Cancer-stromal cell interactions in breast cancer brain metastases induce glycocalyx-mediated resistance to HER2-targeting therapies.

Brain metastatic breast cancer is particularly lethal largely due to therapeutic resistance. Almost half of the patients with metastatic HER2-positive breast cancer develop brain metastases, representing a major clinical challenge. We previously described that cancer-associated fibroblasts are an important source of resistance in primary tumors. Here, we report that breast cancer brain metastasis stromal cell interactions in 3D cocultures induce therapeutic resistance to HER2-targeting agents, particularly to the small molecule inhibitor of HER2/EGFR neratinib. We investigated the underlying mechanisms using a synthetic Notch reporter system enabling the sorting of cancer cells that directly interact with stromal cells. We identified mucins and bulky glycoprotein synthesis as top-up-regulated genes and pathways by comparing the gene expression and chromatin profiles of stroma-contact and no-contact cancer cells before and after neratinib treatment. Glycoprotein gene signatures were also enriched in human brain metastases compared to primary tumors. We confirmed increased glycocalyx surrounding cocultures by immunofluorescence and showed that mucinase treatment increased sensitivity to neratinib by enabling a more efficient inhibition of EGFR/HER2 signaling in cancer cells. Overexpression of truncated MUC1 lacking the intracellular domain as a model of increased glycocalyx-induced resistance to neratinib both in cell culture and in experimental brain metastases in immunodeficient mice. Our results highlight the importance of glycoproteins as a resistance mechanism to HER2-targeting therapies in breast cancer brain metastases.

Pregnancy and neonatal outcomes among women with early-onset colorectal cancer: a nationwide case-control study.

Background: Early-onset colorectal cancer has risen worldwide, leaving more women with colorectal cancer at reproductive ages. We aimed to investigate the risk of adverse pregnancy and neonatal outcomes among women with early-onset colorectal cancer. Methods: We conducted a nationwide, matched case-control study of maternal/pregnancy outcomes including pre-eclampsia and Cesarean delivery (C-section) as well as neonatal outcomes including preterm birth among 207 births in women with early-onset colorectal cancer (ages 18-49) and 1019 births in women without colorectal cancer in Sweden (1992-2019). Early-onset colorectal cancer cases were identified through the Cancer Register, and outcome data were retrieved through linkage to Medical Birth Register and National Patient Register. Using conditional logistic regression, we estimated multivariable-adjusted odds ratios (ORs) and 95% confidence intervals (CIs). Findings: Between Jan 1, 1992, and Dec 31, 2019, women with early-onset colorectal cancer who gave birth had increased odds of pre-eclampsia (7.2% vs 3.2%; OR = 2.52, 95%CI = 1.25-5.08), any C-section (24.6% vs 19.4%; OR = 1.43, 95%CI = 1.00-2.06), particularly emergency C-section (17.4% vs 10.5%; OR = 1.79, 95%CI = 1.17-2.75), after adjustment for maternal education level, country of birth, body mass index and smoking in early pregnancy, and comorbidities. Maternal history of early-onset colorectal cancer was also associated with offspring preterm birth (12.1% vs 5.2%; OR = 2.31, 95%CI = 1.34-3.99), delineated as spontaneous (OR = 1.06, 95%CI = 0.47-2.39) or medically-indicated preterm birth (OR = 4.48, 95%CI = 2.05-9.79). There was no increased risk of congenital malformation or small for gestational age birth. Interpretation: In this population-based study, maternal history of early-onset colorectal cancer was associated with risk of both adverse pregnancy (pre-eclampsia, C-section) and neonatal outcomes (preterm birth). Funding: US National Institutes of Health, Swedish Society of Medicine, Swedish Cancer Foundation.

Clonal Hematopoiesis and Risk of Incident Lung Cancer.

