Lactaptin induces p53-independent cell death associated with features of apoptosis and autophagy and delays growth of breast cancer cells in mouse xenografts

PLoS One. 2014 Apr 7;9(4):e93921. doi: 10.1371/journal.pone.0093921. eCollection 2014.

Abstract

Lactaptin, the proteolytic fragment of human milk kappa-casein, induces the death of various cultured cancer cells. The mechanisms leading to cell death after lactaptin treatment have not been well characterized. In this study the in vivo and in vitro effects of a recombinant analogue of lactaptin (RL2) were examined. Following treatment with the recombinant analogue of lactaptin strong caspase -3, -7 activation was detected. As a consequence of caspase activation we observed the appearance of a sub-G1 population of cells with subdiploid DNA content. Dynamic changes in the mRNA and protein levels of apoptosis-related genes were estimated. No statistically reliable differences in p53 mRNA level or protein level were found between control and RL2-treated cells. We observed that RL2 constitutively suppressed bcl-2 mRNA expression and down regulated Bcl-2 protein expression in MDA-MB-231 cells. We demonstrated that RL2 penetrates cancer and non-transformed cells. Identification of the cellular targets of the lactaptin analogue revealed that α/β-tubulin and α-actinin-1 were RL2-bound proteins. As the alteration in cellular viability in response to protein stimulus can be realized not only by way of apoptosis but also by autophagy, we examined the implications of autophagy in RL2-dependent cell death. We also found that RL2 treatment induces LC3-processing, which is a hallmark of autophagy. The autophagy inhibitor chloroquine enhanced RL2 cytotoxicity to MDA-MB-231 cells, indicating the pro-survival effect of RL2-dependent autophagy. The antitumour potential of RL2 was investigated in vivo in mouse xenografts bearing MDA-MB-231 cells. We demonstrated that the recombinant analogue of lactaptin significantly suppressed the growth of solid tumours. Our results indicate that lactaptin could be a new molecule for the development of anticancer drugs.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Antineoplastic Agents / pharmacology*
  • Apoptosis / drug effects*
  • Autophagy / drug effects*
  • Breast Neoplasms / metabolism
  • Caseins / pharmacology*
  • Caspases / metabolism*
  • Cell Death / drug effects
  • Cell Line, Tumor
  • Cell Survival / drug effects
  • Chloroquine / pharmacology
  • DNA Fragmentation / drug effects
  • Heterografts
  • Humans
  • Mice
  • Proto-Oncogene Proteins c-bcl-2 / metabolism

Substances

  • Antineoplastic Agents
  • Caseins
  • Proto-Oncogene Proteins c-bcl-2
  • lactaptin, human
  • Chloroquine
  • Caspases

Grant support

The study was supported by the Siberian Branch of the Russian Academy of Sciences under the Program “Science to Medicine” (Project no. 13) and Integration Project (no. 94), by grants from the Russian Foundation for Basic Research (Grant nos. 13-04-01313a and 13-04-011457a) and by a grant from Russian Federal Target Programs (Grant no. 16.N08.12.1009). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.