Two major theories have been proposed to describe the stem cells in cancer: In the stochastic model, every tumor cell has the capacity for tumor initiation and growth, and depends on some random and varying intrinsic factor. In the hierarchical model, CSCs make up only a very small subpopulation of cells within the whole tumor. Uniquely, these cells have a greater capacity for self-renewal, differentiation, and tumorigenicity, and may be more resistant to chemotherapy and radiotherapy.

The CSC hypothesis has received support, since it can explain, in part, the heterogenous nature of hepatocellular carcinoma (HCC), and is one potential mechanistic explanation for its high degree of relapse, metastasis, and resistance to therapies.

Identification of Liver CSCs and Their Role in HCC

Attempts at identifying liver CSC (LCSC) markers in established cell lines have shown a high degree of heterogeneity in expression patterns. Given the variability, it is unlikely that a single, universal LCSC marker for HCC exists. Many cell surface antigens have been identified in LCSCs including CD24, CD44, CD90, CD133, and the epithelial cell adhesion molecule (EpCAM; CD326).

While HCC screening and diagnosis is dependent on imaging and serum alpha-fetoprotein (AFP) levels, their use is limited in some contexts, such as with disease recurrence and postoperative detection. Expression of progenitor stem cell markers may serve as alternative indicators of disease assessment and prognosis.

CD24 is associated with early-onset HCC and one mutation, P170T/T, is more common in patients with hepatitis B virus (HBV) than in those without infection. CD44 expression is associated with recurrence after local ablation therapy and metastasis. Tumors that co-express CD44 with CD90 and CD133 are more aggressive than tumors that express CD90 or CD133 alone. Similar to CD24, CD90 expression correlates with HBV-associated HCC and is linked to poor prognosis. In comparison with patients with CD133-negative tumors, patients with CD133-positive tumors have a shorter overall survival (OS) and higher recurrence rate. EpCam-positive, AFP-positive HCCs are associated with poor prognosis and a high metastatic rate.

The search for novel CSC markers in HCC is ongoing. The identification of a novel marker that can be easily detected by blood is especially of interest. In a recent study, transcription factor sex-determining region Y-box 9 (SOX9) was identified as a novel CSC marker in HCC, according to results published in Scientific Reports by Takayuki Kawai and colleagues3 from Kyoto University, Japan.

SOX9 plays a vital role in the development of many different tissues and organs, including the heart, lungs, testes, pancreas, and central nervous system. During development, SOX9 helps to keep cells in an undifferentiated state, and is regulated by various cell signaling pathways such as Wnt/beta-catenin, Notch, and transforming growth factor beta (TGFb)/Smad. As with many of the other LCSC markers, SOX9 expression is activated during organogenesis, but then normally switched off in adult hepatocytes. Kawai et al3 sought to determine whether aberrant SOX9 expression was an indicator of CSC in human HCC.

Using flow cytometry, investigators identified greater proportions of SOX9+ cells than SOX9- cells from multiple human HCC cell lines, including Huh7, HLF, PLC/PRF/5, and Hep3B. In Huh7 cells, those that expressed SOX9 demonstrated higher proliferative activity, colony formation, and sphere formation, which are all indicative of a tumorigenic phenotype. Additionally, these cells were more resistant to chemotherapy using 5-fluorouracil (5-FU), with resistance being a hallmark property of CSCs.

SOX9 expression also was linked to epithelial-mesenchymal transition (EMT), another hallmark of tumorigenesis. Epithelial-mesenchymal transition is regulated, in part, by the TGFb/Smad signaling pathway. Compared with the SOX9- population, SOX9+ cells showed greater and more efficient TGFb/Smad signaling as well as greater cell motility. Furthermore, SOX9 activated the Wnt/beta-catenin pathway, which plays a critical role in CSC maintenance.

Additional analysis determined that osteopontin (OPN), one downstream target of Wnt/beta-catenin, was regulated by SOX9 in vitro. Importantly, OPN was highly expressed in the serum of patients with SOX9+ HCC, indicating that OPN could serve as a surrogate marker for SOX9. Furthermore, OPN was a more sensitive indicator of SOX9 expression in primary tumors and was superior to the conventional markers, AFP and protein induced by vitamin K antagonists-II (PIVKA-II). Immunohistochemical analysis of HCC tissues showed that patients with SOX9+ HCC had greater serum OPN levels, greater venous invasion, and poorer recurrence-free survival (RFS). Serum OPN was significantly correlated with regression-free survival.

Other circulating biomarkers have been identified. A 2015 study by Robin Kelley and colleagues4 with the Helen Diller Family Comprehensive Cancer Center, University of California, San Francisco, detected CTCs from whole blood by EpCAM enrichment in metastatic HCC. In the pilot study of 20 patients with HCC, CTC detection was associated with elevated AFP levels (&ge;400 ng/mL) and vascular invasion. Next-generation sequencing of DNA revealed a number of known HCC mutations.

&ldquo;The ability to detect and characterize malignant cells in circulation holds enormous promise as a biomarker in HCC, a grim cancer challenged by the inability of conventional noninvasive diagnostic and staging modalities to encompass its great clinical and biological heterogeneity, as well as by a scarcity of tumor tissue available for diagnostic or research purposes,&rdquo; the authors reported. &ldquo;With increasingly sensitive and precise technologies for the detection and molecular profiling of rare cells, the genomic interrogation of CTCs may offer a powerful new tool to characterize, and someday to target, the dominant tumor subclones responsible for treatment resistance or metastatic progression.&rdquo;

Others have examined the presence of LCSC markers in blood and their correlation with HCC prognosis. In one study by Fan et al,5 patients with >0.01% circulating CD45+CD90+CD44+ LCSCs had a lower 2-year RFS rate (23% vs 64%) and OS rate (59% vs 94%) than patients with <0.01% circulating LCSCs.

Isolation of CTCs from circulation is critically important. Since taking tumor biopsies is not the standard of care for HCC, serum analysis can function as a noninvasive method for determining the presence of metastatic tumor tissue in real time and allow for assessment of molecular profiling using a &ldquo;liquid biopsy.&rdquo; This is particularly vital in HCC, given the associated risks with tumor biopsy and the acceptance of radiographic diagnosis in the absence of tissue confirmation, which also contributes to a lack of untreated tumor samples for research efforts.

Despite the large degree of marker heterogeneity among HCCs, therapeutic targeting of LCSC surface markers and signaling pathways that regulate their expression could be an efficacious approach, given their role in tumor progression, recurrence, and metastasis. Although mostly in preclinical development, some studies have shown promise for strategies that target CSCs.

Therapeutic Strategies Targeting CSCs in HCC

CSCs are inherently resistant to chemotherapy and ionizing radiation, enhancing the need for alternative and novel approaches that interfere with the self-renewal and survival characteristics of CSCs. Such therapies include antibody therapy, epigenetic therapy, and molecular-targeted therapy.

Antibody therapy using monoclonal antibodies (mAbs) against CSC-specific antigens has shown some efficacy in vitro and in vivo. In a mouse xenograft model, the combination of a CD13 antibody bound to 5-FU significantly reduced tumor volume compared with single treatment with either agent.6 A human CD133 mAb conjugated to monomethyl auristatin F (MMAF) inhibited the growth of hepatocellular and gastric cancer cell lines in vitro and in a xenograft model.

