# Diagnostic value of the wnt target and cancer-associated blood biomarker hPG80: ONCOPRO case-control prospective study

Benoit You, Sebastien Couraud, Philippe Ceruse, Lionel Badet, Philippe Paparel, Alice Durand, Marielle Guillet, Philippe Merle, Gaelle Lescuyer,

Charles-André Philip, et al.

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Benoit You, Sebastien Couraud, Philippe Ceruse, Lionel Badet, Philippe Paparel, et al. Diagnostique value of the wnt target and cancer-associated blood biomarker hPG80: ONCOPRO case-control prospective study. Biomarker Research, 2025, 13 (1), pp.91. 10.1186/s40364-025-00793-z. hal-05223450

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# Diagnostic value of the wnt target and cancer-associated blood biomarker hPG80: ONCOPRO case-control prospective study

Benoit You<sup>1,2,19*</sup>, Sebastien Couraud<sup>1,3</sup>, Philippe Ceruse<sup>1,4</sup>, Lionel Badet<sup>1,5</sup>, Philippe Paparel<sup>1</sup>, Alice Durand<sup>1,6</sup>,
Marielle Guillet<sup>1,6</sup>, Philippe Merle<sup>1,7</sup>, Gaelle Lescuyer<sup>1,8</sup>, Charles-André Philip<sup>1,9</sup>, Francois Ducray<sup>1,10</sup>, Mathieu Pioche<sup>1,6</sup>,
Lionel Karlin<sup>1,11</sup>, Jean-Christophe Lifante<sup>1,12</sup>, Olivier Glehen<sup>1,13</sup>, Pierre-Adrien Bolze<sup>1</sup>, Marion Chauvenet<sup>1,14</sup>,
Carole Langlois-Jacques<sup>1,15,16</sup>, Fabien Subtil<sup>1,15,16</sup>, Marie Piecyk<sup>1,8</sup>, Aurore Carrot<sup>1</sup>, Dominique Joubert<sup>17</sup>,
Alexandre Prieur<sup>17</sup>, Berengere Vire<sup>17</sup>, Sara Calattini<sup>1</sup> and Lea Payen<sup>1,8,18</sup>

**Abstract**

**Background** hPG<sub>80</sub> (circulating progastrin), initially recognized for its oncogenic properties due to its direct link to the Wnt signaling pathway, is secreted by cancer cells and detectable in the blood of cancer patients. The ONCOPRO centralized case-control study (NCT03787056) was designed to prospectively evaluate the diagnostic utility of hPG<sub>80</sub> in patients with 16 different types of cancer.

**Methods** hPG<sub>80</sub> levels were measured in 421 patients with 16 newly diagnosed cancers (median age 65.6 years old) using the DxPG80.Lab kit (Biodena Care). The diagnostic performance of hPG<sub>80</sub> (the primary endpoint) was assessed by comparing baseline hPG<sub>80</sub> levels in cancer patients with those of 330 asymptomatic aged-matched healthy subjects from the general population.

**Results** Between 2018 and 2022, a total of 506 cancer patients were enrolled in the study, with 421 assessable across 16 distinct cancer cohorts. hPG<sub>80</sub> concentrations were significantly higher in cancer patients compared to the healthy population (median 3.8 [IQR: 1.0-11.1] vs. 1.9 [IQR: 0.6-4.2] pM, *P* < 0.0001). hPG80 levels were not impacted by renal failure, liver dysfunction, or cancer-associated inflammation. The diagnostic accuracy in cancer patients was ROC AUC 0.63, 95% CI = [0.59-0.67]. The highest diagnostic accuracy was seen in patients with lung cancer (AUC 0.75, 95% CI [0.68-0.82]); specificity = 88% for hPG<sub>80</sub> > 7.73 pM in patients aged > 58 years old) and hepatocellular carcinoma HCC (ROC AUC 0.75, 95% CI [0.66-083]); specificity = 88% for hPG<sub>80</sub> > 7.73 pM in patients aged > 58 years old).

**Conclusions** This large prospective study confirms that cancer patients have significantly higher hPG<sub>80</sub> blood concentration compared to the healthy population. Incorporating this straightforward ELISA assay into screening programs is warranted.

**Trial registration** NCT03787056.

Full list of author information is available at the end of the article

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BMC

© The Author(s) 2025. **Open Access** This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated in a credit line to the data.

<footnote>*Correspondence:
Benoit You
benoit.you@chu-lyon.fr</footnote>
**Keywords** Biomarkers, tumor, Early detection of Cancer, Wnt signaling pathway, Prospective studies

**Introduction**

According to a study done by the World Health Organi-
zation in 2021, cancer is one of the top two leading causes
of death globally [1]. Among various strategies aiming at
improving cancer prognosis, one promising approach
involves the development of screening programs capable
of detecting cancer at an early stage when the disease is
still curable [2].

