Curcumin and Gingerol Protocol for MDS

Objective

To test the efficacy of the natural compounds curcumin and gingerol in improving the cytopenias of patients with myelodysplastic syndromes.

Background and Rationale

Unlike the past era of chemotherapeutic agents with a broad spectrum of activity, the types of drugs being developed for cancer treatment today have a specific and well defined target and are less toxic than previously used chemotherapies. With improved enabling technologies and cumulative knowledge of molecular genetics, immunology, pharmacology, and experimental therapeutics, translational research is no longer a pie in the sky. With the identification of specific biologic targets, it is also possible to consider the use of some of the natural, non-toxic compounds that have known activity against the targets. Another inflection point is represented by the tendency to test the newer agents in early rather than end stage disease, and to use surrogate molecular/biochemical markers to document tumor progression or regression.

The myelodysplastic syndromes (MDS) represent one of the most rapidly evolving areas of translational research in cancer today. At least a third of these patients eventually transform to acute leukemia, but while they are in the MDS state, they represent an early form of acute leukemia where novel targeted therapies as well as the role of non-toxic natural compounds can be tested. In the current proposal, we intend to test the efficacy of two natural compounds, curcumin and gingerol in patients with MDS. The rationale for this proposed therapy can be best understood if we begin by examining the biology of MDS and the novel therapeutic targets that have been recently identified in this disease. This will be followed by a detailed description of the chemopreventive and biologic properties of curcumin and gingerol that could be exploited for use against the therapeutic targets identified in MDS.

As the name implies, MDS is not a single disease but a collection of heterogeneous syndromes that share in common, a dysplastic morphology and the clinical presentation of variable cytopenias despite a generally cellular bone marrow (BM). Consequently, no single drug is likely to be of universal benefit for all MDS patients. The focus of clinical research has been directed at identifying subgroups of patients who might benefit from a given treatment strategy. Early attempts used cytogenetic abnormalities to stratify patients into subcategories of biologic homogeneity, however, these could not be translated into clinical successes in any consistent manner. Subsequent molecular and biologic studies have identified a number of potential signaling pathways that may serve as excellent therapeutic targets.

Pathology of Cytopenia in MDS

MDS is a monoclonal disease in which an early BM stem cell is the site of transformation, giving rise to daughter cells at least some of whom retain the ability to undergo terminal differentiation, but do not always make it out of the marrow to the periphery. The clonal nature of these disorders has been repeatedly demonstrated (1-7).  The conceptual conundrum of pancytopenia in the presence of a hypercellular marrow was addressed initially by detailed studies of proliferation in a large number of MDS patients. Following infusions of the thymidine analogs iodo- and bromodeoxyuridine, it was demonstrated that in fact there are more cells synthesizing DNA in MDS marrows and cycling faster than both cells of normal acute myeloid leukemia (AML) patients (8). The fact that these patients experience a variable cytopenia therefore could not be explained on the basis of a failing BM. If anything, the hematopoietic cells appear to be hyper-proliferative in MDS. It was then hypothesized that it may be a loss of cells via premature death that accounts for the ineffectiveness of hematopoiesis (9). In fact, using a variety of techniques to document the death of cells, it was next shown that there is evidence of excessive intramedullary apoptosis of hematopoietic cells in the majority of MDS marrows, and that in some patients, this can affect more than half the marrow cells at any given time (10,11). Some unique features of this apoptosis include the observation that parenchymal cells belonging to all three lineages and at every stage of maturation are found to be apoptotic in MDS (11).  S-phase cells can simultaneously undergo apoptosis in MDS and this phenomenon has been referred to as “signal antonymy” (12). An interesting association was found between signal antonymy and an alteration in E2F1 transcript size versus BM cellularity (13).    MDS patients with a hypercellular BM had low signal antonymy and a normal sized E2F1 transcript size while those with hypocellular BM had a high level of signal antonymy and a shorter E2F1 mRNA in addition to a normal sized one (14). 

Mediators of Apotosis in MDS

A role for pro-inflammatory cytokines was suspected in the excessive apoptosis since the pro-inflammatory cytokines tumor necrosis factor alpha (TNF-a), transforming growth factor beta (TGF-b), interleukin-1 beta (IL-1b) were found to be increased in the majority of MDS patients (19-29). It is possible that the pro-inflammatory cyotkines exert a differential effect on the immature versus maturing cells, since the MDS clone could not predominate in the marrow unless it was relatively resistant to pro-apoptotic and inhibitory actions of cytokines. TNF-a appears to be acting as the master switch responsible for turning the inflammatory cascade on or off, which makes it a possible therapeutic target. A 145 patient study in MDS showed high correlation between TNF-aand TGF-b; and between macrophages, TNF-a, and high levels of apoptosis (26). Based on these observations, the cytokine profile in MDS appears more like that seen in chronic inflammatory diseases such as Crohn’s disease or rheumatoid arthritis. It is therefore logical to try anti-TNF/anti-inflammatory therapy similar to that used in these other diseases. Thus, biologic characteristics of MDS include: excessive proliferation in bone marrow, which is matched by excessive apoptosis, mediated by proinflammatory cytokines. 

