The potent oncogene NPM-ALK mediates malignant transformation of normal human CD4(+) T lymphocytes
Authors: James L Riley
Michael Kalos
Fang Wei
Mariusz A Wasik
Gwenn Danet-Desnoyers
Darshan Roy
Andrew Medvec
Hong Yi Wang
Qian Zhang
Xiaobin Liu
Ewa Tomczak
Christopher Watt
Shuguang Jiang
Qun-Bin Xiong
Journal: American Journal Of Pathology (2013)
Discipline: CLINICAL MEDICINE
Keywords: NPM-ALK; CD4(+) T lymphocytes; malignant transformation; anaplastic large-cell lymphomas; immortalized cells

Abstract:  With this study we have demonstrated that in vitro transduction of normal human CD4(+) T lymphocytes with NPM-ALK results in their malignant transformation. The transformed cells become immortalized and display morphology and immunophenotype characteristic of patient-derived anaplastic large-cell lymphomas. These unique features, which are strictly dependent on NPM-ALK activity and expression, include perpetual cell growth, proliferation, and survival; activation of the key signal transduction pathways STAT3 and mTORC1; and expression of CD30 (the hallmark of anaplastic large-cell lymphoma) and of immunosuppressive cytokine IL-10 and cell-surface protein PD-L1/CD274. Implantation of NPM-ALK-transformed CD4(+) T lymphocytes into immunodeficient mice resulted in formation of tumors indistinguishable from patients' anaplastic large-cell lymphomas. Our findings demonstrate that the key aspects of human carcinogenesis closely recapitulating the features of the native tumors can be faithfully reproduced in vitro when an appropriate oncogene is used to transform its natural target cells; this in turn points to the fundamental role in malignant cell transformation of potent oncogenes expressed in the relevant target cells. Such transformed cells should permit study of the early stages of carcinogenesis, and in particular the initial oncogene-host cell interactions. This experimental design could also be useful for studies of the effects of early therapeutic intervention and likely also the mechanisms of malignant progression. 

Retracted: No

PubPeer comments:
 July 2017 correction, which reads: *In Fig 4B, the 5th and 7th fluorescent microscopy images for HA-Core191 + miR-345 inhibitor (10nM) and HA-Core173 + miR-345 inhibitor (10nM) are duplicates of each other. The authors have provided a new set of cell images form the same experiment done at a different time for Fig 4B. Please see the correct Fig 4 and its caption below.* *The authors would like to clarify that for Fig 3C, although the last two FACS Calibur images appear to have identical sections, these images are acquired directly from the computer machine in their original form. They have provided the images in S1 Dataset.* , 
 Here is the original Figure 4 with the duplicated panels marked in red.  Also, the HA-Core173 panels appears to have a perfect fit with the HA-Core173+miR-345 inhibitor (5 nM) panel. Here are the two panels fitted on top of each other:  , 
 For Figure 3C, the correction states: *The authors would like to clarify that for Fig 3C, although the last two FACS Calibur images appear to have identical sections, these images are acquired directly from the computer machine in their original form. They have provided the images in S1 Dataset.* Here are Figure 3C s panels compared, with boxes of the same color focusing attention to areas that look similar. It appears hard to believe that these are raw data as generated by two flow cytometry measurements from 2 different samples, as the authors state.  Could the authors please share the original FCS files instead of the  raw tif files, which look exactly like the data in Figure 3C? , 
 Figure S4. * Blue boxes: The 6.25 uM Curcumin and 12.5 uM Curcumin panels appear to look similar, although at different exposures. Contrast enhanced to bring out details.  , 
 2020 retraction. https: journals.plos.org plosone article?id=10.1371 journal.pone.0235375 After this article [1] and its subsequent correction [2] were published, additional concerns were raised: The PLOS ONE Editors re-reviewed the concerns about the 4th and 5th FACS panels in Fig 3C, which were discussed previously in [2]. Based on our reassessment, we do not consider that the prior correction [2] resolves the concerns about similarities between the lower quadrants of these panels. We regret that this was not fully addressed before the correction was published. The authors provided additional dot plots for four of the five experimental conditions but noted that the raw data (.fcs files) are no longer available. As such, the concerns about this figure remain unresolved and call into question the validity of the results reported for the 25 and 50 nM miR-345 mimic conditions. The u03b2-actin panels in Fig 2C and S4 Fig appear similar, although they represent different experiments. This image issue has implications for the quantitative data reported for the corresponding western blot panels. The authors provided results from an independent replicate for each experiment and noted that an error may have been made in preparing S4 Fig, but they did not provide uncropped images of the original raw blots to support the published results. In the original version of Fig 4B [1], the lower edge of the HA-Core173 panel and the upper edge of the HA-Core173 + miR-345 inhibitor (5 nM) panels align in a manner suggesting that the two panels may show neighboring fields of the same data. In addition, as noted in [2], duplicate images were shown in Fig 4B as representing HA-Core191 + miR-345 inhibitor (10nM) and HA-Core173 + miR-345 inhibitor (10nM). The authors confirmed these issues and noted they may have resulted from mislabeling of images and or errors in figure preparation. In the previous correction [2], Fig 4B images were replaced with replicate data. The authors provided image data in support of the original published results and replication results but these did not resolve the concerns about the original figure. The u03b2-actin blots are overexposed in Figs 3A, 4A and 4B, such that they do not provide a clear representation of the relative protein abundance across lanes. This calls into question the reliability of quantitative results reported for the corresponding western blot experiments. The authors provided replication results for each of these experiments along with associated quantitative data, but did not provide uncropped images of the original raw blots reported in the the published figures. The quantitative data provided relied on the saturated u03b2-actin blots, did not resolve the concern about the reliability of the densitometric analyses, and indicated that the published densitometry results represent results from a single sample per condition rather than summarizing data across multiple experimental replicates. The 6.25 and 12.5 u03bcM Curcumin panels shown for the Annexin V-FITC experiment in S4 Fig appear similar when brightness is adjusted. The authors commented that this was due to an error in figure preparation and they provided results from replicate experiments to support this figure. The above concerns call into question the reliability of the reported results and conclusions, and so the PLOS ONE Editors retract this article. The corresponding author (TYH) notified the journal that TYS and TYH agreed with retraction and apologize for the issues with the published article, but stand by the articleu2019s findings. The other authors either could not be reached or did not respond directly. References 1.Shiu T-Y, Huang S-M, Shih Y-L, Chu H-C, Chang W-K, Hsieh T-Y (2013) Hepatitis C Virus Core Protein Down-Regulates p21Waf1 Cip1 and Inhibits Curcumin-Induced Apoptosis through MicroRNA-345 Targeting in Human Hepatoma Cells. PLoS ONE 8(4): e61089. https: doi.org 10.1371 journal.pone.0061089 ,