Biochemistry III
Dr. Julin
Merle Zimmermann
May 12, 2003

Using a Virus to Correct a Genetic Defect [Internet Version]

The Story:

Research, a virus, X-SCID, treatments.

A Spontaneous Recovery:

A chance mutation, the goal of the experiment.

The Experiment:

Extraction, Infection, and Reintroduction.

How was the recombinant virus designed?

A wild virus, different genes, reverse transcription, modification, and a trick. The actual sequence.

The wild-type MLV contains three genes which, when expressed, make proteins that wrap the virus for transportation and reverse-transcribe it into the host DNA once it reaches the host cell. Reverse transcription is a process in which a protein translates RNA into DNA and inserts it into a preexisting DNA molecule. The name comes from the fact that DNA is usually translated into RNA before protein expression, and in this case the opposite process occurs. Reverse transcription is not uncommon; most of the human genome consists of copies of old reverse transcriptase genes, and recently genes of similar (non)function have appeared in fruit flies as well (4).

The wild-type virus isn't very useful, however. It does not do anything beneficial by itself, and it is impossible to directly insert a human gene sequence of any length into the viral code without rendering the virus inoperative. The key to the problem lies in the structure of the virus itself. It is composed of three major parts: the binding sites that its proteins recognize, the three genes mentioned earlier, and a terminal sequence that appears on both ends (which are used by the viral reverse transcriptase). For the virus to be effective in treating X-SCID, it only needs to add the corrected gamma-c cytokine receptor gene, and not any of the genes needed to reproduce itself. If the addition of the viral gene makes the cell hardier, then the cell's natural division will ensure spread of the added DNA. The problem then becomes to make a functional virus that doesn't contain any of the code used to replicate itself.

The solution described in Viral Vectors for Gene Transfer proceeds as follows: If a virus is manufactured that does not contain the binding site for the viral proteins, then it can be used to infect a cell, which will then produce the viral wrapper proteins. The proteins will not attach to the virus in the cell, because it doesn't have the binding site. However, if the cell also contains a sequence composed of the viral binding site followed by a copy of the desired gene, then the proteins produced by the defective virus will find the binding site on the other sequence and produce "viruses" which insert the desired gene rather than the viral ones (5).

In the experiment described in (1), the custom virus used to treat the patients' cells was in fact ordered from a commercial supplier. This is not the only option, however. Since the sequences of the MLV and the correct gamma-c cytokine receptor genes are readily available online (6, 7), it would be easy to create the necessary viral vectors in an academic laboratory.

After the Experiment:

A surprise, Leukemia, LMO-2, the FDA, an Observation.

Sadly, this caused the FDA to halt similar trials in the US, even though the cancer did not appear until more than three years after the initial treatments (12), and the life expectancy of X-SCID without treatment is less than one year (13).

References: