Wolf, Denise M.

PURPOSE Neoadjuvant immunotherapy (IO) has become the standard of care for early-stage triple-negative breast cancer (TNBC), but not yet for other subtypes. We previously developed a clinical-grade mRNA-based immune classifier (ImPrint) predicting response to IO that is now being used in I-SPY2.2 as part of the response predictive subtypes. We report the performance of ImPrint in hormone receptor–positive and human epidermal growth factor receptor 2–negative (HR+HER2–) patients from five IO arms. METHODS A total of 204 HR+HER2– (MammaPrint high-risk) patients from five IO arms (anti–PD-1, anti–PD-L1/poly [ADP-ribose] polymerase inhibitor combination, anti–PD-1/toll-like receptor 9 dual-IO combination, and anti–PD-1 ± lymphocyte activation gene 3 dual-IO combination) and 191 patients from the chemotherapy-only control arm were included in this analysis. Patients were classified as ImPrint+ (likely sensitive) versus ImPrint– (likely resistant), using pretreatment mRNA. Performance of ImPrint for predicting pathologic complete response (pCR) to IO-containing arms was characterized and compared with tumor grade (III), MammaPrint (ultra) High2 risk (MP2), and estrogen receptor (ER)-low (ER ≤ 10%). RESULTS Overall, the pCR rate across the five IO arms was 33%. 26% of HR+HER2– patients were ImPrint+, and pCR rates with IO were 75% in ImPrint+ versus 17% in ImPrint–, with the highest pCR rate >90% in a dual-IO arm. In the control arm, pCR rates were 33% in ImPrint+ and 8% in ImPrint–. Tumor grade (III), MP2, and ER-low showed pCR rates in IO of 45%, 56%, and 63%, respectively, with lower pCR odds ratios (OR < 7.5) compared with ImPrint (OR = 14.5). CONCLUSION Using an accurate selection strategy, HR+HER2– patients could achieve pCR rates similar to what is seen with best neoadjuvant therapies in TNBC and HER2+ (ie, pCR rate >65%-70%). ImPrint, an Food and Drug Administration IDE-enabled assay, may represent a way to identify HR+HER2– patients for IO that best balances likely benefit versus risk of serious immune-related adverse events.

610 Background: The adverse event (AE) landscape in oncology is changing due to the introduction of immunotherapy and antibody drug conjugates. These AEs come with both short and long-term symptoms that significantly impact patient quality of life. Monitoring for early onset of symptoms could optimize therapy for a particular patient, maximizing potential efficacy while mitigating toxicity. It is also possible that some toxicities are directly associated with drug sensitivity. We sought to identify symptoms associated with pathologic complete response (pCR) using patient-reported outcomes (PROs) in early-stage high-risk breast cancer patients. Methods: Our study population included 288 stage II/III high-risk breast cancer patients enrolled on the I-SPY2 trial from 2021-2024, who received novel neoadjuvant therapies ± standard paclitaxel. pCR was assigned if tumor was absent in breast and nodes at surgery following neoadjuvant treatment. Patients (n = 288, pCR rate = 29%, 89% administered immunotherapy) were sent electronic PROs. 33 patient-reported AEs were measured using NCI's Patient Reported Outcomes - Common Terminology Criteria for Adverse Events (PRO-CTCAE). Each symptom was evaluated using severity, frequency, and interference on a Likert Scale. Presence of early PRO symptoms (cycles 1-3 of treatment) were binarized (at least one of moderate or greater), and odds ratios were computed with pCR as outcome. To assess whether higher grade AEs were enriched in patients that achieved a pCR, we also performed the Wilcoxan rank sum test using maximum (worst) symptom severity. Results: Of 288 patients included in our analysis (median age = 48 years, range = 20-78), 203 (70.5%) were White, 17 (5.9%) were Asian, 33 (11.5%) were Black or African American, and 35 (12.2%) were Hispanic. PRO analysis revealed that patients that