FINAL

Project Title

A Phase III Randomized, Double-Blind, Placebo-Controlled Trial of PRTX007, a TLR7 Agonist, in Combination with Standard Therapy for Hepatitis C Virus–Related Hepatocellular Carcinoma 

1.1 Summary of the project

This project will investigate a novel immunotherapy drug, PRTX007, which activates toll-like receptor 7 (TLR7), a key component of the innate immune system. In patients with chronic hepatitis C virus (HCV) infection, prolonged liver damage and immune suppression frequently lead to hepatocellular carcinoma (HCC). While recent treatments such as immune checkpoint inhibitors and anti-angiogenic agents have improved outcomes for some patients, many fail to respond or relapse after initial benefit, demonstrating the need for additional therapies (Llovet et al., 2021). 

The aim of this study is to evaluate whether adding PRTX007 to standard treatment can improve clinical outcomes in patients with HCV-related HCC. The trial will assess whether PRTX007 delays cancer progression, enhances tumour response rates, improves quality of life, reduces HCV viral load, and stimulates immune activation. By targeting innate immunity, PRTX007 is designed to reawaken suppressed pathways in the tumour microenvironment, potentially overcoming resistance to existing therapies (Smits et al., 2023). 

This will be a phase III, randomised, placebo-controlled clinical trial. Participants will be randomly assigned to receive either PRTX007 plus standard treatment or placebo plus standard treatment. To minimise bias, both participants and clinicians will be blinded to treatment allocation. 

Over six months, patients will undergo CT or MRI scans to monitor tumour size, blood tests to assess viral load and immune activation, and validated questionnaires to measure quality of life. Tumour progression will be assessed using RECIST 1.1 and mRECIST criteria, while quality of life will be evaluated with the EORTC QLQ-C30 and the HCC-specific QLQ-HCC18 (Aaronson et al., 1993; Blazeby et al., 2004). 

The primary outcome is progression-free survival (PFS), defined as the time from randomisation until disease progression or death. Secondary outcomes include tumour response rates, duration of response, changes in quality of life, reductions in viral load, and evidence of immune activation. 

If successful, this trial will provide evidence that PRTX007, by stimulating innate immunity, can improve survival and wellbeing in patients with HCV-related HCC. The findings have the potential to expand treatment options, reduce reliance on late-stage interventions, and lessen the burden of this disease on both patients and the healthcare system. 

1.2 Rationale and proposed impact of study

Pathophysiology of the Disease 

Hepatocellular carcinoma (HCC) is the most common form of primary liver cancer and a leading cause of cancer-related mortality. It most often develops in the context of chronic liver disease, particularly cirrhosis. Chronic hepatitis C virus (HCV) infection has long been one of the strongest risk factors. Although direct-acting antivirals (DAAs) now cure more than 95% of infections, many individuals were infected decades before these treatments were available, leaving them with advanced fibrosis, cirrhosis, or established HCC (Ioannou, 2021). Even after viral eradication, patients with cirrhosis remain at elevated risk of cancer due to persistent structural and molecular alterations within the liver (Kanwal & Singal, 2019). 

The pathogenesis of HCV-related HCC is multifactorial and non-linear. HCV viral proteins disrupt intracellular signalling pathways, leading to cell-cycle dysregulation, uncontrolled proliferation, and impaired apoptosis. Chronic viral replication drives necroinflammatory and regenerative cycles, promoting overall genomic instability and mutations (Ringelhan et al., 2018). Fibrosis and cirrhosis further distort hepatic architecture, creating a microenvironment that favours carcinogenesis. In parallel, immune dysregulation develops: regulatory T cells, myeloid-derived suppressor cells, and cytokines such as TGF-β and IL-10 suppress tumour surveillance and blunt adaptive responses (Llovet et al., 2021). 

