGG is linked to improved prognosis and pCR in early-stage TNBC. The integration of IGG alongside tumor and nodal staging holds promise as an approach to identify patients benefitting from intensified or de-intensified treatments., Funding: This study received funding from: Associació Beca Marta Santamaria, European Union's Horizon 2020 research and innovation and Marie Skłodowska-Curie Actions programs, Fundación FERO, Fundación CRIS contra el cáncer, Agència de Gestó d'Ajuts Universitaris i de Recerca, Instituto de Salud Carlos III, Fundación Contigo, Asociación Cáncer de Mama Metastásico IV, Breast Cancer Research Foundation, RESCUER, Fundación científica AECC and FSEOM., Competing Interests: Declaration of interests B. Conte reports speaker fees from Veracyte and payment for educational events from Medsite and Novartis. A. Prat reports consulting fees from Roche, Pfizer, Novartis, Amgen, BMS, Puma, Oncolytics Biotech, MSD, Guardant Health, Peptomyc and Lilly, lecture fees from Roche, Pfizer, Novartis, Amgen, BMS, Nanostring Technologies and Daiichi Sankyo; patents filed PCT/EP2016/080056, PCT/EP2022/086493, PCT/EP2023/060810, EP23382703 and EP23383369; stockholder and consultant of Reveal Genomics, SL; and institutional financial interests from Boehringer, Novartis, Roche, Nanostring, Sysmex Europa GmbH, Medica Scientia Innovation Research, SL, Celgene, Astellas and Pfizer. F. Schettini has declared consulting fees from Pfizer, honoraria for lectures from Novartis, Gilead and Daiichy Sankyo, and travel expenses from Novartis and Daiichy Sankyo. O. Martínez-Sáez has declared institutional grants from Roche; personal consulting fees from Roche and Reveal Genomics; honoraria for presentations by Daiichi Sankyo, Pierre Fabre, and Reveal Genomics; and travel expenses by Gilead, Pierre Fabre, Novartis, and MSD. A. Vivancos has declared institutional grants from Bristol Meyers Squibb, Incyte, and Roche; personal consulting fees from Bayer, Bristol Meyers Squibb (BMS); Guardant, Incyte, and Roche; and personal stock options from Reveal Genomics. F. Brasó-Maristany has patents filed: PCT/EP2022/086493, PCT/EP2023/060810, EP23382703 and EP23383369. J. Parker as declared individual and institutional royalties from Veracyte–Prosigna, consulting fees from Bristol Meyers Squibb, Reveal Genomics, and GeneCentric, and holds a patent for breast cancer subtyping. Additionally, he has an equity interest in Reveal Genomics. M. Vidal has declared honoraria for presentations from Novartis, Roche, Pfizer, and Daichii, and travel expenses from Roche and Pfizer. Additionally, she has participated on a Data Safety Monitoring Board or Advisory Board for Novartis and Roche. C. Perou has declared consulting fees and personal stock options from Reveal Genomics. T. Pascual has declared consulting fees from Novartis; honoraria from Novartis, Astra-zeneca, Veracyte, and Argenetics. I. Garcia-Fructuoso has declared honoraria for presentations from Novartis, Daiichi Sankyo, Esteve, GSK; and travel expenses from Novartis, Gilead, Daiichi Sankyo, Lilly, and BMS. L. Paré has declared contract from Reveal Genomics, a HER2DX patent filed (PCT/EP2022/086493), and the TNBCDX patent filed (EP23382703.9). M. Marín-Aguilera has declared contract from Reveal Genomics. P. Villagrasa has declared contract and personal stock options from Reveal Genomics, the HER2DX patent filed (PCT/EP2022/086493), and the DNADX patent filed (EP22382387.3). G. Villacampa has received a speaker's fee from Merck Sharp & Dohme, Pfizer, GlaxoSmithKline and Pierre Fabrer, and received consultant fees from Reveal Genomics. C. Falato is currently employed in AstraZeneca. The remaining authors declare no competing interests., (Copyright © 2024 The Author(s). Published by Elsevier B.V. All rights reserved.)

