Research ArticleExperimental Studies
Open Access

Therapeutic Efficacy of Temsirolimus in a Patient-derived Model of Metastatic Fibrolamellar Hepatocellular Carcinoma

JENNIFER L. LEITING, MATTHEW C. HERNANDEZ, JOHN R. BERGQUIST, JENNIFER A. YONKUS, AMRO M. ABDELRAHMAN, MICHAEL S. TORBENSON, NGUYEN H. TRAN, THORVARDUR R. HALFDANARSON, RONDELL P. GRAHAM, RORY L. SMOOT and MARK J. TRUTY
In Vivo September 2023, 37 (5) 1940-1950; DOI: https://doi.org/10.21873/invivo.13290

Abstract

Background/Aim: Fibrolamellar hepatocellular carcinoma (FLHCC) is a rare tumor presenting in younger patients without chronic liver disease. Up to 80-100% develop recurrent disease, necessitating additional surgery or systemic treatment. Systemic options and pre-clinical treatment studies are lacking. We previously described patient-derived xenograft (PDX) development, allowing for pre-clinical studies. Herein, we develop FLHCC PDX models and utilize these to define tumor characteristics and determine the efficacy of systemic agents. Materials and Methods: Primary and lymph node metastatic tumor tissues were obtained at the time of FLHCC resection in two patients. Tumor lysates were screened for protein upregulation. Cell lines were generated from metastatic and primary tumor tissue. The viability of the cell lines was assessed after treatment with temsirolimus, gemcitabine/oxaliplatin, and FOLFIRINOX. Two PDX models were developed from metastatic tissue. For in vivo studies, tumor-bearing mice were treated with temsirolimus, FOLFIRINOX, and Gemcitabine/oxaliplatin. Results: PDX models were successfully generated from metastatic FLHCC, which closely recapitulated the original tumor. Upregulation of mTOR was seen in metastatic tissue compared to primary tumors. Cell lines from metastatic tissue demonstrated significant sensitivity to temsirolimus. In vivo testing of PDX models demonstrated a significant response to single-agent temsirolimus with minimal toxicity. Conclusion: Herein, we demonstrate the feasibility of developing PDX models that closely recapitulate FLHCC. Upregulation of mTOR was seen in metastatic tissue compared to primary tissue. The efficacy of mTOR inhibition with temsirolimus treatment suggests that the upregulation of the mTOR pathway may be a significant mechanism for growth in metastatic lesions and a potential target for therapeutics.

Key Words:

Fibrolamellar hepatocellular carcinoma (FL-HCC) is a rare tumor that accounts for less than 1% of all primary liver cancers (1). In addition to having a distinct morphology that consists of prominent nucleoli and abundant eosinophilic cytoplasm, FL-HCCs have a characteristic gene fusion on one copy of chromosome 19 (2). This fusion, DNAJB1-PRKACA, was first described in 2014 and has been found to be specific for FL-HCC among hepatocellular neoplasms (3, 4).

In contrast to patients with conventional HCC, patients presenting with FL-HCC are generally young and without chronic liver disease (5, 6). The prognosis for patients diagnosed with FL-HCC is largely dependent on whether the tumor can be surgically resected, with 5-year overall survival of 40 to 75% after resection (6, 7). Unfortunately, 80-100% of patients will recur after oncologic resection, necessitating additional surgery or systemic treatment (7, 8). In one large cancer registry database review, the efficacy of systemic treatments was reported to be mixed with no survival benefit with chemotherapy (9). Given the rarity of the diagnosis and difficulty in establishing clinical trials, there are no preferred or recommended treatment regimens, though there have been small retrospective studies and one phase II study (33690232) that have tried to provide treatment recommendations (10, 11).

