**1. Introduction**

Prostate cancer is the most common type of cancer among men in 92 countries and the leading cause of cancer deaths among men in 48 countries [1]. In the United States as well, prostate cancer has been the most commonly diagnosed type of cancer among men, accounting for almost 1 in 5 new diagnoses [2]. While the incidence of latent prostate cancer has been similar between the United States and Japan, the incidence of clinically detected prostate cancer has been lower in Asia, including Japan [3,4]. Of note, the incidence of prostate cancer in Chinese and Japanese men has been reported to increase substantially after migration to the United States [5]. Furthermore, the morbidity and mortality due to prostate cancer in Asia increased remarkably in recent years [6]. Although the etiology of prostate cancer is multifocal, these epidemiological findings, including geographic and ethnic di fferences, sugges<sup>t</sup> that lifestyle and/or environmental factors have a substantial influence on the development and progression of prostate cancer [7]. Epidemiological evidence suggested that among the acquired risk factors for prostate cancer development and progression, diet and obesity have a potential to cause prostate cancer initiation, promotion, and progression [8,9]. Several studies have implicated dietary fats as important factors of prostate cancer risk and its aggressive phenotype [9,10]. A number of clinical and preclinical studies have shown that total fat intake and specific fat composition play a potential role in prostate cancer, although their findings have remained inconclusive.

Considering these backgrounds, this study aimed to summarize previous preclinical studies regarding the relationship between dietary fat and prostate cancer development and progression, focusing on di fferences in preclinical models and dietary fat composition. Furthermore, potential mechanisms on dietary fat-induced prostate carcinogenesis were discussed by updating previous research evidence. To this end, previous preclinical studies investigating dietary fat and prostate cancer were identified using a PubMed search including only studies published in English. This review helps us to understand the current state of diet-induced prostate cancer research in order to guide future works exploring the association between dietary-fat and prostate cancer.

### **2. Various Preclinical Models**

A number of animal models, including those involving prostate cancer cell xenografts and allografts, Transgenic Adenocarcinoma of the Mouse Prostate (TRAMP) mice, and other genetically engineered mice targeting oncogenes and tumor suppressor genes, were tested in order to assess the impact of dietary-fat intake on prostate cancer development and progression (Table 1). First, the models used in the previous studies were summarized as follows.


**Table 1.** Summary of preclinical models on dietary-fat induced prostate cancer development and progression.






restriction, Ab: antibody, SD: standard diet, CD: control diet, PDX: patient-derived xenograft, NWD: new Western-style diet.

**Table 1.** *Cont.*

### *2.1. Human Cancer Cell Xenograft and Allograft Models*

The most experienced models to assess the impact of dietary fats on prostate cancer growth were subcutaneous xenograft models [11,12,15,16,35,52,58].

Nude [16] and severe combined immunodeficient (SCID) mice [15] were frequently used as host mice for human prostate cancer cell xenografts. In 1995, Wand et al. first assessed the impact of five di fferent fat percentages on human prostatic adenocarcinoma (LNCaP) xenograft growth using athymic nude mice [11]. Accordingly, mice who continued to receive a 40.5-kcal% fat diet had substantially greater tumor growth rates, final tumor weights, and final tumor weight to animal weight ratios compared to those whose diets were changed to 2.3 kcal%, 11.6 kcal%, or 21.2 kcal% fat, suggesting that those fed low-fat diets (LFDs) had decreased growth of established LNCaP tumors. An additional study demonstrated that an isocaloric LFD (12 kcal% fat) resulted in significantly slower tumor growth rates and lower serum prostate-specific antigen (PSA) levels compared to a high-fat diet (HFD) using LAPC-4 xenografts on SCID mice [15]. The same group also showed that reduced dietary fat intake delayed conversion from androgen-sensitive to androgen-insensitive prostate cancer and significantly prolonged survival of SCID mice bearing LAPC-4 xenografts [58]. Moreover, we had previously found that Balb/c-nu/nu mice receiving a HFD had significantly higher LNCaP xenograft tumor volumes and serum PSA levels than those receiving an LFD [52]. The impact of a HFD on xenograft tumor growth using other human prostate cancer cell lines, such as 22Rv-1 and PC-3, had also been investigated in previous literatures [32,51]. Although the significance of the e ffect varied, a number of studies proposed that a HFD accelerated tumor growth of human prostate cancer cell xenografts inoculated into immunodeficient mice. Conversely, several studies have found no relationship between a HFD and xenograft growth [22,26]. In a study comparing LAPC-4-xenografted SCID mice receiving an isocaloric Western diet (40% fat and 44% carbohydrate) and those receiving an LFD (12% fat and 72% carbohydrate), the authors found no di fference in tumor growth or survival between both groups when saturated fat was used as the fat source [26]. Another study showed no di fference in LNCaP tumor size between normal (6% fat) and high-fat (14% fat) diets [22]. Taken together, a number of studies involving subcutaneous human prostate cancer cell xenografts in immunodeficient mice suggested an association between HFD and xenograft growth, whereas several other studies showed no such relationship. The lack of standardization in terms of models and duration of specific diet feeding has remained problematic.

