Evaluation of antiangiogenic efficacy in advanced hepatocellular carcinoma: Biomarkers and functional imaging

Table 1 Association between baseline circulating markers and outcome in patients treated with various treatments for hepatocellular carcinoma

Ref.	Markers	Patients (n)	Study design	Treatment	Level values	Clinical impact	Conclusion/comments
Schoenleber et al[85]	VEGF-A	1018	Systemic review and meta-analysis including only serum-based studies	Various (surgery, LRT and systemic therapies)	High serum VEGF level	Poorer OS	Serum VEGF method detection varied among studies
						Poorer DFS	Serum VEGF levels seem more reliable than tissue VEGF for HCC prognosis
Poon et al[115]	bFGF	88	Prospective	Surgery	High serum level > 10.8 pg/mL	Larger tumor > 5 cm	High bFGF serum level before surgery was shown to be an independent factor of early recurrence. No further studies confirmed these findings
						Venous invasion
Vejchapipat et al[105]	HGF	55	Retrospective	BSC	High level (≥ 1.0 ng/mL	Advanced pTNM stage Poorer prognosis Poorer OS	Although a control group was included, results of this small cohort study need confirmation in larger prospective analysis
Chau et al[104]		40	Retrospective	Resection	High portal and serum HGF level (> 699 pg/mL)	Multiple tumor	One limit of this study were the feasibility in routine of intraoperative puncture of the portal vein was difficult
						Poorer prognosis
Mizuguchi et al[106]		100	Retrospective	Resection	High serum level (≥ 0.35 ng/mL)	Postoperative complications	No correlation was observed between HGF level and RFS
						Poorer OS
Kaseb et al[87]	IGF-1	288	Prospective	Various	Low plasma level (26 ng/mL)	High Child-Pugh score	The authors proposed that IGF-1 plasma level to be integrated into the BCLC staging system to predict OS for personal management in patients with HCC. This proposal was not yet adopted in clinical practice
						High AST level
						High tumor size
						Multiple tumor
						Vascular invasion
						Poorer OS

BCLC: Barcelona clinic liver cancer; bFGF: Basic fibroblast growth factor; BSC: Best supportive care; DFS: Disease-free survival; HGF: Hepatocyte growth factor; IGF-1: Insulin growth factors 1; LRT: Loco-regional treatment; OS: Overall survival; RFS: Recurrence-free survival; VEGF: Vascular endothelial growth factors.

Table 2 Prognostic value of baseline circulating factors in patients treated with systemic therapies including antiangiogenic agents for advanced hepatocellular carcinoma

Ref.	Marker	Patient (n)	Study type	Treatment	Levels values	Prognostic value	Conclusion/comments
Kaseb et al[86]	VEGF-A	394	Systemic review including only serum or plasma-based studies	Various (AA alone or combined with CT)	High serum or plasma level	Poorer outcome	Plasma VEGF seemed more relevant than serum VEGF as prognostic factor for HCC
Llovet et al[63]		490	Prospective phase III trial	Sorafenib vs placebo	High plasma level (> 101 pg/mL)	Poor OS	The VEGF level was a prognostic factor for all patient's cohort but surprisingly it did not affect prognosis in patients receiving sorafenib. Moreover, the VEGF level did not predict response
						Better clinical/ demographic parameters
Llovet et al[63]	HGF	251	Prospective phase III trial	Sorafenib vs placebo	High plasma level	Poorer OS	HGF was a prognostic factor for the entire cohort. However, it does not predict response to sorafenib (only a nonsignificant trend)
Miyahara et al[112]	Ang2	30	Prospective?	Sorafenib	High serum level	Shorter PFS Progressive disease	The small cohort and the lack of control arm hamper conclusion on the role of Ang2 as predictive of response to sorafenib
Llovet et al[63]		490	Prospective phase III trial	Sorafenib vs placebo	High plasma level (> 6043.5 pg/mL)	Poorer OS Better clinical/demographic parameters	Ang2 was shown to be a prognostic factor in HCC but did not predict response to sorafenib
Llovet et al[63]	c-KIT	245	Prospective phase III trial	Sorafenib vs placebo	High plasma level (> 11.3 ng/mL)	Trend to a better OS	Soluble c-KIT was shown to be a prognostic factor for HCC. However, it showed only a nonsignificant trend to predict response to sorafenib
						Trend to better TTP
						Better clinical/demographic parameters
Llovet et al[63]	IGF-2	254	Prospective phase III trial	Sorafenib vs placebo	High plasma level (> 797.7 ng/mL)	Better OS	IGF-2 was shown to be prognostic factor in HCC but did not predict response to sorafenib
						Better clinical/demographic parameters
Shao et al[126]	CEC/CECP	40	Prospective	Sorafenib + CT	High CECP level	Poorer PFS Poorer OS	The predictive value of CECP was not confirmed in further investigations

