Hypoxia Selectively Impairs CAR-T Cells In Vitro

Berahovich, Robert; Liu, Xianghong; Zhou, Hua; Tsadik, Elias; Xu, Shuning; Golubovskaya, Vita M.; Wu, Lijun

doi:10.3390/cancers11050602

Cited by 63 publications

(55 citation statements)

References 39 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…These data support our hypothesis that the TSC population can be targeted using a DCLK1-specific CAR-T. Here, we report that, in collaboration with ProMab Inc., we have developed a novel CAR-T based on the DCLK1 ScFv containing a CD28 transmembrane and co-stimulatory domain and CD3ζ activation domain [39][40][41][42][43][44][45][46]. CAR-T cells generated using DCLK1 ScFv (CBT-511) demonstrated~20% CAR expression and significantly induced CRC cell cytotoxicity.…”

Section: Introductionsupporting

confidence: 73%

DCLK1 Monoclonal Antibody-Based CAR-T Cells as a Novel Treatment Strategy against Human Colorectal Cancers

Sureban

Berahovich

Zhou

et al. 2019

Cancers

Self Cite

View full text Add to dashboard Cite

CAR-T (chimeric antigen receptor T cells) immunotherapy is effective in many hematological cancers; however, efficacy in solid tumors is disappointing. Doublecortin-like kinase 1 (DCLK1) labels tumor stem cells (TSCs) in genetic mouse models of colorectal cancer (CRC). Here, we describe a novel CAR-T targeting DCLK1 (CBT-511; with our proprietary DCLK1 single-chain antibody variable fragment) as a treatment strategy to eradicate CRC TSCs. The cell surface expression of DCLK1 and cytotoxicity of CBT-511 were assessed in CRC cells (HT29, HCT116, and LoVo). LoVo-derived tumor xenografts in NOD Scid gamma (NSG™) mice were treated with CBT-511 or mock CAR-T cells. Adherent CRC cells express surface DCLK1 (two-dimensional, 2D). A 4.5-fold increase in surface DCLK1 was observed when HT29 cells were grown as spheroids (three-dimensional, 3D). CBT-511 induced cytotoxicity (2D; p < 0.0001), and increased Interferon gamma (IFN-γ) release in CRC cells (2D) compared to mock CAR-T (p < 0.0001). Moreover, an even greater increase in IFN-γ release was observed when cells were grown in 3D. CBT-511 reduced tumor growth by approximately 50 percent compared to mock CAR-T. These data suggest that CRC cells with increased clonogenic capacity express increased surface DCLK1. A DCLK1-targeted CAR-T can induce cytotoxicity in vitro and inhibit xenograft growth in vivo.

show abstract

Section: Introductionsupporting

confidence: 73%

DCLK1 Monoclonal Antibody-Based CAR-T Cells as a Novel Treatment Strategy against Human Colorectal Cancers

Sureban

Berahovich

Zhou

et al. 2019

Cancers

Self Cite

View full text Add to dashboard Cite

show abstract

“…Another study found the same effect, but attributed the slow growth at physiological DO to a build‐up of cAMP, which in turn, resulted in reduced T‐cell activation due to lymphocyte‐specific protein tyrosine kinase (Lck) inhibition (Larbi, Zelba, Goldeck, & Pawelec, 2010). In contrast, a study by Berahovich et al (2019) found no differences in the expansion of T cells and CD19 CAR‐T cells when cultured at physiological and atmospheric oxygen conditions.…”

Section: Discussionmentioning

confidence: 78%

“…The phenotypical distribution in this study was shown to be highest in CD8 T CM cells when the DO was controlled at 25% and the pH was set to 7.4. Thus far, most studies show no effect of the DO on the differentiation of T cells and CD19 CAR‐T cells (Atkuri et al, 2007; Berahovich et al, 2019). However, it is important to note that the majority of the referenced studies were not done under pH controlled or agitated conditions.…”