Recently, several multi-cancer blood assays have
emerged, attracting significant interest from the scien-
tific and medical communities. These assays hold poten-
tial for integration into screening programs to identify
patients with cancers originating from different tissues.
Most of these assays are based on circulating tumor DNA
(ctDNA) technology, which, despite its promise, faces
limitations regarding complexity, cost, and sensitivity for
routine clinical use [3].

hPG<sub>80</sub> (extracellular circulating progastrin) is emerging as a promising blood-based biomarker for various cancers, with significantly higher levels observed in cancer patients compared to healthy asymptomatic individuals [4]. In healthy individuals, progastrin is produced by G cells of the stomach, and is the precursor of gastrin. In tumor cells, the GAST gene, which encodes hPG<sub>80</sub>, is directly regulated by the WNT/β-catenin oncogenic pathway, a pathway activated in many cancers [5]. Unlike in stomach G cells, tumor cells produce progastrin but do not process it into mature gastrin. This secreted unprocessed form, named hPG<sub>80</sub>, is easily measurable using a cost-effective and straightforward ELISA method [4]. Several studies showed the strong relationships between GAST gene activation, WNT pathway activation, and hPG80 expression in adenoma and cancer cells [6–8].

hPG<sub>80</sub> plays a pivotal role in tumorigenesis by regu-
lating cancer stem cell functions [9, 10], promoting
angiogenesis [11], and influencing apoptosis [12]. Retro-
spective studies have established correlations between
elevated hPG<sub>80</sub> levels and poor survival outcomes in met-
astatic renal cell carcinoma (mRCC) [13], hepatocellular
carcinoma (HCC) [14], breast cancer [10], and glioblas-
toma [15].

However, these findings were derived from retrospec-
tive analyses. A prospective study was necessary to vali-
date these results, identify cancer types with the highest
diagnostic accuracy for hPG<sub>80</sub>, and strengthen its poten-
tial for clinical application.

The ONCOPRO study (NCT03787056) was a prospec-
tive case-control study designed to assess: (1) the diag-
nostic value of hPG<sub>80</sub> by comparing blood concentrations
of this marker in patients with 16 newly diagnosed cancer
types to those in asymptomatic healthy subjects (primary

endpoint); and (2) the monitoring potential of hPG<sub>80</sub> by
evaluating its longitudinal kinetics during treatment (sec-
ondary endpoints).

The results for the primary endpoint are presented
here.

**Patients and methods**

Cancer patient case cohorts

ONCOPRO study (NCT03787056) has been conducted
in 17 medical or surgical cancer departments of a single
institution with a centralized coordination (Lyon Uni-
versity Hospital, HCL, Lyon, France). The patients were
enrolled in 16 cohorts, including 6 large cohorts with
minimum 50 patients (Non-Small Cell lung Carcinoma
(NSCLC) & Small Cell Lung Carcinoma (SCLC); Breast;
Renal; Prostate; Hepatocellular carcinoma (HCC); and
Head-and-Neck carcinomas) and 10 small explorative
cohorts including 10 patients (Colorectal; Thyroid; Pan-
creas; Glioblastoma; Endometrial; Bladder; Superficial
esophageal-gastric; Lymphoma; and Gastric carcinomas).

The selection of targeted cancers was based on the
available data about hPG<sub>80</sub>, and clinician perception
about the unmet medical need for early diagnosis tests.
Large cohorts were meant to assess the diagnostic value
in these selected cancers with sufficient statistical power,
whilst the small cohorts were more explorative.

To be enrolled and considered assessable, the patients
had to be older than 18 years, diagnosed with a histologi-
cally and/or cytologically documented cancer, planned to
be treated with a curative or non-curative intent strategy.
Patients enrolled in “curative intent strategy” cohorts had
to be naïve of anticancer treatment, whilst the patients
enrolled in “non-curative intent strategy” cohorts had
to be naïve of treatment in the non-curative setting (but
could have been previously treated for cancer in curative
setting) (Fig. 1).

The other inclusion criteria included measured cre-
atinine clearance > 30 mL/min or creatinine ≤ 1.5 x ULN;
aspartate aminotransferase (AST) and alanine amino-
transferase (ALT) ≤ 2.5 x ULN (or ≤ 5 x ULN, in case of
liver metastases); and serum bilirubin ≤ 1.5 x ULN. Spe-
cific biological criteria applied for the hepatocellular car-
cinoma cohort (Suppl Material).

Exclusion criteria for curative-intent cohorts com-
prised a history of previous cancers, except for ade-
quately treated non-melanoma skin cancer, curatively
treated in-situ cancer of the cervix with no evidence of
disease for ≥ 5 years. The same criteria applied for non-
curative-intent groups, except that patients with a recur-
rent cancer could have previously been treated with a
curative intent treatment.
A

<figure><img src="image_5.png" /></figure>

B

<figure><img src="image_6.png" /><figcaption>**Fig. 1 ONCOPRO study design with biological sampling strategies for (A) curative intent or non-curative intent treatments, and (B) during systemic or local treatments**</figcaption></figure>

In addition, some patients were proposed to also participate in one of the two kinetic cohorts designed to better understand the longitudinal kinetics of hPG<sub>80</sub> in short time windows: (1) nychthemeral cohort meant to assess the hPG<sub>80</sub> changes during a 24 h time period with respect to meals; (2) a post-operative cohort, proposed to patients treated with initial curative surgery and meant to assess the early dynamics of hPG<sub>80</sub> in the 24 h following the surgical procedure.

All patients gave their consent and signed the informed consent. The protocol, the written information sheet and the consent form for the study obtained a favorable opinion from to ethics committee Ile de France VI on October 3rd 2018. This notification was transmitted to the French health authority ANSM. All Protocol amendments were approved by the same ethic committee.