had moderate to severe muscle pain (27% vs 10% OR = 3.15, p < 0.05), joint pain (22% vs 8% OR = 3.23, p < 0.05), headache (27% vs 12.5% OR = 2.59, p < 0.05), or mouth/throat sores (16% vs 5% OR = 3.56, p < 0.05) within weeks 1-3 had higher odds of achieving a pCR. When we looked at maximum severity between weeks 1-3, patients that achieved a PCR had higher grade muscle pain (p = 0.04), heart palpitations (p = 0.035), and significantly lower grade numbness and tingling (p = 0.002). Beyond 6 weeks, associations were weaker or insignificant. Conclusions: Our study utilizes an analysis framework that was able to determine sentinel symptoms such as muscle and joint pain, mouth/throat sores and palpitations as early as weeks 1-3 associated with increased efficacy. This may suggest an early immune reaction in patients that eventually respond favorably to treatment. Our work can help provide earlier proactive monitoring to mitigate toxicities, treatment redirection if needed, and a potential symptom-based early understanding to personalize treatment efficacy. A similar analysis is underway to predict immune related AEs. Clinical trial information: NCT01042379 .

271 Background: Many studies have shown that fatigue is associated with poor outcomes such as quality of life and physical function in breast cancer survivors.Early treatment of symptoms that result in persistent fatigue may be a strategy to mitigate long-term fatigue.As a result, we studied symptoms and developed risk estimates for patients that are most likely to develop fatigue by pre-surgery and follow up among patients treated on the I-SPY 2 neoadjuvant trial. I-SPY 2 is a phase-II adaptive trial for high-risk early-stage breast cancer patients. Methods: In this retrospective cohort study of 306 women undergoing neoadjuvant chemotherapy +/- targeted therapy, patients complete electronic surveys of symptoms via 34 PRO-CTCAE items and QOL via 9 PROMIS instruments. Fatigue is collected as part of the PROMIS survey and scored using Health Measures. Study timepoints are screening (SC), weekly study treatment (ST), 12 weeks (12w), pre-surgery (PS) and at 1, 6, 12 and 24 months (FU1-FU4) following surgery. Correlation between symptoms and fatigue at equal timepoints was quantified using spearman correlations and BH-adjusted p-values. For symptom odds analysis, symptoms were binarized such that those that had an average grade of mild or above (severity), rarely or above (frequency) or a little bit or above (interference) between ST weeks 1-6 were considered present. Clinically significant (CS) increase in fatigue was defined as an increase >=0.5 standard deviation (SD) in mean from baseline. Logistic regression was used to determine odds ratios of factors associated with fatigue. Results: Baseline PROMIS fatigue mean score was 47.14 (SD 9.31) with a median of 47.45. A CS increase in fatigue from baseline was seen in 55% of patients at 12w, and 61% of patients pre-surgery. Of the patients that had a CS increase in fatigue at any point prior to surgery, 45.7% had persistent fatigue through FU1-4. Symptoms associated with Fatigue at 12w were shortness of breath, headache, and nausea (P< 0.03). At PS, fatigue was significantly correlated with GI symptoms, neuropathy, and shortness of breath (P< 0.03). Risk estimates for patients that filled out symptom surveys at early timepoints (weeks 1-6) and also had fatigue were evaluated. Patients with early GU (genitourinary) symptoms had higher risk of fatigue at IR (OR=9.33, p=0.02, 88.9% in cases vs 33.3% in controls). Patients with early GI symptoms at PS (OR=8.56, p=0.03, 90.1% vs 50%), or FU2-4 (OR= 5.88 and 7.36, p= 0.04 and 0.02, 80% vs 39.1% and 81.8% vs 36.3%) also had a higher risk of developing fatigue. Conclusions: A CS increase in fatigue during treatment and follow up is seen in the majority of patients and is associated with a large number of persistent symptoms.

