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We present a new prognostic classification designated the Van Nuys classification for ductal carcinoma-in-situ (DCIS). The classification combines high nuclear grade and comedo-type necrosis to predict clinical recurrence. Three groups of DCIS patients were defined by the presence or absence of high nuclear grade and comedo-type necrosis: 1--non-high-grade DCIS without comedo-type necrosis, 2--non-high-grade DCIS with comedo-type necrosis, 3--high-grade DCIS with or without comedo-type necrosis. There were 31 local recurrences in 238 patients after breast-conservation surgery 3.8% (3/80) in group 1, 11.1% (10/90) in group 2, and 26.5% (18/68) in group 3. The 8-year actuarial disease-free survivals were 93%, 84%, and 61%, respectively (all p < or = 0.05). The Van Nuys classification defines three distinct and easily recognisable groups, each of which has a different likelihood of local recurrence if treated with breast conservation. [ABSTRACT FROM AUTHOR]

Intrinsic breast cancer molecular subtyping (IBCMS) provides significant prognostic information for patients with breast cancer and helps determine treatment. This study compared IBCMS methods on various gene-expression platforms in PALOMA-2 and PALLET trials. PALOMA-2 tumor samples were profiled using EdgeSeq and nanostring and subtyped with AIMS, PAM50, and research-use-only (ruo)Prosigna. PALLET tumor biopsies were profiled using mRNA sequencing and subtyped with AIMS and PAM50. In PALOMA-2 (n = 222), a 54% agreement was observed between results from AIMS and gold-standard ruoProsigna, with AIMS assigning 67% basal-like to HER2-enriched. In PALLET (n = 224), a 69% agreement was observed between results from PAM50 and AIMS. Different IBCMS methods may lead to different results and could misguide treatment selection; hence, a standardized clinical PAM50 assay and computational approach should be used.Trial number: NCT01740427., (© 2024. The Author(s).)

Clinical trials frequently include multiple end points that mature at different times. The initial report, typically based on the primary end point, may be published when key planned coprimary or secondary analyses are not yet available. Clinical trial updates provide an opportunity to disseminate additional results from studies, published in JCO or elsewhere, for which the primary end point has already been reported. PALOMA-2 demonstrated statistically and clinically significant improvement in progression-free survival with palbociclib plus letrozole versus placebo plus letrozole in estrogen receptor-positive/human epidermal growth factor receptor 2-negative (ER+/HER2-) advanced breast cancer (ABC). Here, we report results for the secondary end point overall survival (OS). Postmenopausal women (N = 666) with ER+/HER2- ABC without previous systemic therapy for ABC were randomly assigned 2:1 to palbociclib plus letrozole or placebo plus letrozole. After a median follow-up of 90.1 months, 405 deaths were observed and 155 patients were known to be alive. The median OS was 53.9 months (95% CI, 49.8 to 60.8) with palbociclib plus letrozole versus 51.2 months (95% CI, 43.7 to 58.9) with placebo plus letrozole (hazard ratio [HR], 0.96 [95% CI, 0.78 to 1.18]; stratified one-sided P = .34). An imbalance in the number of patients with unknown survival outcome between the treatment arms (13.3% v 21.2%, respectively) limited interpretation of OS results. With recovered survival data, the median OS was 53.8 (95% CI, 49.8 to 59.2) versus 49.8 months (95% CI, 42.3 to 56.4), respectively (HR, 0.92 [95% CI, 0.76 to 1.12]; one-sided P = .21). OS was not significantly improved with palbociclib plus letrozole compared with placebo plus letrozole.

Gastric and pancreatic cancers are malignancies of high unmet clinical need. Expression of CLDN18.2 in these cancers, coupled with it's absence from most normal tissues, provides a potential therapeutic window against this target. We present preclinical development and characterization of a novel therapeutic mAb and antibody-drug conjugate (ADC) targeting CLDN18.2. A humanized CLDN18.2 specific mAb, CLDN18.2-307-mAb, was generated through immunization in mice followed by full humanization of the mouse mAb sequences. Antibody clones were screened by flow cytometry for selective binding to membrane bound CLDN18.2. A CLDN18.2-directed ADC (CLDN18.2-307-ADC) was also generated by conjugating MMAE to CLDN18.2 mAb using a cleavable linker. Tissue expression of CLDN18.2 was determined by IHC assay using a CLDN18.2-specific mAb. CLDN18.2-307-mAb binds with high affinity to CLDN18.2-positive (CLDN18.2+) cells and induces antibody-dependent cell-mediated cytotoxicity (ADCC). Treatment with this CLDN18.2-mAb blocked the growth of CLDN18.2+ gastric and pancreas cancer cell line xenograft (CDX) models. Upon binding to the extracellular domain of this target, the CLDN18.2-ADC/CLDN18.2 protein was internalized and subsequently localized to the lysosomal compartment inducing complete and sustained tumor regressions in CLDN18.2+ CDXs and patient-derived pancreatic cancer xenografts (PDX). A screen of human cancer tissues, by IHC, found 58% of gastric, 60% of gastroesophageal junction, and 20% of pancreatic adenocarcinomas to be positive for membrane expression of CLDN18.2. These data support clinical development of the CLDN18.2-307-mAb and CLDN18.2-307-ADC for treatment of CLDN18.2+ cancers. Both are now being investigated in phase I clinical studies., (©2023 American Association for Cancer Research.)

Background: The phase III MONALEESA-3 trial included first- (1L) and second-line (2L) patients and demonstrated a significant overall survival (OS) benefit for ribociclib + fulvestrant in patients with hormone receptor-positive, human epidermal growth factor receptor 2-negative (HR+/HER2-) advanced breast cancer (ABC) in the final protocol-specified and exploratory (longer follow-up) OS analyses. At the time of these analyses, the full OS benefit of 1L ribociclib was not completely characterized because the median OS (mOS) was not reached. As CDK4/6 inhibitor (CDK4/6i) + endocrine therapy (ET) is now a preferred option for 1L HR+/HER2- ABC, we report an exploratory analysis (median follow-up, 70.8 months; 14.5 months longer than the prior analysis) to fully elucidate the OS benefit in the MONALEESA-3 1L population., Methods: Postmenopausal patients with HR+/HER2- ABC were randomized 2:1 to 1L/2L fulvestrant + ribociclib or placebo. OS in 1L patients (de novo disease or relapse > 12 months from completion of [neo]adjuvant ET) was assessed by Cox proportional hazards model and Kaplan-Meier methods. Progression-free survival 2 (PFS2) and chemotherapy-free survival (CFS) were analyzed. MONALEESA-3 is registered with ClinicalTrials.gov (NCT02422615)., Results: At data cutoff (January 12, 2022; median follow-up time, 70.8 months), mOS was 67.6 versus 51.8 months with 1L ribociclib versus placebo (hazard ratio (HR) 0.67; 95% CI 0.50-0.90); 16.5% and 8.6% of ribociclib and placebo patients, respectively, were still receiving treatment. PFS2 (HR 0.64) and CFS (HR 0.62) favored ribociclib versus placebo. Among those who discontinued treatment, 16.7% and 35.0% on ribociclib or placebo, respectively, received a subsequent CDK4/6i. No new safety signals were observed., Conclusions: This analysis of MONALEESA-3 reports the longest mOS thus far (67.6 months) for 1L patients in a phase III ABC trial. These results in a 1L population show that the OS benefit of ribociclib was maintained through extended follow-up, further supporting its use in HR+/HER2- ABC., (© 2023. BioMed Central Ltd., part of Springer Nature.)

Anthracyclines such as doxorubicin (Dox) are effective chemotherapeutic agents; however, their use is hampered by subsequent cardiotoxicity risk. Our understanding of cardiomyocyte protective pathways activated following anthracycline-induced cardiotoxicity (AIC) remains incomplete. Insulin-like growth factor binding protein (IGFBP) 3 (Igfbp-3), the most abundant IGFBP family member in the circulation, is associated with effects on the metabolism, proliferation, and survival of various cells. Whereas Igfbp-3 is induced by Dox in the heart, its role in AIC is ill-defined. We investigated molecular mechanisms as well as systems-level transcriptomic consequences of manipulating Igfbp-3 in AIC using neonatal rat ventricular myocytes and human-induced pluripotent stem cell-derived cardiomyocytes. Our findings reveal that Dox induces the nuclear enrichment of Igfbp-3 in cardiomyocytes. Furthermore, Igfbp-3 reduces DNA damage, impedes topoisomerase IIβ expression (Top2β) which forms Top2β-Dox-DNA cleavage complex leading to DNA double-strand breaks (DSB), alleviates detyrosinated microtubule accumulation-a hallmark of increased cardiomyocyte stiffness and heart failure-and favorably affects contractility following Dox treatment. These results indicate that Igfbp-3 is induced by cardiomyocytes in an effort to mitigate AIC., (© 2023 Federation of American Societies for Experimental Biology.)