Systems for testing chemotherapeutic regimens have been lacking for FL-HCC. One tumor cell line developed from the malignant ascites of a patient with FL-HCC has been described though there have been no reports of chemotherapeutic efficacies (12). Patient-derived xenografts (PDX) closely recapitulate original patient tumor tissue histology, morphology, and genomic characterization (13). Therefore, they can be used to test the efficacy of treatment regimens and predict patient responses (14). Here, we describe the development of PDX models that recapitulate the molecular and histologic characteristics of FL-HCC. Testing of several chemotherapeutic combinations both in vitro and in vivo demonstrated sensitivity to the mTOR inhibitor temsirolimus in these models, developed from metastatic lesions. On target effects on mTOR signaling were noted and evaluation of additional patient samples confirmed upregulation of mTOR, suggesting temsirolimus may have a broader clinical utility in this disease.

Materials and Methods

Reagents. For in vitro experiments, temsirolimus, sorafenib, sunitinib and erlotinib were purchased from Selleckchem (Houston, TX, USA). Gemcitabine, oxaliplatin, 5-fluorouracil, and irinotecan were purchased from Cayman Chemical (Ann Arbor, MI, USA). Drugs were dissolved in dimethyl sulfoxide (DMSO) and stored at −20°C. For in vivo experiments, gemcitabine, oxaliplatin, 5-fluorouracil, and irinotecan were obtained as clinical formulations from the Mayo Clinic Pharmacy (Rochester, MN, USA). Temsirolimus was purchased from Selleckchem and reconstituted in DMSO, poly(ethylene glycol) 300, Tween 80, and purified water.

Establishment of patient-derived xenografts. Establishment of PDX models was performed as previously described (15). All protocols were approved by the Institutional Review Board and the Institutional Animal Care and Use Committee (IACUC) (protocol number: A00003954-18-R21). Informed consent was obtained prior to collection of tissue. Briefly, freshly resected tissue was obtained from the frozen section pathology lab after rapid tissue diagnosis. Primary tumor tissue and lymph node metastases were both diced into approximately 30 mm3 pieces and implanted into the flanks of non-obese diabetic/severe combined immune-deficient (NOD/SCID) mice. Mice were monitored for tumor growth and tumors were harvested when they were at least 125 mm3. Histological verification of the PDX tumor was performed by a liver pathologist.

Immunoblotting. Tumor lysates were resolved by SDS-PAGE and transferred to a nitrocellulose membrane. The primary antibodies used for detection were PKAα cat (A-2) (Santa Cruz, Dallas, TX, USA), phosphorylated mTOR, total mTOR, phosphorylated AKT, total AKT, LC3 I, LC3II, cleaved poly (ADP-ribose) polymerase (PARP), total PARP, cleaved caspase 9, caspase 9, cleaved caspase 3, caspase 3 and β-actin (Cell Signaling Technology, Danvers, MA, USA). Bound antibodies were visualized using the Super Signal West Pico Plus chemiluminescent substrate (ThermoFisher, Waltham, MA, USA).

Immunohistochemistry. Immunohistochemistry was performed on 5-micron thick, formalin fixed, paraffin embedded tissue sections using steam-induced antigen retrieval and standard clinical methods for cytokeratin 7 (Clone OV-TL 12/30; dilution 1:100 Dako, Santa Clara, CA, USA) and CD68 (Clone KP1, dilution 1:50-1:100; Dako) on the Ventana Benchmark (Ventana, Roche, Tucson, AZ, USA) platform.

Fluorescence in-situ hybridization. FISH was performed by the Mayo Cytogenetics Core. Home-brew 5′ PRKACA DNA (clone CTC-548K16) labeled with Spectrum Orange dUTP (Abbott Molecular/Vysis Products) and home-brew 3′ PRKACA DNA (clones RP11-63F22, CTD-2003D17 and CTC-708A18) labeled in Spectrum Green dUTP (Abbott Molecular/Vysis Products) were combined as one probe set. The break-apart (BAP) probe set was applied to individual slides, hybridized, and washed according to the standard interphase protocol. The slides were reviewed and examined as previously described (4)

Tumor dissociation and cell culture. Original patient and PDX tumor tissue was dissociated into single-cell suspensions using the Tumor Dissociation Kit (human) (Miltenyi Biotec, Auburn, CA, USA) and the gentleMACS Dissociator (Miltenyi Biotec) according to the manufacturer’s instructions. Cells were cultured in Dulbecco’s Modified Eagle’s Media (Corning Inc, Corning, NY, USA) with 10% fetal bovine serum (Atlanta Biologicals, Flowery Branch, GA, USA) and 1% antibiotic-antimycotic 100x (ThermoFisher). All cells were cultured at 37°C with 5% CO2.

