Liu, Minetta C.

Introduction Histiocytic neoplasms are rare disorders of the monocyte-macrophage-dendritic cell lineage that pose significant management challenges to the clinicians. Over the last five years, there has been emerging data on the role of MAPK/ERK pathway (RAS-RAF-MEK-ERK) in their pathogenesis. In this study, we report the molecular alterations among patients with histiocytic neoplasms seen at our institution. Methods We pursued target-capture next generation sequencing (NGS) on tissue specimen of patients with various histiocytic neoplasms- Erdheim-Chester disease (ECD), Langerhans cell histiocytosis (LCH), Rosai-Dorfman disease (RDD), histiocytic sarcoma (HS), and Langerhans cell sarcoma (LCS). We used three NGS platforms, Tempus® xO (1,714 genes), Tempus® xT (596 genes), or Foundation One® CDX (324 genes). All platforms used formalin-fixed paraffin-embedded tumor tissue specimens for detection of substitutions, insertion and deletion alterations, and copy number alterations, and gene rearrangements. The Tempus assays also utilized matched blood or saliva samples for germline DNA sequencing in addition to whole-transcriptome RNA sequencing. The tumor cellularity threshold for pursuing RNA sequencing was 30% so the test was not pursued specimens for lower cellularity. Results We included 31 cases with histiocytic neoplasms in this study. The disease distribution was as follows: ECD (n=6), LCH (n=7), RDD (n=12), ECD-RDD overlap (n=3), HS (n=2), and LCS (n=1). Among the 13 patients with ECD and LCH, BRAF-V600E mutations were found in 8 (62%) cases. We found several novel DNA alterations in this study in all disease groups: ECD (BRAFV471F), LCH (MAP2K1 in-frame deletion), and RDD (frameshift mutations of DNMT3A, SMARCA4, KMT2A, and CDC73 truncation). In one LCH patient without any DNA alterations, we found MAPK1 and MAP2K1 overexpression on RNA sequencing. We also had two cases with HS, both of which were found to have KRAS exon 3 mutations. One of these cases had concomitant JAK2V617F+ myelofibrosis and the tissue specimen for HS demonstrated TET2 and SRSF2 mutations as well. Our cohort included one patient with multifocal lymph node LCS that demonstrated NRAS and TP53 mutations, who did not respond to MEK-inhibitor therapy. Alterations in epigenetic regulators (TET2, DMNT3A, KMT2A, ASXL1) were found in 5 (16%), potentially suggesting clonal hematopoiesis in a subgroup of histiocytosis. Conclusions Histiocytic neoplasms demonstrate diverse somatic oncogenic alterations. In our study, we noted several novel alterations beyond the MAPK-ERK pathway in RDD that may point toward other potentially pathogenic mechanisms for these rare disorders. In cases where the DNA sequencing does not reveal pathogenic alterations, transcriptome analysis may aid in the diagnosis and management. OffLabel Disclosure: Trametinib use for Langerhans cell sarcoma

44 Background: A noninvasive cfDNA blood test detecting multiple cancers at earlier stages could decrease cancer mortality. In earlier discovery work, whole-genome bisulfite sequencing outperformed whole-genome and targeted sequencing approaches for multi-cancer detection across stages at high specificity. Here, multi-cancer detection and TOO localization using bisulfite sequencing of plasma cfDNA to identify methylomic signatures was evaluated in preparation for clinical validation, utility, and implementation studies. Methods: 2301 analyzable participants (1422 cancer [ > 20 tumor types, all stages], 879 non-cancer) were included in this prespecified substudy from the Circulating Cell-free Genome Atlas (CCGA) (NCT02889978) study - a prospective, multi-center, observational, case-control study with longitudinal follow-up. Plasma cfDNA was subjected to a targeted methylation sequencing assay using high-efficiency methylation chemistry to enrich for methylation targets, and a machine learning classifier determined cancer status and tissue of origin (TOO). Observed methylation fragments characteristic of cancer and TOO were combined across targeted regions and assigned a relative probability of cancer and of a specific TOO. Results: Performance is reported at 99% specificity (ie, a combined false positive rate across all cancer types of 1%), a level required for population-level screening. Across cancer types, sensitivity ranged from 59 to 86%. Combined cancer detection (sensitivity [95% CI]) was 34% (27-43%) in stage I (n = 151), 77% (70-83%) in stage II (n = 171), 84% (79-89%) in stage III (n = 204), and 92% (88-95%) in stage IV (n = 281). TOO was provided for 94% of all cancers detected; of these, TOO was correct in > 90% of cases. Conclusions: Detection of multiple deadly cancers across stages using methylation signatures in plasma cfDNA was achieved with a single, fixed, low false positive rate, and simultaneously provided accurate TOO localization. This targeted methylation assay is undergoing validation in preparation for prospective clinical investigation as a cancer detection diagnostic. Clinical trial information: NCT02889978.

