Immunohistochemical Biomarkers of Prostatic Carcinoma

RE: QUANTITATIVE POLYMERASE CHAIN REACTION DOES NOT IMPROVE PREOPERATIVE PROSTATE CANCER STAGING: A CLINICOPATHOLOGICAL MOLECULAR ANALYSIS OF 121 PATIENTS

33 Background: Historically, PSA DT can be prognostic for metastasis free survival in BCR (as defined on computed tomography (CT) and bone scan). PSA DT is also an important tool in risk stratification in BCR to identify which patients (pts) require therapy (e.g. pts with a PSA DT>6 months). The emerging use of PSMA imaging creates another tool to assess BCR pts, but there is no data on the association of PSA DT and PSMA findings. Methods: NCT05588128 enrolls BCR pts after definitive +/- salvage therapies. Pts must have a PSA>0.5 ng/ml, testosterone >100 ng/dL, and negative CT/bone scans. Lymph nodes (LNs) up to 1.5 cm and prior therapies are permitted. All patients undergo a baseline PSMA PET scan, along with measurement of PSA and PSA DT. Here we describe the relationship between baseline PSA metrics and baseline PSMA imaging findings in patients with BCR. Results: 130 patients are currently evaluable with a median age= 71 years, baseline PSA of 1.95 ng/dL (range: 0.5 to 71) and PSA DT of 10.6 months (range 1.2 to 132) off therapy. Of all participants, 16.9% (n=22) had findings limited to the prostate bed and 34.6% (n=45) had PSMA+ avidity in the prostate bed with other findings. 42.3% (n=55) had PSMA + LNs, and 13.8% (n=18) had PSMA+ bone lesions. Four patients had serosal deposits with PSMA avidity, and one patient had a PSMA+ lung nodule. Conclusions: These data are the first to compare PSMA imaging results with corresponding PSA levels and PSA DT in a large cohort of patients with BCR. Results show that pts with historically favorable/long PSA DT may have high volume/bone+ findings on PSMA. There is no data to suggest treatment escalation is required in BCR pts with high volume PSMA findings, but long/favorable PSA DT. These results highlight the caveats of using PSMA imaging alone to drive treatment decisions in BCR without further data for how baseline PSMA imaging corresponds with long-term outcomes. Clinical trial information: NCT05588128 . PSMA PET+ lesion locations by PSA doubling time in patients with BCR prostate cancer (total n=130). PSA DT # Pts Median PSA PSMA Neg Prostate+ only Lymph Nodes+ ≥ 4 Lymph Nodes+ Bone+ ≥12 mo 52 3.55 9 (17.3%) 16 (30.8%) 24 (46.2%) 12 (23%) 6 (11.5%) 9 to <12 mo 20 1.56 2 (10%) 4 (20%) 11 (55%) 3 (15%) 3 (15%) 6 to <9 mo 17 0.9 5 (29.4%) 0 (0%) 12 (70%) 8 (47.1%) 2 (11.8*) <6 mo 41 2.1 5 (12.2%) 2 (4.9%) 28 (68.3) 15 (36.6) 7 (17.1)