Two investigational compounds, napabucasin (BBI-608) and amcasertib (BBI-503), have emerged as potential CSC inhibitors and are currently being investigated in clinical trials. Napabucasin is a cancer stemness inhibitor that was developed to inhibit CSC signaling by targeting the STAT3 pathway. Amcasertib is an inhibitor of multiple serine-threonine stemness kinases, which inhibit Nano and other cancer stemness pathways.8 Stemness is a term that was initially used to describe stem cell gene expression, but is now used to express characteristics shared by both embryonic and adult stem cells.

Stemness is measured by a cell&rsquo;s ability to form spheres in culture. Hypermalignant CSCs, or stemness-high CSCs, are highly tumorigenic and metastatic, and are resistant to conventional chemotherapy and radiation. These traditional treatment modalities have been shown to induce stemness genes in cancers, having the negative effective of converting stemness-low CSCs to stemness-high CSCs. Some hypothesize that this is a significant driving factor of relapse in HCC.

Stemness-high CSCs have traditionally been difficult to target effectively due to increased DNA repair ability, overexpression of drug efflux pumps, and the activation of prosurvival and antiapoptotic signaling pathways. An early preclinical study demonstrated that napabucasin inhibited sphere development in stemness-high CSCs and downregulated their stemness gene expression. Additionally, napabucasin inhibited metastasis in a spontaneous liver metastasis model of colorectal cancer.

Ongoing clinical trials will continue to investigate napabucasin and amcasertib in HCC and other tumor types. In a phase I/II open label, 3-arm, dose-escalation study, napabucasin or BBI-503 in combination with sorafenib will be evaluated in patients with advanced HCC who have not received prior systemic chemotherapy (NCT02279719). A phase II, open-label, multicenter study will investigate amcasertib in patients with advanced hepatobiliary cancer, including HCC and cholangiocarcinoma, who have exhausted all currently approved standard anticancer therapies (NCT02232633). Lastly, in a phase I open-label study, napabucasin in combination with sorafenib will be investigated in Japanese patients with advanced HCC (NCT02358395).

Napabucasin and amcasertib are being intensely investigated in several other tumor types. The results of multiple early clinical trials were recently presented at the 2016 ASCO Annual Meeting. The results of these trials have been promising.

&ldquo;The napabucasin data at ASCO this year are exciting because they underscore the potential of cancer stem cell pathway inhibition across multiple tumor types,&rdquo; commented Tanios Bekaii-Saab, MD, FACP, co-leader of the Gastrointestinal Cancer Program at the Mayo Clinic Cancer Center. &ldquo;Additionally, these data demonstrate the potential to combine napabucasin with current standards of care to address unmet needs, including treatment in patients who have had multiple lines of prior therapy.&rdquo;

With regard to the expansion of clinical studies with the stemness inhibitor, Chiang J. Li, MD, FACP, President, CEO, and Chief Medical Officer of Boston Biomedical, and the head of Global Oncology for Sumitomo Dainippon Pharma Group, said in a press release, &ldquo;These studies continue to show napabucasin&rsquo;s safety and early efficacy across doses and in combination with a variety of established agents. We plan to apply these findings as we advance and expand our clinical development program for this first-in-category cancer stemness inhibitor.&rdquo;

Other therapies are also showing promise. In a recent study, Ogawa et al11 utilized VB4-845, an agent consisting of an anti-EpCAM antibody linked to Pseudomonas aeruginosa Exotoxin A. In EpCAM-expressing HCC cell lines, VB4-845 suppressed sphere formation.

Additionally, in combination with 5-FU, VB4-845 significantly reduced tumor growth in a xenograft model. Further studies in preclinical and clinical trials are needed to validate the clinical utility of this approach.

Additional novel alternative strategies involving CSCs have hit some roadblocks. Umbilical cord mesenchymal stem cells (UCMSCs) initially held promise as novel delivery vehicles for therapeutic agents. However, in a recent report, Chang Liu et al12 unveiled potential pro- and antitumorigenic properties of UCMSCs.

Following the birth of a child, UCMSCs can be readily collected and cultured from the umbilical cord. Uniquely, this type of stem cell exhibits a high proliferation rate, wide multipotency, and hypoimmunogenicity.

Generally, the capacity of MSCs to migrate to sites of inflammation or injury is one characteristic that makes them especially attractive as drug carriers. However, studies have demonstrated that both oncogenic and tumor-suppressive properties are present in MSCs, and depend highly on the stem cell origin and tumor model used.

While most in vitro studies of UCMSCs have used 2-dimensional tissue culture models, Liu and colleagues used a 3-dimensional, co-culture system of UCMSCs and HCC cells to more closely mimic in vivo conditions. Cell growth, CSC characteristics, drug resistance, and metastasis were evaluated.

Co-culture of the HCC cell line, HCCLM3, with UCMSCs enhanced many tumorigenic properties. These included an upregulation of matrix metalloproteinase (MMP) and EMT-related genes (N-cadherin and vimentin), as well as increased migration capacity. These characteristics were reversed in the presence of the TGFb receptor inhibitor SB431542, indicating the importance of this pathway in EMT and HCC metastasis.

To conclude, HCC is a heterogenous disease that is difficult to treat with traditional therapies. The cancer stem cell hypothesis is one explanation for therapy resistance observed in advanced disease. Advances in detection methods have identified several markers of liver CSCs and markers that are suitable for noninvasive, liquid biopsy. Ongoing clinical trials will assess inhibitors to stemness factors in combination with sorafenib in patients with advanced HCC. Although UCMSCs have potential as drug carriers, studies have indicated that they exhibit both pro- and antitumorigenic properties, warranting further investigation.

&ldquo;Understanding the mechanisms that underlie CSC maintenance to combat cancer is essential,&rdquo; commented Matilde E. Lleonart, PhD, of the Vall d'Hebron Research Institute in Barcelona, Spain. &ldquo;Determining the mechanisms that govern radio/chemoresistance in CSCs may illuminate the origins of this complex process and improve the treatments of cancer.&rdquo;

Lui SK, Vilchez V, Gedaly R. Liver cancer stem cells: a new paradigm for hepatocellular carcinoma treatment. J Stem Cell Res Ther. 2015;5:283.

Chiba T, Iwama A, Yokosuka O. Cancer stem cells in hepatocellular carcinoma: therapeutic implications based on stem cell biology. Hepatol Res. 2016;46(1):50-57.

Kawai T, Yasuchika K, Ishii T, et al. SOX9 is a novel cancer stem cell marker surrogated by osteopontin in human hepatocellular carcinoma. Sci Rep. 2016;6:30489.

Kelley RK, Magbanua MJ, Butler TM, et al. Circulating tumor cells in hepatocellular carcinoma: a pilot study of detection, enumeration, and next-generation sequencing in cases and controls. BMC Cancer. 2015;15:206.

Fan ST, Yang ZF, Ho DW, et al. Prediction of posthepatectomy recurrence of hepatocellular carcinoma by circulating cancer stem cells: a prospective study. Ann Surg. 2011;254(4):569-576.