Purpose: Claudin-6 (CLDN6) is expressed at elevated levels in multiple human cancers including ovarian and endometrial malignancies, with little or no detectable expression in normal adult tissue. This expression profile makes CLDN6 an ideal target for development of a potential therapeutic antibody-drug conjugate (ADC). This study describes the generation and preclinical characterization of CLDN6-23-ADC, an ADC consisting of a humanized anti-CLDN6 monoclonal antibody coupled to monomethyl auristatin E (MMAE) via a cleavable linker., Experimental Design: A fully humanized anti-CLDN6 antibody was conjugated to MMAE resulting in the potential therapeutic ADC, CLDN6-23-ADC. The antitumor efficacy of CLDN6-23-ADC was assessed for antitumor efficacy in CLDN6-positive (CLDN6+) and -negative (CLDN6-) xenografts and patient-derived xenograft (PDX) models of human cancers., Results: CLDN6-23-ADC selectively binds to CLDN6, versus other CLDN family members, inhibits the proliferation of CLDN6+ cancer cells in vitro, and is rapidly internalized in CLDN6+ cells. Robust tumor regressions were observed in multiple CLDN6+ xenograft models and tumor inhibition led to markedly enhanced survival of CLDN6+ PDX tumors following treatment with CLDN6-23-ADC. IHC assessment of cancer tissue microarrays demonstrate elevated levels of CLDN6 in 29% of ovarian epithelial carcinomas. Approximately 45% of high-grade serous ovarian carcinomas and 11% of endometrial carcinomas are positive for the target., Conclusions: We report the development of a novel ADC, CLDN6-23-ADC, that selectively targets CLDN6, a potential onco-fetal-antigen which is highly expressed in ovarian and endometrial cancers. CLDN6-23-ADC exhibits robust tumor regressions in mouse models of human ovarian and endometrial cancers and is currently undergoing phase I study., (©2023 The Authors; Published by the American Association for Cancer Research.)

Background: Ribociclib has demonstrated a statistically significant overall survival benefit in pre- and postmenopausal patients with hormone receptor positive/human epidermal growth factor receptor 2 negative (HR+/HER2-) advanced breast cancer. New Adjuvant Trial with Ribociclib [LEE011] (NATALEE) is a trial evaluating the efficacy and safety of adjuvant ribociclib plus endocrine therapy (ET) versus ET alone in patients with HR+/HER2- early nonmetastatic breast cancer (EBC)., Methods/design: NATALEE is a multicenter, randomized, open-label, Phase III trial in patients with HR+/HER2- EBC. Eligible patients include women, regardless of menopausal status, and men aged ⩾18 years. Select patients with stage IIA, stage IIB, or stage III disease (per the anatomic classification in the AJCC Cancer Staging Manual , 8th edition) with an initial diagnosis ⩽18 months prior to randomization are eligible. Patients receiving standard (neo)adjuvant ET are eligible if treatment was initiated ⩽12 months before randomization. Patients undergo 1:1 randomization to ribociclib 400 mg/day (3 weeks on/1 week off) +ET (letrozole 2.5 mg/day or anastrozole 1 mg/day [investigator's discretion] plus goserelin [men or premenopausal women]) or ET alone. Ribociclib treatment duration is 36 months; ET treatment duration is ⩾60 months. The primary end point is invasive disease-free survival., Discussion: The 36-month treatment duration of ribociclib in NATALEE is extended compared with that in other adjuvant cyclin-dependent kinases 4 and 6 (CDK4/6) inhibitor trials and is intended to maximize efficacy due to longer duration of CDK4/6 inhibition. Compared with the 600-mg/day dose used in advanced breast cancer, the reduced ribociclib dose used in NATALEE may improve tolerability while maintaining efficacy. NATALEE includes the broadest population of patients with HR+/HER2- EBC of any Phase III trial currently evaluating adjuvant CDK4/6 inhibitor treatment., Trial Registration: ClinicalTrials.gov identifier: NCT03701334 (https://clinicaltrials.gov/ct2/show/NCT03701334)., Competing Interests: Dr. Slamon reports stock ownership from BioMarin, Pfizer, Amgen, Seattle Genetics, TORL BioTherapeutics, 1200 Pharma; travel support from BioMarin, Pfizer, Novartis; personal fees from Novartis, Eli Lilly; grants from Pfizer, Novartis; founder of 1200 Pharma, TORL BioTherapeutics. Dr. Fasching reports personal fees from Novartis, Pfizer, Daiichi Sankyo, AstraZeneca, Eisai, MSD, Lilly, Pierre Fabre, Seagen, F. Hoffmann-La Roche, Hexal, Agendia, Sanofi, Gilead; institutional funding from BioNTech, Pfizer, Cepheid; research grant from Pfizer. Dr. Hurvitz has nothing to disclose. Dr. Chia reports personal fees and grants to institution from Novartis, Pfizer, F. Hoffmann-La Roche, Eli Lilly. Prof. Crown has nothing to disclose. Dr. Martín reports personal fees from Lilly, Pfizer, AstraZeneca, Novartis, Roche/Genentech, GSK, PharmaMar, Taiho Oncology; research grants from Novartis, Roche-Genentech. Dr. Barrios reports institutional research grants from Pfizer, PharmaMar, Polyphor, Henlius Biotech, Shanghai, Merck KGaA, Millennium, LEO Pharma, ImClone Systems, Exelixis, Medivation, Asana Biosciences, AB Science, Abraxis Biosciences, Daiichi Sankyo, Bristol Myers Squibb, BioMarin, Astellas Pharma, AbbVie, Merck (MSD), Merrimack, Mylan, Taiho Pharmaceutical, Sanofi, GSK, Roche/Genentech, Lilly, Boehringer Ingelheim, Novartis, AstraZeneca, Amgen, Pfizer; personal fees from Boehringer Ingelheim, Sanofi, Lilly, Zodiac, AstraZeneca, MSD, Bayer, Eisai, Roche/Genentech, Pfizer, Novartis, GSK, Daiichi Sankyo; stock from MEDSIR, Thummi. Dr. Bardia reports research grants to institution from Genentech, Novartis, Pfizer, Merck, Sanofi, Radius Health, Immunomedics, Mersana, Innocrin; personal fees from Biothermostics, Pfizer, Novartis, Genentech, Merck, Radius Health, Immunomedics, Spectrum Pharma, Taiho, Sanofi, Daiichi Sankyo, Puma. Dr. Im reports personal fees from AstraZeneca, Novartis, Hanmi, Pfizer, Eisai, Amgen, Roche, Lilly, GSK, MSD; research grants from AstraZeneca, Pfizer, Eisai, F. Hoffmann-La Roche, Daewoong Pharmaceuticals. Dr. Yardley reports research funding to institution from Daiichi Sankyo/Lilly, Eisai, Roche/Genentech, Novartis, AbbVie, AstraZeneca, Clovis Oncology, Immunomedics, InventisBio, Lilly, MedImmume, Medivation, Merck, Oncothyreon, Pfizer, Syndax, Tesaro; personal fees from Biotheranostics, Bristol Myers Squibb, Celgene, Daiichi Sankyo/Lilly, Eisai, Roche/Genentech, Novartis, NanoString Technologies. Dr. Untch has nothing to disclose. Dr. Huang reports grants to institution from Novartis, Daiichi Sankyo, AstraZeneca, EirGenix, Eli Lilly, MSD, OBI Pharma, Pfizer, F. Hoffmann-La Roche; personal fees from Novartis, Daiichi Sankyo, AstraZeneca, Eli Lilly, Pfizer, F. Hoffmann-La Roche; nonfinancial support from AstraZeneca, EirGenix, Eli Lilly, OBI Pharma, Roche, Novartis. Dr. Stroyakovskii has nothing to disclose. Dr. Xu reports personal fees from Novartis, AstraZeneca, Pfizer, Roche, Eisai. Dr. Moroose reports personal fees from Gilead, Lilly, Pfizer, Seagen, Genentech, Johnson & Johnson. Dr. Loi reports research funding grants to institution from Novartis, Bristol Myers Squibb, Merck, Puma Biotechnology, Eli Lilly, Nektar Therapeutics, AstraZeneca, Roche/Genentech, Seagen; uncompensated consultant for Seagen, Novartis, Bristol Myers Squibb, Merck, AstraZeneca, Eli Lilly, Pfizer, Gilead Therapeutics, Roche/Genentech; consultant with fees paid to institution from Aduro Biotech, Novartis, GSK, Roche/Genentech, AstraZeneca, Silverback Therapeutics, G1 Therapeutics, PUMA Biotechnologies, Pfizer, Gilead Therapeutics, Seagen, Daiichi Sankyo, Merck, Amunix, Tallac Therapeutics, Eli Lilly, Bristol Myers Squibb. Ms. Visco has nothing to disclose. Ms. Bee-Munteanu has nothing to disclose. Ms. Afenjar has nothing to disclose. Dr. Fresco reports other advising institution is contracted by Novartis as CRO conducting the NATALEE trial. Dr. Taran, Dr. Chakravartty, Dr. Zarate, Dr. Lteif report employment and stock ownership. Dr. Hortobagyi reports personal fees and grants to institution from Novartis., (© The Author(s), 2023.)

Purpose: To evaluate the efficacy and safety of first-line, single-agent trastuzumab in women with HER2 -overexpressing metastatic breast cancer., Patients and Methods: One hundred fourteen women with HER2 -overexpressing metastatic breast cancer were randomized to receive first-line treatment with trastuzumab 4 mg/kg loading dose, followed by 2 mg/kg weekly, or a higher 8 mg/kg loading dose, followed by 4 mg/kg weekly., Results: The objective response rate was 26% (95% confidence interval [CI], 18.2% to 34.4%), with seven complete and 23 partial responses. Response rates in 111 assessable patients with 3+ and 2+ HER2 overexpression by immunohistochemistry (IHC) were 35% (95% CI, 24.4% to 44.7%) and none (95% CI, 0% to 15.5%), respectively. The clinical benefit rates in assessable patients with 3+ and 2+ HER2 overexpression were 48% and 7%, respectively. The response rates in 108 assessable patients with and without HER2 gene amplification by fluorescence in situ hybridization (FISH) analysis were 34% (95% CI, 23.9% to 45.7%) and 7% (95% CI, 0.8% to 22.8%), respectively. Seventeen (57%) of 30 patients with an objective response and 22 (51%) of 43 patients with clinical benefit had not experienced disease progression at follow-up at 12 months or later. The most common treatment-related adverse events were chills (25% of patients), asthenia (23%), fever (22%), pain (18%), and nausea (14%). Cardiac dysfunction occurred in two patients (2%); both had histories of cardiac disease and did not require additional intervention after discontinuation of trastuzumab. There was no clear evidence of a dose-response relationship for response, survival, or adverse events., Conclusion: Single-agent trastuzumab is active and well tolerated as first-line treatment of women with metastatic breast cancer with HER2 3+ overexpression by IHC or gene amplification by FISH.

