Successful Sequential Immunotherapy for Multiple Primary Malignancies (Unresectable Hepatocellular Carcinoma and Esophageal Squamous Cell Carcinoma).

Introduction
Lifestyle factors, such as smoking and excessive alcohol consumption, are well-established risk factors for hepatocellular carcinoma (HCC). They also increase the likelihood of secondary malignancies in other organs. The optimal management of patients with synchronous HCC and esophageal squamous cell carcinoma (ESCC) remains unclear because of the complexities associated with the dual tumor burden and treatment-related toxicities. We herein present a rare case of multifocal HCC and ESCC that was successfully treated with sequential immune checkpoint inhibitors (ICIs), resulting in significant tumor regression and sustained clinical benefits. The development of an optimal treatment strategy for patients with synchronous HCC and ESCC remains a significant clinical challenge. Although both malignancies may respond to immune checkpoint inhibitors, selecting an appropriate ICI regimen requires careful consideration of tumor characteristics, disease burden, and patient-specific factors.

Case Report
The patient was a 78-year-old man with a history of alcoholic liver disease who had been under follow-up at a local clinic since 2004. In October 2023, the patient developed appetite loss and decreased physical activity. Laboratory investigations revealed elevated liver enzyme and alpha-fetoprotein (AFP) levels. Computed tomography (CT) revealed multiple hepatic tumors, raising the suspicion of HCC. The patient was referred to our hospital for further evaluation and treatment.
His medical history included hypertension and type 2 diabetes mellitus, both of which remained untreated. He had no history of blood transfusions or allergies, and no significant family history. He had a 60-year smoking history of 12 cigarettes per day and an alcohol consumption history equivalent to 100-120 g pure ethanol per day.
The laboratory test results are listed in Table. His platelet count was 15.1×104/μL (reference range: 13-36×104/μL) and his glycated hemoglobin value was markedly elevated at 11.4% (reference range: 4.6-6.2%). His serum albumin level was 3.5 g/dL (reference range: 3.8-5.3 g/dL) and his total bilirubin level was 0.7 mg/dL (reference range: 0.4-1.5 mg/dL). The modified albumin-bilirubin (ALBI) score was -2.26, corresponding to grade 2b. Tests for hepatitis B virus surface antigen and hepatitis C virus antibodies were negative. Regarding tumor markers, AFP was 172 ng/mL (reference value: ≤10 ng/mL), and protein induced by vitamin K absence-II (PIVKA-II) was 786 mAU/mL (reference value: ＜40 mAU/mL). Cytokeratin 19 fragment (CYFRA) and squamous cell carcinoma antigen levels were within the normal limits.
Contrast-enhanced CT revealed features indicative of liver cirrhosis, including blunted liver margins, irregular hepatic surface, multiple liver masses with early enhancement (Fig. 1a, b), and subsequent washout (Fig. 1c, d), consistent with HCC. No obvious abdominal lymphadenopathy or distant metastasis was observed. Esophagogastroduodenoscopy (EGD) (Fig. 2) revealed a type 0-I tumor in the middle thoracic esophagus. Histopathological examination confirmed squamous cell carcinoma with programmed death-ligand 1 (PD-L1) immunohistochemistry, with an expression rate of ＜1%. On the basis of these findings, the patient was diagnosed with unresectable multifocal HCC and synchronous ESCC.
Given the differential progression status and prognosis of both malignancies, we prioritized HCC treatment and initiated systemic therapy with atezolizumab and bevacizumab in February 2024. After two cycles, contrast-enhanced CT demonstrated decreased enhancement of the hepatic tumors (Fig. 3a), and EGD revealed a reduction in the esophageal lesion (Fig. 3b), indicating a favorable response in both malignancies.
However, in April, the patient developed right upper limb motor dysfunction. Magnetic resonance imaging (MRI) of the brain revealed an acute cerebral infarction at the bifurcation of the left middle cerebral artery. Given the potential association between bevacizumab and cerebrovascular events, the treatment regimen was modified in May to include a combination of durvalumab and tremelimumab. Following this regimen change, there was a marked decrease in AFP and PIVKA-II (Fig. 4).
