Stefan Steurer

Neurovascular Structure-adjacent Frozen-section Examination (NeuroSAFE) Increases Nerve-sparing Frequency and Reduces Positive Surgical Margins in Open and Robot-assisted Laparoscopic Radical Prostatectomy: Experience After 11 069 Consecutive Patients

BACKGROUND Intraoperative frozen-section analysis allows real-time histologic assessment of surgical margins (SMs) and identification of candidates for nerve-sparing (NS) procedures. OBJECTIVE To examine the efficacy and oncologic safety of a systematic neurovascular structure-adjacent frozen-section examination (NeuroSAFE) during NS radical prostatectomy (RP). DESIGN, SETTING, AND PARTICIPANTS From January 2002 to June 2011, 11 069 consecutive RPs were performed at the University Medical Center Hamburg-Eppendorf. Of these, 5392 (49%) were conducted with NeuroSAFE. SURGICAL PROCEDURE Our NeuroSAFE approach included the whole laterorectal circumference of the prostate to determine the SM status of the complete neurovascular tissue-corresponding prostatic surface. OUTCOME MEASUREMENTS AND STATISTICAL ANALYSIS The impact of NeuroSAFE on NS frequency, SM status, and biochemical recurrence (BCR) was analyzed by chi-square test, and by Kaplan-Meier analyses in propensity score-based matched cohorts. RESULTS AND LIMITATIONS Positive SMs (PSMs) were detected in 1368 (25%) NeuroSAFE RPs, leading to a secondary resection of the ipsilateral neurovascular tissue. Secondary wide resection resulted in conversion to a definitive negative SM (NSM) status in 1180 (86%) patients. In NeuroSAFE RPs, frequency of NS was significantly higher (all stages: 97% vs 81%; pT2: 99% vs 92%; pT3a: 94% vs 72%; pT3b: 88% vs 40%; p<0.0001) and PSM rates were significantly lower (all stages: 15% vs 22%; pT2: 7% vs 12%; pT3a: 21% vs 32%; p<0.0001) than in the matched non-NeuroSAFE RPs. In propensity score-based comparisons, NeuroSAFE had no negative impact on BCR (pT2, p=0.06; pT3a, p=0.17, pT3b, p=0.99), and BCR-free survival of patients with conversion to NSM did not differ significantly from patients with primarily NSM (pT2, p=0.16; pT3, p=0.26). The accuracy of our NeuroSAFE approach was 97% with a false-negative rate of 2.5%. The major limitations of this study are its retrospective nature and relatively short follow-up. CONCLUSIONS NeuroSAFE enables real-time histologic monitoring of the oncologic safety of a NS procedure. Systematic NeuroSAFE significantly increases NS frequencies and reduces PSMs. Patients with a NeuroSAFE-detected PSM could be converted to a prognostically more favorable NSM status by secondary wide resection.

Genomic deletion of MAP3K7 at 6q12-22 is associated with early PSA recurrence in prostate cancer and absence of TMPRSS2: ERG fusions

6q12-22 is the second most commonly deleted genomic region in prostate cancer. Mapping studies have described a minimally deleted area at 6q15, containing MAP3K7/TAK1, which was recently shown to have tumor suppressive properties. To determine prevalence and clinical significance of MAP3K7 alterations in prostate cancer, a tissue microarray containing 4699 prostate cancer samples was analyzed by fluorescence in situ hybridization. Heterozygous MAP3K7 deletions were found in 18.48% of 2289 interpretable prostate cancers. MAP3K7 deletions were significantly associated with advanced tumor stage (P<0.0001), high Gleason grade (P<0.0001), lymph node metastasis (P<0.0108) and early biochemical recurrence (P<0.0001). MAP3K7 alterations were typically limited to the loss of one allele as homozygous deletions were virtually absent and sequencing analyses revealed no evidence for MAP3K7 mutations in 15 deleted and in 14 non-deleted cancers. There was a striking inverse association of MAP3K7 deletions and TMPRSS2:ERG fusion status with 26.7% 6q deletions in 1125 ERG-negative and 11.1% 6q deletions in 1198 ERG-positive cancers (P<0.0001). However, the strong prognostic role of 6q deletions was retained in both ERG-positive and ERG-negative cancers (P<0.0001 each). In summary, our study identifies MAP3K7 deletion as a prominent feature in ERG-negative prostate cancer with strong association to tumor aggressiveness. MAP3K7 alterations are typically limited to one allele of the gene. Together with the demonstrated tumor suppressive function in cell line experiments and lacking evidence for inactivation through hypermethylation, these results indicate MAP3K7 as a gene for which haploinsufficency is substantially tumorigenic.

