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Background: Cyclin-dependent kinase 4/6 inhibitors (CDK4/6i) in combination with endocrine therapy (ET) provide significant clinical benefit in patients with HR+/HER2- metastatic breast cancer (MBC) and have become a standard of care treatment. Prior insights from tumor profiling and preclinical analyses suggest that AKT1 activation can induce CDK4/6i resistance. We hypothesized that targeting AKT1 following CDK4/6i progression may be an effective therapeutic strategy and conducted a clinical trial to evaluate both doublet (ET+AKTi) and triplet (ET+AKTIi+CDK 4/6i) therapy in the ≥ 2nd line MBC setting. Methods: TAKTIC is an open-label phase Ib clinical trial (clinicaltrials.gov NCT03959891) evaluating the combination of the AKT inhibitor ipatasertib (ipat) with fulvestrant (Arm A), an aromatase inhibitor (Arm B), or the triplet combination (Arm C) with fulvestrant + palbociclib (palbo). The primary objective is to evaluate the safety (NCI CTCAE 5.0) and tolerability of ipat in combination with endocrine therapy +/- CDK4/6i. Secondary objectives include clinical efficacy, as determined by objective response rate (RECIST v1.1), clinical benefit rate (CBR), progression-free survival (PFS), and overall survival (OS). Key inclusion criteria include unresectable HR+/HER2- MBC; at least 1 prior therapy for MBC including any CDK4/6i; up to 2 prior lines of chemotherapy for MBC (no limit on prior endocrine therapy). Here, we present an updated interim analysis from all study arms. Results: The trial completed accrual with 77 pts enrolled from June 2019 – February 2022, including 19 on Arm A, 16 on Arm B, and 42 on Arm C. Median age was 62 (range 32-88) and 65/77 pts (84%) received prior CDK4/6i (median no. of prior lines = 3, range 1-13). 56/77 pts (73%) had measurable disease at baseline and 50/77 pts (65%) had visceral metastases in the liver/lung (68% Arm A, 44% Arm B, 71% Arm C). Pts enrolled on Arms A and B received ipat at 400mg in combination with fulvestrant or an aromatase inhibitor, respectively. In Arm C, 27/42 pts enrolled into the dose escalation phase and received ipat + palbo at varying doses in combination with fulvestrant. Two DLTs were observed in the 300mg ipat + 125mg palbo cohort (grade 4 neutropenia ≥ 7 days). ET+400mg ipat + 100mg palbo was determined to be the recommended phase 2 dose (R2PD), and the remaining 15/42 pts on Arm C were treated at this dose level in the expansion phase. Treatment was well tolerated in all arms. Grade 3 and 4 toxicities included neutropenia (39/77, 50.6%), leukopenia (15/77, 19.5%), diarrhea (11/77, 14/3%), transaminitis (7/77, 9.1%), lymphopenia (6/77, 7.8%), rash (6/77, 7.8%), and thrombocytopenia (3/77, 3.9%). As of 6/28/2022, 16/77 pts remain on treatment. The median treatment duration for all pts is estimated at 6 months (range 0.5-39). Among the 56 pts with measurable disease, 11 had partial response (PR) and 32 had stable disease (SD) as the best response. CBR, defined as percentage of pts who achieved PR or SD > 6 months, was 48% across the study (53% Arm A, 31% Arm B, 57% Arm C). The median PFS was 5.5 months (95% confidence interval [CI]: 3.8 – 7.4) and the median OS was 24.5 months (95% CI: 17.1 – 33.9). Conclusions: The combination of ipat with endocrine therapy +/- palbo is well tolerated in heavily pre-treated pts, with preliminary evidence of clinical activity. This trial demonstrates how molecular insights related to CDK4/6i resistance inform potential therapy combinations. Further studies are needed to evaluate AKTi-based combinations in pts with HR+ MBC. Citation Format: Seth A. Wander, Jennifer C. Keenan, Andrzej Niemierko, Dejan Juric, Laura M. Spring, Jeffrey Supko, Neelima Vidula, Steven J. Isakoff, Lianne Ryan, Sarah Padden, Elizabeth Fisher, Amber Newton, Beverly Moy, Leif Ellisen, Douglas S. Micalizzi, Aditya Bardia. PD13-07 Combination therapy with the AKT inhibitor, ipatasertib, endocrine therapy, and a CDK4/6 inhibitor for hormone receptor positive (HR+)/HER2 negative metastatic breast cancer (MBC): results from the phase I TAKTIC trial [abstract]. In: Proceedings of the 2022 San Antonio Breast Cancer Symposium; 2022 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2023;83(5 Suppl):Abstract nr PD13-07.

