PRMT5 is an actionable therapeutic target in CDK4/6 inhibitor-resistant ER+/RB-deficient breast cancer.

CDK4/6 inhibitors (CDK4/6i) have improved survival of patients with estrogen receptor-positive (ER+) breast cancer. However, patients treated with CDK4/6i eventually develop drug resistance and progress. RB1 loss-of-function alterations confer resistance to CDK4/6i, but the optimal therapy for these patients is unclear. Through a genome-wide CRISPR screen, we identify protein arginine methyltransferase 5 (PRMT5) as a molecular vulnerability in ER+/RB1-knockout breast cancer cells. Inhibition of PRMT5 blocks the G1-to-S transition in the cell cycle independent of RB, leading to growth arrest in RB1-knockout cells. Proteomics analysis uncovers fused in sarcoma (FUS) as a downstream effector of PRMT5. Inhibition of PRMT5 results in dissociation of FUS from RNA polymerase II, leading to hyperphosphorylation of serine 2 in RNA polymerase II, intron retention, and subsequent downregulation of proteins involved in DNA synthesis. Furthermore, treatment with the PRMT5 inhibitor pemrametostat and a selective ER degrader fulvestrant synergistically inhibits growth of ER+/RB-deficient cell-derived and patient-derived xenografts. These findings highlight dual ER and PRMT5 blockade as a potential therapeutic strategy to overcome resistance to CDK4/6i in ER+/RB-deficient breast cancer.

PLK4 as a potential target to enhance radiosensitivity in triple-negative breast cancer.

Radioresistance is one of the barriers to developing more effective therapies against the most aggressive, triple-negative, breast cancer (TNBC) subtype. In our previous studies, we showed that inhibition of Polo-like Kinase 4 (PLK4) by a novel drug, CFI-400945 significantly enhances the anticancer effects of radiotherapy (RT) compared to single treatment alone. Here we further investigate the role of PLK4 in enhancing radiation effects in TNBC and explore mechanisms of PLK4 inhibition and radiation combinatorial antiproliferative effects. To assess cellular proliferation in response to treatments, we used colony formation assays in TNBC cell lines and patient-derived organoids (PDOs). Downregulation of PLK4 expression was achieved using siRNA silencing in TNBC cell lines. Immunofluorescence against centrin was used to assess the alteration of centriole amplification in response to treatments. We observed that inhibition of PLK4 by CFI-400945 or Centrinone B or its downregulation by siRNA, when combined with RT, resulted in a significant increase in antiproliferative effect in TNBC cells lines and PDOs compared to untreated or single-treated cells. Anticancer synergy was observed using a response matrix in PDOs treated with CFI-400945 and RT. We show that the overamplification of centrioles might be involved in the combined antiproliferative action of RT and PLK4 inhibition. Our data suggest that PLK4 is a promising target for enhancing the anticancer effects of RT in TNBC that, at least in part, is modulated by the overamplification of centrioles. These results support further mechanistic and translational studies of anti-PLK4 agents and RT as an anticancer combination treatment strategy.

Seize the engine: Emerging cell cycle targets in breast cancer.

Breast cancer arises from a series of molecular alterations that disrupt cell cycle checkpoints, leading to aberrant cell proliferation and genomic instability. Targeted pharmacological inhibition of cell cycle regulators has long been considered a promising anti-cancer strategy. Initial attempts to drug critical cell cycle drivers were hampered by poor selectivity, modest efficacy and haematological toxicity. Advances in our understanding of the molecular basis of cell cycle disruption and the mechanisms of resistance to CDK4/6 inhibitors have reignited interest in blocking specific components of the cell cycle machinery, such as CDK2, CDK4, CDK7, PLK4, WEE1, PKMYT1, AURKA and TTK. These targets play critical roles in regulating quiescence, DNA replication and chromosome segregation. Extensive preclinical data support their potential to overcome CDK4/6 inhibitor resistance, induce synthetic lethality or sensitise tumours to immune checkpoint inhibitors. This review provides a biological and drug development perspective on emerging cell cycle targets and novel inhibitors, many of which exhibit favourable safety profiles and promising activity in clinical trials.

The Clinical, Genomic, and Transcriptomic Landscape of BRAF Mutant Cancers.

