**2. Cellular and Molecular Progression of Prostate Cancer**

The human prostate is a walnut-size glandular organ that develops from the embryonic urogenital sinus [15]. Its primary function is to produce seminal fluid containing zinc, citric acid, and various enzymes, including a protease named prostate-specific antigen (PSA). Histologically, the prostate can be divided into central, peripheral, and transition zones comprised of a secretory ductal-acinar structure located within a fibromuscular stroma [16,17]. The ductal-acinar structure is formed of tall columnar secretory luminal cells, a flattened basal epithelium attached to the basement membrane, and scattered neuroendocrine cells (Figure 1). Luminal epithelial cells express cytokeratins (CK) 8 and 18, NKX3.1, androgen receptor (AR), and PSA, whereas basal epithelial cells express CK5, CK14, glutathione S-transferase Pi 1 (GSTP1), p63, and low levels of AR [18,19].

The cellular origin of prostate cancer is not very clear, partly because of the lack of well-characterized prostate epithelial lineage [20–22]. PCa develops from normal prostate epithelium through a multistep histological transformation process, governed by various underlying molecular changes [23] (Figure 2). Low-grade and high-grade prostate intraepithelial neoplasia (PIN) lesions develop from normal prostate epithelium through the loss of phosphatase and the tensin homolog *(PTEN)*, NK3 Homeobox 1 (*NKX3.1)*, overexpression of *MYC* proto-oncogene, B-cell lymphoma 2 (*BCL-2),* and the glutathione S-transferase pi 1 gene (*GSTP1),* accompanied with Speckle Type BTB/POZ Protein (*SPOP)* mutation and Transmembrane Serine Protease 2- ETS-related gene (*TMPRSS2-ERG)* fusion [24–36]. Further loss of the retinoblastoma protein (*RB1)*, along with telomerase activation and frequent Forkhead Box A1 (*FOXA1)* mutation, leads to the development of prostate adenocarcinoma from the advanced PIN lesion [37–43]. Further molecular aberrations including the loss of SMAD Family Member 4 (*SMAD4)*, AR corepressors, mutations in AR, *FOXA1, BRCA1*/*2, ATM, ATR,* and *RAD51* accompanied with the gain of function of the AR coactivator, *CXCL12, CXCR4, RANK-RANKL,* EMT, *BAI1*, and *EZH2* lead to the development of metastatic prostate cancer [44–59].

**Figure 1.** The location and architecture of the human prostate gland. The prostate gland is located below the bladder and consists of a central, a peripheral, and a transition zone. Histologically, it is comprised of secretary luminal, basal, and rare intermediate and neuroendocrine cells. The prostatic epithelium is separated from the stromal cells by the basement membrane as indicated. Preneoplastic or neoplastic cellular transformation can initiate from either basal or luminal cells.

**Figure 2.** Histopathological and molecular progression of human prostate cancer. Metastatic prostate cancer develops via progression through prostate intraepithelial neoplasia (PIN) and invasive adenocarcinoma through the acquirement of various molecular alterations as depicted. The invasive adenocarcinoma cells and androgen-deprivation therapy resistant cancer cells metastasize to the bone, lymph node, lung, and liver.

As evident from the PCa progression model (Figure 2), inactivation of *PTEN* appears to be a critical event in PCa carcinogenesis and associated with aggressive disease manifestation. *PTEN* alterations occur in various ways in prostate cancer, such as genomic deletion and rearrangement, intragenic breakage, or translocation. The loss of *PTEN* is linked with an upregulation of PI3K/AKT/mTOR signaling that regulates cell survival, proliferation, and energy metabolism [60,61]. Another critical determinant of PCa tumorigenesis is *SMAD4*, a tumor suppressor gene (18q21.1), which mediates the transforming growth factor β (TGF-β) signaling pathway and suppresses epithelial cell growth. Transcriptome analysis revealed significantly lower levels of *SMAD4* in PCa tissues compared to adjacent non-cancerous tissues [46]. Of note, in a mouse model, prostate specific ablation of *Smad4* and *Pten* leads to the development of an invasive and metastatic potential of PCa (discussed below) [45].

In the PCa initiation and progression cascade, tumor suppressor *NKX3.1* (8p21) plays a pivotal role and found to be frequently lost due to the loss of heterozygosity (LOH) [62,63]. Of note, LOH at 8p21 appears to be an early event in PCa tumorigenesis [63–65]. Thus, it is likely that the genes that reside within these frequently deleted regions are associated with PCa initiation. Under the normal condition, *NKX3.1* drives growth-suppressing and differentiating effects on the prostatic epithelium [66]. *Nkx3.1* heterozygous mice develop abnormal prostate morphology with the dysplastic epithelium [67,68]. Importantly, *Nkx3.1*-null mice show changes in prostate epithelial morphology with severe dysplasia [67]. Kim et al. demonstrated that the loss of function of *Pten* and *Nkx3.1* in mice cooperated in PCa development. Importantly, *Pten;Nkx3.1* compound mutant mice showed a higher incidence of High-grade prostatic intraepithelial neoplasia (HGPIN) [69]. In addition to the critical tumor suppressor genes described above, the *MYC* proto-oncogene is also amplified in PCa [70–72]. *MYC* encodes a transcription factor that regulates the expression of several genes involved in cell proliferation, metabolism, mitochondrial function, and stem cell renewal [73–75]. Several studies suggest that *MYC* is activated through overexpression, amplification, rearrangement, Wnt/β-catenin pathway activation, germline *MYC* promotor variation, and loss of *FOXP3* in PCa [76–79], and is a critical oncogenic event driving PCa initiation and progression [71,80].

Other than *MYC, TMPRSS2:ERG* gene fusion, resulting from the chromosomal rearrangement, is also reported in approximately 45% of PCa. This alteration leads to the expression of the truncated *ERG* protein under the control androgen-responsive gene promoter of *TMPRSS2* [81–85]. *ERG* belongs to the *ETS* family of transcription factors (*ERG, ETV1*, and *ETV4*), and its activation is associated with PCa progression in both early- and late-stages [82,83,86]. *MYB*, another gene encoding a transcription factor, is also reported to be amplified in PCa and exhibits an increased amplification frequency in castration resistant PCa (CRPC) [87]. Research from our laboratory has shown that *MYB* plays a vital role in PCa growth, malignant behavior, and androgen-depletion resistance [56].