Background: Clinical outcomes in breast cancer differ across racial and ethnic populations. We have previously demonstrated that receipt of genotype-matched therapy targeted to an actionable mutation may potentially improve patient outcomes (Vidula, CCR, 2021). We evaluated the impact of race on clinical, socioeconomic, and genomic characteristics, clinical trial participation, and receipt of genotype-matched therapy among patients with metastatic breast cancer (MBC). Methods: We conducted a retrospective study of patients with MBC at an academic institution who underwent cell-free DNA testing (cfDNA, Guardant360, 74 gene panel) as part of routine clinical care from 11/29/2016-11/2/2020. Patient demographics (including self-reported race and ethnicity) and clinical trial enrollment (at same institution) were determined by retrospective data collection. Mutations identified in cfDNA were characterized as actionable based on the variant interpretation performed by Guardant360 using vetted genomic databases, and receipt of genotype-matched therapy targeted to an actionable mutation was determined as previously described (Vidula, CCR, 2021). Pearson’s chi-squared and Wilcoxon rank-sum tests were used to compare categorical and continuous variables between groups, with p< 0.05 indicating statistical significance. Results: Four hundred and twenty-five patients with MBC and cfDNA results were identified, of which 369 were White (87%), 27 Black (6.4%), 15 Hispanic (3.5%), and 14 Asian (3.3%). There were no significant differences in median age at MBC diagnosis (p=0.064), disease subtype distribution (p=0.74), proportions of de-novo/recurrent MBC (p=0.95), presence of visceral metastases (p=0.84), Charleston comorbidity index (p=0.93), menopausal status (p=0.3), and level of education (p=0.44) across racial groups. Higher proportions of non-primary English speakers were seen in Hispanic (80%) and Asian (29%) races (p< 0.001). Median distance traveled to the institution also varied based on race, with White patients traveling further (White: 39.1 miles, Black: 21.8 miles, Hispanic 9.4 miles, Asian 9.1 miles, p< 0.001). In addition, type of insurance varied based on race, with White patients having the highest rates of commercial insurance and Medicare, Black patients having the highest rate of state-supported insurance, and Asian patients having the highest uninsured rates (p< 0.001). Clinical trial enrollment rates did not significantly differ by race (White: 44%, Black: 37%, Hispanic: 47%, and Asian 21%, p=0.34), but patients without insurance were significantly less likely to be enrolled on a trial than those with commercial insurance (p=0.03). The proportion of patients with ≥1 actionable mutation in cfDNA did not vary significantly by race (White: 78%, Black: 56%, Hispanic: 73%, Asian 86%, p=0.18) and the median number of actionable mutations found in cfDNA was similar across races (p=0.31). However, receipt of genotype-matched therapy targeted to an actionable mutation varied by race, with the highest rates of matched therapy in White patients (White: 28%, Black: 11%, Hispanic 13%, Asian 14%, p< 0.001). After multivariable logistic regression adjusting for subtype, commercial insurance versus other insurance types, and proximity to the center, White patients remained significantly more likely to receive matched therapy (p=0.029). Conclusions: We observed significant race-based differences in non-English speaking status, insurance type, and median distance traveled to the institution. Racial/ethnic minority patients were less likely to receive genotype-matched therapy than White patients. Further research is needed to identify barriers and reduce disparities in access to precision medicine. Citation Format: Rupali Sood, Lianne Ryan, Andrzej Niemierko, Laura M. Spring, Dejan Juric, Steven J. Isakoff, Seth A. Wander, Jennifer Shin, Naomi Ko, Leif Ellisen, Beverly Moy, Aditya Bardia, Neelima Vidula. Impact of Race on Clinical, Socioeconomic, and Genomic Characteristics, Clinical Trial Participation, and Receipt of Genotype-matched Therapy Among Patients with Metastatic Breast Cancer [abstract]. In: Proceedings of the 2022 San Antonio Breast Cancer Symposium; 2022 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2023;83(5 Suppl):Abstract nr PD1-10.