Recurrent deletion of 3p13 targets multiple tumour suppressor genes and defines a distinct subgroup of aggressive ERG fusion-positive prostate cancers

Deletion of 3p13 has been reported from about 20% of prostate cancers. The clinical significance of this alteration and the tumour suppressor gene(s) driving the deletion remain to be identified. We have mapped the 3p13 deletion locus using SNP array analysis and performed fluorescence in situ hybridization (FISH) analysis to search for associations between 3p13 deletion, prostate cancer phenotype and patient prognosis in a tissue microarray containing more than 3200 prostate cancers. SNP array analysis of 72 prostate cancers revealed a small deletion at 3p13 in 14 (19%) of the tumours, including the putative tumour suppressors FOXP1, RYBP and SHQ1. FISH analysis using FOXP1‐specific probes revealed deletions in 16.5% and translocations in 1.2% of 1828 interpretable cancers. 3p13 deletions were linked to adverse features of prostate cancer, including advanced stage (p < 0.0001), high Gleason grade (p = 0.0125), and early PSA recurrence (p = 0.0015). In addition, 3p13 deletions were linked to ERG+ cancers and to PTEN deletions (p < 0.0001 each). A subset analysis of ERG+ tumours revealed that 3p13 deletions occurred independently from PTEN deletions (p = 0.3126), identifying tumours with 3p13 deletion as a distinct molecular subset of ERG+ cancers. mRNA expression analysis confirmed that all 3p13 genes were down regulated by the deletion. Ectopic over‐expression of FOXP1, RYBP and SHQ1 resulted in decreased colony‐formation capabilities, corroborating a tumour suppressor function for all three genes. In summary, our data show that deletion of 3p13 defines a distinct and aggressive molecular subset of ERG+ prostate cancers, which is possibly driven by inactivation of multiple tumour suppressors. Copyright

Clinical Utility of Quantitative Gleason Grading in Prostate Biopsies and Prostatectomy Specimens

BACKGROUND Gleason grading is the strongest prognostic parameter in prostate cancer. Gleason grading is categorized as Gleason ≤ 6, 3 + 4, 4 + 3, 8, and 9-10, but there is variability within these subgroups. For example, Gleason 4 components may range from 5-45% in a Gleason 3 + 4 = 7 cancer. OBJECTIVE To assess the clinical relevance of the fractions of Gleason patterns. DESIGN, SETTING, AND PARTICIPANTS Prostatectomy specimens from 12823 consecutive patients and of 2971 matched preoperative biopsies for which clinical data with an annual follow-up between 2005 and 2014 were available from the Martini-Klinik database. OUTCOME MEASUREMENTS AND STATISTICAL ANALYSIS To evaluate the utility of quantitative grading, the fraction of Gleason 3, 4, and 5 patterns seen in biopsies and prostatectomies were recorded. Gleason grade fractions were compared with prostatectomy findings and prostate-specific antigen recurrence. RESULTS AND LIMITATIONS Our data suggest a striking utility of quantitative Gleason grading. In prostatectomy specimens, there was a continuous increase of the risk of prostate-specific antigen recurrence with increasing percentage of Gleason 4 fractions with remarkably small differences in outcome at clinically important thresholds (0% vs 5%; 40% vs 60% Gleason 4), distinguishing traditionally established prognostic groups. Also, in biopsies, the quantitative Gleason scoring identified various intermediate risk groups with respect to Gleason findings in corresponding prostatectomies. Quantitative grading may also reduce the clinical impact of interobserver variability because borderline findings such as tumors with 5%, 40%, or 60% Gleason 4 fractions and very small Gleason 5 fractions (with pivotal impact on the Gleason score) are disclaimed. CONCLUSIONS Quantitative Gleason pattern data should routinely be provided in addition to Gleason score categories, both in biopsies and in prostatectomy specimens. PATIENT SUMMARY Gleason score is the most important prognostic parameter in prostate cancer, but prone to interobserver variation. The results of our study show that morphological aspects that define the Gleason grade in prostate cancer represent a continuum. Quantitation of Gleason patterns provides clinically relevant information beyond the traditional Gleason grading categories ≤ 3 + 3, 3 + 4, 4 + 3, 8, 9 -1 0. Quantitative Gleason scoring can help to minimize variations between different pathologists and substantially aid in optimized therapy decision-making.