AA: Antiangiogenic; Ang2: Angiopoietin 2; CEC: Circulating endothelial cells; CECP: Circulating endothelial cell progenitors; c-KIT: Stem-cell factor receptor; CT: Chemotherapy; HCC: Hepatocellular carcinoma; HGF: Hepatocyte growth factor; IGF-2: Insulin growth factor 2; OS: Overall survival; PFS: Progression-free survival; TTP: Time to progression; VEGF: Vascular endothelial growth factors.

Table 3 Treatment-induced changes in biomarkers levels and association with outcome in patients with hepatocellular carcinoma

Ref.	Marker	Patient (n)	Study design	Treatment	Marker treatment-induced changes	Impact value	Comments
Llovet et al[63]	VEGF-A	490	Prospective phase III trial	Sorafenib vs placebo	Increase	No association with OS and ORR	The VEGF-A could serve as pharmacodynamic marker of exposure to sorafenib but did not have prognostic or predictive value
Harmon et al[93]		37	Prospective single arm phase II	Sunitinib	Reversible Increase	Better DCR	Inconsistent results were observed in these trials. The value of VEGF-A to predict response to sunitinib could be confirmed in larger trial
						Better PFS
						Better OS
Zhu et al[91]	VEGF-C	34	Prospective single arm phase II	Sunitinib	Sustained increase	No predictive value
Harmon et al[93]		37	Prospective single arm phase II	Sunitinib	Decrease	Better DC	The predictive value of VEGF-C was not shown for sorafenib probably because of its limited action against the VEGFR-3
						Better ORR
Harmon et al[93]	sVEGFR-2/ sVEGFR-3	37	Prospective single arm phase II	Sunitinib	Reversible decrease	Better OS (for sVEGFR-2)	The small cohort did not allow a definite conclusion
Zhu et al[91]		34	Prospective single arm phase II	Sunitinib	Decrease	No predictive value
Llovet et al[63]	Ang2	490	Prospective phase III trial	Sorafenib vs placebo	No significant change (for sorafenib) Increase (for placebo)	Shorter TTP Shorter OS (for patients who experienced increase)	Ang2 was probably a prognostic biomarker than predictive of response to sorafenib
Llovet et al[63]	c-KIT	245	Prospective single arm phase II	Sorafenib vs placebo	Decrease (sorafenib) no change (placebo)	No predictive value	Tumor expression of KIT was considered as low in HCC, and the role of soluble KIT remains unclear
Zhu et al[91]		34	Prospective single arm phase II	Sunitinib	Decrease	Better TTP
						Better OS
Harmon et al[93]		37	Prospective single arm phase II	Sunitinib	Decease	Better TTP
Boige et al[98]	CEC	36	Prospective single arm phase II	Bevacizumab	Early increase	Better OR	CEC level was not associated with prognosis in this study. However, it could predict response to bevacizumab. The rarity of CEC level and non-standardized measurement methods limited the use of CEC as a predictive marker of response to treatment in HCC
						Better DCR
Zhu et al[91]	CECP	34	Prospective single arm phase II	Sunitinib	Decrease	Progression

Ang2: Angiopoietin 2; CEC: Circulating endothelial cells; CECP: Circulating endothelial cell progenitors; c-KIT: Stem-cell factor receptor; DCR: Disease control; HCC: Hepatocellular carcinoma; ORR: Objective response; OS: Overall survival; PFS: Progression-free survival; sVEGFR: Soluble vascular endothelial growth factors receptor; TTP: Time to progression; VEGF: Vascular endothelial growth factors.

