Citations

To evaluate the clinical utility of cell‐free DNA (cfDNA), we performed whole‐genome sequencing to systematically examine plasma cfDNA copy number variations (CNVs) in a cohort of patients with colorectal cancer (CRC, n = 80), polyps (n = 20), and healthy controls (n = 35). We initially compared cfDNA yield in 20 paired serum–plasma samples and observed significantly higher cfDNA concentration in serum (median = 81.20 ng, range 7.18–500 ng·mL−1) than in plasma (median = 5.09 ng, range 3.76–62.8 ng·mL−1) (P < 0.0001). However, tumor‐derived cfDNA content was significantly lower in serum than in matched plasma samples tested. With ~10 million reads per sample, the sequencing‐based copy number analysis showed common CNVs in multiple chromosomal regions, including amplifications on 1q, 8q, and 5q and deletions on 1p, 4q, 8p, 17p, 18q, and 22q. Copy number changes were also evident in genes critical to the cell cycle, DNA repair, and WNT signaling pathways. To evaluate whether cumulative copy number changes were associated with tumor stages, we calculated plasma genomic abnormality in colon cancer (PGA‐C) score by summing the most significant CNVs. The PGA‐C score showed predictive performance with an area under the curve from 0.54 to 0.84 for CRC stages I‐IV. Locus‐specific copy number analysis identified nine genomic regions where CNVs were significantly associated with survival in stage III‐IV CRC patients. A multivariate model using six of nine genomic regions demonstrated a significant association of high‐risk score with shorter survival (HR = 5.33, 95% CI = 6.76–94.44, P < 0.0001). Our study demonstrates the importance of using plasma (rather than serum) to test tumor‐related genomic variations. Plasma cfDNA‐based tests can capture tumor‐specific genetic changes and may provide a measurable classifier for assessing clinical outcomes in advanced CRC patients.

152 Background: Plasma cfDNA ARcopy number variations (CNV) have been suggested to have prognostic significance in mCRPC stage. We compared plasma AR amplification ( ARamp) with CTC counts (the only FDA approved prognostic marker in mCRPC stage) in a prospective trial of patients (pts) progressing on androgen deprivation therapy. Methods: Plasma and CTC counts were obtained concurrently as part of a prospective clinical trial in mCRPC stage pts initiating abiraterone acetate (NCT# 01953640). Plasma cfDNA was purified using DNA Blood Mini Kit (Qiagen, Valencia, CA) and AR CNVs were detected using QuantStudio3D digital PCR system (Life Technologies, Carlsbad, CA, USA). CTC enumeration was performed using the FDA cleared CellSearch platform. The distributions of overall survival (OS) time based on ARamp status and CTC counts ( < 5 vs ≥ 5 cells) were estimated using the Kaplan-Meier method. Log-rank tests were used to test association of ARamp status and CTC counts with overall survival (OS). Univariate and multivariate logistic regression models and ROC evaluated association of OS at 24 months by ARamp status and CTCs ≥ 5. Results: Between 05/2013 and 09/2015, 92 pts were enrolled of which complete CTC, plasma and outcome data was available for 70 pts. Median age of the cohort (range) was 72 yrs (39-91); Median PSA–16.2 ng/ml (range: 0.9-2026.0); Median CTC count was 2.0 (range: 0-372); 58/70 pts had bone metastases; 38/70 had high volume and 32/70 had low volume metastatic disease at the time of sample collection. The median (range) follow up time at the time of this analysis was: 26.5 (7.9-37.0) months at which time 28/70 patients had died. ARamp was present in 19/70 pts while 51/70 did not have amplification. 24/70 had CTCs ≥ 5. A significant association of increased ARamp status with OS (p < 0.0001, log-rank test; AUC: 0.66) as well as with CTC counts ≥ 5 (p = 0.001, log-rank test; AUC: 0.67) was observed. Combining CTC status and plasma ARamp status the AUC was 0.75. Conclusions: Plasma cfDNA ARamp status and CTC counts have comparable and clinically meaningful prognostic marker value in mCRPC stage patients.