MALDI mass spectrometric imaging based identification of clinically relevant signals in prostate cancer using large-scale tissue microarrays.

To identify molecular features associated with clinico‐pathological parameters and TMPRSS2‐ERG fusion status in prostate cancer, we employed MALDI mass spectrometric imaging (MSI) to a prostate cancer tissue microarray (TMA) containing formalin‐fixed, paraffin‐embedded tissues samples from 1,044 patients for which clinical follow‐up data were available. MSI analysis revealed 15 distinct mass per charge (m/z)‐signals associated to epithelial structures. A comparison of these signals with clinico‐pathological features revealed statistical association with favorable tumor phenotype such as low Gleason grade, early pT stage or low Ki67 labeling Index (LI) for four signals (m/z 700, m/z 1,502, m/z 1,199 and m/z 3,577), a link between high Ki67LI for one signal (m/z 1,013) and a relationship with prolonged time to PSA recurrence for one signal (m/z 1,502; p = 0.0145). Multiple signals were associated with the ERG‐fusion status of our cancers. Two of 15 epithelium‐associated signals including m/z 1,013 and m/z 1,502 were associated with detectable ERG expression and five signals (m/z 644, 678, 1,044, 3,086 and 3,577) were associated with ERG negativity. These observations are in line with substantial molecular differences between fusion‐type and non‐fusion type prostate cancer. The signals observed in this study may characterize molecules that play a role in the development of TMPRSS2‐ERG fusions, or alternatively reflect pathways that are activated as a consequence of ERG‐activation. The combination of MSI and large‐scale TMAs reflects a powerful approach enabling immediate prioritization of MSI signals based on associations with clinico‐pathological and molecular data.

A pathogenic role for T cell–derived IL-22BP in inflammatory bowel disease

Interleukin-22 binding protein (IL-22BP) drives inflammatory bowel disease by sopping up the tissue-protective protein IL-22. Intestinal inflammation can impair mucosal healing, thereby establishing a vicious cycle leading to chronic inflammatory bowel disease (IBD). However, the signaling networks driving chronic inflammation remain unclear. Here we report that CD4+ T cells isolated from patients with IBD produce high levels of interleukin-22 binding protein (IL-22BP), the endogenous inhibitor of the tissue-protective cytokine IL-22. Using mouse models, we demonstrate that IBD development requires T cell–derived IL-22BP. Lastly, intestinal CD4+ T cells isolated from IBD patients responsive to treatment with antibodies against tumor necrosis factor–α (anti–TNF-α), the most effective known IBD therapy, exhibited reduced amounts of IL-22BP expression but still expressed IL-22. Our findings suggest that anti–TNF-α therapy may act at least in part by suppressing IL-22BP and point toward a more specific potential therapy for IBD.

SPINK1 expression is tightly linked to 6q15- and 5q21-deleted ERG-fusion negative prostate cancers but unrelated to PSA recurrence.

The serine peptidase inhibitor, Kazal type 1 (SPINK1) has been suggested to define an aggressive molecular subtype of ERG‐fusion negative prostate cancer. It was the aim of this study to further study the clinical relevance of SPINK1 expression and its relationship with other key genomic alterations of prostate cancer.

TMPRSS2-ERG Fusions Are Strongly Linked to Young Patient Age in Low-grade Prostate Cancer