Antisense oligonucleotides (oligos) have been administered against in vivo and in vitro prostate cancer models employing LNCaP and PC-3 cell lines. While most oligos consist of a single mRNA binding site targeting a single gene product or those with sequence homology, our lab has developed bispecific oligos directed toward two unrelated proteins. In LNCaP cells, we initially identified bispecifics that increased the expression of prostate-specific membrane antigen (PSMA) while not affecting secreted prostate-specific antigen (PSA). We postulated that surface antigen expression is increased by bispecifics able to form double-stranded regions, acting as interferon (IFN-γ) inducers. In other systems, when induced, IFN-γ promotes cell surface antigen expression, including HLA and receptors for tumor necrosis factor. To test this hypothesis, we measured the effect of oligo treatment on both IFN-γ induction and the expression of another secreted product of differentiated prostate cells, prostatic acid phosphatase (PAP). This study initially evaluated the inhibition of in vitro propagating LNCaP cells employing mono- and bispecific oligos directed against bcl-2 (the second bispecific binding site was against the epidermal growth factor receptor). Employing RT-PCR, the expression of non-targeted proteins encoded by mRNA for PSMA, PSA, PAP, and IFN-γ was subsequently valuated. When LNCaP prostate tumor cells were incubated with oligos and compared to lipofectin-containing controls significant growth inhibition resulted. Employing RT-PCR, the levels of mRNA encoding PSMA were unexpectedly found to be elevated following treatment with the bispecific oligos but not with a monospecific directed solely against bcl-2. No differences were detected in mRNA levels encoding PSA following treatment with either oligo type. IFN-γ was significantly induced only by bispecific oligos, and PAP expression was similar to PSA. These data support the hypothesis that double strand-forming bispecific oligos induce IFN-γ that enhances cell surface PSMA expression. Expression of tumor-associated surface antigens could increase their recognition and targeting by immunologic defense mechanisms and increase the effectiveness of tumor vaccines.

Over the past two decades, monoclonal antibody technology has had an increasing impact on clinical diagnostic and therapeutic options, and this is true in the realm of managing prostate cancer. Several targets such as prostate-specific antigen and prostatic acid phosphatase as well as, more recently, angiogenic antigens such as vascular endothelial growth factor have been examined for therapy. Prostate-specific membrane antigen, a type II integral membrane glycoprotein initially characterized by the monoclonal antibody 7E11, has shown promise. Recent evidence suggests that prostate-specific membrane antigen is also expressed in tumor-associated neovasculature of a wide variety of malignant neoplasms. With its expression in prostate secretory-acinar epithelium and the prostate and in the neovasculature associated with tumors, prostate-specific membrane antigen represents an excellent antigenic target for monoclonal antibody diagnostic and therapeutic options. As research continues, the role of monoclonal antibody imaging and therapy will become increasingly important in the management of prostate cancer.

At present, increased early detection of prostate cancer appears to be the most feasible way to reduce cancer-related mortality. As a result significant efforts have been made to identify more men with curable cancer. This article reviews the role of serum prostate-specific antigen in an early detection or screening strategy and describes efforts to enhance the specificity of prostate-specific antigen testing.

We have investigated the expression of prostate-specific membrane antigen (PSMA) and prostate-specific antigen (PSA) transcripts in androgen-dependent (LNCaP) and androgen-independent (22Rv1) prostate cancer cell lines. We also enquired whether Q640X CTE-truncated androgen receptor (AR) has an impact on transcription of mRNA for PSMA and PSA in transfected androgen-sensitive prostate cancer LNCaP cells. Wild type LNCaP, 22Rv1 prostate cancer cells, prostate stromal cells (PrSC) and LNCaP cells transfected with p-Q640X AR, p-WT AR or p-C3 empty plasmids were studied. The expression of PSMA and PSA were detected by real-time PCR after transfection for 4 and 7 days. Expression of mRNAs for PSA was sixfold greater than PSMA in wild type LNCaP cells. In contrast, the wild type androgen refractory 22Rv1 cell line reacted almost exactly the opposite way reverse to LNCaP cells, since the transcription of mRNA for PSMA almost twofold greater than PSA. Non-transfected human PrSC responded similarly to PSMA mRNA and PSA mRNA was not detected in these cells. Q640X AR transfected LNCaP cells downregulated the expression of PSMA and PSA genes after 7 days. Our results demonstrate that Q640X mutated AR may have an important regulatory role in mediating the PSMA and PSA genes expression during the progression of prostate cancer from androgen-dependence to androgen-independence. Understanding their functional properties and mechanisms by which ARs involved in regulation of PSMA and PSA expression will allow the identification of new target therapies for the treatment of hormone-resistant prostate cancer.