Abstract Introduction: Given the established role of PD-1 in mediating immune suppression in chronic lymphocytic leukemia (CLL), we tested and reported the efficacy of PD-1 blocking antibody pembrolizumab in relapsed and transformed CLL patients. A selective response of pembrolizumab (~40%) in patients with RT, particularly after prior ibrutinib, was observed (Ding, Blood, 129:3419). Correlative analysis showed PD-1 expression in tumor B-cells of patients with RT and aggressive CLL after progression on ibrutinib. PD-1, an inhibitory receptor expressed on CLL T-cells, inhibits the immune synapse and cytotoxic T cell functions via the interactions with its ligands. However, the expression pattern and role of PD-1 in tumor B-cells is not well defined. In this study we investigated the functional implication of the PD-1 signaling axis in B-cell pathobiology in CLL and RT patients. Methods: 26 CLL-involved lymph node (LN) and 20 RT-involved LN were tested for PD-1 expression by immunohistochemistry (mouse clone NAT105, Abcam). For in vitro study, we checked PD-1 expression in 11 lymphoma cell lines and 1 CLL cell line by both flow cytometry and Western blot (WB) analysis. Effect of PD-1 knockdown using CRISPR/Cas9 system (Addgene) and over-expression of PD-1 using pLEX-lentiviral (Thermoscientific) or pRetro-retroviral (Clonetech) system were evaluated on pro-survival and apoptotic signaling pathways by Western blot analysis. Gene expression signatures in CLL and RT patients were also evaluated by Illumina-based RNA sequencing using FFPE-nodal tissue obtained by clinical biopsy (Tempus Labs; Chicago, IL). Results: The expression of PD-1 was significantly increased in RT-LN compared to CLL-LN. (mean ± SEM in RT vs. CLL, 30.6 ± 4.7 vs. 11.5 ± 2.8, p < 0.001). PD-1 expression was highest in patients with RT where the immediate prior CLL therapy was ibrutinib (Figure 1A). Among all cell lines tested for PD-1 expression, the expression of PD-1 by WB and flow was highest in Mino (mantle cell lymphoma line), followed by moderate expression in Jvm2 (B-PLL line) and Mec1 (CLL line), and very low-level expression in both Jeko-1 (B-NHL line) and lymphoma line 'Karpas299'. CRISPR/Cas9 mediated depletion of PD-1 in Mino cells inhibited constitutively active Akt, p70S6K and mTOR pathway, accompanied by significant downregulation of the anti-apoptotic proteins, Bcl-2, Mcl-1 and XIAP, but P-ERK1/2 was not affected. Constitutive lentiviral (pLEX-PD-1)-mediated overexpression of PD-1 in Jeko-1 and doxycycline regulated inducible retroviral (pRetro-PD-1) mediated overexpression of PD-1 in Karpas299 activated Akt, mTOR and p70S6K pathway. Overexpression of PD-1 in Jeko-1 significantly increased Bcl-2 and Mcl-1 and in Karpas299 increased Bcl-2, Mcl-1 and XIAP expression (Figure 1B). A parallel genetic analysis using RNA sequencing was performed on 5 nodal tissues involved by either RT or progressive CLL after these patients developed clinical progression after prior ibrutinib therapy. In all 5 patients, overexpression of PD-1 was associated with increased expression of Bcl-2 and mTOR regardless of the genetic mutations detected (including TP53, ATM, BTK, NOTCH1, XPO1, SF3B1, TET2 etc). However, in 2 patients who received prior chemoimmunotherapy, similar overexpression of gene signature was not observed by RNA sequencing analysis, alternative pathways including Met or NFkB overexpression was detected. Given these clinical and laboratory findings, we have treated 2 RT patients with a combination of BTK and Bcl-2 inhibitors (ibrutinib and venetoclax, respectively) whose CLL transformed after prior ibrutinib. Significant reduction of tumor burden was observed in both cases with one complete response and one mixed response. Conclusion: An increased expression of PD-1, Akt/mTOR and Bcl-2 gene signature was first observed in RT patients after prior ibrutinib therapy. PD-1 overexpression in the tumor B-cells of RT and progressive CLL