Background: CDK4/6 inhibitors (CDK4/6i) paired with endocrine therapy (ET) are considered first-line (1L) therapy for patients (pts) with HR+ HER2- advanced breast cancer (aBC). A minority of pts will demonstrate primary resistance to CDK4/6i, as characterized by early progression. Thymidine kinase 1 (TK1) is a cell-cycle regulated enzyme downstream of CDK4/6 and involved in nucleotide metabolism during DNA synthesis. Prior studies have shown TK1 may serve as a biomarker of response to CDK4/6i, with early TK1 activity (TK1a) suppression after initiation of CDK 4/6i therapy associated with improved PFS. Lack of TK1a suppression may be associated with primary resistance to CDK4/6i. In this study, we aim to analyze response to subsequent lines of therapy and overall survival (OS) of pts with early progression on 1L CDK4/6i. Methods: Pts with HR+ HER2- aBC from a phase II trial of an alternative schedule of palbociclib (palbo alt dosing trial NCT 3007979) and from a retrospective palbociclib study were included in this analysis. Pts in the palbo alt dosing trial underwent baseline and C1D15 TK1a analysis after initiation on CDK4/6i. C1D15 TK1a suppression was defined at TK1a < 30 Du/L. Pts in the retrospective palbociclib study included pts receiving palbo as part of their standard of care 1L therapy for HR+ HER2- aBC at Washington University in Saint Louis from 2016 to 2021. Clinical information, including treatment start and stop dates on each of the next-line therapies, were collected from the electronic medical record. PFS was estimated by the treatment duration on a specified treatment regimen. Early progression on CDK4/6i was defined as PFS < 6 mo. Best response was defined as next line of therapy with the numerically longest PFS. OS was defined as time to death from the initiation of CDK4/6i. Results: Of the 54 pts enrolled on the palbo alt dosing trial, 51 pts were evaluable for clinical benefit and 46 pts were evaluable for TK1a suppression rate at C1D15. 7 pts (15.2%) were found without TK1a suppression at C1D15. This lack of TK1a suppression on palbo was associated with a significantly shorter PFS (median PFS=3.1 mo) compared to not reached in pts with TK1a suppression at C1D15. We conducted clinical analysis on N=26 pts who exhibited early progression on CDK4/6i which included 10 pts from the palbo alt dosing trial and 16 from the retrospective study. The average subsequent line of therapies in this cohort was 3, with the most common second line (2L) therapy being chemotherapy (N=17, 65.4%) and ET (N=8, 30.8%). The median PFS for pts receiving 2L chemotherapy and ET was 4.09 mo and 3.64 mo, respectively. 10 pts received both chemotherapy and ET with 7 (70.0%) achieving best response with chemotherapy compared to 3 pts (30.0%) who achieved best response with ET. The median OS for the cohort was 14.6 mo. Conclusions: Early progression on CDK4/6i is associated with a particularly poor prognosis. In our cohort, the median OS was far below the expected median OS for pts receiving 1L palbo as reported in the PALOMA-2 trial (14.6 mo vs 53.9 mo). Early progression on CDK4/6i is associated with more aggressive disease which may respond more favorably to chemotherapy, as demonstrated by best response to therapy. Further prospective studies are warranted to explore this treatment approach. Citation Format: Katherine K. Clifton, Shana N. Thomas, Jingqin Luo, Jing Xi, Nusayba A. Bagegni, Foluso O. Ademuyiwa, Rama Suresh, Ashley Frith, Andrew A. Davis, Ron Bose, Katherine Weilbaecher, Whitney L. Hensing, Timothy Pluard, Massimo Cristofanilli, Hyo S. Han, Adam M. Brufsky, Kevin Kalinsky, Shom Goel, Seth A. Wander, Lindsay L. Peterson, Cynthia Ma. PD13-09 Clinical outcomes of patients with HR+ HER2- advanced breast cancer with early progression on CDK4/6 inhibitors [abstract]. In: Proceedings of the 2022 San Antonio Breast Cancer Symposium; 2022 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2023;83(5 Suppl):Abstract nr PD13-09.

