Downregulation of LRP/LR with siRNA inhibits several cancer hallmarks in lung cancer cells

Abstract

The incidence and mortality rates of cancer are growing rapidly worldwide, with lung cancer being the most commonly occurring cancer in males. Human carcinomas circumvent the inhibitory pathways induced by DNA damage and senescence through the upregulation of telomerase activity. The 37 kDa/67 kDa laminin receptor (LRP/LR) is a cell surface receptor which plays a role in several cancer hallmarks, including metastasis, angiogenesis, cell viability maintenance, apoptotic evasion, and mediating telomerase activity. We have previously shown that the knockdown of LRP/LR with an LRP‐specific siRNA significantly impedes adhesion and invasion, induces apoptosis, and inhibits telomerase activity in various cancer cell lines in vitro. Here, we investigated the effect of downregulating LRP/LR with LRP‐specific siRNA in A549 lung cancer cells. Downregulation of LRP/LR resulted in a significant decrease in cell viability, migration potential, and telomerase activity, as well as a significant increase in apoptosis. Proteomic analysis further suggested the re‐establishment of immune control over the lung cancer cells, a previously unidentified facet of LRP downregulation in cancer. Altogether, we suggest that targeting LRP/LR for downregulation may have therapeutic potential for inhibiting several cancer hallmarks.

PLGA nanocapsules as a delivery system for a recombinant LRP‐based therapeutic

“…It has been shown that once LRP/LR has bound to laminin, LRP/LR can then interact with focal adhesion kinase and this interaction then activates the PI3‐kinase/AKT and MEK/ERK1/2 pathways, which are directly related to proliferation and longevity [ 31 ]. It has further been established that LRP/LR plays a key role in mediating telomerase activity [ 4 , 6 , 7 , 8 ], whereby LRP/LR increases telomerase activity to promote cell viability and prevent cellular senescence, as shown by the reduction in gamma H2AX (which marks sites of DNA damage and DNA breaks) and the senescence biomarker β‐galactosidase [ 6 , 9 ].…”

“…In a 96-well plate, the TRAP reaction mixture and 2 lL of the standardised protein samples solution, for a total of 1 lg, were loaded into each well to a final volume of 12.5 lL. This reaction Mastermix consisted of LunaÒ Universal qPCR Mastermix (New England Biolabs, Ipswich, MA, USA), TS and ACX primers (as described by Bignoux et al [8]), EGTA and PCR-grade water. The CFX96 TM Touch qPCR System (Bio-Rad) was employed to perform qPCR analysis on all samples, using the following cycling parameters: One cycle of 30 min at 37 °C (telomere extension step), and 2 min at 95 °C, followed by 45 cycles of amplification with denaturation at 95 °C for 15 s, annealing at 59 °C for 60 s, and extension at 45 °C for 10 s. Minus telomerase (consisting only of CHAPS lysis buffer), no template (PCR-grade water) and heat-treated (incubated at 90 °C for 10 min to inactivate any telomerase activity) negative controls were included.…”

“…It has further been established that LRP/LR plays a key role in mediating telomerase activity [ 4 , 6 , 7 , 8 ], whereby LRP/LR increases telomerase activity to promote cell viability and prevent cellular senescence. Further evidence shows that the overexpression of LRP::FLAG increases the telomerase catalytic subunit (hTERT) protein levels as well as telomerase activity, and subsequently telomere length within MRC5 and HEK‐293 cell lines.…”

Emerging roles of the cellular prion protein (PrPC) and 37/67 kDa laminin receptor (RPSA) interaction in cancer biology

“…Flow cytometric analysis revealed that melanoma cells displayed high levels of RPSA on their cell surface and the treatment with IgG1–iS18 antibody determined a significant reduction of the adhesive potential to LM and the invasive capacity of malignant melanoma cells, confirming previous results reported in several types of cancer cells [ 75 – 78 ]. For instance, in A549 lung cancer cell line, downregulation of RPSA with specific siRNA, inhibited cellular migration activity, as evaluated by scratch motility assay [ 79 ]. From a mechanistic point of view, proteomic analysis performed in pancreatic cancer cells, revealed that RPSA interacted with Integrin alpha 6 (ITGA6) and that they colocalized on the plasma membrane as showed by immunofluorescence analysis.…”

“…The authors postulated that RPSA upregulation may enhance telomerase activity [ 122 , 123 ]. Reversely, siRNA-mediated RPSA knock-down impeded telomerase activity in breast and lung cancer cells [ 79 , 121 ]. Likewise, a significant decrease in telomerase activity and hTERT expression levels was observed in late-stage CRC cells via RPSA siRNA technology, resulting in genetic instability and activation of DDR pathway [ 54 , 124 ].…”