### hPG<sub>80</sub> sampling times for cancer cohorts

Circulating hPG<sub>80</sub> sampling strategy was individually adjusted to the patient treatment schedule (Fig. 1). The general principle was that hPG<sub>80</sub> was sampled at baseline before any treatment (for the primary objective), and then at each hospital visit for those treated with intravenous systemic treatment, before and after surgery in the case of curative surgery, before and after radiation therapy when applied, at each consultation visit for oral treatment, and during the follow-up period. For curative-intent cohort patients, hPG<sub>80</sub> was monitored during treatment, follow-up, and at relapse if any, for a

maximum of 5 years. For non-curative-intent cohort patients, hPG<sub>80</sub> was monitored on 3 consecutive treatment lines (Fig. 1).

In addition to hPG<sub>80</sub>, the following biomarkers were measured at the same times: AST; ALT; serum bilirubin (in addition to albumin, factor V et prothrombin time for HCC), serum creatinine, creatinine clearance, C-Reactive Protein (CRP); serum tumor markers for specific cohorts (CA15-3 and Carcinoembryonic antigen (CEA) for breast cancer, Prostate-specific antigen (PSA) for prostatic cancer, alpha foetoprotein (AFP) for HCC, CA19-9 and CEA for colorectal, gastric, superficial oesophago-gastric and pancreatic cancers, thyroglobulin for thyroid cancer, and CA125 for ovarian cancer and endometrial cancers).

In the nychthemeral cohort, hPG<sub>80</sub> was sampled before start of treatment on a 24-hour period, starting on day 1 at 11 am, 12 pm, 2 pm, 5 pm, 6 pm, 9 pm, 9 to 10 pm, and on day 2 at 7 am and 9 am, to assess a potential impact of meal intakes on hPG<sub>80</sub> concentrations (Suppl Fig. 1). In the post-operative kinetic cohort, hPG<sub>80</sub> was sampled just at the end of surgery, + 30 min, + 60 min, + 2 h, + 4 h, + 6 h, + 12 to 24 h (Suppl Fig. 1).

### Asymptomatic healthy subjects control group

In a parallel prospective study, asymptomatic healthy subjects with no history of cancer were recruited to provide data on hPG<sub>80</sub> blood levels, constituting the control group. Their characteristics were matched to those of recruited cancer patients for age and gender. The
exclusion criteria included: any known personal history
of cancers, or pre-cancerous lesions such as atypical
dysplasia, in situ carcinomas; active smoking or previ-
ous smoking habit > 5 Pack-Year); a medical daily treat-
ment (except for contraceptive, endocrine treatment for
menopause, anti-hypertensive treatment, psychotropic
treatment and anti-lipemic treatment provided that the
inherent disease had been under control for at least 3
months). All patients gave their consent and signed the
informed consent. Ethical approval was granted by the
French health authority ANSM and the ethics commit-
tee CPP (ID-RCB N°0.2021-A02356-35). All Protocol
amendments were approved by the same authorities and
explained in the protocol.

Centralized hPG<sub>80</sub> level measurements in the blood
samples

hPG<sub>80</sub> blood levels were centrally measured in the mul-
tisite laboratory of medical biology and pathology, in the
"Cancer Pathology Medical Unit" of Biochemistry and
Molecular Biology department of Lyon University Hos-
pital. The ELISA DxPG<sub>80</sub>-Lab kit (Biodena Care, Grabels,
France) was used to measure hPG<sub>80</sub> levels in all blood
samples according to the manufacturer instructions
described previously [16]. Briefly, the limit of Detection
(LoD) was set at a hPG<sub>80</sub> concentration of 1.2 pM and
the limit of Quantitation (LoQ) at a hPG<sub>80</sub> concentra-
tion of 3.3 pM. The inter- and intra-assay coefficients of
variation (CV%) were below 10%. No cross-reactivity was
detected with gastrin-17, Gastrin-Gly or CTFP (C-Termi-
nus Flanking Peptide). No cross-reactivity was detected
with other blood biomarkers such as CA125, CEA or
PSA. No interference was detected with chemicals and
endogenous compounds such as SN-38, 5-FU or triglyc-
erides, cholesterol or hemoglobin.

Objectives and statistical analyses

The primary objective was to characterize the cancer
diagnostic accuracy of hPG<sub>80</sub> assay at baseline, before
any treatment, in cancer patients with respect to control
cases, using Receiver Operating Characteristics (ROC)
curves. Considering 421 enrolled cancer patients (pooled
cancer cohorts) and 330 asymptomatic healthy subjects,
the expected precision for a 95% confidence interval of
the area under the ROC curve of the hPG<sub>80</sub> was 0.022 for
an expected AUC of 0.9. Calculations were performed
assuming that plasma biomarker value distribution
would follow an exponential distribution in the diseased
and non-diseased subjects [17]. Under the same assump-
tions (AUC = 0.9, exponential distribution of the hPG<sub>80</sub>),
the large cohort of 50 patients will enable to obtain a pre-
cision of 0.06 for the 95% confidence interval of this area.

