Background: Lifestyle factors are important determinants for the development and progression of colorectal cancer (CRC) (1). An estimated 50-60% of new CRC cases and deaths in the U.S. could be potentially prevented through modification of lifestyle, including diet (2). Fish oil, a rich source of marine omega-3 polyunsaturated fatty acid (MO3PUFA), is one of the most popular natural dietary supplements used by U.S. adults (3). MO3PUFA has potent anti-inflammatory activity, and substantial experimental data support its beneficial effect for CRC prevention and treatment (4). However, the relevance of these findings to human cancer has been largely unknown. I have led a series of studies linking MO3PUFA to CRC, leveraging data from two large prospective cohort studies, the Nurses’ Health Study (NHS) and the Health Professionals Follow-up Study (HPFS) that have provided detailed information on lifestyle and diet in relation to CRC.

Study setting: The NHS cohort enrolled 121,701 female nurses aged 30-55 years in 1976, and the HPFS cohort enrolled 51,529 men aged 40-75 years in 1986. In both studies, we have followed up participants through a biennial questionnaire to collect their updated lifestyle and medical information. We have also collected dietary data every 4 years through the food frequency questionnaire, which has been validated for MO3PUFA assessment. CRC diagnoses are first self-reported by participants on the questionnaires and then confirmed through review of medical records. We also retrieve formalin-fixed paraffin embedded (FFPE) tissue blocks from hospitals where participants undergo surgical resection. We identify deaths through family members or the postal system in response to the follow-up questionnaires as well as by search of the National Death Index. Study physicians confirm the cause of death from death certificates or review of medical records. Taken together, the two cohorts provide an ideal setting for studying the relationship of long-term MO3PUFA intake with CRC, due to the large sample size, prospective design with more than 20 years of follow-up, repeated dietary assessments using validated instruments, and detailed data on additional cancer risk factors. Moreover, by integrating the rich epidemiologic data with the tumor molecular features characterized in the FFPE tissue, I am able to study how MO3PUFA is associated with specific subtypes of CRC arising from distinct mechanisms.

Main findings: In our first study (5), we showed that the relationship between MO3PUFA and CRC risk differed according to anatomic subsite of the tumor. While MO3PUFA intake was not associated with incidence of distal colon cancer, a nonsignificant protective association was found for proximal colon cancer and rectal cancer. Moreover, we found that high MO3PUFA intake assessed 12-16 years before diagnosis tended to be associated with lower CRC risk, suggesting that MO3PUFA may start exerting its protective effect at an early stage of CRC development. These findings provided thus far the most comprehensive data about the MO3PUFA-CRC relationship.

Given the large variation of genetic and epigenetic alterations in cancers along anatomic subsite, we further examined whether the differential association of MO3PUFA observed in proximal versus distal colon tumors was attributable to differences in tumor molecular features. Through molecular pathologic assessment of tumor specimens, we classified cases into distinct subtypes according to common CRC molecular characteristics. By linking MO3PUFA intake to these CRC subtypes (6), we found that high MO3PUFA intake was primarily associated with lower risk of microsatellite instable (MSI) tumors, a CRC subtype arising from defective DNA mismatch repair mechanisms with predominance in the proximal colon, whereas no association was found for microsatellite-stable (MSS) tumors. These findings are consistent with experimental data supporting the involvement of inflammation in MSI development (7,8), and generate novel hypotheses for the potential anticancer activity of MO3PUFA.

Considering the long-recognized role of MO3PUFA in the immune system and the close link between immune dysregulation and MSI, we further assessed whether the variation in tumor immune infiltrates might contribute to the preferential benefit of MO3PUFA for MSI tumors (9). We first quantified the density of CD3+, CD8+, CD45RO+, and FOXP3+ T cells in the tumor microarrays by immunohistochemistry and Ariol image analysis system. We then classified CRC cases into subtypes with low-level versus high-level infiltrate of each of the T-cell markers, and tested whether the association of MO3PUFA intake with CRC risk differed according to T-cell densities in tumor tissues. We found that the beneficial associations of high MO3PUFA intake with CRC appeared confined to tumors with high infiltrate of FOXP3+ T cells. Although FOXP3+ T cells were more enriched in MSI tumors, the inverse association of MO3PUFA with FOXP3+ T-cell-high tumors was independent of the MSI status, suggesting that MSI may be a surrogate marker for tumors with high FOXP3+ T-cell infiltration. FOXP3 is a prerequisite transcription factor for T regulatory (Treg) cells, which have potent immunosuppressive activity and may facilitate tumor evasion by suppressing T effector cell-mediated antitumor immune surveillance (10). Therefore, it is possible that MO3PUFA may protect against development of CRC, especially those with MSI, by ameliorating Treg-mediated immune suppression and unleashing antitumor immunity. Indeed, this hypothesis is supported by our functional experiment, in which MO3PUFA decreased the T-cell suppressive activity of intestinal FOXP3+ Treg cells (9).

