Molecular Basis of Prostate Cancer

Family history is one of the strongest risk factors for the development of prostate cancer, with a two- to eightfold higher risk of prostate cancer in men with an affected first-degree relative. ⁶ Prostate cancer associated with familial clustering and high incidence of cancer among multiple first-degree relatives with a diagnosis before age 60 is considered hereditary and/or familial prostate cancer (HPC/FPC). Approximately 9% of all cases are attributable to hereditary prostate cancer following an autosomal dominant susceptibility pattern. Prostate cancer susceptibility genes have been identified using lineage analysis of affected families. Significant linkage between chromosome 1q24-25, the HPC1 locus, and hereditary prostate cancer has been established. RNASEL, which lies within the HPC1 locus, encodes an endoribonuclease that mediates the activities of an interferon-inducible RNA degradation pathway. Polymorphisms of the RNASEL gene have been associated with increased prostate cancer risk. However, not all studies have confirmed these findings. Mutations in the RNASEL gene do not occur at a greater frequency in patients with familial prostate cancer compared with patients with sporadic prostate cancer. Recent genome-wide association studies (GWAS) consistently identified that several single-nucleotide polymorphisms (SNPs) in the 8q24 locus are associated with risk of HPC/FPC22. Genetic variants caused by polymorphisms or mutations in other genes, such as PALB2, BRCA2, the androgen receptor (AR), 5-α-reductase type II (SRD5A2), and CYP17, have also been implicated in the development of HPC/FPC.

Early candidate gene approaches have implicated many different genes in prostate cancer. Germline mutations involving ELAC2 (HPC2), MSR1, and RNASEL genes have been reported in familial prostate cancer. The most common somatic mutations found in sporadic prostate cancer include TP53, PTEN, and AR. Recent whole-genome exon sequencing analyses identified significant mutated genes, including TP53, AR, ZFHX3, RB1, PTEN, APC, MLL2, OR5L1, and CDK12. Of these genes, MLL2, OR5L1, and CDK12 have unknown tumor suppressor functions in prostate cancer. ⁷ The most commonly affected signaling pathways by genetic alterations are the WNT signaling (TP53, APC, CTNNB1, MYC, and SMAD4) and the PTEN interaction network (PTEN, MAGI3, and HDAC11). ⁷

DNA copy number variation (CNV) is DNA structure alteration involving relatively large (at least 1 kb) regions. CNV can manifest as loss or gain of chromosomal regions. Earlier studies using fluorescence in situ hybridization (FISH) and comparative genomic hybridization (CGH) identified common losses at 1p, 6q, 8p, 10q, 13q, 16q, and 18q and gains at 1q, 2p, 7, 8q, 18q, and Xq. For example, MSR1, NKX3.1, and N33 are candidate tumor suppressor genes in prostate cancer lying within the most commonly deleted regions on chromosome 8p. MSR1 encodes a receptor on the macrophage cell surface that induces binding of oxidized low-density lipoprotein and other polyanionic ligands. Mutations, polymorphisms, or loss of the MSR1 gene may compromise global macrophage function, thereby exposing organs, including the prostate, to oxidative stress and damage. Although this gene does not code for prostatic proteins directly, oxidative stress has been implicated in the initiation of prostate carcinogenesis.

The prostate cancer genome can harbor an average of 90 chromosomal rearrangements involving many genes. One of the most frequent gene fusion events in prostate cancer is the fusion of the TMPRSS2 and ERG genes, which are located 3 Mb apart on chromosome 21q22.2. ⁹ TMPRSS2 is an androgen-responsive gene, and ERG encodes an erythroblast transformation-specific (ETS) transcription factor. Their fusion is believed to be stimulated by androgens that recruit AR and TOP2B topoisomerase to chromosomal sites where TOP2B introduces double-strand breaks in DNA. TMPRSS2-ERG fusion results in overexpression of ETS genes in prostate cancer. TMPRSS2-ERG has been identified in 40% to 70% of prostate cancer and correlated with metastasis and disease-specific mortality. Gene fusion events such as TMPRSS2-ERG may be used for prostate cancer diagnosis through simple PCR detection of gene fusions in urine sediment.

The Wnt signaling pathway plays a key role in embryonic development and is essential for the maintenance of stem cells. Wnt is an extracellular protein that interacts with the membrane-bound frizzled receptor to initiate its biologic activity. Wnt signaling leads to stabilization of CTNNB1 and its nuclear accumulation. Nuclear CTNNB1 converts the TCF/LEF DNA-binding protein complex from a transcriptional repressor into a transcriptional activator. Inappropriate activation of the Wnt pathway is observed in many cancers and is putatively associated with tumor development.

