CAR T Cells Targeting B7-H3, a Pan-Cancer Antigen, Demonstrate Potent Preclinical Activity Against Pediatric Solid Tumors and Brain Tumors.

Patients with relapsed pediatric solid tumors and CNS malignancies have few therapeutic options and frequently die of their disease. Chimeric antigen receptor (CAR) T cells have shown tremendous success in treating relapsed pediatric acute lymphoblastic leukemia, but this has not yet translated to treating solid tumors. This is partially due to a paucity of differentially expressed cell surface molecules on solid tumors that can be safely targeted. Here, we present B7-H3 (CD276) as a putative target for CAR T-cell therapy of pediatric solid tumors, including those arising in the central nervous system. We developed a novel B7-H3 CAR whose binder is derived from a mAb that has been shown to preferentially bind tumor tissues and has been safely used in humans in early-phase clinical trials. We tested B7-H3 CAR T cells in a variety of pediatric cancer models. B7-H3 CAR T cells mediate significant antitumor activity in vivo, causing regression of established solid tumors in xenograft models including osteosarcoma, medulloblastoma, and Ewing sarcoma. We demonstrate that B7-H3 CAR T-cell efficacy is largely dependent upon high surface target antigen density on tumor tissues and that activity is greatly diminished against target cells that express low levels of antigen, thus providing a possible therapeutic window despite low-level normal tissue expression of B7-H3. B7-H3 CAR T cells could represent an exciting therapeutic option for patients with certain lethal relapsed or refractory pediatric malignancies, and should be tested in carefully designed clinical trials.

Majzner RG ，Theruvath JL ，Nellan A ，Heitzeneder S ，Cui Y ，Mount CW ，Rietberg SP ，Linde MH ，Xu P ，Rota C ，Sotillo E ，Labanieh L ，Lee DW ，Orentas RJ ，Dimitrov DS ，Zhu Z ，Croix BS ，Delaidelli A ，Sekunova A ，Bonvini E ，Mitra SS ，Quezado MM ，Majeti R ，Monje M ，Sorensen PHB ，Maris JM ，Mackall CL ... -  《-》

B7-H3-redirected chimeric antigen receptor T cells target glioblastoma and neurospheres.

The dismal survival of glioblastoma (GBM) patients urgently calls for the development of new treatments. Chimeric antigen receptor T (CAR-T) cells are an attractive strategy, but preclinical and clinical studies in GBM have shown that heterogeneous expression of the antigens targeted so far causes tumor escape, highlighting the need for the identification of new targets. We explored if B7-H3 is a valuable target for CAR-T cells in GBM. We compared mRNA expression of antigens in GBM using TCGA data, and validated B7-H3 expression by immunohistochemistry. We then tested the antitumor activity of B7-H3-redirected CAR-T cells against GBM cell lines and patient-derived GBM neurospheres in vitro and in xenograft murine models. B7-H3 mRNA and protein are overexpressed in GBM relative to normal brain in all GBM subtypes. Of the 46 specimens analyzed by immunohistochemistry, 76% showed high B7-H3 expression, 22% had detectable, but low B7-H3 expression and 2% were negative, as was normal brain. All 20 patient-derived neurospheres showed ubiquitous B7-H3 expression. B7-H3-redirected CAR-T cells effectively targeted GBM cell lines and neurospheres in vitro and in vivo. No significant differences were found between CD28 and 4-1BB co-stimulation, although CD28-co-stimulated CAR-T cells released more inflammatory cytokines. We demonstrated that B7-H3 is highly expressed in GBM specimens and neurospheres that contain putative cancer stem cells, and that B7-H3-redirected CAR-T cells can effectively control tumor growth. Therefore, B7-H3 represents a promising target in GBM. FUND: Alex's Lemonade Stand Foundation; Il Fondo di Gio Onlus; National Cancer Institute; Burroughs Wellcome Fund.

Nehama D ，Di Ianni N ，Musio S ，Du H ，Patané M ，Pollo B ，Finocchiaro G ，Park JJH ，Dunn DE ，Edwards DS ，Damrauer JS ，Hudson H ，Floyd SR ，Ferrone S ，Savoldo B ，Pellegatta S ，Dotti G ... -  《EBioMedicine》

Directing B7-H3 chimeric antigen receptor T cell homing through IL-8 induces potent antitumor activity against pediatric sarcoma.