Given the increasing data linking the tumor immune response to cancer prognosis, I led further studies assessing the relationship of MO3PUFA intake with CRC survival. We found that patients with high MO3PUFA intake after CRC diagnosis had a significantly longer survival than those with low intake 11), suggesting that MO3PUFA may exert a beneficial effect across the continuum of colorectal carcinogenesis. We have recently validated these findings in the cohort of CALGB 89803 (Van Blarigan EL et al., manuscript in preparation).

Future plan: Based on this work, I now propose to better understand the intricate relationship between MO3PUFA, immune response, and CRC through the development of an independent research program. First, given the increasing data supporting a pivotal role of gut microbes in integrating dietary cues with host immunity and potentially mediating the anticancer effect of MO3PUFAs, I have proposed to study how MO3PUFAs modulate the gut microbial composition and function, thereby shaping the gut immune response and suppressing CRC. The findings from this study will provide translational insights into novel dietary and microbial strategies for CRC prevention. Second, to better understand the immunomodulatory effect of MO3PUFA in the tumor microenvironment, I plan to characterize the MO3PUFA-asssociated tumor immune signature and study whether MO3PUFA improves CRC survival by ameliorating the immunosuppressive microenvironment. Because the immunosuppressive mechanisms have been implicated in the development of resistance to powerful cancer immunotherapeutic agents, our findings may open new avenues for incorporating natural products into existing cancer immunotherapy paradigms for optimized clinical benefit. In both these areas, I will take an integrated approach that combines the power of population science and clinical research to establish causality and facilitate clinical translation. This approach will also provide a foundation for my research program as I transition into independence.

References

1. Song M, Garrett WS, Chan AT. Nutrients, foods, and colorectal cancer prevention. Gastroenterology 2015;148(6):1244-60.

2. Song M, Giovannucci E. Preventable incidence and mortality of carcinoma associated with lifestyle factors among white adults in the United States. JAMA Oncol 2016;2(9):1154-61.

3. Kantor ED, Rehm CD, Du M, White E, Giovannucci EL. Trends in dietary supplement use among US adults from 1999-2012. JAMA 2016;316(14):1464-74.

4. Cockbain AJ, Toogood GJ, Hull MA. Omega-3 polyunsaturated fatty acids for the treatment and prevention of colorectal cancer. Gut 2012;61(1):135-49.

5. Song M, Chan AT, Fuchs CS, et al. Dietary intake of fish, omega-3 and omega-6 fatty acids and risk of colorectal cancer: A prospective study in U.S. men and women. Int J Cancer 2014;135(10):2413-23.

6. Song M, Nishihara R, Wu K, et al. Marine omega-3 polyunsaturated fatty acids and risk of colorectal cancer according to microsatellite instability. J Natl Cancer Inst 2015;107(4);djv007.

7. Llosa NJ, Cruise M, Tam A, et al. The vigorous immune microenvironment of microsatellite instable colon cancer is balanced by multiple counter-inhibitory checkpoints. Cancer Discov 2015;5(1):43-51.

8. Colotta F, Allavena P, Sica A, Garlanda C, Mantovani A. Cancer-related inflammation, the seventh hallmark of cancer: links to genetic instability. Carcinogenesis 2009;30(7):1073-81.

9. Song M, Nishihara R, Cao Y, et al. Marine omega-3 polyunsaturated fatty acid intake and risk of colorectal cancer characterized by tumor-infiltrating T cells. JAMA Oncol 2016;2(9):1197-1206.

10. Betts G, Jones E, Junaid S, et al. Suppression of tumour-specific CD4(+) T cells by regulatory T cells is associated with progression of human colorectal cancer. Gut 2012;61(8):1163-71.

11. Song M, Zhang X, Meyerhardt JA, et al. Marine omega-3 polyunsaturated fatty acid intake and survival after colorectal cancer diagnosis. Gut 2016. [Epub ahead of print.]

Citation Format: Mingyang Song, Kana Wu, Shuji Ogino, Jeffrey A. Meyerhardt, Charles S. Fuchs, Wendy Garrett, Edward L. Giovannucci, Andrew T. Chan. Marine omega-3 polyunsaturated fatty acid and colorectal cancer prevention and treatment [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr NG03. doi:10.1158/1538-7445.AM2017-NG03