Background: CDK4/6 inhibitors (CDK4/6i) are standard of care for the management of HR+/HER2- metastatic breast cancer (MBC). Genomic alterations that drive resistance to CDK4/6i are diverse, and while the molecular landscape is heterogeneous, several mechanisms of CDK4/6i resistance converge on the RAS/MAPK and PI3K/AKT/mTOR signaling pathways. NF1 downregulates RAS and dampens cellular proliferation. Laboratory-based models demonstrate that loss of NF1 is associated with resistance to endocrine therapy (ET), and emergence of NF1 mutations (NF1m) are correlated with progressive disease (PD) in circulating tumor DNA (ctDNA). While NF1m may diminish CDK4/6i susceptibility, a clear relationship has not been elucidated. The primary objective of this study was to characterize patient (pt) response to CDK4/6i in NF1m HR+/HER2- MBC. Methods: We identified 47 pts with NF1m via a database with one or more ctDNA samples sequenced at variable time-points as part of routine care for MBC. NF1m were categorized as pathogenic (p)NF1m or variants of uncertain significance (VUS) based on their associated Guardant report. We identified 27 pts with HR+/HER2- MBC and NF1m that received at least 1 line of CDK4/6i in the metastatic setting. Intrinsic resistance was defined as PD < 6 months on a CDK4/6i regimen, and acquired resistance was defined as PD >6 months. Pts with intrinsic resistance or acquired resistance and NF1m detected post-PD were categorized as having a resistance phenotype potentially driven by NF1m. Pts with NF1m detected prior to therapy and >6 months clinical response on a CDK4/6i were categorized as having NF1m tumors sensitive to CDK4/6i. Results: The NF1m cohort (n = 27) had 9 pts with pNF1m, while 18 pts expressed VUS. The median age at MBC diagnosis was 54 years, and 67% had visceral metastasis at ctDNA collection. Pts received a median of 1 prior line (range: 0 - 6) of ET or chemotherapy in the metastatic setting before CDK4/6i. Amongst pts with pathogenic variants (n = 9), we found 3 pts with pNF1m were intrinsically resistant to CDK4/6i. Acquired resistance was seen in 1 pt with pNF1m detected post-PD, and 2 pts had evidence of both acquired and subsequent intrinsic resistance to a later line of CDK4/6i. Overall, 67% (6/9) of pNF1m pts demonstrated a CDK4/6i resistance phenotype; mutant allele fraction (AF) ranged from 0.2% - 29.9%, and the mean maximum allele fraction (MAF) was 6.0%. Pre- and post-treatment samples were available on 3 pts with pNF1m, and 1 of these pts had an AF rise from 2.7% to 12.3% when comparing ctDNA pre- and post-CDK4/6i. ctDNA from 4 of 6 resistant tumors harbored other putative drivers including alterations in FGFR, KRAS, PTEN, and RB. We identified 2 counter-examples of pNF1m tumors sensitive to CDK4/6i. These pts expressed relatively low NF1m AF, ranging 0.1% - 0.5% with a mean MAF 0.3%. Another pNF1m pt had intrinsic resistance to initial CDK4/6i but was sensitive to later-line CDK4/6i. In the subgroup of pts with VUS-NF1m (n = 18), a more mixed picture of resistance and sensitivity was seen. 8 pts had intrinsic or acquired resistance, 8 pts had NF1m tumors sensitive to CDK4/6i, and 1 pt had evidence of both; 61% (n = 11) of pts expressed alterations in other resistance mediating genes. 1 pt stopped therapy due to toxicity rather than PD. Conclusions: Our work demonstrates that tumor expression of pNF1m may be associated with CDK4/6i resistance in pts with HR+/HER2- MBC, and allele fraction could be predictive of drug susceptibility. Tumors harboring VUS had varied sensitivity, suggesting that some of these mutations may not be pathogenic, and counter-examples of pNF1m MBC benefiting from CDK4/6i plus ET highlight the complexities in predicting drug response based on single gene alteration. Future effort is warranted to explore the potential impact of NF1 on CDK4/6i resistance, as well as the potential role for therapies targeting the MAPK pathway in this patient population. Citation Format: Maxwell R. Lloyd, Lianne Ryan, Arielle J. Medford, Jennifer C. Keenan, Laura M. Spring, Neelima Vidula, Beverly Moy, Dejan Juric, Leif Ellisen, Aditya Bardia, Seth A. Wander. Investigating NF1 Mutations in Circulating Tumor DNA of Patients with Hormone-receptor Positive (HR+) Breast Tumors Resistant to CDK4/6 Inhibition (CDK4/6i): A Retrospective Clinical Analysis [abstract]. In: Proceedings of the 2022 San Antonio Breast Cancer Symposium; 2022 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2023;83(5 Suppl):Abstract nr P1-13-07.

