Currently, no targeted therapies are available for metastatic triplenegative breast cancer (mTNBC). We evaluated the safety, efficacy, and biomarkers of response to cabozantinib, a multikinase inhibitor, in patients with mTNBC. We conducted a single arm phase II and biomarker study that enrolled patients with measurable mTNBC. Patients received cabozantinib (60 mg daily) on a 3-week cycle and were restaged after 6 weeks and then every 9 weeks. The primary endpoint was objective response rate. Predefined secondary endpoints included progression-free survival (PFS), toxicity, and tissue and blood circulating cell and protein biomarkers. Of 35 patients who initiated protocol therapy, 3 (9% [95% confidence interval (CI): 2, 26]) achieved a partial response (PR). Nine patients achieved stable disease (SD) for at least 15 weeks, and thus the clinical benefit rate (PR+SD) was 34% [95% CI: 19, 52]. Median PFS was 2.0 months [95% CI: 1.3, 3.3]. The most common toxicities were fatigue, diarrhea, mucositis, and palmar-plantar erythrodysesthesia. There were no grade 4 toxicities, but 12 patients (34%) required dose reduction. Two patients had TNBCs with MET amplification. During cabozantinib therapy, there were significant and durable increases in plasma placental growth factor, vascular endothelial growth factor (VEGF), VEGF-D, stromal cell-derived factor 1a, and carbonic anhydrase IX, and circulating CD3 + cells and CD8 + T lymphocytes, and decreases in plasma soluble VEGF receptor 2 and CD14+ monocytes (all p < .05). Higher baseline concentrations of soluble MET (sMET) associated with longer PFS (p = .03). In conclusion, cabozantinib showed encouraging safety and efficacy signals but did not meet the primary endpoint in pretreated mTNBC. Exploratory analyses of circulating biomarkers showed that cabozantinib induces systemic changes consistent with activation of the immune system and antiangiogenic activity, and that sMET should be further evaluated a potential biomarker of response.

Cancer cell survival is dependent on oxidative-stress defenses against reactive oxygen species (ROS) that accumulate during tumorigenesis. Here, we show a non-canonical oxidative-stress defense mechanism through TRPA1, a neuronal redox-sensing Ca2+-influx channel. In TRPA1-enriched breast and lung cancer spheroids, TRPA1 is critical for survival of inner cells that exhibit ROS accumulation. Moreover, TRPA1 promotes resistance to ROS-producing chemotherapies, and TRPA1 inhibition suppresses xenograft tumor growth and enhances chemosensitivity. TRPA1 does not affect redox status but upregulates Ca2+-dependent anti-apoptotic pathways. NRF2, an oxidant-defense transcription factor, directly controls TRPA1 expression, thus providing an orthogonal mechanism for protection against oxidative stress together with canonical ROS-neutralizing mechanisms. These findings reveal an oxidative-stress defense program involving TRPA1 that could be exploited for targeted cancer therapies.

Previous studies of fatty acids and breast cancer risk have shown mixed results, which may be due in part to tumor heterogeneity. Prior research has also illustrated an important role of specific fatty acids in immune regulation, T cell function, and inflammation, indicating that the effects of specific fatty acids on breast cancer risk may vary by tumor expression of immuno-inflammatory markers. We therefore aimed to evaluate the relationships between prediagnostic erythrocyte membrane fatty acids and breast cancer risk by tumor tissue expression of immuno-inflammatory markers (CD4, CD8, CD20, CD163, COX-2) and fatty acid synthase (FAS).

Whole-exome sequencing of cell-free DNA (cfDNA) could enable comprehensive profiling of tumors from blood but the genome-wide concordance between cfDNA and tumor biopsies is uncertain. Here we report ichorCNA, software that quantifies tumor content in cfDNA from 0.1× coverage whole-genome sequencing data without prior knowledge of tumor mutations. We apply ichorCNA to 1439 blood samples from 520 patients with metastatic prostate or breast cancers. In the earliest tested sample for each patient, 34% of patients have ≥10% tumor-derived cfDNA, sufficient for standard coverage whole-exome sequencing. Using whole-exome sequencing, we validate the concordance of clonal somatic mutations (88%), copy number alterations (80%), mutational signatures, and neoantigens between cfDNA and matched tumor biopsies from 41 patients with ≥10% cfDNA tumor content. In summary, we provide methods to identify patients eligible for comprehensive cfDNA profiling, revealing its applicability to many patients, and demonstrate high concordance of cfDNA and metastatic tumor whole-exome sequencing.