In this study, we performed genomic analyses of cell cycle and tumor microenvironment changes during and after ribociclib and letrozole or chemotherapy in the CORALLEEN trial. 106 women with untreated PAM50-defined Luminal B early breast cancers were randomly assigned to receive neoadjuvant ribociclib and letrozole or standard-of-care chemotherapy. Ki67 immunohistochemistry, tumor-infiltrating lymphocytes quantification, and RNA sequencing were obtained from tissue biopsies pre-treatment, on day 14 of treatment, and tumor specimens from surgical resection. Results showed that at surgery, Ki67 and the PAM50 proliferation scores were lower after ribociclib compared to chemotherapy. However, consistent reactivation of tumor cell proliferation from day 14 to surgery was only observed in the ribociclib arm. In tumors with complete cell cycle arrest (CCCA) at surgery, PAM50 proliferation scores were lower in the ribociclib arm compared to chemotherapy (p < 0.001), whereas the opposite was observed with tumor cellularity (p = 0.002). Gene expression signatures (GES) associated with antigen-presenting cells (APCs) and innate immune system activity showed increased expression post-chemotherapy but decreased expression post-ribociclib. Interferon-associated GES had decreased expression with CCCA and increased expression with non-CCCA. Our findings suggest that while both treatment strategies decreased proliferation, the depth and the patterns over time differed by treatment arm. Immunologically, ribociclib was associated with downregulated GES associated with APCs and the innate immune system in Luminal B tumors, contrary to existing preclinical data. Further studies are needed to understand the effect of CDK4/6 inhibition on the tumor cells and microenvironment, an effect which may vary according to tumor subtypes., (© 2024. The Author(s).)

Background: HER2DX, a multianalyte genomic test, has been clinically validated to predict breast cancer recurrence risk (relapse risk score), the probability of achieving pathological complete response post-neoadjuvant therapy (pCR likelihood score), and individual ERBB2 messenger RNA (mRNA) expression levels in patients with early-stage human epidermal growth factor receptor 2 (HER2)-positive breast cancer. This study delves into the comprehensive analysis of HER2DX's analytical performance., Materials and Methods: Precision and reproducibility of HER2DX risk, pCR, and ERBB2 mRNA scores were assessed within and between laboratories using formalin-fixed paraffin-embedded (FFPE) tumor tissues and purified RNA. Robustness was appraised by analyzing the impact of tumor cell content and protocol variations including different instruments, reagent lots, and different RNA extraction kits. Variability was evaluated across intratumor biopsies and genomic platforms [RNA sequencing (RNAseq) versus nCounter], and according to protocol variations., Results: Precision analysis of 10 FFPE tumor samples yielded a maximal standard error of 0.94 across HER2DX scores (1-99 scale). High reproducibility of HER2DX scores across 29 FFPE tumors and 20 RNAs between laboratories was evident (correlation coefficients >0.98). The probability of identifying score differences >5 units was ≤5.2%. No significant variability emerged based on platform instruments, reagent lots, RNA extraction kits, or TagSet thaw/freeze cycles. Moreover, HER2DX displayed robustness at low tumor cell content (10%). Intratumor variability across 212 biopsies (106 tumors) was <4.0%. Concordance between HER2DX scores from 30 RNAs on RNAseq and nCounter platforms exceeded 90.0% (Cohen's κ coefficients >0.80)., Conclusions: The HER2DX assay is highly reproducible and robust for the quantification of recurrence risk, pCR likelihood, and ERBB2 mRNA expression in early-stage HER2-positive breast cancer., Competing Interests: Disclosure MMA is an employee at Reveal Genomics; GV has received a speaker’s fee from MSD, Pfizer, GSK, and Pierre Fabrer, has held an advisory role with AstraZeneca, and received consultant fees from Reveal Genomics; FBM has a HER2DX patent application EP21383165; PG is an employee at Reveal Genomics; OMS has declared travel expenses and consulting fees from Roche and Reveal, and speaker fees from Eisai, Daiichi, and Novartis; KP serves on the Scientific Advisory Board of Novartis, Ideaya Biosciences, and Scorpion Therapeutics, holds