Table 4 Clinical side effects induced by sorafenib in patients with advanced hepatocellular carcinoma and association with outcome

Ref.	Side effect	Patients (n)	Study design	Impact on survival	Impact on other parameters	Predictive value
Otsuka et al[42]	Skin reaction	94	Retrospective	Better OS	No impact on ORR, DCR, and TTP	No
Vincenzi et al[45]		65	Retrospective	Trend to a better OS	Better DCR	Early skin toxicity could predict efficacy of sorafenib
					Better TTP
Di Costanzo et al[43]		65	Retrospective	Better OS	Not reported	Skin toxicity could predict survival
Shomura et al[44]		37	Retrospective	Better OS	Better DCR	Skin toxicity could predict efficacy
Reig et al[46]		147	Prospective	Better OS	Better TTP	Early skin reaction could predict efficacy of sorafenib and survival
Otsuka et al[42]	Arterial hypertension	94	Retrospective	No impact	No impact	No
Estfan et al[55]		41	Retrospective	Better OS	Trend to better TTP

DCR: Disease control rate; OS: Overall survival; ORR: Objective response rate; TTP: Time to progression.

Table 5 Prognostic value of baseline and increase of alpha-fetoprotein for hepatocellular carcinoma in patients who underwent resection or transplantation

Ref.		Patient (n)	Study design	Treatment	Level values	Impact value	Comments
Liu et al[57]	AFP	2034	Retrospective	Resection (79.2%) NA (20.8)	High AFP levels (> 20 μg/L)	Large tumors (≥ 10 cm)	This large cohort study showed that High AFP level was associated with poor prognosis and poor clinicopathological features of HCC
						Higher vascular invasion
						Lower differentiated tumor
Wang et al[139]		160	Retrospective	Resection	High AFP level (> 4000 UI/L)	Shorter median TTR	In this study, the value of AFP levels to predict recurrence is limited since only a few numbers of patients (9%) have AFP level higher than the cutoff level
Ma et al[58]		108	Retrospective	Resection	High AFP level (> 20 ng/mL)	Lower differentiated tumor	This study demonstrated the negative impact of high AFP levels on surgery benefit and the need to closely screen patients after resection for recurrence
						Higher vascular invasion
						Higher postoperative 2-yr recurrence rate
						Lower 24-mo survival rate
Ikai et al[59]		12118	Japanese nationwide	Resection	High AFP level (≥ 20 ng/mL)	Worsen OS after surgery	This large cohort study showed better outcome of patient resected for HCC in the last decade but the persistence of the negative impact of high AFP level on prognosis
			Analysis
			Comparative study
Vibert et al[60]		153	Retrospective	LT	AFP level increase > 15 μg/L per month	Lower OS	This study showed the negative impact on the outcome of AFP levels increases in patients undergoing LT
						Lower RFS
						Higher recurrence rate
Hakeem et al[61]		12159	Systemic review	LT	AFP > 1000 ng/mL (based on the majority of study included in the review)	Poorer OS	The authors stressed the poor quality of previous studies and the need for high-quality evidence on outcomes to use AFP levels as a prognostic indicator for patients undergoing LT
						Poorer DFS
						Higher vascular invasion
						Poorer differentiated tumor
Duvoux et al[62]		972	Prospective/retrospective	LT	High AFP level	Tumor recurrence	A new score model including AFP level was proposed to select patients for LT
						Vascular invasion
						Poor differentiation

AFP: Alpha-fetoprotein; DFS: Disease-free survival; HCC: Hepatocellular carcinoma; LT: Liver transplantation; NA: Not available; RFS: Recurrence-free survival; TTR: Time to recurrence.

