Studies of NKX3.1
mRNA and protein expression in human prostate cancer and prostatic intraepithelial neoplasia (PIN) have provided somewhat contradictory results. Xu et al43
reported that in prostatic adenocarcinomas NKX3.1
mRNA was overexpressed in 31%, decreased in 21% and was similar to normal epithelium in 48% of cases. Also, a higher fraction of tumor samples showed NKX3.1
mRNA overexpression in nonorgan confined tumors (40%) versus organ confined disease (22%). In contrast, Ornstein et al did not find a change in NKX3.1
mRNA levels by quantitative in situ hybridization in prostatic adenocarcinomas compared with normal prostate in their study of early-stage prostate cancers.30
In fact, these researchers suggested that as NKX3.1
was expressed nearly exclusively in the prostate in adult tissues, it could prove to be a useful marker of malignant prostate epithelium.
Bowen et al, reported that loss of NKX3.1 protein expression, as assessed by immunohistochemistry (IHC), correlated with prostate cancer progression9
; specifically, they reported complete loss of NKX3.1 staining in 20% of high-grade PIN, 6% of stage T1a/b samples, 22% of stage T3/4 samples, 34% of hormone-refractory prostate cancers, and 78% of metastases. By contrast, Korkmaz et al24
conducted in situ hybridization for mRNA expression and IHC for protein staining on adjacent TMA slides and reported that a vast majority of all prostatic adenocarcinoma cases were positive for both the mRNA and protein and that there was no correlation between NKX3.1
mRNA or protein expression and tumor grade or clinical stage. Gelmann et al,16
reported that NKX3.1 protein was present in prostatic adenocarcinomas by IHC in 66% of primary untreated tumors, 44% of untreated metastatic tumors, and 27.3% of castrate resistant/hormone refractory tumors. Asatiani et al showed that although there was reduced intensity of staining for NKX3.1 protein, quantified by image analysis, complete loss in NKX3.1 protein was observed in only 4.6% of primary tumor samples.5
Bethel et al observed reduced levels of NKX3.1 protein in PIN and in primary prostatic adenocarcinomas, although virtually all cases retained moderate to high amounts of NKX3.1 staining.6
Using the same rabbit polyclonal antibody as used by Bethel et al6
(and this study), Chuang et al11
recently reported that NKX3.1 protein staining was a highly specific and relatively sensitive marker when used as a diagnostic aid as part of a panel of IHC markers in helping to distinguish high-grade prostate carcinoma from high-grade bladder urothelial carcinoma. However, the study by Chuang et al11
did not systematically examine the expression of NKX3.1 in metastatic prostatic adenocarcinomas, nor did it determine the specificity for prostate carcinoma beyond examination of urinary bladder cancers.
Although there is some controversy, the prevailing model of NKX3.1 protein expression in human prostatic adenocarcinoma is that the levels are reduced in primary prostate cancers and further reduced and often lost in metastatic lesions.2
Contrary to this view, we found that NKX3.1 expression was retained in metastatic prostatic adenocarcinomas in the vast majority of cases. In fact, in some distant metastases, NKX3.1 staining was higher in both staining intensity and the percentage of tissue stained compared with PSA. The genetic data are consistent with retained expression of NKX3.1 in most cases—whereas loss of one allele of the NKX3.1
gene is common, no mutations have been described in the remaining NKX3.1
allele. It is highly likely that some of the differences found to date in the literature regarding the presence of retained NKX3.1 protein in prostatic adenocarcinomas are related to the performance of the different antibodies that have been employed. The antibody used in this study is a relatively newly developed rabbit polyclonal antibody raised against a recombinant polypeptide corresponding to the NH2-terminal 123 amino acids of human NKX3.1,6
and we found this antibody to be highly specific and to perform well by IHC.6,11
This antibody is currently undergoing licensing by a commercial vendor such that it should be available commercially (Athena Environmental Sciences Inc.) by the time this manuscript is published.
Immunohistochemical studies have been employed in two main areas in which establishing the prostatic origin of a neoplasm is imperative: transurethral resection or biopsy specimens, in which the most common scenario is the need to distinguish poorly differentiated high-grade primary prostatic adenocarcinoma from high-grade urothelial carcinoma11,31,39
and metastatic adenocarcinomas of unknown origin, in which the differential diagnosis is much wider.39
When considering metastatic malignant neoplasms of unknown origin, PSA and PSAP are the most commonly employed markers in identifying prostatic carcinomas. However, despite their high selectivity for prostate tissue, PSA and PSAP have been known to be expressed at a significantly lesser extent in metastatic prostate carcinomas, and may at times be expressed at least somewhat in a number of non-prostate tumors.39
The most well-studied examples of this are PSA expression in breast carcinomas and salivary gland neoplasms and PSAP staining in carcinoid tumors.39
Nevertheless, although PSA and PSAP positivity should be interpreted somewhat cautiously when examining tumor specimens of unknown primary, and certainly in the context of histopathologic and clinical findings, most pathologists consider these markers to be relatively sensitive, specific and useful for distinguishing prostatic origin in the context of metastatic lesions. In this study, we found NKX3.1 staining to be highly specific for prostate epithelium as it was positive in only 1 of 383 nonprostatic tumors, and this tumor was a lobular carcinoma of the breast, which does not enter into the differential diagnosis when dealing with prostatic adenocarcinoma except in very rare cases of potential metastatic breast carcinoma in males. Another relatively recently discovered protein, P501S (also known as prostein), that is selectively expressed in prostatic epithelium, has also been reported to be effective in identifying prostatic origin in metastatic carcinomas.44
In addition, a earlier study employing the metastatic prostate carcinoma TMA used in this study, found 68 of the 69 metastatic prostate carcinoma cases and 15 of the 15 distant metastasis cases to be positive for p501s.34
Like PSA and PSAP, however, P501s staining is exclusively cytoplasmic. Thus, the potential usefulness of NKX3.1 is also highlighted by the finding that its localization is predominantly nuclear, which can add additional diagnostic confidence in cases in which there is only relatively weak staining for one or more of the cytoplasmic markers such as PSA, PSAP, or P501s.
In summary, we report for the first time that IHC staining for NKX3.1 protein is retained in most primary untreated metastatic carcinomas of prostatic origin. The major difference in our study that would seem to account for the higher sensitivity for metastatic prostatic adenocarcinoma is the use of a relatively novel antibody against NKX3.1.6,11
Taking this study together with another recent study also showing strong staining for NKX3.1 in the majority of poorly differentiated primary prostate cancers,11
the addition of NKX3.1 protein staining to a panel of markers, if applied in the appropriate clinicopathologic context, may add diagnostic value in the diagnosis of metastatic lesions of unknown primary origin.