The liver is inherently an immune-tolerant organ, constantly exposed to dietary and microbial antigens from the gut. In chronic HCV infection, this tolerance is exacerbated, resulting in tumours with minimal T-cell infiltration. These immune “cold” tumours often respond poorly to current immunotherapies. This immune biology provides strong justification for therapies such as PRTX007, which aim to reinvigorate innate immune responses. 

Existing Treatments – Pros and Cons 

Treatment for HCC has advanced significantly in the past 15 years. Sorafenib, a multikinase inhibitor, was the first systemic agent to demonstrate a survival benefit in advanced disease, establishing a cornerstone for management. Lenvatinib has since been approved as a first-line alternative with comparable efficacy but a distinct toxicity profile. Both drugs, however, are associated with side effects including fatigue, hypertension, diarrhoea, and hand–foot syndrome, and offer only modest improvements in survival. 

The greatest progress has been achieved with immunotherapy-based combinations. The IMbrave150 trial showed that atezolizumab plus bevacizumab significantly improved overall survival and progression-free survival compared to sorafenib, and it is now considered a standard first-line regimen (Finn et al., 2020). Similarly, durvalumab combined with tremelimumab has demonstrated durable responses in unresectable disease and represents another frontline option (Abou-Alfa et al., 2022). 

Despite these advances, substantial challenges remain. A large proportion of patients do not respond at all, largely due to tumours with poor immune infiltration or defective antigen presentation. Among responders, resistance frequently emerges within 12–18 months, driven by tumour adaptation and immune escape (Llovet et al., 2021). Treatment-related toxicities also pose barriers: bevacizumab increases bleeding risk in cirrhotic patients, while checkpoint inhibitors can cause immune-related adverse events such as hepatitis, colitis, and endocrinopathies. Later-line agents including regorafenib, cabozantinib, and ramucirumab provide only incremental benefits and are often poorly tolerated. 

Thus, while immunotherapy has reshaped the treatment landscape, survival outcomes remain unsatisfactory for many patients. There is an urgent need for therapies that act upstream in the immune cascade, such as PRTX007, to enhance initial responsiveness and extend the durability of benefit. 

The Impact on the Individual & Health System 

For individuals, advanced HCC is clinically devastating. Even with modern therapies, median survival typically remains under two years, during which patients experience debilitating symptoms such as ascites, jaundice, hepatic encephalopathy, abdominal pain, fatigue, and anorexia. These symptoms severely compromise physical, emotional, and social wellbeing. Psychological distress, including depression and anxiety, is common due to the uncertainty of prognosis and the aggressive course of disease (Llovet et al., 2021). 

The social consequences extend beyond the patient. Many individuals lose employment and independence, resulting in financial strain and increased reliance on carers. Families often shoulder the burden of organising daily care, managing medical appointments, and providing emotional support, creating ripple effects across households and communities. 

The healthcare system also bears a significant burden. In Australia, HCC incidence and mortality continue to rise despite widespread DAA use, as patients with cirrhosis remain at lifelong risk (The Kirby Institute, 2023). Systemic therapies are costly, and cirrhotic patients require frequent hospitalisations for complications such as variceal bleeding, hepatic decompensation, and encephalopathy. Liver transplantation offers a potential cure, but eligibility criteria exclude most patients and donor organ shortages further limit access. As a result, HCC places sustained pressure on oncology, hepatology, surgical, palliative, and public health services. 

Overall Burden of Disease 

HCC is the third leading cause of cancer-related death globally, with more than 800,000 deaths annually (Sung et al., 2021). Incidence is highest in regions with endemic viral hepatitis, including East Asia and sub-Saharan Africa. In Western countries, rising rates of HCV infection, alcohol-related liver disease, and non-alcoholic fatty liver disease (NAFLD) have contributed to increasing incidence. 