Purpose: Overexpression of the HER2 protein occurs in 25% to 30% of human breast cancers and leads to a particularly aggressive form of the disease. Efficacy and safety of recombinant humanized anti-HER2 monoclonal antibody as a single agent was evaluated in women with HER2-overexpressing metastatic breast cancer that had progressed after chemotherapy for metastatic disease., Patients and Methods: Two hundred twenty-two women, with HER2-overexpressing metastatic breast cancer that had progressed after one or two chemotherapy regimens, were enrolled. Patients received a loading dose of 4 mg/kg intravenously, followed by a 2-mg/kg maintenance dose at weekly intervals., Results: Study patients had advanced metastatic disease and had received extensive prior therapy. A blinded, independent response evaluation committee identified eight complete and 26 partial responses, for an objective response rate of 15% in the intent-to-treat population (95% confidence interval, 11% to 21%). The median duration of response was 9.1 months; the median duration of survival was 13 months. The most common adverse events, which occurred in approximately 40% of patients, were infusion-associated fever and/or chills that usually occurred only during the first infusion, and were of mild to moderate severity. These symptoms were treated successfully with acetaminophen and/or diphenhydramine. The most clinically significant adverse event was cardiac dysfunction, which occurred in 4.7% of patients. Only 1% of patients discontinued the study because of treatment-related adverse events., Conclusion: Recombinant humanized anti-HER2 monoclonal antibody, administered as a single agent, produces durable objective responses and is well tolerated by women with HER2-overexpressing metastatic breast cancer that has progressed after chemotherapy for metastatic disease. Side effects that are commonly observed with chemotherapy, such as alopecia, mucositis, and neutropenia, are rarely seen.

Background: IGF signaling has been implicated in the pathogenesis and progression of ovarian carcinoma (OC). Single agent activity and safety of ganitumab (AMG 479), a fully human monoclonal antibody against IGF1R that blocks binding of IGF1 and IGF2, were evaluated in patients with platinum-sensitive recurrent OC., Methods: Patients with CA125 progression (GCIG criteria) or measurable disease per RECIST following primary platinum-based therapy received 18 mg/kg of ganitumab q3w. The primary endpoint was objective response rate (ORR) assessed per RECIST 1.1 by an independent radiology review committee (IRC) and/or GCIG CA125 criteria. Secondary endpoints included clinical benefit rate (CBR), progression free survival (PFS) and overall survival (OS)., Results: 61 pts. were accrued. Objective responses were seen in 5/61 patients (ORR 8.2%, 95% CI, 3.1-18.8) with 1 partial response (PR) by RECIST and 2 complete responses (CR) as well as 2 PR by CA125 criteria. CBR was 80.3% (95% CI, 67.8-89.0%). The median PFS according to RECIST by IRC was 2.1 months (95% CI, 2.0-3.1). The median PFS per RECIST IRC and/or CA125 was 2.0 months (95% CI, 1.8-2.2). The median OS was 21 months (95% CI, 19.5-NA). The most common overall adverse events were fatigue (36.1%) and hypertension (34.4%). Grade 1/2 hyperglycemia occurred in 30.4% of patients. Hypertension (11.5%) and hypersensitivity (8.2%) were the most frequent grade 3 adverse events., Conclusions: IGF1R inhibition with ganitumab was well-tolerated, however, our results do not support further study of ganitumab as a single agent in unselected OC patients., Competing Interests: Declaration of Competing Interest The authors declare no potential conflicts of interest., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Background: KRISTINE is an open-label, phase III study of trastuzumab emtansine + pertuzumab (T-DM1 + P) versus docetaxel + carboplatin + trastuzumab + pertuzumab (TCH + P) in patients with HER2-positive, stage II-III breast cancer. We investigated the association of biomarkers with clinical outcomes in KRISTINE., Methods: Patients were randomized to receive neoadjuvant T-DM1 + P or TCH + P and assessed for pathologic complete response (pCR; ypT0/is, ypN0). HER2 status (per central assessment), hormone receptor status, PIK3CA mutation status, HER2/HER3 mRNA levels, tumor-infiltrating lymphocyte levels, PD-L1 status, and NanoString data were analyzed. pCR rates by treatment arm were compared across biomarker subgroups. Analyses were descriptive., Results: Biomarker analyses included data from all 444 patients (T-DM1 + P, n = 223; TCH + P, n = 221) enrolled in KRISTINE. Biomarker distribution was balanced across treatment arms. All subgroups with higher HER2 amplification/expression and immune marker levels showed numerically higher pCR rates in both arms. Mutated versus non-mutated PIK3CA tumors were associated with numerically lower pCR rates in the T-DM1 + P arm but not in the TCH + P arm. In a multivariate analysis, Prediction Analysis of Microarray with the 50-gene classifier (PAM50) HER2-enriched subtype, HER2 gene ratio ≥ 4, and PD-L1-positive status positively influenced the pCR rate. Biomarkers associated with lower pCR rates (e.g., low HER2 levels, positive hormone receptor status, mutated PIK3CA) were more likely to co-occur. Dynamic on-treatment biomarker changes were observed. Differences in the treatment effects for T-DM1 + P versus TCH + P were similar to those observed in the intent-to-treat population for the majority of the biomarker subgroups., Conclusions: Although our biomarker analysis did not identify a subgroup of patients that benefited from neoadjuvant T-DM1 + P versus TCH + P, the data revealed that patients with higher HER2 amplification/expression and immune marker levels had improved response irrespective of treatment arm. These analyses confirm the role of HER2 tumor biology and the immune microenvironment in influencing pCR in the neoadjuvant setting and reaffirm the molecular diversity of HER2-positive breast cancer., Trial Registration: ClinicalTrials.gov NCT02131064. Registered 06 May 2014., (© 2023. The Author(s).)

Purpose: On the basis of preclinical data, we hypothesized that low doses of chemotherapy (10% of therapeutic doses) with full dose of a PARP inhibitor could have improved efficacy and tolerability., Patients and Methods: In this phase I dose-escalation study, patients with BRCA-normal advanced malignancies were assigned to either talazoparib/temozolomide or talazoparib/irinotecan. Talazoparib was dose-escalated from 500 mcg to 1 mg daily before dose escalation of temozolomide/irinotecan. The starting dose of temozolomide was 25 mg/m2/day orally on days 1 to 5 and irinotecan was 25 mg/m2/day intravenously on days 1 and 15. The primary objectives of this trial were safety and tolerability, dose-limiting toxicities (DLT), and maximum tolerated dose (MTD)., Results: Of 40 patients enrolled, 18 (mean: 7 prior therapies) were enrolled in talazoparib + temozolomide and 22 in talazoparib + irinotecan. DLTs were hematologic in both arms, but all hematologic adverse events resolved with either treatment interruption and/or dose reductions of talazoparib. The MTDs were talazoparib 1 mg + temozolomide 37.5 mg/m2 and talazoparib 1 mg + irinotecan 37.5 mg/m2. There were four partial responses in the talazoparib + temozolomide arm and five in the talazoparib + irinotecan arm for a response rate of 23% (9/40). The pharmacokinetic profiles of talazoparib + temozolomide/irinotecan were similar to that of talazoparib monotherapy. Responses were seen independent of homologous recombination (HR) status and HR deficiency score., Conclusions: These results show that talazoparib with low-dose temozolomide or irinotecan is reasonably well tolerated and demonstrates clinical activity in a wide range of cancers. Randomized trials of talazoparib with or without low-dose chemotherapy are ongoing in small cell lung cancer and ovarian cancer., (©2022 American Association for Cancer Research.)

Pharmacologic inhibitors of cyclin-dependent kinases 4 and 6 (CDK4 and 6) are approved for the treatment of subsets of patients with hormone receptor positive (HR+) breast cancer (BC). In metastatic disease, strategies involving endocrine therapy combined with CDK4 and 6 inhibitors (CDK4 and 6i) improve clinical outcomes in HR+ BCs. CDK4 and 6i prevent retinoblastoma tumor suppressor protein phosphorylation, thereby blocking the transcription of E2F target genes, which in turn inhibits both mitogen and estrogen-mediated cell proliferation. In this review, we summarize preclinical data pertaining to the use of CDK4 and 6i in BC, with a particular focus on several of the unique chemical, pharmacologic, and mechanistic properties of abemaciclib. As research efforts elucidate the novel mechanisms underlying abemaciclib activity, potential new applications are being identified. For example, preclinical studies have demonstrated abemaciclib can exert antitumor activity against multiple tumor types and can cross the blood-brain barrier. Abemaciclib has also demonstrated distinct activity as a monotherapeutic in the treatment of BC. Accordingly, we also discuss how a greater understanding of mechanisms related to CDK4 and 6 blockade highlight abemaciclib's unique in-class properties, and could pave new avenues for enhancing its therapeutic efficacy., (© The Author(s) 2022. Published by Oxford University Press.)