11113 Background: Patient-reported outcomes (PROs) are valuable for understanding treatment tolerability and toxicity. I-SPY2 is a phase-II neoadjuvant clinical trial for breast cancer with electronic PROs and quality of life (QOL) assessments. We sought to understand how well the GP5 self-reported tolerability item reflected toxicity burden and QOL during treatment and follow-up in I-SPY 2 participants. Methods: Our study included 306 patients who completed the FACIT Item GP5 and answered PROMIS surveys at two or more timepoints. Analysis timepoints included screening (SC), inter-regimen (IR), pre-surgery (PS), 1-month post-surgery (FU1), and 6-months post-surgery (FU2). Symptoms were assessed weekly using PRO-CTCAE and CTCAEv4.0. QOL was evaluated using PROMIS. The GP5: “I am bothered by side effects of treatment,” was used to estimate tolerability, where worse tolerability is a higher score. Odds ratios, Spearman correlation, and regularized multi-variate regression were used to evaluate the associations between individual symptoms and adverse events with GP5 scores. Results: Demographic information was available for 243 of the 306 (70.2%) participants (median age=47,73.7% white). In the 306, tolerability scores worsened over study treatment, with a maximum mean score of 1.71 at Cycle 11. Tolerability improved after treatment cessation at FU1(1.09). Mean tolerability at week 6 was 1.53. The only clinical factor that was associated with early change in tolerability (week 6 to PS) was age >50 (mean score ranged from 1.55-1.70 vs 1.29-1.45 (age<50), p<0.05). PRO-CTCAE and GP5 correlative analysis suggested that higher grade abdominal pain, decreased appetite, dizziness and blurry vision significantly associated with higher GP5 scores at multiple timepoints (p<0.05). Symptoms not associated with tolerability were nail loss, acne, hair loss, and rash (p>0.05). Additionally, the total number of low-grade (mild to moderate) symptoms a patient experienced during study treatment was significantly correlated to poor tolerability (p<0.00001). Among patients with CTCAE data, 9% had adrenal insufficiency, and higher GP5 was found across all study timepoints in those with (compared to without) adrenal insufficiency (mean GP5 = [1.7-1.9] vs [1.3-1.5] respectively). Similarly, among patients with neuropathy (14%), higher GP5 scores were reported across all study timepoints (mean GP5 = [1.8-2.3] vs [1.3-1.5] respectively). Higher GP5 scores during treatment were correlated with higher fatigue scores at PS and FU1 (p<0.02 and p<0.001) and lower physical functioning at PS (p<0.01). Conclusions: TheGP5 measure is associated with self-reported symptoms, higher grade adverse events and quality of life scores. Furthermore, greater accumulation of lower grade symptoms can lead to poor tolerability. The single measure GP5 composite score reflects multiple components of treatment toxicity.

