58/0008-5472.CAN-18-2096 Author Manuscripts Have Been Peer Reviewed and Accepted for Publication but Have Not Yet Been Edited

Author Manuscript Published OnlineFirst on July 22, 2019; DOI: 10.1158/0008-5472.CAN-18-2096 Author manuscripts have been peer reviewed and accepted for publication but have not yet been edited.

Granados et al, 2019

Selective Targeting of Myoblast Fusogenic Signaling and Differentiation-Arrest

Antagonizes Rhabdomyosarcoma Cells

Valerie A. Granados1, Usha Avirneni-Vadlamudi1, Pooja Dalal1, Samuel R.

Scarborough1, Kathleen A. Galindo1, Priya Mahajan2, and Rene L. Galindo1,2,3

Departments of Pathology1, Pediatrics2, and Molecular Biology3, University of Texas

Southwestern Medical Center, 5323 Harry Hines, Dallas, TX.

Correspondence: Rene L. Galindo, M.D., Ph.D.

Phone: 214.648.4116, Fax: 214.648.4070

E-mail: [email protected]

Classification/Manuscript Info: Priority Report

Word Count: Abstract- 240; Manuscript Length- 2,500 (not including References),

References - 27

Running Title: Selective Targeting of myoblast fusion signaling in RMS

Key Words: Rhabdomyosarcoma / PAX-FOXO1 / Myoblast Fusion / Akt / EGFR

The authors declare NO conflict of interest

Downloaded from cancerres.aacrjournals.org on September 28, 2021. © 2019 American Association for Cancer Research. Author Manuscript Published OnlineFirst on July 22, 2019; DOI: 10.1158/0008-5472.CAN-18-2096 Author manuscripts have been peer reviewed and accepted for publication but have not yet been edited.

Granados et al, 2019

ABSTRACT

Rhabdomyosarcoma (RMS) is an aggressive soft tissue malignancy comprised

histologically of skeletal muscle-lineage precursors that fail to exit the cell-cycle and

fuse into differentiated syncytial muscle - the underlying pathogenetic mechanisms

for which remain unclear. In contrast to myogenic transcription factor signaling,

the molecular machinery that orchestrates the discrete process of myoblast fusion in

mammals is poorly understood, and unexplored in RMS. The fusogenic machinery

in Drosophila, however, is understood in much greater detail, where myoblasts are

divided into two distinct pools: Founder Cells (FCs) and fusion competent myoblasts

(fcms). Fusion is heterotypic and only occurs between FC and fcms. Here, we

interrogated a comprehensive RNA-seq database and found that human RMS

diffusely demonstrates an FC-lineage gene signature, revealing that RMS is a

disease of FC-lineage rhabdomyoblasts. We next exploited our Drosophila RMS-

related model to isolate druggable FC-specific fusogenic elements underlying RMS,

which uncovered the Epidermal Growth Factor Receptor (EGFR) pathway. Using

RMS cells, we showed that EGFR inhibitors successfully antagonized RMS RD cells,

while other cell lines were resistant. EGFR inhibitor-sensitive cells exhibit decreased

activation of the EGFR intracellular effector Akt, while Akt activity remained

unchanged in inhibitor-resistant cells. We then demonstrate that Akt inhibition

antagonizes RMS – including RMS resistant to EGFR inhibition – and sustained

activity of the Akt1 isoform preferentially blocks rhabdomyoblast differentiation

potential in cell culture and in vivo. These findings point towards selective targeting

Granados et al, 2019

of fusion- and differentiation-arrest via Akt as a broad RMS therapeutic

vulnerability.

PRECIS

EGFR and its downstream signaling mediator AKT1 play a role in the fusion and

differentiation processes of rhabdomyosarcoma (RMS) cells, representing a therapeutic

vulnerability of RMS.

Granados et al, 2019

INTRODUCTION

Rhabdomyosarcoma (RMS) is a well-known problem in pediatric oncology, as children

with high-risk RMS endure a 3-year event-free survival rate of only 20% (1).

Histologically, RMS is comprised of neoplastic skeletal muscle-lineage precursors that

fail to exit the cell cycle and terminally differentiate. Rare for a somatic tissue, skeletal

muscle requires that precursor cells not only undergo lineage-specific differentiation, but

also fuse and form a syncytium. Though critical effort has been giving to deciphering

myogenic signaling in the settings of both muscle development and RMS, the

mechanisms orchestrating mammalian myoblast fusion are poorly understood.