Purpose: To conduct an updated exploratory analysis of overall survival (OS) with a longer median follow-up of 73.3 months and evaluate the prognostic value of molecular analysis by circulating tumor DNA (ctDNA)., Patients and Methods: Patients with hormone receptor-positive/human epidermal growth factor receptor 2-negative (HR+/HER2-) advanced breast cancer (ABC) were randomized 2:1 to receive palbociclib (125 mg orally/day; 3/1 week schedule) and fulvestrant (500 mg intramuscularly) or placebo and fulvestrant. This OS analysis was performed when 75% of enrolled patients died (393 events in 521 randomized patients). ctDNA analysis was performed among patients who provided consent., Results: At the data cutoff (August 17, 2020), 258 and 135 deaths occurred in the palbociclib and placebo groups, respectively. The median OS [95% confidence interval (CI)] was 34.8 months (28.8-39.9) in the palbociclib group and 28.0 months (23.5-33.8) in the placebo group (stratified hazard ratio, 0.81; 95% CI, 0.65-0.99). The 6-year OS rate (95% CI) was 19.1% (14.9-23.7) and 12.9% (8.0-19.1) in the palbociclib and placebo groups, respectively. Favorable OS with palbociclib plus fulvestrant compared with placebo plus fulvestrant was observed in most subgroups, particularly in patients with endocrine-sensitive disease, no prior chemotherapy for ABC and low circulating tumor fraction and regardless of ESR1, PIK3CA, or TP53 mutation status. No new safety signals were identified., Conclusions: The clinically meaningful improvement in OS associated with palbociclib plus fulvestrant was maintained with >6 years of follow-up in patients with HR+/HER2- ABC, supporting palbociclib plus fulvestrant as a standard of care in these patients., (©2022 The Authors; Published by the American Association for Cancer Research.)

Pharmacologically targeting the HER2 oncoprotein with therapeutics such as the mAb, trastuzumab, provides clinical benefit for patients with HER2-positive (HER2+) cancers. However, a significant number of patients eventually progress on these therapies. Efforts to overcome therapeutic resistance through combination therapy with small-molecule inhibitors of HER2 have been limited by toxicities associated with off-target activity and/or limited efficacy. In this preclinical study, we explore single-agent and combined activity of tucatinib, a novel HER2-selective small-molecule inhibitor. Tucatinib demonstrated potent, selective activity in a panel of 456 human cancer cell lines, with activity restricted to cell lines (breast and non-breast) with HER2-amplification, including models of acquired resistance to trastuzumab. Within the HER2+ population, tucatinib response tracked strongly with HER2-driven signaling. Single-agent tucatinib induced tumor regressions in xenograft models of HER2+ breast cancer and combination with trastuzumab induced a complete and sustained blockade of HER2/PI3K/AKT signaling. Efficacy of the tucatinib/trastuzumab combination matched that induced by current standard-of-care trastuzumab/pertuzumab/docetaxel combination, with the exception that the chemotherapy-sparing tucatinib/trastuzumab combination did not require a dosing holiday to achieve the same efficacy. In xenograft models of HER2+ breast cancer that also express estrogen receptor (ER; HER2+/ER+), tucatinib showed combined efficacy with inhibitors of CDK4/6 and ER, indicating potential novel therapeutic strategies for difficult-to-treat subtypes of HER2+ breast cancer. These data support expanded clinical investigations of tucatinib as a combination partner for other novel and approved targeted therapies for HER2-driven malignancies., (©2022 American Association for Cancer Research.)

Purpose: Cardiac toxicity is a serious potential complication of HER2-directed therapies and anthracyclines. HER2 codon 655 and SLC28A3 gene polymorphisms have been reported to be associated with cardiac toxicity from anti-HER2 and anthracycline therapy, respectively. Association of the polymorphism at HER2 codon 655 with prognosis has also been reported., Experimental Design: Whole blood samples from patients treated on a randomized adjuvant breast cancer trial (BCIRG-006) that compared chemotherapy with or without trastuzumab plus either anthracycline or nonanthracycline chemotherapy were tested for genetic polymorphisms in HER2 codon 655 and SLC28A3. Genotypes were correlated with cardiac function and disease-free survival (DFS) outcomes., Results: Of 3,222 patients enrolled in BCIRG-006, 662 patient samples were successfully genotyped for the rs1136201 allele in HER2 (codon 655): 424 (64%) were AA, 30 (4.5%) were GG, and 208 (31%) were AG genotype. In addition, 665 patient samples were successfully genotyped for the rs7853758 allele in the SLC28A3 gene: 19 (3%) were AA, 475 (71%) were GG, and 171 (26%) were AG genotype. Follow-up time was 10 years. No correlation between DFS, cardiac event rate, or mean left ventricular ejection fraction (LVEF) and rs1136201 genotype was seen in the trastuzumab-treated or non-trastuzumab-treated patients. Moreover, mean LVEF and cardiac event rates were similar in all rs7853758 genotype groups treated with anthracycline-based therapy., Conclusions: In the largest study to date to evaluate whether two polymorphisms are associated with DFS and/or cardiac toxicity in HER2-positive breast cancer treated with trastuzumab and/or anthracyclines, we observed no correlation., (©2022 American Association for Cancer Research.)

Purpose: Insulin-like growth factor (IGF) signaling is implicated in pathogenesis and chemotherapy resistance of epithelial ovarian cancer (EOC). We explored efficacy and safety of adding ganitumab, a monoclonal antibody targeting IGF-1R, to carboplatin/paclitaxel (CP) chemotherapy in patients with primary EOC., Design: Patients were randomly assigned to receive CP/ganitumab (18 mg/kg q3w) or CP/placebo for 6 cycles followed by 6 cycles of single agent ganitumab/placebo maintenance therapy as front-line therapy. Primary endpoint was progression free survival. Secondary endpoints were time to progression and overall survival. Pretreatment samples were prospectively collected for retrospective biomarker analyses., Results: 170 patients enrolled. 165 patients assessable for toxicity. Median PFS was 15.7 months with CP/ganitumab and 16.7 months with CP/placebo (HR 1.23; 95% CI 0.82-1.83, P = 0.313). All grade neutropenia (84.1% vs 71.4%), thrombocytopenia (75.3% vs 57.1%) and hyperglycemia (15.9% vs 2.6%) were more common in the ganitumab group compared to the placebo group. Ganitumab/placebo related serious adverse events were reported in 26.1% of the patients with ganitumab and in 6.5% with placebo. Non-progression related fatal events were more common with ganitumab (5 versus 2 patients). The ganitumab group experienced more dose delays which resulted in lower relative dose intensity of chemotherapy in the experimental group. In an exploratory model IGFBP2 expression was predictive of ganitumab response (treatment interaction; PFS, P = 0.03; OS, P = 0.01)., Conclusion: Addition of ganitumab to CP chemotherapy in primary EOC did not improve PFS. Our results do not support further study of ganitumab in unselected EOC patients., Competing Interests: Declaration of Competing Interest The authors declare no potential conflicts of interest., (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)

It has been over three decades since anthracyclines took their place as the standard chemotherapy backbone for breast cancer in the curative setting. Though the efficacy of anthracycline chemotherapy is not debatable, potentially life-threatening and long-term risks accompany this class of agents, leading some to question their widespread use, especially when newer agents with improved therapeutic indices have become available. Critically assessing when to incorporate an anthracycline is made more relevant in an era where molecular classification is enabling not only the development of biologically targeted therapeutics but also is improving the ability to better select those who would benefit from cytotoxic agents. This comprehensive analysis will present the problem of overtreatment in early-stage breast cancer, review evidence supporting the use of anthracyclines in the pre-taxane era, analyze comparative trials evaluating taxanes with or without anthracyclines in biologically unselected and selected patient populations, and explore published work aimed at defining anthracycline-sensitive tumor types., (© 2021. The Author(s).)

Purpose: Cyclin-dependent kinase 4 (CDK4) and CDK6 inhibitors (CDK4/6i) are highly effective against estrogen receptor-positive (ER + )/HER2 - breast cancer; however, intrinsic and acquired resistance is common. Elucidating the molecular features of sensitivity and resistance to CDK4/6i may lead to identification of predictive biomarkers and novel therapeutic targets, paving the way toward improving patient outcomes., Experimental Design: Parental breast cancer cells and their endocrine-resistant derivatives (EndoR) were used. Derivatives with acquired resistance to palbociclib (PalboR) were generated from parental and estrogen deprivation-resistant MCF7 and T47D cells. Transcriptomic and proteomic analyses were performed in palbociclib-sensitive and PalboR lines. Gene expression data from CDK4/6i neoadjuvant trials and publicly available datasets were interrogated for correlations of gene signatures and patient outcomes., Results: Parental and EndoR breast cancer lines showed varying degrees of sensitivity to palbociclib. Transcriptomic analysis of these cell lines identified an association between high IFN signaling and reduced CDK4/6i sensitivity; thus an "IFN-related palbociclib-resistance Signature" (IRPS) was derived. In two neoadjuvant trials of CDK4/6i plus endocrine therapy, IRPS and other IFN-related signatures were highly enriched in patients with tumors exhibiting intrinsic resistance to CDK4/6i. PalboR derivatives displayed dramatic activation of IFN/STAT1 signaling compared with their short-term treated or untreated counterparts. In primary ER + /HER2 - tumors, the IRPS score was significantly higher in lumB than lumA subtype and correlated with increased gene expression of immune checkpoints, endocrine resistance, and poor prognosis., Conclusions: Aberrant IFN signaling is associated with intrinsic resistance to CDK4/6i. Experimentally, acquired resistance to palbociclib is associated with activation of the IFN pathway, warranting additional studies to clarify its involvement in resistance to CDK4/6i., (©2021 American Association for Cancer Research.)