In the BrighTNess trial, carboplatin added to neoadjuvant chemotherapy (NAC) was associated with increased pathologic complete response (pCR) rates in patients with stage II/III triple-negative breast cancer (TNBC). In this matched cohort study, cases with a germline BRCA1/2 mutation (gBRCA; n = 75) were matched 1:2 with non-gBRCA controls (n = 150) by treatment arm, lymph node status, and age to evaluate pCR rates and association of benefit from platinum/PARP inhibitors with validated RNA expression-based immune, proliferation, and genomic instability scores among gBRCA with the addition of carboplatin ± veliparib to NAC. Among the well-matched cohorts, odds of pCR were not higher in gBRCA cancers who received standard NAC with carboplatin (OR 0.24, 95% CI [0.04-1.24], p = 0.09) or with carboplatin/veliparib (OR 0.44, 95% CI [0.10-1.84], p = 0.26) compared to non-gBRCA cancers. Higher PAM50 proliferation, GeparSixto immune, and CIN70 genomic instability scores were each associated with higher pCR rate in the overall cohort, but not specifically in gBRCA cases. In this study, gBRCA carriers did not have higher odds of pCR than non-gBRCA controls when carboplatin ± veliparib was added to NAC, and showed no significant differences in molecular, immune, chromosomal instability, or proliferation gene expression metrics.

Approximately 50% of patients with metastatic HER2-positive (HER2+) breast cancer develop brain metastases (BCBMs). We report that the tumor suppressor p16INK4A is deficient in the majority of HER2+ BCBMs. p16INK4A-deficiency as measured by protein immunohistochemistry predicted response to combined tucatinib and abemaciclib in orthotopic patient-derived xenografts (PDXs) of HER2 + BCBMs. Our findings establish the rationale for a biomarker-driven clinical trial of combined CDK4/6- and HER2-targeted agents for patients with HER2 + BCBM.

Proliferation is a fundamental trait of cancer cells, but its properties and spatial organization in tumours are poorly characterized. Here we use highly multiplexed tissue imaging to perform single-cell quantification of cell cycle regulators and then develop robust, multivariate, proliferation metrics. Across diverse cancers, proliferative architecture is organized at two spatial scales: large domains, and smaller niches enriched for specific immune lineages. Some tumour cells express cell cycle regulators in the (canonical) patterns expected of freely growing cells, a phenomenon we refer to as 'cell cycle coherence'. By contrast, the cell cycles of other tumour cell populations are skewed towards specific phases or exhibit non-canonical (incoherent) marker combinations. Coherence varies across space, with changes in oncogene activity and therapeutic intervention, and is associated with aggressive tumour behaviour. Thus, multivariate measures from high-plex tissue images capture clinically significant features of cancer proliferation, a fundamental step in enabling more precise use of anti-cancer therapies.

Clonal hematopoiesis of indeterminate potential (CHIP), an emerging biomarker for personalized risk-directed interventions, is increased in cancer survivors. However, little is known about patient preferences for CHIP testing. We surveyed participants in a prospective cohort study of young women with breast cancer (BC). The emailed survey included an introduction to CHIP and a vignette eliciting participants' preferences for CHIP testing, considering sequentially: population-based 10-year risk of BC recurrence, hematologic malignancy, and heart disease; increased CHIP-associated risks; current CHIP management; dedicated CHIP clinic; and hypothetical CHIP treatment. Preference changes were evaluated using the McNemar test. The survey response rate was 82.2% (528/642). Median age at time of survey was 46 years and median time from diagnosis was 108 months. Only 5.9% had prior knowledge of CHIP. After vignette presentation, most survivors (87.1%) recommended CHIP testing for the vignette patient. Presented next with CHIP-independent, population-based risks, 11.1% shifted their preference from testing to not testing. After receiving information about CHIP-associated risks, an additional 10.1% shifted their preference to testing. Preference for testing increased if vignette patient was offered a CHIP clinic or hypothetical CHIP treatment, with 7.2% and 14.1% switching preferences toward testing, respectively. Finally, 75.8% of participants desired CHIP testing for themselves. Among participants, 28.2% reported that learning about CHIP caused at least moderate anxiety. Most young survivors favored CHIP testing, with preferences influenced by risk presentation and potential management strategies. Our findings highlight the importance of risk communication and psychosocial support when considering biomarkers for future risk in cancer survivors. This trial has been registered at www.clinicaltrials.gov as #NCT01468246.