Based on next-generation sequencing of early-onset prostate cancer (PCa), we earlier demonstrated that PCa in young patients is prone to rearrangements involving androgen-regulated genes-such as transmembrane protease, serine 2 (TMPRSS2)-v-ets avian erythroblastosis virus E26 oncogene homolog (ERG) fusion-and provided data suggesting that this situation might be caused by increased androgen signaling in younger men. In the same study, an accumulation of chromosomal deletions was found in cancers of elderly patients. To determine how age-dependent molecular features relate to cancer phenotype, an existing data set of 11,152 PCas was expanded by additional fluorescence in situ hybridization analyses of phosphatase and tensin homolog (PTEN), 6q15 and 5q21. The results demonstrate that the decrease in TMPRSS2-ERG fusions with increasing patient age is limited to low-grade cancers (Gleason ≤3+4) and that the significant increase in the deletion frequency with age was strictly limited to ERG-negative cancers for 6q15 and 5q21 but to ERG-positive cancers for PTEN. These data suggest that the accumulation of non-androgen-linked genomic alterations with advanced patient age may require an appropriate microenvironment, such as a positive or negative ERG status. The strong link of ERG activation to young patient age and low-grade cancers may help to explain a slight predominance of low-grade cancers in young patients.

Cysteine-rich secretory protein 3 overexpression is linked to a subset of PTEN-deleted ERG fusion-positive prostate cancers with early biochemical recurrence.

The aim of this study was to determine whether cysteine-rich secretory protein 3 (CRISP3) expression is linked to clinically or molecularly relevant subgroups of prostate cancer. A tissue microarray representing samples from >10 000 prostate cancers from radical prostatectomy specimens with clinical follow-up data were analyzed for CRISP3 expression by immunohistochemistry. CRISP3 expression was also compared with key genomic alterations of prostate cancer. CRISP3 staining was found as weak in 15%, moderate in 8.5%, and strong in 7.2% of prostate cancers, whereas no expression was detected in normal prostate. Strong CRISP3 expression was linked to advanced tumor stage, high Gleason score, and positive surgical margin status (P<0.0001 each). There was a marked accumulation of high CRISP3 expression in PTEN-deleted ERG-positive tumors (P<0.0001). A total of, 21.7% of ERG-positive and PTEN-deleted cancers had strong CRISP3 expression, but only 10.4% of ERG-positive cancers without PTEN deletion (P<0.0001). The rate of high CRISP3 expression was 2.5% in ERG-negative cancers (P=0.0001; vs ERG-positive cancers). Accordingly, CRISP3 overexpression was associated with early prostate-specific antigen recurrence in all tumors (P=0.0013) as well as in ERG-negative (P=0.004) and ERG-positive cancers (P=0.0318). CRISP3 expression did not retain prognostic significance in models also involving PTEN deletions. Strong CRISP3 expression is associated with unfavorable tumor phenotype and early recurrence in prostate cancers. The tight link of strong CRISP3 expression to the ERG fusion-positive prostate cancers with PTEN deletions provides further evidence for the existence of molecularly distinct subgroups of prostate cancers.

A tertiary Gleason pattern in the prostatectomy specimen and its association with adverse outcome after radical prostatectomy.

PURPOSE The prognostic significance of a tertiary Gleason pattern in the radical prostatectomy specimen is controversial. We tested the impact of a tertiary Gleason pattern on adverse histopathological features and biochemical recurrence rates after radical prostatectomy. MATERIALS AND METHODS We assessed data on 11,226 consecutive patients treated with radical prostatectomy at our institution between June 2007 and February 2013. We compared 2,396 patients with (22.4%) and 8,260 without (77.5%) a tertiary Gleason pattern for adverse histopathological features (extraprostatic extension, seminal vesicle invasion, positive surgical margins and lymph node invasion) using the chi-square test. The effect of a tertiary Gleason pattern on biochemical recurrence was tested in univariable and multivariable models. Subanalyses were then done for different radical prostatectomy Gleason groups (6 or less, 3 + 4 and 4 + 3). RESULTS A tertiary Gleason pattern was statistically significantly associated with all evaluated histopathological parameters (each p <0.001). It was an independent predictor of biochemical recurrence (HR 1.43, p <0.001). On subanalysis only a tertiary Gleason pattern independently predicted biochemical recurrence in the patient cohort with a radical prostatectomy Gleason score of 3 + 4 and 4 + 3. However, it failed to attain independent predictor status in patients with a radical prostatectomy Gleason score of 6 or less. CONCLUSIONS A tertiary Gleason pattern is a significant and independent predictor of biochemical recurrence after radical prostatectomy with the strongest prognostic effect in cases with Gleason scores 3 + 4 and 4 + 3. Therefore, a tertiary Gleason pattern should be recorded in the pathological report.