Clinical Trial Details
— Status: Completed
Administrative data
| NCT number |
NCT03155113 |
| Other study ID # |
NR-DAA-HCV |
| Secondary ID |
|
| Status |
Completed |
| Phase |
N/A
|
| First received |
|
| Last updated |
|
| Start date |
August 30, 2017 |
| Est. completion date |
December 31, 2019 |
Study information
| Verified date |
July 2021 |
| Source |
Azienda Ospedaliero-Universitaria di Parma |
| Contact |
n/a |
| Is FDA regulated |
No |
| Health authority |
|
| Study type |
Interventional
|
Clinical Trial Summary
Anti-viral, hepatitis C virus (HCV)-specific immune T cell responses are functionally
defective in patients with chronic hepatitis C and this functional impairment is believed to
contribute to virus persistence. Persistent exposure to high virus loads is likely involved
in the pathogenesis of T cell dysfunction. The underlying hypothesis of the project is that
the level of anti-viral immune dysfunction in chronic HCV infection is a causal factor which
can influence non-response to therapy.
Although the rate of response to direct anti-viral agent (DAA) therapy, in untreated,
non-cirrhotic, patients is between 95% and 100%, however, the response rate is lower in
specific subgroups of patients, including genotype 3 cirrhotics and patients with
decompensated cirrhosis, irrespective of the infecting genotype.
Aim of the present study will be thus to understand whether non-response to therapy is
associated with a wider and deeper anti-viral immune dysfunction, by comparing individual
HCV-specific T cell responses in two groups of responder and non-responder patients.
Characterization of protective immunity in non-responder patients could allow to identify
baseline predictors of non-response to therapy to be used in the daily clinical practice.
Objective of the study will be to compare the features (intensity and quality) of the overall
HCV-specific immune T cell response in patients non-responder and responder to DAA therapy.
To achieve this goal, T lymphocytes (either CD4 or CD8) isolated from the peripheral blood of
the patients, before starting DAA therapy, will be stimulated with HCV proteins to evaluate
the capacity of those cells to expand, produce cytokines and express cytotoxic capacity.
Description:
Study objective and Hypothesis The underlying hypothesis of the project is that the level of
anti-viral immune dysfunction in chronic HCV infection is a causal factor which can influence
non-response to therapy. However, information about the features of individual anti-viral T
cell responses expressed by naïve chronic HCV patients before starting DAA therapy in
relation to the subsequent outcome of treatment and their possible impact on the failure to
control infection is not available. The few available results on anti-HCV immune responses in
chronic HCV infection have been generated in patients treated with DAA regimens which are not
included in the present guidelines without evaluation according to the outcome. Aim of the
present study will be thus to understand whether non-response to therapy is associated with a
wider and deeper anti-viral immune dysfunction, by comparing individual HCV-specific T cell
responses in two groups of responder and non-responder patients. The assumption made is that
possible defects of T cell responses associated with the failure to respond to DAA therapy
should not be substantially modified by an ineffective treatment allowing to characterize the
immunological background of non responder patients after the end of an ineffective cycle of
therapy and to compare results with baseline pretreatment responses of naïve, viremic
patients who subsequently clear the virus. A pre-treatment analysis also for non responder
patients would require to enroll a huge number of naive patients in order to identify a
sufficient number of non responder patients, making the study unfeasible and exceedingly
long. By this strategy, recruitment will be instead very rapid because most patients enrolled
at baseline are expected to be responder, while non responder patients who will be studied
after the end of therapy and after an adequate wash-out period of 6 to 12 months and before
starting re-treatment have already been identified in the collaborating Centers and will be
readily available for the study as soon as the protocol is approved. Based on the results,
possible objective of future analyses could be to characterize further at a molecular level
the T cell functional defects of non-responder patients to define whether and by which
strategies their correction can represent a feasible approach to induce/restore an efficient
anti-viral response to complement and strengthen the effect of last generation DAA. Finally,
characterization of protective immunity in non-responder patients could allow to identify
baseline predictors of non-response to therapy to be used in the daily clinical practice.
Study background The rate of sustained viral response (SVR) in naïve non-cirrhotic patients
treated with IFN-free DAAs is between 95% and 100%, but the response rate is lower in
specific subgroups of patients, including genotype 3 cirrhotics and patients with
decompensated cirrhosis, irrespective of the infecting genotype. The high efficacy of
DAA-based therapies is confirmed in real-world cohorts, which show a rate of SVR only
slightly lower than registration studies. Although the rate of failure to DAA therapy is
quite limited (around 5%), the overall number of non-responder patients is expected to be
high because of the high number of chronically HCV infected patients who need treatment.
Treatment failure most frequently results in relapse and less often in on-treatment viral
breakthrough. Different factors are believed to be implicated in non-response to therapy,
including emergence of resistance mutations, suboptimal treatment due to incorrect genotype
definition and advanced liver disease.
The role of baseline RAVs in determining treatment failure is still debated and baseline
resistance testing appears to have limited clinical utility. Instead, emergence of RAVs
during DAA-based regimens and its role in determining virological failure is well documented.
Even if drug-resistant variants are detected in a large proportion of non-responder patients,
their role in impairing treatment efficacy is however not totally clear.