The identification of prognostic markers in patients receiving neoadjuvant therapy is crucial for treatment optimization in HER2-positive breast cancer, with the immune microenvironment being a key factor. Here, we investigate the complexity of B and T cell receptor (BCR and TCR) repertoires in the context of two phase III trials, NeoALTTO and CALGB 40601, evaluating neoadjuvant paclitaxel with trastuzumab and/or lapatinib in women with HER2-positive breast cancer. BCR features, particularly the number of reads and clones, evenness and Gini index, are heterogeneous according to hormone receptor status and PAM50 subtypes. Moreover, BCR measures describing clonal expansion, namely evenness and Gini index, are independent prognostic factors. We present a model developed in NeoALTTO and validated in CALGB 40601 that can predict event-free survival (EFS) by integrating hormone receptor and clinical nodal status, breast pathological complete response (pCR), stromal tumor-infiltrating lymphocyte levels (%) and BCR repertoire evenness. A prognostic score derived from the model and including those variables, HER2-EveNT, allows the identification of patients with 5-year EFS > 90%, and, in those not achieving pCR, of a subgroup of immune-enriched tumors with an excellent outcome despite residual disease.

The molecular underpinnings of HER2-low and HER2-0 (IHC 0) breast tumors remain poorly defined. Using genomic findings from 1039 patients with HER2-negative metastatic breast cancer undergoing next-generation sequencing from 7/2013-12/2020, we compare results between HER2-low (n = 487, 47%) and HER2-0 tumors (n = 552, 53%). A significantly higher number of ERBB2 alleles (median copy count: 2.05) are observed among HER2-low tumors compared to HER2-0 (median copy count: 1.79; P = 2.36e-6), with HER2-0 tumors harboring a higher rate of ERBB2 hemideletions (31.1% vs. 14.5%). No other genomic alteration reaches significance after accounting for multiple hypothesis testing, and no significant differences in tumor mutational burden are observed between HER2-low and HER2-0 tumors (median: 7.26 mutations/megabase vs. 7.60 mutations/megabase, p = 0.24). Here, we show that the genomic landscape of HER2-low and HER2-0 tumors does not differ significantly, apart from a higher ERBB2 copy count among HER2-low tumors, and a higher rate of ERBB2 hemideletions in HER2-0 tumors.

A large collaborative analysis of data from 47 epidemiological studies concluded that longer duration of breastfeeding reduces the risk of developing breast cancer. Despite the strong epidemiological evidence, the molecular mechanisms linking prolonged breastfeeding to decreased risk of breast cancer remain poorly understood.

MET expression and activation appear to be important for initiation and progression of triple-negative breast cancer. Tivantinib (ARQ 197) is an orally administered agent that targets MET, although recent preclinical data suggests the agent may have mechanisms of action that are independent of MET signaling. We conducted a phase 2 study of tivantinib monotherapy in patients with metastatic triple-negative breast cancer.

Cyclin-dependent kinases 4 and 6 (CDK4/6) are fundamental drivers of the cell cycle and are required for the initiation and progression of various malignancies. Pharmacological inhibitors of CDK4/6 have shown significant activity against several solid tumours. Their primary mechanism of action is thought to be the inhibition of phosphorylation of the retinoblastoma tumour suppressor, inducing G1 cell cycle arrest in tumour cells. Here we use mouse models of breast carcinoma and other solid tumours to show that selective CDK4/6 inhibitors not only induce tumour cell cycle arrest, but also promote anti-tumour immunity. We confirm this phenomenon through transcriptomic analysis of serial biopsies from a clinical trial of CDK4/6 inhibitor treatment for breast cancer. The enhanced anti-tumour immune response has two underpinnings. First, CDK4/6 inhibitors activate tumour cell expression of endogenous retroviral elements, thus increasing intracellular levels of double-stranded RNA. This in turn stimulates production of type III interferons and hence enhances tumour antigen presentation. Second, CDK4/6 inhibitors markedly suppress the proliferation of regulatory T cells. Mechanistically, the effects of CDK4/6 inhibitors both on tumour cells and on regulatory T cells are associated with reduced activity of the E2F target, DNA methyltransferase 1. Ultimately, these events promote cytotoxic T-cell-mediated clearance of tumour cells, which is further enhanced by the addition of immune checkpoint blockade. Our findings indicate that CDK4/6 inhibitors increase tumour immunogenicity and provide a rationale for new combination regimens comprising CDK4/6 inhibitors and immunotherapies as anti-cancer treatment.

