US National Cancer Institute–China Collaborative Studies on Chinese Medicine and Cancer

Abstract

Since 2007, the US National Cancer Institute (NCI) Office of Cancer Complementary and Alternative Medicine (OCCAM), together with the Cancer Institute of the China Academy of Chinese Medical Sciences (CICACMS), institutes at China Academy of Sciences and Chinese Academy of Medical Sciences, have engaged in collaborations on Chinese medicine (CM) and cancer research. Through these collaborations, CM drugs and compounds have been studied at NCI labs. This paper summarizes the discoveries and progress on these research projects, exploring the aspects of cancer prevention, botanical drug mechanisms of action and component analysis/quality control (QC), and anticancer activity screening. These and other related projects have been presented in various jointly convened workshops and have provided the backdrop for establishing a new organization, the International Consortium for CM and Cancer, to promote international collaborations in this field.

The National Cancer Institute (NCI) is interested in exploring different types of complementary and alternative medicine (CAM) interventions with regard to cancer prevention, treatment, and symptom management. Chinese medicine has a long history over thousands of years, and it is a completely developed medical system encompassing physiology, pathology, prevention, diagnosis, and treatment of diseases including cancer. PubMed records more than 20 000 publications on CM and cancer since 1990. In order to systematically understand CM regarding its theory, practice, development, and limitation and communicate with the CM community in the field of cancer, in April 2006 OCCAM hosted a conference titled “Traditional Chinese Medicine and Cancer Research: Fostering Collaborations; Advancing the Science” at the National Institutes of Health (NIH), which almost 200 scientists and physicians attended, including more than 40 from China (1). This conference served as an incubator for establishing new collaborative relationships between Chinese medicine and Western medicine practitioners and researchers for cancer prevention, treatment, and palliation. As a result of this conference, new collaborations have been established between OCCAM, various intramural NCI laboratories, and CICACMS, as well as other institutes in China. From January 2007 to November 2016, seven fellows from CICAMS came to NCI Center for Cancer Research (CCR) labs to study the mechanisms of action of certain CM herbal formulations and purified natural products from CM herbs used in China to treat patients with cancer.

Two CM anticancer herbal medicines clinically used in China were studied in the NCI intramural research program by Drs. Joost Oppenheim and Zack Howard at the Laboratory of Molecular Immunoregulation, CCR, along with visiting fellows from a hospital associated with CACMS, Guanganmen Hospital.

Sheng Qi Formula

Sheng Qi formula (SQF) contains hydrophilic herbal extracts of Astragalus mongholicus Bge (91.8%) and Panax notoginseng (8.2%) and is used clinically to reduce chemotherapy side effects and enhance the efficacy of conventional cancer therapy (2). Drs Jie Li and Huiting Fan from CICACMS worked on this project with Drs Zack Howard and Joost Oppenheim of CCR, NCI. Standardized and genotyped root of Astragalus mongholicus Bge (Huang Qi) in dried form and preeminent grade, grown in the Neimenggu Province, and the powder of Panax notoginseng (San Qi), grown in the Yunnan Province of mainland China, were purchased from Da Xing Chinese herbal medicines store (Washington, DC); 450 grams of Huang Qi was homogenized with a Waring blender, and then soaked in six liters of double-distilled water for 24 hours. Following this soaking, the preparation of swollen roots was boiled for one hour. The decoction was then filtered through a coarse (3 mm) filter, and roots were reboiled with the same amount of water. This step was repeated twice. Collected filtrates were then pooled and thoroughly mixed with San Qi powder (a dried hot water extract) to form SQF mixture, which was then filtered through Whatman #1 filter paper. The dissolved solids of SQF are in solution/suspension form for testing. All in vitro experiments on SQF (and FKI, see below) were performed at least three times, and the means ± SEMs of representative results are presented. All animal studies in this paper were in accord with the NIH animal study guidelines. Significant differences among the groups were determined using two-way analysis of variance. A comparison of the rate of animal survival was analyzed by Kaplan-Meier survival analysis method. P values of less than .05 were considered statistically significant.