The use of currently available second-generation commercial assays for HCV genotyping has
reduced the risk of genotype misclassification, but the possibility of mixed infections with
a percentage of different genotypes/subtypes below the sensitivity of the methods applied in
the clinical practice is still a possible cause of non response to therapy due to suboptimal
treatment.
Emergence of resistant strains and suboptimal treatment due to incorrect genotype detection
can however explain only part of treatment failure cases and host-related factors may play a
role, in particular the anti-viral immune response. Indeed, innate and adaptive immune
responses are known to be deeply impaired in chronic HCV infection but very limited
information is available about the possible contribution that background immune responses can
give to the final outcome of DAA treatment. In contrast to PegIFN-based therapies, recent
studies in DAA treated patients indicate that frequency and function of HCV-specific CD8
cells can increase under IFN-free therapies with partial reversal of their exhausted
phenotype. Moreover, DAA therapy can modulate the NK cell compartment correcting the NK cell
activated phenotype which is typical of chronic HCV patients. Thus, the level of baseline
impairment of anti-viral immunity might influence the subsequent likelihood of immune
restoration upon therapy with more chances of resistance to DAA treatment when baseline
immune inhibition is deeper and wider. This hypothesis requires to be tested.
Primary Endpoints Objective of the study will be to compare intensity (total levels of
anti-viral functions) and quality (multi-specificity and multi-functionality) of the overall
HCV-specific T cell response in patients non-responder (with and without detectable
resistance associated variants - RAVs) and responder to DAA therapy. To achieve this goal,
CD4- and CD8-mediated responses will be assessed by using overlapping synthetic peptides
covering the overall HCV proteome of genotype 1 in order to characterize T cell reactivity to
all HCV proteins, in terms of cytokine production (IL2, IFN-g and TNF-a) and cytotoxic
potential (CD107 degranulation).
- As a primary endpoint, the overall intensity of T cell responses will be assessed in
responder and non responder patients; the total intensity of individual T cell
reactivity will be defined by summing the different analyzed T cell parameters (cytokine
production and cytotoxicity, as detected in total CD3 cells and CD4/CD8 subsets) for
each individual responder and non-responder patient; the resulting values will be then
compared in the two groups of patients.
- At a second level of analysis, qualitative differences in T cell reactivity between
responder and non-responder patients will be assessed in terms of multi-functionality
and multi-specificity of HCV-specific T cell responses, by comparing the expression of
each individual function separately (IL2, IFN-g, TNF-a) and the capacity of each
individual HCV protein to induce T cell responses in the two groups of patients.
Characterization of HCV-specific T cell responses.
1. To analyze global CD4+ and CD8+ reactivity against structural and non-structural HCV
proteins, a comprehensive panel of overlapping 15-mer peptides covering the entire HCV
(genotypes 1) sequence will be used; T cell responses will be analyzed by
flow-cytometric intracellular cytokine staining (ICS) for IFN-g, IL-2 and TNF-a and for
degranulation (up-regulation of CD107) in vitro (after 10 days of peptide stimulation)
and by Elispot for IFN-g ex-vivo (after short-term peptide stimulation); synthetic
peptide epitopes with HLA class I- and class II-restricted specificities and known to be
the targets of CD8+ and CD4+ responses against different HCV-unrelated viruses and
pathogens (CMV, EBV, FLU) will be used as controls.
2. To further analyze CD8+ T cell reactivity, HLA-A2/peptide dextramers containing some of
the most widely recognized HLA-A2 restricted epitopes of HCV will be used in HLA-A2+
patients to quantify circulating virus-specific CD8+ cells and to measure expansion
capacity by comparing dextramer-positive CD8+ cell frequencies ex-vivo and after 10 days
of peptide stimulation.
3. In non-responder patients showing the emergence of resistance mutations, specific
peptides corresponding to the variant sequences will be synthesized and used to analyze
whether emerging mutations can influence HCV-specific T cell activation and function.
For this purpose, the stimulatory effect on different T cell functions of prototype and
variant peptides (cytokine production, cytotoxicity, capacity of expansion) will be
compared
Secondary Endpoints
1. Analysis of additional immune populations and serum factors known to be relevant with
respect to control of virus infection and modulation of T cell responses.
- Natural Killer (NK) cell analysis: NK cell phenotype will be studied by assessing
the expression of specific markers, such as CD16, NKG2A/D, TRAIL, NKP46/NKP30,
Ki67, CD38/HLA-DR by flow cytometry; NK cell function will be studied by testing
IFN-g/TNF-a production and CD107 degranulation upon overnight PBMC incubation with
appropriate stimuli.
- Analysis of T regulatory cell (Treg): frequency, phenotype and function will be
studied on whole PBMCs co-stained with CD3, CD4, CD25, FoxP3 and CD45RA.
- Serum concentration of cytokines, chemokines and ISG, including IL15, IL6, CXCL9,
CXCL10, IFN-g and IL28, will be analyzed in patients' sera by the Luminex
technology.
2. Identification of baseline predictors of non-response to DAA therapy Elucidation of the
impact that the anti-viral immune response can have in non-response to therapies with or
without protease inhibitors
