Xenograph models to help children resist cancer

Acute lymphoblastic leukaemia (ALL) is the most common cancer in children and despite the remarkable successes with current treatment regimes, resistance to therapy remains a problem that cancer researchers are trying desperately to understand.

Dr Richard Lock is known internationally in the cancer field for his work on cell cycle control and drug resistance mechanisms. Since arriving in Australia eight years ago, Lock has focused on developing a laboratory model for the in vivo growth of human ALL cells - the first such model in Australia.

Lock joined the Children's Cancer Institute Australia for Medical Research as head of its leukaemia biology program in 1998. He hails originally from the UK where he did his undergraduate studies and PhD, but came to Sydney after a stint at the University of Louisville, Kentucky.

The long-term goal of his work is to develop treatment strategies for childhood leukaemias specifically able to overcome drug resistance and thereby enhance survival.

As an invited speaker at the 19th Lorne Cancer conference earlier this month, Lock spoke about two aspects of his work - identifying novel mechanisms of drug resistance in childhood leukaemia using xenograft models, and an international collaboration for prioritising treatment for childhood malignancies.

Good news, bad news

More than 95 per cent of children with ALL respond to initial chemotherapy and achieve complete remission in the short term. Happily, more than 70 per cent will remain free of disease. Less happily, 25 to 30 per cent will relapse, and three in four of those will ultimately succumb to the disease.

This is due to the leukaemia developing resistance to the chemotherapy drugs, with the best treatment option left an allogenic bone marrow transplant.

The anticancer drugs that Lock focused on in his talk are the glucocorticoids. These agents are critical components of many chemotherapy regimens and are among the most effective drugs used in the treatment of childhood ALL.

Resistance to glucocorticoid therapy remains an important factor in achieving a 100 per cent cure rate and strategies are needed to overcome these resistance mechanisms. Lock's team has made significant advances over many years in delineating ways that the leukaemia cells develop resistance to glucocorticoids and other drugs, using a preclinical xenograft model of ALL.

At Lorne, Lock discussed this model and the insights it is delivering into resistance to standard drugs used to treat malignancies, in a project funded by the Leukaemia Foundation of Australia.

In vitro discrepancies

The xenograft model was developed to overcome inherent deficiencies in the in vitro cell line model systems available, Lock says.

"In the past, the standard method for testing anticancer agents was to take a cell line and treat it with your drug of choice to analyse mechanisms of resistance," he says. These findings then had to be translated back to the clinical disease, where there was often a 'disconnect' in results.

In fact, Lock says, "discrepancies have been consistently noted in resistance mechanisms between in vitro studies and clinical biopsy specimens".

Lock's laboratory has been developing the ALL xenograft models in stages since 1998. Setting up such a model involves taking primary biopsies from children with ALL and engrafting them into highly immunodeficient (NOD/SCID) mice. This mouse strain appears superior to previous strains tried in its ability to engraft ALL cells, and Lock's group reproducibly achieved high engraftment rates and dissemination of the disease to the same sites as in the patient.

"The systemic disease in the human is simulated in the mouse," he says. "We observe the same primary organ affected, the same pattern of dissemination and importantly, the same resistance mechanisms as in the patient."

The ALL xenografts therefore represent a continuously available, sensitive and accurate model of the clinical disease. They have now established continuous xenograft panels representing all the ALL subtypes and a range of leukaemias showing resistance, even some xenografts derived from long-term survivors.

This bank of samples is always ready for testing, he says. "All we have to do is take the cells out of the freezer and put them back into NOD/SCID mice for screening." It is important to emphasise that these grafts are inherently resistant to drugs in the same way as in the patient from whom the cells were originally derived, he says.

Prioritising therapies

Lock will also discuss a major project he is involved in with several groups in the US to prioritise new therapeutic agents for inclusion into trials in children with cancer, the Pediatric Preclinical Testing Program (PPTP), an initiative organised through and funded by the US National Cancer Institute (NCI).

Lock is one of the six project leaders and the only one participating from outside the US. "Novel drugs are being developed all the time, especially with the range of new genetic and biochemical targets being identified, and advances in technology to produce candidate drugs," Lock says.

"There are just not enough children to test all the drugs in clinical trials, so we need a pre-clinical prioritisation mechanism."

Across all the groups directly involved and other collaborators, the project already has large panels of xenograft models available that represent almost all common childhood malignancies. The program is testing 12-16 novel drugs per year with pre-planned stages of testing.

In the initial stage, new drugs are screened as single agents. The best ones out of that round will be tested in combination with selected standard chemotherapy drugs, and finally, promising candidates will be put through pharmacokinetic testing in preparation for clinical trials.

The project was initially set up to run over five years and is now into its third. Its goal is to generate the kind of information that will allow paediatric oncologists to make educated, reliable decisions on which new agents should be tested in children with specific cancers so that only those with the best chance of success go on to clinical trials in patients.

The overall importance of his work is that "it offers real hope that in the future we will be able to design specific combination therapies to overcome drug resistance in children with leukaemia", Lock says.

In the short term, researchers are continuing the huge amount of pre-clinical testing required in the ALL model systems (as well as trying to maintain the sanity of those doing all the mouse work).

In the longer term, "we want to get as many promising compounds into clinical trials as quickly as possible - that is what we are here for". According to Lock, there are some very exciting compounds coming through the pipeline and that is very good news.