FLLL12 induces apoptosis in lung cancer cells through a p53/p73-independent but death receptor 5-dependent pathway
Authors: Dong M Shin
James R Fuchs
Abedul Haque
Mohammad A Rahman
Zhuo Georgia Chen
A R M Ruhul Amin
Fadlo R Khuri
Journal: Cancer Letters (2015)
Discipline: CLINICAL MEDICINE
Keywords: Apoptosis; Death receptor-5; FLLL12; Lung cancer; Protein tyrosine phosphatases (PTPs).
Affiliations: Department of Hematology and Medical Oncology, Winship Cancer Institute of Emory University, Atlanta, GA 30322, USA., Deaprtment of Medicinal Chemistry and Pharmacognosy, College of Pharmacy, Ohio State University, Columbus, OH, USA., Department of Hematology and Medical Oncology, Winship Cancer Institute of Emory University, Atlanta, GA 30322, USA. Electronic address: .

Abstract:  Unlike chemotherapy drugs, the safety of natural compounds such as curcumin has been well established. However, the potential use of curcumin in cancer has been compromised by its low bioavailability, limited tissue distribution and rapid biotransformation leading to low in vivo efficacy. To circumvent these problems, more potent and bioavailable analogs have been synthesized. In the current study, we investigated the mechanism of anti-tumor effect of one such analog, FLLL12, in lung cancers. IC50 values measured by sulforhodamine B (SRB) assay at 72 h and apoptosis assays (annexin V staining, cleavage of PARP and caspase-3) suggest that FLLL12 is 5-10-fold more potent than curcumin against a panel of premalignant and malignant lung cancer cell lines, depending on the cell line. Moreover, FLLL12 induced the expression of death receptor-5 (DR5). Ablation of the expression of the components of the extrinsic apoptotic pathway (DR5, caspase-8 and Bid) by siRNA significantly protected cells from FLLL12-induced apoptosis (p &lt; 0.05). Analysis of mRNA expression revealed that FLLL-12 had no significant effect on the expression of DR5 mRNA expression. Interestingly, inhibition of global phosphatase activity as well as protein tyrosine phosphatases (PTPs), but not of alkaline phosphatases, strongly inhibited DR5 expression and significantly inhibited apoptosis (p &lt; 0.05), suggesting the involvement of PTPs in the regulation of DR5 expression and apoptosis. We further showed that the apoptosis is independent of p53 and p73. Taken together, our results strongly suggest that FLLL12 induces apoptosis of lung cancer cell lines by posttranscriptional regulation of DR5 through activation of protein tyrosine phosphatase(s). 

Retracted: No

PubPeer comments:
 Some unexpectedly similar bands in Figure 6. Could the authors comment? *** Inverse images  *** Figure 6 showing caption  , 
 Thanks for the bring my attention of this matter and appreciate the reviewer s comment. I communicated with the corresponding author who moved to a different institution and he will directly handle this comment and please contact him directly. ARM Ruhul Amin email: amina@marshall.edu , 
 #2 Thank you for your reply and for contacting the corresponding author at his current email address. , 
 From the corresponding author Dr. Amin I am replying through Dr. Shin s link since I have not received any direct correspondence. The experiment for Fig. 6D and F was conducted as a single experiment with set of 6 samples, run on the same gel and developed (see the original figure attached). Lane 1 and 2 are common for both inhibitors. During making figures for publication, two different figures were made (6D with lanes 1-4 and 6F with lanes 1, 2, 5 and 6 cutting out lane 3 and 4). The article was published almost 6 years earlier. Neither me nor the postdoc exactly recall the reason for making two separate figures instead of one since it does not affect the conclusion except making it simpler for the readers.  Ruhul Amin , 
 #4 Thank you for providing the originals. Perhaps the image below will help illustrate the issue with 6D. The actin band doesnu2019t appear to line up, at least to me. And if Iu2019m understanding this correctly, then 6F should show a splice in order to show the non-adjacent lanes. ***  , 
 From Dr. Amin After looking carefully, I agree with you that there are issues with actin although all lanes are there. It appears to me that actin for 6F should be for 6D (first four lanes). The actin for 6D was made using the last four lanes without cutting out the middle two lanes. I am acknowledging that I was not as careful for the actin as for other proteins during looking figures made by the postdoc. At this point, to ensure integrity I am thinking to send a correction to the journal combining Fig. 6D and F as a single figure. If you have any suggestion, let us know. Ruhul Amin , 
 #6 Dr. Amin: Thank you for your reply. I agree that contacting the journal to discuss the authorsu2019 desire to make correction seems like a good next step. Best wishes. , 
 To add more clarity on the comment by Actinopolyspora Biskrensis in #5 I have the labelled bands in all the three gels with rectangles of specific colours. Please look at the analysis below. It seems that the bands labels in figures 6D and 6F do not match with the composite figure provided by the author in #4.  Are the authors confident about the composite gel they posted in #4? The composite image that the authors show leaves out the DR5, cleaved caspase-3, and cleaved PARP bands from the NT and F4 lanes of figure 6F. Is there a mistake in the arrangement of the Beta-actin panel while preparing the composite figure in #4 or Figures 6D and F in the paper? One possible reason that I can think off is that both the figures 6D and 6F show similar band profiles for NT and F4 panels so that is why the authors chose to remove the entire NT and F4 panel from figure 6F while preparing the composite figure in #4 and only the NT and F4 panels in the presence of IM are shown. If that is the case then the authors should also select the corresponding beta-actin bands so that the correct loading control matches along with the correct samples. I am sure that the authors might want to review the composite gel that they have provided with great seriousness. , 
 Any update from the authors? Were you successful at contacting the journal? , 
 Correction has been submitted. , 
 Corrected Figure  ,