Dana-Farber Cancer Institute researchers and their colleagues have demonstrated that a genetic error so scarce it can't be detected with some standard screening equipment is often responsible for the loss of effectiveness of front-line drugs against non-small cell lung cancer.

Investigators led by Pasi J?nne, MD, PhD, found that many non-small cell lung cancer (NSCLC) patients who become resistant to targeted drugs such as Iressa® and Tarceva® have a mutation in a single building block of the EGFR protein. The study, which will be published by the Journal of Clinical Investigation, demonstrates that even a minute mutation, present in tiny quantities, is sufficient to cause drug resistance in some cancers. The findings are currently available on the journal's Web site and will be published in its October print edition.

"The implication of our study is that perhaps many more patients than previously thought have this as their mechanism of resistance," says J?nne. "Identifying those patients will be important in the next generation clinical studies of drugs for NSCLC, which accounts for about 85 percent of all cases of lung cancer in the United States."

Iressa and Tarceva are used to treat NSCLC in patients whose cancer cells have abnormalities in the EGFR gene. It's estimated that 10-15 percent of NSCLC patients in the U.S. have such mutations, and up to 40 percent of those in Asia. In virtually all cases, however, the drugs lose their effectiveness with time – between six and 14 months, depending on the type of mutation a patient has.

Scientists have suggested that about half of NSCLC patients who develop resistance to Iressa-like drugs have a specific mutation in one of the chemical components of EGFR – amino acid 790 (designated T790M) – but it wasn't known whether this secondary mutation was enough to cause the resistance. To find out, J?nne and his colleagues did an experiment involving NSCLC cells from patients who responded well to Iressa. Researchers exposed the cells to Iressa in vitro for six months. As expected, the cells became resistant to the drug. Surprisingly, though, no T790M mutations turned up when the cells were analyzed with standard gene-sequencing techniques.

When researchers used a more sensitive technique, called HPLC (High Performance Liquid Chromatography), which J?nne and his colleagues had previously developed, they found that this cell line did have mutated T790M, but only in a few copies of the gene. Further, they found that the mutated gene – scarce as it was – was sufficient to confer Iressa resistance on the cells.

The reason that mutated T790M was so hard to find is because the gene for EGFR was not only mutated in the cells, it was also amplified – copied in numbers far beyond normal. The amplification basically diluted the evidence, but not the effect, of the T790M mutation.

The investigators also found that in lung cancer patients with EGFR mutations who had become resistant to Iressa, the T790M mutation was not identified by standard gene-sequencing procedures, but by the more advanced HPLC technique.

The discovery offers hope to some NSCLC patients who have relapsed after taking Iressa or Tarceva, remarks J?nne, who is also an assistant professor of medicine at Harvard Medical School.

"Drugs capable of blocking the activity of mutated T790M may be effective therapies in NSCLCs that have become resistant to Iressa and Tarceva. The key is to accurately identify patients who harbor the T790M mutation. Because the mutated gene is present in such small numbers, sophisticated tests like HPLC will be needed to find it," says J?nne. He added that similar challenges in identifying mechanisms of resistance may exist in other malignancies, such as breast cancer, where cancer-gene amplification is common.

Dana-Farber currently is conducting a clinical trial of a novel drug in NSCLC patients who have relapsed from Iressa and have the T790M mutation. "As we're learning to overcome drug resistance in other types of cancer, there is a real possibility we'll be able to make similar progress with this form of NSCLC," explains J?nne.

Jeffrey Engelman, MD, PhD, from Massachusetts General Hospital, is the lead author of the study. The study's co-authors are Toru Mukohara, MD, Kreshnik Zejnullahu, Jason Sun, Sean Tracy, Xiaojun Zhao, PhD, Ana Borras, PhD, and Bruce Johnson, MD, of Dana-Farber; Lewis Cantley, PhD from Beth Israel Deaconess Medical Center; and associates from Children's Hospital Boston and University of Texas M.D. Anderson Cancer Center.

The research was funded in part by grants from the National Institutes of Health and the National Cancer Institute.

Dana-Farber Cancer Institute (www.dana-farber.org) is a principal teaching affiliate of the Harvard Medical School and is among the leading cancer research and care centers in the United States. It is a founding member of the Dana-Farber/Harvard Cancer Center, designated a comprehensive cancer center by the National Cancer Institute.