Purpose: Evaluate the efficacy of an SN-38-anti-Trop-2 antibody-drug conjugate (ADC) against several human solid tumor types, and to assess its tolerability in mice and monkeys, the latter with tissue crossreactivity to hRS7 similar to humans.Experimental Design: Two SN-38 derivatives, CL2-SN-38 and CL2A-SN-38, were conjugated to the anti-Trop-2-humanized antibody, hRS7. The immunoconjugates were characterized in vitro for stability, binding, and cytotoxicity. Efficacy was tested in five different human solid tumor-xenograft models that expressed Trop-2 antigen. Toxicity was assessed in mice and in Cynomolgus monkeys.Results: The hRS7 conjugates of the two SN-38 derivatives were equivalent in drug substitution ($6), cell binding (K d $ 1.2 nmol/L), cytotoxicity (IC 50 $ 2.2 nmol/L), and serum stability in vitro (t/ 1/2 $ 20 hours). Exposure of cells to the ADC demonstrated signaling pathways leading to PARP cleavage, but differences versus free SN-38 in p53 and p21 upregulation were noted. Significant antitumor effects were produced by hRS7-SN-38 at nontoxic doses in mice bearing Calu-3 (P 0.05), Capan-1 (P < 0.018), BxPC-3 (P < 0.005), and COLO 205 tumors (P < 0.033) when compared to nontargeting control ADCs. Mice tolerated a dose of 2 Â 12 mg/kg (SN-38 equivalents) with only short-lived elevations in ALT and AST liver enzyme levels. Cynomolgus monkeys infused with 2 Â 0.96 mg/kg exhibited only transient decreases in blood counts, although, importantly, the values did not fall below normal ranges.Conclusions: The anti-Trop-2 hRS7-CL2A-SN-38 ADC provides significant and specific antitumor effects against a range of human solid tumor types. It is well tolerated in monkeys, with tissue Trop-2 expression similar to humans, at clinically relevant doses, and warrants clinical investigation. Clin Cancer Res; 17(10); 3157-69. Ó2011 AACR.

Sacituzumab govitecan (IMMU-132) is an antibody-drug conjugate (ADC) made from a humanized anti-Trop-2 monoclonal antibody (hRS7) conjugated with the active metabolite of irinotecan, SN-38. In addition to its further characterization, as the clinical utility of IMMU-132 expands to an ever-widening range of Trop-2-expressing solid tumor types, its efficacy in new disease models needs to be explored in a nonclinical setting. Unlike most ADCs that use ultratoxic drugs and stable linkers, IMMU-132 uses a moderately toxic drug with a moderately stable carbonate bond between SN-38 and the linker. Flow cytometry and immunohistochemistry disclosed that Trop-2 is expressed in a wide range of tumor types, including gastric, pancreatic, triple-negative breast (TNBC), colonic, prostate, and lung. While cell-binding experiments reveal no significant differences between IMMU-132 and parental hRS7 antibody, surface plasmon resonance analysis using a Trop-2 CM5 chip shows a significant binding advantage for IMMU-132 over hRS7. The conjugate retained binding to the neonatal receptor, but it lost greater than 60% of the antibody-dependent cell-mediated cytotoxicity activity compared to that of hRS7. Exposure of tumor cells to either free SN-38 or IMMU-132 demonstrated the same signaling pathways, with pJNK1/2 and p21(WAF1/Cip1) upregulation followed by cleavage of caspases 9, 7, and 3, ultimately leading to poly-ADP-ribose polymerase cleavage and double-stranded DNA breaks. Pharmacokinetics of the intact ADC in mice reveals a mean residence time (MRT) of 15.4 h, while the carrier hRS7 antibody cleared at a similar rate as that of the unconjugated antibody (MRT ∼ 300 h). IMMU-132 treatment of mice bearing human gastric cancer xenografts (17.5 mg/kg; twice weekly × 4 weeks) resulted in significant antitumor effects compared to that of mice treated with a nonspecific control. Clinically relevant dosing schemes of IMMU-132 administered either every other week, weekly, or twice weekly in mice bearing human pancreatic or gastric cancer xenografts demonstrate similar, significant antitumor effects in both models. Current Phase I/II clinical trials ( ClinicalTrials.gov , NCT01631552) confirm anticancer activity of IMMU-132 in cancers expressing Trop-2, including gastric and pancreatic cancer patients.

The antibody-drug conjugate (ADC), IMMU-130, of the moderately cytotoxic topoisomerase I inhibitor, SN-38, and the CEACAM5-targeted humanized antibody (mAb), labetuzumab, was evaluated in model systems of human colon carcinoma and in phase I clinical trials of heavily pretreated patients with metastatic colorectal cancer. The conjugate, designed with a near-homogeneous drug substitution of 7-8 SN-38/mAb and with a linker that released 50% of the drug in ∼20 h, showed significant antitumor effects compared to a nontargeted ADC in human tumor xenografts, which could be augmented in combination with bevacizumab. The advantage of fractionated dosing was demonstrated, with potential implications for the clinical dosing schedule. Biodistribution comparing IMMU-130 with labetuzumab showed that the conjugate cleared somewhat faster from the blood, but this did not affect tumor uptake and retention. The use of an ultrastable linker in the conjugate design abrogated antitumor effects. A tolerability study in rabbits showed a high safety margin, with no-observed-adverse-effect level (NOAEL) corresponding to a cumulative human-equivalent protein dose of 40-60 mg/kg. The preclinical findings appear to be corroborated in two phase I clinical trials, with high tolerability and evidence of antitumor activity, including objective responses. The impact of the ADC design on the utility of IMMU-130, tailored to a poorly internalizing target, is discussed.

Summary Chemotherapy plus rituximab has been the mainstay of treatment for follicular lymphoma ( FL ) for two decades but is associated with immunosuppression and relapse. In phase 2 studies, lenalidomide combined with rituximab (R 2 ) has shown clinical synergy in front‐line and relapsed/refractory FL . Here, we show that lenalidomide reactivated dysfunctional T and Natural Killer ( NK ) cells ex vivo from FL patients by enhancing proliferative capacity and T‐helper cell type 1 (Th1) cytokine release. In combination with rituximab, lenalidomide improved antibody‐dependent cellular cytotoxicity in sensitive and chemo‐resistant FL cells, via a cereblon‐dependent mechanism. While single‐agent lenalidomide and rituximab increased formation of lytic NK cell immunological synapses with primary FL tumour cells, the combination was superior and correlated with enhanced cytotoxicity. Immunophenotyping of FL patient samples from a phase 3 trial revealed that R 2 treatment increased circulating T‐ and NK ‐cell counts, while R‐chemotherapy was associated with reduced cell numbers. Finally, using an in vitro model of myeloid differentiation, we demonstrated that lenalidomide caused a reversible arrest in neutrophil maturation that was distinct from a cytotoxic chemotherapeutic agent, which may help explain the lower rates of neutropenia observed with R 2 versus R‐chemotherapy. Taken together, we believe these data support a paradigm shift in the treatment of FL – moving from combination immunochemotherapy to chemotherapy‐free immunotherapy.