Background: Addition of PI3K/mTOR inhibitor after progression on CDK4/6 inhibitor (CDK4/6i) and endocrine therapy (ET) can potentially restore sensitivity to CDK4/6i and prevent adaptive activation of the PI3K/mTOR pathway. To evaluate this hypothesis, we conducted a Phase Ib study of gedatolisib (G), a dual inhibitor of PI3K/mTOR, palbociclib (P) a CDK4/6i, and ET (with letrozole [LET] or fulvestrant [FUL]) in women with hormone receptor positive (HR+)/HER2- advanced breast cancer (ABC). Manageable toxicity and preliminary antitumor activity were observed in 35 patients(pts) enrolled in the dose escalation portion of the study (Forero-Torres, ASCO 2018) and 103 pts enrolled in the expansion portion of the study (Layman, SABCS 2021). Here, we report updated efficacy and safety data and sub-group analysis by PIK3CA mutation status in the four expansion study arms with a March 3, 2022, data cut-off. Methods: Pts with HR+/HER2- ABC were treated in four expansion arms: A) G+P+LET as first-line treatment, B) G+P+FUL as 2nd line treatment in pts without prior CDK4/6i; C & D) G+P+FUL as 2nd or 3rd line therapy in pts with prior CDK4/6i. P, LET, and FUL were administered at standard doses. G 180 mg was intravenously administered weekly in Arms A, B, and C and three weeks on/one week off in Arm D. The primary endpoint was investigator assessed objective response rate (ORR). Secondary endpoints included safety, duration of response and progression free survival (PFS). Results: Of the 103 pts treated with G+P+ ET in the expansion arms (A-D), 100% had measurable disease at baseline, 71% (73/103) lacked PIK3CA mutations (wild type; WT), 27% (28/103) had PIK3CA-mutations (MT), 70% (72/103) had evidence of bone metastases, and 59% (61/103) had liver metastases. The most frequent grade 3 and 4 treatment related AEs (TRAE) with G+P+ET included neutropenia (63%), stomatitis (27%), rash (20%), fatigue (11%) and hyperglycemia (7%). Treatment discontinuation due to TRAEs was 6.5% in Arm A, 15.4% in Arm B, 9.4% in Arm C and 3.7% in Arm D. Efficacy data for each arm is presented in Table 1. Promising ORR and PFS were seen in all arms regardless of PIK3CA mutation status. In Arm D, ORR was 63% overall, 73% in PIK3CA-MT pts, and 60% in PIK3CA-WT pts. Median PFS in Arm D was 12.9 months with a median follow up of 29 months. 60% and 48% of pts in the PIK3CA-MT and PIK3CA-WT Arm D sub-groups, respectively, were progression free at 12 months. Conclusions: These preliminary data demonstrate promising activity of G+P+ET combination in pts who were CDK4/6i-naïve and in those whose disease progressed on or after CDK4/6i therapy regardless of PIK3CA mutation status. Encouraging results in CDK4/6i treatment naïve patients warrant further evaluation of gedatolisib in combination with CDK4/6i treatment in the front-line setting. Arm D results provide a strong basis for the initiated Phase 3 study (VIKTORIA-1) in pts whose disease progressed on or after CDK4/6i therapy. Table 1. Efficacy Data by Expansion Arms Citation Format: Robert Wesolowski, Hope Rugo, Erica Stringer-Reasor, Hyo S. Han, Jennifer M. Specht, E. Claire Dees, Peter Kabos, Ulka Vaishampayan, Seth A. Wander, Janice Lu, Keerthi Gogineni, Alexander I. Spira, Anne F. Schott, Maysa Abu-Khalaf, Pratima Nayak, Brian F. Sullivan, Igor Gorbatchevsky, AND Rachel M. Layman. PD13-05 Updated results of a Phase 1b study of gedatolisib plus palbociclib and endocrine therapy in women with hormone receptor positive advanced breast cancer: Subgroup analysis by PIK3CA mutation status [abstract]. In: Proceedings of the 2022 San Antonio Breast Cancer Symposium; 2022 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2023;83(5 Suppl):Abstract nr PD13-05.