Table 6 Prognostic and predictive value of baseline or changes of alpha-fetoprotein level for patients with hepatocellular carcinoma treated with antiangiogenic therapies alone or combined with systemic therapies

Ref.		Patients (n)	Study design	Treatment	Level values	Clinical impact	Comments
Shim et al[160]	AFP	57	Retrospective	Sorafenib	High level ≥ 400 ng/mL	Shorter TTP	This study suffers from some limits: a retrospective study, a small cohort including only hepatitis B patients, short median follow-up duration, lack of correlation with OS or ORR
Shao et al[69]		72	Prospective	Various AA + CT	AFP response (> 20% decrease from baseline within the first four weeks)	Better DCR	The magnitude of AFP decline (20% or 50%) from baseline was not clearly defined. Similarly, the time point for evaluation of AFP level was not clear also (4 wk? 7 wk?). Limits: a small number of patients with heterogeneous treatment
						Better ORR
						Better PFS
						Better OS
Yau et al[70]		94	Retrospective	Sorafenib	AFP response (> 20% decrease from baseline within the first six weeks)	Clinical benefit rate	The cutoff value to define AFP response was inconsistent between various studies
						Better PFS
						Marginal better OS
Personeni et al[71]		85	Retrospective	Sorafenib	AFP response (> 20% decrease from baseline within the first six weeks)	Better DCR	The authors used the landmark method to limit the potential favorable outcome due to tumor features than to AFP response
						Better TTP
						Better OS
Køstner et al[72]		76	Retrospective	Sorafenib	AFP response (> 20% decrease from baseline within the first four weeks)	Better ORR	No correlation was observed between AFP response and OS probably because of the limited number of patients evaluated and the unusual poor OS seen in all cohort (5.4 mo)
Kuzuya et al[73]		48	Retrospective	Sorafenib	AFP response (decrease from baseline within 2 and 4 wk)	Better DCR	Limits of the study: retrospective design and the small number of patients included
						Better TTP
						Better OS
Nakazawa et al[74]		59	Retrospective	Sorafenib	AFP response (increase from baseline within four weeks)	Progressive disease	Limits of the study: a small number of patients was enrolled in this and retrospective study. No association between AFP level before treatment and tumor response was observed
						Shorter PFS
						Shorter OS
Llovet et al[63]		491	Prospective Phase III trial	Sorafenib vs placebo	High plasma level > 200 ng/mL	Poorer OS	The impact of baseline AFP on survival was observed in both groups of patients treated with placebo or sorafenib
Hsu et al[64]		53	Prospective single-arm Phase II trial	Sorafenib + mT/U	> 400 ng/mL	Poorer OS?	The prognostic value of baseline AFP level was shown only in univariate analysis and only score CLIP ≥ 3 was an independent prognostic factor of poor OS
Baek et al[65]		201	Retrospective	Sorafenib	≥ 400 ng/mL	Shorter FFS	Baseline AFP level, tumor size, PS, albumin and bilirubin levels were the independent factor associated with OS in this study
						Poorer OS
Lin et al[66]		156	Systemic review of the prospective phase II trials	Various systemic therapies	≥ 400 ng/mL	No impact	Limits of the study: heterogeneous population
Shao et al[119]		45	Pooled analysis of single-arm phase II trials	Sorafenib + mT/U and beva + C	> 400 ng/mL	No impact	This study especially focused on the impact of IGF factors on outcome and the small cohort analyzed limits the interpretation of the effect of AFP levels on survival

AA: Antiangiogenic; AFP: Alpha-fetoprotein; Beva: Bevacizumab; C: Capecitabine; CLIP: Cancer of the liver Italian program[161]; CT: Chemotherapy; DCR: Disease control rate; FFS: Failure-free survival; mT/U: Metronomic tegafur/uracil; ORR: Objective response rate; OS: Overall survival; PFS: Progression-free survival; PS: Performance status; TTP: Time to progression.