Non-genetic mechanisms have recently emerged as important drivers of cancer therapy failure 1 , where some cancer cells can enter a reversible drug-tolerant persister state in response to treatment 2 . Although most cancer persisters remain arrested in the presence of the drug, a rare subset can re-enter the cell cycle under constitutive drug treatment. Little is known about the non-genetic mechanisms that enable cancer persisters to maintain proliferative capacity in the presence of drugs. To study this rare, transiently resistant, proliferative persister population, we developed Watermelon, a high-complexity expressed barcode lentiviral library for simultaneous tracing of each cell's clonal origin and proliferative and transcriptional states. Here we show that cycling and non-cycling persisters arise from different cell lineages with distinct transcriptional and metabolic programs. Upregulation of antioxidant gene programs and a metabolic shift to fatty acid oxidation are associated with persister proliferative capacity across multiple cancer types. Impeding oxidative stress or metabolic reprogramming alters the fraction of cycling persisters. In human tumours, programs associated with cycling persisters are induced in minimal residual disease in response to multiple targeted therapies. The Watermelon system enabled the identification of rare persister lineages that are preferentially poised to proliferate under drug pressure, thus exposing new vulnerabilities that can be targeted to delay or even prevent disease recurrence., (© 2021. The Author(s), under exclusive licence to Springer Nature Limited.)

Background: Ribociclib plus fulvestrant demonstrated significant progression-free survival (PFS) and overall survival (OS) benefits in patients with hormone receptor-positive, human epidermal growth factor receptor 2-negative (HR+/HER2-) advanced breast cancer (ABC). Here we present a new landmark in survival follow-up for a phase III cyclin-dependent kinases 4 and 6 inhibitor clinical trial in patients with ABC (median, 56.3 months)., Patients and Methods: This phase III, randomized, double-blind, placebo-controlled trial was conducted at 174 sites (30 countries). Patients were men and postmenopausal women (age ≥18 years) with histologically/cytologically confirmed HR+/HER2- ABC. Patients could have received ≤1 line of endocrine therapy (ET) but no chemotherapy for ABC. Patients, assigned 2:1, were stratified by the presence/absence of liver/lung metastases and previous ET. Patients received intramuscular fulvestrant (500 mg, day 1 of each 28-day cycle plus day 15 of cycle 1) with oral ribociclib (600 mg/day, 3 weeks on, 1 week off) or placebo. Efficacy analyses were by intention to treat. Safety was assessed in patients receiving ≥1 dose study treatment. OS was a secondary endpoint. MONALEESA-3 is registered with ClinicalTrials.gov (NCT02422615; no longer enrolling)., Results: Between 18 June 2015 and 10 June 2016, 726 patients were randomly assigned (484, ribociclib; 242, placebo). At data cut-off (30 October 2020), median OS (mOS) was 53.7 months (ribociclib) versus 41.5 months (placebo) [hazard ratio (HR), 0.73; 95% confidence interval (CI) 0.59-0.90]. Subgroup analyses were consistent with overall population. In the first-line setting, most patients in the ribociclib arm (∼60%) lived longer than median follow-up; mOS was 51.8 months in the placebo arm (HR, 0.64; 95% CI 0.46-0.88). In the second-line setting, mOS was 39.7 months (ribociclib) versus 33.7 months (placebo) (HR, 0.78; 95% CI 0.59-1.04). No apparent drug-drug interaction between ribociclib and fulvestrant or new safety signals were observed., Conclusions: This analysis reported extended OS follow-up in MONALEESA-3. mOS was ∼12 months longer in patients with HR+/HER2- ABC treated with ribociclib plus fulvestrant compared with fulvestrant monotherapy., Competing Interests: Disclosure DJS reports board of directors (stock) and travel expenses from BioMarin; stock ownership, research funding, and travel expenses from Pfizer; advisory board, consulting, research funding, and travel expenses from Novartis; consulting from Eli Lilly; and stock ownership from Amgen and Seattle Genetics. SC reports that his institution received grants and personal fees for advisory boards from Novartis, Pfizer, Hoffmann-La Roche, and Eli Lilly, outside the submitted work. GJ reports personal fees from Novartis, during the conduct of the study; grants, personal fees, and non-financial support from Novartis, Roche, and Pfizer; personal fees and non-financial support from Lilly, Amgen, BMS, and AstraZeneca; personal fees from AbbVie and Daiichi-Sankyo; and non-financial support from Med-immune and Merck KGaA, outside the submitted work. MDL reports personal fees for speaker honoraria and advisory board honoraria from Novartis, AstraZeneca, Eli Lilly, and Pierre Fabre, outside the submitted work. SI reports grants from AstraZeneca, Pfizer, Eisai, and Daewoong; advisory and personal fees from AstraZeneca, Novartis, Hanmi, Pfizer, Eisai, Amgen, MediPacto, Roche, Eli Lilly, MSD, and GlaxoSmithKline; and non-financial support from Novartis. KP reports personal fees for advisory board from Novartis, AstraZeneca, Roche, Pfizer, and BMS, outside the submitted work. GVB reports personal fees for advisory board from Novartis and Eli Lilly, outside the submitted work. MM reports personal fees for speaker honoraria and honoraria for participation in advisory boards from Lilly and Pfizer; honoraria for participation in advisory boards from AstraZeneca, GlaxoSmithKline, Pharmamar, and Taiho Oncology; and research grants and honoraria for participation in advisory boards from Novartis and Roche-Genentech, outside the submitted work. AN reports consulting/advisory role, travel/accommodation/expenses, and research funding from Novartis and consulting/advisory role from Amgen during the conduct of the study. GSS reports institutional reimbursement for patient accrual and education and steering committee activities from Novartis and institutional research support from Merck, AstraZeneca, and Roche, outside the submitted work. LDLC-M reports personal fees for consultant/advisory role, research funding, and speaking engagements from MSD-Merck; consultant/advisory role, research funding, speaking engagements, and grant support from Roche Farma; consultant/advisory role, speaking engagements, and grant support from Bristol Myers Squibb; consultant/advisory role from Novartis and Pierre Fabre; consultant/advisory role and speaking engagements from Amgen; and research funding from Celgene, outside the submitted work. JTB reports grants for institutional funding for doing research from AbbVie, Alliance, Amgen, Ascentage Pharma Group, AstraZeneca, Bayer, Boston Biomedial, Bristol Myers Squibb, Celgene, Eli Lilly, Genentech-Roche, Hutchison, Immunomedics, Janssen, MT Group, Nektar, Pfizer, Polynoma, Seattle Genetics, Serono-EMD, Tesaro, TG Therapeutics, Biodesix, Exact Sciences, Boehringer Ingleheim, Laekna, Novocure, Daiichi-Sankyo, Mirati Therapeutics, and Tarveda Therapeutics, during the conduct of the study. YJ, CW, UD, AC, JPZ, and TT report employment and stock ownership from Novartis. PAF reports personal fees for advisory board from Celgene, Merck Sharp & Dohme, Macrogenics, Eisai, Puma, Lilly, and AstraZeneca; lectures from Daiichi-Sankyo, Merck Sharp & Dohme, and Lilly; and research support from Cepheid and Novartis, outside the submitted work. PN has declared no conflicts of interest. Data sharing Novartis made the study protocols available for MONALEESA-3 at the time of primary publications. Individual participant data will not be made available., (Copyright © 2021 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Our previous pre-clinical work defined BCL-2 induction as a critical component of the adaptive response to lapatinib-mediated inhibition of HER2. To determine whether a similar BCL-2 upregulation occurs in lapatinib-treated patients, we evaluated gene expression within tumor biopsies, collected before and after lapatinib or trastuzumab treatment, from the TRIO-B-07 clinical trial (NCT#00769470). We detected BCL2 mRNA upregulation in both HER2+/ER- as well as HER2+/ER+ patient tumors treated with lapatinib or trastuzumab. To address whether mRNA expression correlated with protein expression, we evaluated pre- and post-treatment tumors for BCL-2 via immunohistochemistry. Despite BCL2 mRNA upregulation within HER2+/ER- tumors, BCL-2 protein levels were undetectable in most of the lapatinib- or trastuzumab-treated HER2+/ER- tumors. BCL-2 upregulation was evident within the majority of lapatinib-treated HER2+/ER+ tumors and was often coupled with increased ER expression and decreased proliferation. Comparable BCL-2 upregulation was not observed within the trastuzumab-treated HER2+/ER+ tumors. Together, these results provide clinical validation of the BCL-2 induction associated with the adaptive response to lapatinib and support evaluation of BCL-2 inhibitors within the context of lapatinib and other HER2-targeted receptor tyrosine kinase inhibitors., Competing Interests: I have read the journal’s policy and the authors of this manuscript have the following competing interests: S.A. Hurvitz Contracted Research: Ambrx Amgen Astra Zeneca Arvinas Bayer Daiichi-Sankyo Genentech/Roche Gilead GSK Immunomedics Lilly Macrogenics Novartis Pfizer OBI Pharma Pieris PUMA Radius Sanofi Seattle Genetics Dignitana Zymeworks Phoenix Molecular Designs, Ltd. Stock Options: NK Max Travel: Lilly M.F. Press Consulting or Advisory Role: AstraZeneca Biocartis SA CEPHEID Clinical Care Options, LLC CME Outfitters, LLC Lilly USA, LLC Merck & Co. Puma Biotechnology Zymeworks Inc. Eli Lilly & Company Private Equity Role: TORL BIOTHERAPEUTICS LLC All other authors have declared that no competing interests exist. We have confirmed that the declared competing interests do not alter adherence to all PLOS ONE policies on sharing data and materials.

