Even though further clarification is required regarding the structure and generation of GcMAF, Yamamoto’s body of work has given rise to two key propositions that may have extraordinary importance if sustained by future research: namely, that serum nagalase can serve as a virtually universal marker for cancer, and that parenteral administration of pre-formed GcMAF can exert a profound immunostimulant effect in cancer patients, enabling the effective control and eradication of many cancers when they are in a micrometastatic or very early nascent form.

In 1996, Yamamoto and colleagues published a study in which they assayed serum nagalase in 20 patients with a wide range of cancers, as well as in 5 healthy subjects.10 Whereas the serum nagalase activity in the healthy subjects averaged 0.23 (nmoles/mg/min), it ranged in the cancer patients from a low of 0.64 to a high of 5.21. In several patients who were receiving radiation therapy, the serum nagalase activity declined progressively, coming close to the normal range in some. Conversely, the ability of Gc protein derived from the patients to give rise to GcMAF activity in vitro (assessed by stimulation of superoxide production by macrophages) tended to be low in cancer patients, and vary inversely with serum nagalase; this precursor activity increased progressively during radiation therapy.

A year later, Yamamoto published a study confirming these findings in 36 patients with oral cancer; whereas serum nagalase averaged 0.29 in twelve healthy controls, this activity was above 1 in all except one of the cancer patients (0.74), and was above 6 in four of them.11 Once again, serum precursor activity for GcMAF generation tended to be subnormal and correlated inversely with serum nagalase in the cancer patients. Surgical excision of primary tumors or metastasized nodes was followed by a rapid decline in serum nagalase, coming close to the normal range in 3 patients.

Yamamoto’s assessment of serum nagalase in patients with micrometastatic breast, colorectal, and prostate cancers appears to provide further confirmation for the utility of nagalase as a cancer marker; these findings are discussed below.

 

To date, two independent clinical groups have published assessments of serum nagalase in cancer patients. Researchers at the University of Madras measured serum nagalase in 85 healthy controls, and in 210 patients with squamous cell carcinoma of the uterine cervix, both before and after a course of radiotherapy.18 In the controls, nagalase averaged 0.26 – very similar to Yamamoto’s control values.

Prior to treatment, nagalase averaged 0.72, 1.54, 3.78, and 5.05 in patients with Stage 1, 2, 3, and 4 cancers, respectively. After radiotherapy, these values averaged 0.39, 0.64, 1.38, and 3.14, respectively. These findings appear wholly confirmative of Yamamoto’s proposition that serum nagalase can serve as a cancer marker and can be used to assess the effectiveness of therapy. More recently, Greek researchers have examined nagalase in patients with pre-surgical melanoma.19 Nagalase levels were found to be significantly elevated relative to controls in patients with stage 3 disease, but not in patients with earlier lesions. Nagalase correlated directly with tumor thickness (Breslow index); in patients with stage 2 or 3 disease, it declined significantly after surgery. 

Hence, whereas much further clinical research is needed to confirm that nagalase is the nearly universal cancer marker that Yamamoto proposes it to be, the published clinical evidence so far is consistent with this proposition – although it may not always be elevated in early cancer.

Several studies in rodent models likewise confirm an elevation of serum nagalase in tumor-bearing animals. Moreover, they suggest that this elevation of nagalase tends to be proportional to tumor mass.

However, whereas one such study – in nude mice implanted with human squamous cell carcinoma11 – found that nagalase correlated directly with tumor mass, a subsequent study in mice carrying Ehrlich ascites tumors found that serum nagalase varied directly with the log of tumor mass.20 This latter finding might explain why patients with advanced cancer can display serum nagalase that isn’t orders of magnitude higher than those with micrometastatic disease (as discussed below).

If, as appears likely, serum nagalase is elevated in a very high proportion of patients with cancer, what might account for this? So far, the molecular biology behind these elevations remains virtually unexplored – as noted, the gene responsible for tumor-associated nagalase hasn’t yet been characterized.

However, a study focusing on the molecular biology of Ehrlich ascites cancer is of intriguing relevance. Segura and colleagues, noting that increased expression of glutaminase is a common feature of cancer thought to contribute to its malignant behavior, used anti-sense technology to impede the expression of glutaminase in Ehrlich ascites cells.21 In this altered cell line (0.28-AS-2), the expression of two proteins thought to influence the susceptibility of cancer cells to immune rejection, mucin-1 and nagalase, was notably decreased. (Nagalase expression was assessed by its enzymatic activity.) 

Whereas wild-type Ehrlich ascites cells grew prolifically in immunocompetant mice, the altered cells could not produce tumors in these mice, and a far higher number of activated macrophages were found in the ascitic cavity of the mice injected with the 0.28-As-2 cells. However, these cells readily formed tumors in immunodeficient nude mice, killing them within 20 days. 

These findings suggest that glutaminase knockdown, likely in part by blocking nagalase expression, abrogates mechanisms which Ehrlich ascites cells employ to evade immune rejection – mechanisms which are superfluous when cancer is implanted in immunodeficient mice. It remains unclear why glutaminase expression or activity might influence nagalase expression, and no subsequent research has examined this intriguing lead. Glutaminase is now viewed as an oncogene, and its increased expression is often a reflection of increased myc activity – so perhaps myc is the prime driver of nagalase overexpression in cancer.

The possibility that nagalase might be elevated in other pathologies – raising the possibility of false positives when nagalase is used as a cancer marker – requires much further study. Certain viruses, including the HIV virus, also express a nagalase-like activity, and serum nagalase activity is increased in HIV patients.24 An elevation of serum nagalase has also been reported in systemic lupus erythematosis; the basis and source of this increase remains obscure, but Yamamoto intriguingly proposes that this  increase may responsible for a decrease in phagocytic activity that contributes to the elevation of circulating immune complexes in this disorder.