equity options in Scorpion Therapeutics and Ideaya Biosciences, and receives sponsored research funding from Novartis through DFCI; and has patents on S100A7 antibody and BET inhibitor resistance held by DFCI; JM is an employee at Reveal Genomics; WB is an employee at Reveal Genomics; CMP is an equity stockholder and consultant of BioClassifier LLC, and for Reveal Genomics, is also listed as an inventor on patent applications for the Breast PAM50 assay; PVG is one of the stockholders of Reveal Genomics; AP reports advisory and consulting fees from Roche, Pfizer, Novartis, Amgen, BMS, Puma, Oncolytics Biotech, MSD, Guardant Health, Peptomyc, and Lilly, lecture fees from Roche, Pfizer, Novartis, Amgen, BMS, and Daiichi Sankyo, institutional financial interests from Novartis, Roche, and Pfizer, stockholder and consultant of Reveal Genomics, SL; AP is also listed as an inventor on patent applications for the HER2DX assay; JSP is an equity stockholder and consultant for Reveal Genomics and is also listed as an inventor on patent applications for the Breast PAM50 assay; LP is listed as an inventor on HER2DX patent PCT/EP2021/070788 and is an employee at Reveal Genomics. All other authors have declared no conflicts of interest., (Copyright © 2024 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Background: The HER2DX risk-score has undergone rigorous validation in prior investigations involving patients with early-stage human epidermal growth factor receptor 2 (HER2)-positive (HER2+) breast cancer. In this study, we present the outcomes of the HER2DX risk-score within the most recent release of the Sweden Cancerome Analysis Network-Breast (SCAN-B) HER2+ cohort. This updated examination benefits from a larger patient sample, an extended follow-up duration, and detailed treatment information., Materials and Methods: Clinical and RNAseq data from the SCAN-B dataset were retrieved from Gene Expression Omnibus (GSE81538). Among the 6600 patients, 819 had HER2+ breast cancer, with 757 individuals with research-based HER2DX risk-scores and corresponding survival outcomes. The HER2DX risk-score was evaluated (i) as a continuous variable and (ii) using predefined cut-offs. The primary endpoint for this study was overall survival (OS). The Kaplan-Meier method and Cox models were used to estimate OS and a multistate model with four states was fitted to better characterize patients' follow-up., Results: The median follow-up time was 7.5 years (n = 757). The most common systemic therapy was chemotherapy with trastuzumab (82.0%) and most tumors were classified as T1-T2 (97.1%). The HER2DX risk-score as a continuous variable was significantly associated with OS after adjustment for clinical variables and treatment regimen [hazard ratios (HR) per 10-unit increment = 1.31, 95% confidence interval (CI) 1.13-1.51, P < 0.001] as well as within predefined risk groups (high versus low; HR = 2.57, 95% CI 1.36-4.85, P < 0.001). Patients classified as HER2DX high-risk also had higher risk of (i) breast cancer recurrence and (ii) death without previous recurrence. Within the subgroup of HER2+ T1N0 tumors (n = 297), those classified as high-risk demonstrated inferior OS compared to low-risk tumors (7-year OS 77.8% versus 96.8%, P < 0.001). The HER2DX mRNA ERBB2 score was associated with clinical HER2 status (area under the receiver operating characteristic curve = 0.91)., Conclusions: In patients with early-stage HER2+ breast cancer, HER2DX risk-score provides prognostic information beyond clinicopathological variables, including treatment regimen with or without trastuzumab., Competing Interests: Role of the funder In addition to financial support, the funders actively contributed to various aspects of the research, including study design, data collection, analysis, interpretation, and report writing. Disclosure GV has received a speaker’s fee from MSD, Pfizer, GSK, and Pierre Fabre; has held an advisory role with AstraZeneca; and received consultant fees from Reveal Genomics. TP has received honoraria for speaker activities from AstraZeneca, Pfizer, Novartis, Veracyte, and Argenetics, and has held an advisory role with Novartis. FBM has a patent application (EP21383165). LP is listed as an inventor on patent PCT/EP2021/070788. OMS has declared travel expenses and consulting fees from Roche and Reveal, and speaker fees from Eisai, Daiichi-Sankyo, and