In Australia, HCV-related HCC remains a significant health challenge. Approximately 10% of liver transplants are attributable to HCV, despite the success of DAAs (ASHM, 2023). Surveillance reports estimate that thousands remain at risk due to advanced fibrosis and cirrhosis (The Kirby Institute, 2023). The disease also exacerbates health inequities, disproportionately affecting people with a history of injecting drug use, Aboriginal and Torres Strait Islander peoples, and migrants from high-prevalence regions (Butt et al., 2022). 

The socioeconomic burden is equally profound. Patients frequently rely on social welfare systems due to loss of employment, while families face financial strain from caregiving responsibilities. Healthcare costs continue to rise due to advanced surveillance, hospitalisations, and the expense of novel systemic therapies. Taken together, HCV-related HCC represents not only a clinical but also a substantial social and economic burden. 

Rationale for Investigating PRTX007 

The limitations of current therapies underscore the need for novel strategies. PRTX007 offers a promising solution by targeting innate immunity at its source. As a TLR7 agonist, it activates plasmacytoid dendritic cells and induces type I interferon production, enhancing antigen presentation and priming T-cell responses (Smits et al., 2023). By converting immune “cold” tumours into “hot” tumours, PRTX007 may improve responsiveness to existing immunotherapies and provide benefit in resistant cases. 

Early studies of related TLR7 agonists suggest a manageable safety profile, with transient flu-like symptoms being the most common adverse events (Wolska et al., 2020). The mechanism of action is distinct yet complementary to checkpoint inhibitors, supporting potential utility both as monotherapy and in combination regimens. 

If effective, PRTX007 could extend survival, preserve quality of life, and reduce reliance on costly late-stage interventions such as transplantation. Furthermore, because HCV-related HCC disproportionately affects disadvantaged groups, an effective new therapy could help reduce health inequities. 

Taken together, the pathophysiology of HCV-related HCC, the shortcomings of current treatments, and the ongoing global and national burden demonstrate a strong need for innovative therapies. By stimulating innate immunity, PRTX007 has the potential to address this gap, offering meaningful benefits for patients, healthcare systems, and society. 

2.1 Research Plan - Background

. Hepatocellular carcinoma (HCC) has been shown to be linked to hepatitis C infections. Hepatitis C is a single-stranded RNA (ssRNA) virus. These types of viruses are recognized by the endosomal Toll-Like Receptor TLR7 (REN et al., 2016). 

TLR7 activation in cases of viral infection is essential for immune and inflammatory responses. Downstream activation of TLR7 activates both innate and adaptive immune cells via the recruitment of multiple types of IL-1-associated receptor kinases. The proinflammatory NF-κB pathway is also activated via MyD88. NF-ΚB activation is essential to the immune response to infection as the inflammation is regulated, contributing to the activation of immune cells, and guidance of immune cells to the infected tissue. (Mifsud, Tan and Jackson, 2014) 

Issues arise in instances of prolonged infection. Long-term activation of NF-κB results in excessive production of reactive oxygen species (ROS) and reactive nitrogen intermediates (RNI) (Mao, Zhao and Sun, 2025). Both of these have a mutagenic effect on DNA. When working properly, this can cause infected cells to be killed and viral DNA to become damaged. However, some of the mutations caused by ROS and RNI can be oncogenic. 

Because cancer formation and sustained periods of inflammation, NF-ΚB is a target for cancer treatment as an overarching regulator of inflammation. Research into NF-ΚB inhibition or inactivation has previously been conducted as a potential method of cancer treatment. 

Imiquimod is a TLR7 agonist that has been tested for its effect on NF-ΚB recruitment in cell cultures. In this study, cultures were inoculated with TLR7 ligand and then treated with the drug. Cell growth and inflammation were then measured among different treatment groups which received a different number of treatments with Imiquimod. The results showed suppression of growth of HCC cells in culture (REN et al., 2016). This indicates a lack of NF-ΚB activation, demonstrating that TLR7 agonists can be used as a potential treatment method for HCC. 