PurposeIn diffusion MRI (dMRI), fractional anisotropy derived from the single‐tensor model (FADTI) is the most widely used metric to characterize white matter (WM) microarchitecture, despite known limitations in regions with crossing fibers. Due to time constraints when scanning patients in clinical settings, high angular resolution diffusion imaging acquisition protocols, often used to overcome these limitations, are still rare in clinical population studies. However, the tensor distribution function (TDF) may be used to model multiple underlying fibers by representing the diffusion profile as a probabilistic mixture of tensors.MethodsWe compared the ability of standard FADTIand TDF‐derived FA (FATDF), calculated from a range of dMRI angular resolutions (41, 30, 15, and 7 gradient directions), to profile WM deficits in 251 individuals from the Alzheimer's Disease Neuroimaging Initiative and to detect associations with 1) Alzheimer's disease diagnosis, 2) Clinical Dementia Rating scores, and 3) average hippocampal volume.ResultsAcross angular resolutions and statistical tests, FATDFshowed larger effect sizes than FADTI, particularly in regions preferentially affected by Alzheimer's disease, and was less susceptible to crossing fiber anomalies.ConclusionThe TDF “corrected” form of FA may be a more sensitive and accurate alternative to the commonly used FADTI, even in clinical quality dMRI data. Magn Reson Med 78:2322–2333, 2017. © 2017 International Society for Magnetic Resonance in Medicine.

463 Background: The genomic assessment of cancer has been revolutionized by next-generation sequencing and is increasingly being applied in the clinic to guide therapeutic decision-making. Time to reporting of results, specimen quantity, and analyte quality have constrained initial clinical applications to gene panels and whole exome based strategies. Methods: Patients underwent surgical resection, excisional or core biopsies, or bone marrow biopsy. Samples were analyzed by whole genome or exome sequencing and RNA sequencing, bioinformatics analysis, and therapeutic target prioritization by a multi-disciplinary Clinical Genomics Board. All prioritized targets were CLIA validated using Sanger sequencing, RT-qPCR, FISH, or IHC as appropriate. Treatment was delivered using off-label FDA approved drugs, clinical trials, or single patient INDs. Results: We have enrolled 40 patients with advanced, treatment-refractory cancers of whom sequencing data is available on 33. The initial 6 patients were evaluated in a non-CLIA pilot phase and 27 in the CLIA enabled phase. Upon availability of the initial report, identified targets of putative therapeutic relevance were then prioritized by the CGB in 22/27 patients (81%) for subsequent CLIA validation. Eleven patients have been treated with genomically selected therapy with partial response in 3/10 assessed patients. A testicular cancer patient had aberrations in two testes specific genes, a copy number gain in TSSK6 and a novel gene fusion between thyroid hormone receptor associated protein 3 (THRAP3) and Tektin 2 (TEKT2). Additionally, a case of papillary renal cell carcinoma had an amplification of YAP1 and a previously unreported P287T mutation in CCND1, suggesting potential benefit with a CDK4/6 inhibitor. Treatment is ongoing and results will be reported. Conclusions: Integrated whole genome analysis in a CLIA setting is feasible. Integration of SNV, copy number and transcriptional data may allow for selection of putative driver genes to enhance targeted therapy decisions. Barriers for future broader implementation include the need for reduced time from biopsy to report and availability of therapies.

260 Background: The purpose of this study was to evaluate cell free nucleic acid based biomarkers for prognosis in mCRPC stage. Methods: Uniformly collected blood from mCRPC patients was processed for plasma which was used for exosomal RNA extraction. RNA sequencing was applied to examine microRNA (miRNA) profiles initially in a discovery cohort of 23 mCRPC patients. The sequences from fastq files were mapped to human miRBase (release 19) for known miRNA read counts. A software package (miRDeep2) was used for novel miRNA prediction. Cox regression was used for association of candidate miRNAs with overall survival (OS), defined as time from development of mCRPC to death or last follow up after adjusting for Gleason Score (GS) and time on androgen deprivation (AD) for hormone sensitive disease. miRNAs significantly associated with OS (FDR<0.05) were then validated in an independent cohort of 100 mCRPC subjects using univariate Cox regression. Results: RNA sequencing generated >10 million mappable reads per specimen in the discovery cohort. Among these reads ~50% were known miRNAs which included 1237 unique RNA sequences. Due to low abundance of most RNA transcripts, only miRNAs (n=250) with >/= 32 reads (on average) were analyzed. Four mRNAs (miR-1290, -1246, -375 and a predicted miRNA on chr. 12) were associated with OS(FDR<0.05). These were evaluated as prognotic markers in the independent cohort (N=100) using real time RT-PCR. Median time on AD for this cohort prior to progression to mCRPC was 35.7 (range: 3.5-136) months; median GS was 7; median follow up after mCRPC development was 12.5 (1-41) months at which time 28 patients had died. Elevated expression for miR-1290 and -375 were associated with shorter OS after adjusting for GS and AD time with a Hazard Ratio (HR) for miR-375 of 1.19 (CI: 1.006-1.18); (p=0.04) and a HR for miR1290 of 1.58 (CI:1.01-2.4); (p=0.04). A stronger association with shorter OS was observed for elevated expression of both miRs-HR of 2.02 (CI: 1.1-3.6) (p=0.01). Conclusions: Blood-based nucleic acid assays are easily accessible for biomarker development in mCRPC. Results from this study suggest specific exosomal miRs are of prognostic value in mCRPC and should be validated in larger cohorts.