BACKGROUND: BRAF mutations are classified into four molecularly distinct groups, and Class 1 (V600) mutant tumors are treated with targeted therapies. Effective treatment has not been established for Class 2/3 or BRAF Fusions. We investigated whether BRAF mutation class differed according to clinical, genomic, and transcriptomic variables in cancer patients. METHODS: Using the AACR GENIE (v.12) cancer database, the distribution of BRAF mutation class in adult cancer patients was analyzed according to sex, age, primary race, and tumor type. Genomic alteration data and transcriptomic analysis was performed using The Cancer Genome Atlas. RESULTS: BRAF mutations were identified in 9515 (6.2%) samples among 153,834, with melanoma (31%), CRC (20.7%), and NSCLC (13.9%) being the most frequent cancer types. Class 1 harbored co-mutations outside of the MAPK pathway (TERT, RFN43) vs. Class 2/3 mutations (RAS, NF1). Across all tumor types, Class 2/3 were enriched for alterations in genes involved in UV response and WNT/ß-catenin. Pathway analysis revealed enrichment of WNT/ß-catenin and Hedgehog signaling in non-V600 mutated CRC. Males had a higher proportion of Class 3 mutations vs. females (17.4% vs. 12.3% q = 0.003). Non-V600 mutations were generally more common in older patients (aged 60+) vs. younger (38% vs. 15% p < 0.0001), except in CRC (15% vs. 30% q = 0.0001). Black race was associated with non-V600 BRAF alterations (OR: 1.58; p < 0.0001). CONCLUSIONS: Class 2/3 BRAFs are more present in Black male patients with co-mutations outside of the MAPK pathway, likely requiring additional oncogenic input for tumorigenesis. Improving access to NGS and trial enrollment will help the development of targeted therapies for non-V600 BRAF mutations.

A Phase I/II Study of GSK525762 Combined with Fulvestrant in Patients with Hormone Receptor-positive/HER2-negative Advanced or Metastatic Breast Cancer.

PURPOSE: Endocrine-based therapy is the initial primary treatment option for hormone receptor-positive and human epidermal growth factor receptor 2-negative (HR+/HER2-) metastatic breast cancer (mBC). However, patients eventually experience disease progression due to resistance to endocrine therapy. Molibresib (GSK525762) is a small-molecule inhibitor of bromodomain and extraterminal (BET) family proteins (BRD2, BRD3, BRD4, and BRDT). Preclinical data suggested that the combination of molibresib with endocrine therapy might overcome endocrine resistance. This study aimed to investigate the safety, tolerability, pharmacokinetics, pharmacodynamics, and efficacy [objective response rate (ORR)] of molibresib combined with fulvestrant in women with HR+/HER2- mBC. PATIENTS AND METHODS: In this phase I/II dose-escalation and dose-expansion study, patients received oral molibresib 60 or 80 mg once daily in combination with intramuscular fulvestrant. Patients enrolled had relapsed/refractory, advanced/metastatic HR+/HER2- breast cancer with disease progression on prior treatment with an aromatase inhibitor, with or without a cyclin-dependent kinase 4/6 inhibitor. RESULTS: The study included 123 patients. The most common treatment-related adverse events (AE) were nausea (52%), dysgeusia (49%), and fatigue (45%). At a 60-mg dosage of molibresib, >90% of patients experienced treatment-related AE. Grade 3 or 4 treatment-related AE were observed in 47% and 48% of patients treated with molibresib 60 mg and molibresib 80 mg, respectively. The ORR was 13% [95% confidence interval (CI), 8-20], not meeting the 25% threshold for proceeding to phase II. Among 82 patients with detected circulating tumor DNA and clinical outcome at study enrollment, a strong association was observed between the detection of copy-number amplification and poor progression-free survival (HR, 2.89; 95% CI, 1.73-4.83; P < 0.0001). CONCLUSIONS: Molibresib in combination with fulvestrant did not demonstrate clinically meaningful activity in this study.

Health-Related Quality of Life With Pembrolizumab+Chemotherapy Versus Placebo+Chemotherapy for Advanced Triple-Negative Breast Cancer: KEYNOTE-355.