Cell viability assay. Human and PDX tumor cells were plated on 96-well plates at a concentration of 300 cells/well. After 24 h, cells were treated with one of six chemotherapeutic regimens at concentrations of 1 nM, 10 nM, 100 nM, and 1 μM. Drug dilutions were prepared immediately before administration and all tests were performed in triplicate. After 72 h of treatment, cell viability was assessed using the CellTiter-Glo Luminescent Cell Viability Assay (Promega, Madison, WI, USA) according to the manufacturer’s instructions with a microplate reader (Synergy H1, BioTek, Winooski, VT, USA).

Patient-derived xenograft in vivo experiment. For the in vivo treatment study, male NOD/SCID mice were implanted with PDX tumor tissue in the left flank. When the tumors had reached a palpable size of approximately 60 mm3, mice were randomized into four groups of five mice each. The first group was treated with a vehicle. The second group was treated with gemcitabine [70 mg/ml, intraperitoneally (IP) once weekly] and oxaliplatin (4 mg/ml, IP once weekly). The second group was treated with FOLFIRINOX consisting of 5-fluorouracil (75 mg/kg, IP once weekly), irinotecan (30 mg/kg, IP once weekly), and oxaliplatin (4 mg/kg, IP once weekly). The fourth group was treated with temsirolimus (25 mg/kg, IP four times weekly). Twice weekly, mice were weighed, and tumor volumes were measured with a digital caliper. After 29 days, the mice were sacrificed, and tumors were harvested.

Human tissue. Immunohistochemistry was performed for mTOR on the human primary and metastatic lesions from the pathology database. The staining distribution (0, none to less than 5%; 1, 5-30%; 2, 30-60%, 3, >60%) and intensity (0, none; 1, weak; 2, moderate, 3, strong) were evaluated by a liver pathologist (Rondell P. Graham) blinded to tissue origin. The composite score was calculated by multiplying these two values. The differences in composite score between the primary and the metastatic lesions were compared using the paired Student’s t-test.

Statistical analysis. Statistical analysis for in vitro cell viability was determined using the Wilcoxon rank sum test. Statistical analysis of change in tumor volume and mouse weight was determined initially by the Kruskal Wallis test given the small sample sizes (n=5). Individual Wilcoxon rank sum tests were then performed for each pair using the Bonferroni correction. A p-value of <0.05 was considered significant and all tests were 2-sided. All statistical analysis was performed using JMP software (JMP Pro, Version 13.0.0, SAS Institute Inc., Cary, NC, USA).

Results

Patient-derived xenografts can be generated from metastatic FL-HCC lesions. The first patient, Patient #1, was an 18-year-old male who presented with an acute pulmonary embolism. During the medical evaluation of the patient, a large right-sided hepatic mass with large adjacent lymph nodes was discovered on cross-sectional imaging. This was biopsied and determined to be FL-HCC (Figure 1A and B). The patient underwent an extended right hepatectomy with a radical lymphadenectomy which revealed a well-differentiated FL-HCC measuring 8×8×7 cm (Figure 1C). Five of 16 lymph nodes were involved, and he was started on gemcitabine and oxaliplatin post-operatively. The patient progressed on this treatment regimen and was switched to nivolumab after clinical molecular analysis found the tumor to be PD-L1 positive. He went on to have a complete radiographic response though this only lasted for approximately 8 to 9 months. He has since died of disease progression.

Figure 1.
Figure 1.