Novartis. JC has ownership interests in MedSIR, Nektar, and Leuko; has held consulting or advisory roles in Celgene, Cellestia Biotech, AstraZeneca, Roche, Seattle Genetics, Daiichi-Sankyo, ERYTECH Pharma, Polyphor, Athenex, Lilly, Servier, Merck Sharp & Dohme, GlaxoSmithKline, Leuko, Clovis Oncology, Bioasis, Boehringer Ingelheim, Ellipses Pharma, HiberCell, Bioinvent, GEMoaB, Gilead Sciences, Menarini, Zymeworks, and Reveal Genomics; has received research funding from ARIAD, AstraZeneca, Baxalta, Bayer, Eisai, Guardant Health, Merck Sharp & Dohme, Pfizer, Puma Biotechnology, Queen Mary University of London, Roche, and Piqur; has patents, royalties, or other intellectual property from pharmaceutical combinations of a Pi3k Inhibitor and a microtubule destabilizing agent (Javier Cortés Castán, Alejandro Piris Giménez, Violeta Serra Elizalde. WO 2014/199294 A, Her2 as a predictor of response to dual HER2 blockade in the absence of cytotoxic therapy. Aleix Prat, Antonio Llombart, Javier Cortés. US 2019/ 0338368 A1); and has received travel expenses from Roche, Pfizer, Eisai, Novartis, Daiichi-Sankyo, and Gilead Sciences. EC has received honoraria for advisory roles from Pfizer, AstraZeneca, Daiichi-Sankyo, Roche, Novartis, Lilly, and MSD; and has received a speaker’s fee from Roche and Lilly. MM reports research grants from Puma; consulting/advisory fees from Roche, Novartis, AstraZeneca, Daiichi-Sankyo, Seagen, Lilly, and Sanofi; speakers’ honoraria from Seagen, Lilly, AstraZeneca, Pfizer, Daiichi-Sankyo, and Roche; a leadership or fiduciary role as Chairman, GEICAM and as a member of the Board of Directors for TRIO. PC reports speakers’ bureaus with AstraZeneca, GlaxoSmithKline, Novartis, and Roche; travel, accommodation, and expenses from Celgene, GlaxoSmithKline, and Novartis; and research funding from Merck Serono, Novartis, and Roche. LAC reports participation on a Data Safety Monitoring Board or Advisory Board of Sanofi Aventis, Novartis, Genentech/Roche, GSK, AstraZeneca/Daiichi-Sankyo, and Aptitude Health; and has a spouse serving on the board of Falcon Therapeutics and spouse involvement in a neural stem cell therapy patent. NH received consulting fees from Pierre Fabre and Roche. AV has the DNADX patent filed (EP22382387.3). GC served as consultant or advisor for Roche, Lilly, and Bristol-Myers Squibb; served on the speaker’s bureau for Roche, Pfizer, and Lilly; received travel funding from Pfizer and Roche; and received honoraria from Roche, Pfizer, Lilly, Novartis, and Seagen, all outside the submitted work. PV is one of the stockholders of Reveal Genomics, and also reports personal fees from NanoString. JSP is an equity stockholder and consultant for Reveal Genomics and is listed as an inventor on patent applications for the Breast PAM50 assay. CMP is an equity stockholder and consultant of BioClassifier LLC and Reveal Genomics, and is listed as an inventor on patent applications for the Breast PAM50 assay. AP reports advisory and consulting fees from Roche, Pfizer, Novartis, Amgen, Bristol-Myers Squibb, Puma, Oncolytics Biotech, MSD, Guardant Health, Peptomyc, and Lilly; lecture fees from Roche, Pfizer, Novartis, Amgen, Bristol-Myers Squibb, NanoString Technologies, and Daiichi-Sankyo; institutional financial interests from Boehringer, Novartis, Roche, NanoString, Sysmex Europa GmbH, Medica Scientia Innovation Research, SL, Celgene, Astellas, and Pfizer; is a stockholder and consultant of Reveal Genomics and SL; and is listed as an inventor on patent applications for the HER2DX assay. SMT reports consulting or advisory roles for Novartis, Pfizer, Merck, Eli Lilly, AstraZeneca, Genentech/Roche, Eisai, Sanofi, Bristol-Myers Squibb, Seattle Genetics, CytomX Therapeutics, Daiichi-Sankyo, Gilead, OncXerna, Zymeworks, Zentalis, Blueprint Medicines, Reveal Genomics, ARC Therapeutics, Infinity Therapeutics, Sumitovant Biopharma, Zetagen, Umoja Biopharma, Artios Pharma, Menarini/Stemline, Aadi Biopharma, Bayer, Incyte Corp, Jazz Pharmaceuticals, Natera, Tango Therapeutics, Systimmune, eFFECTOR, and Hengrui USA; research funding from Genentech/Roche, Merck, Exelixis, Pfizer, Lilly, Novartis, Bristol-Myers Squibb, Eisai, AstraZeneca, Gilead, NanoString Technologies, Seattle Genetics, and OncoPep; and travel support from Eli Lilly, Sanofi, Gilead, and Pfizer. All other authors have declared no conflicts of interest., (Copyright © 2024 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Background: Even with contemporary treatment strategies, more than 10% of HER2-positive early stage breast cancer patients may experience distant metastasis as first event during follow-up. Tools for predicting unique patterns of metastatic spread are needed to plan personalized surveillance. We evaluated how molecular heterogeneity affects the pattern of distant relapse in HER2-positive breast cancer., Methods: A total of 677 HER2-positive stage I-III breast cancer patients from ShortHER trial, Cher-LOB trial, and 2 institutional cohorts were included. PAM50 molecular subtypes and research-based HER2DX scores were evaluated. The cumulative incidence of distant relapse as the first event (any site and site specific) was evaluated using competing risk analysis. Median follow-up was 8.4 years. Tests of statistical significance are 2-sided., Results: Stage III and high HER2DX risk score identified patients at the highest risk of distant relapse as first event (10-year incidence 24.5% and 19.7%, respectively). Intrinsic molecular subtypes were associated with specific patterns of metastatic spread: compared with other subtypes, HER2-enriched tumors were more prone to develop brain metastases (10-year incidence 3.8% vs 0.6%, P = .005), basal-like tumors were associated with an increased risk of lung metastases (10-year incidence 11.1% vs 2.6%, P = .001), and luminal tumors developed more frequently bone-only metastases (10-year incidence 5.1% vs 2.0%, P = .042). When added to stage or HER2DX risk score in competing risk regression models, intrinsic subtype maintained an independent association with site-specific metastases., Conclusions: The integration of intrinsic molecular subtypes with stage or HER2DX risk score predicts site-specific metastatic risk in HER2-positive breast cancer, with potential implications for personalized surveillance and clinical trials aimed at preventing site-specific recurrence., (© The Author(s) 2023. Published by Oxford University Press.)

Background: Restorative regeneration, the capacity to reform a lost body part following amputation or injury, is an important and still poorly understood process in animals. Annelids, or segmented worms, show amazing regenerative capabilities, and as such are a crucial group to investigate. Elucidating the molecular mechanisms that underpin regeneration in this major group remains a key goal. Among annelids, the nereididae Platynereis dumerilii (re)emerged recently as a front-line regeneration model. Following amputation of its posterior part, Platynereis worms can regenerate both differentiated tissues of their terminal part as well as a growth zone that contains putative stem cells. While this regeneration process follows specific and reproducible stages that have been well characterized, the transcriptomic landscape of these stages remains to be uncovered., Results: We generated a high-quality de novo Reference transcriptome for the annelid Platynereis dumerilii. We produced and analyzed three RNA-sequencing datasets, encompassing five stages of posterior regeneration, along with blastema stages and non-amputated tissues as controls. We included two of these regeneration RNA-seq datasets, as well as embryonic and tissue-specific datasets from the literature to produce a Reference transcriptome. We used this Reference transcriptome to perform in depth analyzes of RNA-seq data during the course of regeneration to reveal the important dynamics of the gene expression, process with thousands of genes differentially expressed between stages, as well as unique and specific gene expression at each regeneration stage. The study of these genes highlighted the importance of the nervous system at both early and late stages of regeneration, as well as the enrichment of RNA-binding proteins (RBPs) during almost the entire regeneration process., Conclusions: In this study, we provided a high-quality de novo Reference transcriptome for the annelid Platynereis that is useful for investigating various developmental processes, including regeneration. Our extensive stage-specific transcriptional analysis during the course of posterior regeneration sheds light upon major molecular mechanisms and pathways, and will foster many specific studies in the future., (© 2023. BioMed Central Ltd., part of Springer Nature.)