The secondary objectives were (1) to assess the influ-
ence of patient covariates (renal function, liver function,

age, CRP...) on hPG₈₀ values before any treatment as
well as during multi-modal treatments, during follow
up and at relapse; (2) to explore the variability of hPG₈₀
after meals (nycthemeral cohort), and after surgery; (3)
to characterize the diagnostic accuracy by cancer origin
within the large cohorts, and explore the hPG₈₀ concen-
tration associated with a 90% specificity; (4) to estimate
the prognostic value of hPG₈₀ modeled kinetics for recur-
rence free, progression free and overall survivals; and
(5) to characterize the kinetics of hPG₈₀ during multi-
modal treatments (chemotherapy, surgery, radiotherapy,
other...), during follow-up and at relapse, in localized or
metastatic cancers.

Five main amendments were made between June 2018
and January 2023. Two of them introduced significant
changes in the composition of cohorts to optimize the
recruitment rate (e.g. suppression of a melanoma cohort
replaced by a NSCLC cohort treated with immunother-
apy; increase of the head-and-neck cancer cohort up to
50 patients; suppression of the “weekly cohort” meant
to assess the hPG<sub>80</sub> kinetics on a weekly basis during the
first month of treatment) (Suppl Material).

As a consequence, the number of patients to enroll
increased up to 420 (compared to the 410 initially
planned).

In the present manuscript, we present the primary
objective, and some secondary objectives related to the
baseline hPG<sub>80</sub> values.

Differences in hPG₈₀ levels between cases and con-
trols were evaluated using the Wilcoxon rank-sum test.
The diagnosis discriminative accuracy of hPG₈₀ levels in
patients with cancer compared to asymptomatic healthy
subjects was quantified by the Area under the ROC curve
(AUC) with its 95% confidence interval. The correlation
between hPG₈₀ and baseline characteristics was quanti-
fied by the Spearman correlation coefficient. Analyses
were performed using SAS version 9.4 and R version
4.1.2. The level of significance was set at *P* < 0.05.

Results

Clinical characteristics of the enrolled cancer patient cases
and asymptomatic healthy subjects

Between December 2018 and June 2022, 421 patients
out of 506 patients enrolled were assessable as per study
criteria (Fig. 2). The main characteristics of assessable
patients are presented in Table 1. The median age was
65.6 [IQR: 58.2–72.9], and the gender distribution was
59.6% men/ 40.4% women.

Between January and April 2022, 330 asymptomatic
healthy subjects were enrolled in parallel, with matched
characteristics to cancer cases for age and gender
distribution.

Compared to asymptomatic healthy subjects, the can-
cer patients had statistically higher baseline value of
CANCER PATIENT COHORT

<figure><img src="image_7.png" /><figcaption>**Fig. 2** Flowchart of ONCOPRO study</figcaption></figure>

HEALTHY SUBJECT COHORT

<figure><img src="image_8.png" /></figure>

hPG$_{80}$ when all cohorts were pooled together (median, 3.8 [IQR: 1.0-11.1] pM vs. 1.9 [IQR: 0.6-4.2] pM, P < 0.0001) (Fig. 3A). The area under the ROC curve was 0.63 (95% CI = [0.59-0.67]), and was statistically different from 0.5 (Fig. 3B).

hPG$_{80}$ concentrations were not homogenously elevated in all the cohorts (Suppl Fig. 2, and Suppl Fig. 3). As shown on the Fig. 4, displaying the baseline values of hPG$_{80}$ in the large cohorts, the patients with lung cancers and with HCC had the highest baseline titers.

There was a significant and weak correlation between increasing patient age and higher hPG$_{80}$ concentrations in both the cancer patient cases ($R$ = 0.24, $P$ < 0.001) (Suppl Fig. 4) and asymptomatic healthy subjects ($R$ = 0.22, $P$ < 0.001) (Suppl Fig. 5). Men tended to have higher hPG$_{80}$ concentrations compared to women, but the difference was not statistically significant (median, 4.2 [IQR: 1.1-13.4] vs. 3.4 [IQR: 0.7-9.9] pM, $P$ = 0.087) (Suppl Fig. 6).

No clinically relevant correlation was found between hPG$_{80}$ concentrations and liver function (ALT, $R$ = 0.03, $P$ = 0.51; AST, $R$ = 0.11, $P$ = 0.029; bilirubin, $R$ = -0.02, $P$ = 0.67), renal function based on serum creatinine levels ($R$ = 0.23, $P$ < 0.001), or inflammatory status assessed by CRP ($R$ = 0.11, $P$ = 0.14) (Suppl Figs. 7 to 9).

In the nychthemeral cohort, 15 patients were enrolled, including 5 patients with breast cancers, 4 with renal cancers, 4 with bladder cancers, 1 with ovarian cancer, and 1 with endometrial cancer. hPG$_{80}$ concentration was not impacted by meal intakes (Suppl Fig. 10). In the post-operative cohort, 12 patients were enrolled, exclusively head-and-neck cancers. Out of 6 patients with elevated

pre-operative hPG$_{80}$ > LoQ, a decline to concentration < LoQ was observed in 5 patients 2 h post-surgery (83%) (Suppl Fig. 11).

Diagnostic value of hPG$_{80}$ in the large cohorts of interest

Among the 6 large cohorts, the diagnostic accuracy of hPG$_{80}$ was particularly more promising in patients with lung cancers and HCC aged > 1st quartile (58 years on the cancer population), an age at which cancer risk increases.