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.

PURPOSE As the continuation beyond progression (BP) of cyclin-dependent kinase 4/6 inhibitors (CDK4/6i) is becoming increasingly attractive for the treatment of patients with hormone receptor (HR)–positive, human epidermal growth factor receptor 2 (HER2)–negative metastatic breast cancer (MBC), the definition of resistance factors is crucial. The aim of the study was to investigate the impact of CDK 4/6i BP and to explore potential genomic stratification factors. MATERIALS AND METHODS We retrospectively analyzed a multi-institutional cohort of patients with HR-positive HER2-negative MBC characterized for circulating tumor DNA through next-generation sequencing before treatment start. Differences across subgroups were analyzed by chi-square test, and survival was tested by univariable and multivariable Cox regression. Further correction was applied by propensity score matching. RESULTS Among the 214 patients previously exposed to CDK4/6i, 172 were treated with non–CDK4/6i-based treatment (non-CDK) and 42 with CDK4/6i BP. Multivariable analysis showed a significant impact of CDK4/6i BP, TP53 single-nucleotide variants, liver involvement, and treatment line on both progression-free survival (PFS) and overall survival (OS). Propensity score matching confirmed the prognostic role of CDK4/6i BP both for PFS and OS. The favorable impact of CDK4/6i BP was consistent across all subgroups, and a differential benefit was suggested for ESR1-mutated patients. ESR1 and RB1 mutations were more represented in the CDK4/6i BP subgroup with respect to CDK4/6i upfront. CONCLUSION The study highlighted a significant prognostic impact of the CDK4/6i BP strategy with a potential added benefit in patients with ESR1 mutations suggesting the need for an extensive biomarker characterization.