In Drosophila, myoblast fusion is understood in greater detail (2), where myoblasts

are divided into two pools: Founder Cells (FCs) and fusion competent myoblasts (fcms).

FCs are seminal, establishing the position of each myofiber, while fcms seek out FCs and

fuse. FC-fcm recognition is mediated by IgSF receptors – the Kirre subfamily, unique for

FCs, and the fcm-specific Nephrin subfamily. Upon FC-fcm adhesion, the FC lineage-

restricted adaptor molecule Rols links the transmembrane signal to the cytoskeleton and

drives downstream fusion events. We and others have shown that Kirrel, Nephrin, and

Rols orthologs participate in vertebrate myoblast fusion (3-5). We additionally have

found that overexpression of mammalian Rols, named TANC1, influences RMS

pathobiology (5).

Unknown, however, is whether misexpression of the FC program broadly underlies

RMS. Additionally, identifying fusion regulators relevant to RMS for which inhibitors

are available would suggest new therapeutic opportunities.

Granados et al, 2019

MATERIALS AND METHODS

Additional information can be found in Supplemental Methods.

Drosophila Genetics

Transgenes and screen methods were as previously described (6, 7).

Cell culture and reagents

Cells lines were handled as previously described (5). RMS lines were used up to 30

passages, C2C12 up to 20. STR profiles for all lines were obtained at UTSW’s

sequencing core and verified as authentic. Mycoplasma testing regularly was negative.

Lines were obtained from: C2C12, ATCC; Rh30, M. Hatley (St. Jude); RD, E. Olson

(UTSW); SMS-CTR, C. Linardic (Duke). Please see Supplemental Methods for

information regarding drugs and vehicles used, and shRNA (Dharmacon) sequences.

MTT assays were performed using the Vibrant™ MTT Assay Kit (V-13154) (Molecular

Probes/Invitrogen). TUNEL was performed using the DeadEnd™ Fluorometric TUNEL

kit (Promega).

Indexes were calculated from cells cultured for six days in differentiation medium

(see Supplemental Methods) from three independent experiments. For differentiation,

the percentage of nuclei in MHC-positive tissue were scored. For fusion, the percentage

of nuclei present in MHC-positive bi- or multi-nucleated myotubes were counted. For

proliferation, the percentage of Ki67-positive cells were scored. For tumor sections,

mitotic figures or Ki67-positive nuclei were scored.

Granados et al, 2019

Xenografts

Studies were supervised and approved by UTSW’s Institutional Animal Care and Use

Committee. Drugs were administered (see Supplemental Methods for dosing) when

tumor size reached ~100 mm3. An event was based on Pediatric Preclinical Testing

Program criteria: tumor volume quadrupling from a base volume [here, 200 mm3 for RD

(slower growing), 250 mm3 for RH30 (faster growing)].

Statistics

Type I error was evaluated by two-tailed Student’s t test. P values less than 0.05 was

considered significant. Type II was evaluated by Achieved Statistical Power (post hoc)

analysis, values greater than 0.80 was considered significant. Data are mean ± SEM.

Software used were Excel (Microsoft), Prism 7 (GraphPad), and G*Power 3 (Heinrich-

Heine-Universität).

Granados et al, 2019

RESULTS

To probe for RMS FC/fcm-gene expression levels, we surveyed the Oncogenomics RNA-

seq database, derived from an extensive collection of human RMS specimens (8). As

mentioned above, each Drosophila myofiber forms from one FC cell, with the remainder

of the syncytium comprised of sequentially fused fcms. Thus, fcms dramatically

outnumber FCs. However, when querying FC-marker expression levels in the panel, we

found that TANC1 and KIRREL1 transcripts were broadly abundant in RMS negative

and positive for the PAX-FOXO (PF) oncoprotein, while KIRREL3 [the encoding gene

for which possesses a PF transcriptional activation site] is overexpressed in PF-positive

RMS (Fig. 1A) (9). In contrast, expression levels of the fcm-NEPRIN orthologs, NPHS1

and NPHS2, are downregulated (Fig. 1A). These findings are consistent with our

previous immunohistochemical analysis of RMS tumors, which showed diffuse positivity

for TANC1 (5). These data show that RMS associates with the FC-signature.

