CAR-T's for Solid Tumors
Safety Assessment
Regina Kelder

CAR-T's for Solid Tumors

Scientists study the immunotherapy strategy in brain cancers, plus a real pissing match in the mouse cage. Day Three from SOT.

Chimeric antigen receptor (CAR) therapy is an immunotherapy that harvests a patient’s T cells and then edits the cells to recognize antigens on lab-adapted tumors. The experimental strategy has eliminated blood cancers in some clinical trials, but it hasn’t been tested much in patients with solid tumors.

That seems to be changing. At an SOT session Tuesday on immune-oncology, Laura Johnson, who directs the Solid Tumor Immunology Laboratory at the University of Pennsylvania, described work that her group has been doing on glioblastoma, an aggressive brain cancer with a median survival rate of about 14 months. “This is one of the worst tumors imaginable,” said Johnson. “People are always surprised when I tell them that it kills more people per year than melanoma,” says Johnson. (13,000 deaths compared to 9,700 deaths respectively).

About a third of these brain lesions express a mutated tumor-specific protein called epidermal growth factor receptor variant three (EGFRvIII). Johnson’s group and others have targeted it using CAR T-cells in syngeneic and xenogeneic (cross-species) mouse models of glioblastoma.

Using CARs in attacking solid tumors is tricky because most of the receptors aren’t specific to tumors. This means that the CAR T-cell therapy can induce unintended responses in healthy tissue as well, triggering severe adverse reactions and even patient deaths. EGFRvIII has an advantage over other receptors because it is only represented on tumors. Johnson showed, in two very quick time-lapsed videos, the CAR therapy “literally mopping up the tumor cells.”

Before progressing to the clinic, her lab conducted a battery of in vitro and in vivo studies to rule out any cross-reactivity to EGFR—the unmutated form of the protein— and found no toxicity or cross-recognition of normal EGFR by EGFRvIII CARs. UPENN is now treating glioblastoma patients with the EGFRvIII CARs.

Johnsons said syngeneic mice—mice with intact murine immune systems—showed that that an infusion of EGFRvIII mCAR T cells would cure the brain tumors. In xenogeneic mice, the CAR-T therapy was able to control tumor growth and indicate that intravenous administration is likely a suitable route to target CNS tumors with T cells.

But Johnson said a fully immune-competent syngeneic mouse would be the ideal in vivo candidate to evaluate the impact of EGFRvIII CARs on human tissue.

A Real Pissing Match

A study conducted at Charles River’s Safety Assessment site in Montreal finds that mice pee well in the sandbox! At the request of a client, Principal Scientist Nancy Doyle and her colleagues examined an alternative method of collecting urine from group-housed mice. They lined the metabolic cages with the sand—a sterile, hydrophobic product that keeps the urine on top until it evaporates—and collected the miniscule amounts of urine, between 1/6 to 8 drops per mouse, every five hours.

The sterile, hyrophobic sand is not a new tool, but in large research studies where urine collection is needed to collect data, using this strategy can be a bit challenging. Still, a drug developer preparing to test their compound in 240 male and female mice that were being group-housed in cages of three had a goal. The company did not want the animals separated in the course of the study because it made the animals cranky. “They [the client] wanted to keep the fighting to a minimum,” says Aurore Varela, the Scientific Director of the Musculoskeletal Research & Imaging at Charles River in Montreal who presented the poster at SOT on behalf of her colleague.

So rather than remove the animals to obtain the urine using minimally invasive methods, a common method in research studies, the client wondered, would it be possible to gather the urine in a group-housed setting without compromising the toxicology data? The answer, it turns out, is yes.

The Montreal site first needed to pilot the idea to verify whether the sand technique would interfere with the urinalysis and urine chemistry parameters. The study compared results from female singly housed mice in metabolic cages to animals group-housed in cages lined with the sand. By and large, the lab results were comparable, but Varela said they did learn some things to help prevent potential evaporation of the urine. Collecting the urine in the afternoon is best. So is transferring the pee in a capped polypropylene tube. Multiple sampling approaches is also better than collecting once a day.

End of story: The client opted to use the sand technique for their study and it worked very well.  

Dietary Findings

Laboratories like Charles River have switched to lower protein diets for their lab rats to keep the animals healthier and expand life expectancy, but findings presented at SOT seemed to refute this theory. A large retrospective study led by Rina Massarelli, a Research Scientist in Toxicology at Charles River’s Montreal site, found that among the 2,400 rats used in various studies between 2008 and 2016, eating a higher protein diet didn’t have much impact on survival .