BACKGROUND: In KEYNOTE-355 (NCT02819518), addition of pembrolizumab to chemotherapy led to statistically significant improvements in progression-free survival and overall survival in patients with advanced triple-negative breast cancer (TNBC) with tumor PD-L1 combined positive score (CPS) ≥10. We report patient-reported outcomes (PROs) from KEYNOTE-355. METHODS: Patients were randomized 2:1 to pembrolizumab 200 mg or placebo every 3 weeks for up to 35 cycles plus investigator's choice chemotherapy (nab-paclitaxel, paclitaxel, or gemcitabine/carboplatin). QLQ-C30, QLQ-BR23, and EQ-5D visual analogue scale (VAS) were prespecified. PROs were analyzed for patients who received ≥1 dose of study treatment and completed ≥1 PRO assessment. Change in PRO scores from baseline were assessed at week 15 (latest time point at which completion/compliance rates were ≥60%/≥80%). Time to deterioration (TTD) in PROs was defined as time to first onset of ≥ 10-point worsening in score from baseline. RESULTS: PRO analyses included 317 patients with tumor PD-L1 CPS ≥10 (pembrolizumab plus chemotherapy; n = 217; placebo plus chemotherapy, n = 100). There were no between-group differences in change from baseline to week 15 in QLQ-C30 global health status/quality of life (GHS/QoL; least-squares mean difference, -1.81 [95% CI, -6.92 to 3.30]), emotional functioning (-1.43 [-7.03 to 4.16]), physical functioning (-1.05 [-6.59 to 4.50]), or EQ-5D VAS (0.18 [-5.04 to 5.39]), and no between-group difference in TTD in QLQ-C30 GHS/QoL, emotional functioning, or physical functioning. CONCLUSIONS: Together with the efficacy and safety findings, PRO results from KEYNOTE-355 support pembrolizumab plus chemotherapy as a standard of care for patients with advanced TNBC with tumor PD-L1 (CPS ≥10).

Topoisomerase 1 Inhibition in MYC-Driven Cancer Promotes Aberrant R-Loop Accumulation to Induce Synthetic Lethality.

MYC is a central regulator of gene transcription and is frequently dysregulated in human cancers. As targeting MYC directly is challenging, an alternative strategy is to identify specific proteins or processes required for MYC to function as a potent cancer driver that can be targeted to result in synthetic lethality. To identify potential targets in MYC-driven cancers, we performed a genome-wide CRISPR knockout screen using an isogenic pair of breast cancer cell lines in which MYC dysregulation is the switch from benign to transformed tumor growth. Proteins that regulate R-loops were identified as a potential class of synthetic lethal targets. Dysregulated MYC elevated global transcription and coincident R-loop accumulation. Topoisomerase 1 (TOP1), a regulator of R-loops by DNA topology, was validated to be a vulnerability in cells with high MYC activity. Genetic knockdown of TOP1 in MYC-transformed cells resulted in reduced colony formation compared with control cells, demonstrating synthetic lethality. Overexpression of RNaseH1, a riboendonuclease that specifically degrades R-loops, rescued the reduction in clonogenicity induced by TOP1 deficiency, demonstrating that this vulnerability is driven by aberrant R-loop accumulation. Genetic and pharmacologic TOP1 inhibition selectively reduced the fitness of MYC-transformed tumors in vivo. Finally, drug response to TOP1 inhibitors (i.e., topotecan) significantly correlated with MYC levels and activity across panels of breast cancer cell lines and patient-derived organoids. Together, these results highlight TOP1 as a promising target for MYC-driven cancers. SIGNIFICANCE: CRISPR screening reveals topoisomerase 1 as an immediately actionable vulnerability in cancers harboring MYC as a driver oncoprotein that can be targeted with clinically approved inhibitors.

Differential DNA damage repair and PARP inhibitor vulnerability of the mammary epithelial lineages.

It is widely assumed that all normal somatic cells can equally perform homologous recombination (HR) and non-homologous end joining in the DNA damage response (DDR). Here, we show that the DDR in normal mammary gland inherently depends on the epithelial cell lineage identity. Bioinformatics, post-irradiation DNA damage repair kinetics, and clonogenic assays demonstrated luminal lineage exhibiting a more pronounced DDR and HR repair compared to the basal lineage. Consequently, basal progenitors were far more sensitive to poly(ADP-ribose) polymerase inhibitors (PARPis) in both mouse and human mammary epithelium. Furthermore, PARPi sensitivity of murine and human breast cancer cell lines as well as patient-derived xenografts correlated with their molecular resemblance to the mammary progenitor lineages. Thus, mammary epithelial cells are intrinsically divergent in their DNA damage repair capacity and PARPi vulnerability, potentially influencing the clinical utility of this targeted therapy.