patients likely regulate AKT/mtOR to upregulate Bcl-2. Targeting both BTK and Bcl-2 pathways in addition to PD-1 blockade appear to be a promising strategy to treat these aggressive diseases. Disclosures Parikh: Gilead: Honoraria; Janssen: Research Funding; Abbvie: Honoraria, Research Funding; Pharmacyclics: Honoraria, Research Funding; MorphoSys: Research Funding; AstraZeneca: Honoraria, Research Funding. Kenderian:Novartis: Patents & Royalties; Tolero Pharmaceuticals: Research Funding; Humanigen: Research Funding. Ansell:Takeda: Research Funding; Trillium: Research Funding; Affimed: Research Funding; Celldex: Research Funding; Merck & Co: Research Funding; Regeneron: Research Funding; LAM Therapeutics: Research Funding; Bristol-Myers Squibb: Research Funding; Seattle Genetics: Research Funding; Pfizer: Research Funding. Kay:Infinity Pharm: Membership on an entity's Board of Directors or advisory committees; Celgene: Membership on an entity's Board of Directors or advisory committees; Acerta: Research Funding; Agios Pharm: Membership on an entity's Board of Directors or advisory committees; Cytomx Therapeutics: Membership on an entity's Board of Directors or advisory committees; Tolero Pharmaceuticals: Membership on an entity's Board of Directors or advisory committees, Research Funding; Gilead: Membership on an entity's Board of Directors or advisory committees; Morpho-sys: Membership on an entity's Board of Directors or advisory committees; Pharmacyclics: Membership on an entity's Board of Directors or advisory committees, Research Funding; Janssen: Membership on an entity's Board of Directors or advisory committees. Ding:Merck: Research Funding.

Abstract Introduction Histiocytic neoplasms are rare hematological malignancies that have protean clinical manifestations and can pose significant management challenges. Recently, vemurafenib was approved by the US-FDA for treatment of BRAF-V600-mutant Erdheim-Chester disease (ECD). However, there is a lack of FDA-approved therapies for other histiocytic neoplasms such as Langerhans cell histiocytosis (LCH) and Rosai-Dorfman disease (RDD). Over the last 5 years, immune checkpoint inhibitors such as programmed death-1 (PD-1) and programmed death-ligand 1 (PD-L1) inhibitors have shown significant improvement in outcomes among patients with several hematological and solid organ malignancies. In order to identify appropriate treatment candidates for these therapies, predictive biomarkers have been developed in various cancers. Evidence from solid tumors has suggested a favorable response to checkpoint inhibitor therapy with higher tumor mutational burden (TMB), defined as the number of mutations within a tumor genome. Next generation sequencing (NGS) of various tumors has shown an association between TMB of < 5 mutations/megabase (mut/Mb) to be associated with an absence of benefit from checkpoint inhibitors. In addition, high levels of PD-1/PD-L1 expression and microsatellite instability (MSI) are also correlated with response to therapy. The latter may be a result of somatic or germline alterations in DNA mismatch repair genes. In this study, we report the results for these biomarkers using NGS in patients with histiocytic neoplasms. Methods We utilized TempusTM NGS platform to analyze the tissue specimen of patients with histiocytic neoplasm. The Tempus xO Assay (Tempus Labs; Chicago, IL) combines a 1,711 gene targeted somatic and germline DNA sequencing panel with RNA-sequencing to detect both germline and somatic single nucleotide polymorphisms, indels, copy number variants, and gene rearrangements causing chimeric mRNA transcript expression in a wide array of solid tumor types. The assay utilizes formalin-fixed paraffin-embedded tumor samples and matched blood samples. TMB was calculated and reported as somatic mutations in tumor tissue per million base-pairs or mut/Mb. RNA sequencing was utilized to assess for PD-L1 and PD-1 gene expressions as compared