Angiogenic Activity in MDS Marrows

It is interesting to note that the maximum supply of blood to the bone marrow has been shown to be present in patients with MDS and myelofibrosis. This has been accomplished through total body arteriograms using nuclear medicine techniques. Our own studies have found excessive levels of two angiogenesis related cytokines TNFa and transforming growth factor beta (TGFb) to be  greatly increased in the marrows of the majority of MDS patients. Given these findings, it is logical to consider agents that would inhibit TNFa and TGFb for the treatment of MDS. Furthermore, clonal expansion in MDS is not an isolated activity, but arises from an interaction between the cells and their microenvironment. Both components of the immune system and blood vessels along with stromal cells and macrophages contribute to this vascularization. Increased neo-vascularization, increased micro-vessel density and particularly, increased vascular endothelial growth factor-A (VEGF-A) expression have been demonstrated in MDS marrows (30-31). The molecular processes that control angiogenesis are comprised of a balance between pro- and anti- angiogenesis factors. Cytokine production by both the stromal cells and the MDS cells stimulate endothelial growth. One of the key stimulatory cytokines as already mentioned, is VEGF, which controls proliferation, differentiation and survival of the endothelial cells. The high levels of VEGF found in many tumors, accompanied by receptor expression on the adjacent endothelial cells, results in continuous angiogenesis stimulation (32). The cell adhesion molecules, especially the integrins (cell surface receptors of the extracellular matrix), not only mediate migration of endothelial cells, they also transduce intracellular signals that control cell proliferation and survival.

Increased NFkb Activity in MDS

Another pivotal regulatory protein in the cell is the transcription factor NF-kB. Transcription factors bind to DNA, initiating the transcription of genes, which ultimately leads to the production of proteins encoded by those genes. NF-kB regulated gene products include various growth factors (IL6, VEGF), cell adhesion molecules (intracellular adhesion molecule-1 or ICAM-1, and VCAM-1) and anti-apoptotic factors which together play an important role in promoting the growth of transformed cell and its progeny. NF-kB activation in transformed cells prevents it from undergoing apoptosis (33-35). In quiescent cells, NF-kB is located in the cytoplasm in an inactive form, bound to the molecule called “inhibitor of NF-kB” (IkB). Stimulation of cells by a variety of mechanisms such as viruses, growth factors, antigens, or chemotherapeutic drugs, triggers a cascade of signaling events that ultimately results in the degradation of IkB by the 26S proteasome (36). This releases NF-kB, which then translocates into the cell nucleus, where it binds to specific DNA sequences at its target genes. Target genes are transcribed, resulting in protection against apoptosis. The proteasome inhibitor PS-341 now known as Velcade, inhibits the proteasome, preventing its degradation of the ubiquitinated NF-kB-IkB complex, and thus abrogating the effects of NF-kB on growth, proliferation and gene expression (37). This inhibition of the proteasome leads to transformed cell cycle arrest and apoptosis (38). We have used Velcade in MDS patients and found encouraging results with both ant-leukemic activity and improvement in the variable cytopenia of a subset of patients (39).

Rationale for Using Curcumin and Gingerol in MDS

There are several important biologic and clinical reasons to expect encouraging responses with curcumin and gingerol use in MDS. The disease predominates in an elderly population, the median age being approximately 70 years. Many of these patients also have numerous co-morbid conditions which make the use of natural compounds with little or no toxicity highly desirable. The disease also follows a chronic and insidious phase in many patients, and only a third of the patients actually progress to acute leukemia. Despite this less aggressive course of the disease, it is an incurable and fatal illness for which a stem cell transplant is the only potential curative option. It is conceivable that intervention with natural compounds in patients whose disease is marked mostly by a variable cytopenia may not only improve the counts, but may also retard the progression of the disease towards acute leukemia. Equally important are the recent insights into the biology of the disease which suggest that therapies directed at suppression of cytokines such as TNFa and TGFb, anti-angiogenic and immune modulatory approaches can be highly successful in producing complete and durable responses in subsets of MDS patients. A prime example is the use of thalidomide in MDS, producing a ~20% response rate by virtue of its ant-TNF, anti-angiogenic and immunemodulatory effects, and then the more recent use of the thalidomide analog Revlimid which produced complete hematologic and cytogenetic remissions in almost all MDS patients with a 5q- abnormality, and half of those without (40,41). Our group has also used the mitochondrially coded coenzyme Q10 (coQ10) in high doses of 1200mg/day for 6-12 months on the basis of finding mt-DNA mutations and found that 7/28 patients with low or intermediate-1 risk MDS achieved hematologic improvement. Furthermore, as already mentioned above, inhibition of the transcription factor NFkB has also been found to be effective in a subset of MDS patients. Based upon these clinical and biologic observations, we propose to explore the efficacy of the natural compounds curcumin and gingerol in MDS because these two agents possess many of the effects that are desirable in MDS as described below.