Indeed, in 56 patients with NSCLC or SCLC cancers aged > 58 years, the ROC AUC was 0.75, 95% CI = [0.67-0.82] with a 49% sensitivity and 88% specificity for hPG$_{80}$ > 7.73 pM (Fig. 5A). hPG$_{80}$ was found elevated in the 5 patients with early stage I disease (median, 26.5 pM [IQR: 8.1-28.5]). In the NSCLC cancer patients from the non-curative cohort where the data were available, the baseline value of hPG$_{80}$ was not significantly different according to the smoking status (active, $n$ = 20; former, $n$ = 22; never, $n$ = 5, $P$ = 0.83), or the molecular status regarding TP53, K-RAS, or B-RAF mutations in the NSCLC cancer patients on tumor tissue or circulating tumor DNA (Suppl Tables 1, and Suppl Figs. 12 and 13).

Similarly, in the 47 patients with HCC aged > 58 years, the ROC AUC was 0.74, 95% CI = [0.65-0.83], with a 51% sensitivity and 88% specificity for hPG$_{80}$ > 7.73 pM (Fig. 5B). The diagnostic value of AFP was overlapping compared to hPG$_{80}$ (ROC AUC, 0.77, 95% CI = [0.69-0.85]) (Suppl Fig. 14). Among the 39 patients with AFP-negative HCC at diagnosis (< 20 ng/mL), 51.2% patients had hPG$_{80}$ > 7.73 pM, meaning that hPG$_{80}$ could potentially compensate the false-negative AFP. The baseline value of hPG$_{80}$ was not different according to the
<table><caption>**Table 1** Characteristics of cancer patient cases</caption><thead><tr><th rowspan="2"></th><th rowspan="2">N</th><th colspan="3">Age</th><th rowspan="2">Gender males (%)</th></tr><tr><th>Median</th><th>Q1</th><th>Q3</th></tr></thead><tbody><tr><td><strong>TOTAL</strong></td><td><strong>421</strong></td><td></td><td></td><td></td><td><strong>251 (59.6)</strong></td></tr><tr><td>Breast cancer</td><td>50</td><td>54.3</td><td>46.1</td><td>67.1</td><td>0 (0%)</td></tr><tr><td>Curative intent</td><td>30</td><td>49.5</td><td>41.2</td><td>56.6</td><td>0</td></tr><tr><td>Non-curative intent</td><td>20</td><td>67</td><td>54.4</td><td>76.7</td><td>0</td></tr><tr><td>Renal cancer</td><td>50</td><td>61.6</td><td>55.5</td><td>71.2</td><td>33(66%)</td></tr><tr><td>Curative intent</td><td>25</td><td>62.6</td><td>54.1</td><td>69.8</td><td>17</td></tr><tr><td>Non-curative intent</td><td>25</td><td>60.4</td><td>58</td><td>72.3</td><td>16</td></tr><tr><td>Prostate cancer</td><td>50</td><td>69.6</td><td>64.8</td><td>74.6</td><td>50(100%)</td></tr><tr><td>Curative intent</td><td>25</td><td>66.1</td><td>64.6</td><td>69.7</td><td>25</td></tr><tr><td>Non-curative intent</td><td>25</td><td>73.3</td><td>65.5</td><td>77.8</td><td>25</td></tr><tr><td>Lung cancer</td><td>70</td><td>66.5</td><td>60.5</td><td>71.7</td><td>42(60%)</td></tr><tr><td>Curative intent</td><td>20</td><td></td><td></td><td></td><td>10 (50%)</td></tr><tr><td>Stage I</td><td>5</td><td>63.3</td><td>63.1</td><td>68.4</td><td>2</td></tr><tr><td>Stage II</td><td>1</td><td>66.3</td><td>66.3</td><td>66.3</td><td>1</td></tr><tr><td>Stage III</td><td>7</td><td>67.3</td><td>65.3</td><td>72.6</td><td>4</td></tr><tr><td>Unknown</td><td>7</td><td>72.8</td><td>67.4</td><td>74.1</td><td>3</td></tr><tr><td>Non-curative intent</td><td>50</td><td></td><td></td><td></td><td>32 (64%)</td></tr><tr><td>Stage III</td><td>1</td><td>66.3</td><td>66.3</td><td>66.3</td><td>1</td></tr><tr><td>Stage IV</td><td>48</td><td>67.3</td><td>65.3</td><td>72.6</td><td>30</td></tr><tr><td>Unknown</td><td>1</td><td>69.9</td><td>69.9</td><td>69.9</td><td>1</td></tr><tr><td>Hepatocellular Carcinoma</td><td>50</td><td>71.4</td><td>64.5</td><td>75</td><td>39(78%)</td></tr><tr><td>Curative intent</td><td>40</td><td>71.9</td><td>65</td><td>75.5</td><td>31</td></tr><tr><td>BCLC score 0</td><td></td><td>60.9</td><td>59.7</td><td>62.1</td><td>2</td></tr><tr><td>BCLC score A</td><td></td><td>72.3</td><td>65.5</td><td>748</td><td>20</td></tr><tr><td>BCLC score B</td><td></td><td>75</td><td>69.1</td><td>767</td><td>9</td></tr><tr><td>Non-curative intent</td><td>10</td><td>66.9</td><td>62.9</td><td>71.9</td><td>8</td></tr><tr><td>BCLC score B</td><td></td><td>64.1</td><td>62.6</td><td>69.7</td><td>7</td></tr><tr><td>BCLC score C</td><td></td><td>71.9</td><td>71.9</td><td>71.9</td><td>1</td></tr><tr><td>Head and Neck cancer</td><td>50</td><td>63.0</td><td>56.9</td><td>70.2</td><td>36(72%)</td></tr><tr><td>Curative intent</td><td>30</td><td>62</td><td>56.3</td><td>70.2</td><td>19</td></tr><tr><td>Non-curative intent</td><td>20</td><td>63.2</td><td>58.4</td><td>69.9</td><td>17</td></tr><tr><td>Gastric cancer</td><td>10</td><td>75.3</td><td>64.9</td><td>79.6</td><td>7(70%)</td></tr><tr><td>Colorectal cancer</td><td>10</td><td>61.3</td><td>56.9</td><td>70.7</td><td>4(40%)</td></tr><tr><td>Pancreatic Cancer</td><td>10</td><td>67.7</td><td>56</td><td>74.5</td><td>6(60%)</td></tr><tr><td>Ovarian Cancer</td><td>10</td><td>66.6</td><td>46.9</td><td>69.2</td><td>0(0%)</td></tr><tr><td>Glioblastoma</td><td>10</td><td>64.4</td><td>55.6</td><td>72.2</td><td>7(70%)</td></tr><tr><td>Endometrial cancer</td><td>10</td><td>69.7</td><td>65.6</td><td>73.3</td><td>0(0%)</td></tr><tr><td>Bladder Cancer</td><td>10</td><td>69.7</td><td>59.5</td><td>73</td><td>5(50%)</td></tr><tr><td>Superficial oeso-gastric cancer</td><td>11</td><td>73.3</td><td>63.9</td><td>81.4</td><td>11(100%)</td></tr><tr><td>Diffuse large B-cell lymphoma</td><td>10</td><td>67.5</td><td>64</td><td>72.2</td><td>10(100%)</td></tr><tr><td>Thyroid cancer</td><td>10</td><td>34.3</td><td>32.5</td><td>45.3</td><td>1(10%)</td></tr></tbody></table>