TROPION-Breast02: Datopotamab deruxtecan for locally recurrent inoperable or metastatic triple-negative breast cancer.

Despite recent treatment advances, the prognosis for patients with locally recurrent inoperable or metastatic triple-negative breast cancer (TNBC) remains poor. The antibody-drug conjugate datopotamab deruxtecan (Dato-DXd) is composed of a humanized anti-TROP2 IgG1 monoclonal antibody linked to a topoisomerase I inhibitor payload via a stable, cleavable linker. The phase III TROPION-Breast02 trial in patients previously untreated for locally recurrent inoperable or metastatic TNBC, who are not candidates for PD-1/PD-L1 inhibitors is evaluating efficacy and safety of Dato-DXd versus investigator's choice of chemotherapy (ICC). Approximately 600 patients will be randomized 1:1 to Dato-DXd 6 mg/kg iv. every 3 weeks or ICC (paclitaxel, nab-paclitaxel, carboplatin, capecitabine or eribulin mesylate). Dual primary end points are progression-free survival by blinded independent central review and overall survival.

Triple-negative breast cancer (TNBC) is a subtype of breast cancer that is hard to treat. Tumors lack receptors for estrogen and progesterone, which means that standard endocrine therapy is ineffective, and it does not express HER2, so HER2 therapies are also not appropriate. However, the majority of TNBC tumors do possess a cell surface protein called TROP2 which provides a way of directing treatment inside tumor cells that is more selective than traditional chemotherapy. Datopotamab deruxtecan (Dato-DXd) is a drug that consists of two parts: datopotamab (an antibody) and DXd (the cancer-cell killing toxic component), which are joined via a stable linker. Datopotamab binds to the TROP2 protein found on TNBC tumors and is taken into the cell. The linker is then broken and releases DXd, which kills the tumor cell. By binding to cancer cells before releasing the payload, treatment is directed to the tumor, minimizing side effects in the rest of the body. The TROPION-Breast02 study aims to discover whether Dato-DXd is more effective than standard-of-care chemotherapy, allowing patients with TNBC to live longer without their breast cancer getting worse. This study is also looking at how Dato-DXd may affect patients' overall functioning and quality of life. TROPION-Breast02 will recruit approximately 600 patients who: Have cancer that has spread from the original site (metastatic), or cancer that returned to the same site (locally recurrent) that cannot be surgically removed Have not received any prior treatment for this stage of cancer Cannot receive an alternative type of anticancer treatment called PD-(L)1 inhibitors Had any length of time between their last treatment with the aim of cure and return of their disease Eligible patients will be randomly assigned to a treatment group in equal numbers to either Dato-DXd or an appropriate chemotherapy (one of five available options, chosen by the treating doctor). Each patient will generally continue to receive their designated treatments if the tumor is controlled by the drug, there are no unacceptable side effects, or the patient chooses to stop treatment. Clinical Trial Registration: NCT05374512 (ClinicalTrials.gov).

Protein arginine methyltransferase 5 (PRMT5) is an actionable therapeutic target in CDK4/6 inhibitor-resistant ER+/RB-deficient breast cancer.

CDK4/6 inhibitors (CDK4/6i) have improved survival of patients with estrogen receptor-positive (ER+) breast cancer. However, patients treated with CDK4/6i eventually develop drug resistance and progress. RB1 loss-of-function alterations confer acquired resistance to CDK4/6i, but the optimal therapy for these patients is unclear. Using a genome-wide CRISPR screen, we identified protein arginine methyltransferase 5 (PRMT5) as a molecular vulnerability in ER+/RB1-knockout (RBKO) breast cancer cells. PRMT5 inhibition blocked cell cycle G1-to-S transition independent of RB, thus arresting growth of RBKO cells. Proteomics analysis uncovered fused in sarcoma (FUS) as a downstream effector of PRMT5. Pharmacological inhibition of PRMT5 resulted in dissociation of FUS from RNA polymerase II (Pol II), Ser2 Pol II hyperphosphorylation, and intron retention in genes that promote DNA synthesis. Treatment with the PRMT5i inhibitor pemrametostat and fulvestrant synergistically inhibited growth of ER+/RB-deficient patient-derived xenografts, suggesting dual ER and PRMT5 blockade as a novel therapeutic strategy to treat ER+/RB-deficient breast cancer.