Abstract Background: Neoadjuvant immunotherapy (IO) has become standard of care for early-stage triple negative breast cancer (TNBC). I-SPY2 was the first randomized trial to examine the efficacy of IO therapy in high-risk HR+HER2- breast cancer, where most IO arms showed improved efficacy relative to control. We also previously showed immune-gene expression signatures associate with pathologic complete response (pCR) in HR+HER2- treated with IO; and developed a clinically applicable Immune classifier (ImPrint) predicting response to IO that is now being used in I-SPY2.2 as part of the Response Predictive Subtypes. Here we report the performance of ImPrint in HR+HER2- patients from 5 IO arms. Methods: 204 HR+HER2- (MammaPrint high risk) patients from 5 IO arms (anti-PD1, anti-PDL1/PARPi, anti-PD1/TLR9 dual-IO, and anti-PD1 +/- LAG3 dual-IO) and 191 patients from the control arm were included in this analysis. Patients were classified ImPrint+ (likely sensitive) vs. ImPrint- (likely resistant), by Agendia, Inc, using pre-treatment mRNA. Performance of ImPrint for predicting pCR to IO was characterized and compared to that of MammaPrint (ultra) High2 risk (MP2) and tumor grade (III). Based on the DRFS of pCR vs. non-pCR HR+HER2-/ImPrint+ patients (median follow up ~4.5yr), we predicted the DRFS in the IO vs. control arms using an exponential model as a function of the pCR rates. Results: Overall, the pCR rate over the 5 IO arms was 33%. 28% of HR+HER2- patients were ImPrint+; and pCR rates with IO were 76% in ImPrint+ vs. 16% in ImPrint-, with the highest pCR rate &gt;90% in a dual-IO arm. In the control arm, pCR rates were 33% in ImPrint+ and 8% in ImPrint-. Based on the pCR rates in the IO vs. control (76% vs. 33%), the predicted DRFS of HR+HER2-/ImPrint+ patients are 91% vs 80% at 5 years, respectively. This translates to a risk reduction of 52% for the IO arm relative to standard chemotherapy in HR+HER2-/ImPrint+ patients. We also compared MP2 and tumor grade (III) to ImPrint+ as pCR predictors, as these markers have been used or proposed as selection markers for neoadjuvant IO trials in HR+HER2-. The pCR rates for MP2, Grade III, at 56%, 45%, respectively, are much smaller than that for ImPrint+ at 75% pCR; thus ImPrint appears to be a more precise predictive biomarker for neoadjuvant IO therapy in the HR+HER2- breast cancers. Conclusion: These results suggest that a subset of high risk HR+HER2- breast cancers is highly sensitive to immunotherapy, and by using a specific and sensitive selection strategy patients could achieve pCR rates similar to what is seen with best neoadjuvant therapies in TNBC and HER2+ (i.e., pCR rate &gt; 65-70%). ImPrint, an FDA IDE-enabled assay currently being further evaluated in I-SPY2, may represent the way to identify patients for IO that best balances likely benefit vs risk of serious immune-related adverse events. Citation Format: Denise M. Wolf, Christina Yau, Lajos Pusztai, Rita Nanda, Jo Chien, Erica Stringer-Reasor, Rebecca Shatsky, Claudine Isaacs, MInetta Liu, Hyo Han, Hatem Soliman, Michael Campbell, Annuska Glas, Andrei Barcaru, Lorenza Mittempergher, Midas Kuilman, Lamorna Brown Swigart, Gillian Hirst, Hongmei Yu, Philip Beineke, Amrita Basu, I-SPY2 Investigators, Douglas Yee, W. Fraser Symmans, Angela DeMichele, Jane Perlmutter, Amy Delson, Laura Huppert, Hope Rugo, Amy Clark, Nola Hylton, Paula Pohlmann, Laura Esserman, Laura van 't Veer. Immune subtyping identifies a subset of HR+HER2- early-stage breast cancer patients with a very high likelihood of response to neoadjuvant immunotherapy (IO): Results from 5 IO arms of the I-SPY2 TRIAL [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1207.

