"Something to do with mice, and antibodies, and making them human, or something" was usually the best I could do.
So when the subject appeared yet again during this years course, I decided to finally look it up properly and work out just what was going on.
Antibodies look like this:
The two variable regions recognise bind to antigen (parts of invading bacteria) leading to the invading bacteria being destroyed. Antibodies produced in the body are polyclonal, because each one has a different variable region and can target a different antigen (until a threat is realised in which case they massively overproduce the relavent antibody).The idea of monoclonal antibody therapy is to produce a large number of essentially the same antibody, that can find and potentially destroy a specific target. The idea was to produce a kind of 'magic bullet' that went through the body picking out the specifically ill parts and removing them. And antibodies are very specific, and can be targeted to lots of different proteins.
The problem with producing them is that a single B cell (antibody-producing cell) will only last a few generations before dying. Not long enough to produce the large amounts of specifically-target antibody needed for therapy. The original solution to this problem was to use a technique known as hybridoma. Individual B cells that had been grown in mice and produced antibodies that destroyed whatever target the therapy was being designed to remove were fused with immortal myeloma cell lines. The B cell could then propagate for much longer, secreting monoclonal antibodies. The main problems with this technique were that is was slow and laborious and created problems for purifying the antigen.
The most modern technique I know of (although others are being developed) is called SLAM, which stands for Selected Lymphocyte Antibody Method. B cells are isolated from mice (or rabbits, other animals can potentially be used as well) and grown in little plastic wells until they start secreting antibodies. Single B cells are then isolated, and screened for activity. The relevant antibody genes are then cloned through PCR and expressed as recombinant antibodies. This technique is a lot faster and produces high affinity antibodies from a number of species.
Monoclonal antibodies are used in various drugs currently on the market. Lymphomas (cancerous B cells) can be treated with Zevalin (R) or Bexxar (R). Apparently on 3 February 2005, the New England Journal of Medicine reported that 59% of patients with a B-cell lymphoma were disease-free 5 years after a single treatment with Bexxar.
The thing is though, I'm being taught this as a biochemist student/researcher, not as a medical researcher. Which means that I have very little idea how useful, common, or applicable most of these techniques and products are. Academic researchers and medical researchers seem to live a world apart, something that hit me particularly hard during the conference. You could almost always tell, about half way through a talk, whether the speaker was a medical or academic researcher. There doesn't seem to be a whole lot of cross-talk between them either, which is a pity because academic research does often come up with the odd useful medical application, but of course they aren't in any position to implement it.
I'm not sure that the hybridoma approach is especially slower than SLAM, when you take into account that in a clinical setting you will want to select for multiple properties (affinity, function, immune response, etc). The problem with SLAM is that, because you never culture the B cells, you can't really introduce selection beyond "the antibody forms a plaque in the presence of the antigen". With a hybridoma, you can introduce an arbitrary amount of selection, as you have an immortal cell culture.
ReplyDeleteThere are more modern SLAM-like methods that also introduce a culture stage, prior to cDNA-ing the variable region, but they are still in pretty early stages, and AFAIK virtually all monoclonal antibodies are produced using hybridoma in practice.
The most interesting story with monoclonal antibodies, I think, is the increasing chimerisation and humanization of them. If you inject a patient with a perfectly pure mouse monoclonal antibody, as traditionally was done, you get a strong immune response. This is alright when you use them as they were originally used for, like to prevent clotting during an operation, but if you want to use them in cancer treatment or that sort of thing it becomes a problem.
In some cases you can use human B-cells, which works well enough, but you cannot elicit self responses (as is required for cancer or autoimmune problems), and you can't inject humans with dangerous things. So instead, we have this interesting progression, where we start by adding human heavy-chains to mouse light chains, and then go on to move just the variable region of the mouse antibody onto a human one.
Of course the problem with all this is that we have to do all these transgenics every time we to create a new antibody. Which leads us to the xenomice, which are really the logical progression of this story. Instead of fiddling around with post-recombination B cells, instead we move the entire human antibody-generating region (with all it's little genetic capsules to be shuffled during B cell maturity) into a mouse, and then disable the mouse's response.
If you want antibodies, you don't need to extract variable regions, instead you just inject your mouse with your antigen, screen the resulting B cells, and hybridoma-ize the results - giving purely human antibodies with from a purely mousey mouse.
It's funny, but I've never noticed the gap between medical and academic researchers. My lab is kind of a hybrid between the two, and my bio friends are pretty evenly split. I guess since in the US we get so much common background at the upper levels that we're a lot more interdisciplinary in our professional approaches, as well.
ReplyDeleteAnd with antibodies: Dr. Mitchell explained epitopes to us by pretending that the lecture theatre had been invaded by overhead projectors and that he was the B cell. 2nd most entertaining lecture of my life.
To Luke: Thanks for the information! You see our notes just say "hybridoma = slow uncertain generally bad; SLAM = fast exciting generally good". They really don't focus much on actual usage or clinical applications.
ReplyDeleteWe have covered humanisation, although I don't think I've heard of the xenomouse. I'll have to look it up, it sounds really interesting :)
To Rhan: hmm, that's interesting that it's different in the US. Here, in my experiance at least, medical researchers get different funding, look for different things, and seem to approach things from a different mindset. In microbiology it least, there really doesn't seem to be enough communication or colaboration. A hybrid lab sounds really great; is it just medical researchers and academic researchers working on the same site? Or do you all recieve the same funding and share resources and things>