PURPOSE: To prospectively examine the association between clonal hematopoiesis (CH) and subsequent risk of lung cancer. METHODS: Among 200,629 UK Biobank (UKBB) participants with whole-exome sequencing, CH was identified in a nested case-control study of 832 incident lung cancer cases and 3,951 controls (2006-2019) matched on age and year at blood draw, sex, race, and smoking status. A similar nested case-control study (141 cases/652 controls) was conducted among 27,975 participants with whole-exome sequencing in the Mass General Brigham Biobank (MGBB, 2010-2021). In parallel, we compared CH frequency in published data from 5,003 patients with solid tumor (2,279 lung cancer) who had pretreatment blood sequencing performed through Memorial Sloan Kettering-Integrated Mutation Profiling of Actionable Cancer Targets. RESULTS: In UKBB, the presence of CH was associated with increased risk of lung cancer (cases: 12.5% v controls: 8.7%; multivariable-adjusted odds ratio [OR], 1.36; 95% CI, 1.06 to 1.74). The association remained robust after excluding participants with chronic obstructive pulmonary disease. No significant interactions with known risk factors, including polygenic risk score and C-reactive protein, were identified. In MGBB, we observed similar enrichment of CH in lung cancer (cases: 15.6% v controls: 12.7%). The meta-analyzed OR (95% CI) of UKBB and MGBB was 1.35 (1.08 to 1.68) for CH overall and 1.61 (1.19 to 2.18) for variant allele frequencies ≥ 10%. In Memorial Sloan Kettering-Integrated Mutation Profiling of Actionable Cancer Targets, CH with a variant allele frequency ≥ 10% was enriched in pretreatment lung cancer compared with other tumors after adjusting for age, sex, and smoking (OR for lung v breast cancer: 1.61; 95% CI, 1.03 to 2.53). CONCLUSION: Independent of known risk factors, CH is associated with increased risk of lung cancer.

Disagreement in Readmission Rates After Total Hip and Knee Arthroplasty Across Data Sets.

BACKGROUND: In 2014, the Affordable Care Act Hospital Readmissions Reduction Program began penalizing hospitals for excessive readmission rates 30 days after total hip arthroplasty (THA) and total knee arthroplasty (TKA). Various data sets with nonstandardized validation processes report readmission data, which may provide conflicting outcome values for the same patient populations. METHODS: We queried 4 separate data sets: the American Joint Replacement Registry, Centers for Medicare and Medicaid Services billing data, the Vizient data set, and an advanced analytics integration (Cognos) report from our electronic medical record. We identified 2763 patients who underwent primary TKA and THA at a single academic medical center from June 2016 to June 2019. We then matched 613 surgery encounters in all 4 databases. Our primary outcome metric was 30-day readmissions. Fleiss' Kappa was used to measure agreement among the different data sets. RESULTS: Of the 613 THA and TKA patients, there were 45 (7.3%) readmissions noted. Data collected from the Centers for Medicare and Medicaid Services flagged 41 (6.7%) readmissions, Vizient flagged 11 (1.8%) readmissions, and the American Joint Replacement Registry and Cognos report both flagged 6 (0.98%) readmissions each. None of the readmissions were identified by all 4 data sets. There was significant disagreement among data sets using Fleiss' Kappa (kappa = -0.1318, P = .03). CONCLUSION: There is disagreement in readmission rates in databases receiving the same patient data after THA and TKA. Care must be taken to establish standard validation processes and reporting methods and scrutiny applied when interpreting readmission rates from various data sets.

Spinal anesthesia in a designated block bay for total joint arthroplasty: improving operating room efficiency.

BACKGROUND: Creating highly efficient operating room (OR) protocols for total joint arthroplasty (TJA) is a challenging and multifactorial process. We evaluated whether spinal anesthesia in a designated block bay (BBSA) would reduce time to incision, improve first case start time and decrease conversion to general anesthesia (GA). METHODS: Retrospective cohort study on the first 86 TJA cases with BBSA from April to December 2018, compared with 344 TJA cases with spinal anesthesia performed in the OR (ORSA) during the same period. All TJA cases were included if the anesthetic plan was for spinal anesthesia. Patients were excluded if circumstances delayed start time or time to incision (advanced vascular access, pacemaker interrogation, surgeon availability). Data were extracted and analyzed via a linear mixed effects model to compare time to incision, via a Wilcoxon rank-sum test to compare first case start time, and via a Fisher's exact test to compare conversion to GA between the groups. RESULTS: In the mixed effect model, the BBSA group time to incision was 5.37 min less than the ORSA group (p=0.018). The BBSA group had improved median first case start time (30.0 min) versus the ORSA group (40.5 min, p<0.0001). There was lower conversion to GA 2/86 (2.33%) in the BBSA group versus 36/344 (10.47%) in the ORSA group (p=0.018). No serious adverse events were noted in either group. CONCLUSIONS: BBSA had limited impact on time to incision for TJA, with a small decrease for single OR days and no improvement on OR days with two rooms. BBSA was associated with improved first case start time and decreased rate of conversion to GA. Further research is needed to identify how BBSA affects the efficiency of TJA.

Inhibition of PI3K by copanlisib exerts potent antitumor effects on Merkel cell carcinoma cell lines and mouse xenografts.