To isolate potentially targetable FC-signaling elements, we turned to our Drosophila

PAX-FOXO1 model (6, 7), which we exploit to uncover new influential RMS genetic

elements (5, 10)]. We identified two chromosomal deletions, Df(2L)pr-A16 and

Df(2R)Excel6076, that suppress PAX-FOXO1-based lethality (Fig. 1B). Df(2L)pr-A16

deletes EGFR, while Df(2R)Excel6076 deletes EGF (named spitz in flies). As EGFR

signaling is known to drive naïve Drosophila myoblasts to the FC-differentiation program

we tested individual loss-of-function alleles in EGFR or spitz, which similarly suppressed

PAX-FOXO1 lethality (Fig. 1B). These data identify EGFR signaling – a druggable

pathway – as a potential FC-based RMS target.

Granados et al, 2019

To extrapolate this finding to mammals, we established that EGFR signaling is active

and regulated in wild-type cultured myoblasts (Fig. S1A). We questioned whether the

EGFR inhibitors Erlotinib (EGFR tyrosine kinase inhibitor) or Cetuximab (humanized

monoclonal interfering antibody) antagonize RMS, utilizing the PF-negative RD and

RH36 cell lines, and the RH30 PF-positive line (Fig. S1B). We profiled viability for each

line against each agent (Fig. S1C) [IC50 values were similar to human carcinoma cells

(11, 12)] and demonstrated that each inhibitor antagonized EGFR. Both inhibitors

interfered with RD cell proliferation, increased Myosin Heavy Chain (MHC)-positive

terminal differentiation (Fig. 2A & B) (Fig. S1D and E), and blocked anchorage

independent growth (Fig. S1F and G). TUNEL assay for apoptotic cell death was

negative (Fig. S1H). In vivo, tumorigenesis was inhibited, event-free survival increased,

mitotic activity decreased, and MHC-expression enhanced (Fig. 2C & D) (Fig. S1I-L).

RH36 and RH30 cells, however, were not antagonized (Fig. S1M).

As the RD line carries an oncogenic N-RAS mutation (Q61H), we hypothesized that

an intracellular signaling pathway other than RAS must be antagonized upon

Erlotinib/Cetuximab treatment. We analyzed inhibitor-treated cells and observed

downregulated Akt activation, whereas MEK/MAPK or STAT3 activation levels showed

no decrease (Fig. S2A). We next found that Akt activation levels remained unaltered in

EGFR-inhibitor resistant RH36 and RH30 cells (Fig. S2B). These results infer that Akt is

a critical RMS effector.

To test this notion, we treated the RMS cell lines with an allosteric Akt inhibitor,

MK-2206 [IC50 values were similar to human carcinoma cells (Fig. S3A) (13)]. We

additionally included a fourth line, SMS-CTR (PF-negative), found to be EGFR inhibitor

Granados et al, 2019

resistant (Fig. S3B). We observed potent blockage of Akt activation in each cell line (Fig.

S3C), with all lines now exhibiting decreased proliferative activity and anchorage

independent growth (Fig. 3A and B) (Fig. S3D) (note - RH36 cells did not form colonies

in soft agar). TUNEL assays performed on RD and RH30 MK-2206-treated cells were

negative (Fig. S3E). In vivo, we observed inhibited tumorigenesis, increased event-free

survival, and decreased proliferation (Fig. 3C) (Fig. S3F and G). No difference in MHC-

positivity was observed, however (Fig. S3H). Together, these findings point towards Akt

as a broadly targetable RMS vulnerability.

Though transcripts for Akt1/2/3 are detectable in human skeletal muscle, only Akt1

and 2 protein are detected (14). As Akt1 has been shown to function early in myogenesis

and promote myoblast proliferation, while Akt2 downstream directs myoblast fusion and

differentiation (15), we hypothesized that the MK-2206 Akt inhibitor, though promoting

RMS cell cycle exit, failed to induce RMS rhabdomyoblast fusion and differentiation due

to dual blockage of Akt1/2. As differing roles for Akt1/2 in RMS are unexplored, we

silenced Akt1 or Akt2 (Fig. S4A) and found that Akt1-silenced RD cells exhibited a

marked rescue of fusion and differentiation potential when compared to control or Akt2-

silenced cells (Fig. 4A and B). In vivo, tumorigenesis was inhibited, event-free survival

increased, mitotic activity decreased, and MHC-expression enhanced (Fig. 4C) (Fig.

