Citations

703 Background: Clinical and pathologic factors alone have limited prognostic ability in patients with metastatic clear cell renal cell carcinoma (ccRCC). We sought to determine if tissue biomarkers, along with our previously reported clinical metastases score, can be used to predict cancer specific survival (CSS) in patients with metastatic ccRCC. Methods: Patients with metastatic ccRCC who underwent nephrectomy between 1990-2004 were identified using the Mayo Clinic Nephrectomy Registry. Sections from paraffin-embedded primary tumor tissue blocks were used for immunohistochemistry staining for PD-1, B7-H1 (PD-L1), B7-H3, Bim (downstream pro-apoptotic signaling molecule in PD-1 pathway), CA-IX, IMP3, Ki67, and survivin. CSS was the primary outcome. Biomarkers that were significantly associated with CSS after adjusting for the metastases score were used to develop a biomarker-specific multivariable model using a bootstrap resampling approach and forward selection. Predictive ability was summarized using a bootstrap-corrected c-index. Results: The cohort included 602 patients, 192 (32%) with metastases at diagnosis and 410 (68%) who developed metastases after nephrectomy. Median follow-up among survivors was 9.6 years (IQR 4.2,12.8) and 504 patients died of RCC. Bim, IMP3, Ki67, and survivin expression were significantly associated with CSS after adjusting for the metastases score and were used to develop a biomarker-specific model. High Bim (HR 1.44; 95%CI 1.16-1.78; p < 0.001), high survivin (HR 1.35; 95%CI 1.08-1.68; p = 0.008), and the metastases score (HR 1.13 per one point; 95%CI 1.10-1.16; p < 0.001) were retained as independent predictors in the final multivariable model (c-index 0.69). Conclusions: We created a prognostic model combining the clinical metastases score and two primary tissue expression biomarkers, Bim and survivin, for patients with metastatic RCC who underwent nephrectomy. External validation will be required prior to clinical use.

We studied 57 patients with isolated lunotriquetral injuries treated by arthrodesis, direct ligament repair, or ligament reconstruction. The outcomes were compared by using written questionnaires, the Disabilities of the Arm, Shoulder and Hand (DASH) score, range of movement, strength, morbidity and rates of reoperation. Isolated lunotriquetral injury was confirmed by arthroscopy or arthrotomy. The mean age of the patients was 30.7 years (15.4 to 53.7) and the injuries were subacute or chronic in 98.2%. Eight patients underwent lunotriquetral reconstruction using a distally-based strip of the tendon of extensor carpi ulnaris, 27 had lunotriquetral repair and 22 had lunotriquetral arthrodesis. The mean follow-up was 9.5 years (2 to 22). The probability of remaining free from complications at five years was 68.6% for reconstruction, 13.5% for repair, and less than 1% for arthrodesis. Of the lunotriquetral arthrodeses, 40.9% developed nonunion and 22.7% developed ulnocarpal impaction. The probability of not requiring further surgery at five years was 68.6% for reconstruction, 23.3% for repair and 21.8% for arthrodesis. The DASH scores for each group were not significantly different. Objective improvements in strength and movement, subjective indicators of pain relief and satisfaction were significantly higher in the lunotriquetral repair and reconstruction groups than in those undergoing arthrodesis.

AbstractProtein phase diagrams have a unique potential to identify the presence of additional thermodynamic states even when non‐2‐state character is not readily apparent from the experimental observables used to follow protein unfolding transitions. Two‐state analysis of the von Willebrand factor A3 domain has previously revealed a discrepancy in the calorimetric enthalpy obtained from thermal unfolding transitions as compared with Gibbs‐Helmholtz analysis of free energies obtained from the Linear Extrapolation Method (Tischer and Auton, Prot Sci 2013; 22(9):1147‐60). We resolve this thermodynamic conundrum using a Clausius‐Clapeyron analysis of the urea‐temperature phase diagram that defines how and the urea m‐value interconvert through the slope of cm versus T, . This relationship permits the calculation of at low temperature from m‐values obtained through iso‐thermal urea denaturation and high temperature m‐values from obtained through iso‐urea thermal denaturation. Application of this equation uncovers sigmoid transitions in both cooperativity parameters as temperature is increased. Such residual thermal cooperativity of and the m‐value confirms the presence of an additional state which is verified to result from a cooperative phase transition between urea‐expanded and thermally‐compact denatured states. Comparison of the equilibria between expanded and compact denatured ensembles of disulfide‐intact and carboxyamidated A3 domains reveals that introducing a single disulfide crosslink does not affect the presence of the additional denatured state. It does, however, make a small thermodynamically favorable free energy (∼–13 ± 1 kJ/mol) contribution to the cooperative denatured state collapse transition as temperature is raised and urea concentration is lowered. The thermodynamics of this “cooperative collapse” of the denatured state retain significant compensations between the enthalpy and entropy contributions to the overall free energy.

Abstract The trend toward precision-based therapeutic approaches dictated by molecular alterations offers substantial promise for men with metastatic castration-resistant prostate cancer (mCRPC). However, current approaches for molecular characterization are primarily tissue based, necessitating serial biopsies to understand changes over time and are limited by the challenges inherent to extracting genomic material from predominantly bone metastases. Therefore, a circulating tumor cell (CTC)-based assay was developed to determine gene expression across a panel of clinically relevant and potentially actionable prostate cancer–related genes. CTCs were isolated from the whole blood of mCRPC patients (n = 41) and multiplex qPCR was performed to evaluate expression of prostate cancer–related target genes (n = 78). A large fraction of patients (27/41, 66%) had detectable CTCs. Increased androgen receptor (AR) expression (70% of samples) and evidence of Wnt signaling (67% of samples) were observed. The TMPRSS2:ERG fusion was expressed in 41% of samples, and the aggressive prostate cancer–associated long noncoding RNA SChLAP1 was upregulated in 70%. WNT5a [HR 3.62, 95% confidence interval (CI), 1.63–8.05, P = 0.002], AURKA (HR 5.56, 95% CI, 1.79–17.20, P = 0.003), and BMP7 (HR 3.86, 95% CI, 1.60–9.32, P = 0.003) were independently predictive of overall survival (FDR &lt; 10%) after adjusting for a panel of previously established prognostic variables in mCRPC (Halabi nomogram). A model including Halabi, WNT5a, and AURKA expression, termed the miCTC score, outperformed the Halabi nomogram alone (AUC = 0.89 vs. AUC = 0.70). Understanding the molecular landscape of CTCs has utility in predicting clinical outcomes in patients with aggressive prostate cancer and provides an additional tool in the arsenal of precision-based therapeutic approaches in oncology. Implications: Analysis of CTC gene expression reveals a clinically prognostic “liquid biopsy” signature in patients with metastatic castrate-resistance prostate cancer. Mol Cancer Res; 16(4); 643–54. ©2018 AACR.