The collaborative studies have shown that SQF can slow 4T1 inflammatory breast tumor growth and prolong survival time when given orally to mice alone or combined with chemotherapeutic drugs such as paclitaxel, cyclophosphamide, and gemcitabine. One mechanism of action of SQF is the induction of apoptosis through the mitochondrial pathway. SQF delivered in the drinking water of Balb/c mice bearing 4T1 inflammatory breast tumors resulted in reduction in all the following factors: tumor growth, numbers of tumor-induced myeloid-derived suppressor cells (MDSC), functionality of the remaining MDSC, and circulating levels of IL-1 and G-CSF. Combination therapy with SQF and paclitaxel or cyclophosphamide resulted in additive reduction in tumor growth and mixed effects on MDSC. In the 4T1 inflammatory breast tumor model, 100% of tumor-challenged but untreated animals developed tumors by day 32. Similarly, 90% of mice receiving only gemcitabine had developed tumors by day 32. In contrast, only 30% of mice receiving both gemcitabine and SQF had developed tumors by day 32.

A liquid chromatography (LC)/mass spectrometry (MS) method was developed to quantify saponins in SQF. In vitro studies of ginsenoside Rg1, Re, Rb1, Rd, and astragaloside II (AsII) alone and in combination demonstrated that the saponins are not the only functional components of SQF. LC/MS standardization determined functional quantities of these saponins: Per 100 g SQF, it contains: Ginsenoside Rg1 100 ± 5 mg; Ginsenoside Rb1 80 ± 3 mg; Ginsenoside Rd 25 ± 3 mg; Ginsenoside Re 15 ± 2 mg; Astragaloside II 7 ± 1 mg. Combining both a chemical fingerprint of saponins found in active SQF and quantitative polymerase chain reaction (PCR) of cytokines induced by SQF treatment of hepatocyte cell culture resulted in an in vitro assay that was predictive of in vivo efficacy. Rg1, Re, Rb1, Rd, and AsII have been chosen as fingerprint chemicals because of their degree of separation in LC/MS. Retrospective studies have correlated the chemical fingerprint of batches of SQF extract and tumor growth characteristics of tumors in mice treated with SQF. The antitumor activities of the individual chemical constituents of SQF are not clear and need to be tested in the future. In vivo metabolic studies of Balb/c-naïve and tumor-bearing mice treated with SQF showed that SQF has a profound effect of glycolysis and fatty acid metabolism (3,4).

Fufang Kushen Injection

Fufang Kushen injection (FKI; a standardized commercial Chinese herbal medicine from Shanxi Zhendong Pharmaceutical, Shanxi, China) is a mixture of extracts from the roots of two herbs—Sophora flavescens and Rhizoma Smilacis glabrae. FKI is used alone or combined with conventional chemotherapy to treat lung and other cancers and to relieve cancer-related pain. In order to fully explore the preclinical effects of FKI, gain a better understanding of the mechanism of action of FKI, and determine effective dosing in combination with chemotherapy, Dr. Zack Howard of CCR/NCI and Dr. Zhizheng Zhao, visiting fellow from CICACMS, began a systematic study of FKI. FKI was found to reduce tumor growth and tumor-induced hyperalgesia in a murine sarcoma model. A summary of the results of this work is as follows: 1) FKI has a selective in vitro cytotoxic effect on various sarcoma cell lines; 2) low-dose FKI (25 mg/kg) reduces tumor-induced temperature sensitivity; high-dose FKI (50 mg/kg) reduces tumor-induced temperature sensitivity and sarcoma growth; 3) FKI’s effect on hyperalgesia involves the pain sensor pathway protein TRPV1 and ERK phosphorylation; 4) in an animal model, twice-a-day dosing of FKI was more effective than single dosing (5). This research was supported in part through a Cooperative Research and Development Agreement (CRADA) between NCI and the Cancer Institute, CACMS.