Barcelona-Clinic Liver Cancer (BCLC)

Barcelona-Clinic Liver Cancer (BCLC) score (0, n = 2; A, n = 28; B, n = 19; C, n = 1; P = 0.59) (Suppl Fig. 15), or alcohol consumption (No, n = 38; Yes, n = 12; P = 0.68).

In other large cohorts where serum biomarkers are routinely used—such as PSA for prostate cancer and CA 15-3 or CEA for breast cancer—the added diagnostic value of hPG<sub>80</sub> was also evaluated. In prostate cancer, the diagnostic performance of PSA alone was already high (ROC AUC: 0.97; 95% CI: 0.94–0.99), and the addition

of hPG<sub>80</sub> did not further improve the overall diagnostic accuracy (combined ROC AUC: 0.97; 95% CI: 0.94–0.99).

Similarly, in breast cancer, the diagnostic values of both CA 15-3 and CEA were limited (ROC AUC: 0.65; 95% CI: 0.55–0.75 and 0.61; 95% CI: 0.52–0.72, respectively), and the addition of hPG<sub>80</sub> provided no improvement (CA 15-3 + hPG<sub>80</sub>: ROC AUC 0.67; 95% CI: 0.57–0.76; CEA + hPG<sub>80</sub>: ROC AUC 0.61; 95% CI: 0.52–0.72).
<figure><img src="image_9.png" /><figcaption>**Fig. 3** Diagnostic value of hPG<sub>80</sub> in cancer patient cases compared to asymptomatic healthy subjects. (A) Box-plots of hPG<sub>80</sub> concentration. (B) ROC curve area under the curve</figcaption></figure>

Discussion

The development of multi-cancer early detection (MCED) tests capable of screening for multiple cancers from a simple blood sample has become a major area of interest within the scientific community [18, 19]. Most of these assays are based on circulating tumor DNA. Their clinical validity and utility have been assessed in large prospective clinical trials. The DETECT-A study enrolled approximately 10,000 women without a known cancer diagnosis to evaluate a panel of 16 genes using circulating cell-free DNA (cfDNA), along with 8 circulating proteins, followed by confirmation through positron emission tomography (PET)-computed tomography (CT) scanning. The results demonstrated a sensitivity of 15.6% with a specificity of 99.6% [20]. Similarly, the PATHFINDER study assessed a targeted methylation sequencing assay using cfDNA, detecting cancer signals in 1.4% (92/6621) of prospectively recruited participants, with a specificity of 99.1% and a positive predictive value (PPV) of 38.0% [21]. Other prospective clinical trials are currently underway to evaluate not only the benefit-to-harm ratio, but also the cost-effectiveness of these expensive MCED technologies which is still unclear [22–25].

In this context, hPG<sub>80</sub> has been developed as a novel blood-based multi-cancer diagnostic biomarker, offering significant advantages over existing assays due to its simpler implementation. The hPG<sub>80</sub> assay, based on an ELISA test, can be performed in any standard medical laboratory, making it a far more cost-effective option [16]. Following the highly promising results from

several retrospective studies demonstrating the diagnostic potential of hPG<sub>80</sub> across multiple cancer types, evaluating its clinical utility through a prospective case-control study was warranted [4, 10].