Section: Discussionmentioning

confidence: 99%

Bioprocess considerations for T‐cell therapy: Investigating the impact of agitation, dissolved oxygen, and pH on T‐cell expansion and differentiation

Amini

Patel

Veraitch

et al. 2020

Biotech & Bioengineering

View full text Add to dashboard Cite

Adoptive T‐cell therapy (ACT) has emerged as a promising new way to treat systemic cancers such as acute lymphoblastic leukemia. However, the robustness and reproducibility of the manufacturing process remains a challenge. Here, a single‐use 24‐well microbioreactor (micro‐Matrix) was assessed for its use as a high‐throughput screening tool to investigate the effect and the interaction of different shaking speeds, dissolved oxygen (DO), and pH levels on the growth and differentiation of primary T cells in a perfusion‐mimic process. The full factorial design allowed for the generation of predictive models, which were used to find optimal culture conditions. Agitation was shown to play a fundamental role in the proliferation of T cells. A shaking speed of 200 rpm drastically improved the final viable cell concentration (VCC), while the viability was maintained above 90% throughout the cultivation. VCCs reached a maximum of 9.22 × 106 cells/ml. The distribution of CD8+ central memory T cells (TCM), was found to be largely unaffected by the shaking speed. A clear interaction between pH and DO (p < .001) was established for the cell growth and the optimal culture conditions were identified for a combination of 200 rpm, 25% DO, and pH of 7.4. The combination of microbioreactor technology and Design of Experiment methodology provides a powerful tool to rapidly gain an understanding of the design space of the T‐cell manufacturing process.

show abstract

“…34 Furthermore, antigen-specific CAR-T cells that were mobilised into hypoxic tumour areas were more proliferative than CAR-T cells in normoxic areas, when evaluated in two in vivo models. 34,35 Berahovich et al 36 also explored the effects of hypoxia on CAR-T cells targeted against B-cell maturation antigen (BCMA) and CD19. They observed that hypoxia diminished the expansion of both CAR-and non-transduced T cells.…”

Section: Car-t Cells and Hypoxiamentioning

confidence: 99%

Editor's Pick: Tumour-Associated Hypoxia: Can We Give Chimeric Antigen Receptor T Cells More Breathing Space?

Martinez

Kosti

Schurich

et al. 2020

EMJ

View full text Add to dashboard Cite

Immunotherapy using chimeric antigen receptor (CAR)-engineered T cells has encountered important limitations in the transition of their use from liquid to solid tumours. Success is dependent upon T-cell trafficking to, and functional persistence within, tumours that often present a metabolically and immunologically hostile microenvironment. Moreover, CAR targets that are tumour specific are extremely scarce. To address these issues, several strategies have been proposed to improve both tumour selectivity and safety. One approach involves the engineering of CAR-T cells that only deploy their effector function at tumour sites. Conceptually, a solution for this exploits the oxygen-limited nature of advanced tumour deposits through the engineering of CAR that are exclusively expressed or activated under conditions of profound hypoxia. T cells have a complex inter-relationship with oxygen, which also needs to be factored into the refinement of these technologies. Ideally, oxygen-sensing CAR should only function when oxygen tension is below 2%, as is commonly the case in solid tumours but rare in healthy tissue. Successful advancement of such technologies presents opportunities for solid tumour immunotherapy because it should broaden the target repertoire that may safely be exploited in this context.

show abstract

Hypoxia Selectively Impairs CAR-T Cells In Vitro

Cited by 63 publications

References 39 publications

DCLK1 Monoclonal Antibody-Based CAR-T Cells as a Novel Treatment Strategy against Human Colorectal Cancers

DCLK1 Monoclonal Antibody-Based CAR-T Cells as a Novel Treatment Strategy against Human Colorectal Cancers

Bioprocess considerations for T‐cell therapy: Investigating the impact of agitation, dissolved oxygen, and pH on T‐cell expansion and differentiation

Editor's Pick: Tumour-Associated Hypoxia: Can We Give Chimeric Antigen Receptor T Cells More Breathing Space?

Contact Info

Product

Resources

About