General Immuno-Oncology Questions
Q: Are humanized mouse models appropriate for cancer vaccine research?
To get sufficient vaccine response, you need interaction between antigen presenting cells and T cells. You might want functional B cells. There is suboptimal interaction between antigen presenting cells and T cells in the HSC humanized models because the T cells are trained on the mouse program, while the antigen-presenting cells are trained on a human program. This results in a limited response.
A better model would offer training for human T cells on the human thymus. Several models that accomplish this have been described in the literature; the BLT mouse, for example, in which a piece of human thymus is engrafted into a NOG mouse. They can be challenging to use because of the potential for graft vs. host disease. HLA transgenic models may also be useful for cancer vaccine research.
Q: To what extent are humanized mice being used for immunotherapy toxicology research?
Cytokine release syndrome ("cytokine storm") manifests within six hours in PBMC-engrafted mice at clinically-relevant doses of TGN1412, a monoclonal antibody associated with cytokine storm in humans. Helping to further illustrate the importance of this result to immuno-toxicology research, TGN1412 at doses 500-times higher than toxic doses in humans had no ill-effects in GLP-compliant non-human primate studies.
Beyond cytokine release syndrome, FDA researchers have begun to explore the utility of humanized NOG and NOG-EXL mice for evaluating adverse events associated with immunotherapy treatment. Their preliminary results indicate humanized mice can experience profound and dose-dependent adverse autoimmunity in response to checkpoint inhibitor therapy.
While humanized mice are relevant as toxicology tools, the chimeric nature of humanized models means they likely cannot be leveraged to evaluate safety events associated with off-target cross-reactivity for biologics. Similar to pharmacodynamics studies, the utility of humanized models in tox analyses will depend on properties of the therapy and its putative mechanism.
Q: What are some other research applications for humanized mice, outside of oncology?
There are established uses of humanized mice to study HIV, IBD, and other conditions in which immune response is relevant to preclinical research. The utility of humanized models is most thoroughly demonstrated in immuno-oncology, but its relevance to evaluating immunotherapy candidates suggests a much wider scope of application.
Q: What sort of PBMCs are used to make huPBMC-NOG mice?
Once identified, you can purchase whole lots from a single donor.
Q: Have PBMCs from cancer patients been engrafted and studied in NOG mice?
One recent example used hIL-2 NOG mice to create an autologous tumor/tumor-infiltrating lymphocyte model that successfully reproduced the patient-donors' responses to adoptive cell therapy.
Q: Can NK cell bispecifics be studied in the huPBMC-NOG model?
In part II of this webinar series, we'll discuss next generation NOG models, like hIL-15 NOG, in which PBMCs injection provides successful human NK cell engraftment.
Q: What are the relevant differences between engrafting human PBMCs and engrafting purified CD3+ T-cells in NOG mice and would there be an impact on tumor growth?
The added costs and labor associated with purifying CD3+ T-cells is one drawback to their use. Clearly, some component of PBMCs contributes to T-cell function in the model and loss of certain functionality in purified CD3+ T-cells is a valid concern. However, I'm unaware of data describing relevant functional differences that impact tumor growth or response to therapy in mice engrafted with PBMCs vs. purified CD3+ T-cells.
Q: Is the huPBMC-NOG model appropriate to study either checkpoint inhibitor or checkpoint inhibitors in combination with a traditional chemotherapy agent?
Prior to using huPBMC-NOG mice for these types of studies, it's important to consider the typical four- to five-week study window provided in the model before graft vs. host disease typically manifests. If pilot data suggest tumor-growth kinetics or resolution of efficacy may require longer than three weeks, researchers should consider CD34+ HSC-engrafted huNOG mice as an alternative model.
Q: How many huNOG mice can be generated from a single hematopoietic stem cell (HSC) donor?
Q: Whole body irradiation is critical for engraftment success in the huNOG model. If a research doesn't have access to an irradiator, is there an alternative preconditioning method which can be used prior to HSC engraftment?
Q: How was 25% humanization chosen as the cut-off specification? Is there any correlation between humanization ratio and immuno-oncology (IO) agent efficacy in huNOG studies?
Q: Use of huNOG mice from multiple donors in a single study is a best practice. How many donors are recommended for studies with checkpoint inhibitors?
The only strong recommendation is to recognize the additional layer of variability humanized immune system models can bring and to use more than one donor for your study. Furthermore, even if a therapy fails to inhibit the growth of a tumor, consider still evaluating the important immune cell(s) targeted by the therapy. This is certainly true for checkpoint inhibitors where CD8+ T-cell expansion and increased tumor infiltration may still occur despite no significant effects on tumor growth.
Q: Why is HLA matching between HSCs and the tumor unnecessary? Does this mean graft vs. host disease contributes some of the efficacy in humanized-mouse studies?
Accordingly, several independent groups have illustrated HLA matching of engrafted tumors and CD34+ cells are dispensable for checkpoint-mediated tumor-growth inhibition in humanized mice. The etiology of checkpoint-mediated efficacy in huNOG mice potentially involves mechanisms similar to allograft rejection.
GvHD is not thought to play a role, because huNOG mice do not suffer from GvHD. However, huPBMC-NOG mice do manifest GvHD. Hypothetically, GvHD could contribute to efficacy, especially if the administered therapeutic is associated with increased T-cell infiltration into engrafted tumors.
Q: What is the mechanism through which T cells in mice engrafted with CD34+ cells show anti-tumor efficacy in response to a checkpoint inhibitor?
However, the HLA/MHC mismatch does not completely exclude the possibility of some antigen-specific recognition of tumors by T-cells. A study evaluating the tumor-infiltrating T-cell repertoire following checkpoint inhibition could shed light on whether some antigen-specific responses do occur and the extent to which they contribute to efficacy.
Q: Is administration of recombinant human cytokines an effective strategy to enhance certain human immune cell lineages that don't establish well in huNOG mice?
This approach has some strong limitations. Human cytokines tend to have short half-lives in mice, which translates to multiple administrations, increased experimental difficulty, and increased costs. For basic studies, delivering recombinant human proteins is a great option, but preclinical studies involving many more animals may require a transgenic model.