Real-World Use of Highly Sensitive Liquid Biopsy Monitoring in Metastatic Breast Cancer Patients Treated with Endocrine Agents after Exposure to Aromatase Inhibitors.

Endocrine-resistant, hormone receptor-positive, and HER2-negative (HR+/HER2-) metastatic breast cancer (mBC) is largely governed by acquired mutations in the estrogen receptor, which promote ligand-independent activation, and by truncal alterations in the PI3K signaling pathway, with a broader range of gene alterations occurring with less prevalence. Circulating tumor DNA (ctDNA)-based technologies are progressively permeating the clinical setting. However, their utility for serial monitoring has been hindered by their significant costs, inter-technique variability, and real-world patient heterogeneity. We interrogated a longitudinal collection of 180 plasma samples from 75 HR+/HER2- mBC patients who progressed or relapsed after exposure to aromatase inhibitors and were subsequently treated with endocrine therapy (ET) by means of highly sensitive and affordable digital PCR and SafeSEQ sequencing. Baseline PIK3CA and TP53 mutations were prognostic of a shorter progression-free survival in our population. Mutant PIK3CA was prognostic in the subset of patients receiving fulvestrant monotherapy after progression to a CDK4/6 inhibitor (CDK4/6i)-containing regimen, and its suppression was predictive in a case of long-term benefit with alpelisib. Mutant ESR1 was prognostic in patients who did not receive concurrent CDK4/6i, an impact influenced by the variant allele frequency, and its early suppression was strongly predictive of efficacy and associated with long-term benefit in the whole cohort. Mutations in ESR1, TP53, and KRAS emerged as putative drivers of acquired resistance. These findings collectively contribute to the characterization of longitudinal ctDNA in real-world cases of HR+/HER2- mBC previously exposed to aromatase inhibitors and support ongoing studies either targeting actionable alterations or leveraging the ultra-sensitive tracking of ctDNA.

Utility of ctDNA in predicting relapse in solid tumors after curative therapy: a meta-analysis.

BACKGROUND: Presence of circulating tumor DNA (ctDNA) is prognostic in solid tumors treated with curative intent. Studies have evaluated ctDNA at specific "landmark" or multiple "surveillance" time points. However, variable results have led to uncertainty about its clinical validity. METHODS: A PubMed search identified relevant studies evaluating ctDNA monitoring in solid tumors after curative intent therapy. Odds ratios for recurrence at both landmark and surveillance time points for each study were calculated and pooled in a meta-analysis using the Peto method. Pooled sensitivity and specificity weighted by individual study inverse variance were estimated and meta-regression using linear regression weighted by inverse variance was performed to explore associations between patient and tumor characteristics and the odds ratio for disease recurrence. RESULTS: Of 39 studies identified, 30 (1924 patients) and 24 studies (1516 patients) reported on landmark and surveillance time points, respectively. The pooled odds ratio for recurrence at landmark was 15.47 (95% confidence interval = 11.84 to 20.22) and at surveillance was 31.0 (95% confidence interval = 23.9 to 40.2). The pooled sensitivity for ctDNA at landmark and surveillance analyses was 58.3% and 82.2%, respectively. The corresponding specificities were 92% and 94.1%, respectively. Prognostic accuracy was lower with tumor agnostic panels and higher with longer time to landmark analysis, number of surveillance draws, and smoking history. Adjuvant chemotherapy negatively affected landmark specificity. CONCLUSIONS: Although prognostic accuracy of ctDNA is high, it has low sensitivity, borderline high specificity, and therefore modest discriminatory accuracy, especially for landmark analyses. Adequately designed clinical trials with appropriate testing strategies and assay parameters are required to demonstrate clinical utility.

Association Between Biomarkers and Clinical Outcomes of Pembrolizumab Monotherapy in Patients With Metastatic Triple-Negative Breast Cancer: KEYNOTE-086 Exploratory Analysis.