Only a small fraction of the transcription output in human genome encodes for proteins. Noncoding RNAs (ncRNAs) are arbitrarily classified into two major classes based on their size: small (microRNA) and long ncRNA (lncRNA).

Small ncRNAs, exemplified by microRNA (miRNA), are known to regulate diverse biological processes in stem cells, development, differentiation, metabolism, and disease such as cancer. Several hundred miRNAs have been identified in human cells, and these are transcribed from the genome as long, primary miRNAs ranging in size from hundreds to thousands of nucleotides. miRNAs depend on multiple proteins for their biogenesis and function. Aberrations in any of these proteins will affect miRNA-mediated gene regulation. Expression of Dicer, a key gene in the biosynthesis of miRNA, is upregulated in a significant fraction of prostate cancer and is associated with aggressive cancer features. Knockout of Dicer in the mouse prostate impairs prostate stem cell activity and causes prostate atrophy. Some other components of the miRNA machinery (XPO5, Ago1, Ago2, HSPCA, MOV10, and TNRC6B) are also upregulated in prostate cancer.

High-throughput miRNA expression profiling studies have found altered miRNA expression in prostate cancer, providing evidence for the involvement of miRNAs in this disease. miRNAs that have increased expression or amplification in cancer can act as oncogenes to enhance cell proliferation and survival by the inhibition of protein-coding tumor suppressor genes. They are thus called oncomiRs. Several such oncomiRs have been identified in prostate cancer cell lines including miR-21, miR-291, miR-221, miR-222, and the well-known miR-17-92 cluster, which contains six members: miR-17, miR-18a, miR-19a, miR-20a, miR-19b-1, and miR-92a-1. Some common target genes of these oncomiRs include PTEN, BIM, RB1, p21, and p27. On the other hand, miRNAs that normally target and suppress are regarded as tumor suppressor miRNAs (ts-miRNAs). In prostate cancer, the miR-15a/16-1 cluster that targets BCL2 and CCND1 is downregulated in prostate cancer. Forced expression of these miRNAs can lead to suppression of cell proliferation and induction of apoptosis. ¹⁰ Other commonly identified tumor suppressor miRNAs include miR-34a/b/c, miR-145, miR-205, and let-7.

The AR signaling pathway and miRNAs are engaged in reciprocal regulation via multiple interaction points. miRNA can suppress AR expression through the canonical miRNA pathway or affect AR transcriptional activity by modulating other AR cofactors such as MYC. A systemic analysis of miRNAs identified a number of miRNAs that could suppress AR expression post-transcriptionally by targeting the AR 3′UTR element. Similar to protein coding genes, miRNA genes are also under the regulation of many transcription factors, including AR, which binds to ARE in the promoter of many miRNAs. As the first example, AR can bind to the ARE of miR-21, a prostate oncomiR, and directly regulate its expression as a downstream signaling effector. ¹¹ Several other miRNAs were later found to be also under the regulation of AR, including miR-101, miR-141, miR-27a, miR-32, and miR-148a.

miRNAs derived from cancer cells can exist as circulating extracellular or cell-free RNA and are remarkably stable in body fluids such as plasma/serum. These extracellular miRNAs can be detected and quantified using either PCR-based methods or other high-throughput approaches, including miRNA microarray and next-generation sequencing. A number of studies have performed miRNA profiling in body fluids including serum, plasma, and urine and have identified several miRNAs whose levels can serve as diagnostic and/or prognostic markers for prostate cancer patients. ^12,13 Some of the miRNAs that consistently exist in prostate cancer patients across different studies include miR-141 and miR-375.

The longer class of ncRNAs is known as long intergenic RNA (lncRNA), with sizes ranging from a few hundred to thousands of nucleotides. lncRNAs can regulate protein-encoding genes by affecting transcription and chromatin state by mechanisms distinct from those used by small ncRNAs. Thus genetic alterations and aberrant expression of lncRNAs can be a causal factor in disease. Several lncRNAs pertinent to prostate cancer have been identified. A 3.7-kb lncRNA known as PCA3 (DD3) has been mapped to chromosome 9q21-22 and shown to be highly overexpressed in prostate cancer samples. Genetic variation in lncRNAs has been found to affect prostate cancer risk. Microarray profiling of intronic transcripts identified many ncRNAs expressed in prostate cancer samples, and the expression levels of some correlate with the extent of prostate tumor cell differentiation. ¹⁴ By high-throughput RNA sequencing of 102 prostate tissues and cell lines, Prensner and colleagues identified 121 unannotated prostate cancer–associated ncRNA transcripts (PCATs) and have characterized one of them, PCAT-1, as a prostate cancer–specific ncRNA functionally implicated in disease progression.