Haraguchi N, Ishii H, Mimori K, et al. CD13 is a therapeutic target in human liver cancer stem cells. J Clin Invest. 2010;120(9):3326-3339.

Smith LM, Nesterova A, Ryan MC, et al. CD133/prominin-1 is a potential therapeutic target for antibody-drug conjugates in hepatocellular and gastric cancers. Br J Cancer. 2008;99(1):100-109.

Boston Biomedical Featured Seven Clinical Posters on Cancer Stem Cell Pathway Inhibitors Napabucasin and Amcasertib in Multiple Cancer Types at ASCO 2016. Cambridge, MA: Boston Biomedical. June 2016. http://www.bostonbiomedical.com/boston-biomedical-featured-seven-clinical-posters-cancer-stem-cell-pathway-inhibitors-napabucasin-amcasertib-multiple-cancer-types-asco-2016/#_edn 1. Accessed August 12, 2016.

Li Y, Rogoff HA, Keates S, et al. Suppression of cancer relapse and metastasis by inhibiting cancer stemness. Proc Natl Acad Sci. 2015;112(6):1839-1844.

Boston Biomedical&rsquo;s Investigational Cancer Stem Cell Pathway Inhibitor, Napabucasin (BBI608), Featured at the ASCO 2016 Gastrointestinal Cancers Symposium [press release]. Cambridge, MA: Boston Biomedical. January 25, 2016. http://www.bostonbiomedical.com/boston-biomedicals-investigational-cancer-stem-cell-pathway-inhibitor-napabucasin-bbi608-featured-at-the-asco-2016-gastrointestinal-cancers-symposium. Accessed August 20, 2016.

Ogawa K, Tanaka S, Matsumura S, et al. EpCAM-targeted therapy for human hepatocellular carcinoma. Ann Surg Oncol. 2014;21(4):1314-1322.

Liu C, Liu Y, Xu XX, et al. Mesenchymal stem cells enhance the metastasis of 3D-cultured hepatocellular carcinoma cells. BMC Cancer. 2016;16:566.

Fong CY, Chak LL, Biswas A, et al. Human Wharton&rsquo;s jelly stem cells have unique transcriptome profiles compared to human embryonic stem cells and other mesenchymal stem cells. Stem Cell Rev. 2011;7(1):1-16.

Bergfeld SA, DeClerck YA. Bone marrow-derived mesenchymal stem cells and the tumor microenvironment. Cancer Metastasis Rev. 2010;29(2):249-261.

Articles

Clinical Articles

Technological advances and greater understanding of stem cell biology have contributed to the identification and characterization of the tissues and organs involved with these cells.

Cancer Stem Cell Therapies on the Horizon for HCC

Several types of immune cells are present in the liver, including macrophages, natural killer (NK) cells, natural killer T cells (NKT cells), and CD4+ and CD8+ T cells. These immune cells not only regulate liver homeostasis and help to fight microorganisms but also maintain liver regeneration through the growth of hepatic progenitor cells and hepatocytes.

As with many other tumors, inflammation is also a key contributor to carcinogenesis for those with HCC. Continuous inflammation and the corresponding proliferative stimuli not only promote liver fibrosis and cirrhosis, but also the transformation of hepatocytes to a malignant phenotype.

Studies conducted on in vivo models have found that lymphocytes are critical regulators of hepatocyte damage, liver fibrosis, and tumor development, according to findings reported by Jessica Endig, PhD, with the Hannover Medical School in Hannover, Germany, and colleagues in Cancer Cell.

For this study, the investigators utilized a fumarylacetoacetate hydrolase (Fah)-deficient murine model, which reliably mimics the inflammatory environment of chronic liver disease. Liver injury was induced with fumarylacetoacetate (FAA), a toxic metabolite that accumulates with Fah deficiency. This model has been used extensively to delineate hepatocellular carcinogenesis and regeneration.

Repeated flares of liver injury with FAA resulted in strong increases in CD3+ CD4+ and CD3+ CD8+ T cells. In contrast, treatment with the drug NTBC (2-[2-nitro-4-(trifluoromethyl)benzoyl] cyclohexane-1,3-dione, or Nitisinone), which blocks FAA formation and prevents liver injury in Fah-deficient mice, abrogated this effect. Chronic, long-term FAA-induced liver injury in Fah-/- mice resulted in progressive biliary fibrosis.

Generation of mice lacking mature B and T lymphocytes or CD8+ T and NKT cells resulted in significantly decreased median overall survival (OS; 18.5 days and 20 days vs 31 days for Fah-/- mice), indicating that CD8+ T cells protect Fah-/- mice from acute-on-chronic liver failure.

Importantly, blockage of lymphotoxin-beta (LT&beta;), a member of the tumor necrosis family, using a lymphocyte-beta receptor (LT&beta;R) immunoglobulin fusion protein resulted in significantly suppressed, FAA-induced tumor formation compared with controls (50% vs 8.3%; P &le;.05) and reduced tumor count (2.0 vs 1.0) in Fah-/- mice.

Alternating liver injury flares with recovery and regeneration, a process that mimics human liver diseases, produced multifocal HCCs in Fah-/- mice within 4 months. Array comparative genomic hybridization (aCGH) analysis of tumors showed elevated chromosomal aberrations that reflected increased genomic instability, and were consistent with chromosomal gains and losses in human HCCs.

To identify important gene expression changes associated with the immune response, microarray analysis was performed. In comparison with controls, the most significant expression changes in Fah-/- mice were associated with the cell cycle.

&ldquo;Our results reveal that lymphocytes significantly contribute to liver damage and hepatocarcinogenesis, but also protect mice from acute-on-chronic liver failure and support liver progenitor cell proliferation during chronic liver injury,&rdquo; the authors concluded. &ldquo;These findings emphasize the fundamental requirement that the immune system needs to be tightly regulated in a context-specific fashion to balance immune surveillance and cancer risk. Moreover, inhibiting LT&beta;R signaling might be an interesting approach to prevent tumor development in patients with chronic liver diseases accompanied by high levels of LT&beta;.&rdquo;

A Novel Immunomodulatory Effect of Sorafenib

One hallmark of the tumorigenic phenotype is the expression of immune checkpoints, which typically stop an immune response after antigen stimulation. The immune checkpoint receptor programmed death-1 (PD-1) is highly expressed on T cells with chronic stimulation, and functions to downregulate the antitumor T-cell response when stimulated by its ligand PD-L1. This mechanism is beginning to be explored more thoroughly in HCC, using direct inhibitors and available therapies.

In a recent report, Suresh Kalathil, PhD, and coauthors with the Roswell Park Cancer Institute, Buffalo, NY, showed for the first time that sorafenib (Nexavar) has an immunomodulatory effect that reduces the number of PD-1+ T cells and regulatory T cells (Tregs), resulting in a survival benefit. The results were published in JCI Insight.

&ldquo;From the literature, we knew that both immune cells and tumor cells expressed the VEGF receptor, which is a target of sorafenib,&rdquo; Yasmin Thanavala, PhD, the senior author and professor of Oncology, described. &ldquo;We then rationalized there is the possibility that sorafenib could also modulate immune cells via the same VEGF receptor. That is the question that we addressed.&rdquo;

Re-activating the immune response has been of much interest in a number of tumor types, and novel antibody therapies have targeted the immune checkpoint pathways that inhibit T cells.4,5 Although the impact of PD-1 inhibition in HCC was not clearly understood, studies have indicated that the antitumor T-cell response is significantly impaired in advanced HCC.