PURPOSE: In the two-cohort phase II KEYNOTE-086 study (ClinicalTrials.gov identifier: NCT02447003), first-line and second-line or later pembrolizumab monotherapy demonstrated antitumor activity in metastatic triple-negative breast cancer (mTNBC; N = 254). This exploratory analysis evaluates the association between prespecified molecular biomarkers and clinical outcomes. METHODS: Cohort A enrolled patients with disease progression after one or more systemic therapies for metastatic disease irrespective of PD-L1 status; Cohort B enrolled patients with previously untreated PD-L1-positive (combined positive score [CPS] ≥ 1) metastatic disease. The association between the following biomarkers as continuous variables and clinical outcomes (objective response rate [ORR], progression-free survival [PFS], and overall survival [OS]) was evaluated: PD-L1 CPS (immunohistochemistry), cluster of differentiation 8 (CD8; immunohistochemistry), stromal tumor-infiltrating lymphocyte (sTIL; hematoxylin and eosin staining), tumor mutational burden (TMB; whole-exome sequencing [WES]), homologous recombination deficiency-loss of heterozygosity, mutational signature 3 (WES), mutational signature 2 (apolipoprotein B mRNA editing catalytic polypeptide-like; WES), T-cell-inflamed gene expression profile (TcellinfGEP; RNA sequencing), and 10 non-TcellinfGEP signatures (RNA sequencing); Wald test P values were calculated, and significance was prespecified at α = 0.05. RESULTS: In the combined cohorts (A and B), PD-L1 (P = .040), CD8 (P < .001), sTILs (P = .012), TMB (P = .007), and TcellinfGEP (P = .011) were significantly associated with ORR; CD8 (P < .001), TMB (P = .034), Signature 3 (P = .009), and TcellinfGEP (P = .002) with PFS; and CD8 (P < .001), sTILs (P = .004), TMB (P = .025), and TcellinfGEP (P = .001) with OS. None of the non-TcellinfGEP signatures were associated with outcomes of pembrolizumab after adjusting for the TcellinfGEP. CONCLUSION: In this exploratory biomarker analysis from KEYNOTE-086, baseline tumor PD-L1, CD8, sTILs, TMB, and TcellinfGEP were associated with improved clinical outcomes of pembrolizumab and may help identify patients with mTNBC who are most likely to respond to pembrolizumab monotherapy.

Exploring immune interactions in triple negative breast cancer: IL-1ß inhibition and its therapeutic potential.

Triple negative breast cancer (TNBC) has poor prognosis when compared to other breast cancer subtypes. Despite pre-clinical data supporting an immune targeted approach for TNBCs, immunotherapy has failed to demonstrate the impressive responses seen in other solid tumor malignancies. Additional strategies to modify the tumor immune microenvironment and potentiate response to immunotherapy are needed. In this review, we summarise phase III data supporting the use of immunotherapy for TNBC. We discuss the role of IL-1ß in tumorigenesis and summarize pre-clinical data supporting IL-1ß inhibition as a potential therapeutic strategy in TNBC. Finally, we present current trials evaluating IL-1ß in breast cancer and other solid tumor malignancies and discuss future studies that may provide a strong scientific rationale for the combination of IL-1ß and immunotherapy in the neoadjuvant and metastatic setting for people with TNBC.

Changes in circulating tumor DNA and outcomes in solid tumors treated with immune checkpoint inhibitors: a systematic review.

BACKGROUND: Quantification of circulating tumor DNA (ctDNA) levels is a reliable prognostic tool in several malignancies. Dynamic changes in ctDNA levels in response to treatment may also provide prognostic information. Here, we explore the value of changes in ctDNA levels in response to immune checkpoint inhibitors (ICIs). METHODS: We searched MEDLINE (host: PubMed) for trials of ICIs in advanced solid tumors in which outcomes were reported based on change in ctDNA levels. ctDNA reduction was defined as reported in individual trials. Typically, this was either >50% reduction or a reduction to undetectable levels. We extracted HRs and related 95% CIs and/or p values comparing ctDNA reduction versus no reduction for progression-free survival (PFS) and/or overall survival (OS). Data were then pooled in a meta-analysis. Variation in effect size was examined using subgroup analyses. RESULTS: Eighteen trials were included in the meta-analysis. ctDNA levels were detectable in all participants in all studies prior to initiation of ICIs. A reduction in ctDNA measured 6-16 weeks after starting treatment was associated with significantly better PFS (HR 0.20; 95% CI, 0.14 to 0.28; p<0.001). Similarly, OS was superior in patients with reduced ctDNA levels (HR 0.18; 95% CI, 0.12 to 0.26; p<0.001). The results were consistent across all disease sites, lines of treatment, magnitude of change (to undetectable vs >50% reduction) and whether treatment exposure comprised single or combination ICIs. CONCLUSIONS: In advanced solid tumors, a reduction in ctDNA levels in response to ICIs is associated with substantial improvements in outcome. ctDNA change is an early response biomarker which may allow for de-escalation of cross-sectional imaging in patients receiving ICIs or support treatment de-escalation strategies.