The addition of HER2-targeted agents to neoadjuvant chemotherapy has dramatically improved pathological complete response (pCR) rates in early-stage, HER2-positive breast cancer. Nonetheless, up to 50% of patients have residual disease after treatment, while others are likely overtreated. Here, we performed multiplex spatial proteomic characterization of 122 samples from 57 HER2-positive breast tumors from the neoadjuvant TRIO-US B07 clinical trial sampled pre-treatment, after 14-21 d of HER2-targeted therapy and at surgery. We demonstrated that proteomic changes after a single cycle of HER2-targeted therapy aids the identification of tumors that ultimately undergo pCR, outperforming pre-treatment measures or transcriptomic changes. We further developed and validated a classifier that robustly predicted pCR using a single marker, CD45, measured on treatment, and showed that CD45-positive cell counts measured via conventional immunohistochemistry perform comparably. These results demonstrate robust biomarkers that can be used to enable the stratification of sensitive tumors early during neoadjuvant HER2-targeted therapy, with implications for tailoring subsequent therapy.

Objective: There is an immunoreactive subtype of ovarian cancer with a favorable prognosis, but the majority of ovarian cancers have limited immune reactivity. The reason for this is poorly understood. This study aimed to approach this question by identifying prognostically relevant genes whose prognostic mRNA expression levels correlated with a genomic event., Methods: Expression microarray and 5-year survival data on 170 ovarian tumors and aCGH data on 45 ovarian cancer cell lines were used to identify amplified/deleted genes associated with prognosis. Three immune-response genes were identified mapping to epigenetically modified chromosome 6p21.3. Genes were searched for roles in epigenetic modification, identifying KANSL1. Genome-wide association studies were searched to identify genetic variants in KANSL1 associated with altered immune profile. Sensitivity to HDAC inhibition in cell lines with KANSL1 amplification/rearrangement was studied., Results: Expression of 196 genes was statistically significantly associated with survival, and expression levels correlated with copy number variations for 82 of them. Among these, 3 immune-response genes (HCP5, PSMB8, PSMB9) clustered together at epigenetically modified chromosome 6p21.3 and their expression was inversely correlated to epigenetic modification gene KANSL1. KANSL1 is amplified/rearranged in ovarian cancer, associated with lymphocyte profile, a biomarker for response to HDAC inhibition, and may drive expression of immune-response genes., Conclusion: This study identifies 82 genes with prognostic relevance and genomic alteration in ovarian cancer. Among these, immune-response genes have correlated expression which is associated with 5-year survival. KANSL1 may be a master gene altering immune-response gene expression at 6p21.3 and drive response to HDAC inhibitors. Future research should investigate KANSL1 and determine whether targeting it alters the immune profile of ovarian cancer and improves survival, HDAC inhibition, and/or immunotherapy response., Competing Interests: Declaration of Competing Interest The authors have no conflicts of interest to declare., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Purpose of Review: The field of HER2-positive breast cancer has seen tremendous advances in the last 2 years with largest number of new drugs in decades. The present review aims to summarize the cutting-edge research of the past 2 years and future directions., Recent Findings: This review will go over four new drugs, three of which have gained FDA approval within the past 18 months, in the treatment of HER2-positive breast cancer. We will go over early and mature clinical data on these therapeutics and ongoing clinical trials further exploring their role in the treatment of patients with advanced HER2-positive breast cancer and HER2 low breast cancer. Will also discuss ongoing trials using immunotherapy and CDK4/6 inhibitors in the advanced HER2-positive setting., Summary: : The therapies described in this review have quickly become standard of care for patients with HER2-positive breast cancer. Furthermore, they have the potential to change the landscape of breast cancer therapy further to include even patients with HER2 low breast cancer., (Copyright © 2020 Wolters Kluwer Health, Inc. All rights reserved.)

Perturbation biology is a powerful approach to modeling quantitative cellular behaviors and understanding detailed disease mechanisms. However, large-scale protein response resources of cancer cell lines to perturbations are not available, resulting in a critical knowledge gap. Here we generated and compiled perturbed expression profiles of ∼210 clinically relevant proteins in >12,000 cancer cell line samples in response to ∼170 drug compounds using reverse-phase protein arrays. We show that integrating perturbed protein response signals provides mechanistic insights into drug resistance, increases the predictive power for drug sensitivity, and helps identify effective drug combinations. We build a systematic map of "protein-drug" connectivity and develop a user-friendly data portal for community use. Our study provides a rich resource to investigate the behaviors of cancer cells and the dependencies of treatment responses, thereby enabling a broad range of biomedical applications., Competing Interests: Declaration of Interests R.A. is a bioinformatics consultant for the University of Houston. G.B.M. is on the scientific advisory board or a consultant for AstraZeneca, ImmunoMet, Lilly, Nuevolution, PDX Pharmaceuticals, Symphogen, and Tarveda; has stock options with Catena Pharmaceuticals, ImmunoMet, SignalChem, and Tarveda; and has licensed technologies to Myriad and Nanostring. H.L. is a shareholder and scientific advisor of Precision Scientific Ltd., (Copyright © 2020 Elsevier Inc. All rights reserved.)

In this multicenter, open-label, randomized phase II investigator-sponsored neoadjuvant trial with funding provided by Sanofi and GlaxoSmithKline (TRIO-US B07, Clinical Trials NCT00769470), participants with early-stage HER2-positive breast cancer (N = 128) were recruited from 13 United States oncology centers throughout the Translational Research in Oncology network. Participants were randomized to receive trastuzumab (T; N = 34), lapatinib (L; N = 36), or both (TL; N = 58) as HER2-targeted therapy, with each participant given one cycle of this designated anti-HER2 therapy alone followed by six cycles of standard combination chemotherapy with the same anti-HER2 therapy. The primary objective was to estimate the rate of pathologic complete response (pCR) at the time of surgery in each of the three arms. In the intent-to-treat population, we observed similar pCR rates between T (47%, 95% confidence interval [CI] 30-65%) and TL (52%, 95% CI 38-65%), and a lower pCR rate with L (25%, 95% CI 13-43%). In the T arm, 100% of participants completed all protocol-specified treatment prior to surgery, as compared to 69% in the L arm and 74% in the TL arm. Tumor or tumor bed tissue was collected whenever possible pre-treatment (N = 110), after one cycle of HER2-targeted therapy alone (N = 89), and at time of surgery (N = 59). Higher-level amplification of HER2 and hormone receptor (HR)-negative status were associated with a higher pCR rate. Large shifts in the tumor, immune, and stromal gene expression occurred after one cycle of HER2-targeted therapy. In contrast to pCR rates, the L-containing arms exhibited greater proliferation reduction than T at this timepoint. Immune expression signatures increased in all arms after one cycle of HER2-targeted therapy, decreasing again by the time of surgery. Our results inform approaches to early assessment of sensitivity to anti-HER2 therapy and shed light on the role of the immune microenvironment in response to HER2-targeted agents.

Purpose: This analysis evaluated the relationship between treatment-free interval (TFI, in PALOMA-2)/disease-free interval (DFI, in PALOMA-3) and progression-free survival (PFS) and overall survival (OS, in PALOMA-3), treatment effect in patients with bone-only disease, and whether intrinsic subtype affects PFS in patients receiving palbociclib., Methods: Data were from phase 3, randomized PALOMA-2 and PALOMA-3 clinical studies of hormone receptor‒positive/human epidermal growth factor receptor 2‒negative (HR+ /HER2-) advanced breast cancer (ABC) patients receiving endocrine therapy plus palbociclib or placebo. Subpopulation treatment effect pattern plot (STEPP) analysis evaluated the association between DFI and PFS and OS. PFS by luminal subtype and cyclin-dependent kinase (CDK) 4/6 or endocrine pathway gene expression levels were evaluated in patients with bone-only disease; median PFS and OS were estimated by the Kaplan-Meier method., Results: Median durations of TFI were 37.1 and 30.9 months (PALOMA-2) and DFI were 49.2 and 52.0 months (PALOMA-3) in the palbociclib and placebo groups, respectively. Among the PALOMA-2 biomarker population (n = 454), 23% had bone-only disease; median PFS was longer with palbociclib versus placebo (31.3 vs 11.2 months; hazard ratio, 0.41; 95% CI 0.25‒0.69). The interaction effect of bone-only versus visceral disease subgroups on median PFS with palbociclib was not significant (P = 0.262). Among the PALOMA-3 biomarker population (n = 302), 27% had bone-only disease. STEPP analyses showed that palbociclib PFS benefit was not affected by DFI, and that palbociclib OS effect may be smaller in patients with short DFIs. Among patients who provided metastatic tumor tissues (n = 142), regardless of luminal A (hazard ratio, 0.23; 95% CI 0.11‒0.47; P = 0.0000158) or luminal B (hazard ratio, 0.26; 95% CI 0.12‒0.56; P = 0.000269) subtype, palbociclib improved PFS versus placebo., Conclusions: These findings support palbociclib plus endocrine therapy as standard of care for HR+ /HER2- ABC patients, regardless of baseline TFI/DFI or intrinsic molecular subtype, including patients with bone-only disease., Trial Registration: Pfizer (clinicaltrials.gov:NCT01740427, NCT01942135).

