Characteristics of Prostate Cancers Missed by Biopsies : Evaluation of Cumulative Tumor Volume Missed According to Cancer True Prevalence *

Purpose: To characterize missed prostate tumors and their cumulative volume with various biopsy regimens to determine optimal biopsy schemes. Methods: We performed 6, 12 and 18-core needle biopsies on 165 and 36-core biopsies on 47 autopsy prostates, respectively. The 6-core biopsy included 6 cores from the mid peripheral zone (MPZ), the 12-core biopsy included 6 cores from the MPZ and lateral PZ (LPZ), and the 18-core biopsies included 6 cores from the MPZ, LPZ and central zone (CZ). The 36-core biopsies included 12 cores in each of these 3 areas. We analyzed the sensitivity of biopsies at each site and evaluated the cumulative volume of cancers and tumor foci missed. Results: Whole-mount analysis identified 59 cancers, 110 tumor foci, and a total cumulative tumor volume of 43 cm. The percentage of tumor foci and corresponding cumulative volume missed with 6, 12, 18 and 36-core biopsies were of 79% and 58%, 64% and 48%, 57% and 26%, and 42% and 17%, respectively (p < 0.05). 12-core biopsies from the MPZ and LPZ performed best for clinically significant cancers detection. However, increasing the number of cores over the 6-core biopsy cutoff increased solely the detection of tumor foci < 0.5 cm. Conclusion: Twelve biopsies from the MPZ and LPZ detected most of the clinically significant cancers while missing most of the tumor foci. These missed tumors represented only a small amount of the overall cancer volume.


Introduction
The trend among clinicians has been performed more prostate biopsies to detect more prostate cancers.Sextant biopsies have been all but abandoned, and most studies recommend extended biopsy protocols of 12 cores [1][2][3].Although these efforts have undoubtedly led to increased detection of cancer, they have also led to the over diagnosis of small and well-differentiated tumors, designated as clinically insignificant [4,5].Even undetected, they may not be of immediate threat, and should they be detected, not to be managed with radical and possibly morbid treatment.This issue recently came under increased scrutiny as a consequence of the recent recommendations by the Task Force for Preventive Medicine [6].Notwithstanding the evidence that PSA screening followed by early stage treatment may reduce mortality [7][8][9], they concluded that the use of PSA was harmful because it caused increased morbidity and over treatment of indolent tumors.
While recommendations from the Task Force for Preventive Medicine focused on the adverse effects of PSA screening, they did not address the role of prostate biopsy as a staging tool.Also, we have to be concerned not only with the ability of the biopsy to detect cancer, but also with the potential characteristics of cancers missed.We previously reported cancer detection rates obtained with different ex-vivo biopsy protocols performed on autopsied prostates [10].Based on the cancer true prevalence, we found that 12 biopsies targeting the mid peripheral zone (MPZ) and lateral peripheral zone (LPZ) performed best for cancer detection.In the current study, we performed post hoc sub-analyses of our data to evaluate the performance of sextant, 12-core, 18-core and 36-core biopsies to detect cancer, with emphasis on the characterization of missed tumor foci and their cumulative volume.

Tissue Collection
We prospectively collected 261 consecutive prostate glands from deceased men that were provided by the University Hospital, Syracuse, NY, the Onondaga County Medical Examiner, Syracuse, NY and by the National Disease Research Interchange, Philadelphia, PA.This study was approved by the Institutional Review Board, and the tissue suppliers obtained informed consent from the next of kin.The decedents had no known history of prostate cancer.At autopsy, the entire prostate gland, together with the seminal vesicles, were excised within 24 hours post-mortem by an experienced medical examiner or pathologist, en-block, and placed in 10% neutral buffered formalin.Prostatic tissue was not entirely removed in 49 (20%) of the prostates autopsied; these subjects were excluded, leaving 212 prostate glands available for analysis.

Prostate Biopsy
All biopsies were performed in a manner that mimicked clinical biopsy with a standard 18F spring-loaded biopsy gun.The needle was inserted through the posterior surface of the hand-held gland, and bilateral samples were taken from the apex, mid gland, and base.The first 165 autopsied glands were biopsied using an 18-core biopsy protocol (Figure 1).The first six cores, corresponding to the sextant biopsy protocol, were taken from the mid peripheral zone (MPZ).The next six cores were taken with the needle inserted into the central zone (CZ).The last six cores were taken from the lateral peripheral zone (LPZ).The last 47 autopsied glands were biopsied using a saturation biopsy protocol of 36 cores: the first 18 cores were taken in a similar fashion and 6 additional cores were taken in each of the 3 locations (Figure 1).

Whole Mount Prostate Processing and Histological Evaluation
After the biopsies were taken, the glands were fixed in formalin for at least 72 hours.The glands were cut into 4-mm sections perpendicular to the posterior plane, labelled, embedded in paraffin, and further sectioned to produce 5-μm whole-mount sections that were stained with hematoxylin and eosin.A single pathologist ana- lyzed the biopsies and whole-mount slides in a blinded fashion.The total number of tumor foci and their locations were recorded.An area of carcinoma was considered to be a separate focus if it was separated from the nearest adjacent focus by a low-power field diameter (4.5 mm), as previously described [11].Each tumor focus was graded according to the modified Gleason grading system [12].