Inhibition of the MNK1/2-eIF4E Axis Augments Palbociclib-Mediated Antitumor Activity in Melanoma and Breast Cancer.

Aberrant cell-cycle progression is characteristic of melanoma, and CDK4/6 inhibitors, such as palbociclib, are currently being tested for efficacy in this disease. Despite the promising nature of CDK4/6 inhibitors, their use as single agents in melanoma has shown limited clinical benefit. Herein, we discovered that treatment of tumor cells with palbociclib induces the phosphorylation of the mRNA translation initiation factor eIF4E. When phosphorylated, eIF4E specifically engenders the translation of mRNAs that code for proteins involved in cell survival. We hypothesized that cancer cells treated with palbociclib use upregulated phosphorylated eIF4E (phospho-eIF4E) to escape the antitumor benefits of this drug. Indeed, we found that pharmacologic or genetic disruption of MNK1/2 activity, the only known kinases for eIF4E, enhanced the ability of palbociclib to decrease clonogenic outgrowth. Moreover, a quantitative proteomics analysis of melanoma cells treated with combined MNK1/2 and CDK4/6 inhibitors showed downregulation of proteins with critical roles in cell-cycle progression and mitosis, including AURKB, TPX2, and survivin. We also observed that palbociclib-resistant breast cancer cells have higher basal levels of phospho-eIF4E, and that treatment with MNK1/2 inhibitors sensitized these palbociclib-resistant cells to CDK4/6 inhibition. In vivo we demonstrate that the combination of MNK1/2 and CDK4/6 inhibition significantly increases the overall survival of mice compared with either monotherapy. Overall, our data support MNK1/2 inhibitors as promising drugs to potentiate the antineoplastic effects of palbociclib and overcome therapy-resistant disease.

Polo-like kinase 4 inhibitor CFI-400945 suppresses liver cancer through cell cycle perturbation and eliciting antitumor immunity.

BACKGROUND AND AIMS: Prognosis of HCC remains poor due to lack of effective therapies. Immune checkpoint inhibitors (ICIs) have delayed response and are only effective in a subset of patients. Treatments that could effectively shrink the tumors within a short period of time are idealistic to be employed together with ICIs for durable tumor suppressive effects. HCC acquires increased tolerance to aneuploidy. The rapid division of HCC cells relies on centrosome duplication. In this study, we found that polo-like kinase 4 (PLK4), a centrosome duplication regulator, represents a therapeutic vulnerability in HCC. APPROACH AND RESULTS: An orally available PLK4 inhibitor, CFI-400945, potently suppressed proliferating HCC cells by perturbing centrosome duplication. CFI-400945 induced endoreplication without stopping DNA replication, causing severe aneuploidy, DNA damage, micronuclei formation, cytosolic DNA accumulation, and senescence. The cytosolic DNA accumulation elicited the DEAD box helicase 41-stimulator of interferon genes-interferon regulatory factor 3/7-NF-κß cytosolic DNA sensing pathway, thereby driving the transcription of senescence-associated secretory phenotypes, which recruit immune cells. CFI-400945 was evaluated in liver-specific p53/phosphatase and tensin homolog knockout mouse HCC models established by hydrodynamic tail vein injection. Tumor-infiltrated immune cells were analyzed. CFI-400945 significantly impeded HCC growth and increased infiltration of cluster of differentiation 4-positive (CD4 + ), CD8 + T cells, macrophages, and natural killer cells. Combination therapy of CFI-400945 with anti-programmed death-1 showed a tendency to improve HCC survival. CONCLUSIONS: We show that by targeting a centrosome regulator, PLK4, to activate the cytosolic DNA sensing-mediated immune response, CFI-400945 effectively restrained tumor progression through cell cycle inhibition and inducing antitumor immunity to achieve a durable suppressive effect even in late-stage mouse HCC.

