Prostate-Specific Membrane Antigen-Targeted Imaging and Its Correlation with HOXB13 Expression

Abstract

Homeobox 13 (HOXB13) is an oncogenic transcription factor that directly regulates expression of folate hydrolase 1, which encodes prostate-specific membrane antigen (PSMA). HOXB13 is expressed in primary and metastatic prostate cancers (PCs) and promotes androgen-independent PC growth. Since HOXB13 promotes resistance to androgen receptor (AR)-targeted therapies and regulates the expression of folate hydrolase 1, we investigated whether SUVs on PSMA PET would correlate with HOXB13 expression. Methods: We analyzed 2 independent PC patient cohorts who underwent PSMA PET/CT for initial staging or for biochemical recurrence. In the discovery cohort, we examined the relationship between HOXB13, PSMA, and AR messenger RNA (mRNA) expression in prostate biopsy specimens from 179 patients who underwent PSMA PET/CT with ¹⁸F-piflufolastat. In the validation cohort, we confirmed the relationship between HOXB13, PSMA, and AR by comparing protein expression in prostatectomy and lymph node (LN) sections from 19 patients enrolled in ¹⁸F-rhPSMA-7.3 PET clinical trials. Correlation and association analyses were also used to confirm the relationship between the markers, LN positivity, and PSMA PET SUVs. Results: We observed a significant correlation between PSMA and HOXB13 mRNA (P < 0.01). The association between HOXB13 and ¹⁸F-piflufolastat SUVs was also significant (SUV_max, P = 0.0005; SUV_peak, P = 0.0006). Likewise, the PSMA SUV_max was significantly associated with the expression of HOXB13 protein in the ¹⁸F-rhPSMA-7.3 PET cohort (P = 0.008). Treatment-naïve patients with LN metastases demonstrated elevated HOXB13 and PSMA levels in their tumors as well as higher PSMA tracer uptake and low AR expression. Conclusion: Our findings demonstrate that HOXB13 correlates with PSMA expression and PSMA PET SUVs at the mRNA and protein levels. Our study suggests that the PSMA PET findings may reflect oncogenic HOXB13 transcriptional activity in PC, thus potentially serving as an imaging biomarker for more aggressive disease.

Keywords: HOXB13; PSMA; PSMA PET; androgen receptor; metastasis; prostate cancer.

Graphical abstract

FIGURE 1.

Study design of PSMA PET/CT and biomarker expression profiling in PC patients. Schematic diagram of patient stratification for retrospective analysis is shown. ADT = androgen-deprivation therapy; IHC = immunohistochemical; RP = radical prostatectomy; RT = radiation therapy.

FIGURE 2.

Association among HOXB13, PSMA, and AR with PSMA SUVs in PyL cohort. (A) Correlation among HOXB13, PSMA, and AR mRNA expression for each patient (n = 179). Lines represent linear regression. (B and C) Correlation among HOXB13, PSMA, and AR mRNA expression vs. prostate PSMA SUV_max (B) or prostate PSMA SUV_peak (C) for measurable lesions. Line represents linear regression (note logarithmic scale) (n = 105; initial staging PSMA PET–positive, n = 94; biochemical recurrence, n = 11).

FIGURE 3.

Correlation of HOXB13, PSMA, and AR protein expression in advanced PC. (A) Immunohistochemical (IHC) analysis shows representative normal and prostate tumor sections from individual FFPE patient specimens stained for AR, HOXB13, and PSMA. Digital quantification (DQ) analysis of marker expression is given below each panel. Scale bar = 200 μm. (B) Expression of HOXB13, AR, and PSMA in normal vs. tumor sections obtained by IHC and DQ determination was compared by Student t test (****P < 0.0001; n = 19). (C) Correlation analysis compares expression of each marker by IHC and DQ determination. Line represents linear regression, and r indicates Pearson correlation coefficient. H&E = hematoxylin and eosin; + = positive; (-)ve = negative.

FIGURE 4.

HOXB13 PSMA coexpression in recurrent and metastatic PC. (A) Representative normal and prostate tumor FFPE sections from treatment-naïve or recurrent PC immunohistochemically (IHC) stained for AR, HOXB13, and PSMA protein expression. Digital quantification (DQ) of staining is shown below each panel. Left panel: treatment-naïve. Right panel: recurrence with radiation plus androgen-deprivation therapy. (B) LN sections (GGG3 T3aN1 for LN 1 or GGG5 T3bN1 for LN 2). (C) Pearson correlation analysis comparing HOXB13 and PSMA protein expression in LNs. Line represents linear regression, and r indicates Pearson correlation coefficient. H&E = hematoxylin and eosin; + = positive; (-)ve = negative.

FIGURE 5.

Detection of primary and metastatic prostate tumors by ¹⁸F-rhPSMA-7.3 PSMA PET. (A) Schematic of screening of PC patients with PSMA PET. (B) ¹⁸F-rhPSMA-7.3 PET/CT images of PSMA-positive prostate tumor and LN metastasis. Immunohistochemical (IHC) analysis shows representative FFPE-fixed prostate and LN patient sections stained for AR, HOXB13, and PSMA with corresponding hematoxylin and eosin (H&E) staining. Scale bar = 200 μm. (C and D) Pearson correlation analysis of the individual biomarker expression by IHC and digital quantification and prostate PSMA SUV_peak (C) (n = 19) and prostate PSMA SUV_max (D) (n = 19). Line represents linear regression, and r indicates Pearson correlation coefficient.