Abstract Purpose: The neutralizing peptibody trebananib prevents angiopoietin-1 and angiopoietin-2 from binding with Tie2 receptors, inhibiting angiogenesis and proliferation. Trebananib was combined with paclitaxel±trastuzumab in the I-SPY2 breast cancer trial. Patients and Methods: I-SPY2, a phase II neoadjuvant trial, adaptively randomizes patients with high-risk, early-stage breast cancer to one of several experimental therapies or control based on receptor subtypes as defined by hormone receptor (HR) and HER2 status and MammaPrint risk (MP1, MP2). The primary endpoint is pathologic complete response (pCR). A therapy “graduates” if/when it achieves 85% Bayesian probability of success in a phase III trial within a given subtype. Patients received weekly paclitaxel (plus trastuzumab if HER2-positive) without (control) or with weekly intravenous trebananib, followed by doxorubicin/cyclophosphamide and surgery. Pathway-specific biomarkers were assessed for response prediction. Results: There were 134 participants randomized to trebananib and 133 to control. Although trebananib did not graduate in any signature [phase III probabilities: Hazard ratio (HR)-negative (78%), HR-negative/HER2-positive (74%), HR-negative/HER2-negative (77%), and MP2 (79%)], it demonstrated high probability of superior pCR rates over control (92%–99%) among these subtypes. Trebananib improved 3-year event-free survival (HR 0.67), with no significant increase in adverse events. Activation levels of the Tie2 receptor and downstream signaling partners predicted trebananib response in HER2-positive disease; high expression of a CD8 T-cell gene signature predicted response in HR-negative/HER2-negative disease. Conclusions: The angiopoietin (Ang)/Tie2 axis inhibitor trebananib combined with standard neoadjuvant therapy increased estimated pCR rates across HR-negative and MP2 subtypes, with probabilities of superiority &gt;90%. Further study of Ang/Tie2 receptor axis inhibitors in validated, biomarker-predicted sensitive subtypes is warranted.

611 Background: Patient reported outcomes (PROs) capture direct feedback from patients in clinical trials. The NIH Patient-Reported Outcomes Measurement Information System (PROMIS) is an open source and validated PRO platform. Data on PROMIS score trajectories over time for breast cancer patients undergoing neoadjuvant chemotherapy (NAC) are limited. Methods: We administered paper surveys to patients enrolled on the I-SPY2 trial, a multicenter, randomized phase 2 trial comparing novel agents to standard NAC in high-risk breast cancer. PRO domains included PROMIS Physical Function, Anxiety, Depression, Applied Cognition and Fatigue. Surveys were administered at baseline (post-consent), C1D1, inter-regimen (~12 wks into NAC), pre-surgery and at 1, 6, 12, and 24 months post-surgery. Responses were scored using the HealthMeasures Scoring Service to generate a standardized T score, with a score of 50 representing the general US population mean. Higher Anxiety, Depression and Fatigue scores represent worse symptoms; higher Physical Function and Applied Cognition scores represent better symptoms. We calculated average scores by time-point and compared changes from baseline to various grouped time points using a paired t-test. Results: From 1/2012 to 7/2020, 1631 patients enrolled in I-SPY 2 with 959 completing the baseline survey. 62% had at least two “on-treatment” surveys, and 52% had at least one follow-up time-point. Average baseline PROMIS scores for Physical Function, Anxiety, Depression, Applied Cognition and Fatigue were 46.7, 55.1, 47.2, 53.3 and 46.0 respectively. Between baseline and the latest on-treatment time point we observed improvement in Anxiety (-5.2) and worsening in Physical Function, Applied Cognition and Fatigue (-5.4, -6.9 and +8.2 respectively). Between baseline and the latest post-surgery time point, we saw improved Anxiety (-5.9) and worsened Physical Function, Applied Cognition and Fatigue (-5.9, -3.6 and +4.3). Depression changed minimally. Conclusions: Using PROMIS, patients on I-SPY2 experienced worse Physical Function, Applied Cognition and Fatigue but improved Anxiety during and after treatment compared to baseline. While changes in Applied Cognition and Fatigue recovered partially after treatment, Physical Function scores did not, suggesting potential for longer term impact on quality of life. Clinical trial information: NCT01042379 . [Table: see text]