In any case, it is clear that nagalase may have great clinical potential as a cancer marker – perhaps a near universal cancer marker that could be of great utility for guiding cancer therapy, particularly when cancer is present in micrometastatic or early form, and hence difficult or impossible to assess with radiological techniques. 

It is distressing that, 17 years after Yamamoto’s first pertinent clinical publications, so few independent groups have chosen to study this issue – and so little progress has been made on defining the molecular biology of tumor-associated nagalase.

The Need for Confirmatory Research

Five years after the publication of these quite provocative studies, no further clinical trials with GcMAF have emerged. Apparently, pharmaceutical companies are attempting to develop more-readily-patentable synthetic analogs of GcMAF as drugs; there appears to be no current effort to register natural GcMAF for therapeutic use. Several biotech companies have recently stepped into this void to sell GcMAF, packaged in sterile vials suitable for use in injections, via mail order internationally, without explicit drug claims; a number of cancer patients, and a few “integrative” doctors, are availing themselves of this resource. It seems likely that the majority of cancer patients currently using GcMAF have advanced lesions that are unlikely to respond definitively to immunotherapies; GcMAF is still a little-known option, and the patients who seek it out are likely to be searching for options after standard therapies have failed them.

Yamamoto’s clinical reports have no evident implications for those with advanced cancer – albeit the possibility that GcMAF therapy might sometimes help to slow the growth and spread of progressive cancers, as suggested by some of the rodent studies, certainly merits consideration. The anecdotal experience of cancer patients currently using GcMAF may cast some light on this issue.

It is clear that two research initiatives are urgently needed: the utility of nagalase as a cancer marker should be evaluated aggressively in a wide range of cancers, correlating it, when feasible, with other cancer markers that are more broadly accepted; and surgical oncologists should evaluate weekly injections of GcMAF as an adjuvant strategy in post-surgical patients who are radiologically cancer free but remain nagalase positive, in conjunction with whatever adjuvant chemotherapy or radiotherapy that is considered indicated. Unless and until GcMAF is formally registered as an investigational drug, legal consideration may make clinical trials with this agent problematic; nonetheless, this issue is of such potential importance that avenues for evading these legal restrictions should be explored – particularly in light of the fact that no serious side effects of GcMAF therapy have been reported to date. Notably, nothing currently prevents cancer patients, on their own initiative and at their own expense, from acquiring GcMAF for personal use. Currently, GcMAF acquired from a European source costs about $600 per month to use if 100 ng GcMAF is injected weekly.

As noted, rodent studies suggest that daily administration of GcMAF may have important potential as an anti-angiogenic agent; this strategy might prove to be of some utility in advanced cancers unlikely to becontrollable by immunotherapy per se. To date, no clinical trials have evaluated this strategy; the current expense of GcMAF would discourage most cancer patients from trying this. Moreover, if GcMAF does have the potential to evoke significant adverse effects, such effects might be more likely with a daily administration regimen. Nonetheless, this strategy merits clinical evaluation. A proviso is that antiangiogenic therapies to date, while they often temporarily slow cancer spread, have had little impact on overall survival, probably because the hypoxic tumor environment induced by suppression of angiogenesis tends to select for highly aggressive cancer variants.33-35

 

Monitoring Nagalase for Early Cancer Detection?

The intriguing possibility that nagalase assessment and GcMAF treatment might enable early detection and elimination of nascent cancers, has been raised in an insightful monograph posted online by Tim Smith.61 If indeed many incipient cancers raise serum nagalase levels, the inclusion of nagalase in standard health profiles might enable detection of cancers before they become symptomatic – and in an early phase where they might be susceptible to elimination, not only by surgery (if a source can be established), but also conceivably by a course of GcMAF treatment. Of course, it is still unclear whether very early cancers are prone to raise serum nagalase; Yamamoto’s data pertain to patients with well established, often pre-treated, cancers, and recall that the evaluation of nagalase in melanoma patients failed to note an elevation of serum nagalase in patients with phase 1 or 2 disease. But, if nagalase proves to be a genuine and near-universal marker for cancer, the possibility that it might be employed to achieve early detection at a curable stage of at least some cancers is quite exciting.

Questions to Address in Future Research

Clearly, the interrelated research literature on GcMAF and nagalase shows very intriguing promise, but it is currently underdeveloped, and far too isolated from major research trends in cancer immunology.

Questions that merit consideration in future research include:

Is nagalase the nearly universal cancer marker that it has been purported to be?

If so, why – what oncogenes drive its expression?

Is serum nagalase elevated in a high proportion of early tumors, so that it could be monitored as an “early warning system” for cancer?

Precisely how does nagalase impair the generation or maintenance of GcMAF activity in tumors?

What is the molecular biology that underlies GcMAF’s ability to boost superoxide production and phagocytic activity in macrophages?11

Does GcMAF work in other ways to aid the cancer-killing activity of macrophages?

Does GcMAF influence M1/M2 polarization in tumor-associated macrophages?

Does GcMAF treatment notably boost the tumoricidal activity of M1 polarized macrophages?

Is weekly GcMAF administration as effective for eliminating microscopic metastatic disease as suggested by Yamamoto’s published clinical trial – at least for certain major cancers?

If so, which types of cancer respond most effectively?

Can GcMAF administration slow the growth of more advanced cancers?

Would addition of certain M1-polarizing agents to GcMAF therapy boost its efficacy and make it more useful for control of macroscopic tumors?

Would daily administration of GcMAF have utility as an anti-angiogenic strategy in cancer therapy – and would such regimens be safe?