Cross-sectional imaging and gross pathologic specimen images from fibrolamellar hepatocellular carcinoma (FLHCC) lesions. (A) Cross-sectional image of primary tumor from Patient #1. (B) Cross-sectional imaging of enlarged lymph node from Patient #1. (C) Gross pathologic specimen from Patient #1. (D) Cross-sectional image of primary tumor from Patient #2. (E) Cross-sectional imaging of enlarged lymph node from Patient #2. (F) Gross pathologic specimen from Patient #2.

The second patient, Patient #2, was a 16-year-old male who presented with abdominal fullness. An ultrasound revealed a left-sided hepatic lesion and biopsy confirmed FL-HCC (Figure 1D and E). The patient proceeded directly to surgery and underwent resection of segments IVA, IVB, V, and VIII with lymphadenectomy. Final pathology showed a moderately-differentiated FL-HCC measuring 13×13×10 cm (Figure 1F). Nine of 18 lymph nodes were involved. The last MRI showed lesions in the remaining liver; unfortunately, the patient has since been lost to follow-up.

Fresh tumor tissue was obtained from both patients after evaluation and confirmation of the diagnosis in the frozen section pathology laboratory. Both primary tumor tissue and involved lymph node tissue were available and implanted into NOD/SCID mice. For patient #1, the primary tumor tissue did not grow, and the mice were sacrificed after 277 days. In contrast, tumor tissue from the lymph node metastasis grew quickly with a time to tumor formation (TTF) of 27 days and a time to tumor harvest (TTH) of 67 days. Similarly, for patient #2, the primary tumor again did not grow, and the mice were sacrificed after 310 days. Tissue implanted from the lymph node metastasis grew successfully with a TTF of 22 days and TTF of 127 days.

Patient-derived xenografts recapitulate molecular and morphologic features of FL-HCC. Histological examination with hematoxylin and eosin (H&E) stains of both PDX models revealed the characteristic ample granular cytoplasm, open chromatin, and conspicuous nucleoli with close recapitulation of the original patient tumor morphology (Figure 2). Immunohistochemistry of PDX tumors was performed, which showed positivity for CD68 and cytokeratin 7, which are characteristic of FL-HCCs (Figure 2) (16).

Figure 2.
Figure 2.

Morphology and immunohistochemistry of original patient tumor tissue and patient-derived xenograft tumor tissue. All images are at 10´ magnification. F0 and F1 slides in the first three columns are prepared with H&E staining. The final two columns are stained with CD68 and CK7, as listed. Met, Metastasis; PDX, patient-derived xenograft; CK, cytokeratin.

Immunoblot was consistent with the presence of the DNAJB1-PRKACA fusion protein with demonstration of two PRKACA bands including one at approximately 46 kDa, the anticipated mass of the fusion protein. For both patients, the original patient primary tumor, patient lymph node metastasis, and PDX tumors showed evidence of the fusion protein (Figure 3A and C), while normal adjacent liver did not. FISH analysis of the PDX tumors also showed evidence of rearrangement of the PRKACA locus at the genomic level with the loss of the PRKACA 5′ prove as evidenced by solitary green probes seen in evaluation of both models (Figure 3B and D).

Figure 3.
Figure 3.

Immunoblot for the PRKACA protein was performed on both patient and patient-derived xenograft tissue (A and C). Fluorescent in situ hybridization was also performed to detect the rearrangement of the PRKACA locus (B and D). Met, Metastasis; PDX, patient-derived xenograft.

Elevated mTOR activity can be targeted in xenografts. In order to assess in vitro sensitivities, dissociated cells from Patient #1 PDX tumors were treated with several chemotherapeutic regimens. Cell viability was determined after 72 hours of treatment (Figure 4). There was a significant decrease in relative cell viability following exposure to three chemotherapeutic regimens: gemcitabine and oxaliplatin (Gem/Ox), FOLFIRINOX, and the m-TOR inhibitor temsirolimus with IC50s of 1 μM, 100 nM, and 100 nM respectively. Given the significant response to temsirolimus in vitro, immunoblots were performed to assess the degree of mTOR activity in the original patient tumor tissue. The patient lymph node metastasis and the PDX tumor tissue from both patients had increased levels of activated mTOR when compared to both the primary patient tumor and the normal liver tissue (Figure 5A and D), suggesting that the upregulation of the mTOR pathway may be a significant mechanism for growth in these metastatic lesions. Cryopreserved cells from both the primary liver tumor and the lymph node metastasis from both patients were thawed, dissociated, and treated with temsirolimus to assess the response to mTOR inhibition in vitro. The cells from the primary tumor were not responsive to temsirolimus treatment compared to cells from the lymph node metastasis that had a significant response to mTOR inhibition (Figure 5).