Abstract Background: Pembrolizumab (Pembro), an anti-PD-1 immune checkpoint inhibitor, has been approved for the treatment of a variety of cancers including melanoma, non-small cell lung cancer, head and neck squamous cell carcinoma, and urothelial carcinoma. Pembro was recently evaluated in HER2- breast cancer patients in the neoadjuvant I-SPY 2 TRIAL and graduated in the triple negative (TN), HR+HER2-, and HER2- signatures. HER2- patients were randomized to receive Pembro+paclitaxel followed by doxorubicin/cyclophosphamide (P+T -&gt; AC) vs. T -&gt; AC. We and others have shown that TN breast cancers tend to have high numbers of immune infiltrates, including T cells and tumor associated macrophages (TAMs). We evaluated expression signatures representing 14 immune cell types (TILs, T cells, CD8 T cells, exhausted T cells, Th1, Tregs, cytotoxic cells, NK, NK CD56dim, dendritic cells, mast cells, B cells, macrophages, and neutrophils) as specific predictors of response to Pembro. Methods: Data from 248 patients (Pembro: 69; controls: 179) were available. Pre-treatment biopsies were assayed using Agilent gene expression arrays. Signature scores are calculated by averaging cell type specific genes. All I-SPY 2 qualifying biomarker analyses follow a pre-specified analysis plan. We used logistic modeling to assess biomarker performance. A biomarker is considered a specific predictor of Pembro response if it associates with response in the Pembro arm but not the control arm, and if the biomarker x treatment interaction is significant (likelihood ratio test, p&lt;0.05). This analysis is also performed adjusting for HR status as covariates, and within receptor subsets. For successful biomarkers, we use Bayesian modeling to estimate the pCR rates of 'predicted sensitive' patients in each arm. Our statistics are descriptive rather than inferential and do not adjust for multiplicities of other biomarkers outside this study. Results: 10 out of the 14 cell-type signatures tested are associated with response in the Pembro arm. Higher expression levels of 9 of these cell-type signatures are associated with higher pCR rates (T cells, exhausted T cells, Th1, cytotoxic cells, NK, NK CD56dim, dendritic cells, B cells, and macrophages), whereas higher mast cell signature expression is associated with non-pCR. Interestingly, many of these same signatures also associate or trend towards association with response in the control arm; and in a model adjusting for HR status, only 3 of these signatures (Th1, B cells and dendritic cells) show significant interaction with treatment. Within the whole population and the TN subtype, the dendritic cell signature is the strongest predictor of specific response to Pembro (OR/1SD: 4.04 and 4.4, LR p &lt; 0.001 overall and in TN). Although other immune signatures (T cells, exhausted T cells, NK, and macrophages) also associate with response in the Pembro arm in the TN subtype, only the dendritic cell and Th1 signatures have a significant interaction with treatment. In contrast, in the HR+HER2- subtype, only 3 signatures (Th1, B cells, and mast cells) associate with response to Pembro; but none of these signatures have significant interaction with treatment. Of note, in both the Pembro and control arms, HR+HER2- patients with higher average mast cell marker expression have lower pCR rates (OR/1SD: 0.33 and 0.51, LRp: 0.006 and 0.04 in Pembro and control arm). Conclusion: As expected, multiple immune cell expression signatures are predictive of response in the Pembro arm; but only dendritic cells and Th1 cells are specific to Pembro in both the population as a whole and the TN subtype. Interestingly, the presence of mast cells may impede response, especially in HR+HER2- patients. Correlation of these signatures with multiplex-IF immune markers is pending. Citation Format: Campbell M, Yau C, Borowsky A, Vandenberg S, Wolf D, Rimm D, Nanda R, Liu M, Brown-Swigart L, Hirst G, Asare S, van't Veer L, Yee D, DeMichele A, Berry D, Esserman L. Analysis of immune infiltrates (assessed via multiplex fluorescence immunohistochemistry) and immune gene expression signatures as predictors of response to the checkpoint inhibitor pembrolizumab in the neoadjuvant I-SPY 2 trial [abstract]. In: Proceedings of the 2017 San Antonio Breast Cancer Symposium; 2017 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2018;78(4 Suppl):Abstract nr PD6-08.

Abstract Background New treatment strategies for metastatic breast cancer (mBC) are mainly driven by therapies against specific targets. BRCA mutations are one of the few established actionable targets, with PARP-Inhibitors and Platinum showing high efficacy in mBC. Hereditary cancer testing panels are now broadly used for identification of individuals with BRCA1/2 mutations who may benefit from these therapies. Many of these panels also contain other predisposition genes involved in BRCA-related DNA repair pathways, but the clinical relevance of mutations in these genes remain unclear. The aim of this study was to describe the mutation rates of BRCA1/2 and panel-based predisposition genes, and the associated clinical characteristics of individuals with these mutations, in a prospective cohort of mBC patients. Methods The PRAEGNANT mBC registry (NCT02338167) is a prospective registry for metastatic breast cancer patients with a focus on molecular biomarkers. Patients receiving any therapeutic regimen are eligible for this registry. Germline DNA was collected at study entry and genotyped for 37 cancer predisposition genes including BRCA1 and BRCA2. The frequency of mutations in each gene was determined, and associations between mutations and patient and tumor characteristics, metastatic pattern, and overall survival were assessed. Results Mutations in established high (odds ratio (OR)&gt;5.0) and moderate risk (OR&gt;2.0) breast cancer genes (BRCA1/2, PALB2, CHEK2, ATM, RAD51D, BARD1, and MSH6) were seen in 123 of 1462 tested patients with mBC (8.4%). BRCA1 and BRCA2 mutations were seen in 1.4% and 2.9% of patients respectively. Most frequently mutated non-BRCA panel genes were CHEK2, PALB2 and ATM with 2.8%, 0.8% and 0.6% of patients. Mutation frequency varied with regard to patients who developed brain metastases, visceral metastases or bone only metastases. BRCA1 or BRCA2 mutations were seen frequently in patients with brain (5.3%) or visceral metastases (5.2%), but were present in only 2.5% patients with bone only metastases and 1.5% of patients with lesions in other locations. Panel genes were equally distributed among all metastatic patients. PALB2 mutations (n=11) were only seen in patients with brain (1.9%) and visceral metastases (0.9%), but not in patients with bone metastases or other locations. 36.4% (N=4) of all patients with PALB2 mutations developing a brain metastasis. When adjusted for other mBC prognostic factors, a mutation (all genes) was associated with an unfavorable prognosis (HR: 1.50; 95%CI: 1.04 to 2.30, p=0.03). Mutation frequencies were similar according to therapy lines. All other associations with molecular subtypes and risk factors were similar to primary breast cancer cases. Conclusion Mutations in high and moderate risk breast cancer genes were frequent in metastatic breast cancer patients. The frequency was substantially higher than the 4 to 5% mutation frequency observed among unselected primary breast cancer patients, but is consistent with recent results from studies of metastatic prostate cancer patients. Patients with brain and visceral metastases had the highest BRCA mutation rates. Results suggested that PALB2 mutations may be more frequent in patients with brain metastases. Citation Format: Fasching PA, Hu C, Hart SN, Polley EC, Lee KY, Gnanolivu RD, Lilyquist J, Hartkopf AD, Taran FA, Janni W, Hadji P, Tesch H, Haeberle L, Ettl J, Lux MP, Lueftner D, Wallwiener M, Mueller V, Beckmann MW, Belleville E, Wimberger P, Untch M, Kolberg H-C, Fehm TN, Overkamp F, Wallwiener D, Brucker SY, Schneeweiss A, Couch F. Cancer predisposition genes in metastatic breast cancer – Association with metastatic pattern, prognosis, patient and tumor characteristics [abstract]. In: Proceedings of the 2017 San Antonio Breast Cancer Symposium; 2017 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2018;78(4 Suppl):Abstract nr PD1-02.

