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Phytochemicals &


Beta Glucan, Complement and Rituxan

Date: January 23, 2003

by Chaya Venkat

Make Like a Yeast Cell

Related Articles:
Rituxan Therapy

yeast cell

You will have to be patient with me on this one, the story is long winded, but I found it fascinating.

Let us say your body is invaded by yeast. That's right, you guys, you can get yeast infections too. Any one who has ever made bread knows that given the right conditions, yeast cells multiply and proliferate rapidly, colonizing the entire eco-niche available to them. Over the millennia, the human body has developed very strong defense mechanisms against such very dangerous invading organisms.

One of the body's defenses when it is under attack is circulating antibodies in the blood that latch on the antigens on the surface of the yeast cell. We are not talking about just one antigen and one antibody attached to it, per yeast cell. There can be literally hundreds of such pairs per yeast cell. Presence of such Ag-Ab pairs gets the complement systems going, and pretty soon the cell is coated with literally thousands of particles of complement. (Please read my previous article on "Complement (no typo) to your health" for background information on complement system and how it works).

Cells which are covered by complement fragments are said to be "opsonized". Such cells are quickly killed. Phagocytic cells (cells that are literally capable of eating up other cells) such as macrophages and neutrophils are attracted to the site of the opsonized cells, and quickly eat them up. This is called the classic pathway by which complement attacks pathogens such as yeast cells.

Tumor cells grow in the body in the first place because the body does not recognize them as "foreign and dangerous", as are yeast cells. Part of the problem is that tumor cells do not have a real ugly antigen on their surface that triggers activation of the immune system. If this were not the case, there would be sufficient home grown antibodies against the cancer cells, and the tumor would be killed before it had a chance to grow. The fact you have CLL says your body is not able to mount a sufficiently effective antibody response to the antigens on the tumor cells.

OK. So we have CLL, but no antigen that the body recognizes, and therefore no antibody. Not to worry, IDEC comes to the rescue, we define CD20 marker on B-cells as an antigen, and make an antibody against it called Rituxan. Once this drug is administered to patients with CLL, especially those with a large percentage of b-cells that are positive for CD20, and have a large number of CD20 markers per cell, we have met the first condition for killing the cell. Namely, antigen-antibody complexes are formed on the surface of the cancer cells.

Scientists have actually followed a patient undergoing Rituxan therapy (see ASH abstract below). They could see that the Rituxan was co-located with CD20 markers, and that a huge amount of complement fragment C3b was also co-located with the Ag-Ab pairs. So far, so good. Antibody has attached itself to the antigen, and complement has coated the surface of the cell.

Here is where the problems start. In the case of diseases like Follicular Lymphoma, cells which have Ag-Ab pairs, and are covered ("opsonized") with complement fragments are quickly killed and phagocyted, gobbled up by things like macrophages. This is why Rituxan is so effective in the case of Follicular lymphoma. But in the case of CLL, even though Rituxan-CD20 pairs (i.e., Ag-Ab pairs) are created, and complement fragments cover the cell, only a small percentage of the cells actually die. Below is a PubMed abstract of an article in Blood, which suggests that there are other markers on the surface of CLL cells, namely CD59 and CD55 etc, which prevent the next step from happening. These markers are somehow able to prevent the phagocytic cells from getting into a killing mode. You can get this whole article free of charge, if you are so inclined, by searching for it at the Blood-on-line site.

Here is a politically-incorrect analogy that might make it easier to understand. The country is infiltrated with a group of terrorists (CLL cells). Problem is they look just like innocent citizens, no particular way of saying who is a terrorist (CLL cells do not have strong and ugly antigens), so the police force (complement) can't do much about it. Along comes the CIA (IDEC Pharmaceuticals) that discovers all terrorists wear black turbans (CD20 markers), and is able to create a smart snitch (Rituxan) that can home in on the black turbans, lock on to them and send out an all-points alarm (form the CD20-Rituxan pair). The alarm gets the desired response from the cops (complement) and pretty soon the bad guy (CLL cell) is surrounded by cop cars (CLL cell is covered by complement fragments). The assassination squad (macrophages and neutrophils) move in, and we all hold our breath, waiting for the bad guy to get blown away and taken apart.

Small problem, the bad guys have very smart lawyers (the CLL cells have CD55 and CD59 markers) who start quoting chapter and verse from the Bill of Rights and the assassination squad is thoroughly confused. They mill around, not knowing quite what to do, and many of the bad guys escape as a result (CLL cells can evade killing even after getting opsonized, because of inhibitory markers such as CD55 and CD59). All this fuss and bother, and not much to show for it. Country is still stuck with terrorists (CLL cells), and only a few them have been killed (only partial remission). Unless we do things differently, they will soon recruit new members and grow back to their original numbers (relapse).

Light bulb goes off. If the problem is that the assassin squad is not motivated, and easily fooled, how about giving these Rambos a quick course in remedial killing 101, get them all riled up and ready  for action, before letting the smart snitches (Rituxan) loose on the guys in the black turbans (cells with CD20 marker). This way, once the terrorists are identified, tagged by the snitches and surrounded by cops, they cannot escape death from Rambo.

