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Is There a Role for Fatty Acid Synthase in the Diagnosis of Prostatic Adenocarcinoma?
A Comparison With AMACR

Xiaojun Wu MD, PhD, Majd Zayzafoon MD, PhD, Xinzhi Zhang MD, PhD, Omar Hameed MBChB
DOI: http://dx.doi.org/10.1309/AJCP0Y5QWWYDKCJE 239-246 First published online: 1 August 2011


Our aim was to compare the usefulness of fatty acid synthase (FASn) with that of α-methylacyl coenzyme-A racemase (AMACR) in the diagnosis of prostatic adenocarcinoma. The expression of these 2 markers was compared in a tissue microarray containing 62 foci of benign glands and 36 foci of prostatic adenocarcinoma. Similar to AMACR, there was significantly higher FASn expression in adenocarcinoma compared with that in benign glands. The optimal accuracy rate and area under curve (AUC) by receiver operating characteristic analysis for FASn were not significantly different from those for AMACR (accuracy, 80% vs 87%; AUC, 0.942 vs 0.956; P for both, > .05). Moreover, in cases with coexistent malignant and benign glands on the same core, FASn could selectively distinguish a proportion of cases (17/21 [81%]) similar to using AMACR. We conclude that FASn may aid in the diagnosis of prostatic adenocarcinoma, at least to supplement AMACR as another positive marker of carcinoma and potentially increase diagnostic accuracy.

Key Words:
  • α-Methylacyl coenzyme-A racemase
  • Fatty acid synthase
  • Prostate
  • Prostatic adenocarcinoma
  • Immunohistochemistry

Prostatic adenocarcinoma is the most common noncutaneous malignancy and the second most common cause of cancer death in men.1,2 It is estimated that about 1 in 6 men in the United States will be diagnosed with prostate cancer during their lifetime, and 1 in 36 will die of this disease. It was projected that in 2010, 217,730 new cases of prostate cancer would be diagnosed in the United States.3

Histologic examination is the “gold standard” for diagnosing prostatic adenocarcinoma. Although this examination is mostly based on routine H&E-stained sections, it is often supplemented by immunohistochemical staining. One immunostain that is frequently used in this situation is α-methylacyl coenzyme-A racemase (AMACR) that, as a “positive” marker of malignancy, is of great value in confirming the presence of adenocarcinoma.35 However, AMACR is not 100% sensitive, and its expression is not limited to prostatic adenocarcinoma but may also be seen in several of its histologic mimics,6,7 resulting in many potential caveats in its use.813 Accordingly, evaluation of potential new markers of prostatic adenocarcinoma is needed.

Fatty acid synthase (FASn), a regulator of the de novo synthesis of long-chain fatty acids, has been shown to have elevated expression levels in many human carcinomas.1417 In the prostate, FASn has been reported to be up-regulated in prostatic intraepithelial neoplasia and prostatic adenocarcinoma.16,1821 These findings suggest that FASn could potentially be useful as a diagnostic marker for prostatic adenocarcinoma; however, to our knowledge, there are only 2 studies22,23 that specifically examine its diagnostic usefulness, including 1 available only in abstract form.22

Accordingly, the aim of this study was to evaluate the diagnostic usefulness of FASn in the diagnosis of prostatic adenocarcinoma by specifically comparing it with that of AMACR, the only diagnostically useful positive marker of malignancy currently available.

Materials and Methods

To investigate the effectiveness of FASn compared with AMACR in distinguishing prostatic adenocarcinoma from benign prostatic glands, the immunohistochemical expression of these 2 markers was compared side by side. A tissue microarray containing foci of 62 benign prostatic glands and 36 prostatic adenocarcinomas was constructed with diagnoses confirmed by evaluation of H&E-stained sections and immunohistochemical studies using a p63/34βE12/AMACR antibody cocktail. By using an automated immunostainer (Autostainer 720, Labvision, Fremont, CA) and following heat-induced (98°C) epitope retrieval with a 0.02-mol/L concentration of citrate buffer (pH 6.0) for 20 minutes, we immunostained newly cut tissue microarray sections with antibodies (30-minute incubation at room temperature) directed at AMACR (clone 13H4, dilution 1:50; DAKO, Carpinteria, CA) and FASn (clone 23, dilution 1:3,200; BD Biosciences, San Diego, CA). The UltraVision LP polymer system (Labvision) and the chromogen diaminobenzidine tetrachloride were used to amplify and visualize the antibody-antigen complex. The tissues were then counterstained with hematoxylin. The newly stained sections were reviewed to determine the intensity of AMACR and FASn expression (0–3+) and percentage of positive cells (0%-100%) at each intensity level; these were then used to calculate an H score (0–300) as previously described.24

The expression levels of these 2 markers were compared using the paired t test and the χ2 test as statistically appropriate. In addition, their discriminatory powers were compared by receiver operating characteristic (ROC) analysis; by calculating the sensitivity, specificity, and accuracy rates of these 2 markers at different intensity cutoffs; and by studying cases with coexistent benign prostatic glands and prostatic adenocarcinoma on the same core. Statistical analyses were performed by using STATA (version 10, StataCorp, College Station, TX). A P value of less than .05 was considered statistically significant.