Studies of Purified Compounds Derived From Chinese Herbal Medicines

Berberine

Dr. Weidong Li from CICACMS has worked with Drs Nancy Colburn and Matthew R. Young at the Gene Regulation Section in the Laboratory of Cancer Prevention/CCR at the NCI to study the effects of berberine and other compounds extracted from CM herbs on colon cancer prevention. In these studies, berberine (Molecular Weight [MW] = 336.36 g/mol., >90% purity by HPLC, provided by China National Institute for the Control of Pharmaceutical and Biological Products), a natural compound extracted from the herbal plant Rizoma coptidis that has been used in Chinese medicine for many years, was found to suppress colon epithelial proliferation and tumorigenesis by activating AMP-activated protein kinase (AMPK) and inhibiting mammalian target of rapamycin (mTOR) activity in a dose-dependent and time-dependent manner. Berberine inhibited the growth of three different colorectal cancer cell lines (HCT116, SW480, and LOVO). The downstream targets of mTOR–4EBP1 and p70S6 kinase were also downregulated by berberine treatment in three colon cancer cell lines. In the azoxymethane-initiated/dextran sulfate sodium (AOM/DSS)–induced mouse model of colitis-induced colorectal cancer, berberine administration activated the AMPK signaling pathway, inhibited NF-κB, and reduced significantly the number and size of tumors compared with vehicle-treated mice. This study demonstrates that berberine might offer a safe and promising candidate for chemoprevention of colon cancer and other metabolic disorders that might be regulated by AMPK, such as type 2 diabetes (6,7).

Resveratrol

Mr. Shakir M. Saud, Drs Weidong Li, Nancy Colburn, and Matthew Young, and other members of the Laboratory of Cancer Prevention at the NCI also jointly studied resveratrol, another botanical compound. Resveratrol (MW = 228.25 g/mol, product from Sigma-Aldrich, Inc., Catalog Number R5010, >99% [HPLC], CAS Number 501-36-0, PubChem Substance ID 24278055) is a naturally occurring polyphenolic compound found in various plants including berries and red wine grapes, and it has been found to have broad-spectrum beneficial health effects including anti-oxidant, anti-inflammatory, and anticancer activities. Using a genetically engineered mouse model of sporadic colon cancer with an active Kras, it was found that feeding mice a diet supplemented with resveratrol before tumors were visible by colonoscopy resulted in 60% inhibition of tumor production, and in the 40% of mice that did develop tumors, Kras expression was lost. Feeding mice a resveratrol-containing diet after tumors developed resulted in a complete remission in 33% of these mice. In the remaining mice, the average tumor size was 97% smaller than that in mice fed the control diet. The expression of miR-96, a Kras regulator, was increased in nontumoral and tumoral colonic tissue of resveratrol-treated mice, suggesting that resveratrol might prevent the formation and growth of sporadic colorectal cancer in mice by downregulating Kras expression (8).

Cryptotanshinone

A collaborative study was established between Dr. William Farrar’s group at the Laboratory of Cancer Prevention/CCR of NCI and CICAMS using compounds isolated from Chinese medicinal herbs to target human prostate cancer cells and prostate cancer–initiating cells.

Dr. Farrar’s group and visiting fellow Dr. Ying Zhang studied cryptotanshinone (CT) (MW = 296.36, > 99 purity by HPLC, provided by the China National Institute for the Control of Pharmaceutical and Biological Products), a compound derived from the herb Salvia miltiorrhiza (Danshen), and found that it could significantly affect various cellular characteristics including cellular proliferation, cell cycle status, migration, viability, colony formation, and sphere formation of prostate tumor–initiating cells. CT was found to target the LNCaP CD24+ CD44+ cell population that is representative of prostate tumor–initiating cells and downregulate stemness genes, such as Nanog, Oct4, Sox2, β-catenin, and CXCR4 in prostate tumor–initiating cells (9).

In a separate group of in vitro studies, Drs Weidong Li, Matthew R. Young, Nancy Colburn, and Baojin Hua and Mr. Shakir M. Saud determined the effects cryptotanshinone (CPT) on colorectal cancer cell proliferation and growth. These studies showed that CPT inhibited the growth and cell viability of multiple colorectal cancer cell lines (HCT116, SW480, and LOVO). CPT inhibited activation of signal transducer and activator of transcription 3 (STAT3), a transcription factor that mediates expression of many genes known to regulate tumor growth (10).