S4C). These results reveal that sustained Akt1 activity preferentially influences the

failure of RMS cells to complete the myogenic developmental program, and that

targeting of Akt1 is sufficient to rescue RMS cell differentiation-arrest.

Granados et al, 2019

DISCUSSION

We previously reported that correcting FC-lineage TANC1 overexpression induces RMS

cells to terminally differentiate (5). Here, utilizing the Oncogenomics database, we now

reveal that RMS broadly demonstrate the FC-program gene signature. Collectively, these

findings argue that misexpression of FC-programming is a common RMS mechanism.

Since it remains unclear the extent to which the lineage-restricted process of Drosophila

myoblast fusion is precisely conserved in mammals, unknown is whether human RMS

tumor initiation occurs in FC-lineage cells, or whether misexpression of the FC-program

occurs downstream during tumor progression.

Utilizing our Drosophila model, we probed for FC-elements that possess druggable

human orthologs and uncovered EGFR, which [though studied in cultured RMS cells (16,

17)] has not been functionally probed in vivo. Though EGFR inhibitors demonstrated

efficacy against RD cells, EGFR inhibition was ineffective against the remaining lines

tested. As numerous receptor kinases (e.g., FGFR4, c-MET) have been shown to

influence RMS (1), we speculate that the RMS cells resistant to EGFR inhibition do not

rely upon EGFR for growth-promoting signaling. Interestingly, we note that a subset of

fly myoblasts utilizes an FGFR4 ortholog for FC program activation. We thus speculate

that PF-positive RMS – FGFR4 is a direct target of the PF transcription factor – and the

subset of PF-negative RMS possessing activating FGFR4 mutations [~10% (8, 18)]

instead rely upon FGFR4 for FC-program dysregulation. Whether EGFR inhibitor

sensitivity is common or limited in PF-negative RMS remains an open question.

MK-2206, however, broadly antagonized RMS, including RMS driven by oncogenic

N-RAS (RD cells), mutationally activated FGFR4 (RH36), PAX3-FOXO1 (RH30), and

Granados et al, 2019

oncogenic H-RAS (SMS-CTR) (19). Though Akt-mediated phosphorylation has been

shown to inhibit the activity of PF (20), other studies have demonstrated that synthetic

lethality in RMS can be induced by dual inhibition of the PI3K pathway (PI3K/mTOR or

TORC1/2 inhibition) and either RAS or Hedgehog signaling (21-24), and that

PI3K/mTOR inhibitors demonstrate efficacy against FGFR4-driven RMS (25). Here we

newly reveal that mono-targeting of Akt is both effective and sufficient to antagonize

RMS and point towards Akt as a critical RMS nodal point. We additionally found that

sustained Akt1 activity preferentially incites RMS differentiation-arrest, suggesting that

inhibitors specific for Akt1 would be similarly effective against RMS, and presumably

with less overall toxicity than pan-Akt inhibitors.

Focusing on the mechanisms that underlie RMS cell differentiation-arrest, the read-

outs surveyed in these studies focus on myoblast maturation, and not cytotoxicity. Thus,

we suggest that scoring for tumor regression (which requires cytotoxicity) is not the most

appropriate metric, and that event-free-survival is a better preclinical gauge. This notion

differs in part from the Pediatric Preclinical Testing Program (PPTP), which tests for

cytotoxicity and tumor regression, and thus reported MK-2206 as not inducing greater

than 50% tumor volume regression. Neither Erlotinib or Cetuximab has been tested by

the PTPP. We next anticipate testing EGFR and Akt inhibitors in the context of

conventional chemotherapy agents, as an emphasis has been placed on identifying agents

that enhance outcomes in combination with current therapeutic protocols. As both MK-

2206 and Erlotinib have been successfully Phase I tested in pediatric patients (26, 27), we

speculate that these agents, when combined with established protocols, will improve

RMS outcomes.

Granados et al, 2019

ACKNOWLEDGEMENTS

We are grateful to S. Skapek, J. Amatruda, C. Linardic, L. Crose for critical review of

data and manuscript. Studies were supported by: RLG - American Cancer Society

(124717-RSG-13-194-01-DDC), Cancer Prevention Research Institute of Texas

(RP120685), NIH/NCI (R01CA193339), Wipe-out Kid’s Cancer Foundation, Live Like

Bella Foundation; VAG- Pharmacology Training Grant (T32GM007062). We apologize

to the studies and authors that we were unable to discuss or cite due to space limitations.