View this table:
Table 1

The study was approved by the University of Alabama at Birmingham Institutional Review Board.


Expression of FASn and AMACR in Benign and Malignant Prostatic Glands

In comparing the expression of FASn with that of AMACR in prostatic adenocarcinoma, we found a higher percentage of positive cells, intensity of expression, and H score with FASn; however, only the H score was significantly different Table 1. A representative focus of prostatic adenocarcinoma with AMACR and FASn immunostains is shown in Image 1.

These findings were replicated in benign prostatic glands, in which there was also a higher percentage of positive cells, intensity of expression, and H score with FASn, with all differences being statistically significant Table 2. A representative focus of benign prostatic glands with AMACR and FASn immunostains is shown in Image 2.

Diagnostic Usefulness of FASn and AMACR for the Diagnosis of Prostatic Adenocarcinoma

Similar to AMACR, FASn showed higher expression in foci of prostatic adenocarcinoma than that in foci of benign prostatic glands, as evidenced by higher averages of intensities, percentages of positive cells, and H scores Figure 1. This differential expression of FASn suggested that FASn has potential clinical usefulness for distinguishing prostatic adenocarcinoma from benign prostatic glands. Furthermore, the ROC analysis of the H score Figure 2 showed that the area under the curve of AMACR was not significantly different from that of FASn (0.9558 vs 0.9423; P = .71).

A comparison of the sensitivity, specificity, and accuracy rates of both antibodies for the diagnosis of prostatic adenocarcinoma at different intensity cutoffs (at least 10% cells positive at cutoff intensity) showed that a stronger intensity cutoff (3+) was needed for FASn to achieve the highest rates Table 3. Given the overlapping 95% confidence intervals, however, the differences in the sensitivity, specificity, and accuracy rates of FASn and AMACR were not statistically significant.

Image 1

α-Methylacyl coenzyme-A racemase (AMACR) and fatty acid synthase (FASn) in prostatic adenocarcinoma. A focus of packed malignant glands (A, H&E, ×200) with circumferential, predominantly luminal, 1+ to 2+ granular cytoplasmic AMACR expression (B, ×200) and diffuse 3+ cytoplasmic FASn expression (C, ×200).

Discriminatory Power of FASn and AMACR in Distinguishing Malignant Glands From Adjacent Benign Glands

To better assess the discriminatory power of FASn in comparison with AMACR, cases with coexistent benign and malignant prostatic glands on the same core were evaluated. A total of 16 such cases on the AMACR and 21 on the FASn tissue microarray slide were identified. Only when there was higher intensity of expression in malignant glands compared with benign was the immunostain considered to be discriminatory. Based on that, FASn could distinguish carcinoma in 17 (81%) of 21 cases compared with 13 (81%) of 16 cases by AMACR Image 3. Of the 4 cases that were “missed” by FASn, 2 were labeled by AMACR, whereas 1 of the 3 cases missed by AMACR was labeled by FASn Image 4.

View this table:
Table 2


In this study, we found that the expression levels of FASn in prostatic adenocarcinoma were significantly different from the levels in benign glands and could potentially be used as an aid in the diagnosis. Although FASn did not seem to be superior to AMACR, the findings suggested that FASn could potentially supplement AMACR as another positive marker of malignancy. Additional studies are warranted to further investigate this possibility.

Image 2

α-Methylacyl coenzyme-A racemase (AMACR) and fatty acid synthase (FASn) in benign prostatic glands. A focus of unremarkable benign prostatic glands (A, H&E, ×200) that lack AMACR expression (B, ×200) display 1+ to 2+ diffuse cytoplasmic FASn expression (C, ×200).

Prostatic adenocarcinoma is mostly diagnosed by H&E evaluation; however, immunohistochemical staining has become indispensable in the evaluation of challenging cases.9 Although several markers are available to demonstrate the lack of basal cells in malignant glands and confirm the diagnosis in difficult cases,2528 AMACR is currently the only positive marker of adenocarcinoma.9,29 Based on this, AMACR has become an indispensable tool in diagnostic urologic pathology. There are, however, several inherent downsides associated with the use of AMACR. First, AMACR expression is not always present in prostatic adenocarcinoma and may also be seen in benign prostatic tissue, thus decreasing its sensitivity and specificity, respectively. Second, the expression of AMACR is not limited to prostatic tissue, and expression in some nonprostatic mimickers of prostatic adenocarcinoma, such as nephrogenic adenoma, is a potential diagnostic pitfall.7 Last but not the least, the occasional heterogeneous pattern of AMACR expression in prostatic adenocarcinoma can sometimes make it difficult to interpret.4 Accordingly, identification of additional markers of malignancy would be desirable to further increase diagnostic accuracy when AMACR is not beneficial.

