WHG - I am currently holding a very small position in Ariad. I had had a large position in the warrants; 85% were sold @ $33-38, the rest @ $2, in that order. As with most selling decisions (IMO) that appear to be extremely wise, I can assure you this windfall was the result of very lucky timing.
Your questions are excellent and appear to at least partly arise from the lack of clarity of my prior post.
Which promoter are you comparing it to? Is it a viral promoter (e.g. CMV, SV40, etc)? An endogenous promoter? Does the promoter also contain all of the promoter enhancement regions?
My post discussed, in effect, replacing Wilson's construct (which probably used one of the constitutive promoters you mentioned [SV40 etc.]) with one of Ariad's constructs containing their proprietary promoter. Thus I was comparing Ariad's inducible promoter to the construct used by Wilson. I'm not sure if the efficacy of these two approaches has ever been compared. RAPID allows one to conduct an in vivotitration of the transgene: give a low level of the dimerizer to start and increase it until the desired outcome is achieved. My point was that this testing could begin at a significantly lower viral particle titer; if the desired result was NOT observed when RAPID was fully induced, higher dosing of the viral vector would THEN be mandated. At a minimum, the patient's risk of a potentially lethal immune reaction to viral load would be reduced.
What if the transgene needs to be constitutively expressed? What if the timing or the length of expression is unknown?
As I noted in a post around the time of the Science paper (http://www.siliconinvestor.com/readmsg.aspx?msgid=12795587), the 'leakiness' of the RAPID system can be pre-set by altering the number of Fm repeats in the cassette. Thus, as described, if only one CAD is formed then the system is extremely leaky i.e. it is essentially constitutive (and also essentially unregulatable). The number of Fm motifs is determined for each application of RAPID based on the specific parameters of the clinical issue being addressed. Regarding the timing of expression: I was somewhat surprised at this question since Ariad has produced a ream of statistics over the years showing blood levels of gene products before and after dimerizer dosing. In fact the entire dimerizer concept is based upon the idea of tightly regulating gene expression. The fact is that with both ARGENT and RAPID (the non-leaky construct) there is virtually no expression in the absence of the dimerizer. If I am misinterpreting your question - that you're actually asking about the timing of expression of the normal, endogenous gene - that would fall into the category of a problem faced by everyone in gene therapy....a problem that can only be addressed by tightly regulated clinical trials.
Or why not use a spliceosome instead of replacing the whole gene, thus you would need a constitutive promoter to drive the transgene...and would not need RAPID as the cells endogenous promoter would be used?
I think there are a number of misunderstandings here. In distinction to the approaches of others, Ariad does not produce transgenes to interact with endogenous genes. Recall ARGENT/RAPID are episome-like, inducible cassettes that are activated by the dimerized transcription factors previously infected into the cell. Ariad's constructs are self contained systems with all the elements necessary (in the cellular environment) to transcribe and translate a gene. Ariad's approach never involves replacing anything. ARGENT and RAPID supplant or supplement endogenous genes rather than deleting or interacting with them. I cannot comment on the advantages and disadvantages of gene replacement approaches such as spliceosomes. My impression is that there are clear scenarios where replacement therapy is indicated (such as when an endogenous gene is mutated to produce a deleterious gene product). While I cannot claim that the gene therapy world is divided in to nice niches, that would not appear to be Ariad's space.
Your subsequent comments about RAPID's 'main usefulness' is, IMO, a limited perception. RAPID could be used in cases where there is a complete lack of expression of a regulated gene product (the Science paper was, after all, addressing a diabetes model), or if there was over expression of a gene, or if there was no mutation in the promoter/enhance/coding sequence (for example, a dysfunctional transcription factor can completely quench transcription).
And let's not forget that it's not so easy to target endogenous promoters. There's much more than the sequence of bases - secondary/ tertiary structure(s), recognition and activation from histone complexes, etc. - that makes this approach difficult at best. The last time I checked, Sangamo is addressing only one class of structures (zinc finger) involved in promoter activation. What if a specific endogenous promoter has an extremely complex or unidentifiable promoter/enhancer structure? Mother nature's work is always the best; it is also often very difficult to replicate and/or 'correct' when it goes wrong.
Please explain the VLTS system to me. Perhaps it is better than ARGENT and RAPID. My last contact with Tet-based systems indicated they had significant leakiness problems. Is that still the case? Is Tet-off tight?
I studied the regulation of expression of nerve growth factor in rodent glial cells (astrocytes and oligodendrocytes) and microglia for my thesis work (thanks for asking!). This was during the time when folks thought that NGF would save the world from Alzheimer's. The project was mostly a humbling lesson in the complexity of the brain: if anyone ever claims a growth factor, cytokine, prostaglandin, antioxidant, radical gas, etc. will 'cure' a neurological pathology, please, please, please ignore them. Neuroscience can currently moderately ameliorate the symptoms of the major brain-related illnesses and can do nothing about the underlying causes. In addition, always check the specialty of the training of those providing neuroscience information - I can't tell you how often I've read outright nonsense relating to neuroscience on SI from those considered to be, in theory, biotech gurus.
Needless to say be very, very careful of biotechnology companies in this field. Neuroscience is at the stage of providing purely palliative treatments, despite the seemingly rarefied tone of those claiming so much more. Trust me on this one.
Take care,
Scott. |