Epigenetic alterations contribute to the malignant transformation and progression of prostate cancer. Figure 38-2 shows the potential contribution of epigenetic events to the development of prostate cancer and its progression to advanced and castration-resistant cancer. Initially DNA methylation was regarded as a new type of promising biomarker for prostate cancer diagnosis and prognosis, and a therapeutic target. Despite intensive research efforts in the past decade, results from clinical studies evaluating DNA methylation as a biomarker have been disappointing, and so far no biomarker has advanced into the clinical arena.

DNA hypermethylation is one of the most common and best-characterized epigenetic abnormalities in prostate cancer. Genes including classic and putative tumor suppressor genes as well as genes involved in a number of cellular pathways, such as hormonal responses, cancer cell invasion/tumor architecture, cell cycle control, and DNA damage repair, can be hypermethylated in prostate cancer. For many of these genes, promoter hypermethylation is often the mechanism responsible for their functional loss in prostate cancer. Inappropriate silencing of these genes can contribute to cancer initiation, progression, invasion, and metastasis. Some commonly hypermethylated genes in prostate cancer are discussed next (Table 38-2 ).

Antioxidants and DNA repair pathways protect the genome and maintain genome stability during replication or following DNA damage. Hypermethylation of genes important for such processes, such as glutathione S-transferase Pi (GSTP1) and O-6-methylguanine DNA methyltransferase (MGMT), has been frequently documented in prostate cancer. GSTP1, located at chromosome 11q13, belongs to a supergene family of glutathione S-transferases (GSTs) that play an important role in the detoxification of carcinogens and cytotoxic drugs by catalyzing their conjugation to glutathione. GSTP1 inactivation may lead to increased cell vulnerability to oxidative DNA damage and the accumulation of DNA base adducts, which can precede carcinogenesis.

AR mediates testosterone and dihydrotestosterone activity, which is essential for the development and maturation of the prostate gland and prostate cancer. Most prostate cancer is initially androgen dependent, but eventually becomes androgen independent after androgen-deprivation therapy. Androgen-independent prostate cancers are characterized by a heterogeneous loss of AR expression. Genetic alterations that alter the sensitivity of the receptor to androgen, such as AR gene mutation and, more commonly, amplification without loss of AR expression, are thought to play key roles in the development of androgen-independent prostate cancer. The prostate expresses two types of estrogen receptors (ERs): ERα (ESR1) and ERβ (ESR2), whose functional role in the prostate and prostate cancer remains controversial. Hypermethylation of ERs leading to decreased expression increases with aging and can contribute to tumor progression in prostate cancer patients.

An important characteristic of tumor cells is unbalanced proliferation due to impaired regulation of the cell cycle. The multiple checkpoints that control the cell cycle include the retinoblastoma protein, cyclins, cyclin-dependent kinases (CDKs), and CDK inhibitors (CDKIs). CDKIs are potential tumor suppressor genes that act as molecular brakes on cell cycle progression. Failure of cell cycle arrest due to alterations in CDKI expression has been implicated in prostate cancer. CDKIs are grouped into two families: the INK4 family and the CIP/KIP (kinase inhibitor protein) family. The INK4 family includes CDKN2A (p16), CDKN2B (p15), CDKN2C (p18), and CDKN2D (p19) and inhibits cyclin D–associated kinases (CDK4 and CDK6). The CIP/KIP family, which includes CDKNIA (p21), CDKN1B (p27), and CDKN1C (p57), inhibits most CDKs.

The cadherin-catenin adhesion system is critical for the preservation of normal tissue architecture and is regulated by a family of proteins collectively termed cell adhesion molecules (CAMs). Decreased expression of CDH1 and other CAMs has been reported to have prognostic significance in various human cancers, including prostate cancer. In prostate cancer, expression of CDH1 is markedly suppressed and its promoter is methylated to varying degrees. In addition, methylation of the CDH1 promoter is increased in advanced prostate cancer, making it a potential biomarker for cancer progression.