After seven cycles, contrast-enhanced CT demonstrated further regression of the multifocal HCC (Fig. 5a), and EGD confirmed complete resolution of the ESCC lesion (Fig. 5b). The patient has continued treatment without experiencing severe immune-related adverse events (irAEs).

Discussion
Systemic therapy for unresectable HCC (urHCC) has advanced significantly in recent years, with ICIs emerging as a new standard of care. The IMbrave150 trial demonstrated that the combination of atezolizumab and bevacizumab significantly prolonged overall survival and progression-free survival relative to sorafenib treatment (1). In addition, the HIMALAYA trial established the superiority of the STRIDE regimen (durvalumab plus tremelimumab) over sorafenib. Tremelimumab, a cytotoxic T-lymphocyte antigen-4 (CTLA-4) inhibitor, enhances T cell activation and potentiates antitumor immune responses (2).
This case represents a rare instance in which combination ICI therapy was effective for both urHCC and superficial ESCC. HCC and ESCC typically require distinct treatment strategies, and the efficacy of ICIs for synchronous malignancies remains largely unexplored. Notably, despite the absence of PD-L1 expression, durvalumab plus tremelimumab achieved remarkable tumor regression in both cancers. This suggests that ICI therapy may be a viable treatment option, irrespective of the PD-L1 expression status.
Multiple primary malignancies (MPMs) are defined as the occurrence of two or more distinct malignancies in the same patient, either synchronously or metachronously, accounting for approximately 0.7-11% of all cancer cases (3). The risk factors for HCC include chronic hepatitis B or C infection, obesity, smoking, and excessive alcohol consumption. Concurrent HCC and gastric cancer are particularly prevalent in Japan (4-9). Currently, there is no established standard first-line therapy for MPMs, and treatment selection is typically guided by the biological characteristics and prognosis of each malignancy.
Atezolizumab plus bevacizumab and durvalumab plus tremelimumab are both approved first-line treatments for urHCC, based on the IMbrave150 and HIMALAYA trials, respectively. Although no head-to-head trials have directly compared these regimens, atezolizumab plus bevacizumab has demonstrated a favorable trend in the overall response rate (30.0% vs. 20.1%), median overall survival (19.2 vs. 16.4 months), and progression-free survival (6.9 vs. 3.8 months). In the present case, the patient had multiple bilobar lesions and a substantial intrahepatic tumor burden. Therefore, atezolizumab plus bevacizumab was selected as the first-line therapy to achieve a better tumor response and improved survival outcomes. At initiation, the patient had a preserved liver function (Child-Pugh class A, ALBI grade 2b), with no evidence of gastrointestinal bleeding or other high-risk hemorrhagic lesions that would contraindicate anti-vascular endothelial growth factor therapy. However, an acute infarction in the left middle cerebral artery territory was later confirmed by brain MRI, prompting reassessment of the treatment plan. Despite this event, the patient's Eastern Cooperative Oncology Group performance status remained stable (score 1), and his liver function did not significantly deteriorate. Following consultation with the neurology team, bevacizumab was discontinued owing to its known association with thromboembolic complications. Rather than switching to monotherapy, we initiated combination therapy with durvalumab and tremelimumab based on the efficacy of the HIMALAYA trial in HCC. Given the patient's stable performance status, preserved hepatic function, and absence of autoimmune comorbidities, this intensive immunotherapeutic approach was considered appropriate. Although the STRIDE regimen carries a higher risk of immune-related toxicities, the potential benefits outweigh the risks in this clinical context. The patient was closely monitored for irAEs and was able to continue therapy without serious complications, ultimately achieving sustained tumor regression.
Several cases of MPMs involving HCC and other malignancies have been previously reported. Aoki et al. (10) described a case of urHCC with synchronous esophageal cancer treated with first-line atezolizumab plus bevacizumab, in which a partial response for HCC and stable disease for esophageal cancer were achieved, with a survival duration of approximately nine months. Similarly, Suga et al. (11) reported a case of HCC and gastric cancer, and Inaba et al. (12) described a case of HCC and rectal cancer that were successfully treated with atezolizumab plus bevacizumab. Notably, in the case reported by Inaba et al., a complete response was achieved for both malignancies, with a 28-month survival duration. Additionally, Ghammem et al. (13) reported a case of synchronous HCC and pulmonary squamous cell carcinoma treated with durvalumab plus tremelimumab, which resulted in a partial response.