Abstract Background: We compared the predictive and prognostic value of ctDNA dynamics in high-risk hormone receptor-positive/HER2-negative (HR+/HER2-) and triple negative breast cancer (TNBC) receiving neoadjuvant chemotherapy (NAC) enrolled in the I-SPY 2 trial (NCT01042379). To our knowledge, this is the largest ctDNA study in breast cancer in the neoadjuvant setting. Methods: Blood samples were collected at pre-treatment (T0), during treatment (T1 at 3 weeks, and T2 at 12 weeks) and after NAC (T3 at 24 weeks) from 106 HR+/HER2- and 97 TNBC patients. Plasma samples (n=734) were analyzed using a personalized and tumor-informed mPCR NGS-based ctDNA test (SignateraTM). Patients, all high risk for recurrence by MammaPrint, received paclitaxel-based treatment +/- experimental therapy followed by anthracycline. The median follow-up was 3.0 years (0.5 to 6.5). The predictive and prognostic value of ctDNA dynamics and status at different timepoints were examined. Our analysis is exploratory and does not adjust for other biomarkers. Results: Pretreatment ctDNA positivity (Fisher p&lt;0.0001) and levels (mean tumor molecules/mL, MTM/mL, t test p=0.0062) were significantly higher in TNBC (90.7%, 14.7 MTM/mL) than in high risk HR+/HER2- (66.0%, 5.5 MTM/mL). Early and late ctDNA clearance during treatment (3 and 12 weeks of NAC) was predictive of pathologic complete response (pCR) and residual cancer burden (RCB), class 0-III, in TNBC but not HR+/HER2- (Table). In both subtypes: (1) ctDNA was a significant negative prognostic factor for distant recurrence-free survival (DRFS) at all timepoints (p&lt;0.05) except at pretreatment; (2) all patients who achieved pCR were ctDNA-negative after NAC; (3) among non-responding patients, ctDNA-negativity after NAC was associated with improved DRFS (Table). Conclusions: The predictive value of ctDNA for prediction of pCR and RCB differed between subtypes (HR+/HER2- vs. TNBC), while similar prognostic value was observed. In TNBC, early clearance of ctDNA at 3 weeks was a significant predictor of favorable response to NAC. Compared to patients who were ctDNA-positive after NAC, ctDNA-negative status in both subtypes was associated with improved DRFS even in patients with residual cancer (no pCR or RCB-II/III). These findings could inform on the design of future studies that seek to demonstrate the utility of ctDNA in the curative setting. Predictive and prognostic significance of ctDNA in early breast cancer in the neoadjuvant setting HR+HER2- (n=106) TNBC (n=97) Predictive value for prediction of pCR and RCB Fisher p-value Fisher p-value Early ctDNA clearance (between T0 and T1) and pCR 0.4521 &lt;0.0001 Late ctDNA clearance (between T0 and T2) and pCR 0.8071 0.0004 Early ctDNA clearance (between T0 and T1) and RCB (0-III) 0.1360 &lt;0.0001 Late ctDNA clearance (between T0 and T2) and RCB (0-III) 0.4869 0.0004 Early ctDNA clearance at T1 and pCR rates pCR rate pCR rate ctDNA clearance (ctDNA+ at T0/ctDNA- at T1) 21% 67% Late ctDNA clearance (betweeNo early clearance (ctDNA+ at T0/ctDNA+ at T1) 13% 14% Prognostic value for prediction of DRFS Log rank p-value Log rank p-value ctDNA at T3 and pCR vs no PCR 0.0002 &lt;0.0001 ctDNA at T3 and RCB (0-I vs II-III) 0.0110 &lt;0.0001 Timepoints: T0 - pretreatment; T1 - three weeks after treatment initiation; T2 - at 12 weeks, between paclitaxel-based and anthracycline regimens; T3- after neoadjuvant chemotherapy prior to surgery Citation Format: Mark Jesus Mendoza Magbanua, Lamorna Brown Swigart, Derrick Renner, Svetlana Shchegrova, Gillian L. Hirst, Christina Yau, Denise M. Wolf, Hsin-Ta Wu, Ekaterina Kalashnikova, Amy L. Delson, A. Jo Chien, Debu Tripathy, Smita Asare, Raheleh Salari, Angel Rodriguez, Bernhard Zimmermann, Himanshu Sethi, Alexey Aleshin, Paul Billings, Rita Nanda, Hope S. Rugo, Laura J. Esserman, Minetta C. Liu, Angela DeMichele, Laura van 't Veer. Comparison of the predictive and prognostic significance of circulating tumor DNA in patients with high risk HER2-negative breast cancer receiving neoadjuvant chemotherapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr LB111.