Merkel cell carcinoma (MCC) is a highly aggressive neuroendocrine skin cancer with steadily increasing incidence and poor prognosis. Despite recent success with immunotherapy, 50% of patients still succumb to their diseases. To date, there is no Food and Drug Administration-approved targeted therapy for advanced MCC. Aberrant activation of phosphatidylinositide-3-kinase (PI3K)/AKT/mTOR pathway is frequently detected in MCC, making it an attractive therapeutic target. We previously found PI3K pathway activation in human MCC cell lines and tumors and demonstrated complete clinical response in a Stage IV MCC patient treated with PI3K inhibitor idelalisib. Here, we found that both PI3K-α and -δ isoforms are abundantly expressed in our MCC cell lines and clinical samples; we therefore examined antitumor efficacy across a panel of five PI3K inhibitors with distinctive isoform-specificities, including idelalisib (PI3K-δ), copanlisib (PI3K-α/δ), duvelisib (PI3K-γ/δ), alpelisib (PI3K-α), and AZD8186 (PI3K-ß/δ). Of these, copanlisib exerts the most potent antitumor effects, markedly inhibiting cell proliferation, survival, and tumor growth by suppressing PI3K/mTOR/Akt activities in mouse models generated from MCC cell xenografts and patient-derived tumor xenografts. These results provide compelling preclinical evidence for application of copanlisib in advanced MCC with aberrant PI3K activation for which immunotherapy is insufficient, or patients who are unsuitable for immunotherapy.

Rapamycin inhibits the secretory phenotype of senescent cells by a Nrf2-independent mechanism.

Senescent cells contribute to age-related pathology and loss of function, and their selective removal improves physiological function and extends longevity. Rapamycin, an inhibitor of mTOR, inhibits cell senescence in vitro and increases longevity in several species. Nrf2 levels have been shown to decrease with aging and silencing Nrf2 gene induces premature senescence. Therefore, we explored whether Nrf2 is involved in the mechanism by which rapamycin delays cell senescence. In wild-type (WT) mouse fibroblasts, rapamycin increased the levels of Nrf2, and this correlates with the activation of autophagy and a reduction in the induction of cell senescence, as measured by SA-ß-galactosidase (ß-gal) staining, senescence-associated secretory phenotype (SASP), and p16 and p21 molecular markers. In Nrf2KO fibroblasts, however, rapamycin still decreased ß-gal staining and the SASP, but rapamycin did not activate the autophagy pathway or decrease p16 and p21 levels. These observations were further confirmed in vivo using Nrf2KO mice, where rapamycin treatment led to a decrease in ß-gal staining and pro-inflammatory cytokines in serum and fat tissue; however, p16 levels were not significantly decreased in fat tissue. Consistent with literature demonstrating that the Stat3 pathway is linked to the production of SASP, we found that rapamycin decreased activation of the Stat3 pathway in cells or tissue samples from both WT and Nrf2KO mice. Our data thus suggest that cell senescence is a complex process that involves at least two arms, and rapamycin uses Nrf2 to regulate cell cycle arrest, but not the production of SASP.

Dual mTOR inhibitor MLN0128 suppresses Merkel cell carcinoma (MCC) xenograft tumor growth.

Merkel cell carcinoma (MCC) is an aggressive neuroendocrine skin cancer. Pathologic activation of PI3K/mTOR pathway and elevated expression of c-Myc are frequently detected in MCC. Yet, there is no targeted therapy presently available for this lethal disease. Recently, MLN0128, a second-generation dual TORC1/2 inhibitor is shown to have therapeutic efficacy in preclinical studies. MLN0128 is currently in clinical trials as a potential therapy for advanced cancers. Here we characterize the therapeutic efficacy of MLN0128 in the preclinical setting of MCC and delineate downstream targets of mTORC1/2 in MCC cellular systems. MLN0128 significantly attenuates xenograft MCC tumor growth independent of Merkel cell polyomavirus. Moreover, MLN0128 markedly diminishes MCC cell proliferation and induces apoptosis. Further investigations indicate that senescence does not contribute to MLN0128-mediated repression of xenograft MCC tumor growth. Finally, we also observe robust antitumor effects of MLN0128 when administered as a dual therapy with JQ1, a bromodomain protein BRD4 inhibitor. These results suggest dual blockade of PI3K/mTOR pathway and c-Myc axis is effective in the control of MCC tumor growth. Our results demonstrate that MLN0128 is potent as monotherapy or as a member of combination therapy with JQ1 for advanced MCC.