Purpose: Palbociclib is a cyclin-dependent kinase 4/6 (CDK4/6) inhibitor, approved in combination with endocrine therapy for the treatment of women and men with hormone receptor-positive, human epidermal growth factor receptor 2-negative advanced breast cancer (HR+/HER2- ABC). In the phase 2, open-label, PALOMA-1 trial, palbociclib plus letrozole significantly prolonged progression-free survival (PFS) versus letrozole alone (hazard ratio, 0.488; 95% CI 0.319‒0.748; P = 0.0004; median PFS, 20.2 vs 10.2 months, respectively) in postmenopausal women with estrogen receptor-positive (ER+)/HER2- ABC. Here, we present the final overall survival (OS) and updated safety results., Methods: Postmenopausal women with ER+/HER2- ABC were randomized 1:1 to receive either palbociclib (125 mg/day, 3/1 schedule) plus letrozole (2.5 mg/day, continuous) or letrozole alone (2.5 mg/day, continuous). The primary endpoint was investigator-assessed PFS; secondary endpoints included OS and safety., Results: A total of 165 patients were randomized. At the data cutoff date of December 30, 2016 (median duration of follow-up, 64.7 months), the stratified hazard ratio for OS was 0.897 (95% CI 0.623-1.294; P = 0.281); median OS in the palbociclib plus letrozole and letrozole alone arms was 37.5 and 34.5 months, respectively. The median time from randomization to first subsequent chemotherapy use was longer with palbociclib plus letrozole than letrozole alone (26.7 and 17.7 months, respectively). The most frequently reported adverse event in the palbociclib plus letrozole arm was neutropenia (any grade, 75%; grade 3 or 4, 59%)., Conclusions: Palbociclib plus letrozole treatment led to a numerical but not statistically significant improvement in median OS. Pfizer Inc (NCT00721409).

Background: Combined targeting of CDK4/6 and ER is now the standard of care for patients with advanced ER+/HER2- breast cancer. However, acquired resistance to these therapies frequently leads to disease progression. As such, it is critical to identify the mechanisms by which resistance to CDK4/6-based therapies is acquired and also identify therapeutic strategies to overcome resistance., Methods: In this study, we developed and characterized multiple in vitro and in vivo models of acquired resistance to CDK4/6-based therapies. Resistant models were screened by reverse phase protein array (RPPA) for cell signaling changes that are activated in resistance., Results: We show that either a direct loss of Rb or loss of dependence on Rb signaling confers cross-resistance to inhibitors of CDK4/6, while PI3K/mTOR signaling remains activated. Treatment with the p110α-selective PI3K inhibitor, alpelisib (BYL719), completely blocked the progression of acquired CDK4/6 inhibitor-resistant xenografts in the absence of continued CDK4/6 inhibitor treatment in models of both PIK3CA mutant and wild-type ER+/HER2- breast cancer. Triple combination therapy against PI3K:CDK4/6:ER prevented and/or delayed the onset of resistance in treatment-naive ER+/HER2- breast cancer models., Conclusions: These data support the clinical investigation of p110α-selective inhibitors of PI3K, such as alpelisib, in patients with ER+/HER2- breast cancer who have progressed on CDK4/6:ER-based therapies. Our data also support the investigation of PI3K:CDK4/6:ER triple combination therapy to prevent the onset of resistance to the combination of endocrine therapy plus CDK4/6 inhibition.

Palbociclib is a cyclin-dependent kinase 4/6 inhibitor indicated for treatment of hormone receptor-positive/human epidermal growth factor receptor 2-negative advanced breast cancer in combination with endocrine therapy. We investigated the efficacy and safety of palbociclib in patients enrolled in North America during two-phase 3 trials: PALOMA-2 (n = 267, data cutoff: May 31, 2017) and PALOMA-3 (n = 240, data cutoffs: April 13, 2018, for overall survival, October 23, 2015, for all other outcomes). In PALOMA-2, treatment-naïve postmenopausal patients with advanced breast cancer were randomized 2:1 to palbociclib (125 mg/d; 3 weeks on/1 week off [3/1]) plus letrozole (2.5 mg/d, continuous) or placebo plus letrozole. In PALOMA-3, patients who progressed on prior endocrine therapy were randomized 2:1 to palbociclib (125 mg/d; 3/1) plus fulvestrant (500 mg, per standard of care) or placebo plus fulvestrant; pre/perimenopausal patients received ovarian suppression with goserelin. Palbociclib plus endocrine therapy prolonged median progression-free survival vs placebo plus endocrine therapy in North American patients (PALOMA-2: 25.4 vs 13.7 months, hazard ratio, 0.54 [95% CI, 0.40-0.74], P < .0001; PALOMA-3: 9.9 vs 3.5 months, hazard ratio, 0.52 [95% CI, 0.38-0.72], P < .0001). Objective response and clinical benefit response rates were greater with palbociclib vs placebo in North American patients in both trials. While overall survival data are not yet mature for PALOMA-2, median overall survival was increased in PALOMA-3 (32.0 vs 24.7 months, hazard ratio, 0.75 [95% CI, 0.53-1.04]), though this did not reach statistical significance (P = .0869). Safety profiles in North American patients were similar to those of the overall populations; neutropenia was the most common treatment-emergent adverse event. No new safety signals were observed. In summary, palbociclib plus endocrine therapy is an effective treatment option for North American women with hormone receptor-positive/human epidermal growth factor receptor 2-negative advanced breast cancer., (© 2019 The Authors. The Breast Journal published by Wiley Periodicals, Inc.)

Background: In an earlier analysis of this phase 3 trial, ribociclib plus fulvestrant showed a greater benefit with regard to progression-free survival than fulvestrant alone in postmenopausal patients with hormone-receptor-positive, human epidermal growth factor receptor 2 (HER2)-negative advanced breast cancer. Here we report the results of a protocol-specified second interim analysis of overall survival., Methods: Patients were randomly assigned in a 2:1 ratio to receive either ribociclib or placebo in addition to fulvestrant as first-line or second-line treatment. Survival was evaluated by means of a stratified log-rank test and summarized with the use of Kaplan-Meier methods., Results: This analysis was based on 275 deaths: 167 among 484 patients (34.5%) receiving ribociclib and 108 among 242 (44.6%) receiving placebo. Ribociclib plus fulvestrant showed a significant overall survival benefit over placebo plus fulvestrant. The estimated overall survival at 42 months was 57.8% (95% confidence interval [CI], 52.0 to 63.2) in the ribociclib group and 45.9% (95% CI, 36.9 to 54.5) in the placebo group, for a 28% difference in the relative risk of death (hazard ratio, 0.72; 95% CI, 0.57 to 0.92; P = 0.00455). The benefit was consistent across most subgroups. In a descriptive update, median progression-free survival among patients receiving first-line treatment was 33.6 months (95% CI, 27.1 to 41.3) in the ribociclib group and 19.2 months (95% CI, 14.9 to 23.6) in the placebo group. No new safety signals were observed., Conclusions: Ribociclib plus fulvestrant showed a significant overall survival benefit over placebo plus fulvestrant in patients with hormone-receptor-positive, HER2-negative advanced breast cancer. (Funded by Novartis; MONALEESA-3 ClinicalTrials.gov number, NCT02422615.)., (Copyright © 2019 Massachusetts Medical Society.)

Purpose: neoMONARCH assessed the biological effects of abemaciclib in combination with anastrozole in the neoadjuvant setting., Patients and Methods: Postmenopausal women with stage I-IIIB HR + /HER2 - breast cancer were randomized to a 2-week lead-in of abemaciclib, anastrozole, or abemaciclib plus anastrozole followed by 14 weeks of the combination. The primary objective evaluated change in Ki67 from baseline to 2 weeks of treatment. Additional objectives included clinical, radiologic, and pathologic responses, safety, as well as gene expression changes related to cell proliferation and immune response., Results: Abemaciclib, alone or in combination with anastrozole, achieved a significant decrease in Ki67 expression and led to potent cell-cycle arrest after 2 weeks of treatment compared with anastrozole alone. More patients in the abemaciclib-containing arms versus anastrozole alone achieved complete cell-cycle arrest (58%/68% vs. 14%, P < 0.001). At the end of treatment, following 2 weeks lead-in and 14 weeks of combination therapy, 46% of intent-to-treat patients achieved a radiologic response, with pathologic complete response observed in 4%. The most common all-grade adverse events were diarrhea (62%), constipation (44%), and nausea (42%). Abemaciclib, anastrozole, and the combination inhibited cell-cycle processes and estrogen signaling; however, combination therapy resulted in increased cytokine signaling and adaptive immune response indicative of enhanced antigen presentation and activated T-cell phenotypes., Conclusions: Abemaciclib plus anastrozole demonstrated biological and clinical activity with generally manageable toxicities in patients with HR + /HER2 - early breast cancer. Abemaciclib led to potent cell-cycle arrest, and in combination with anastrozole, enhanced immune activation., (©2019 American Association for Cancer Research.)

Purpose: Preclinical data identified the cyclin-dependent kinase 4/6 (CDK4/6) inhibitor palbociclib as synergistic with antiestrogens in inhibiting growth of hormone receptor-positive/human epidermal growth factor receptor 2-negative (HR+/HER2-) human breast cancer models. This observation was validated clinically in the randomized, placebo-controlled, phase III PALOMA-2 study., Experimental Design: To determine markers of sensitivity and resistance to palbociclib plus letrozole, we performed comprehensive biomarker analyses, investigating the correlation with progression-free survival (PFS), on baseline tumor tissues from PALOMA-2., Results: Despite a broad biomarker search, palbociclib plus letrozole demonstrated consistent PFS gains versus placebo plus letrozole, with no single biomarker or cassette of markers associated with lack of benefit from combination treatment. Palbociclib plus letrozole confers efficacy on both luminal A and B patients. Higher CDK4 levels were associated with endocrine resistance which was mitigated by the addition of palbociclib, whereas lower PD-1 levels were associated with greater palbociclib plus letrozole benefit. Tumors with more active growth factor signaling, as exemplified by increased expression of FGFR2 and ERBB3 mRNA, appeared to be associated with greater PFS gain from the addition of palbociclib., Conclusions: These data underscore the importance of CDK4/6 signaling in HR+/HER2- breast cancer and suggest that the interplay between steroid hormone and peptide growth factor signaling could drive dependence on CDK4/6 signaling. See related commentary by Anurag et al., p. 3 ., (©2019 American Association for Cancer Research.)

