Cell & Gene Therapy Answers: Preclinical Oncology

Your source for answers on how to assess the preclinical efficacy of cell therapies.

As the landscape of cell therapy continues to evolve, so are the tools to assess the efficacy of cell therapy in the preclinical space. To better understand Labcorp’s preclinical oncology role in cell therapy development, we spoke with Sheri Barnes, associate director of scientific development for Labcorp preclinical oncology, who assists drug development sponsors in study design and model selection for preclinical oncology studies, along with David Draper, associate director of scientific development for Labcorp preclinical oncology who helps sponsors understand how in vitro services can be used effectively to address their preclinical investigational needs.
 

How did preclinical oncology get started in the cell therapy space?

The preclinical oncology site in Ann Arbor, Michigan, was acquired by Labcorp in 2019 but was originally founded as an imaging-based CRO in 2003. At the time, the site was the first CRO with an in vivo imaging system (IVIS) for bioluminescent imaging, which led to extensive experience with monitoring disease progression of systemic luciferase-enabled cancer models by bioluminescence. In 2013, the site had more than 50 heme cell lines established, many of which were luciferase enabled. Because the development of CAR-T cell therapy initially targeted heme malignancies, the preclinical oncology site was well positioned to help develop cell therapies for cancer and address significant industry needs for these models.

With several established heme malignancy models, the preclinical oncology site ran its first cell therapy study in 2015 for a CAR-T product. By 2017, the site had supported over 100 CAR-T and TCR cell therapy studies for 12 sponsors, a count that has now grown to more than 500 cell therapy studies—including both heme malignancies and solid tumors—for more than 30 unique sponsors. 

The development of solid tumor models in the NOD.Cg-Prkdc^scid Il2rg^tm1Wjl/SzJ (NSG) mouse helped the site explore solid tumor efficacy of cell therapies. Because the preclinical oncology site already had expertise with these models, it was easy to adapt these models to the NSG mouse. Furthermore, the team also has several specific flow panels geared toward persistence and activation states of these cell therapies, which can help enable molecular characterization of cell therapies in vivo following tumor engagement. Together, these services help meet industry needs to develop solutions for cell therapy development.

What tools are currently available to assess efficacy of cell therapy in the preclinical space?

The preclinical oncology site has a range of in vivo and in vitro tools to assess efficacy of cell therapies preclinically. Beyond human tumor models, the site has characterized both heme and solid tumor modeling in the NSG mouse. The site’s specialists also develop sponsors’ cell lines for their use in an applicable mouse strain. Both solid and liquid models have been used successfully for screening CAR-T,TCR, NK-CAR, and iPSC constructs, and manufacturing lots, assessing donor-to-donor variability as well as cell therapy combination studies.

While these models have great utility, they are not without their limitations. The human immune system is not reconstituted in these mice, so while they are very good for assessing T-cell related activity against a tumor, they are missing the myeloid compartment, which could introduce immunosuppressive elements that are not present in this system. As a result, these studies do not typically experience related toxicities, which limits the predictive nature of these studies.

Scientists are working to provide better modeling for human cell therapies. For example, mice expressing human IL-2 or IL-15 can support cell therapy health in the model while major histocompatibility complex (MHC) class I/II knockout mice can help reduce the onset of characteristics associated with graft versus host disease.

In addition to in vivo capabilities, the preclinical oncology site has a strong team of scientists performing in vitro studies. The most common cell-based assays at the site assess the cytotoxicity, or cell killing, of a cell therapy product against a target tumor cell. These assays are a great way to get an early read on activity, and the site has demonstrated that these assays can be predictive of activity in an in vivo study. These assays can be run in a 2D model system as well as a 3D model system. The 3D model system is supported by Predictive Oncology to model interactions between a tumor and its surrounding tissue, and the site has demonstrated cell therapy activity against an acute lymphoblastic leukemia (ALL) model in a 3D bone marrow matrix using this platform.

The preclinical oncology site also performs in vitro assays to assess cytokine release to ensure that the CAR-T cells can produce proinflammatory cytokines in response to a target cell. Through these in vitro and in vivo assays, they can produce critical decision-driving data for the early development of cell therapies.

What ex vivo tools are available to assess cell therapies?

Ex vivo assessment tools analyze cell therapies after they’ve had time to exert their activity in vivo. The data complement the efficacy evaluation to provide a clearer picture of the mechanistic action of the treatment.

In the preclinical space, the tools most frequently utilized can be broadly described as immune monitoring tools. They monitor the quantity and the persistence of the product in the host over time. For years, flow cytometry has been the gold standard tool for this analysis. Using CAR-T therapy as an example, flow cytometry is used to measure the number of CAR T-cells circulating in the blood of recipient models but can also be applied to other tissues, including solid tumor analysis. A benefit of using flow cytometry is its ability to characterize the various subpopulations within the product and their activation state. Conventional CAR-T therapies are heterogeneous in nature. Using flow cytometry to examine the persistence and phenotype of these subsets can provide valuable mechanistic insight into therapeutic activity.