PURPOSE For patients with hormone receptor–positive (HR+), human epidermal growth factor receptor 2–negative (HER2–) metastatic breast cancer (MBC), first-line treatment is endocrine therapy (ET) plus cyclin-dependent kinase 4/6 inhibition (CDK4/6i). After disease progression, which often comes with ESR1 resistance mutations (ESR1-MUT), which therapies to use next and for which patients are open questions. An active area of exploration is treatment with further CDK4/6i, particularly abemaciclib, which has distinct pharmacokinetic and pharmacodynamic properties compared with the other approved CDK4/6 inhibitors, palbociclib and ribociclib. We investigated a gene panel to prognosticate abemaciclib susceptibility in patients with ESR1-MUT MBC after palbociclib progression. METHODS We examined a multicenter retrospective cohort of patients with ESR1-MUT MBC who received abemaciclib after disease progression on ET plus palbociclib. We generated a panel of CDK4/6i resistance genes and compared abemaciclib progression-free survival (PFS) in patients without versus with mutations in this panel (CDKi-R[–] v CDKi-R[+]). We studied how ESR1-MUT and CDKi-R mutations affect abemaciclib sensitivity of immortalized breast cancer cells and patient-derived circulating tumor cell lines in culture. RESULTS In ESR1-MUT MBC with disease progression on ET plus palbociclib, the median PFS was 7.0 months for CDKi-R(–) (n = 17) versus 3.5 months for CDKi-R(+) (n = 11), with a hazard ratio of 2.8 ( P = .03). In vitro, CDKi-R alterations but not ESR1-MUT induced abemaciclib resistance in immortalized breast cancer cells and were associated with resistance in circulating tumor cells. CONCLUSION For ESR1-MUT MBC with resistance to ET and palbociclib, PFS on abemaciclib is longer for patients with CDKi-R(–) than CDKi-R(+). Although a small and retrospective data set, this is the first demonstration of a genomic panel associated with abemaciclib sensitivity in the postpalbociclib setting. Future directions include testing and improving this panel in additional data sets, to guide therapy selection for patients with HR+/HER2– MBC.

Figure S9

Mechanisms driving resistance to cyclin-dependent kinase 4/6 inhibitors (CDK4/6i) in hormone receptor–positive (HR+) breast cancer have not been clearly defined. Whole-exome sequencing of 59 tumors with CDK4/6i exposure revealed multiple candidate resistance mechanisms including RB1 loss, activating alterations in AKT1, RAS, AURKA, CCNE2, ERBB2, and FGFR2, and loss of estrogen receptor expression. In vitro experiments confirmed that these alterations conferred CDK4/6i resistance. Cancer cells cultured to resistance with CDK4/6i also acquired RB1, KRAS, AURKA, or CCNE2 alterations, which conferred sensitivity to AURKA, ERK, or CHEK1 inhibition. Three of these activating alterations—in AKT1, RAS, and AURKA—have not, to our knowledge, been previously demonstrated as mechanisms of resistance to CDK4/6i in breast cancer preclinically or in patient samples. Together, these eight mechanisms were present in 66% of resistant tumors profiled and may define therapeutic opportunities in patients.Significance:We identified eight distinct mechanisms of resistance to CDK4/6i present in 66% of resistant tumors profiled. Most of these have a therapeutic strategy to overcome or prevent resistance in these tumors. Taken together, these findings have critical implications related to the potential utility of precision-based approaches to overcome resistance in many patients with HR+ metastatic breast cancer.This article is highlighted in the In This Issue feature, p. 1079

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Supplementary material

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The CDK4/6 inhibitor abemaciclib is a mainstay of treatment for hormone receptor-positive breast cancer. However, increased venous thromboembolism (VTE) rates in multiple clinical trials resulted in a black-box warning for this agent. Thrombosis rates in unselected real-world populations receiving abemaciclib remain ill defined.A multicenter observational cohort study was conducted of patients with metastatic breast cancer receiving abemaciclib. The primary end point was thrombosis during treatment or within 30 days of discontinuation. Multivariable logistic models assessed predictors of VTE, and a multivariable Cox proportional hazards model assessed mortality.A total of 364 patients were included, with a median treatment duration of 5.5 months. Twenty-six patients developed 27 (7.4%) thrombotic events (17 VTE, nine arterial thrombosis, and one with both events). No baseline characteristics were associated with increased VTE risk in multivariable modeling. Patients developing VTE during therapy had a higher risk of death than those who did not (hazard ratio, 2.09; 95% CI, 1.07-4.13). Median survival in patients who developed VTE compared with those who did not was 9.6 vs 25.8 months, respectively. The rate of VTE and any thrombosis during abemaciclib therapy was 9.1 and 13.7 events per 100 person-years, respectively, which is notably higher than rates observed in clinical trials.In a real-world setting, abemaciclib was associated with a VTE rate approximately two-fold greater than the already elevated rates reported in the MONARCH trials. Patients developing thrombosis on abemaciclib had a significantly higher risk of death. Given these findings, studies evaluating the role of thromboprophylaxis in patients receiving abemaciclib are needed.