Figure 4.
Figure 4.

In vitro treatment efficacy against patient-derived xenograft tumor cells. In vitro viability of cells dissociated from a patient-derived xenograft tumor developed from a lymph node metastasis. Cells were treated with increasing concentrations of chemotherapeutic agents. Cell viability was assessed 72-hours post-treatment. Treatment with gemcitabine/oxaliplatin (A), FOLFIRINOX (B), and temsirolimus (F) resulted in significant responses (p<0.05). There was no significant response to 5-Flourourocil (C), Sorafenib (D), or Erlotinib (E).

Figure 5.
Figure 5.

Assessment of mTOR activity and effect of inhibition on original patient tumors. Immunoblots were performed to assess for phosphorylated mTOR and total mTOR. In both patients, the metastatic lesions showed substantially increased mTOR expression when compared to the primary liver tumor and normal liver tissue (A, D). Beta-actin was used as a loading control. Cells from the primary tumors did not respond to mTOR inhibition with temsirolimus (B, E) while cells from the lymph node metastasis showed a significant response (C, F).

Given these in vitro results, an in vivo study was designed to test the efficacy of these chemotherapeutic regimens in a preclinical PDX mouse model. Mice were implanted with PDX tumor tissue and once the tumor was approximately 1,000 mm3, mice were treated for 29 days with either vehicle, temsirolimus, Gem/Ox, or FOLFIRINOX. Mice treated with temsirolimus and FOLFIRINOX showed a significant reduction in tumor volume over the course of the study when compared to the vehicle-treated group (Figure 6A). The Gem/Ox group initially showed some tumor growth inhibition but by the beginning of the second week, began to show continued tumor growth at a rate similar to the vehicle-treated group (Figure 6A). This correlates to the subsequent disease progression seen in the first patient after treatment with Gem/Ox. Mice in the FOLFIRINOX groups lost a significant amount of weight when compared to the vehicle treated group, as well as when compared to the temsirolimus group (Figure 6B). The only mouse death in the study was in the FOLFIRINOX group and occurred on treatment day 26. Final gross tumor images are demonstrated in Figure 6C.

Figure 6.
Figure 6.

In vivo patient-derived xenograft treatment study. Mice treated with FOLFIRINOX and temsirolimus had significantly smaller tumors when compared to the vehicle-treated group (A). Additionally, mice treated with FOLFIRINOX had weights that were significantly lower than both the vehicle-treated group and the temsirolimus-treated group (B). Only one mouse died during the experiment from the FOLFIRINOX group (C). All p-values have been adjusted for multiple comparisons. Analysis was performed using the Kruskal Wallis test initially followed by individual Wilcoxon rank sum tests for each pair using the Bonferroni correction.

To assess the effect of treatment on the PDX tumors, immunoblot was performed on the treated tumor tissue (Figure 7A). Tumors from all treatment groups showed very limited activated mTOR and no activated AKT, suggesting that proliferation and cell growth was inhibited in all treated tumor tissues. Autophagy was assessed which showed increased expression of LC3 II in tumors treated with temsirolimus as well as FOLFIRINOX. In the treated tumors, an increase in apoptosis in the temsirolimus and the FOLFIRINOX-treated groups was also seen, as evidenced by increased expression of cleaved caspase 3, cleaved caspase 9, and cleaved PARP.

Figure 7.
Figure 7.

