The La antigen is over-expressed in lung cancer and is a selective dead cancer cell target for radioimmunotherapy using the La-specific antibody APOMAB®

Abstract

BackgroundThe lupus-associated (La)-specific murine monoclonal antibody DAB4 (APOMAB®) specifically binds dead cancer cells. Using DAB4, we examined La expression in human lung cancer samples to assess its suitability as a cancer-selective therapeutic target. We evaluated the safety and effectiveness of radioimmunotherapy (RIT) using DAB4 radiolabeled with Lutetium-177 (177Lu) in the murine Lewis Lung (LL2) carcinoma model, and determined whether combining RIT with DNA-damaging cisplatin-based chemotherapy, a PARP inhibitor (PARPi), or both alters treatment responses.MethodsThe expression of La mRNA in human lung cancer samples was analysed using the online database Oncomine, and the protein expression of La was examined using a TissueFocus Cancer Survey Tissue Microarray. The binding of DAB4 to cisplatin-treated LL2 cells was assessed in vitro. LL2 tumour-bearing mice were administered escalating doses of 177Lu-DAB4 alone or in combination with chemotherapy, and tumour growth and survival measured. Biodistribution analysis was used to determine tissue uptake of 177Lu-DAB4 or its isotype control (177Lu-Sal5), when delivered alone or after chemotherapy. PARPi (rucaparib; AG-014699) was combined with chemotherapy and the effects of combined treatment on tumour growth, tumour cell DNA damage and death, and intratumoural DAB4 binding were also analysed. The effect of the triple combination of PARPi, chemotherapy and 177Lu-DAB4 on tumour growth and survival of LL2 tumour-bearing mice was tested.ResultsLa was over-expressed at both mRNA and protein levels in surgical specimens of human lung cancer and the over-expression of La mRNA conferred a poorer prognosis. DAB4 bound specifically to cisplatin-induced dead LL2 cells in vitro. An anti-tumour dose response was observed when escalating doses of 177Lu-DAB4 were delivered in vivo, with supra-additive responses observed when chemotherapy was combined with 177Lu-DAB4. Combining PARPi with chemotherapy was more effective than chemotherapy alone with increased tumour cell DNA damage and death, and intratumoural DAB4 binding. The combination of PARPi, chemotherapy and 177Lu-DAB4 was well-tolerated and maximised tumour growth delay.ConclusionsThe La antigen represents a dead cancer cell-specific target in lung cancer, and DAB4 specifically targeted tumour tissue in vivo, particularly after chemotherapy. Tumour uptake of DAB4 increased further after the combination of PARPi and chemotherapy, which generated new dead tumour cell-binding targets. Consequently, combining 177Lu-DAB4 with PARPi and chemotherapy produced the greatest anti-tumour response. Therefore, the triple combination of PARPi, chemotherapy and RIT may have broad clinical utility.

APOMAB Antibody–Drug Conjugates Targeting Dead Tumor Cells are Effective In Vivo

“…We have previously shown that DAB4 binds within the necrotic and apoptotic areas of solid tumors including LL2 tumors (7,13,17). We also saw a similar tumor uptake pattern of chDAB4 in Supplementary Fig.…”

“…3B9 was the gift of Prof. Tom Gordon, SA Pathology, Flinders Medical Centre, Adelaide, Australia. DAB4 and the isotype-specific control mAb (Sal5, targeting the irrelevant Salmonella antigen) were produced and purified as previously described (7). All cell lines were used for fewer than 3 months after receipt or resuscitation from cryopreservation, and no further cell authentication was performed.…”

“…As La/SSB is highly conserved between mice and humans, DAB4 binds both mouse and human forms of La/SSB. By radiolabeling DAB4 with aor b-emitting radionuclides, we have been able to specifically target syngraft and xenograft tumors in mice, particularly after prior treatment with chemotherapy, effectively irradiating the tumor from the inside out (7,13,17). These studies demonstrate that radiation cross-fire dose, the dose delivered to cells surrounding the antibody-bound target cells, can have potent antitumor effects.…”

“…APOMAB Ò (DAB4) is a mouse mAb, which targets the Lupusassociated (La)/Sj€ ogren Syndrome-B (SSB) antigen, which is overexpressed in a variety of different cancers (7)(8)(9)(10)(11). La/SSB only becomes available for antibody binding in cells that have lost membrane integrity, particularly in apoptotic and necrotic cancer cells, making DAB4 a dead tumor cell-targeting mAb (7,(12)(13)(14)(15)(16)(17). As La/SSB is highly conserved between mice and humans, DAB4 binds both mouse and human forms of La/SSB.…”

“…Nevertheless, characteristics of the in vitro binding of the DAB4 mAb to dead tumor cells compared with a TNT mAb help to explain tumor accumulation of DAB4. In vitro, DAB4 preferentially binds to dead tumor cells, which die because of DNA-damaging antitumor treatment; this binding is saturable, rapid, high-avidity, irreversible, and involves protein-protein crosslinking of DAB4 with its target antigen, La/SSB, during tumor cell death (7,(12)(13)(14)(15)(16). TNT3 has been used preclinically as an effective vehicle for the delivery of immunostimulatory CpG to the tumor site in mouse xenograft tumor models (21) and as a fusion protein for tumor delivery of IFNg (22).…”