Background: The HER2DX genomic test predicts pathological complete response (pCR) and survival outcome in early-stage HER2-positive (HER2+) breast cancer. Here, we evaluated the association of HER2DX scores with (i) pCR according to hormone receptor status and various treatment regimens, and (ii) survival outcome according to pCR status., Materials and Methods: Seven neoadjuvant cohorts with HER2DX and clinical individual patient data were evaluated (DAPHNe, GOM-HGUGM-2018-05, CALGB-40601, ISPY-2, BiOnHER, NEOHER and PAMELA). All patients were treated with neoadjuvant trastuzumab (n = 765) in combination with pertuzumab (n = 328), lapatinib (n = 187) or without a second anti-HER2 drug (n = 250). Event-free survival (EFS) and overall survival (OS) outcomes were available in a combined series of 268 patients (i.e. NEOHER and PAMELA) with a pCR (n = 118) and without a pCR (n = 150). Cox models were adjusted to evaluate whether HER2DX can identify patients with low or high risk beyond pCR status., Results: HER2DX pCR score was significantly associated with pCR in all patients [odds ratio (OR) per 10-unit increase = 1.59, 95% confidence interval 1.43-1.77; area under the ROC curve = 0.75], with or without dual HER2 blockade. A statistically significant increase in pCR rate due to dual HER2 blockade over trastuzumab-only was observed in HER2DX pCR-high tumors treated with chemotherapy (OR = 2.36 (1.09-5.42). A statistically significant increase in pCR rate due to multi-agent chemotherapy over a single taxane was observed in HER2DX pCR-medium tumors treated with dual HER2 blockade (OR = 3.11, 1.54-6.49). The pCR rates in HER2DX pCR-low tumors were ≤30.0% regardless of treatment administered. After adjusting by pCR status, patients identified as HER2DX low-risk had better EFS (P < 0.001) and OS (P = 0.006) compared with patients with HER2DX high-risk., Conclusions: HER2DX pCR score and risk score might help identify ideal candidates to receive neoadjuvant dual HER2 blockade in combination with a single taxane in early-stage HER2+ breast cancer., (Copyright © 2023 European Society for Medical Oncology. Published by Elsevier Ltd. All rights reserved.)

Background: Patritumab deruxtecan (HER3-DXd) is a human epidermal growth factor receptor 3 (HER3)-directed antibody-drug conjugate composed of a fully human anti-HER3 monoclonal antibody (patritumab) covalently linked to a topoisomerase I inhibitor payload via a stable, tumor-selective, tetrapeptide-based cleavable linker. TOT-HER3 is a window-of-opportunity study designed to assess the biological activity, measured by CelTIL score [= -0.8 × tumor cellularity (in %)  +  1.3  × tumor-infiltrating lymphocytes (TILs) (in %)], and clinical activity of HER3-DXd during short-term (21 days) pre-operative treatment in patients with primary operable HER2-negative early breast cancer., Patients and Methods: Patients with previously untreated hormone receptor-positive/HER2-negative tumors were allocated to one of four cohorts according to baseline ERBB3 messenger RNA expression. All patients received one dose of HER3-DXd 6.4 mg/kg. The primary objective was to evaluate change from baseline in CelTIL score., Results: Seventy-seven patients were evaluated for efficacy. A significant change in CelTIL score was observed, with a median increase from baseline of 3.5 (interquartile range, -3.8 to 12.7; P = 0.003). Among patients assessable for clinical response (n = 62), an overall response rate of 45% was observed (tumor measurement by caliper), with a trend toward an increase in CelTIL score among responders compared with non-responders (mean difference, +11.9 versus +1.9). Change in CelTIL score was independent of baseline ERBB3 messenger RNA and HER3 protein levels. Genomic changes occurred, including switching toward a less proliferative tumor phenotype based