In this study, we have clearly shown that FASn is one such potential marker. Although there was more FASn than AMACR expression in benign prostatic glands, FASn expression in prostatic adenocarcinoma was sufficiently distinct from that in benign glands to potentially aid in the diagnosis. In fact, the discriminatory power of FASn was very similar to that of AMACR as demonstrated by comparable areas under the curve in ROC analysis of the H score and comparable sensitivity, specificity, and accuracy rates calculated at optimal cutoff intensities. It is interesting that the sensitivity and specificity of FASn for the diagnosis of adenocarcinoma reported herein were lower than those reported by Zhang et al.22 Although a more detailed comparison cannot be made because of the limited nature of their report (abstract only), these differences clearly suggest that additional studies evaluating the diagnostic usefulness of FASn for the diagnosis of prostatic adenocarcinoma are warranted.

Figure 1

Side-by-side comparison of α-methylacyl coenzyme-A racemase (AMACR) and fatty acid synthase (FASn) expression in benign (black bars) and malignant (white bars) glands. Similar to AMACR, FASn immunostaining showed higher averages of intensities (A), percentages of positive cells (B), and H scores (C) in prostatic adenocarcinoma than in benign prostatic glands. Data are given as mean with standard error bar (*P < .05, benign vs malignant glands).

Figure 2

Receiver operating characteristic curves for α-methylacyl coenzyme-A racemase (AMACR) and fatty acid synthase (FASn) H scores. The area under curve for FASn (0.9423) is comparable to that for AMACR (0.9558).

View this table:
Table 3
Image 3

A focus of adenocarcinoma adjacent to benign glands (A, H&E, ×200) with strong expression of α-methylacyl coenzyme-A racemase (AMACR; B, ×200) and fatty acid synthase (FASn; C, ×200). Note that the benign glands are completely negative for AMACR but express FASn.

At this point, it should be noted that the diagnostic usefulness of FASn was comparable to that of AMACR only when the FASn antibody was optimally titrated. In fact, we repeated the entire study using lower (1:1,600) and higher (1:6,400) FASn dilutions than the one (1:3,200) presented herein, both of which resulted in a significantly less discriminatory power (data not shown). This should come as no surprise because it is well known that alteration of this, as well as other immunohistochemical variables (eg, antigen retrieval, incubation times, and detection methods) can clearly lead to significantly different results in research settings and clinical practice.

A potential source of variability in our study (and many others using tissue microarrays) was the fact that the tissue microarray was constructed from different cases with potentially different specimen handling procedures that might have affected warm ischemia time, time to fixation, and fixation time. Because all of these factors have the potential to affect immunohistochemical findings, we wanted to minimize this potential bias by performing a subset analysis on cases with coexistent malignant and benign glands on the same tissue microarray core (which is also how these antibodies would be used in clinical practice). This analysis showed that an identical proportion of cases (81%) were identified by FASn and AMACR. Moreover, FASn was able to identify a case of adenocarcinoma that was not labeled by AMACR. The latter finding is somewhat similar to the only other study that we are aware of comparing AMACR with FASn expression in the prostate gland.23 In this recent study, the discriminatory power of FASn was not analyzed by ROC curves or by comparing its sensitivity, specificity, or accuracy rates with those of AMACR. Nevertheless, the authors showed that FASn expression was seen in 19 of 21 cases of AMACR– prostatic adenocarcinoma, including 14 cases with 2+ to 3+ FASn intensity. This latter finding (which was present in 1 of the 3 AMACR– adenocarcinomas in our study) suggests that FASn immunohistochemical staining, if nothing else, could at least supplement AMACR as a positive marker for prostatic adenocarcinoma.

Image 4

A focus of adenocarcinoma (A, H&E, ×100) that completely lacked α-methylacyl coenzyme-A racemase expression (B, ×100) showed 1+ to 2+ fatty acid synthase expression (C, ×100) that contrasts with the predominantly negative adjacent benign glands.

We have found that FASn, despite significant expression in benign prostatic glands, could still potentially be used as an aid in the diagnosis of prostatic adenocarcinoma, especially if used to complement AMACR. As is the case with any other immunostain, choosing the correct dilution was crucial for a satisfactory performance. Additional, more comprehensive investigations using more samples, needle biopsy and resection specimens, and examples of benign histologic mimickers of adenocarcinoma are needed to further characterize and validate the diagnostic usefulness of this marker.


Upon completion of this activity you will be able to:

  • recognize the role of immunohistochemistry in the diagnosis of prostatic adenocarcinoma.

  • discuss the main pros and cons of commonly used immunostains in the evaluation of prostate needle biopsy material.

  • compare the diagnostic utilities and limitations of α-methylacyl coenzyme-A racemase (AMACR) and fatty acid synthase (FASn) immunostains for the diagnosis of prostatic adenocarcinoma.

The ASCP is accredited by the Accreditation Council for Continuing Medical Education to provide continuing medical education for physicians. The ASCP designates this journal-based CME activity for a maximum of 1 AMA PRA Category 1 Credit ™ per article. Physicians should claim only the credit commensurate with the extent of their participation in the activity. This activity qualifies as an American Board of Pathology Maintenance of Certification Part II Self-Assessment Module.

The authors of this article and the planning committee members and staff have no relevant financial relationships with commercial interests to disclose.

Questions appear on p 330. Exam is located at www.ascp.org/ajcpcme.


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