Advances in pediatric oncology have occurred for some cancers; however, new therapies for sarcoma have been inadequate. Cellular immunotherapy using chimeric antigen receptor (CAR) T cells has shown dramatic benefits in leukemia, lymphoma, and multiple myeloma but has been far less successful in pediatric solid tumors such as rhabdomyosarcoma (RMS) and osteosarcoma (OS). Balancing issues of "on-target, off-tumor toxicity", investigators have identified B7-H3 as a broadly expressed tumor antigen with otherwise restricted expression on normal tissues. We hypothesized that rapid homing via a chemokine receptor and CAR engagement through B7-H3 would enhance CAR T cell efficacy in solid tumors. We generated B7-H3 CAR T cells that also express the Interleukin-8 (IL-8) receptor, CXCR2. Cytokine production, flow cytometry, Seahorse assays and RNA sequencing were used to compare the B7-H3 CXCR2 (BC2) CAR T cells with B7-H3 CAR T cells. We developed an IL-8 overexpressing human RMS mouse model to test homing and cytotoxicity in vivo. We demonstrate that IL-8 is expressed by RMS and OS and expression significantly increases after radiation. Overexpression of an IL-8 receptor, CXCR2, on B7-H3 CAR T cells enhances homing into IL-8 expressing tumors, augments T cell metabolism and leads to significant tumor regression. These findings warrant further investigation into the use of BC2 CAR T cells as a treatment for patients with RMS, OS and other B7-H3-expressing, IL-8 producing solid tumors.

Lake JA ，Woods E ，Hoffmeyer E ，Schaller KL ，Cruz-Cruz J ，Fernandez J ，Tufa D ，Kooiman B ，Hall SC ，Jones D ，Hayashi M ，Verneris MR ... -  《Journal for ImmunoTherapy of Cancer》

Locoregionally administered B7-H3-targeted CAR T cells for treatment of atypical teratoid/rhabdoid tumors.

Theruvath J ，Sotillo E ，Mount CW ，Graef CM ，Delaidelli A ，Heitzeneder S ，Labanieh L ，Dhingra S ，Leruste A ，Majzner RG ，Xu P ，Mueller S ，Yecies DW ，Finetti MA ，Williamson D ，Johann PD ，Kool M ，Pfister S ，Hasselblatt M ，Frühwald MC ，Delattre O ，Surdez D ，Bourdeaut F ，Puget S ，Zaidi S ，Mitra SS ，Cheshier S ，Sorensen PH ，Monje M ，Mackall CL ... -  《-》

Tandem CAR-T cells targeting CD70 and B7-H3 exhibit potent preclinical activity against multiple solid tumors.

Purpose: Given that heterogeneous expression and variants of antigens on solid tumors are responsible for relapse after chimeric antigen receptor (CAR)-T cell therapy, we hypothesized that combinatorial targeting two tumor-associated antigens would lessen this problem and enhance the antitumor activity of T cells. Methods: The co-expression level of CD70 and B7-H3 was analyzed in multiple tumor tissue samples. Further, two putative antigens were identified in The Cancer Genome Atlas and Gene Expression Profiling Interactive Analysis database. Two CD70 targeted CARs with different antigen binding domain, truncated CD27 and CD70 specific single-chain antibody fragment (scFv), were designed to screen a more suitable target-antigen binding moiety. Accordingly, we designed a bivalent tandem CAR (TanCAR) and further assessed the anti-tumor efficacy of TanCAR-T cells in vitro and in vivo. Results: Our results indicated that co-expression of CD70 and B7-H3 was observed on multiple tumor types including kidney, breast, esophageal, liver, colon cancer, glioma as well as melanoma. The CD70 targeted CAR-T cells with binding moiety of CD70 specific scFv exhibit a higher affinity and antitumor effect against CD70+ tumor cells. TanCAR-T cells induced enhanced ability of cytolysis and cytokine release over unispecific CAR-T cells when encountering tumor cells expressing two target-antigens. Further, low doses of TanCAR-T cells could also effectively control the lung cancer and melanoma xenografts and improved overall survival of the treated animals. Conclusion: TanCAR-T cells targeting CD70 and B7-H3 exhibit enhanced antitumor functionality and improve the problem of antigenic heterogeneity and variant in the treatment against solid tumor and melanoma.

Yang M ，Tang X ，Zhang Z ，Gu L ，Wei H ，Zhao S ，Zhong K ，Mu M ，Huang C ，Jiang C ，Xu J ，Guo G ，Zhou L ，Tong A ... -  《Theranostics》