Other Compounds

In 2008, OCCAM started collecting pure compounds and medicinal herb extracts through international collaborations performing various collaborative research agreements with institutes in China. Up to 349 pure compounds and 200 extracts have been collected from three institutes in China (Kunming Institute of Botany; Key Laboratory of Chemistry for Natural Products [KLCNP] in Guizhou Province; and Institute of Matera Medica, Chinese Academy of Medical Sciences).

Seventy-four pure compounds have been screened (one-dose, 10⁻⁵ Mole) on NCI 60 human cancer cell lines (11). When these cancer cell lines’ growth is inhibited by compounds at this level, it will be screened further at more widely concentrated levels; 14 of them showed inhibition at this level and entered five-dose (10⁻³, 10⁻⁴, 10⁻⁵, 10⁻⁶, 10⁻⁷ Mole each) screens. Also, 26 compounds and 152 medicinal herb extracts have been studied in various cell target assays [GP78 (12), PLK1 (13), NF1 (14), EpCAM (15), MALT1 (16), META (17), SUMO (18), TRAIL (19), p38 (20)] with certain hits.

International Consortium for Chinese Medicine and Cancer

In 2011 and 2013, OCCAM and CICACMS cohosted two international conferences in Beijing, China, on CM and cancer research. In keeping with the 2006 conference mentioned above, these meetings continued to explore the intersection of CM and Western medicine in cancer research. With attendees from the United States, China, South Korea, Australia, Taiwan, Singapore, and Hong Kong, these meetings further established the potential value of more dialogues among scientists and health care practitioners from different countries and areas, as well as the potential for international scientific collaborations to aid in the modernization of CM for oncology applications.

In February 2014, CICACMS presented OCCAM with a proposal for an international consortium that would continue the work of improving scientific communications and collaborations, improving the quality of research on CM for oncology applications, finding potential promising CM drugs, and modernizing the research of CM for cancer. OCCAM agreed to work with CICACMS to explore the feasibility of establishing an international consortium for Chinese medicine and cancer.

On November 3, 2014, at the NIH campus in Bethesda, Maryland, OCCAM held a meeting to examine the potential utility and feasibility of establishing an International Consortium for Chinese Medicine and Cancer (ICCMC). At the meeting, participants from China, the United States, Canada, Taiwan, Australia, and South Korea discussed issues in Chinese medicine and cancer research, treatment, and management, including potential mechanisms of action, proof of efficacy, side effects, regulatory issues, and the need for improving the quality of randomized clinical trials of CM treatments and supportive care interventions (21). A follow-up meeting was held in Dalian, China, on October 16–18, 2015, and subsequently a development committee was established for the ICCMC. The ICCMC will be an international CM and Cancer research platform to promote and enhance basic research and clinical trials, as well as symptom management of cancer all over the world.

Some proposed initial projects for the consortium include 1) developing guidelines for CM oncology clinical trial design, implementation, and reporting; 2) developing standard treatment protocols and evaluation procedures; 3) quantification of phytoestrogens from CM herbs; 4) developing curricula to standardize oncology education for CM physicians; and 5) starting an electronic journal on CM and cancer.

Conclusions

Collaborations between the US NCI and various institutes in China have enhanced the dialog and understanding between two medical systems—Chinese medicine and Western medicine—on cancer prevention, treatment, and symptom management. These joint efforts have provided opportunities for scientists and clinicians of both countries to explore systemically the components of CM herbal extracts and compounds for their potential activities and applications either alone or in combination with Western medicine with regards to cancer. The anticancer mechanisms of action of certain of herbal medicines and their components have been elucidated through such collaborative projects. Other studies have confirmed, in preclinical models, the clinically observed analgesic effects of the herbal mixture FKI. Findings such as these from SQF, FKI, berberine, resveratrol, and cryptotanshinone studies may lead to further work to optimize the demonstrated effects and better determine how to integrate these therapies with Western medicine approaches to cancer prevention and management.