Monocyte-derived cytokines IL-1 / IL-6 contribute to CAR-T cell-induced cytokine-release syndrome and neurotoxicity

This week Nature Medicine published two papers showing in mouse CAR-T model that cytokine-release syndrome (in both studies) and neurotoxicity (in one study only) were primarily driven by IL-1 and IL-6 cytokines released by monocytes following interaction with infused CAR-T cells. 

In the first study led by Michel Sadelain at Sloan Kettering Institute, New York, immunodeficient SCID-beige mice transplanted intra-peritoneally with human B cell tumor cell line and later infused with CD19 CAR-T cells develop cytokine-release syndrome (CRS) that were reversible by anti-IL-6 antibody,  injection.  

Injection of anti-IL-1 antibody, Anakinra, had similar protective effect against CRS-driven mortality without compromising anti-tumor effectiveness. 

However, due to some limitation of their mouse tumor model where CAR-T cells are of human origin and non-T cells such as monocytes are of mouse origin, the authors acknowledged that not all features of CAR-T cell toxicity could be reproduced here since "None of the reported pathologic findings indicative of neuropathology or associated with neurotoxicity (cortical laminar necrosis, hemorrhages, disseminated intravascular coagulation (DIC), gliosis or vasogenic, neurotoxic or interstitial edema) in human patients were observed in any of the mice examined in the present study".   

Fortunately, the second study is more extensive and fills much of holes of the first study. Here, the authors led by Attilio Bondanza at San Raffaele Hospital Scientific Institute, Milano, "transplanted human cord blood (CB) hematopoietic stem and progenitor cells (HSPCs) through intrahepatic injection into sublethally irradiated newborn NSG or triple transgenic NSG (SGM3) mice expressing human stem cell factor, granulocytemacrophage colony-stimulating factor (GM-CSF) and IL-3 and initially profiled lymphohematopoietic reconstitution." Basically, this mouse model, referred as newborn humanized SGM3 (nHuSGM3), had both T cell and non-T cell components of hematopoietic system mostly of human origin. In allogeneic tests, human T cells developed in nHuSGM3 mice showed expected functionality. 

Staining of cells in nHuSGM3 challenged with tumor and CAR-T cells that induced CRS showed that monocytes were producing IL-1 early on, followed by IL-6. 

Depletion of monocytes/macrophages with liposomal clodronate (LC) could rescue mice from CRS.

Importantly, while both anti-IL-1 and anti-IL-6 antibody injection could significantly reduce CRS, only anti-IL-1 antibody were able to reduce neurotoxicity in nHuSGM3 mice. Similar effect were seen with CD44v6 CAR-T cells as well.

In all, these two studies showed that in addition of anti-IL-6 injection, anti-IL-1 antibody therapy could significantly reduce complications of CAR-T cell immunotherapy in humans. They provided a strong evidence to suggest that monocytes were primarily responsible for CRS and neurotoxicity complications of CAR-T cell therapy. 

However, it is not clear why are monocytes getting activated after CAR-T cell infusion in the first place. There is interaction, at certain level, between CAR-T cells and monocytes, but is it antigen-specific via anti-CD19 CAR-T or endogenous TCR, or is it non-specific, is unknown presently. 

posted by David Usharauli 

New mouse model to mimic immune response to Flu virus in elderly

This week Science published several research papers in immunology. One of these papers is the study from Yale School of Medicine showing that Tlr7^–/– × Mavs^–/– double KO mice expressing human anti-viral molecule Mx1 display susceptibility to influenza A virus in a caspase 1/11-dependent manner.

It is known that elderly individuals show susceptibility to influenza A virus. One reason for this susceptibility is a reduced ability of elderly immune system to produce type I IFNs in response to influenza A virus.

In contrast, aging mice do not show such susceptibility to influenza A virus. Mouse does not express Mx1, a dynamin-like guanosine triphosphatase that blocks primary transcription of influenza in humans. To make mouse model "usable" for  studying human response influenza A virus, the authors created Mx1⁺ mice. Mx1⁺ mice were resistant to experimental influenza A virus infection and this resistance was dependent on Tlr7 and Mavs (both molecules are involved in anti-viral response).

Interestingly, while Mx1⁺ mice double deficient for Tlr7^–/– × Mavs^–/– were susceptible to influenza A virus infection,  Mx1⁺ mice triple deficient for Tlr7^–/– × Mavs^–/–× Casp1/11^–/– were resistant to influenza A virus infection even though both showed similar viral burden (Of note, Tlr7^–/– × Mavs^–/–× Casp1/11^–/– mice were eventually cleared the virus by 30 days after infection).

Moreover, Mx1⁺ mice triple deficient for Tlr7^–/– × Mavs^–/–× Casp1/11^–/– were resistant to influenza A virus infection even though they too showed similar secondary bacterial "bloom" in their airways.

These data suggested that caspase 1/11 signaling reduced "tissue tolerance" [rather than increased anti-viral response] to influenza A virus infection and secondary bacterial "bloom" in Mx1⁺ mice deficient for anti-viral innate signaling via Tlr7^–/– × Mavs^–/–.

In summary, this study showed that intact caspase 1/11 signaling compromises tissue tolerance to acute influenza A virus infection and secondary bacterial "bloom" in host with a weakened anti-viral signaling.

David Usharauli

Specialized monocyte subset protects lung vasculature from tumor metastases

Immune system's cellular lineage diversity is expanding and quite rapidly. Now we have several types of T cells, several types of NK cells, whole new family of innate lymphoid cells (ILCs), cytokine-secreting B cells, specialized subset of neutrophils using sticky DNA nets to trap the microbes. 

Here comes a paper in Science showing a new subset of orphan nuclear receptor Nur77-positive monocytes specializing in protection of lung vasculature from tumor metastases.

First, using reporter mice that tracks Nur77⁺monocytes (encoded by Nr4a1, a Nr4a1-GFP green mice), the authors showed that after intravenous injection of red fluorescent Lewis Lung Carcinoma cells (LLC-RFP), Nur77⁺monocytes are rapidly accumulating in the lung vasculature, then slowing down their speed in close proximity to tumor cells.

To confirm that Nur77⁺monocytes play a role in tumor defense, the authors first repeated the same experiment with total Nr4a1-KO mice, which according to the authors, selectively lack Nur77⁺monocytes. Nra1-KO mice appeared to harbor more lung metastasis (but not in liver), even if tumor cells [B16 melanoma] were injected subQ.

Similarly, increased tumor metastases (but not primary tumors) were seen with mouse model of spontaneous mammary tumor, MMTV-PyMT, receiving Nr4a1-KO bone marrow transfer, supporting the role of Nur77⁺ hematopoietic cells in prevention of tumor metastases.

To more specifically address the role of Nur77⁺monocytes in tumor protection, the authors have used two different myeloid-specific Nr4a1 conditional knockout models, CSF1R-Cre⁺ Nr4a1^fl/fl and LysM-Cre⁺ Nr4a1^fl/fl. In both models, there was an increase in tumor metastases to the lung. Interestingly, T cell-specific Nur77 deletion showed no effect on tumor metastases.

Finally, the authors found that receptor CX3CR1 could play an important role in tumor recognition by Nur77⁺monocytes.

In summary, this study suggest that there is a specialized monocytes lineage defined by Nr4a1 expression that specifically patrol and protects the lung vasculature against tumor metastases.

David Usharauli