This trial proposal for PRTX007 suggests that a similar effect can be achieved in humans with this drug. PRTX007 is a prodrug for PRX034, which is another TLR7 agonist. The drug is administered as PRTX007 as the drug is given orally, and PRX034 has been shown to cause activation of gastrointestinal immune cells, which would cause further inflammation of the GIT and is not favourable for this treatment (Appleman & Webber, 2020). 

PRTX007 is a guanosine analogue that binds to TLR7 in place of ssRNA, initiating the IRAK mediated sequestering of innate and adaptive immune cells without the typically associated activation of the NF-ΚB pathway.  

PRTX007 has been tested within an animal study on Cynomolgus Monkeys. The study found a lack of side effects related to the drug, and that the drug was able to activate an immune response without activation of the NF-κB pathway (Appleman & Webber, 2020). 

PRTX007 has also been tested in a phase 1 trial, which demonstrated favourable results. The trial included 130 participants, 104 of which were administered the drug. Within the multiple test groups on different dosing schedules, expression of interferon-encoding genes activated by TLR7 was found to be as expected for what would typically occur in a viral infection. Expression of some intermediary proteins was higher in the test group than the control group. Members of the test group also demonstrated increased proliferation of T cells and NK cells as would be expected in a viral infection (Lemech et al., 2023). 

There was also a lack of detected proinflammatory molecules regulated by NF-ΚB in participants such as IL-6 and TNF-a. This, paired with the presence of immune cells and immune cell activating molecules, indicates that PRTX007 was successful in activating an immune response via TLR7 activation without activating the NF-ΚB pathway (Lemech et al., 2023). 

The phase 1 trial also found the drug to be safe with only minimal minor side effects. This includes headaches, not related to dosage, and dose-related fevers that subsided once dosage was adjusted (Lemech et al., 2023). 

The activated immune response is has anti-cancer properties alongside being used to combat the underlying viral infection which causes HCC. Activation of TLR7 via PRTX007 induced plasmacytoid dendritic cells to express CD40, CD80, and CD86. These receptors are involved in the activation of B and T cells. These adaptive immune cells can target and kill tumour cells while they form (Lemech et al., 2023). 

The combined inactivation of the NF-ΚB pathway and activation of an anti-tumour immune response via adaptive immune cells both serve to combat cancer growth and spread. As such, PRTX007 achieving both of these makes it a prime candidate for further study in the treatment of HCC. 

2.2 Research Plan - Aims

This trial would aim to measure the effectiveness of PRTX007 in treating hepatitis C mediated hepatocellular carcinoma. It would also test the safety of the drug for its use in cancer treatment schedules. 

It is hypothesised that, based on phase 2 trials, the use of PRTX007 in a cancer treatment schedule would result in a reduction of inflammation via the NF-KB pathway, contributing to a reduction in cancer growth and spread with minimal side effects. 

2.3 Impact statement

A successful trial will show a reduction in growth and spread of hepatocellular carcinoma in trial patients. This reduced rate of growth will ease the physical burden of the disease and treatment on patients.  

Reducing inflammation caused by hepatitis C infection will reduce the mutagenic effect of viral infection on liver cells, lowering the rate of mutation within hepatic cells and reducing their risk of becoming cancerous. Simultaneously, PRTX007 activates the immune response that would typically result in the aforementioned inflammation. This means that in the absence of the typical immunosuppressive effect of many cancer drugs the body is still able to combat the hepatitis C infection while receiving treatment for their hepatocellular carcinoma. 

Since growth of the cancer will be reduced, there is likely to be less liver damage in patients receiving PRTX007. This means that patients are likely to retain more functional liver tissue. In cases where damaged tissue must be removed to avoid patients developing sepsis, that patients will retain more of their liver. This will increase quality of life for patients and reduce the possibility that effected patients will require liver transplants. 