In the JCI Insight analysis, blood samples from 19 patients with HCC were obtained before treatment initiation and after 4 weeks of sorafenib. The frequency and phenotype of PD-1+ T cells, Tregs, and myeloid-derived suppressor cells (MDSCs) were analyzed by flow cytometry.

Following sorafenib, a reduction in the absolute number of CD4+PD-1+ T cells and CD8+PD-1+ T cells was observed. Reductions in these cell types after therapy correlated with better OS compared with a lesser decline. Similarly, patients with greater numbers of CD4+PD-1+ T cells and CD8+PD-1+ T cells prior to sorafenib treatment also showed greater improvements in OS compared with patients who had lower numbers of these phenotypes, suggesting that patients with higher levels of PD-1+ T cells were more responsive to therapy.

In addition to a decrease in the number of T cells, the frequency and absolute number of Foxp3+ Tregs was significantly decreased, while the ratio of CD4+CD127+PD-1 T effector cells to CD4+Foxp3+PD-1+ Tregs was significantly increased after sorafenib therapy. The decrease in Foxp3+ Tregs also correlated with a significant improvement in OS.

In vitro analysis demonstrated a reduction in frequency of CD4+CD127+PD-1+ T cells, with a concomitant increase in the frequency of CD4+CD127+PD-1 T cells with sorafenib, indicating that the sorafenib benefit was due to specific downregulation of PD-1 expression on CD4+ T cells rather than nonspecific cytotoxicity.

Ultimately, the results indicate that PD-1+ T cells and Tregs may act as predictive biomarkers, and could be used to identify patients who may benefit the most from sorafenib.

&ldquo;Checkpoint inhibitors bring down the level of immunosuppression and allow the patient&rsquo;s own immune system to either target the tumor or make a more favorable environment for other kinds of immunotherapy, such as vaccination,&rdquo; said Thanavala. &ldquo;If you have a therapy that reduces immunosuppression, such as sorafenib, and bring in another therapy that activates the patient&rsquo;s immune system, then patients may experience a greater therapeutic benefit.&rdquo;

Ongoing clinical trials are evaluating the antiPD-1 antibody nivolumab (Opdivo) in patients with HCC. Although approved by the FDA for a number of tumor types, including unresectable or metastatic melanoma, metastatic non&ndash;small cell lung cancer, and advanced renal cell carcinoma, nivolumab has not been extensively studied in HCC. Nivolumab blocks the interaction between PD-1 and its ligand PD-L1, enhancing the antitumor immune response.

Early clinical trials of nivolumab in HCC have been promising. In an interim analysis of the phase I/II, dose-expansion CheckMate-040 study, 206 patients with Child-Pugh class A HCC were evaluated. Bruno Sangro, MD, PhD, from the Clinica Universitaria de Navarra in Spain and colleagues presented findings from the dose escalation cohorts at the 2016 ASCO Annual Meeting.

Patient cohorts consisted of: (1) those who were uninfected sorafenib-na&iuml;ve/intolerant; (2) those who were uninfected sorafenib progressors; (3) those with hepatitis C virus (HCV); and (4) those with hepatitis B virus (HBV), with dose expansion of nivolumab at 3 mg/kg. Over half of patients (64%) had prior sorafenib treatment, 75% had extrahepatic metastasis, and 7% had vascular invasion. The primary endpoint was confirmed overall response rate (ORR) by RECIST v1.1 criteria, and secondary endpoints were OS, progression-free survival (PFS), time to progression, and biomarker assessments.

Of the 174 evaluable patients, 39% had a reduction in tumor burden. In a preliminary analysis, 55% had at least 18 weeks of follow-up and/or progressive disease. Among those patients, the ORR was 9% across etiologies and the 6-month OS rate was 69%. Nivolumab response was seen in patients with and without quantifiable PD-L1 expression as measured by immunohistochemistry.

Patients received a median of 5 to 6 doses of nivolumab (range, 1-19). Treatment-related adverse events (trAEs) occurred in half of patients, with the most frequent being fatigue (17%) and pruritus (12%). Grade 3/4 trAEs were observed in 14% of patients, and increased alanine aminotransferase (ALT) and aspartate aminotransferase (AST) levels were the most common (3% each).

A further analysis of patients from the CheckMate-040 study with 3 years of follow-up was also presented at the 2016 ASCO Annual Meeting by Anthony El-Khoueiry, MD, phase I program director at the University of Southern California Norris Comprehensive Cancer Center, and colleagues.

Eligible patients had a Child-Pugh Score &le;7 and had previously failed or were intolerant of sorafenib. Nivolumab dosing ranged from 0.1 to 10 mg/kg for up to 2 years. The primary endpoint was safety, and secondary endpoints were antitumor activity and duration of response. Among the 51 patients who were enrolled and treated with nivolumab, 73% had prior sorafenib therapy, 76% had extrahepatic metastasis, and 12% had vascular invasion. Most discontinuations (35/41) were due to progressive disease.

Nivolumab treatment exhibited durable responses and disease stabilization across all dosages and cohorts. The median duration of response was 23.7 months, and median OS was 15.1 months. The OS rates at 6, 9, 12, and 18 months were 67%, 67%, 59%, and 48%, respectively. The complete response (CR) and partial response (PR) rates were 6% and 8%, respectively. Half of patients achieved stable disease.

Nivolumab exhibited a manageable safety profile. Treatment-related AEs occurred in 77% of patients, most commonly rash and AST increase (20% each). Grade 3/4 trAEs occurred in 20% of patients, most commonly increases in AST (10%), lipase (6%), or ALT (6%). The maximum tolerated dose was not reached.

&ldquo;Hepatocellular carcinoma is an aggressive and fatal cancer, comprising 90% of all liver cancer in adults worldwide with limited therapeutic options for patients with advanced stage disease; no treatment advances have been made for patients who fail to respond or progress on the current standard of care,&rdquo; said El-Khoueiry in a press release.

&ldquo;These preliminary data are encouraging and support the ongoing evaluation of nivolumab in this patient population, as they show promising preliminary survival data, and durable partial or complete response in one out of five nivolumab-treated patients, with many others experiencing stable disease.&rdquo;

A randomized phase III study, CheckMate-459, is currently recruiting participants and will evaluate nivolumab as a first-line therapy in patients with advanced HCC (NCT02576509). In the open-label study, the efficacy of nivolumab will be compared with sorafenib. Eligible patients have 1 or more lesions, Child-Pugh class A liver function, ECOG status of 0 or 1, and no prior systemic therapy.

Over 700 patients will be randomized in a 1:1 ratio to receive nivolumab or sorafenib until disease progression or unacceptable toxicity. The primary objectives are time to progression and OS. The secondary endpoints are ORR, PFS, and PD-L1 expression. Patient data will be stratified by vascular invasion and/or extrahepatic spread, etiology, and geography. The estimated primary completion date is July 2017.

