In response to your questions.
It's nice to see that the subscribers at SI are interested in challenging critical discussions. Being challenged facilitates good decision making. This is refreshing compared to some other boards (eg. Yahoo) a critical comment has the interesting consequence of restricting the intelligence of those who post to a prehistoric version dominated by the limbic system with very limited frontal lobe activity.
In response to your question regarding the nature of CAT's Ig library, my research indicates that the library is composed of VH and VL loci cloned into a filamentous phage vector. The library of VH and VL genes was obtained from a large and heterogeneous population of human donors and contains greater than 100 billion different functional V regions. This library of greater than 1.2 x 10 to the 11 is now approaching the limit of human Ig diversity which is estimated somewhere between 10 to 13 and 10 to the 15. The resulting phage population expresses scFv on their surface allowing "panning like" screening in multiwell plates coated with target antigen. This is a very easy selection process compared to those often employed with hybridomas. Several rounds of screening are performed within a span of 2 to 3 weeks to select for the VH and VL of highest affinity. scFv fragments are then engineered from the highest affinity heavy and light chains selected. The number of promising clones obtained in this matter does not seem to me to be a limiting factor. I think the production of anti TGFB2 antibodies using this technology offers a good example (J Immunol Methods 227:17-29, 1999). Thompson et al obtained 43 promising scFv fragments after several rounds of scVH and scVL selection and random ligation of VL loci to promising VH loci. After further screening, 4 scFv fragments were selected for further engineering into full Ig based on in vitro bioassay activity. In some cases, where Fab or Fab'2 fragments may be of greater clinical desirability than full Ig (eg. When activation of complement cascade or opsonization is not desirable), these Ig fragments may be assessed for bioactivity directly. In such cases, CAT may have an advantage over murine transgenic systems where further manipulation would be required to obtain Ig fragments. To produce full Ig, CAT ligates the Fc portion of their choice to the selected Fv genes. CAT can then assess many different isotypes of the same specificity for bioactivity. Phage display also offers the possibility of specifically engineering superantibodies. Superantibodies are fascinating because they posses intrinsic proteolytic activity. Superantibody applications in the literature include the clearance of poisonous drugs in patients experiencing drug overdoses. Phage display does not appear to have any advantage over the murine transgenics in terms of the final number of promising full Ig clones obtained. Proper glycosylation of the full Ig products is achieved via expression cloning in a eukaryotic expression system. The mAbs produced here are then subjected to bioactivity selection which can be performed via many different methods including animal models and biossays. Mutagenesis may be performed on the full Ig in any of the 3 CDR sites and the best mAbs selected for based on bioactivity. Of course, Abgenix and Medarex can do this too but I believe CAT has broad patents protecting this final process as well as agreements with Ixsys by which CAT can utilize similar in vitro affinity maturation technology without infringement.
In terms of the quality of the human mAbs produced against specific known targets, I don't think there is much to choose between phage display and transgenic mice as long as the phage display library is sufficiently large. See Paul Carter's comments in Nature Biotechnology on a study by Vaughan et al (Nature Biotechnology 14: 309-314, 1996) CAT has impressed me because of their careful selection of target antigens. They seem to understand that it is better to select targets with limited risk in terms of efficacy than to choose targets which are high risk, high gain (a business strategy that almost always fails). CAT is performing preclinical studies in other undisclosed and most certainly higher risk areas but it has impressed me that they are being careful to bring the lower risk ones to the clinic first even if their preclinical data is very promising and even if it means assessing an indication which may require very lengthy clinical trials, may not have an orphan indication or as large a market. They are risking the impatience of the market, in my opinion the major fault of most biotech "stock traders". This strategy may not be terrific for short term stock price performance but it is a far better strategy in building a successful company that will maintain long term success. My analogy for the day is the Montreal Canadians. They have won more championships than any other team in hockey; not by risking team chemistry by signing one or two superstars but rather by maintaining a solid team of quality players that play well together year after year. The current Canadian's team aside which sweet victory for me growing up in Toronto and all. (GO LEAFS GO!) From what I can tell the businessmen at CAT must actually "listen" to the scientists!
While performing my DD on CAT, I have been impressed with the fact that they are not just an antibody factory. They have two other major applications which include an interesting technology for identifying molecules associated with desired targets (basically a more advanced immunoprecipitation technique which includes the concomitant selection of antibodies against the associated proteins) and of particular interest to me a bioinformatics/genomics division. Like many other companies in the genomics field (eg. VRTX, AXPH) CAT searches genetic databases for genes that might be associated with certain diseases. When a target gene is found, CAT employs their phage display system to produce high affinity Fv fragments against recombinant peptides produced from the gene of interest. These Fv fragments are then used to screen tissue samples from normal and test populations for potential differences in gene product expression pattern. This data is then compiled into a functional genomics database. Murine transgenics could also produce antibodies against gene products of interest but the selection process of hybridomas is quite cumbersome and I believe would limit its usefulness for this application. For genomics research, speed really is an advantage. This strategy, then, has the potential to streamline bioinformatics, functional genomics and drug discovery into one simple system.
With regard to the anti TNF story, I have not seen Celltech Chiroscience's data but I did see data on Remicade at last year's PANLAR conference. The data was very similar to that seen with Enbrel except for the clear disadvantage of rejection which was a common finding within 6 months of drug initiation in the absence of any immunosupression. I am not a big fan of humanized antibodies. They are still immunogenic, it just takes a little longer to reject them. For some indications, this is OK but not for RA and so I would rank the anti TNF reagents in the following order: fully human anti TNF mAbs (CAT)>recombinant human TNF receptors(IMNX)>humanized anti TNF mAbs(CCH, JNJ).
Actually, anti-CD4 mAbs (MEDX) have been plagued with efficacy difficulties as well. Although the data is often presented in a glorifying way, upon close inspection anti CD4 antibodies are not very effective clinical therapeutics for autoimmune disease. When given in combination with anti cytokine reagents they appear to have some benefit but as a stand alone therapy, I'm not impressed.
The CTLA-4 (MEDX) story is an interesting one. The dogma goes like this. T cells require two signals to be properly activated. A TCR-MHC interaction and costimulation. The major costimulatory pathway is the B7/CD28 pathway. CTLA-4 is upregulated on activated T cells. Like CD28, its ligand is B7 on APC but CTLA-4 is associated with T cell regulatory activity including cellular arrest and apoptosis. CTLA-4 Ig was designed to block the signal through CTLA-4, thus rescuing tumour specific cells from "senescence". It is an intriguing idea and might work. The caveat here is that in many cases, tumour specific T cells may have undergone peripheral deletion in patients with advanced cancer. Many of these patients are on very toxic drugs and it is very contraversial whether therapies that rely on immunostimulation will work in this patient group. MEDX has partnered up with IDM to combine anti CTLA-4 therapy with dendritic cell therapy. Dendritic cell therapy is also very useful in immunocompetent hosts but the data in real cancer populations has been anything but outstanding. I'm not saying that this type of therapy is not going to be a miracle for a handful of patients, I just think that without improving the overall immunocompetence of the patient, it will never achieve tremendous efficacy.
All the best,
Paradigm |