Background: Luminespib (AUY922) is a second-generation heat shock protein 90 (HSP90) inhibitor with demonstrated activity in non-small cell lung cancer (NSCLC). Since luminespib reduces levels of dihydrofolate reductase (DHFR), a key enzymatic target of pemetrexed, we assessed the safety and tolerability of luminespib in combination with pemetrexed in patients with previously treated metastatic non-squamous non-small cell lung cancer (NSCLC). We also sought to study the pharmacokinetics and correlate tumor dihydrofolate reductase (DHFR) expression with clinical response., Methods: Patients received weekly luminespib at either 40 mg/m 2 , 55 mg/m 2 , or 70 mg/m 2 according to a standard 3 + 3 dose-escalation design along with pemetrexed at 500 mg/m 2 followed by an expansion at the maximum tolerated dose (MTD)., Results: Two-dose limiting toxicities (DLTs) were experienced in the 70 mg/m2 cohort, therefore the MTD was determined to be 55 mg/m 2 . 69% (N = 9) of patients experienced ophthalmologic toxicity related to luminespib. Maximum serum concentration (Cmax) of luminespib was associated with increased grade 2 drug related adverse events (DRAEs) (r s  = 0.74, P <  0.01), with volume of distribution (V D ) inversely associated with the number of DRAEs (r s  = - 0.81, P =  0.004) and ophthalmologic related DRAEs (r s  = - 0.65, P =  0.04). The best response was partial response in one patient for 20 months, prior to expiration of all luminespib. Amongst patients treated at the MTD, the objective response rate was 14%., Conclusion: In patients with previously treated metastatic NSCLC, the MTD of luminespib in combination with pemetrexed was 55 mg/m 2 per week. The combination of luminespib and pemetrexed demonstrated clinical activity. Tolerability of luminespib with pemetrexed is limited by ocular toxicity., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Background: CFI-400945 is a first-in-class oral inhibitor of polo-like kinase 4 (PLK4) that regulates centriole duplication. Primary objectives of this first-in-human phase 1 trial were to establish the safety and tolerability of CFI-400945 in patients with advanced solid tumours. Secondary objectives included pharmacokinetics, pharmacodynamics, efficacy, and recommended phase 2 dose (RP2D)., Methods: Continuous daily oral dosing of CFI-400945 was evaluated using a 3+3 design guided by incidence of dose-limiting toxicities (DLTs) in the first 28-day cycle. Safety was assessed by CTCAE v4.0. ORR and CBR were evaluated using RECIST v1.1., Results: Forty-three patients were treated in dose escalation from 3 to 96 mg/day, and 9 were treated in 64 mg dose expansion. After DLT occurred at 96 and 72 mg, 64 mg was established as the RP2D. Neutropenia was a common high-grade (19%) treatment-related adverse event at ≥ 64 mg. Half-life of CFI-400945 was 9 h, with C max achieved 2-4 h following dosing. One PR (45 cycles, ongoing) and two SD ≥ 6 months were observed (ORR = 2%; CBR = 6%)., Conclusions: CFI-400945 is well tolerated at 64 mg with dose-dependent neutropenia. Favourable pharmacokinetic profiles were achieved with daily dosing. Response rates were low without biomarker pre-selection. Disease-specific and combination studies are ongoing., Trial Registration: Clinical Trials Registration Number - NCT01954316 (Oct 1st, 2013).

The mitotic protein polo-like kinase 4 (PLK4) plays a critical role in centrosome duplication for cell division. By using immunofluorescence, we confirm that PLK4 is localized to centrosomes. In addition, we find that phospho-PLK4 (pPLK4) is cleaved and distributed to kinetochores (metaphase and anaphase), spindle midzone/cleavage furrow (anaphase and telophase), and midbody (cytokinesis) during cell division in immortalized epithelial cells as well as breast, ovarian, and colorectal cancer cells. The distribution of pPLK4 midzone/cleavage furrow and midbody positions pPLK4 to play a functional role in cytokinesis. Indeed, we found that inhibition of PLK4 kinase activity with a small-molecule inhibitor, CFI-400945, prevents translocation to the spindle midzone/cleavage furrow and prevents cellular abscission, leading to the generation of cells with polyploidy, increased numbers of duplicated centrosomes, and vulnerability to anaphase or mitotic catastrophe. The regulatory role of PLK4 in cytokinesis makes it a potential target for therapeutic intervention in appropriately selected cancers., Competing Interests: Conflict of interest statement: The intellectual properties of the PLK4 inhibitor CFI400945 is owned by the University Health Network (Toronto) and is in the process of being transferred to a company in which T.W.M. has equity. Observations for a role of polo-like kinase 4 on cytokinesis supported by CFI-400945 inhibition were disclosed to the University of Southern California Stevens Institute in March 2016. D.J.S. is an employee of BioMarin and has stock or other ownership in Pfizer. Research grants from the following companies have been provided to authors’ institutions: Cepheid (M.F.P.), Eli Lilly & Company (M.F.P.), Novartis Pharmaceuticals (M.F.P. and D.J.S.), F. Hoffmann La-Roche Ltd. (M.F.P. and J.S.B.), Pfizer (D.J.S.). M.F.P. holds a consulting or advisory role with honoraria with Karyopharm Therapeutics, Puma Biotechnology, Biocartis, Eli Lilly & Company, Novartis Pharmaceuticals, and F. Hoffmann–La Roche. D.J.S. holds a consulting or advisory role with honoraria with Eli Lilly & Company, and Novartis Pharmaceuticals. Pfizer, BioMarin, and Novartis also provided travel, accommodations, or expenses to D.J.S. S.D., Y.Z., R.G., G.P.N., N.O., and M.P. declare no potential conflict of interest., (Copyright © 2019 the Author(s). Published by PNAS.)

Purpose: Several clinical trials have investigated the prognostic and predictive usefulness of molecular markers. With limited predictive value, molecular markers have mainly been used to identify prognostic subgroups in which the indication for chemotherapy is doubtful and the prognosis is favorable enough for chemotherapy to be avoided. However, limited information is available about which groups of patients may benefit from additional therapy. This study aimed to describe the prognostic effects of Ki-67 in several common subgroups of patients with early breast cancer., Methods: This retrospective study analyzed a single-center cohort of 3140 patients with HER2-, hormone receptor-positive breast cancer. Five-year disease-free survival (DFS) rates were calculated for low (< 10%), intermediate (10-19%), and high (≥ 20%) Ki-67 expression levels, as assessed by immunohistochemistry, and for subgroups relative to age, body mass index, disease stage, tumor grade, and (neo-)adjuvant chemotherapy. It was also investigated whether Ki-67 had different effects on DFS in these subgroups., Results: The 5-year DFS rates for patients with low, intermediate, and high levels of Ki-67 expression were 0.90, 0.89, and 0.77, respectively. Ki-67 was able to further differentiate patients with an intermediate prognosis into different prognostic groups relative to common clinical parameters. Patients with stage II breast cancer had 5-year DFS rates of 0.84, 0.88, and 0.79 for low, intermediate, and high levels of Ki-67 expression. Ki-67 had different prognostic effects in subgroups defined by age and tumor grade., Conclusions: Ki-67 may help identify patients in intermediate prognostic groups with an unfavorable prognosis who may benefit from further therapy.

One of the most recent advances in the treatment of HER2 + breast cancer is the development of the antibody-drug conjugate, T-DM1. T-DM1 has proven clinical benefits for patients with advanced and/or metastatic breast cancer who have progressed on prior HER2-targeted therapies. However, T-DM1 resistance ultimately occurs and represents a major obstacle in the effective treatment of this disease. Because anti-apoptotic BCL-2 family proteins can affect the threshold for induction of apoptosis and thus limit the effectiveness of the chemotherapeutic payload, we examined whether inhibition of BCL-2/X L would enhance the efficacy of T-DM1 in five HER2-expressing patient-derived breast cancer xenograft models. Inhibition of BCL-2/X L via navitoclax/ABT-263 significantly enhanced the cytotoxicity of T-DM1 in two of three models derived from advanced and treatment-exposed metastatic breast tumors. No additive effects of combined treatment were observed in the third metastatic tumor model, which was highly sensitive to T-DM1, as well as a primary treatment-exposed tumor, which was refractory to T-DM1. A fifth model, derived from a treatment naïve primary breast tumor, was sensitive to T-DM1 but markedly benefited from combination treatment. Notably, both PDXs that were highly responsive to the combination therapy expressed low HER2 protein levels and lacked ERBB2 amplification, suggesting that BCL-2/X L inhibition can enhance sensitivity of tumors with low HER2 expression. Toxicities associated with combined treatments were significantly ameliorated with intermittent ABT-263 dosing. Taken together, these studies provide evidence that T-DM1 cytotoxicity could be significantly enhanced via BCL-2/X L blockade and support clinical investigation of this combination beyond ERBB2 -amplified and/or HER2-overexpressed tumors., (©2019 American Association for Cancer Research.)