Vaccine Therapy Under Exploration for HCC

Other novel approaches are also being explored that target immunoregulatory cells, with one such trial exploring a vaccine therapy in combination with sorafenib. In a case study reported by Yongxiang Yi et al in Molecular Clinical Oncology,

a patient with metastatic HCC achieved a CR after treatment with sorafenib, focused ultrasound knife therapy, and dendritic cell (DC) DRibbles vaccine.

DRibbles are autophagosomes containing a variety of antigens from antigen donor cells. They stimulate antigen-specific CD8 T cells ex vivo and induce a T-cell response against HCC in vivo. The case report described a 42-year-old male with HBV-associated HCC. The patient underwent curative resection, but in less than 2 months after surgery, multiple intrahepatic and extrahepatic lesions were detected, and the patient was treated with sorafenib. Because alpha-fetoprotein (AFP) levels continued to rise, focused ultrasound knife therapy was initiated, as was DC-DRibbles therapy.

Eight months after combination therapy, AFP levels returned to normal and remission of the liver and lung metastases was observed. Two years after the combination treatment, metastatic lesions were undetectable by computed tomography (CT) scan. No serious AEs were reported.

&ldquo;Our results demonstrated that antigen-specific T-cell response aimed to treat recurrent HCC may be induced through stimulation by the DC-DRibbles vaccine,&rdquo; the authors concluded. The case supports a combination strategy for patients with recurrent HCC after resection.

An ongoing, open-label, phase II trial will investigate the safety and efficacy of the individualized anticancer vaccine CRCL-AlloVax in approximately 30 patients with advanced HCC after prior sorafenib therapy (NCT02409524).

The novel AlloVax therapy is a vaccine consisting of chaperone-rich cell lysate (CRCL) antigens, derived from a patient&rsquo;s own tumor cells, in combination with AlloStim cells, which are activated allogenic Th1 memory T cells that contain cytolytic granules.

The primary outcome measure is survival compared with historical controls. The estimated primary completion date is July 2017.

In conclusion, recent studies support the vital importance of immunosuppression in HCC and key molecular components that contribute to hepatocarcinogenesis. While novel immunomodulatory therapies are on the horizon, sorafenib may reduce the immunosuppressive network in HCC. Further work is needed to fully capitalize on the immune system as an anticancer therapy for HCC.

Tumanov AV, Koroleva EP, Christiansen PA, et al. T cell-derived lymphotoxin regulates liver regeneration. Gastroenterology. 2009;136(2):694-704.e4.

Wolf MJ, Adili A, Piotrowitz K, et al. Metabolic activation of intrahepatic CD8+ T cells and NKT cells causes nonalcoholic steatohepatitis and liver cancer via cross-talk with hepatocytes. Cancer Cell. 2014;26(4):549-564.

Endig J, Buitrago-Molina LE, Marhenke S, et al. Dual role of the adaptive immune system in liver injury and hepatocellular carcinoma development. Cancer Cell. 2016;30(2):308-323.

Topalian SL, Hodi FS, Brahmer JR, et al. Safety, activity, and immune correlates of anti-PD-1 antibody in cancer. N Engl J Med. 2012;366(26):2443-2454.

Hamid O, Robert C, Daud A, et al. Safety and tumor responses with lambrolizumab (anti-PD-1) in melanoma. N Engl J Med. 2013;369(2):134-144.

El-Khoueiry A, Sangro B, Yau T, et al. Phase I/II safety and antitumor activity of nivolumab (nivo) in patients (pts) with advanced hepatocellular carcinoma (HCC): Interim analysis of the CheckMate-040 dose escalation study. J Clin Oncol. 2016;34(suppl; abstr 4012).

Bristol-Meyers Squibb. Phase I/II Opdivo (nivolumab) Trial Shows Bristol-Myers Squibb&rsquo;s PD-1 Immune Checkpoint Inhibitor is First to Demonstrate Anti-Tumor Activity In Patients With Hepatocellular Carcinoma [press release]. May 29, 2015.http://news.bms.com/press-release/phase-iii-opdivo-nivolumab-trial-shows-bristol-myers-squibbs-pd-1-immune-checkpoint-in. Accessed August 21, 2016.

Yi Y, Han J, Fang Y, et al. Sorafenib and a novel immune therapy in lung metastasis from hepatocellular carcinoma following hepatectomy: a case report. Mol Clin Oncol. 2016;5(2):337-341.

Immunovative Therapies Ltd. http://www.immunovative.com. Accessed August 15, 2016.

The immune system performs a complex role in the development and growth of hepatocellular carcinoma (HCC), according to evidence from preclinical and clinical studies that are beginning to unravel the cellular components that link cytokines and inflammation with hepatocarcinogenesis.

New Evidence Supports a Key Role of the Immune System in HCC

In a recent retrospective study of patients with inoperable, nonmetastatic HCC, 1- and 2-year freedom from local progression (FFLP) rates were greater with SBRT versus radiofrequency ablation (RFA). The results of the study were reported in the

by Daniel Wahl, MD, PhD, and colleagues with the University of Michigan Medical Center.

Prior to this publication, data comparing SBRT and RFA were lacking.

While surgical resection is the preferred first-line therapeutic intervention for patients with a single HCC lesion and preserved liver function, many patients with multifocal disease, poor liver function, or major vascular invasion need to be managed with other locoregional therapy. These alternative therapeutic strategies may include RFA, SBRT, microwave ablation, percutaneous ethanol injection, or various embolization techniques.

Previous reports have shown similar local recurrence rates between SBRT and RFA, ranging from 10% to 30% for small tumors.

However, specific outcomes and treatment failure in various patient and tumor populations, especially those with large tumors, was unknown. With retrospective results in hand, the next step will be a prospective comparison of the two approaches, if it is deemed feasible.

Support of SBRT as a Frontline Therapy in HCC

In the study by Wahl et al, 249 lesions in 161 patients were treated with RFA, and 83 lesions in 63 patients were treated with SBRT. Compared with patients who had received SBRT, those treated with RFA had significantly lower alpha-fetoprotein levels, higher rates of cirrhosis, fewer previous liver-directed treatments, and longer follow-up. Median maximum tumor diameter was similar between treatment groups.

Freedom from local progression rates were greater for tumors treated with SBRT compared with those treated with RFA. The 1-year and 2-year FFLP rates were 97.4% versus 83.6% and 83.8% versus 80.2% for SBRT and RFA, respectively.

To account for any imbalances in treatment assignment, authors used an inverse probability of treatment weighting (IPTW) to the Kaplan-Meier and Cox models for FFLP. Using the IPTW univariate analysis, patients who received RFA had a significant association with local progression compared with SBRT (HR, 2.63

= .016). Among all of the variables analyzed, only tumor size was predictive of local progression (HR, 1.36

= .029). Larger tumor size was predictive of treatment failure with RFA but not with SBRT.

Further stratification of data was performed to identify additional differences between treatments. Although FFLP was similar in tumors smaller than 2 cm, significantly worse FFLP was associated with RFA (HR, 3.35; 95% Cl, 1.17-9.62,

.025). One- and 2-year overall survival (OS) rates were similar between treatment groups.