Given the variations in the genetic background of mouse strains, it is important to consider the importance of the immune system in tumor progression [59]. Several studies have investigated the impact of dietary fat on allografts using immunocompetent mice and mouse-derived prostate cancer cells [31,41,45,49,55]. Several groups have shown that a HFD significantly increased allograft tumor growth of TRAMP-derived prostate cancer cells, such as TRAMP-C1 and TRAMP-C2, in C57BL6 mice [31,41,55]. The study involving the largest number of allografts (low-fat; *n* = 40, high-fat; *n* = 134) revealed that mice receiving AIN-93M-high-fat diet had significantly heavier and significantly larger TRAMP-C2 allografts compared to those receiving AIN-93M, whereas no di fferences in prostate weight were observed among the groups [31]. This result suggests that TRAMP allografts derived from C57BL6 mice can be one of the promising allograft models when studying HFD-induced prostate cancer progression.

A unique study involving a peritoneal dissemination model established through intracorporeal injection of PC-3M-luc cells detected using the Xenogen IVIS ™ system reported that a HFD increased tumor formation rates and total metastasis rates in the peritoneal organs [53].

In summary, given that most of the xenograft and allograft studies were performed using subcutaneous xenograft models, studies involving metastatic models and human patient-derived xenografts (PDXs) have been lacking. Although several studies using xenografts and allografts have shown that a HFD accelerated tumor growth, further validation is warranted.

### *2.2. TRAMP Mouse Models*

Since its generation in 1996, the TRAMP mouse model has been one of the most widely used models in prostate cancer research [60]. This model represents a transgene comprising the minimal probasin promoter driving viral SV40 large-T and small-t antigens, which lead to prostate-specific inactivation of pRb and p53, specifically in the prostatic epithelium [61]. TRAMP mice develop prostatic intraepithelial neoplasia (PIN) by the time they are 6 weeks old; this progresses to high-grade PIN by the age of 12 weeks and poorly di fferentiated and invasive adenocarcinoma by the age of 24 weeks, with nearly 100% penetrance [61]. The impact of a HFD on the growth of TRAMP mouse tumors had been frequently evaluated [25,31,39,41–43,57,62]. Accordingly, Llaverias et al. showed that mice consuming a Western-type diet enriched in both fat and cholesterol had higher prostate tumor incidence and greater tumor burden compared to those fed a control chow diet [25]. After necropsy at 28 weeks, 33% of TRAMP mice fed a Western diet showed grossly evident spherical prostate tumors, whereas only 17% of TRAMP mice fed a chow diet exhibited the same [25]. In another study on TRAMP mice, Xu et al. revealed that the HFD group had significantly higher mortality than the normal diet group (23.81% and 7.14%, respectively, *p* = 0.035). Moreover, HFD-fed TRAMP mice had significantly higher tumor incidence at 20 weeks, as compared to the normal diet group (78.57% and 35.71%, *p* = 0.022, respectively) [43]. The same group also showed that HFD-fed mice su ffered higher rates of extracapsular extension (20 weeks, 16.7% vs. 8.3%; 28 weeks, 66.7% vs. 50.0%, respectively) and distant metastasis (e.g., retroperitoneal lymph nodes or lung metastasis) (28 weeks, 41.7% vs. 25.0%, respectively) [62]. Bonorden et al. conducted a unique study involving the largest number of mice (*n* = 25 each) to assess the direct e ffect of diet and body weight on prostate tumors. TRAMP mice received low- and high-fat diets with the latter being divided into three groups: obesity-prone (the heaviest third), overweight (the middle third), and obesity-resistant (the lightest third). Accordingly, their results showed that body weight or diet had no e ffect of on either age at tumor detection, neuroendocrine status, or age at death [31]. Taken together, the impact of a HFD on tumor incidence and survival of TRAMP mice still remains controversial. The timing of diet change, selection of control diet, and diet ingredients may be important in establishing HFD-accelerated orthotopic prostate tumor models in TRAMP mice.