Imaging of Prostate Specific Membrane Antigen Targeted Radiotracers for the Detection of Prostate Cancer Biochemical Recurrence after Definitive Therapy: A Systematic Review and Meta-Analysis.

PREDICTORS OF SUBSEQUENT PROSTATE CANCER IN MEN WITH A PROSTATE SPECIFIC ANTIGEN OF 2.6 TO 4.0 NG/ML AND AN INITIALLY NEGATIVE BIOPSY

236 Background: Conventional imaging techniques infrequently detect the site of prostate cancer disease in men with biochemical recurrence following radical prostatectomy (RP). We examine the use of Gallium-68 Prostate-Specific Membrane Antigen (PSMA) positron emission tomography (PET) to determine the site of disease recurrence in these men. Methods: We retrospectively reviewed men with persistently detectable prostate specific antigen (PSA) and those with formal biochemical recurrence (PSA ≥ 0.2 ng/mL) following RP who underwent PSMA PET scan under our institution's prospective trials (NCT02918357 & NCT02611882). We assessed the location of detected recurrences, and examined the distribution of recurrence location (prostate bed, pelvic lymph nodes (LN), and extra-pelvic disease) according to PSA at time of PET scan. Results: One hundred and fifty-eight men underwent PSMA PET after RP, 46% of men had previously undergone either adjuvant or salvage radiation therapy. Eleven men (7%) had the scan for persistently detectable PSA < 0.2 and 93% of men had biochemical recurrence. At time of RP, 66% of men had ≥pT3a disease, 40% had positive surgical margins, and 18% had positive lymph nodes. Gleason grade at RP was ≤3+3 in 5% of men, 3+4 in 27%, 4+3 in 32% and ≥4+4 in 36%. Overall, 77% of men had a lesion detectable on PSMA PET. Men with higher PSA at time of scan had a higher prevalence of lesions, with all men PSA > 6.0 having at least one lesion on PSMA PET (Table). Overall, we found extra-pelvic lesions in 44% of all men, and 58% of men with PSA ≥ 1.0. These are areas not typically covered by salvage radiation fields. Conclusions: PSMA PET detects lesions in a high proportion of men with biochemical recurrence following RP, with many lesions outside of the pelvis. This may facilitate more precise targeting of treatment areas for these patients. Clinical trial information: NCT02918357. Clinical trial information: NCT02611882. [Table: see text]

In patients with biochemical recurrence (BCR), prostate-specific membrane antigen (PSMA) positron emission tomography/computed tomography (PET/CT) can detect oligometastatic prostate cancer (PCa). However, the optimal treatment approach for oligometastatic disease remains unclear. This study aims to assess the oncological outcomes of metachronous oligometastatic PCa patients treated with metastasis-directed radiotherapy (MDRT). We retrospectively evaluated patients with hormone-sensitive, metachronous oligometastatic PCa who underwent MDRT for BCR (from July 2012 to September 2022). Patients had one to four lymph nodes and/or bone metastases on PSMA PET/CT and were irradiated with 5×7 or 3×10 Gy. The biochemical response to MDRT was assessed as undetectable prostate-specific antigen (PSA) at follow-up, PSA response (PSA≤pretreatment level), or biochemical progression (PSA>pretreatment level). Biochemical progression-free survival (bPFS) and local remission of disease (absence of disease at the MDRT site on follow-up PSMA PET/CT or undetectable PSA) were evaluated. Eighty patients underwent MDRT for 105 PSMA PET/CT-confirmed lesions. The median time from curative treatment to MDRT was 55 mo (interquartile range [IQR] 31-103). At a median follow-up of 32 mo after MDRT (IQR 21-64), 10% of the patients were PSA free, 10% had a PSA response, and 80% experienced biochemical progression. The bPFS rates at 1 and 2 yr were 54% and 38%, respectively. A total of 87% of patients had local control of disease after MDRT, whereas 72% had new metastatic lesions on repeated PSMA PET/CT. Limitations include the retrospective design and a small cohort. MDRT for oligometastatic disease shows high local efficacy. However, disease progression is observed in a substantial proportion of patients.