The rates of grade &ge;3 adverse events (AEs) were 11% for the RFA cohort and 5% for the SBRT cohort. Radiofrequency ablation complications included bleeding (n = 3), duodenal and colonic perforation (n = 2), sepsis (n = 2), and pneumothorax (n = 1). Stereotactic body radiation therapy complications included radiation-induced liver disease (RILD; n = 1), gastrointestinal bleeding (n = 1), and worsening ascites (n = 1). Radiofrequency ablation-associated complications resulted in 2 deaths, 1 from hemothorax and 1 from gastrointestinal bleeding, while no deaths resulted from SBRT.

&ldquo;These results suggest that both SBRT and RFA are excellent choices for smaller tumors but that SBRT may be preferred for larger tumors,&rdquo; the authors concluded. &ldquo;Prospective, randomized clinical trials are needed to compare these two modalities, especially for larger tumors, although we are unaware of any such ongoing trials.&rdquo;

Not everyone is in agreement regarding the frontline use of SBRT in HCC, despite the growing evidence of its clinical utility. In response to the retrospective study by Wahl et al, international clinicians have voiced their concerns over the endorsement of SBRT as the &ldquo;preferred treatment for larger HCC.&rdquo;

In one commentary, Berardino De Bari, MD, and colleagues, with Centre Hospitalier Universitaire Vaudois, Lausanne, Switzerland, have argued that although the authors have shown that SBRT is safe and efficient, the European Society for Medical Oncology and European Society of Digestive Oncology (EMSO-ESDO) and European Association for the Study of the Liver and European Organization for Research and Treatment of Cancer (EASL-EORTC) clinical practice guidelines largely support the use of radiotherapy (RT) as a palliative treatment option. Furthermore, De Bari and coauthors questioned the potential of some methodologies from the Wahl study that may have influenced the results.

De Bari et al pointed out that efficacy assessment within the retrospective study used RECIST criteria to determine the FFLP rate. They argued that others have shown that RECIST and modified RECIST (mRECIST) are not optimal for examining HCC response to SBRT, and that the evaluation of response or recurrence itself is challenging because of limitations in imaging technologies.

Additionally, De Bari and colleagues highlighted patient heterogeneity, particularly in follow-up duration, as one contributing influence on the results. Follow-up was 20 versus 13 months for RFA and SBRT, respectively, which could have influenced efficacy in terms of late relapses and/or late complete responses. The commentators suggested that more homogeneous patient cohorts and comparisons with radioembolization or transarterial chemoembolization are needed in future randomized trials.

In another commentary, Jen-Fu Yang, MD, and coauthors, with the Tri-Service General Hospital, Taipei, Taiwan, voiced 2 main concerns with the Wahl et al study.

First, Yang and colleagues pointed to the assessment of treatment response. Many ablative techniques, including RT and RFA, result in tumor necrosis rather than tumor shrinkage. As such, evaluation criteria that rely only on measurements of tumor size may not accurately reflect tumor response. While EASL and mRECIST criteria incorporate tumor necrosis into their evaluation guidelines, the original RECIST, as used by Wahl et al did not, and could have contributed to distorted data.

Second, Yang et al discussed the ambiguity surrounding the timing of liver transplantation in each treatment cohort. When performed before treatment, transplantation has the potential to significantly impact OS rates. The commentators suggested censorship of patients with subsequent liver transplantation from the time of the procedure onward for survival estimates. Similarly to De Bari et al, Yang and colleagues advocated for the need for future prospective randomized trial for SBRT.

Wahl and colleagues have issued a reply in response to the commentaries. They clarify that because their multidisciplinary board evaluates tumor growth as a part of disease progression, the original RECIST criteria most closely aligned with their clinical practice. Further, they note that RECIST is the standard assessment of the Radiation Therapy Oncology Group.

With regard to liver transplantation, Wahl et al reiterate that few patients underwent the procedure in the original study, and that pathologic complete response (pCR) rates were similar (SBRT 44% vs RFA 33%) and consistent with other published studies.

To address the need for additional studies, Wahl et al outline plans for a prospective clinical trial. &ldquo;We are in the process of opening a clinical trial comparing RFA with SBRT to test the results of this study in both larger and smaller HCCs,&rdquo; Wahl and colleagues stated. &ldquo;Given the difficulties of imaging-based treatment response for HCC, one of the goals of this trial will be to assess the pathologic treatment response of both modalities at the time of transplantation. This trial will also address the differences in follow-up between the two modalities seen in our current study and allow for the prospective determination of acute toxicity rates.&rdquo;

Additional Studies on the Use of SBRT in the Frontline Setting

Although met with some criticism, additional studies have supported the use of SBRT in inoperable or advanced HCC. Alexander Kirichenko, MD, from Allegheny General Hospital, Pittsburgh, PA, and colleagues found that SBRT alone or in combination with surgery was safe and effective for inoperable liver tumors. The investigators presented their findings at the 2016 ASCO Gastrointestinal Cancers Symposium.

In the retrospective study, patients with primary (n = 43) or isolated metastatic tumors (n = 48) received from 4 to 6 SBRT treatments. Local control was the primary endpoint of the study, and toxicity and survival were the secondary endpoints.

After a median follow-up of 20.3 months, the 2-year local control rates were 96% and 94% for primary and metastatic tumors, respectively. For tumors ˂4 cm, local control rate was 100%. Of the 14 patients that underwent liver transplant and who received SBRT as a bridging therapy, 10 patients had pCR. In 8 patients, liver resection was performed in conjunction with SBRT for unresectable tumors. After 2 years, the OS rates were 82% and 64% for primary and metastatic disease, respectively.

The toxicity with SBRT was minimal. No incidence of accelerated Child-Pugh class migration, grade &ge;3 AEs, or operative or perioperative complications were reported with SBRT.

&ldquo;Surgery remains the gold standard for treating liver cancer, but unfortunately more than 80% of patients with liver cancer are not eligible for surgery,&rdquo; said Kirichenko, lead author of the study. &ldquo;SBRT offers these patients a safe option that can stand on its own as a treatment or serve as an effective bridge to surgical removal of the tumor or a liver transplant.&rdquo;

Not only has SBRT been shown to be a viable option for those with inoperable tumors, but it also has shown promise for those with portal vein tumor thrombosis (PVTT) or inferior vena cava tumor thrombosis (IVCTT). In the

, Yoshiro Matsuo and colleagues with the Kobe University Graduate School of Medicine, Japan, reported that SBRT had favorable outcomes over 3-dimensional conformal RT (3DCRT) in patients with HCC with PVTT/IVCTT.

Portal vein tumor thrombosis and IVCTT are conditions that plague up to 40% of patients with HCC at the time of diagnosis. If left untreated, patients with PVTT/IVCTT have a median survival of just 2 to 4 months. Portal vein tumor thrombosis and IVCTT can lead to aggressive complications that worsen prognosis, including extensive intrahepatic tumor spreading, portal vein hypertension, and ischemic liver damage. Treatment of patients with PVTT/IVCTT has remained challenging, as most patients are unsuitable for surgery.

In the retrospective analysis, 43 patients were treated with SBRT (either with CyberKnife or TrueBeam), and 54 patients were treated with 3DCRT. In comparison with 3DCRT, rates of tumor response and OS were greater for total SBRT, CyberKnife, and TrueBeam while 1-year local progression rates were lower. Additionally, higher median biologically effective dose 10 (BED

) was achieved with SBRT compared with 3DCRT.