Microglia are a major cellular component of gliomas, and abundant in the centre of the tumour and at the infiltrative margins. While glioma is a notoriously infiltrative disease, the dynamics of microglia and glioma migratory patterns have not been well characterized. To investigate the migratory behaviour of microglia and glioma cells at the infiltrative edge, we performed two-colour time-lapse fluorescence microscopy of brain slices generated from a platelet-derived growth factor-B (PDGFB)-driven rat model of glioma, in which glioma cells and microglia were each labelled with one of two different fluorescent markers. We used mathematical techniques to analyse glioma cells and microglia motility with both single cell tracking and particle image velocimetry (PIV). Our results show microglia motility is strongly correlated with the presence of glioma, while the correlation of the speeds of glioma cells and microglia was variable and weak. Additionally, we showed that microglia and glioma cells exhibit different types of diffusive migratory behaviour. Microglia movement fit a simple random walk, while glioma cell movement fits a super diffusion pattern. These results show that glioma cells stimulate microglia motility at the infiltrative margins, creating a correlation between the spatial distribution of glioma cells and the pattern of microglia motility.

ObjectiveCurrently, there are no reliable biomarkers for predicting therapeutic response in patients with rheumatoid arthritis (RA). The synovium may unlock critical information for determining efficacy, since a reduction in the numbers of sublining synovial macrophages remains the most reproducible biomarker. Thus, a clinically actionable method for the collection of synovial tissue, which can be analyzed using high‐throughput strategies, must become a reality. This study was undertaken to assess the feasibility of utilizing synovial biopsies as a precision medicine‐based approach for patients with RA.MethodsRheumatologists at 6 US academic sites were trained in minimally invasive ultrasound‐guided synovial tissue biopsy. Biopsy specimens obtained from patients with RA and synovial tissue from patients with osteoarthritis (OA) were subjected to histologic analysis, fluorescence‐activated cell sorting, and RNA sequencing (RNA‐seq). An optimized protocol for digesting synovial tissue was developed to generate high‐quality RNA‐seq libraries from isolated macrophage populations. Associations were determined between macrophage transcriptional profiles and clinical parameters in RA patients.ResultsPatients with RA reported minimal adverse effects in response to synovial biopsy. Comparable RNA quality was observed from synovial tissue and isolated macrophages between patients with RA and patients with OA. Whole tissue samples from patients with RA demonstrated a high degree of transcriptional heterogeneity. In contrast, the transcriptional profile of isolated RA synovial macrophages highlighted different subpopulations of patients and identified 6 novel transcriptional modules that were associated with disease activity and therapy.ConclusionPerformance of synovial tissue biopsies by rheumatologists in the US is feasible and generates high‐quality samples for research. Through the use of cutting‐edge technologies to analyze synovial biopsy specimens in conjunction with corresponding clinical information, a precision medicine–based approach for patients with RA is attainable.