End of silly analogy. It turns out that one of the most effective ways of riling up the macrophages and other phagocytic cells is to make them think they are under attack by an invasion of yeast cells. Now we don't really want to add to the problems of our poor CLL patient with a real yeast infection, we just want to trick the macrophages into thinking there is such an infection. It turns out that many yeast and fungal cells have on their surfaces these long polymeric strands of sugars called beta-glucans. Over many many eons the immune system says to itself "Yeast Attack!!" the moment it sees a molecule of beta-glucan. So here is a possibility for getting around the confused and unmotivated macrophages and neutrophils, the remedial-killing 101 course needs to be no more than letting our Rambos see a lot of beta-glucan molecules floating around. Interesting stuff. See the second Ash Abstract below, which says that co-administration of beta-glucan increases the efficiency of monoclonal antibodies.

If you type beta-glucan into the Google search engine, you will get literally thousands of hits, with dozens of companies willing to sell you their version of beta-glucan, good for everything that ails you except an empty wallet. Even the FDA says it is non-toxic, and no one needs a prescription to buy this stuff at local drug store or GNC outlet. What is the harm in trying it?

Plenty, if you don't know what you are doing. The least of the problems is that unless the beta-glucan is pure and free of a lot of attached fat and protein, it will pass right through your system, and you have just lost a few dollars down the toilet, literally. There can be other and more serious consequences. Getting back to our analogy, let us say the country has a problem with out of control assassin squads running amuck, killing innocent citizens (you have some form of autoimmune disease), without caring whether or not they  are bad guys. Last thing you would want is to further rile up these homicidal Rambos on steroids. Beta-glucan supplements can be real dangerous for people with autoimmune disease.

Another problem can be severe tumor lysis syndrome. If the cell kill is happening fast enough after administration of Rituxan, and by adding beta-glucan to the mix you further increase the speed with which it is happening, your body might have trouble disposing off of the debris from the killing. Tumor lysis syndrome can be potentially fatal, with serious complications of the liver and other organs. Seriously folks, I don't want to hear that any off you are rushing off to buy this stuff, and self medicating yourself, without learning about it, and getting help with using it if you think this concept is worth exploring.


The references below are to two long and very detailed papers from Dr. Gordon Ross of the University of Louisville, Kentucky. The titles are quite descriptive. I would strongly urge you to at least browse through these articles using the links provided to the full-text pdfs.

Beta Glucan Review;
Beta Glucan as Adjuvant to Monoclonal Antibody Treatment.


Complement (C) Activation Is Required for Rituximab (RTX) Mediated Killing of CD20 Positive Cells, and a High Tumor Burden May Decrease Therapeutic Efficacy Due to C Depletion as a Consequence of Therapy. In Vitro and In Vivo Studies.

Ronald Taylor, Adam Kennedy, Michael Solga, Paul Beum, Patricia Foley, Margaret Lindorfer, Charles Hess, John Densmore.

Biochemistry, Universtiy of Virginia School of Medicine, Charlottesville, VA, USA; Comparative Medicine, Universtiy of Virginia School of Medicine, Charlottesville, VA, USA; Hematology/Oncology, Universtiy of Virginia School of Medicine, Charlottesville, VA.

We investigated C activation, C3bi deposition, and cell killing when RTX was bound to Raji or DB cells, in serum (NHS) as a C source. In the presence of RTX and C, large numbers of C3bi molecules deposit per cell, and fluorescence microscopy revealed that C3bi co-localized with bound RTX. Both cell types are killed by RTX in the presence of NHS, and use of mAb 3E7, specific for C3bi, enhanced killing. However, in the absence of NHS, little or no killing was demonstrable. RTX was infused into monkeys and we found that it rapidly bound to circulating B cells and activated C; 2 min after RTX infusion, C3bi was co-localized with RTX bound to the B cells. A similar pattern of co-localization of C3bi and RTX was obtained in vitro in opsonization experiments with blood samples taken from patients with B cell lymphomas. A patient with CLL was treated with the standard 4 week course of RTX therapy. Analyses of blood samples taken during this time revealed the following: Immediately after the first infusion, flow cytometry measurements and fluorescence microscopy indicated that RTX and deposited C3bi were associated with both B cells and cellular debris, and a high degree of co- localization of C3bi with RTX was evident. After the first infusion, CH50 assays demonstrated that the patient's C titer, normal before treatment, had been substantially reduced (~ 5 fold). Although C levels were partially restored before the second infusion, the same pattern of C depletion occurred after this infusion, and by week three and four the patient's C levels were reduced ~ 10-fold, but returned to baseline 3 weeks later. We found, however, that in vitro supplementation of the C-depleted sera with C component C2 markedly increased C activity, leading to levels that were at least 50% of the pre-therapy baseline. In vitro studies with RTX, B cell lines, and NHS gave rise to similar findings. That is, at high cell concentrations RTX-mediated C3bi deposition and killing is limited by serum C, and both of these activities can be enhanced by supplementation with purified human C2. Our results indicate that the primary mechanism of action of RTX in killing CD20 positive cells is mediated through C, and it is likely that the in vivo form of this mAb bound to target cells has covalently incorporated C3bi. We suggest that if an anti-tumor mAb such as RTX requires robust C activation for therapeutic efficacy, then insuring an adequate level of C activity in a patient, by supplementation with either fresh plasma or a purified C component such as C2, may provide an important approach for improving the therapeutic efficacy of a C-fixing mAb.