3.1 Objectives

The primary purpose of this study is to find out if PRTX007, when added to standard treatment, can delay cancer progression and improve short-term clinical outcomes within 6 months in patients with HCV-related hepatocellular carcinoma (HCC). The trial focuses on progression-free survival (PFS), tumour response rates, and immune/viral activity as outcomes, that can be measured within the 6-month timeframe. Secondary outcomes include the drug's response rate and the amount of patients attaining complete or partial tumour response (responders vs non-responders). The duration from the initial recorded tumour response to cancer progression will also be assessed (in weeks). Another outcome is improving the participants' quality of life over the 6 month time period which can be measured on a scale. 

PFS is a recognised substitute endpoint in cancer trials, especially when the trial only lasts a short time. In a 6-month period, variations in overall survival may not be evident; however, PFS offers a quantifiable and clinically significant indicator. The tumour response rate adds to this by displaying how many patients benefit from therapy on X-rays, and immunological and viral markers let us see how PRTX007 works in a unique way. 

This balanced set of outcomes ensures that the study does not only focus on tumour control but also incorporates patient-centred outcomes (quality of life) and mechanistic data (immune and viral activity), creating a comprehensive representation of efficacy and tolerability. 

3.2 Data Collection

PFS will be measured in weeks from randomisation to either cancer progression or fatality. RECIST 1.1 and mRECIST for HCC will be used to monitor progression. Blinded radiologists will review this to reduce biases and ensure more objective outcomes. Imaging (CT or MRI) will happen at start of Week 8, Week 16, and Week 24. The size of the tumour will be measured in millimetres (mm), and the total of the diameters of the target lesions will be compared to the baseline. Progression is marked by a minimum 20% increase in tumour size, the emergence of more lesions, or an overall decline of the tumour. Death at any time will also be considered an event. To ensure accuracy, each patient will undergo imaging using the same modality and specific scanning methods. 

Tumour response will be recorded as the percentage of patients achieving Complete Response (CR) or Partial Response (PR) according to RECIST 1.1/mRECIST. This is a binary outcome (responder vs non-responder). Responses must be confirmed on repeat imaging at a minimum of 4 weeks later to reduce false positives. If images are poor quality due to motion or contrast failure, a repeat scan will be requested. 

We will use the EORTC QLQ-C30 and the HCC-specific QLQ-HCC18 to measure QoL. Patients will finish them at the start, Week 8, Week 16, and Week 24. Scores go from 0 to 100. Higher scores on functional scales represent better function, whereas higher scores on symptom scales mean worse symptoms. The result is the difference in the mean score between the start and 6 months later. Scoring manuals will be used to deal with missing items; computerised completion cuts down on lost data. Using both a general cancer QoL instrument and an HCC-specific module lets doctors look at treatment tolerability in more detail, covering symptoms like fatigue, pain, hunger, and liver-related problems. To cut down on missing data, questionnaires will be sent out electronically. If items are missing, validated scoring rules will be used. A predetermined minimal clinically important difference (MCID) of 10 points will be utilised to assess significance. 

We will measure the amount of Hepatitis C virus in the blood using a standard PCR blood test at the start, Week 8, Week 16, and Week 24. We will record how much the virus level drops and how many patients reach “undetectable” levels. 

We will test blood samples to see if the immune system has been activated. This includes checking certain “interferon genes” and looking at markers on immune cells (NK cells) that show they are switched on. Results will be reported as changes in activity levels compared to baseline. 

All scans will be sent to a central core lab for quality control checks on matters such as contrast timing, slice thickness, and motion artefacts. If a scan is not adequate, patients will be asked to come back for another one within a week. To prevent misdiagnosis resulting from "pseudo progression"—a recognised phenomenon in immunotherapy where tumours initially enlarge due to immune infiltration—progression must be confirmed with repeat imaging while patients are clinically stable. 

3.3 Treatment protocol & Controls

Oral PRTX007 once weekly for 24 weeks. To improve tolerability, a lead-in of 100 mg weekly for 4 weeks is followed by 200 mg weekly from Week 5–24, or the closest dose that the protocol says is safe. Participants will continue to use  pre-selected guideline-concordant standard of care (SOC) without change unless toxicity mandates it. Oral placebo once weekly on the same schedule whilst maintaining the same type of SOC chosen before randomisation. 