Large, multidimensional cancer datasets provide a resource that can be mined to identify candidate therapeutic targets for specific subgroups of tumors. Here, we analyzed human breast cancer data to identify transcriptional programs associated with tumors bearing specific genetic driver alterations. Using an unbiased approach, we identified thousands of genes whose expression was enriched in tumors with specific genetic alterations. However, expression of the vast majority of these genes was not enriched if associations were analyzed within individual breast tumor molecular subtypes, across multiple tumor types, or after gene expression was normalized to account for differences in proliferation or tumor lineage. Together with linear modeling results, these findings suggest that most transcriptional programs associated with specific genetic alterations in oncogenes and tumor suppressors are highly context-dependent and are predominantly linked to differences in proliferation programs between distinct breast cancer subtypes. We demonstrate that such proliferation-dependent gene expression dominates tumor transcriptional programs relative to matched normal tissues. However, we also identified a relatively small group of cancer-associated genes that are both proliferation- and lineage-independent. A subset of these genes are attractive candidate targets for combination therapy because they are essential in breast cancer cell lines, druggable, enriched in stem-like breast cancer cells, and resistant to chemotherapy-induced down-regulation.

Triple-negative breast cancer (TNBC) is a highly heterogeneous disease, and molecular subtyping may result in improved diagnostic precision and targeted therapies. Our previous study classified TNBCs into four subtypes with putative therapeutic targets. Here, we conducted the FUTURE trial (ClinicalTrials.gov identifier: NCT03805399), a phase Ib/II subtyping-based and genomic biomarker-guided umbrella trial, to evaluate the efficacy of these targets. Patients with refractory metastatic TNBC were enrolled and stratified by TNBC subtypes and genomic biomarkers, and assigned to one of these seven arms: (A) pyrotinib with capecitabine, (B) androgen receptor inhibitor with CDK4/6 inhibitor, (C) anti PD-1 with nab-paclitaxel, (D) PARP inhibitor included, (E) and (F) anti-VEGFR included, or (G) mTOR inhibitor with nab-paclitaxel. The primary end point was the objective response rate (ORR). We enrolled 69 refractory metastatic TNBC patients with a median of three previous lines of therapy (range, 1-8). Objective response was achieved in 20 (29.0%, 95% confidence interval (CI): 18.7%-41.2%) of the 69 intention-to-treat (ITT) patients. Our results showed that immunotherapy (arm C), in particular, achieved the highest ORR (52.6%, 95% CI: 28.9%-75.6%) in the ITT population. Arm E demonstrated favorable ORR (26.1%, 95% CI: 10.2%-48.4% in the ITT population) but with more high grade (≥ 3) adverse events. Somatic mutations of TOP2A and CD8 immunohistochemical score may have the potential to predict immunotherapy response in the immunomodulatory subtype of TNBC. In conclusion, the phase Ib/II FUTURE trial suggested a new concept for TNBC treatment, demonstrating the clinical benefit of subtyping-based targeted therapy for refractory metastatic TNBC.

CALGB 40603 (NCT00861705), a 2 × 2 randomized phase II trial, demonstrated that adding carboplatin or bevacizumab to weekly paclitaxel (wP) followed by doxorubicin and cyclophosphamide significantly increased the pathologic complete response (pCR) rate in stage II-III triple-negative breast cancer. We now report long-term outcomes (LTOs) and correlative science end points.

The antigenic targets of immunity and the role of vaccination in breast cancer are unknown. We performed a phase I study of an autologous GM-CSF-secreting breast cancer vaccine in patients with metastatic and stage II-III breast cancer.

Sensitivity to endocrine therapy (ET) is critical for the clinical benefit from the combination of palbociclib plus ET in hormone receptor-positive/HER2-negative (HR+/HER2-) advanced breast cancer. Bazedoxifene is a third-generation selective estrogen receptor (ER) modulator and selective ER degrader with activity in preclinical models of endocrine-resistant breast cancer, including models harboring ESR1 mutations. Clinical trials in healthy women showed that bazedoxifene is well tolerated.

Neoadjuvant chemotherapy (NAC) is standard for many females with breast cancer (FBC). The efficacy of NAC in male breast cancer (MaBC) is unclear. The aim of this study was to compare proportions of pathologic complete response (pCR) between MaBC and FBC by tumor subtype (TS).

Treatment options for triple-negative breast cancer remain limited. Activation of the PI3K pathway via loss of PTEN and/or INPP4B is common. Buparlisib is an orally bioavailable, pan-class I PI3K inhibitor. We evaluated the safety and efficacy of buparlisib in patients with metastatic triple-negative breast cancer.