In recent years, a combination of local and systemic therapies for urHCC has gained attention. Local therapy has been shown to modulate the tumor immune microenvironment, thereby enhancing antitumor immune responses (14). Transarterial chemoembolization (TACE) induces tumor necrosis via arterial embolization, leading to the release of tumor antigens and the promotion of inflammatory responses that may potentiate antitumor immunity (15). Montasser et al. demonstrated that the expression levels of Programmed cell Death 1 (PD-1) and PD-L1 increased following TACE in HCC (16).
In the context of MPMs involving HCC, Inaba et al. reported favorable treatment outcomes following TACE prior to atezolizumab plus bevacizumab in a patient with HCC and rectal cancer lung metastases. Although TACE is not expected to directly enhance antigen presentation in colorectal cancer, its effects on specific immune microenvironmental factors have been suggested. Therefore, TACE may be a potential treatment strategy for MPM involving intermediate-stage HCC. However, in cases such as the present case, where the tumor burden exceeds the up-to-seven criteria with bilobar disease and albumin-bilirubin grade 2b hepatic function, systemic therapy should be prioritized.
A key finding in this case was the successful application of ICI therapy in PD-L1-negative ESCC. Although ICI efficacy is generally correlated with the expression of PD-L1, this case suggests that additional immune-related mechanisms may contribute to the treatment response beyond the PD-L1 status.
First, the dual blockade of the CTLA-4 and PD-L1/PD-1 pathway likely provided complementary immune activation, which helped overcome the limitations of PD-L1 negativity. While PD-1/PD-L1 inhibitors primarily reinvigorate exhausted T cells within the tumor microenvironment, CTLA-4 inhibitors act earlier in the immune cascade by enhancing T cell priming and activation in lymphoid tissues. This combination may generate a broader repertoire of tumor-reactive T cells capable of simultaneously recognizing HCC and ESCC.
Second, we hypothesized that coexisting HCC may enhance systemic immune activation, facilitating the response in ESCC. HCC is known to release tumor-associated antigens and damage-associated molecular patterns such as high mobility group protein B1 and ATP, which act as immunogenic signals to promote antigen presentation and T-cell priming (17). This process of immunogenic cell death may contribute to a broader anti-tumor immune response. Moreover, the interaction between the two tumors may trigger an abscopal-like effect, a phenomenon in which immune activation at one tumor site leads to the regression of distant, untreated tumors (18). Although traditionally associated with radiotherapy, growing evidence indicates that the abscopal effect can be enhanced by immune checkpoint blockade (19). The sequential tumor response observed in our case suggests that immune activation against HCC may have contributed to the subsequent eradication of PD-L1-negative ESCC, supporting the concept of systemic immune crosstalk.
Although speculative, this case supports the hypothesis that global immune activation, potentially involving tertiary lymphoid structures (TLSs) and cytotoxic T-cell recruitment, may underlie the responses in tumors traditionally considered less immunogenic or PD-L1-negative. Such responses may reflect enhanced immune activity within the tumor microenvironment, where tumor-infiltrating lymphocytes (TILs), particularly CD8+ T cells, have been associated with an improved prognosis and ICI responsiveness across various malignancies (20-22).
Additionally, TLSs, which are ectopic lymphoid formations within non-lymphoid tissues, are increasingly being recognized as key regulators of antitumor immunity (23). Hayashi et al. reported that in patients with ESCC, the density and maturity of TLSs at the tumor margin may serve as predictors of long-term survival and the response to anti-PD-1 therapy (24).
To further expand the indications for ICI therapy in MPMs, comprehensive clinical data and in-depth analyses of the tumor microenvironment and immune-related factors are essential. The identification of novel biomarkers independent of the expression of PD-L1, such as CD8+ TILs and TLSs, as well as the optimization of ICI combination strategies, remains a key challenge for future research. This case provides novel clinical insights into the immunological interplay between multiple primary tumors and highlights the need for mechanistic studies to explore the systemic immune dynamics underlying ICI-based treatment strategies for PD-L1-negative cancers.

The authors state that they have no Conflict of Interest (COI).