A placebo add-on is suitable due to the absence of a established TLR7 agonist for HCC, the research relates to the additional advantage of innate immune stimulation beyond current best practices. Using SOC in both controls makes sure that ethical integrity is maintained and no one is refused effective therapy and that PRTX007's effect is isolated. 

The trial is double-blind, which means that the capsules, packaging, dosing windows, and visit schedules are all the same. Randomisation employs a central interactive web response including secret, changeable block sizes. Emergency unblinding is only available when medically necessary. 

To reduce bias, the trial requires that the standard of care (SOC) , whether immunotherapy-based or tyrosine kinase inhibitor (TKI)-based, is predetermined prior to randomisation. The trial requires that the standard of care (SOC) backbone, whether immunotherapy-based or tyrosine kinase inhibitor (TKI)-based, is predetermined prior to randomisation. All imaging endpoints will be reviewed in a Blinded Independent Central Review (BICR) with RECIST 1.1 and mRECIST criteria, therefore assuring consistency in tumour assessments. Standardised rules will regulate supportive care and accompanying use of medications, allowing for the use of direct-acting antivirals (DAAs), which will be recorded and included into the analysis. The intention-to-treat (ITT) population will be the primary analysis group, whereas per-protocol analyses will be conducted as sensitivity assessments. Finally, all groups will have the same schedules for tests, evaluations, and imaging, which will make it easier to compare them and lower the chance of performance bias. 

Outcomes in advanced HCC differ based on stage, liver function, and SOC dosage; historical controls may include complicating variables and cannot guarantee comparability. A simultaneous, randomised, double-blind placebo plus standard of care control offers the most unbiased assessment of PRTX007's additional value. 

3.4 Study Design

This is a randomised, double-blind, placebo-controlled Phase 3 trial lasting 6 months. Patients with HCV-related hepatocellular carcinoma will be randomised 1:1 to receive PRTX007 + standard of care (SOC) or Placebo + SOC. 

Duration: 24 weeks of treatment and follow-up. 

Participants: 300-5,000 (even splits per arm). 

Visits and measurements: 

Screening (≤28 days before treatment): Baseline imaging (CT/MRI), labs, HCV RNA, immune markers, QoL questionnaires. 

Treatment Phase (Weeks 1–24): 

·       Week 2 & 4: Safety , side effects. 

·       Week 8 & 16: Imaging, HCV RNA, immune markers, QoL questionnaires, safety checks. 

·       Week 24 (End of Study): Imaging, HCV RNA, immune markers, QoL questionnaires, final safety review. 

Follow-up: Survival status checked up to Week 24; post-trial extension optional. 

3.5 Randomisation

Patients will be randomly assigned in a 1:1 ratio to receive either PRTX007 plus standard of care (SOC) or a placebo plus SOC. Randomisation will take place centrally through an Interactive Web Response System (IWRS), which automatically creates assignments after confirming patient eligibility. This keeps the allocation secret, stops selection bias, and stops predictable sequences that could happen with simpler systems like coin flips. 

To maintain balance between groups, a permuted block design with variable block sizes will be used. This prevents researchers from anticipating the next allocation while ensuring that treatment groups remain approximately equal in size throughout enrolment 

Because this is a multicentre trial, randomisation will also be based on a small number of relevant prognostic criteria. These are: 

·       Baseline HCV RNA status (detectable vs undetectable/cleared). 

·       Type of SOC backbone (immunotherapy-based vs TKI-based). 

·       Disease extent (liver-only vs presence of macrovascular invasion/extrahepatic spread) 

There will be no more than these three strata used, since too many strata can make analysis harder and lower statistical power. The IWRS will make sure that patients are evenly spread among arms and participating centres within every level. This will keep the trial's integrity and comparability.