514 Background: The remarkable increase of novel Immuno-Oncology drugs in many malignancies has led to the need for biomarkers to identify who would benefit. Various predictive biomarkers have been developed (PD-1/PD-L1 expression, mutations in mismatch repair genes and microsatellite instability, tumor mutational burden and immune infiltration), none have consistently predicted efficacy. The I-SPY2 consortium qualified several expression-based immune biology related signatures that predict response to PD1 checkpoint inhibition. Here we assessed whole transcriptome data of high-risk early-breast cancer (EBC) patients who received Pembrolizumab within the neoadjuvant biomarker-rich I-SPY2 trial (NCT01042379), aiming to migrate the I-SPY2 research findings to a robust clinical grade platform signature to predict sensitivity to PD1 checkpoint inhibition. Methods: Whole transcriptome microarray data were available from pre-treatment biopsies of 69 HER2- patients enrolled in the Pembrolizumab (4 cycles) arm of the I-SPY2 trial. All patients had a High-Risk 70-gene MammaPrint profile. Pathologic complete response (pCR) was defined as no residual invasive cancer in breast or nodes at the time of surgery. Of the 69 patients, 31 had a pCR (12 HR (hormonal receptor)+HER2-, 19 Triple Negative (TN)), while 38 (28 HR+HER2-, 10 TN) had residual disease (RD). To identify the most predictive genes associated with pCR, gene selection was performed comparing pCR and RD groups by iteratively splitting the dataset in training and test, balancing for HR status. Due to limited sample size, leave one out cross validation was used for performance assessment. Genes with effect size > 0.45 were considered significant. Results: A signature of 53 genes, named ImPrint, was identified with overall sensitivity and specificity > 90% and > 80% for predicting pCR to pembrolizumab in all patients. Sensitivity and specificity in TN were > 95% and ≥70%, and in HR+HER2- > 80% and > 85%, respectively. The Positive Predictive Value (PPV) is 77% for the HR+HER2- subgroup. Biological annotation of the 53 genes showed that over 90% of the genes have known immune system related functions, of which 63% were previously known to be involved in immune response (including genes coding PD-L1 and PD-1, as well as those identified in I-SPY2). Conclusions: In the signature development phase, ImPrint predicts pCR to Pembrolizumab in a set of 69 high risk EBC with high sensitivity and specificity. The signature features genes with immune-related functions known to be involved in immune response indicating that it might aid identifying patients with an immune-active phenotype. Importantly, ImPrint appears effective in identifying a subset of HR+HER2- patients who could benefit from immunotherapy. External validation in independent dataset(s) is ongoing and will be presented at the time of the meeting.

