Editorial Article
Monday August 23, 2010
by Caitlin Smith
It used to be – back in the days of early cell culture, when things were simpler – that there were cell lines (that had to be tended to and passaged), and there were primary cultures (that were finicky and didn’t last long). Now it seems there is a growing area in between, with cells acquiring immortality at different points along their developmental pathways. In addition, screening technology has made it possible to study more cell lines than ever before in pursuit of cancer therapeutics.
Different types – and degrees – of immortality
At ATCC (American Type Culture Collection), they are excited about the hTERT (human telomerase reverse transcriptase) technology they have licensed from Geron Corporation. Telomerase is an enzyme that maintains telomere length, enabling cells to divide indefinitely while retaining normal function and phenotype. The hTERT technology allows researchers to engineer their own immortalized cell lines. Furthermore, “what makes these hTERT cell lines so fascinating is that [hTERT] enables the immortalization of a particular cell at a unique moment in its differentiation,” says Robin Rothrock, market development director for cell biology and biostandards at ATCC. “So it is possible to immortalize a completely normal cell – one, for example, that is moderately transformed in terms of cancer. And then the cells can be immortalized along a complete differentiation pathway. That is something that we are very excited about.”
As a cell repository, ATCC is an invaluable resource for any researchers using cell lines. They have a newly expanded portfolio of primary cells, an evolving collection of hTERT immortalized cell lines, and over 3400 continuous cell lines (over 900 of which are cancer cell lines). “In addition to the hTERT cell lines, we also have available the hTERT plasmid so that researchers can make their own hTERT cell lines,” says Rothrock. “And then after they make the cells, they have the option of depositing them with ATCC.”
Despite the power of hTERT technology, creating an immortalized cell line can still be a challenge in some cases. “Some cells can achieve this immortality just by using hTERT, and other cells will need to use some of the viral genes as well,” says Rothrock. “So it is definitely not an easy task.”
Screening cell lines
Though there are many cell lines that have become more popular among academic researchers or clinical researchers for their particular properties, many lesser-known cell lines exist that may hold important properties yet to be discovered – especially in connection with new compounds that could hold keys to fighting diseases such as cancer. A 5-year collaboration between the Cancer Genome Project at the Sanger Institute and Massachusetts General Hospital aims to screen 1000 human cancer cell lines against 400 pre-clinical and clinical compounds. The screening process will cover all of the tissue types typically seen in clinics.
“To date, this is the largest-ever screen of this type carried out, not in terms of the number of cell lines but also with respect to the number of pre-clinical and clinical compounds,” says Ultan McDermott, CDF Group Leader for the Cancer Genome Project at the Wellcome Trust Sanger Institute. “Cell lines are incubated in a range of concentrations of each drug, and after a defined time point, cell viability is measured using a fluorescent read-out. Our increasing knowledge of cancer genomics has highlighted the high degree of genetic heterogeneity between cancers, even when they arise from the same tissue type. It is therefore critical to have sufficient scale in any such screen to capture these subsets of cancers. Once we have defined the sensitivity profile of a particular drug against the 1000 cell lines, we use a variety of statistical tools to correlate that pattern with the genomic data we have already captured for these lines (somatic mutations, genomic deletions, and amplifications and rearrangements).” McDermott notes that next generation sequencing is an exciting tool that can aid them in sequencing the genomes and transcriptomes of cancer cell lines – another tool in discovering new biomarkers of cancer therapeutics sensitivity.
One detail that distinguishes this collaborative effort from other screening projects, according to McDermott, is that they release all of their screening data every three months (via their website. “We view this project as providing an important resource to the research community,” says McDermott. But despite the large number of cell lines studied, it is by no means the complete picture. “Although a large number of cell lines have been developed for some of the major cancer types (e.g. lung, breast, colorectal), other cancers are poorly represented and therefore we do not have good in vitro models in which to test the new therapeutic agents under development,” says McDermott. “There is no doubt that recent publicity regarding ownership of cell lines, as well as the ethics of using tissue in this way, is deterring many researchers from actively developing new lines today.”
iPS cells: the reprogramming challenge
One of the most exciting developments in cell lines today is the creation of induced pluripotent stem (iPS) cells. Because these can be derived from adult somatic cells and artificially made pluripotent, the technology generates excitement as it avoids two stem cell problems: it avoids the controversy of using embryonic cells, and it lessens the chance of immune rejection if returned to a patient for therapeutic purposes.
“Millipore offers a novel method to efficiently create induced pluripotent (iPS) cell lines using a polycistronic vector,” says Louise Rollins, product manager for cell biology at EMD Millipore. “The STEMCCA Lentivirus Reprogramming Kit contains all four Yamanaka factors.” Yamanaka factors refer to the genes identified by Shinya Yamanaka’s research team as being important in the production of pluripotent stem cells. “Having all the factors in one vector greatly increases the efficiency of reprogramming and reduces viral integrations when compared to transfecting separate lentiviral vectors,” says Rollins.
While the generation of iPS cells is an exciting area of research today, a major challenge facing the field is to identify a safer method to make iPS cells so that we can take advantage of them for clinical therapeutics. “Scientists are challenged with finding more efficient, safer means of reprogramming cells using non-viral based technologies,” says Rollins. “Improved solutions in these areas will increase the pace of research and increase the possibilities for stem-cell-based therapies.” iPS cells also hold great promise for the biological modeling of different human diseases. “Researchers can now take diseased cells from a patient and create iPS cells that are genetically identical,” says Rollins. “Presumably, this will lead to more predictive models and, ultimately, more effective therapies.”
A valuable tool that many would like to have, according to Rothrock, “is a robust way to make iPS cell lines from a variety of different tissue sources. And also, understanding how iPS cell lines differ from embryonic stem cell lines in terms of what they can, and cannot, do.” Though iPS cells generate a lot of excitement among cell biologists, how many people actually use them? “We are working to better understand how many researchers are exploring the use of iPS cells,” says Rothrock. But we know that it is increasing. As is true for the study of embryonic stem cells, work in mouse iPS model systems is being rapidly adapted to create human iPS cell lines.
The fast-paced development of iPS cell research reflects the quickly evolving technologies surrounding them. “This truly defines cutting edge research,” says Rothrock. “The initial report of the creation of iPS cells in mice, was very quickly followed by publication after publication enhancing that methodology. There is a challenge not only in the components, but also in the efficiency, and then how the efficiency is affected by the choice of what type of cell.” But like a lot of exciting research, it generates more questions than answers. “Do you differentiate back to that pluripotent state?” says Rothrock. “And then understanding what those cells look like and how they behave, what they remember and what they don’t remember... Then the challenge is, what if you want to send them down a differentiation pathway? How do you do that? How do you control that? How do you know that you’ve developed the cell type that you were looking for? Then it becomes again a question of characterization.” Perhaps the next chapter in the iPS story will be solid methodology for characterizing new cell lines.
The image at the top of the page is ATCC's hTERT immortalized mammary epithelial cells.