Digital Reconstruction and Measurement of Tumor Volume
The surface of each tumor focus was determined by computerized planimetry, using an image analysis program [10,13].Tumor volume was calculated by multiplying each tumor surface by the section thickness (4 mm) and by 1.5 to compensate for tissue shrinkage [14].Tumors were considered clinically insignificant if they were organ-confined (<pT3) with an index tumor volume of less than 0.5 cm 3 and Gleason score 6 or less [4,5].The cumulative tumor volume detected was calculated as the sum of the volumes of each individual tumor foci detected by biopsies.The cumulative cancer volume detected was calculated as the sum of the volumes of each cancer identified by biopsies.

Tumors missed by biopsies were compared with those
Copyright © 2013 SciRes.OJU detected using the student t-test and the χ 2 test for quantitative and qualitative variables, respectively.McNemar and χ 2 tests were used to compare the sensitivity of cancer detection, the number of missed foci and their cumulative volume between 6-, 12-, and 18-and 36-core biopsies.Sensitivity of biopsy results were calculated using as the standard either the presence of any prostate cancer or of clinically significant cancer on whole-mount analysis.All statistical tests were two-sided, and P values less than 0.05 were considered to be statistically significant.Statistical analyses were conducted using MedCalc ® .
The sensitivity of 12-core biopsies for cancer detection was significantly higher than for sextant biopsies alone (p < 0.0009).The sensitivities of biopsies from the MPZ for clinically significant and insignificant cancer were 50% and 15%, respectively, compared with 77% and 33% for those from the MPZ and LPZ combined (p = 0.02 and p = 0.04, respectively).Increasing sampling over the 12-core cut-off decreased significantly the number of tumor foci missed as well as their cumulative volume, without increasing any further cancer detection rate (Figure 2).However, as shown in Figure 3, increased sampling over the 6-core cut-off decreased significantly the number of small tumor foci missed (<0.5 cm 3 ) but not that of large (>0.5 cm 3 ).The number of missed tumor foci with a Gleason score > 6 decreased with additional biopsies, but the difference was not statistically significant (p = 0.2).

Discussion
In the large ongoing debate over prostate cancer screening, urologists mainly focus on cancers detected and their management.These cancers represent however only the tip of the iceberg.Estimation of tumors that are "missed" with particular biopsy regimens, the location of such tumors, and their histological characteristics and significance is therefore of critical importance.Our analysis confirmed that performing more than 12 biopsies did not increase cancer detection any further.Although 64% of tumor foci and more than half (54%) of the cancers present were missed by 12-core biopsies, they corresponded to a small amount of the overall cancer volume.The percentage of cumulative tumor volume missed with 12-core biopsies was high (48%), but a significant number of the missed foci were located in glands diagnosed with cancer thanks to other contiguous or distant tumor foci detected by biopsies.As a result, only 14% of the overall cancer volume was truly missed with 12-core biopsies.Moreover, our results suggested that missed foci were significantly smaller than those detected and that they had a lower Gleason score.These findings are in accordance with earlier published studies suggesting that 12 cores directed to the peripheral zone of the prostate seem to have a high performance while minimizing the number of unnecessary cores [1][2][3].Biopsies within the central zone have a low likelihood of detecting cancer in the absence of positive results from the peripheral zone of the gland [1,15].
Once having recognized that 12 biopsies from the MPZ and LPZ performed best and detected most of the significant cancers, the next question would concern the need for additional biopsy sampling for cancer characterization.The potential benefit of additional biopsies would have to be balanced against the increased morbidity of the procedure.Biopsy-related morbidity is also a major issue highlighted by the Task Force for Preventive Medicine [6] and has increased these past 10 years [16].
The rate of hospital admission within 30 days of having prostate biopsies was reported to be as high as 4%, mainly for infection-related reasons [16].In our analysis, 18 and 36-core biopsies allowed a significant increase in tumor foci detection and in their cumulative volume, improving therefore staging for each individual patient.However, this increased detection yielded only tumor foci < 0.5 cm 3 .Additionally, the number of missed poorly differentiated tumor foci (Gleason score > 6) decreased with additional sampling, but the difference was not significant (p = 0.2).These findings support the concept that cancers are not missed by biopsies because of suboptimal technique, but rather because they are smaller and earlier in their development.In clinical practice, these cancers may be detected later with repeated biopsies [17].Knowing the existence and characteristics of such secondary foci is of low value if a radical treatment is immediately intended.Conversely, to develop and increase the possibilities of active surveillance and focal therapies, performing an initial accurate cancer mapping of the prostate may be of crucial importance, especially when 12-core biopsies reveal only small amount of low grade cancer.This initial staging could then serve as a reference for potential re-biopsy strategies.
Our study has several limitations, the most important being the absence of pre-mortem PSA data available.PSA screening is indeed the first step before considering prostate biopsies.Another limitation is our "histological" definition of clinical significance, which did not take into consideration age, comorbidities, and other individual circumstances.

Conclusion
Twelve biopsies from the MPZ and LPZ detected most of  the clinically significant cancers while missing as much as 48% of cancers.These missed cancers represented however only 13% of the overall volume of cancer.

Figure 1 .
Figure 1.Schema of the anatomic sites of prostate biopsies taken from the 212 men postmortem.The first six biopsy cores (1-6) were taken from the mid peripheral zone (MPZ), the next six cores (7-12) from the central zone (CZ), and the last six cores (13-18) from the lateral peripheral zone (LPZ).In the last 47 glands, 6 additional biopsy cores were taken in each of these 3 mentioned areas.

Figure 2 .Figure 3 .
Figure 2. Rates of missed cancers and individual tumor foci, and corresponding volumes, according to biopsy protocol.