Conducting a large-scale prospective study involving nearly 500 patients across 16 different cancer types and 17 cancer departments within a single institution was a significant organizational achievement. Centralized coordination and standardized hPG<sub>80</sub> measurements were implemented to ensure uniformity in both data collection and procedural execution, overcoming the logistical challenges of such a complex study design. The results indicate that hPG<sub>80</sub> levels were significantly higher in cancer patients compared to asymptomatic healthy subjects, and this difference was not attributable to altered renal or liver function, nor to inflammatory status. We conclude that the elevated hPG<sub>80</sub> levels are likely attributable to the presence of cancer. The post-surgery decrease of hPG<sub>80</sub> observed in 83% of patients with initial high levels would support this assumption, as previously reported in glioblastoma and peritoneal carcinomatosis [4, 15]. Although the diagnostic discriminative accuracy was statistically different from 0.5, it was lower than anticipated based on retrospective studies when all cancer cases were evaluated [4, 10].

In this study, the most clinically relevant findings were observed in patients over 58 years of age (age > first quartile in our cohort) with lung cancer and hepatocellular carcinoma (HCC). Indeed, the diagnostic accuracy was high for lung cancer patients, with a ROC curve AUC
<figure><img src="image_10.png" /><figcaption>**Fig. 4** hPG<sub>80</sub> concentrations in the large cancer patient cohorts and the asymptomatic healthy subjects. The number of patients is indicated on the x-axis</figcaption></figure>

of 0.75. This high diagnostic accuracy of hPG<sub>80</sub> may be hypothetically explained by the robust activation of the WNT signaling pathway, which typically occurs through the overexpression of WNT ligands (e.g., Wnt1, Wnt3a) or their receptors (e.g., Frizzled-7), leading to downstream transcriptional activation of WNT target genes, including GAST [26]. In our study, at a cutoff value of 7.73 pM, the specificity was 90%, with a sensitivity of 49%. As a consequence, hPG<sub>80</sub> could be a valuable addition to the current screening strategies, complementing low-dose CT scans. By improving specificity in patients with suspicious lung lesions, hPG<sub>80</sub> may be an interesting biomarker for non-invasive diagnosis of lung nodules [27].

For hepatocellular carcinoma (HCC), the diagnostic accuracy was similarly high, with an ROC curve AUC of 0.74. This observation is consistent with the literature, as HCC is frequently characterized by somatic mutations in CTNNB1, the gene encoding β-catenin. These mutations result in constitutive activation of β-catenin and subsequent uncontrolled transcription of downstream WNT target genes [28]. At a cut-off of 7.73 pM, the specificity

was 90%, with a sensitivity of 48%. Moreover, over 50% of patients with AFP-negative HCC at diagnosis (AFP < 20 ng/mL) exhibited elevated hPG<sub>80</sub> concentrations. This finding aligns with previous reports indicating that AFP is non-informative in 50–70% of HCC cases, and that hPG<sub>80</sub> is independently elevated in more than 60% of these patients [14, 29]. These results support the potential role of hPG<sub>80</sub> as a complementary biomarker to AFP for improving the diagnostic accuracy in HCC.

We believe that hPG<sub>80</sub> could be a valuable complement to AFP measurement in patients suspected of having HCC [29].

While the diagnostic value of hPG<sub>80</sub> shows promise for lung cancer and HCC in patients aged 58 years or older, the study was limited by a small sample size and lower diagnostic accuracy in patients with other cancer types. These findings suggest that the spectrum of cancers detectable by the hPG<sub>80</sub> assay is likely narrower than initially anticipated when it was developed as a multi-cancer early detection biomarker. Similar to other MCED tests currently in development, the accuracy of cancer detection varies depending on the type of cancer [19, 23].
<figure><img src="image_11.png" /><figcaption>**Fig. 5** Diagnostic value of hPG<sub>80</sub> in patients with lung cancer (A) and HCC (B) aged 58 years or older, compared to asymptomatic healthy subjects</figcaption></figure>

These outcomes will help guide the further develop-
ment of hPG<sub>80</sub> as a biomarker. The exclusion of patients
with lung cancer and HCC younger than the first quar-
tile age (58 years) is open to criticism but is justified, as
the cancer risk in younger patients is obviously lower
and less relevant from a macroeconomic perspective.
Some discrepancies were observed between our results
and those of retrospective studies for certain cancers.
For example, hPG<sub>80</sub> levels were low in the exploratory
glioblastoma cohort (median, 0.3 pM (IQR 0.00-1.85),
n = 10) compared to the retrospective study published
by Doucet et al. where the median hPG<sub>80</sub> concentra-
tion was 5.37 pM (IQR 0.00-13.90). These discrepancies
underscore the need for additional data, which is cur-
rently being collected from ongoing prospective studies.
Indeed, blood hPG<sub>80</sub> concentrations are being measured
in patients with high-grade glial tumors (PROGLIO,
NCT05157594), and those with neuroendocrine tumors
(PRO-NEM1, NCT06430021; and NCT05724108). The
high variability of hPG<sub>80</sub> suggests that this biomarker
should likely be developed as a complementary test
rather than as a standalone tool, to overcome the limited
sensitivity and specificity of existing screening meth-
ods. The prognostic value of baseline hPG<sub>80</sub> regarding
progression-free survival and overall survival— a sec-
ondary endpoint in the ONCOPRO study—could not be
assessed due to the insufficient number of events. This
will be addressed in future reports. Additionally, the
present study did not evaluate the monitoring value of
hPG<sub>80</sub>, which is a key secondary endpoint of ONCOPRO.