<div>AbstractPurpose:<p>Prostate-specific membrane antigen (PSMA)–based images, which visually quantify <i>PSMA</i> expression, are used to determine prostate cancer micrometastases. This study evaluated whether a circulating tumor cell (CTC)–based transcript platform, including PSMA mRNA, could help identify potential prognostic markers in prostate cancer.</p>Experimental Design:<p>We prospectively enrolled 21 healthy individuals and 247 patients with prostate cancer [localized prostate cancer (LPCa), <i>n</i> = 94; metastatic hormone–sensitive prostate cancer (mHSPC), <i>n</i> = 44; and metastatic castration-resistant prostate cancer (mCRPC), <i>n</i> = 109]. The mRNA expression of six transcripts [<i>PSMA</i>, prostate-specific antigen (<i>PSA</i>), <i>AR, AR-V7, EpCAM</i>, and <i>KRT 19</i>] from CTCs was measured, and their relationship with biochemical recurrence (BCR) in LPCa and mCRPC progression-free survival (PFS) rate in mHSPC was assessed. PSA-PFS and radiological-PFS were also calculated to identify potential biomarkers for predicting androgen receptor signaling inhibitor (ARSI) and taxane-based chemotherapy resistance in mCRPC.</p>Results:<p>CTC detection rates were 75.5%, 95.3%, and 98.0% for LPCa, mHSPC, and mCRPC, respectively. In LPCa, PSMA [hazard ratio (HR), 3.35; <i>P</i> = 0.028) and PSA mRNA (HR, 1.42; <i>P</i> = 0.047] expressions were associated with BCR. Patients with mHSPC with high PSMA (HR, 4.26; <i>P</i> = 0.020) and PSA mRNA (HR, 3.52; <i>P</i> = 0.042) expressions showed significantly worse mCRPC-PFS rates than those with low expression. Increased PSA and PSMA mRNA expressions were significantly associated with shorter PSA-PFS and radiological PFS in mCPRC, indicating an association with drug resistance.</p>Conclusions:<p>PSMA and PSA mRNA expressions are associated with BCR in LPCa. In advanced prostate cancer, PSMA and PSA mRNA can also predict rapid progression from mHSPC to mCRPC and ARSI or taxane-based chemotherapy resistance.</p></div>

Prostate-specific membrane antigen (PSMA) is an integral cell-surface membrane glycoprotein that is overexpressed in prostate carcinomas rendering it an appropriate target for antibody-based therapeutic strategies. The biosynthesis of PSMA in transfected COS-1 cells reveals a slow conversion of mannose-rich to complex glycosylated PSMA compatible with slow transport kinetics from the endoplasmic reticulum to the Golgi. Importantly, mannose-rich PSMA persists as a trypsin-sensitive protein throughout its entire life cycle, and only Golgi-located PSMA glycoforms acquire trypsin resistance. This resistance, used here as a tool to examine correct folding, does not depend on the type of glycosylation, because different PSMA glycoforms generated in the presence of inhibitors of carbohydrate processing in the Golgi are also trypsin resistant. The conformational transition of PSMA to a correctly folded molecule is likely to occur in the Golgi and does not implicate ER molecular chaperones, such as BiP. We show here that PSMA is not only heavily N-but also O-glycosylated. The question arising is whether glycans, which do not play a role in folding of PSMA, are implicated in its transport to the cell surface. Neither the cell-surface expression of PSMA nor its efficient apical sorting in polarized Madin-Darby canine kidney cells are influenced by modulators of N- and O-glycosylation. The acquisition of folding determinants in the Golgi, therefore, is an essential prerequisite for protein trafficking and sorting of PSMA and suggests that altered or aberrant glycosylation often occurring during tumorigenesis has no regulatory effect on the cell-surface expression of PSMA.