Keywords: Rituximab\ Complement\ Immunotherapy

Blood 2001 Dec 1;98(12):3383-9

CD20 levels determine the in vitro susceptibility to rituximab and complement of B-cell chronic lymphocytic leukemia: further regulation by CD55 and CD59.

Golay J, Lazzari M, Facchinetti V, Bernasconi S, Borleri G, Barbui T, Rambaldi A, Introna M.

Laboratory of Molecular Immunohematology, Istituto Ricerche Farmacologiche Mario Negri, Milano, Italy.

Complement-dependent cytotoxicity is thought to be an important mechanism of action of the anti-CD20 monoclonal antibody rituximab. This study investigates the sensitivity of freshly isolated cells obtained from 33 patients with B-cell chronic lymphocytic leukemia (B- CLL), 5 patients with prolymphocytic leukemia (PLL), and 6 patients with mantle cell lymphoma (MCL) to be lysed by rituximab and complement in vitro. The results showed that in B-CLL and PLL, the levels of CD20, measured by standard immunofluorescence or using calibrated beads, correlated linearly with the lytic response (coefficient greater than or equal to 0.9; P <.0001). Furthermore, the correlation remained highly significant when the 6 patients with MCL were  included in the analysis (coefficient 0.91; P <.0001), which suggests that CD20 levels primarily determine lysis regardless of diagnostic group. The role of the complement inhibitors CD46, CD55, and CD59 was also investigated. All B-CLL and PLL cells expressed these molecules, but at different levels. CD46 was relatively weak on all samples (mean fluorescence intensity less than 100), whereas CD55 and CD59 showed variability of expression (mean fluorescence intensity 20-1200 and 20-250, respectively). Although CD55 and CD59 levels did not permit prediction of complement susceptibility, the functional block of these inhibitors demonstrated that they play an important role in regulating complement-dependent cytotoxicity. Thus, lysis of poorly responding B-CLL samples was increased 5- to 6-fold after blocking both CD55 and CD59, whereas that of high responders was essentially complete in the presence of a single blocking antibody. These data demonstrate that CD20, CD55, and CD59 are important factors determining the in vitro response to rituximab and complement and indicate potential strategies to improve the clinical response to this biologic therapy.

PMID: 11719378

Beta Glucan plus Monoclonals Is Effective in Mice

Here is an interesting PubMed citation from folks at Memorial Sloan-Kettering. By now you know that what Beta-glucan does is prime the immune cells of your system (the CR3 site of the leukocytes) such that they are more effective in killing target cells that have been tagged by monoclonal antibodies and covered with fragments of complement molecules. 

These are mouse studies, and the results suggest that the approach works for a variety of cancers, including lymphoma. Too bad they did not include CLL in the list. By the way, Memorial Sloan-Kettering has a clinical trial for people with neuroblastoma, combining a monoclonal antibody right for that cancer with beta-glucan.

Looks good so far, and worth keeping an eye on these developments. But do remember, mice are easier to cure than humans. 


Cancer Immunol Immunother 2002 Nov;51(10):557-64

Orally administered beta-glucans enhance anti-tumor effects of monoclonal antibodies.

Cheung NK, Modak S, Vickers A, Knuckles B.

Department of Pediatrics, Memorial Sloan-Kettering Cancer Center, 1275 York Avenue, New York, NY. 

Beta-Glucan primes leukocyte CR3 for enhanced cytotoxicity and synergizes with anti-tumor monoclonal antibodies (mAb). We studied readily available (1-->3)-beta- D-glucan using the immune deficient xenograft tumor models, and examined the relationship of its anti- tumor effect an  physico-chemical properties. Established subcutaneous (s.c.) human xenografts were treated for 29 days orally with daily beta-glucan by intragastric injection and mAb intravenously (i.v.) twice weekly. Control mice received either mAb alone or beta-glucan alone. Tumor sizes were monitored over time. beta-Glucans were studied by carbohydrate linkage analysis, and high performance size-exclusion chromatography with multiple angle laser scattering detection. Orally administered beta- D-glucan greatly enhanced the anti-tumor effects of mAb against established tumors in mice. We observed this beta-glucan effect irrespective of antigen (GD2, GD3, CD20, epidermal growth factor-receptor, HER-2), human tumor type (neuroblastoma, melanoma, lymphoma, epidermoid carcinoma and breast carcinoma) or tumor sites (s.c. versus systemic). This effect correlated with the molecular size of the (1-->3),(1-->4)-beta- D-glucan. Orally administered (1-->3),(1-->6)-beta- D-glucans also synergized with mAb, although the effect was generally less marked. Given the favorable efficacy and toxicity profile of oral beta- D- glucan treatment, the role of natural products that contain beta- glucan in cancer treatment as an enhancer of the effect of mAb therapy deserves further study. 

PMID: 12384807



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