This will be assessed on a cohort-by-cohort basis. Finally,
other multi-cancer early detection biomarkers, such as
ctDNA, nucleosomes, and epigenetic circulating markers
will be assessed on the ONCOPRO samples to investigate
their diagnostic and monitoring potential, as well as their
possible complementarity with hPG<sub>80</sub>.

In conclusion, the ONCOPRO case/control prospective
study was successfully conducted across 17 departments
within a single institution, with centralized coordina-
tion. hPG<sub>80</sub> concentrations were effectively measured to
assess its diagnostic value in patients with 16 different
types of cancer. The study confirmed that hPG<sub>80</sub> levels
are significantly higher in cancer patients compared to
asymptomatic healthy subjects, independent of renal or
liver function and inflammatory status. Despite some
variability, the diagnostic accuracy of hPG<sub>80</sub> was statis-
tically significant. The most clinically relevant findings
were observed in patients with lung cancer and HCC,
where high ROC AUC values and high specificity suggest
that hPG<sub>80</sub> could serve as a complementary biomarker to
existing screening tools in these cancers.

**Supplementary Information**

The online version contains supplementary material available at https://doi.org/10.1186/s40364-025-00793-z.

Supplementary Material 1
**Acknowledgements**

The authors would like to thank all patients and their families, the
investigators, study nurses, pharmacists, pathologists and the Biodena Care
team.

**Author contributions**

Conception and design: All authors. Administrative support: BY, DJ, AP, BV, SC. Provision of study materials or patients: BY, SC, PC, LB, PP, AD, MG, PM, GL, CAP, FD, MP, LK, JCL, OG, PAB, MC. Collection and assembly of data: CLJ, FS, MP, AC, SC, LP. Data analysis and interpretation: CLJ, FS, MP, AC, DJ, AP, BV, SC, LP. Manuscript writing: All authors. Final approval of manuscript: All authors.

**Funding**

Biodena Care funded the study and provided the assays.

**Data availability**

No datasets were generated or analysed during the current study.

**Declarations**

**Ethics and consent to participate declarations**

All patients gave their consent and signed the informed consent. The protocol, the written information sheet and the consent form for the study obtained a favorable opinion from to ethics committee Ile de France VI on October 3rd 2018. This notification was transmitted to the French health authority ANSM. All Protocol amendments were approved by the same ethic committee.

**Consent for publication**

Not applicable.

**Competing interests**

DJ, AP, and BV are employed by Biodena Care (Progastrin Manufacturing). All the other coauthors have no conflict of interest to disclose.

**Author details**

Received: 11 March 2025 / Accepted: 25 May 2025

Published online: 01 July 2025

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<footnote n="1">¹Lyon University Hospital, IC-HCL, Lyon University, Lyon, France</footnote>

<footnote n="2">²Medical Oncology, IC-HCL, EPSILYON, Université Claude Bernard, CICLY, Lyon 1, Univ Lyon 1, Lyon, France</footnote>

<footnote n="3">³Pneumology, Lyon-Sud Hospital, URCOT, CICLY, Lyon, France</footnote>

<footnote n="4">⁴Head and Neck Surgery, Croix Rousse Hospital, CICLY, Lyon, France</footnote>

<footnote n="5">⁵Urology, Edouard Herriot Hospital, Lyon, France</footnote>

<footnote n="6">⁶Hépatogastroentérologie et Endoscopie Digestive, E Herriot Hospital, Lyon, France</footnote>

<footnote n="7">⁷Hepatology and Gastroenterology, Croix-Rousse Hospital, Lyon, France</footnote>

<footnote n="8">⁸Center for Innovation in Cancerology of Lyon (CICLY) ER 3738, Faculty of Medicine and Maieutic Lyon Sud, Claude Bernard University Lyon I (UCBL1), Institut of Pharmaceutical and Biological Sciences of Lyon (ISPB), Oullins 69921, France</footnote>

<footnote n="9">⁹Gynecology and Obstetrics, Hopital Femme Mere Enfant, Lyon, France</footnote>

<footnote n="10">¹⁰Neurology, East Hospital, Lyon, France</footnote>

<footnote n="11">¹¹Hematology, Lyon-Sud Hospital, Lyon, France</footnote>

<footnote n="12">¹²Endocrine Surgery, RESHAPE, Lyon, France</footnote>

<footnote n="13">¹³Visceral and Digestive Surgery, Lyon-Sud Hospital, CICLY, Lyon, France</footnote>

<footnote n="14">¹⁴Hepatology and Gastroenterology, Lyon-Sud Hospital, Lyon, France</footnote>

<footnote n="15">¹⁵Service de Biostatistique, Pole Santé Publique, Lyon, France</footnote>

<footnote n="16">¹⁶Université Claude Bernard Lyon 1, LBBE, UMR 5558, CNRS, VAS, Villeurbanne, France</footnote>

<footnote n="17">¹⁷Progastrin Manufacturing, Cap Sigma, 1642 rue de la Valsière, Grabels 34790, France</footnote>

<footnote n="18">¹⁸LBMMS, Université Claude Bernard Lyon 1, Univ Lyon 1, CICLY, Lyon 69003, France</footnote>

<footnote n="19">¹⁹Gynecology and Obstetrics, Lyon-Sud Hospital, CICLY, Lyon, France</footnote>
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