Curcumin

Diets containing garlic, onion, soy, turmeric, ginger, tomatoes, green tea and chillies that are common in

Asia  are associated with a lower risk of a variety of cancers ranging from colon, GI tract, breast, leukemias and lymphomas. Some of these dietary agents have now been extensively studied for their biologic activities and as potential chemopreventive agents for cancer. Curcuma longa or turmeric, responsible for the yellow color of curry powder, is a herb belonging to the ginger family and curcumin is its most active component. Turmeric has been widely used in India for centuries as a panacea for a variety of ailments and curcumin has received greater attention as the active component with over 1000 publications in the Medical archives describing the chemistry, pharmacology and its mechanism of action. In summary, curcumin has been found to have the following properties (42,43):

• Anti-TNF and anti-TGFb
• Suppression of transcription factors such as NFkB and AP-1
• COX-2 inhibition
• Anti-angiogenic
• Glutathione transferase Pi inhibition
• Anti-proliferative
• Pro-apoptotic
• Protease inhibition
• Sensitization of cells to chemo and radiation therapies

Gingerol

Plants of the ginger family have been credited with therapeutic and preventive powers and have been reported to have anti-cancer activity. The substance called [6]-gingerol is the main active compound in ginger root and the one that gives ginger its distinctive flavor. Plants of the ginger (Zingiber officinale Roscoe, Zingiberaceae) family, one of the most heavily consumed dietary substances in the world, have been shown to inhibit tumor promotion in mouse skin. The oleoresin from the root of ginger contains [6]-gingerol, the major pharmacologically active component and lesser amounts of a structurally related vanilloid, [6]-paradol. At least two recent studies suggest that these compounds suppress proliferation of human cancer cells through the induction of apoptosis (44,45). A review of recently published studies indicate the following properties of gingerol (46-48):

• Gingerol is anti leukemic by inducing apoptosis in leukemia cells
• Acts as an anti-bcl-2 agent
• Can prevent the development of colon cancer cells
• Inhibits EGF-induced transformation
• Protects against radiation induced lethality
• Can act as a blood thinner via platelet activation inhibition

Structure of Curcumin and Gingerol Compared to Other Chemopreventive Agents

Clinical Trials with Curcumin and Gingerol

At least 9 clinical studies with curcumin have now been reported in humans in diseases ranging from cancer to rheumatism, uveitis, inflammatory diseases, leukoplakia, metaplasia of the stomach, and as cholesterol-lowering agents. All studies show that curcumin is extremely well tolerated in doses ranging from 4-8 grams/day, although up to 12 Gm/day have also been administered. Clinical responses of varying degrees have been reported in almost all of these clinical trials (for a review, see ref 43). Similarly, gingerol has been widely used for its biologic and chemopreventive effects for centuries, with more controlled clinical trials in recent years (49). Both curcumin and gingerol are well tolerated and without any toxic effects. The only reason that higher doses are not taken related to the large number of pills required (up to 4-8 tablets four times/day).

Proposal for Use of Curcumin and Gingerol in Patients With MDS

In this protocol, we propose the use of a combination of curcumin and gingerol in patients with all categories of MDS. The rationale for using these two compounds is that both are natural, non-toxic substances with anti-angiogenic, anti-TNF/anti-TGb, anti-NFkB, anti-proliferative and chemopreventive properties which are likely to be well tolerated in this elderly population. Since we are using these compounds with therapeutic rather than chemopreventive intent, a combination of the two is likely to be better than use of either alone. The reason for including all types of MDS is that with low to intermediate-1 risk disease patients, the disease is slow growing and will allow for prolonged use of these agents over at least a six month period. For intermediate-2 and higher risk MDS patients, we will use these agents only if the patients cannot tolerate other and more aggressive forms of anti-leukemic and cytotoxic therapies.

Treatment Protocol

All patients with a confirmed diagnosis of myelodysplastic syndromes will be eligible for this trial. No other experimental agent aimed at treating MDS will be allowed during the period of protocol therapy. Commercially available curcumin and gingerol will be used. We propose to start the patients with 1 gm qid of curcumin and ginger extract, 350mg, two tablets twice daily. Both curcumin and gingerol will be increased as tolerated to a maximum dose of 12 Gm/day for curcumin and 2.8 Gm/day for gingerol. Therapy will be continued unless there are signs of disease progression. Supportive care measures including the use of transfusions as well as growth factors as indicated will be permitted during the protocol duration.

The schema of this protocol will be:

Start curcumin at 4Gm po q day and escalate as tolerated to 12 Gm  po qday and gingerol at 1400mg/day and escalate to 2800mg/day

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