on PAM50 subtypes, suppression of cell proliferation genes, and induction of genes associated with immunity. Treatment-emergent adverse events were observed in 96% of patients (14% grade ≥3); most common were nausea, fatigue, alopecia, diarrhea, vomiting, abdominal pain, and neutrophil count decrease., Conclusions: A single dose of HER3-DXd was associated with clinical response, increased immune infiltration, suppression of proliferation in hormone receptor-positive/HER2-negative early breast cancer, and a tolerable safety profile consistent with previously reported results. These findings support further study of HER3-DXd in early breast cancer., (Copyright © 2023 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Importance: Biomarkers to guide the use of pertuzumab in the treatment of early-stage ERBB2 (formerly HER2)-positive breast cancer beyond simple ERBB2 status are needed., Objective: To determine if use of the HER2DX genomic assay (Reveal Genomics) in pretreatment baseline tissue samples of patients with ERBB2-positive breast cancer is associated with response to neoadjuvant trastuzumab-based chemotherapy with or without pertuzumab., Design, Setting, and Participants: This is a retrospective diagnostic/prognostic analysis of a multicenter academic observational study in Spain performed during 2018 to 2022 (GOM-HGUGM-2018-05). In addition, a combined analysis with 2 previously reported trials of neoadjuvant cohorts with results from the assay (DAPHNe and I-SPY2) was performed. All patients had stage I to III ERBB2-positive breast cancer, signed informed consent, and had available formalin-fixed paraffin-embedded tumor specimens obtained prior to starting therapy., Exposures: Patients received intravenous trastuzumab, 8 mg/kg, loading dose, followed by 6 mg/kg every 3 weeks in combination with intravenous docetaxel, 75 mg/m2, every 3 weeks and intravenous carboplatin area under the curve of 6 every 3 weeks for 6 cycles, or this regimen plus intravenous pertuzumab, 840 mg, loading dose, followed by an intravenous 420-mg dose every 3 weeks for 6 cycles., Main Outcome and Measures: Association of baseline assay-reported pathologic complete response (pCR) score with pCR in the breast and axilla, as well as association of baseline assay-reported pCR score with response to pertuzumab., Results: The assay was evaluated in 155 patients with ERBB2-positive breast cancer (mean [range] age, 50.3 [26-78] years). Clinical T1 to T2 and node-positive disease was present in 113 (72.9%) and 99 (63.9%) patients, respectively, and 105 (67.7%) tumors were hormone receptor positive. The overall pCR rate was 57.4% (95% CI, 49.2%-65.2%). The proportion of patients in the assay-reported pCR-low, pCR-medium, and pCR-high groups was 53 (34.2%), 54 (34.8%), and 48 (31.0%), respectively. In the multivariable analysis, the assay-reported pCR score (as a continuous variable from 0-100) showed a statistically significant association with pCR (odds ratio [OR] per 10-unit increase, 1.43; 95% CI, 1.22-1.70; P < .001). The pCR rates in the assay-reported pCR-high and pCR-low groups were 75.0% and 28.3%, respectively (OR, 7.85; 95% CI, 2.67-24.91; P < .001). In the combined analysis (n = 282), an increase in pCR rate due to pertuzumab was found in the assay-reported pCR-high tumors (OR, 5.36; 95% CI, 1.89-15.20; P < .001) but not in the assay-reported pCR-low tumors (OR, 0.86; 95% CI, 0.30-2.46; P = .77). A statistically significant interaction between the assay-reported pCR score and the effect of pertuzumab in pCR was observed., Conclusions and Relevance: This diagnostic/prognostic study demonstrated that the genomic assay predicted pCR following neoadjuvant trastuzumab-based chemotherapy with or without pertuzumab. This assay could guide therapeutic decisions regarding the use of neoadjuvant pertuzumab.

