In the battle against cancer it is very important to kill every tumor cell present; otherwise, the remaining malignant cells will continue to grow and form new tumors. Although adenoviral vectors are one of the most efficient vectors used, one injected dose is thought to infect maximally 10% of the tumor cells. Thus it is necessary to increase the efficiency of adenoviral infection. An approach to increase this efficiency is to allow the virus to replicate, thereby increasing the amount of infectious particles. Replication of the adenovirus inside these cells will eventually lead to cell lysis, thereby releasing newly formed virions within the tumor mass. The subsequent viral generations will continue this cycle of infection, replication, and cell killing, thereby killing more and more tumor cells, resulting in the elimination of the tumor mass. Obviously, adenoviral replication should be confined to the tumor cells. Adenoviruses that can only replicate inside malignant cells are termed conditionally replicating adenoviruses (CRAds).
Two different approaches can be distinguished in constructing CRAds, generating either type I or type II CRAds. Type I CRAds contain a specific deletion in the adenoviral genome based on genes that are differentially expressed between tumor and normal cells. Due to this mutation normal cells are not permissive to adenoviral replication. In tumor cells this host cell defense mechanism is deregulated, thereby creating the perfect environment for the virus to replicate.
A well-known example of a type I CRAd is the adenovirus Onyx-015, also known as d11520. This virus contains a deletion in the adenoviral E1B-region. As mentioned before, the E1A and E1B regions are important for the replication of the adenovirus. The adenoviral E1B 55-kDa gene codes for a protein that inhibits p53 function which is an essential step in viral replication. Onyx-015 is deleted for E1B 55 kDa, and thus unable to inhibit p53. Therefore in normal cells, where p53 is functionally present, Onyx-015 will not be able to replicate as p53 cannot be inhibited by this virus. However, in most tumor cells the function of p53 is already disturbed and the ability of the virus to inactivate p53 is not required anymore. This causes selective adenoviral replication in p53-deficient tumor cells. At this moment, phase I, II, and III clinical trails have been conducted with limited success.
Type II CRAds are adenoviruses where the expression of genes essential for adenoviral replication are under the control of a tumor-specific promoter. When this promoter is active, expression of this gene will result in replication of the virus. These promoters should mainly be active in tumor cells to ensure that replication predominantly occurs inside the malignant tissue. An example of a tissue-specific promoter is the prostate-specific antigen (PSA) promoter which is highly active in PSA-producing prostate cells and shows limited activity in other tissues. Placing a gene essential for replication directly under the control of the PSA promoter directs adenoviral replication primarily to prostate cells that express PSA. Thus this adenovirus will replicate inside prostate (tumor) tissue while sparing the other tissues. At the moment, phase I and II trials are being conducted. Another example of a tumor-specific promoter is the telomerase promoter, which is active in more than 80% of all tumors. An adenovirus which has the expression of a replication essential gene under control of the telomerase promoter can therefore replicate in a broad range of tumor types. Other examples of tissue- or tumor-specific promoters are the epithelial gycoprotein-2 promoter, which is active in most epithelial-derived cancers making it a good candidate for the treatment of many tumor types, and the tyrosinase promoter, which is highly active in melanoma cells.
Although the theoretical idea behind type I and type II CRAds is very attractive, some drawbacks exist. In the case of an adenovirus where the expression of the replication essential gene E1 was restricted to melanoma cells using the tyrosinase promoter, replication was shown to be specific for melanoma cells at low infectious units. However, at a higher dosage replication was also demonstrated in nonmelanoma cells. This loss of specificity might be due to the presence of adenoviral promoter-like sequences upstream of the tyrosinase promoter. These sequences may interfere with the specific regulation of the tyrosinase promoter, causing activation of transcription in cells which lack actual tyrosinase promoter activity. Another possibility is the presence of E1-like proteins inside the cell, compensating for the lack of adenoviral E1 expression in nonmelanoma cells.
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