Abstract Background: I-SPY 2 is a multicenter, phase 2 trial using response-adaptive randomization within molecular subtypes defined by receptor status and MammaPrint (MP) risk to evaluate novel agents as neoadjuvant therapy for women with high-risk breast cancer. Tucatinib is a potent HER2 (ErbB2) tyrosine kinase inhibitor, selective for HER2 vs. epidermal growth factor receptor (EGFR) and is active vs. brain metastases. Safety and efficacy of tucatinib combined with paclitaxel, pertuzumab, and trastuzumab are unknown and were tested in a planned 10 patient (pt) safety run-in of the I-SPY 2 trial. Methods: Women with tumors ≥ 2.5cm were eligible for screening. Only pts with tumors that were HER2+ by FISH were eligible for this treatment. Treatment included tucatinib (max dose 300 mg) BID for 12 weeks with weekly paclitaxel 80 mg/m2 and trastuzumab (2 mg/kg weekly following loading), and pertuzumab (420 mg every 3 weeks following loading), followed by doxorubicin/cyclophosphamide (AC) every 2 weeks x 4. The control arm was weekly paclitaxel and trastuzumab with pertuzumab for 12 weeks followed by AC every 2 weeks x 4. All pts undergo serial MR imaging and response at 3 & 12 weeks is combined with real time pCR data to estimate, and continuously update, the predicted pCR rate for each trial arm. The goal of the trial is to identify/graduate regimens with ≥85.% Bayesian predictive probability of success (i.e. demonstrating superiority to control) in a future 300-patient phase 3 neoadjuvant trial with a pCR endpoint. This run-in arm was conducted to determine safety of combining tucatinib with paclitaxel/trastuzumab/pertuzumab, monitoring special adverse events of interest including LFT elevations and gastrointestinal toxicities.Methods: The I-SPY 2 methods have been previously published. Results: 20 pts were evaluable in tucatinib treatment arm. The control arm included 329 historical controls enrolled since April 2010. The initial tucatinib dose was 300 mg BID. After enrollment of the first 8 pts, there were 3 pts with grade 3 LFT elevations, 2 pts with grade 2/3 diarrhea, 1 pt with grade 2 neutropenia, and 1 pt with grade 3 nausea. After this safety review, the tucatinib dose was lowered to 250 mg BID. Among 5 additional pts enrolled, 3 developed grade 2/3 LFT abnormalities. The protocol was then modified to tucatinib 150 mg BID days 1-28 and then 250 mg BID days 29-84; 7 pts were treated. Safety data were reviewed after 20 pts were enrolled; the arm was then suspended due to similar LFT elevations regardless of tucatinib dose reduction or schedule. 7 of 20 pts (35%) had reversible Grade 3 or higher ALT/AST elevation (Table). No pt met criteria for Hy’s Law. In terms of efficacy, 12 of 14 evaluable pts had &gt; 80% reduction of tumor volume by 12 weeks, measured by MRI assessment of functional tumor volume (FTV). Conclusion: The goal of the run-in arm was to determine the safety of adding tucatinib to the combination of paclitaxel/trastuzumab/pertuzumab. The addition of tucatinib resulted in unacceptable but reversible LFT elevations despite tucatinib dose reduction. Tucatinib containing therapy resulted in &gt;80% decline in tumor volume at 12 weeks in 86% of pts. Tucatinib showed a high level of activity when combined with paclitaxel/trastuzumab/pertuzumab, but the combination is not feasible. Table: Number of pts with grade 2, 3, and 4 LFT elevations by treatment schedule (highest grade per patient per event, ALT or Treatment scheduleGrade 2 LFT elevationGrade 3 LFT elevationGrade 4 LFT elevationTucatinib 300 mg BID030Tucatinib 250 mg BID210Tucatinib 150 mg BID days 1-28 followed by 250 mg BID days 29 to 84112 Citation Format: David A Potter, Erin Roesch, Christina Yau, Ruixiao Lu, Denise Wolf, Susan Samson, Debra Stafford, Kathy S Albain, Claudine Isaacs, Meghana Trivedi, Douglas Yee, Judy Boughey, Alexandra Thomas, A. Jo Chien, Nola Hylton, Wen Li, Angela DeMichele, Jane Perlmutter, W. Fraser Symmans, Dawn L Hershman, Michelle Melisko, Laura J van 't Veer, Amy Wilson, Smita M Asare, Donald A Berry, Richard Schwab, Hope S Rugo, Laura J Esserman. Evaluation of Tucatinib + (Paclitaxel + Pertuzumab + Trastuzumab) followed by AC in high-risk HER2 positive (HER2+) stage II/III breast cancer: Results from the I-SPY 2 TRIAL [abstract]. In: Proceedings of the 2021 San Antonio Breast Cancer Symposium; 2021 Dec 7-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2022;82(4 Suppl):Abstract nr PD8-07.