90Y-Labeled Gold Nanoparticle Depot (NPD) Combined with Anti-PD-L1 Antibodies Strongly Inhibits the Growth of 4T1 Tumors in Immunocompetent Mice and Induces an Abscopal Effect on a Distant Non-Irradiated Tumor.

The effectiveness and normal tissue toxicity of a novel nanoparticle depot (NPD) brachytherapy seed incorporating gold nanoparticles (AuNPs) labeled with ß-particle emitting, 90Y (termed a "radiation nanomedicine"), were studied for the treatment of 4T1 triple-negative murine mammary carcinoma tumors in Balb/c mice and for inducing an abscopal effect on a distant non-irradiated tumor alone or combined with anti-PD-L1 immune checkpoint antibodies. Balb/c mice with two subcutaneous 4T1 tumors─a primary tumor and a distant secondary tumor were implanted intratumorally (i.t.) in the primary tumor with NPD incorporating 3.5 MBq of 90Y-AuNPs (1 × 1014 AuNPs) or unlabeled AuNPs, alone or combined with systemically administered anti-PD-L1 antibodies (200 µg i.p. three times/week for 2 weeks) or received anti-PD-L1 antibodies alone or no treatment. The primary tumor was strongly growth-inhibited over 14 d by NPD incorporating 90Y-AuNPs but only very modestly inhibited by NPD incorporating unlabeled AuNPs. Anti-PD-L1 antibodies alone were ineffective, and combining anti-PD-L1 antibodies with NPD incorporating 90Y-AuNPs did not further inhibit the growth of the primary tumor. Secondary tumor growth was inhibited by treatment of the primary tumor with NPD incorporating 90Y-AuNPs, and growth inhibition was enhanced by anti-PD-L1 antibodies. Treatment of the primary tumor with NPD incorporating unlabeled AuNPs or anti-PD-L1 antibodies alone had no effect on secondary tumor growth. Biodistribution studies showed high uptake of 90Y in the primary tumor [516-810% implanted dose/g (%ID/g)] but very low uptake in the secondary tumor (0.033-0.16% ID/g) and in normal tissues (<0.5% ID/g) except for kidneys (5-8% ID/g). Very high radiation absorbed doses were estimated for the primary tumor (472 Gy) but very low doses in the secondary tumor (0.13 Gy). There was highdose-heterogeneity in the primary tumor with doses as high as 9964 Gy in close proximity to the NPD, decreasing rapidly with distance from the NPD. Normal organ doses were low (<1 Gy) except for kidneys (4 Gy). No normal tissue toxicity was observed, but white blood cell counts (WBC) decreased in tumor-bearing mice treated with NPD incorporating 90Y-AuNPs. Decreased WBC counts were interpreted as tumor response and not toxicity since these were higher than that in healthy non-tumor-bearing mice, and there was a direct association between WBC counts and 4T1 tumor burden. We conclude that implantation of NPD incorporating 90Y-AuNPs into a primary 4T1 tumor in Balb/c mice strongly inhibited tumor growth and combined with anti-PD-L1 antibodies induced an abscopal effect on a distant secondary tumor. This radiation nanomedicine is promising for the local treatment of triple-negative breast cancer tumors in patients, and these therapeutic effects may extend to non-irradiated lesions, especially when combined with checkpoint immunotherapy.

Computational pharmacogenomic screen identifies drugs that potentiate the anti-breast cancer activity of statins.

Statins, a family of FDA-approved cholesterol-lowering drugs that inhibit the rate-limiting enzyme of the mevalonate metabolic pathway, have demonstrated anticancer activity. Evidence shows that dipyridamole potentiates statin-induced cancer cell death by blocking a restorative feedback loop triggered by statin treatment. Leveraging this knowledge, we develop an integrative pharmacogenomics pipeline to identify compounds similar to dipyridamole at the level of drug structure, cell sensitivity and molecular perturbation. To overcome the complex polypharmacology of dipyridamole, we focus our pharmacogenomics pipeline on mevalonate pathway genes, which we name mevalonate drug-network fusion (MVA-DNF). We validate top-ranked compounds, nelfinavir and honokiol, and identify that low expression of the canonical epithelial cell marker, E-cadherin, is associated with statin-compound synergy. Analysis of remaining prioritized hits led to the validation of additional compounds, clotrimazole and vemurafenib. Thus, our computational pharmacogenomic approach identifies actionable compounds with pathway-specific activities.