to matched tumor and normal reference sets. Both PD-L1 and PD-1 gene expressions were reported as percentiles. DNA mismatch repair status was predicted by analysis of alterations in five common mismatch repair genes in somatic and germline DNA (MSH2, MSH6, MLH1, PMS2, and EPCAM). If there were no alterations identified in these genes, the mismatch repair status was predicted as microsatellite stable (MSS). Results A total of 13 patients with histiocytic neoplasms were included in the study. The distribution of individual histiocytic neoplasms was as follows: RDD (n=9), ECD (n=3), and LCH (n=1). The median TMB for RDD and ECD patients was 0.17 mut/Mb. For the one patient with LCH, the TMB level was 0.51 mut/Mb. Individual TMB levels are shown in figure 1. PD-L1 and PD-1 expression levels are depicted in table 1. Compared to normal reference sets, the PD-L1 expression was elevated in one patient each with RDD and ECD, and PD-1 expression was elevated in two patients each with RDD and ECD. For both ECD patients with higher PD-1 expression, NGS also showed presence of BRAF-V600E in the tumor tissue. The LCH patient had a low level of PD-L1 and PD-1 expression. For patients where evaluation of DNA mismatch repair was feasible on the tissue specimen (n=4), none showed related somatic or germline alterations. Conclusions In our series, the histiocytic neoplasms RDD, ECD, and LCH demonstrated low levels of TMB. None of the RDD and ECD patients were found to have alterations in DNA mismatch repair genes. Other markers of immunotherapy such as PD-L1/PD-1 expression appeared to be higher in ECD patients with BRAF-V600E, but in only a small subset of RDD patients. The low TMB seen in our study suggests that these histiocytic neoplasms may be less likely to respond to immune checkpoint inhibitors such as anti-PD-1 and anti-PD-L1 agents as compared to the tumors with high TMB. Disclosures No relevant conflicts of interest to declare.

Background: Anaplastic thyroid cancer (ATC) has poor prognosis with median overall survival (OS) of ∼6 months. We previously reported high PD-1/PDL-1 staining in ATC, raising the possibility of the productive application of the immunotherapeutic pembrolizumab. However, having found pembrolizumab to anecdotally have limited single-agent activity in ATC, we sought to alternatively define whether pembrolizumab might synergistically combine with chemoradiotherapy as initial ATC therapy. Methods: An investigator-initiated therapeutic phase 2 trial of pembrolizumab, 200 mg intravenously (IV) every 3 weeks, combined with chemoradiotherapy (docetaxel/doxorubicin, 20 mg/m 2 each IV weekly plus volumetric modulated arc therapy) was initiated as frontline therapy (with or without surgery) in ATC to assess efficacy and toxicities. Six-month OS was selected as the primary endpoint using a Simon's optimal design with interim analysis (targeting accrual of 25 patients; Cohort A: prior resection, Cohort B: no resection). Based on a prior patient cohort-treated similarly, but without pembrolizumab, the design was such that, if 6-month true survival is 75%, the probability of declaring the approach worthy of further pursuit would be 91%. Results: Three patients were enrolled, two with rapidly enlarging unresectable neck masses. Early tumor responses were favorable in all three, and all three satisfactorily completed: intended radiotherapy, preceding and radiotherapy-concurrent pembrolizumab, and concurrent chemoradiotherapy. However, all three patients died <6 months following therapy initiation—one from pulmonary metastases and two from otherwise unexpected fatal pulmonary complications occurring subsequent to chemoradiotherapy completion—prompting study closure. Conclusions: Although initially tolerated and effective in terms of locoregional disease control, disappointing survival outcomes compared with historical controls raise uncertainty that the piloted approach merits further pursuit in ATC.