Granados et al, 2019

REFERENCES

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15. Heron-Milhavet L, Franckhauser C, Rana V, Berthenet C, Fisher D, Hemmings BA, et al. Only Akt1 is required for proliferation, while Akt2 promotes cell cycle exit through p21 binding. Mol Cell Biol. 2006;26(22):8267-80. 16. De Giovanni C, Landuzzi L, Frabetti F, Nicoletti G, Griffoni C, Rossi I, et al. Antisense epidermal growth factor receptor transfection impairs the proliferative ability of human rhabdomyosarcoma cells. Cancer Res. 1996;56(17):3898-901. 17. Ricci C, Polito L, Nanni P, Landuzzi L, Astolfi A, Nicoletti G, et al. HER/erbB receptors as therapeutic targets of immunotoxins in human rhabdomyosarcoma cells. J Immunother. 2002;25(4):314-23. 18. Chen X, Stewart E, Shelat AA, Qu C, Bahrami A, Hatley M, et al. Targeting oxidative stress in embryonal rhabdomyosarcoma. Cancer cell. 2013;24(6):710- 24. 19. Hinson AR, Jones R, Crose LE, Belyea BC, Barr FG, and Linardic CM. Human rhabdomyosarcoma cell lines for rhabdomyosarcoma research: utility and pitfalls. Front Oncol. 2013;3:183. 20. Wachtel M, and Schafer BW. PAX3-FOXO1: Zooming in on an "undruggable" target. Semin Cancer Biol. 2018;50:115-23. 21. Graab U, Hahn H, and Fulda S. Identification of a novel synthetic lethality of combined inhibition of hedgehog and PI3K signaling in rhabdomyosarcoma. Oncotarget. 2015;6(11):8722-35. 22. Renshaw J, Taylor KR, Bishop R, Valenti M, De Haven Brandon A, Gowan S, et al. Dual blockade of the PI3K/AKT/mTOR (AZD8055) and RAS/MEK/ERK (AZD6244) pathways synergistically inhibits rhabdomyosarcoma cell growth in vitro and in vivo. Clin Cancer Res. 2013;19(21):5940-51. 23. Guenther MK, Graab U, and Fulda S. Synthetic lethal interaction between PI3K/Akt/mTOR and Ras/MEK/ERK pathway inhibition in rhabdomyosarcoma. Cancer Lett. 2013;337(2):200-9. 24. Yohe ME, Gryder BE, Shern JF, Song YK, Chou HC, Sindiri S, et al. MEK inhibition induces MYOG and remodels super-enhancers in RAS-driven rhabdomyosarcoma. Sci Transl Med. 2018;10(448). 25. McKinnon T, Venier R, Yohe M, Sindiri S, Gryder BE, Shern JF, et al. Functional screening of FGFR4-driven tumorigenesis identifies PI3K/mTOR inhibition as a therapeutic strategy in rhabdomyosarcoma. Oncogene. 2018;37(20):2630-44. 26. Jakacki RI, Hamilton M, Gilbertson RJ, Blaney SM, Tersak J, Krailo MD, et al. Pediatric phase I and pharmacokinetic study of erlotinib followed by the combination of erlotinib and temozolomide: a Children's Oncology Group Phase I Consortium Study. J Clin Oncol. 2008;26(30):4921-7. 27. Fouladi M, Perentesis JP, Phillips CL, Leary S, Reid JM, McGovern RM, et al. A phase I trial of MK-2206 in children with refractory malignancies: a Children's Oncology Group study. Pediatr Blood Cancer. 2014;61(7):1246-51.

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FIGURE LEGENDS

Fig. 1. FC-genes in RMS.

(A) RMS demonstrates an FC- signature. Relative abundance levels in a human RMS

tumor cohort profiled by RNA-seq. DESMIN is a muscle-specific intermediate filament,

while GFAP and KRT20 are filament markers for glial and gastrointestinal

adenocarcinoma neoplasms, respectively. PAX3-variant fusion (PAX3-INO80D or -

NCOA1) specimens are shown separately.