Background: Due to its biological properties, Prostate-specific membrane antigen (PSMA) is a good target for prostate cancer molecular imaging. The aim of this work was to evaluate the role of 18F PSMA Tomography–Computed Tomography (PET CT) in the diagnosis and staging of cancer prostate in association with prostate specific antigen serum level. Methods: This prospective cohort work was performed on 30 individuals pathologically proven prostate cancer patients, seven patients were recently diagnosed and were referred for primary staging of the disease, and 27 patients were long term patients who received initial treatment and were referred due to high prostate-specific antigen (PSA) level. Results: There was positive association between PSA values and maximum standardized uptake values (SUV max) of PSMA expression at prostate with insignificant correlations between PSA and age or SUV max of PSMA expression of metastatic lesions of prostate cancer. At 3.4 cut-off value, SUV max of PSMA expression at prostate had 100% sensitivity and specificity win diagnosis of active local cancer prostatic lesions with higher AUC than PSA. PSA was higher among metastatic prostate cancer patients with statistically significant difference. At cut-off value equal to 5.3, PSA had 95% sensitivity and 71.4% specificity in prediction of metastasis.Conclusions: PSMA PET/CT is the modality of choice in recurrent cancer prostate, it can assess the site of recurrence and determine the disease burden. It can detect cancer prostate metastasis in non-enlarged lymph nodes, bony deposits even with no apparent CT morphological changes.

This study compared the radiomic features and quantitative biomarkers of 18 F-PSMA-1007 [prostate-specific membrane antigen (PSMA)] and 18 F-fluorocholine (FCH) PET/computed tomography (CT) in prostate cancer patients with biochemical recurrence (BCR) enrolled in the phase 3, prospective, multicenter BIO-CT-001 trial. A total of 106 patients with BCR, who had undergone primary definitive treatment for prostate cancer, were recruited to this prospective study. All patients underwent one PSMA and one FCH PET/CT examination in randomized order within 10 days. They were followed up for a minimum of 6 months. Pathology, prostate-specific antigen (PSA), PSA doubling time, PSA velocity, and previous or ongoing treatment were analyzed. Using LifeX software, standardized uptake value (SUV) maximum, SUV mean , PSMA and choline total volume (PSMA-TV/FCH-TV), and total lesion PSMA and choline (TL-PSMA/TL-FCH) of all identified metastatic lesions in both tracers were calculated. Of the 286 lesions identified, the majority 140 (49%) were lymph node metastases, 118 (41.2%) were bone metastases and 28 lesions (9.8%) were locoregional recurrences of prostate cancer. The median SUV max value was significantly higher for 18 F-PSMA compared with FCH for all 286 lesions (8.26 vs. 4.99, respectively, P < 0.001). There were statistically significant differences in median SUV mean , TL-PSMA/FCH, and PSMA/FCH-TV between the two radiotracers (4.29 vs. 2.92, 1.97 vs. 1.53, and 7.31 vs. 4.37, respectively, P < 0.001). The correlation between SUV mean /SUV max and PSA level was moderate, both for 18 F-PSMA ( r = 0.44, P < 0.001; r = 0.44, P < 0.001) and FCH ( r = 0.35, P < 0.001; r = 0.41, P < 0.001). TL-PSMA/FCH demonstrated statistically significant positive correlations with both PSA level and PSA velocity for both 18 F-PSMA ( r = 0.56, P < 0.001; r = 0.57, P < 0.001) and FCH ( r = 0.49, P < 0.001; r = 0.51, P < 0.001). While patients who received hormone therapy showed higher median SUV max values for both radiotracers compared with those who did not, the difference was statistically significant only for 18 F-PSMA ( P < 0.05). Our analysis using both radiomic features and quantitative biomarkers demonstrated the improved performance of 18 F-PSMA-1007 compared with FCH in identifying metastatic lesions in prostate cancer patients with BCR.

The pattern of serum markers in patients with androgen-independent adenocarcinoma of the prostate