### *2.3. Other Genetically Engineered*/*Transgenic Mouse Models Targeting Oncogenes and Tumor Suppressor Genes*

Several studies have investigated the e ffect of dietary fat on prostate cancer development and progression using genetically engineered mouse models (GEMMs) targeting oncogenes and tumor suppressor genes [19,29,30,36,40,47,48,54,56]. Designated Hi-myc uses a PB promoter coupled with a sequence of the ARR2 promoter, both of which lie upstream to the human c-Myc gene, in order to drive progression from mouse prostatic intraepithelial neoplasia (mPIN) to invasive adenocarcinoma [63]. Using this animal model, Kobayashi et al. showed that the HFD group (42 kcal% fat) had a greater number of invasive adenocarcinoma and a higher proliferative index in the PIN region compared to the LFD group (12 kcal% fat) [19]. Phosphatase And Tensin Homolog (Pten) alteration has been shown to be an early event in prostate cancer initiation and progression. Moreover, Pten-null mice that develop PIN have among the valuable animal models in prostate cancer research [64]. Kalaany et al. showed that 40% dietary restriction did not have any detectable e ffect on the extent or histological appearance of the PIN in Probasin-Cre; PTEN L/L prostate cancer models but significantly reduced tumor nodules in the lungs of K-RASLA2; P53 LSL/WT lung adenocarcinoma models [23], suggesting that the Phosphoinositide 3-kinase (PI3K)/ protein kinase B (AKT) pathway is critical for diet-induced cancer progression. Conversely, a high-calorie diet (45 kcal% fat) promoted prostate cancer progression in genetically susceptible Pten haploinsu fficient mice with increasing inflammatory response in the presence of enhanced insulin response to chronically elevated insulin levels [40]. Hayashi et al. demonstrated that mice receiving a HFD for 17 weeks starting from an age of 5 weeks had significantly higher prostate weights of than those receiving control [54]. Moreover, HFD-fed model mice had a significantly higher Ki67-positive cell to tumor cell ratio than control mice, while no marked di fference

in glandular structures was observed between the control diet (CD)-fed and HFD-fed model mice [54]. An interesting study involving the basal cell-specific Pten-null model using K14-Pten-mTmG mice showed that HFD intake promoted the initiation and progression of PIN lesions [48]. Although dietary fat could potentially be associated with prostate cancer development of Pten-null mice, the impact may not be extensive. Additionally, the evaluation of prostate pathology in GEMMs needs to be standardized according to the Consensus Report from the Bar Harbor Meeting of the Mouse Models of Human Cancer Consortium Prostate Pathology Committee for accurate comparison among different studies [65].

With regard to other GEMMs, PTP1B (PTPN1), an androgen-regulated phosphatase, acts as a HFD-dependent tumor suppressor in prostate cancer driven by the absence of Pten, such as in the Pten-/-Ptpn1-/- mice model [47]. Deficiency in RXRα (a unique and important member of the nuclear receptor superfamily) in the prostates of mice receiving a new Western-style diet resulted in higher rates of mPIN and prostate cancer [30]. Pommier et al. showed that mice with double knockout of Liver X receptors (LXRa and LXRb), which belong to the nuclear receptor superfamily and are central mediators of cholesterol homeostasis, developed PIN under a diet high in cholesterol [36].

Reports regarding HFD-induced metastatic models using GEMMs have been rare. Chen et al. showed that among mice with Pten deletion and a double deletion of Pten and Promyelocytic Leukemia (PML), a suppressor of pp1α-dependent activation of MAPK signaling, those receiving a lard-based HFD displayed lymph node metastasis and lung metastasis, whereas those receiving a chow diet exhibited limited metastases [56].

Taken together, GEMM studies showed that a HFD enhanced tumor growth through the modulation of several genes, including those related to PTEN. Studies that assess the impact of a HFD using more aggressive, metastatic GEMMs while considering the effect of dual and/or multiple genes may be intriguing.