Table 7 Prognostic and predictive value of tissue biomarkers evaluated in hepatocellular carcinoma

Ref.	Marker	Patient (n)	Origin of specimen	Method assay	Quantification	Marker level	Clinical impact
Mitsuhashi et al[108]	Ang2	46	Resected specimens	RT-PCR and IHC	Quantitative	High tumor Ang2/1 ratio	Tumor portal vein invasion
							Large tumor
							Increase MVD
							Poor OS
Zhang et al[109]		38	Resected specimens	RT-PCR	No	High tumor Ang2/1 ratio	Large tumor
							Portal vein invasion
							Metastasis
Torimura et al[110]		59	Resected specimens (19) and Biopsy (40)	RT-PCR and IHC	Semi-quantitative	High tumor Ang2	Poor differentiated tumor
Abou-Alfa et al[127]	pERK	33	Biopsy before sorafenib	IHC	Semi-quantitative	High tumor pERK	Better TTP
Ozenne et al[128]		20	Biopsy before sorafenib	IHC	Semi-quantitative	High tumor pERK	No impact
Hagiwara et al[131]	JNK	39	Biopsy before sorafenib	IHC and Western Blot	Quantitative	High JNK tumor	Lower ORR
							Poorer TTP
							Poorer OS
Peng et al[134]	pVEGFR-2	35	Resected specimen before sorafenib	RT-PCR and IHC	Semi-quantitative	Low tumor expression	Poorer OS
Poon et al[84]	VEGF	60	Resected specimen	IHC and ELISA	Semi-quantitative	High tumor expression	Advanced HCC stage

Ang2: Angiopoietin 2; ELISA: Enzyme-linked immunoadsorbent assay; IHC: Immunohistochemistry; JNK: C-Jun N-Terminal Kinase; MVD: Microvessel density; ORR: Objective response rate; OS: Overall survival; pERK: Phosphorylated extracellular signal regulated kinase; pVEGFR: Phosphorylated vascular endothelial growth factors receptor; RT-PCR: Real-time polymerase chain reaction; TTP: Time to progression.

Table 8 Value of functional imaging in patients with hepatocellular carcinoma treated with antiangiogenic agents

Ref.	Imaging tools	Patients (n)	Study design	Treatment	Imaging findings and clinical impact	Conclusion/comments
Sugimoto et al[152]	DCE-US	37	Prospective	Sorafenib	Tumor vascularity decreases and blood volume within seven days trends towards better PFS and OS	These studies enrolled small cohort of patients hampering adequate interpretation. However, DCE-US remains a promising noninvasive imaging, but operator dependent, to predict response in patients with HCC treated with sorafenib and larger cohort of patients should be evaluated
Zocco et al[153]		28	Prospective	Sorafenib	An early decrease in AUC and increase of median transit time was associated with better PFS and OS
Zhu et al[91]	DCE-MRI	34	Prospective	Sunitinib	Decrease in vascular permeability was associated with better disease control	The decrease of vascular permeability induced by antiangiogenic agents seems to be a good predictive of tumor response and clinical benefit. These promising findings should be confirmed by largest cohort of patient
Hsu et al[156]		31	Prospective	Sorafenib + mT/U	A ≥ 40% decrease in vascular permeability with 14 d was associated with better PFS and OS
Lee et al[159]	FGD-PET	29	Retrospective	Sorafenib	SUV < 5.00 correlated with longer PFS and OS	Prospective studies are needed to evaluate the predictive value of the FDG-PET in HCC

AUC: Area under the time-intensity curve; DCE-US: Dynamic contrast-enhanced ultrasound; DCE-MRI: Dynamic contrast-enhanced magnetic resonance imaging; FGD-PET: 18F-fluorodeoxyglucose - positron-emission tomography; PFS: Progression-free survival; OS: Overall Survival; mT/U: Metronomic tegafur/uracil.