(A) Immunoblot of treated tumor tissue demonstrating limited activated mTOR and no activated AKT. (B) Median composite mTOR immunohistochemistry (IHC) score for ten patients with both primary and metastatic fibrolamellar hepatocellular carcinoma (FLHCC), demonstrating increased intensity or mTOR staining in metastatic lesions. (C) Median difference in composite score of primary lesions and the corresponding metastatic lesions was +2, indicating an overall increase in extent and intensity of mTOR staining in the metastatic lesions.

Lastly, we investigated whether these differences in mTOR expression were present in a larger cohort of human FL-HCC tissue samples. We identified 10 patients with both primary and metastatic tissue available for evaluation. The median composite score of the primary lesions was 2 compared to 4 for the metastatic lesions (p=0.01) (Figure 7B). Additionally, the median difference in composite score between the primary lesions and the corresponding metastatic lesions was +2, indicating an overall increase in extent and intensity of mTOR staining in the metastatic lesions (p<0.01) (Figure 7C). This correlates to what we observed in the PDX tissue and provides evidence for the presence of mTOR upregulation in metastatic lesions in a larger cohort of patient tumor tissue.

Discussion

The vast majority of patients with FL-HCC will recur and these recurrences become more difficult to treat over time. Though rare, effective treatment options for these patients are desperately needed. This study proposes a treatment specifically for metastatic lesions. Treatment efficacy of single-agent temsirolimus was shown in vitro as well as in vivo with a patient-derived xenograft model of validated metastatic FL-HCC. This treatment was well tolerated by mice without the significant weight loss seen with FOLFIRINOX treatment, the only other treatment that showed in vivo efficacy. The increased expression of activated mTOR seen in the PDX tumor tissue and patient tumor tissue may be the mechanism for this treatment effect.

There have been two reports of a patient-derived model of FL-HCC (12). Here we report the generation and validation of two additional PDX models of FL-HCC developed from metastatic lesions. The primary tumor from both patients presented here did not grow in mice and we were not able to establish a cell line. The first patient-derived model was also generated from metastatic cells (malignant ascites) (12). Though it is unclear why the primary tumors are unable to grow in the NOD/SCID mouse, it is likely more clinically relevant to be able to test the metastatic lesions as these are the major cause for morbidity in these patients.

Although inhibition of mTOR in FL-HCC has been proposed as a possible treatment in the past, there is limited evidence supporting its efficacy (17). Riehle et al. reported increased expression of P-S6, a downstream target of rapamycin complex 1 (mTORC1), in all 13 cases of FL-HCC that were reviewed compared to just 13% in non-FL-HCC (18). These primary tumors additionally showed relatively low levels of AKT activation, suggesting that the dysregulation of mTOR is the driving force, rather than a result of AKT activity (18). We also found very limited AKT activity in our primary patient samples as well as our patient-derived xenograft samples (data not shown). Cornella et al. also reported increased P-S6 in approximately 50% of 78 FL-HCC tumors analyzed (19). A case report described a significant treatment response to everolimus, a mTOR inhibitor, in a single patient with end-stage FL-HCC (20). Another small study reported one patient with FL-HCC that experienced stable disease for at least 7 months after treatment with sirolimus, also a mTOR inhibitor (21). On the other hand, a randomized trial in patients with unresectable or metastatic FL-HCC with 28 patients where they were randomized to everolimus, estrogen deprivation therapy, or both, showed no improved outcomes in any group (22). This study suggests that unselected patients likely do not benefit from mTOR inhibition, though the benefit for patients with confirmed mTOR upregulation, like our two young patients, is yet unknown. Our study adds to the evidence that mTOR inhibition in FL-HCC, particularly metastatic lesions, may be a valuable option in the overall treatment scheme for FL-HCC in those with altered mTOR pathways.