Background: In hormone receptor-positive (HoR+) breast cancer (BC), gene expression analysis identifies luminal A (LumA), luminal B (LumB), human epidermal growth factor receptor 2 (HER2)-enriched (HER2-E), basal-like (BL) intrinsic subtypes and a normal-like group. This classification has an established prognostic value in early-stage HoR+ BC. Here, we carried out a trial-level meta-analysis to determine the prognostic ability of subtypes in metastatic BC (MBC)., Materials and Methods: We systematically reviewed all the available prospective phase II/III trials in HoR+ MBC where subtype was assessed. The primary endpoint was progression-free survival (PFS)/time to progression (TTP) of the LumA subtype compared to non-LumA. Secondary endpoints were PFS/TTP of each individual subtype, according to treatment, menopausal and HER2 status and overall survival (OS). The random-effect model was applied, and heterogeneity assessed through Cochran's Q and I 2 . Threshold for significance was set at P < 0.05. The study was registered in PROSPERO (ID: CRD42021255769)., Results: Seven studies were included (2536 patients). Non-LumA represented 55.2% and was associated with worse PFS/TTP than LumA [hazard ratio (HR) 1.77, P < 0.001, I 2  = 61%], independently of clinical HER2 status [P subgroup difference (P sub ) = 0.16], systemic treatment (P sub  = 0.96) and menopausal status (P sub  = 0.12). Non-LumA tumors also showed worse OS (HR 2.00, P < 0.001, I 2  = 65%), with significantly different outcomes for LumB (PFS/TTP HR 1.46; OS HR 1.41), HER2-E (PFS/TTP HR 2.39; OS HR 2.08) and BL (PFS/TTP HR 2.67; OS HR 3.26), separately (PFS/TTP P sub  = 0.01; OS P sub  = 0.005). Sensitivity analyses supported the main result. No publication bias was observed., Conclusions: In HoR+ MBC, non-LumA disease is associated with poorer PFS/TTP and OS than LumA, independently of HER2, treatment and menopausal status. Future trials in HoR+ MBC should consider this clinically relevant biological classification., (Copyright © 2023 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Importance: Patients with early-stage ERBB2 (formerly HER2)-positive breast cancer (ERBB2+ BC) who experience a pathologic complete response (pCR) after receiving neoadjuvant therapy have favorable survival outcomes. Predicting the likelihood of pCR may help optimize neoadjuvant therapy., Objective: To test the ability of the HER2DX assay to predict the likelihood of pCR in patients with early-stage ERBB2+ BC who are receiving deescalated neoadjuvant therapy., Design, Setting, and Participants: In this diagnostic/prognostic study, the HER2DX assay was administered on pretreatment tumor biopsy samples from patients enrolled in the single-arm, multicenter, prospective phase 2 DAPHNe clinical trial who had newly diagnosed stage II to III ERBB2+ BC that was treated with neoadjuvant paclitaxel weekly for 12 weeks plus trastuzumab and pertuzumab every 3 weeks for 4 cycles., Interventions and Exposures: The HER2DX assay is a classifier derived from gene expression and limited clinical features that provides 2 independent scores to predict prognosis and likelihood of pCR in patients with early-stage ERBB2+ BC. The assay was administered on baseline tumor samples from 80 of 97 patients (82.5%) in the DAPHNe trial., Main Outcomes and Measures: The primary aim was to test the ability of the HER2DX pCR likelihood score (as a continuous variable from 0-100) to predict pCR (ypT0/isN0)., Results: Of 80 participants, 79 (98.8%) were women and there were 4 African American (5.0%), 6 Asian (7.5%), 4 Hispanic (5.0%), and 66 White individuals (82.5%); the mean (range) age was 50.3 (26.0-78.0) years. The HER2DX pCR score was significantly associated with pCR (odds ratio, 1.05; 95% CI, 1.03-1.08; P < .001). The pCR rates in the HER2DX high, medium, and low pCR score groups were 92.6%, 63.6%, and 29.0%, respectively (high vs low odds ratio, 30.6; P < .001). The HER2DX pCR score was significantly associated with pCR independently of hormone receptor status, ERBB2 immunohistochemistry score, HER2DX ERBB2 expression score, and prediction analysis of microarray 50 ERBB2-enriched subtype. The correlation between the HER2DX pCR score and prognostic risk score was weak (Pearson coefficient, -0.12). Performance of the risk score could not be assessed due to lack of recurrence events., Conclusions and Relevance: The results of this diagnostic/prognostic study suggest that the HER2DX pCR score assay could predict pCR following treatment with deescalated neoadjuvant paclitaxel with trastuzumab and pertuzumab in patients with early-stage ERBB2+ BC. The HER2DX pCR score might guide therapeutic decisions by identifying patients who are candidates for deescalated or escalated approaches.