Abstract Background: Pembrolizumab (Pembro), an anti-PD-1 immune checkpoint inhibitor, has been approved for the treatment of a variety of cancers. Pembro was recently evaluated in HER2- breast cancer patients in the neoadjuvant I-SPY 2 TRIAL and graduated in the triple negative (TN), HR+HER2-, and HER2- signatures. TN breast cancers tend to have high numbers of immune infiltrates, including T cells and tumor associated macrophages (TAMs). We utilized multiplex multispectral imaging to evaluate the presence and spatial proximity of various immune cell populations as specific predictors of response to Pembro. Methods: Pre-treatment biopsies were available from 54 patients. FFPE sections were immunostained using Opal fluorescence reagent kits on a fully automated Ventana Discovery platform and imaged with a Vectra automated imaging system. Image analyses were performed with a variety of software packages. Immune biomarkers included CD3, CD8, FoxP3, CD68, PD-1, PD-L1, and cytokeratins. Cell densities were determined per area of tissue. Spatial analyses were performed to quantitate proximity of cell types. We used logistic regression with the likelihood ratio test to evaluate associations between pathologic complete response (pCR) with immune cell counts, ratios, and spatial proximity measurements. This analysis was also performed adjusting for HR status, and within receptor subsets, sample size permitting. Our statistics are descriptive rather than inferential and do not adjust for multiple hypothesis testing or multiplicities of other biomarkers outside this study. Results: Densities of CD3+ T cells ranged from 193-5155 cells/mm2, with CD8+ cytotoxic T cells (Tc) ranging from 103 to 3774 cells/mm2 and FoxP3+ Tregs from 0-416 cells/mm2. Densities of PD-1+ T cells and PD-1+ Tc cells ranged from 0-1341 cells/mm2 and 0-698 cells/mm2, respectively. Macrophage cell densities ranged from 84-2644 cells/mm2. PD-L1+ tumor cell densities ranged from 0-2981 cells/mm2. High PD-L1 expression (&gt;=10% of tumor cells positive for PD-L1) was observed in 12% of the cases. Overall T cell density was positively associated with pathological complete response (pCR), as were Treg cell density and PD-1+ T cell density. The ratio of macrophages to Tc cells was negatively associated with response. Finally, the spatial distribution of CD3+ T cells in proximity to cancer cells correlated positively with pCR. Conclusion: Cell density, immune cell ratios and spatial resolution provide insight to response and resistance. We confirmed the observation that T cell density predicts pCR, but more specifically that Treg and PD1+ T cell densities increase the chance of pCR. An increase in macrophages relative to cytotoxic T cells predicts resistance. The closer the T cells are to the tumor, the better the response. These results demonstrate the utility of characterizing the immune microenvironment, in the context of therapy with immuno-oncology agents, not only to predict response, but to gain insight and learn now to potentiate agents and combinations. Citation Format: Michael J. Campbell, Christina Yau, Jennifer Bolen, Scott Vandenberg, Clifford Hoyt, Lamorna Brown-Swigart, Gillian Hirst, Rita Nanda, Minetta Liu, Smita Asare, Laura van'tVeer, Douglas Yee, Angie DeMichele, Don Berry, Laura Esserman, I SPY 2 TRIAL Investigators. Analysis of immune cell infiltrates as predictors of response to the checkpoint inhibitor pembrolizumab in the neoadjuvant I-SPY 2 TRIAL [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr CT003.