Treatment with temsirolimus resulted in minimal weight loss and no additional observed toxicities. Toxicities commonly observed in patients undergoing temsirolimus treatment include hematological toxicities like anemia and thrombocytopenia, hyperglycemia, and stomatitis (23, 24). On the other hand, treatment with FOLFIRINOX resulted in significant weight loss and a mouse death during the course of the treatment trial, though it was also associated with a significant decrease in tumor volume. The FOLFIRINOX regimen was tested as this was a potential treatment option being considered for one of the patients, though there have been no reports of its use in the treatment of FL-HCC. Gem/Ox has been associated with a significant treatment effect in a small number of case reports (25, 26), though the first patient in this study had substantial disease progression while on this regimen. Treatment with Gem/Ox in vitro showed a significant response and there appeared to be some tumor growth inhibition in the first two weeks of the treatment trial, but the effect was not sustained and the rate of tumor growth in the Gem/Ox group was similar to that of the vehicle-treated group (Figure 6). This direct correlation to the patient’s own clinical response emphasizes the utility of PDX models.

The specificity of the DNAJB1-PRKACA fusion protein for FL-HCC makes it of interest for drug targeting (27). It is crucial for tumor formation and is likely the driving force in the development of FL-HCC (28). This fusion protein is believed to allow PRKACA activity to exceed constraint from protein kinase A regulatory subunits (3). This dysregulated PKA is thought to lead to tumorigenesis in FL-HCC in a cAMP dependent fashion (29). Treatment with protein kinase inhibitors may be able to target this directly, though this has not yet been done clinically (30). Aurora Kinase A and ErbB2 have also been observed to be overexpressed which may provide other targeted treatment options (31).

Limitations of this study include the limited number of viable in vivo models which is partly due to the rarity of the tumor and difficulty in accessing primary patient tissue. Additionally, the patient primary tumor did not grow in vivo so we were not able to directly compare treatment regimens, though we were able to show differing treatment responses in vitro. The treatment study was also limited in duration due to large tumor size, which led to a study endpoint in control animals that were not receiving treatment. Therefore, the sustained ability of temsirolimus to be efficacious over an extended period of time could not be determined.

Conclusion

In conclusion, our findings suggest that mTOR inhibition may be an effective therapeutic in the treatment of metastatic FL-HCC. Our results show increased mTOR activity in metastatic FL-HCC lesions when compared to primary liver lesions with in vitro and in vivo treatment efficacy with temsirolimus administration. Additional studies assessing the clinical application of mTOR inhibition in metastatic FL-HCC are warranted.

Footnotes

  • Authors’ Contributions

    Conceived and designed study: Jennifer L. Leiting, Matthew C. Hernandez, Mark J. Truty. Collected data: Jennifer L. Leiting, Matthew C. Hernandez, John R. Bergquist, Jennifer A. Yonkus, Amro M. Abdelrahman. Analysis and interpretation of data: Jennifer L. Leiting, Matthew C. Hernandez, John R. Bergquist, Jennifer A. Yonkus, Amro M. Abdelrahman, Michael S. Torbenson, Nguyen H. Tran, Thorvardur R. Halfdanarson, Rondell P. Graham, Rory L. Smoot, Mark J. Truty. Drafting and critically revising: Jennifer L. Leiting, Matthew C. Hernandez, John R. Bergquist, Jennifer A. Yonkus, Amro M. Abdelrahman, Michael S. Torbenson, Nguyen H. Tran, Thorvardur R. Halfdanarson, Rondell P. Graham, Rory L. Smoot, Mark J. Truty.

  • Conflicts of Interest

    The Authors have no conflicts of interest to declare.

  • Received April 23, 2023.
  • Revision received June 14, 2023.
  • Accepted June 28, 2023.

This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY-NC-ND) 4.0 international license (https://creativecommons.org/licenses/by-nc-nd/4.0).

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Therapeutic Efficacy of Temsirolimus in a Patient-derived Model of Metastatic Fibrolamellar Hepatocellular Carcinoma
JENNIFER L. LEITING, MATTHEW C. HERNANDEZ, JOHN R. BERGQUIST, JENNIFER A. YONKUS, AMRO M. ABDELRAHMAN, MICHAEL S. TORBENSON, NGUYEN H. TRAN, THORVARDUR R. HALFDANARSON, RONDELL P. GRAHAM, RORY L. SMOOT, MARK J. TRUTY
In Vivo Sep 2023, 37 (5) 1940-1950; DOI: 10.21873/invivo.13290
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