Abstract
Antibody drug conjugate (ADC) showed potent therapeutic efficacy in several types of cancers. The role of autophagy in antitumor effects of ADC remains unclear. In this study, the ADC, Rituximab-monomethyl auristatin E (MMAE) with a Valine-Citrulline cleavable linker, was designed to investigate its therapeutic efficacy against non-Hodgkin lymphoma (NHL) as well as the underlying mechanisms. Methylthiazolyldiphenyl-tetrazolium bromide (MTT) was used to detect growth inhibition in B-cell lymphoma cell lines, Ramos and Daudi cells, which were treated by Rituximab-MMAE alone or combined with autophagy conditioner. Apoptosis was detected by flow cytometry and immunohistochemistry, and apoptosis inhibitor was employed to discover the relationship between autophagy and apoptosis during the Rituximab-MMAE treatment. Autophagy was determined by three standard techniques which included confocal microscope, transmission electron microscope, and western blots. Ramos xenograft tumors in BALB/c nude mice were established to investigate the antitumor effect of combination use of Rituximab-MMAE and autophagy conditioner in B-NHL therapy. Our results showed that Rituximab-MMAE elicited caspase-3-dependent apoptosis in NHL cells and exhibited potent therapeutic efficacy in vivo. Autophagy, which was characterized by upregulated light chain 3-II expression, and accumulation of autophagosomes, was triggered during the Rituximab-MMAE treatment. Meanwhile, inactivation of Akt/mTOR pathway was shown to be involved in the autophagy triggered by Rituximab-MMAE, indicating a probable mechanism of the ADC-initiated autophagy. Importantly, inhibition of autophagy by chloroquine suppressed the Rituximab-MMAE-induced apoptosis, while activating autophagy by rapamycin significantly enhanced the therapeutic effect of Rituximab-MMAE both in vitro and in vivo. Our data elucidated the critical relationship between autophagy and apoptosis in Rituximab-MMAE-based therapy and highlighted the potential approach for NHL therapy by combined administration of the ADC and autophagy activator.
Highlights
Non-Hodgkin lymphoma (NHL) is one of the most common types of lymphomas which accounts for about 90% of all cases [1]
Mice were sacrificed on day 22 and the tumor weight in mice treated with Rituximab-MMAE, and the vehicle controls were 280.00 ± 144.80 and 1,977.14 ± 214.22 mg, respectively
Our data showed that Rituximab-MMAE did not induce significant toxicity in the organs, indicating Rituximab-MMAE is safe for animal studies (Figure S5 in Supplementary Material)
Summary
Non-Hodgkin lymphoma (NHL) is one of the most common types of lymphomas which accounts for about 90% of all cases [1]. Novel therapeutic approaches for NHL are urgently needed. As one of the most promising target therapy strategies, antibody drug conjugate (ADC) was characterized by monoclonal antibody conjugated with small molecule drugs, which has shown significant therapeutic efficacy in the treatment of hematological malignancies and solid tumors [5]. By covalent conjugated to monoclonal antibody, highly potent cytotoxic agents could be delivered to tumors which minimize toxicity to healthy tissue [6]. Despite the concept of ADC is straightforward, the early failures in development of ADC due to high systemic toxicity and therapy resistance suggested more unknown mechanism need to be dug out [7, 8]. Antibody drug conjugate (ADC) showed potent therapeutic efficacy in several types of cancers. The role of autophagy in antitumor effects of ADC remains unclear
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