(B) EGF or EGFR loss-of-function alleles suppress PAX-FOXO1. Based on Mendelian

ratios, the F1 population should be 50% control and 50% PAX7-FOXO1-expressing

adults (“Expected”). PAX7-FOXO1 causes lethality, as PAX7-FOXO1 adults comprise

~20% of F1 adults (“control”; n = 124). Chromosomal deletions Df(2L)pr-A16 (n = 47)

or Df(2R)Excel6076 (n = 77) suppresses PAX-FOXO1 lethality, as do two EGF (named

spitz in Drosophila) [spiDG04705 (n = 55) spis3547 (n = 66)] or two EGFR [Egfrf2 (n = 66)

and Egfrt1 (n = 60)] loss-of-function alleles (though the Egfrt1 allele showed a P value of

0.067). rolsP1729 is the loss-of-function allele previously isolated as a PAX-FOXO1

suppressor. Df(2L)ed1 (n = 160) (third column) and DF(3R)23D1 (n = 74) (red column)

are unrelated chromosomal deletions included as controls to demonstrate examples of a

non-modifier and genetic enhancer, respectively. P values: *P < 0.05, **P < 0.01 versus

Control.

Fig. 2. Erlotinib and Cetuximab block tumorigenicity in RD RMS cells.

Granados et al, 2019

(A,B) Erlotinib- or Cetuximab-treated RD cells show decreased proliferation and

enhanced differentiation. Cells were stained with Ki67 or MHC antibody, and DAPI.

Erlotinib concentration = 10 M, Cetuximab concentration = 1 g/mL.

(C,D) Erlotinib or Cetuximab antagonizes tumorigenesis. Shown are tumor growth and

event-free survival (see Methods) for “Control” (6% Captisol) (n = 3) versus Erlotinib-

treated (n = 4) (Panel C), and “Control” (PBS) (n = 3) versus Cetuximab-treated (n = 3)

tumors (Panel D). Achieved Statistical Power for “tumor volume” and “event-free

analyses were 0.99 and 0.82, respectively, for the Erlotinib study; for Cetuximab, 0.97

and 0.95, respectively. Myosin Heavy Chain IHC shows enhanced differentiation within

Erlotinib- or Cetuximab-treated xenografts.

Scale bar = 100 m. P values: *P < 0.05 versus Control.

Fig. 3. MK-2206 blocks tumorigenicity in PF-negative and -positive RMS.

(A) MK-2206 antagonizes RMS proliferation in culture. RD, RH36, RH30, and SMS-

CTR cells were each cultured with MK-2206 (0.5 uM) and stained with Ki67 antibody

and DAPI.

(B) MK-2206-treated RMS cells show decreased colony formation in soft agar. Shown

are average number of colonies per 20×-objective field.

survival (see Methods) plots for “Control” (15% Captisol) (n = 4) and MK-2206-treated

(n = 4) RD or RH30 tumors. Achieved Statistical Power for RD and RH30 tumor volume

Granados et al, 2019

and event-free survival analyses were 1.00 and 0.95 (RD), and 1.00 and 0.93 (RH30),

respectively.

P values: **P < 0.01, ***P < 0.001 versus Control.

Fig. 4. Akt1 silencing rescues RMS cell differentiation-arrest in vitro and in vivo.

(A,B) Akt1 silencing rescues RD cell fusion- and differentiation-arrest. RD cells

expressing shRNA (transient transfection) against GFP (Control), Akt1, or Akt2 are

shown, stained with MHC antibody and DAPI (panel A). Fusion and differentiation

indexes (panel B) show that Akt1-silenced RD cells exhibited rescue of fusion and

differentiation when compared to control or Akt2-silenced cells.

express shRNA against eGFP, Akt1, or Akt2 upon doxycycline administration were

generated. Shown are tumor growth and event-free survival plots for “Control”

(shRNA>>eGFP) (n = 4), shAkt1 (shRNA>>Akt1) (n = 3), and shAkt2 (shRNA>>Akt2)

expressing (n = 3) RD tumors. Achieved Statistical Power for RD Control versus Akt1

tumor volume and event-free survival analyses were 0.97 and 0.97, respectively.

P values: *P < 0.05, **P < 0.01, *** P < 0.001 versus Control.

Selective Targeting of Myoblast Fusogenic Signaling and Differentiation-Arrest Antagonizes Rhabdomyosarcoma Cells

Valerie A Granados, Usha Avirneni-Vadlamudi, Pooja Dalal, et al.

Cancer Res Published OnlineFirst July 22, 2019.

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