Other tools include immunohistochemistry (IHC) as well as approaches based on polymerase chain reaction (PCR) such as qPCR and digital droplet PCR. PCR is an alternative to flow cytometry that monitors the cell therapy by detecting unique DNA sequences that are engineered into the cells, such as those that code for the CARs in CAR-T products. It is useful for both biodistribution analyses as well as product persistence. Unlike flow cytometry, PCR does not require fresh tissue for analysis, which adds logistical flexibility. But for this, the rich immune subset phenotyping data that can be provided by flow cytometry are sacrificed.  

IHC is a technique used to quantify immune cells in cross sections of preserved tissue and measure adoptive cell therapy biodistribution across tissues in preclinical models. With solid tumor analysis, IHC can help determine the capacity of the product to penetrate the tumor, which is one of the main challenges faced by cell therapies that are designed for solid tumor treatment. IHC is also useful for examining the spatial relationships between the product and other targets in the tumor tissue.

There are also tools used that are not for immune cell measurements but rather the detection of proteins released by the cells as they become activated to inhibit tumor growth. Typically, proinflammatory cytokines, these biomarkers can be measured in the blood from tumor-bearing models after they receive the cell therapy by high-throughput techniques such as Meso Scale Discovery® (MSD) analyses that are able to detect multiple targets simultaneously. While there are certainly other tools available, the ones mentioned here are techniques most often combined to provide a comprehensive profile of the mechanistic action of adoptive cell therapy efficacy.

What are the typical assays required for IND-enabling work for cell therapies?

For adoptive cell therapy IND-enabling work, today most non-GLP safety work is suitable by U.S. FDA and other global agency standards, because oncology is the primary therapeutic area and the fact that patients need access to lifesaving medications. This work consists of in vivo studies in tumor-bearing murine hosts.

When GLP work is requested, study conduct is performed in non-tumor-bearing models or through in vitro assays to assess the persistence or uncontrolled growth of the cell therapy in the presence or absence of a target signal or stimulant.

The in vitro work is often performed in multiwell plates, with and without various positive cytokines to stimulate growth with the goal of measuring and characterizing any uncontrolled expansion. Samples are often collected at various time points to cover expansion and waning phases to support safety profiling “in a vacuum.” Analyses consist of flow cytometry measurements of cell counts and markers of cell identity. Samples are also collected to examine cytokines and other proteins produced by the cells, nucleic acid messages and even genome inspection should genomic rearrangement be considered as a potential safety factor.

In vivo GLP safety studies sometimes consist of similar tumor-bearing work. Most GLP work consists of an assessment of mortality, clinical observations, body weight changes, quantitative food consumption and clinical and anatomic pathology. The biodistribution of the product is also measured either by flow cytometry or PCR in both tumor-bearing and non-tumor-bearing models at dose levels at and exceeding those intended for patient administration. For pluripotent-derived cell therapies, the preclinical oncology site’s approach to safety evaluation is similar, however, the guidance aligns with many of the global agencies: these evaluations require a more thorough in vivo analysis, for a longer duration of time, within similar model systems.

Together, these studies are performed in conjunction with Labcorp’s quality assurance unit oversight of data collection in a part 11 compliant system, and with periodic review of study events. Based on agency guidance and input, these safety evaluations of cell therapies can be accomplished in either tumor-bearing or non-tumor-bearing models, or even in vitro systems that are non-GLP or GLP, depending on agency agreement and the science encompassed around the potential therapeutic.

How does the Labcorp preclinical oncology team see the landscape of cell therapy changing?

The landscape is changing very quickly. Six CAR-T therapies have been approved for oncology indications, and we expect many more cell therapies to follow suit in the coming years. Our preclinical oncology site hasa strong foundation of experience with a number of cell therapies, including CAR-T cells, TCR T cells, NK-CARs and iPSC-derived cell products. These cell therapies use many of the same tumor models as other therapeutic classes, so while each type of cell therapy has their own nuances, the in vitro and in vivo assays that we have established can be utilized across multiple cell therapy types.

In addition to newly approved cell therapies, we’ve witnessed new and interesting ways of using cell therapies. Some of the approaches we have seen are coinjecting multiple cell therapies against different targets, expressing multiple CARs in a single cell as well as using combination strategies with small molecules, oncolytic viruses and biologics, such as antibodies or peptides. These combination strategies can both help support the health and persistence of cell therapies, which can lead to improved efficacy. With our deep experience both in the cell therapy space—as well as in development of combination strategies with two or three or even more agents—our preclinical oncology site is well suited to help sponsors explore these avenues of research.

Our preclinical oncology team realizes that some companies may have potential combination partners to explore with cell therapies but lack specific cell therapy experience to execute these studies. To address this need, we have developed a tool cell therapy directed against CD19 that can be used in such a strategy. We have demonstrated activity of this CAR T-cell therapy both in vitro and in vivo, so it’s ready to go. We are currently exploring other targeted cell therapies for the same approach in solid tumors. Overall, we believe the future is bright for cell therapy, and we’re excited that Labcorp is playing a role in its advancement to address ongoing therapeutic needs and make a difference for patients.

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Hear from our experts in our CGTAnswers blog series on additional topics: ddPCR, CMC, oncology , Cell & Gene Therapy and Rare Disease.