RNA-seq experiment conditions and QC stats

Gene sets enrichment and associations

Evolutionary status data

Supplemental Table S3. Actionable mutations found in cfDNA and tumor tissue genotyping in 12 patients who went on to receive matched therapy after having both forms of testing.

Foundation Medicine cohort

Purpose:Actionable mutations can guide genotype-directed matched therapy. We evaluated the utility of tissue-based and plasma-based genotyping for the identification of actionable mutations and selection of matched therapy in patients with metastatic breast cancer (MBC).Experimental Design:Patients with MBC who underwent tissue genotyping (institutional platform, 91-gene assay) or plasma-based cell-free DNA (cfDNA, Guardant360, 73-gene assay) between January 2016 and December 2017 were included. A chart review of records to identify subtype, demographics, treatment, outcomes, and tissue genotyping or cfDNA results was performed. The incidence of actionable mutations and the selection of matched therapy in tissue genotyping or cfDNA cohorts was determined. The impact of matched therapy status on overall survival (OS) in tissue genotyping or cfDNA subgroups was determined with Cox regression analysis.Results:Of 252 patients who underwent cfDNA testing, 232 (92%) had detectable mutations, 196 (78%) had actionable mutations, and 86 (34%) received matched therapy. Of 118 patients who underwent tissue genotyping, 90 (76%) had detectable mutations, 59 (50%) had actionable mutations, and 13 (11%) received matched therapy. For cfDNA patients with actionable mutations, matched versus nonmatched therapy was associated with better OS [HR 0.41, 95% confidence interval (CI): 0.23–0.73, P = 0.002], and this remained significant in a multivariable analysis correcting for age, subtype, visceral metastases, and brain metastases (HR = 0.46, 95% CI: 0.26–0.83, P = 0.010).Conclusions:Plasma-based genotyping identified high rates of actionable mutations, which was associated with significant application of matched therapy and better OS in patients with MBC.See related commentary by Rugo and Huppert, p. 3275

Purpose:While FGFR1 amplification has been described in breast cancer, the optimal treatment approach for FGFR1-amplified (FGFR1+) metastatic breast cancer (MBC) remains undefined.Experimental Design: We evaluated clinical response to endocrine and targeted therapies in a cohort of patients with hormone receptor–positive (HR+)/HER2− MBC and validated the functional role of FGFR1-amplification in mediating response/resistance to hormone therapy in vitro.Results:In the clinical cohort (N = 110), we identified that patients with FGFR1+ tumors were more likely to have progesterone receptor (PR)-negative disease (47% vs. 20%; P = 0.005), coexisting TP53 mutations (41% vs. 21%; P = 0.05), and exhibited shorter time to progression with endocrine therapy alone and in combination with CDK4/6 inhibitor, but not with a mTOR inhibitor (everolimus), adjusting for key prognostic variables in multivariate analysis. Furthermore, mTOR-based therapy resulted in a sustained radiological and molecular response in an index case of FGFR1+ HR+/HER2− MBC. In preclinical models, estrogen receptor–positive (ER+)/FGFR1-amplified CAMA1 human breast cancer cells were only partially sensitive to fulvestrant, palbociclib, and alpelisib, but highly sensitive to everolimus. In addition, transduction of an FGFR1 expression vector into ER+ T47D cells induced resistance to fulvestrant that could be overcome by added TORC1 inhibition, but not PI3K or CDK4/6 inhibition.Conclusions:Collectively, these findings suggest that while FGFR1 amplification confers broad resistance to ER, PI3K, and CDK4/6 inhibitors, mTOR inhibitors might have a unique therapeutic role in the treatment of patients with ER+/FGFR1+ MBC.

Supplementary figures

GSEA list

FGFR alterations in different cohorts

Supplementary Data from Acquired FGFR and FGF Alterations Confer Resistance to Estrogen Receptor (ER) Targeted Therapy in ER+ Metastatic Breast Cancer

Supplemental Figure S1. Consort diagram of patients included in this study.

clinicopathologic info

Complete exome data

Biopsy purity and ploidy