Studies have also explored outcomes for transarterial chemoembolization (TACE) plus RT, using various delivery methods, for patients with unresectable HCC. In one such analysis, TACE plus RT showed better clinical outcomes compared with TACE alone. Ya Huo, MD, and Guy Eslick, MD, with the University of Sydney, Australia, analyzed combination therapy versus TACE alone, and published their findings in

The meta-analysis of over 2500 patients from 25 clinical trials (11 randomized, controlled trials) looked for differences in survival, tumor response, and AEs using a random effects model. Radiotherapy techniques included in the analysis were 3DCRT, SBRT, and the moving strip technique for whole-liver irradiation.

Compared with TACE alone, patients who were treated with TACE plus RT had significantly better 1-year survival (odds ratio [OR], 1.36; 95% CI, 1.19-1.54) and complete response (OR, 2.73; 95% CI, 1.95-3.81).  There was a corresponding increase in benefits in the 2-, 3-, 4-, and 5-year survival rates (OR, 1.55, 1.91, 3.01, and 3.98, respectively). Gastroduodenal ulcers and elevated alanine aminotransferase and bilirubin levels were observed with a greater incidence in the combination group than in the TACE-alone group.

Importantly, the authors stress, &ldquo;RT for HCC has traditionally not been used because older techniques have not been able to adequately localize the radiation to the tumor. However, recent improvements in RT have allowed increased intratumor radiation and decreased radiation to the adjacent normal liver and organs, thereby reducing the rate of adverse events such as RILD.&rdquo;

Direct Comparisons of SBRT With Other Approaches

Additional studies have directly evaluated SBRT and TACE, the standard locoregional therapy for those with intermediate HCC. In one study, SBRT was found to be a safe alternative to TACE, demonstrating better local control. Eli Sapir, MD, with the Hadassah Hebrew University Medical Center, Jerusalem, Israel, and colleagues presented findings at the 2016 ASCO Annual Meeting for local control, OS, and toxicity between these approaches.

Patients with portal venous involvement, extrahepatic spread, or Child-Pugh class C liver function were excluded from the analysis because they were ineligible for TACE treatment. Outcomes were measured in an 8-year period in 84 patients with 114 lesions treated with TACE and 125 patients with 173 lesions treated with SBRT. The median follow-up was 28 months.

Although OS was not significantly different between treatment groups, 1- and 2-year local control rates both favored SBRT over TACE (97% vs 41% and 91% vs 18%, respectively; HR, 18.8; 95% CI, 6.7-52.7;

<.001). In TACE-treated patients but not SBRT-treated patients, factors that were predictive of worse local control included increasing tumor size and partial PVTT. Grade &ge;3 AEs occurred at a higher rate with TACE than with SBRT (14% vs 7%, respectively;

In the past, the use of RT was limited by potential detrimental AEs, including the risk of RILD and low tolerance in general. However, technological advances, such as with SBRT, allow for higher and more targeted radiation doses to the tumor site, while sparing adjacent tissue and organs for substantial radiation doses. Although met with some criticism, ongoing clinical trials will address the potential of SBRT as the preferred treatment for large HCC lesions.

Wahl DR, Stenmark MH, Tao Y, et al. Outcomes after stereotactic body radiotherapy or radiofrequency ablation for hepatocellular carcinoma. J Clin Oncol. 2016;34(5):452-459.

National Comprehensive Cancer Network. NCCN Clinical Practice Guidelines in Oncology: Hepatobiliary Cancers. Version 2.2016. https://www.nccn.org/professionals/physician_gls/f_guidelines.asp. Accessed August 22, 2016.

Truty MJ, Vauthey J-N. Surgical resection of high-risk hepatocellular carcinoma: patient selection, preoperative considerations, and operative technique. Ann Surg Oncol. 2010;17(5):1219-1225.

Garrean S, Hering J, Saied A, et al. Radiofrequency ablation of primary and metastatic liver tumors: a critical review of the literature. Am J Surg. 2008;195(4):508-520.

Bujold A, Massey CA, Kim JJ, et al. Sequential phase I and II trials of stereotactic body radiotherapy for locally advanced hepatocellular carcinoma. J Clin Oncol. 2013;31(13):1631-1639.

De Bari B, Ozsahin M, Bize P, et al. Can stereotactic body radiotherapy really be considered the preferred treatment in large hepatocellular carcinoma? J Clin Oncol. 2016;34(23):2798-2799.

Yang J-F, Lo C-H, Huang W-Y. Is stereotactic body radiotherapy better than radiofrequency ablation for the treatment of hepatocellular carcinoma? J Clin Oncol. 2016;34(23):2797-2798.

Wahl DR, Tao Y, Schipper M, et al. Reply to Yang et al and De Bari et al. J Clin Oncol. 2016;34(23):2799.

Kirichenko A, Gayou O, Parda D, et al. Stereotactic body radiotherapy (SBRT) with or without surgery for primary and metastatic liver tumors. HPB (Oxford). 2016;18(1):88-97.

GI ASCO: AHN Research Finds SBRT Effective Option for Liver Cancer Patients [press release]. PRWeb. January 22, 2016. http://www.prweb.com/releases/2016/01/prweb13178180.htm. Accessed August 8, 2016.

Matsuo Y, Yoshida K, Nishimura H, et al. Efficacy of stereotactic body radiotherapy for hepatocellular carcinoma with portal vein tumor thrombosis/inferior vena cava tumor thrombosis: evaluation by comparison with conventional three-dimensional conformal radiotherapy [published online April 6, 2016]. J Radiat Res. 2016. pii:rrw028.

Minagawa M, Makuuchi M. Treatment of hepatocellular carcinoma accompanied by portal vein tumor thrombus. World J Gastroenterol. 2006;12(47):7561-7567.

Huo Y, Eslick GD. Transcatheter arterial chemoembolization plus radiotherapy compared with chemoembolization alone for hepatocellular carcinoma: a systematic review and meta-analysis. JAMA Oncol. 2015;1(6):756-765.

Sapir E, Tao Y, Parikh ND, et al. Stereotactic body radiotherapy (SBRT) as an alternative to transarterial chemoembolization (TACE) for hepatocellular carcinoma (HCC). J Clin Oncol. 2016;34(suppl; abstr 4087)

Bai T, Chen J, Xie Z-B, et al. The efficacy and safety of postoperative adjuvant transarterial embolization and radiotherapy in hepatocellular carcinoma patients with portal vein tumor thrombus. Onco Targets Ther. 2016;9:3841-3848.

The jury is still out on whether stereotactic body radiation therapy (SBRT) should truly be considered a preferred frontline treatment modality for patients with hepatocellular carcinoma (HCC) who are not candidates for surgery, despite an ever-growing body of evidence supporting this approach.

Despite Growing Evidence for SBRT, the Jury Is Still Out

The objective response rate (ORR) was 16% among 214 patients who participated in an expansion cohort of the CheckMate-040 study, Bruno Sangro, MD, PhD, reported at the at the 2016 International Liver Cancer Association conference.

Among the 35 patients who responded, 33 had partial responses and 2 experienced a complete response. Another 111 patients had stable disease.