Abstract Background: Triple negative breast cancer (TNBC), generally considered aggressive with few options beyond standard chemotherapy, is currently experiencing a period of optimism, with multiple trials showing increased efficacy of platinum containing regimens (with and without PARP-inhibition), and more recently, immunotherapy. In I-SPY 2, veliparib/carboplatin (VC) and the immune checkpoint inhibitor pembrolizumab (Pembro) graduated in the TN subtype. As part of the I-SPY 2 biomarker program, we previously identified the signature PARPi7, a measure of DNA repair deficiency (DRD), as predicting response to VC. As well, immune signatures, including one representing dendritic cells, predicted response to Pembro. Here we investigate the overlap between these biomarkers to identify patient subgroups more likely to respond to immunotherapy or platinum-based therapy and estimate response rates to VC and Pembro in patients positive for one, both, or neither biomarker. Methods: 153 TNBC patients (Control: 85; VC: 39; Pembro: 29) were considered in this analysis. Continuous DNA repair deficiency and immune signatures assayed at the pre-treatment time point were evaluated as published (Wolf, 2017; Danaher, 2017). To identify optimal dichotomizing thresholds, 2-fold cross-validation was repeated 500 times. Bayesian logistic regression was used to estimate pCR rates by arm for patient subsets defined by biomarker combinations. Our study is exploratory with no claims for generalizability; and this analysis does not adjust for multiplicities. Results: Using the optimal dichotomizing cutpoints, 54% of TNBC patients are classified as DRD+, and 67% Immune+. As expected, DRD+ patients have a high estimated pCR rate to VC (75%); and Immune+ patients have a high estimated pCR rate to Pembro (82%). Combining these biomarkers, 40% of TNBC are positive for both biomarkers, 40% for only one biomarker (26% Immune+/DRD-, 14% Immune-/DRD+), and 20% for neither. The Immune+/DRD+ TN subset predicted sensitive to both VC and Pembro has a higher estimated pCR rate to both VC and Pembro relative to control (estimated pCR rates: VC: 75%, Pembro: 73% vs. control: 18%). In contrast, the Immune+/DRD- group, predicted to be sensitive to Pembro, has the highest pCR rate to Pembro (VC: 48%, Pembro: 83% vs. control 21%), whereas the Immune-/DRD+ group is most responsive to VC (VC: 63%, Pembro: 41% vs. control 29%). For the 20% of Immune-/DRD- TNBC patients, their estimated response rates are lower than the biomarker-positive patients (VC: 27%, Pembro: 43% vs. control: 19%). Conclusion: Molecular phenotypes capturing immune activation and DNA repair deficiency may predict response of TNBC and help prioritize selection of platinum or immunotherapy containing regimens. Citation Format: Denise M. Wolf, Christina Yau, Julia Wulfkuhle, Emanuel Petricoin, Lamorna Brown-Swigart, Gillian Hirst, Smita Asare, I-SPY 2 Consortium, Douglas Yee, Angela DeMichele, Hope Rugo, Olufunmilayo Olopade, Rita Nanda, Minetta Liu, Laura Esserman, Laura van t' Veer. Integration of DNA repair deficiency and immune biomarkers to predict which early-stage triple-negative breast cancer patients are likely to respond to platinum-containing regimens vs. immunotherapy: The neoadjuvant I-SPY 2 trial [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 2679.

AbstractThere is a clear and unmet need for biomarkers in hepatocellular carcinoma (HCC). Circulating cell free deoxyribonucleic acid (cfDNA) is a fragmented DNA subtype, found in the blood circulation. Circulating tumor DNA (ctDNA) is the fraction of total cfDNA, which originates from the primary tumor or metastases in patients with cancer. Earlier studies reported that quantitative measurement cfDNA has diagnostic and prognostic role for HCC. More recently, improvement in next-generation sequencing technology and better understanding of genetic or epigenetic alteration of HCC have allowed comprehensive analysis of mutational and methylation landscape of ctDNA. Hotspot mutation panels and methylation panels have both shown promising performance. None of these tests have yet been validated in longitudinal cohorts for preclinical detection of HCC. In this article, the authors discuss the currently available ctDNA detection technologies, their diagnostic and prognostic performance in HCC, and future research directions.

PURPOSE This focused update addresses the use of Onco type DX in guiding decisions on the use of adjuvant systemic therapy. METHODS ASCO uses a signals approach to facilitate guideline updating. For this focused update, the publication of the Trial Assigning Individualized Options for Treatment (TAILORx) evaluating noninferiority of endocrine therapy alone versus chemoendocrine therapy for invasive disease–free survival in women with Onco type DX scores provided a signal. An expert panel reviewed the results of TAILORx along with other published literature on the Onco type DX assay to assess for evidence of clinical utility. UPDATED RECOMMENDATIONS For patients with hormone receptor–positive, axillary node–negative breast cancer whose tumors have Onco type DX recurrence scores of less than 26, there is little to no benefit from chemotherapy, especially for patients older than age 50 years. Clinicians may recommend endocrine therapy alone for women older than age 50 years. For patients 50 years of age or younger with recurrence scores of 16 to 25, clinicians may offer chemoendocrine therapy. Patients with recurrence scores greater than 30 should be considered candidates for chemoendocrine therapy. Based on informal consensus, the panel recommends that oncologists may offer chemoendocrine therapy to these patients with recurrence scores of 26 to 30. Additional information can be found at www.asco.org/breast-cancer-guidelines .