PURPOSE: We sought to determine the preoperative factors associated with surgical margin status in patients who underwent radical prostatectomy for prostate cancer. PATIENTS AND METHODS: The study group consisted of 339 patients who were treated by radical retropubic prostatectomy and bilateral pelvic lymphadenectomy at the Mayo Clinic. None received preoperative adjuvant therapy. The mean age at the time of surgery was 66 years (range, 45 to 79 years). All specimens were totally embedded and whole-mounted. Positive surgical margin was defined as the presence of cancer cells at the inked margins. Numerous pathologic characteristics in needle biopsies and preoperative clinical findings were analyzed. RESULTS: The overall margin positivity rate was 24%. In univariate analysis, preoperative serum prostate-specific antigen (PSA) level, Gleason score, perineural invasion, percentage of cancer in the biopsy specimens, and number and percentage of biopsy cores involved by cancer were all associated with positive surgical margins. In multivariate analysis, preoperative serum PSA level (odds ratio for a doubling of PSA levels, 1.9; 95% confidence interval, 1.5 to 2.4; P < .001) and percentage of cancer in the biopsy specimens (odds ratio for a 10% increase, 1.3; 95% confidence interval, 1.2 to 1.4; P < .001) were predictive of margin status in radical prostatectomy. With use of preoperative serum PSA level and percentage of cancer in the biopsy as predictors of surgical margins, the overall accuracy as measured by the area under the receiver operating characteristic curve was 0.74. CONCLUSION: Preoperative serum PSA level and percentage of cancer in the biopsy specimens were independently associated with surgical margin status in patients who underwent radical prostatectomy for prostate cancer. The combination of these two factors provides a high level of predictive accuracy for margin status.

PURPOSE: A significant number of patients with stage T1 bladder carcinoma are at risk for cancer progression. We sought to identify factors associated with cancer progression in a series of patients with stage T1 bladder carcinoma treated with a contemporary therapeutic approach. PATIENTS AND METHODS: The study population consisted of 83 consecutive patients in whom stage T1 bladder carcinoma was diagnosed at the Mayo Clinic between 1987 and 1992. All patients underwent transurethral resection of the bladder (TURB) and had histologic confirmation of the diagnosis. The mean age was 71 years (range, 47 to 94 years). The male-to-female ratio was 3.9:1. The mean length of follow-up was 5.2 years (range, 1 day to 10.4 years). The depth of lamina propria invasion in the TURB specimens was measured with an ocular micrometer. Cancer progression was defined as the development of muscle-invasive or more advanced stage carcinoma, distant metastasis, or death from bladder cancer. RESULTS: The overall 5- and 7-year progression-free survival rates were 82% and 80%, respectively. The depth of invasion in the TURB specimens was associated with cancer progression (hazards ratio, 1.6 for doubling of depth of invasion; 95% confidence interval, 1.03 to 2.4; P = .037). The 5-year progression-free survival rate for patients with depth of invasion of ≥ 1.5 mm was 67%, compared with 93% for those with depth of invasion of less than 1.5 mm (P = .009). No other variable, including age, sex, tobacco use, alcohol use, the presence of carcinoma-in-situ, histologic grade, lymphocytic infiltration, or muscularis mucosae invasion, was associated with cancer progression. CONCLUSION: The depth of invasion in the TURB specimens, measured with a micrometer, is predictive of cancer progression in patients with stage T1 bladder carcinoma.