The interim findings are similar to efficacy and safety observations from the dose-escalation portion of the study reported at the 2016 ASCO Annual Meeting,

although the follow-up period is much shorter than the 3-year timeframe in the first phase, said Sangro, who is director of the Liver Unit at Clinica Universidad de Navarra in Pamplona, Spain.

Sangro said the follow-up period is too short to draw conclusions about overall survival (OS). The OS rate for all patients was 82.5% at 6 months and 70.8% at 9 months.

&ldquo;The 6-month and 9-month survival rates are not only highly encouraging but fit very well with the findings in the escalation cohort, in which the follow-up is much longer and in which 14.2-[month] median overall survival was reported at ASCO this year,&rdquo; sand Sangro. &ldquo;While these results are preliminary, the 9-month overall survival rate of 71% is certainly encouraging.&rdquo;

In all, the expansion phase of the trial consisted of 4 groups of patients, all of whom received nivolumab at 3 mg/kg every 2 weeks: patients without hepatitis who had not received or were intolerant to sorafenib (n = 54); participants who had progressed after receiving sorafenib (n = 58); patients who were infected with hepatitis C (n = 51); and individuals who were infected with hepatitis B (n = 51).

In the 2 hepatitis cohorts, participants must have received prior sorafenib, become intolerant to it, or progressed during or after therapy with the agent. Many of the participants across all cohorts were heavily pretreated with therapies including surgical resection (58%), locoregional therapy (59%), and systemic therapy including sorafenib (73%).

The primary endpoint of the trial was ORR according to RECIST v1.1 criteria by blinded independent central review. Secondary endpoints included duration of response, time to response, time to progression, and OS.

During the study, patients received a median of 10 doses of nivolumab (range, 1-27) over a median time on treatment of 20 weeks, Sangro said. &ldquo;Most of the patients who discontinued treatment did so because of progression,&rdquo; he said. &ldquo;Only a minority of 7% of the total population had to discontinue treatment because of study drug toxicity. Treatment was, by and large, very well tolerated.&rdquo;

The most frequent symptomatic treatment-related adverse events (AEs) of any grades included fatigue (21%), pruritus (15%), and rash (12%). Treatment-related grade 3/4 AEs were reported in 39 patients (18%).

In analyzing response rates by cohort, Sangro said ORRs were similar across the groups. The ORR ranged from 12% for patients who had been infected with hepatitis B to 20% for uninfected patients who were sorafenib na&iuml;ve or intolerant. &ldquo;Etiology is not a major factor in defining the response to nivolumab therapy,&rdquo; Sangro said.

He also said that the response rate did not differ by PD-L1 expression status, as assessed by immunohistochemistry on &ge;1% versus <1% of tumor cells.

In terms of durability, Sangro indicated that responses were ongoing in 30 of 35 responders at the time of analysis in March, and the median duration of response had not yet been reached. He also said 29 of the patients responded within 3 months of beginning treatment.

As analysis continues on this group of patients, a phase III randomized trial of nivolumab versus sorafenib has been launched in the frontline setting. The trial, which is seeking to enroll 726 patients, has an estimated primary completion date of July 2017.

Sangro B, Melero I, Yau T, et al. Safety and antitumor activity of nivolumab in patients with advanced hepatocellular carcinoma: interim analysis of dose-escalation cohorts from the phase 1/2 CheckMate 040 study. Presented at: 10th Annual International Liver Cancer Association Conference; September 9-11, 2016; Vancouver, Canada. Abstract O-019.

El-Khoueiry AB, Sangro B, Cheung Yau T, et al. Phase I/II safety and antitumor activity of nivolumab (nivo) in patients (pts) with advanced hepatocellular carcinoma (HCC): interim analysis of the CheckMate-040 dose escalation study. J Clin Oncol. 2016;34(suppl; abstr 4012). .

Nivolumab (Opdivo) continues to post &ldquo;highly encouraging&rsquo;&rdquo; and durable responses in patients with advanced hepatocellular carcinoma (HCC) regardless of whether they had hepatitis B or C or whether they had received prior treatment with sorafenib (Nexavar), according to the lead investigator on the early-phase study.

Nivolumab Maintains Positive Results in Latest HCC Findings

Regorafenib&rsquo;s success is set against a background of continued development of novel agents that may widen the treatment options for patients with advanced disease in the near future, speakers reported at the 10th International Liver Cancer Association (ILCA) Annual Conference.

The near-term potential includes a head-to-head comparison between lenvatinib (Lenvima) and sorafenib (Nexavar) in a first-line setting, and separate phase III studies versus placebo of cabozantinib (Cabometyx; Cometriq) and tivantinib in second-line settings, noted Ann-Lii Cheng, MD, PhD.

He also pointed to phase III trials of 2 PD-1 inhibitors, nivolumab (Opdivo) and pembrolizumab (Keytruda), as possible practice-changers in the next several years. Nivolumab is being tested against sorafenib in patients with advanced HCC who have not received systemic therapy, while pembrolizumab is being compared with placebo in previously treated patients.

Jordi Bruix, MD, PhD, a professor of Medicine at the University of Barcelona and director of the Barcelona Clinic Liver Cancer (BCLC) Group, which he founded, joined Cheng, of the National Taiwan University College of Medicine, on the podium in the discussion of the advanced disease paradigm. Bruix was the lead investigator for the pivotal RESORCE trial on regorafenib.

Both Cheng and Bruix noted that research changes are needed for improved outcomes in HCC. Cheng emphasized that biomarkers, particularly an immune signature, must be identified to help guide therapy while Bruix stressed the importance of revising the standards by which tumor progression is judged.

As it stands now, sorafenib is the only choice of systemic therapy for patients with advanced-stage HCC. Bruix said sorafenib is most effective for patients with stage C disease with symptoms including portal invasion and extrahepatic spread and Child-Pugh A-B scores, according to the BCLC algorithmthe most widely used staging system in the field.

The FDA approved sorafenib in 2007 for the treatment of patients with advanced, unresectable HCC based on a 3-month improvement in median overall survival (OS) compared with placebo (10.7 months vs 7.9 months; HR, 0.69).

Since then, phase III clinical trials have failed to register statistically significant improvements in OS in 5 first-line and 3 second-line studies involving targeted agents and, in 1 instance, chemotherapy regimens, according to Bruix.

That string of disappointments was broken earlier this year when Bruix and colleagues reported that regorafenib, which inhibits VEGF and multiple other kinases, improved median OS by 2.8 months versus placebo in the second-line setting.

Bayer Pharmaceuticals submitted the RESORCE findings to the FDA and the European Medicines Agency for approval.

After 9 years of failed trials for once-promising drugs, regorafenib (Stivarga) has emerged as the clear choice for second-line therapy in advanced hepatocellular carcinoma (HCC) after demonstrating survival improvements for patients whose disease has progressed after systemic treatment, according to liver cancer experts.

September 2016 | HCC Monitor

Cancer Stem Cell Therapies on the Horizon for HCC

New Evidence Supports a Key Role of the Immune System in HCC

Despite Growing Evidence for SBRT, the Jury Is Still Out

Nivolumab Maintains Positive Results in Latest HCC Findings

Regorafenib Moves Ahead in Advanced HCC

Expert Shares Views on RESORCE Data

Conference Marks a Decade of Milestones in HCC