Systemic strategies for osteosarcoma: advances and future directions

Abstract

Osteosarcoma is a common and aggressive bone cancer in children and adolescents, typically affecting the long bones during growth spurts. Treatment involves a combination of surgery, chemotherapy, and radiotherapy. Although surgery has become more limb-preserving, the overall prognosis remains challenging, with a 5-year survival rate of 60–70% for localized disease and much lower for advanced stages. Chemotherapy is the mainstay treatment; however, it can cause severe side effects. Targeted therapies (e.g., IGF-1R inhibitors, TKIs) and immunotherapy (e.g., checkpoint inhibitors, CAR-T therapies) are promising areas of research that aim to attack cancer cells more precisely with lower toxicity. Other novel approaches, such as gene therapy and drug delivery systems, are being explored. With continued research and development of new therapies, we hope to significantly improve the outlook of patients with osteosarcoma in the future.

Introduction

Osteosarcoma is the most common primary bone malignancy affecting children and adolescents and is characterized by its aggressive nature and high likelihood of metastasis, particularly to the lungs (Fig. 1) [1]. This cancer typically arises in the metaphyseal region of long bones, such as the femur, tibia, and humerus, often during periods of rapid bone growth such as puberty [2]. Osteosarcoma originates from mesenchymal stem cells that undergo abnormal differentiation into osteoblasts [3]. Osteoblasts produce osteoid in an uncontrolled manner, leading to tumor formation. Several genetic mutations and alterations in molecular pathways are associated with the development of osteosarcoma. Commonly affected genes include tumour suppressor protein 53 and retinoblastoma 1, and other genes involved in cell cycle regulation, apoptosis, and DNA repair. These genetic changes lead to uncontrolled cell proliferation and survival. Abnormal osteoblasts proliferate rapidly to form a mass of malignant cells that produce osteoids. The tumor grows aggressively, invading nearby bone tissue and potentially spreading to the adjacent soft tissues. Metastasis occurs when cancer cells break away from the primary tumor, travel through the bloodstream or the lymphatic system, and form secondary tumors in distant organs such as lung and bones (Fig. 1) [4, 5].

Fig. 1.

Development and progression of osteosarcoma. Osteosarcoma is an aggressive malignant neoplasm. It typically arises in the metaphyseal region of long bones. Immature osteoblast differentiation forms the atypical hyperplasia, which converts into malignant tumor cells. This tumor cell further gets metastasized to other organs, specifically lung followed by other bone sites. This phenomenon of metastasis results in the tumor progression and development of drug resistance

The management of osteosarcoma requires a multimodal approach, combining surgery, chemotherapy, and occasionally, radiation therapy [6, 7]. Advances in surgical techniques, including limb-salvage procedures, have significantly enhanced patients’ quality of life. However, the prognosis remains challenging, with a 5-year survival rate of 60–70% for localized disease, which drops significantly in cases of metastatic or recurrent osteosarcoma, emphasizing the need for more effective treatments [8]. Chemotherapy, the mainstay of osteosarcoma treatment, involves drugs such as high-dose methotrexate (MTX), doxorubicin, cisplatin, and ifosfamide, which are administered to eliminate micrometastatic disease and shrink tumors for surgical resection [9]. Although this improves survival in localized cases, its effectiveness in metastatic disease is limited, and toxic side effects present significant challenges, driving the search for less toxic targeted therapies. Emerging targeted therapies, focusing on specific molecular pathways such as insulin like growth factor-1 receptor (IGF-1R) and the mammalian target of rapamycin (mTOR), have shown promise but have yielded mixed clinical results, underscoring the complexity of osteosarcoma [10]. Immunotherapy, inspired by successes in other cancers, is being explored through agents targeting the programmed death-1 (PD-1)/programmed death ligand-1 (PD-L1) pathway and adoptive T-cell therapies, although it is still in the early stages of osteosarcoma [11]. Additionally, novel treatments such as gene therapy, oncolytic virotherapy are under investigation. Despite advancements, the prognosis of osteosarcoma, particularly metastatic cases, remains poor, highlighting the critical need for ongoing research and innovative therapeutic strategies to improve patient outcomes (Fig. 2).

Fig. 2.

Current advancement in therapies for the treatment of osteosarcoma

The purpose of this review is to provide a comprehensive overview of the current state of osteosarcoma research and treatment. This review summarizes the latest advancements in chemotherapy protocols, targeted therapies, and immunotherapy and highlights their impact on patient outcomes. Additionally, this review seeks to identify the limitations of existing treatments and underscore the urgent need for novel therapeutic strategies. By consolidating recent findings and ongoing research efforts, this article intends to guide future investigations and foster the development of more effective and less toxic treatments for osteosarcoma, ultimately improving the survival rates and quality of life of osteosarcoma patients.

Historical perspective

The historical perspective of clinical trials on osteosarcoma highlights the significant advancements and challenges in the treatment and management of aggressive bone cancer. Early trials in the 1970s revealed the efficacy of systemic chemotherapy with agents such as high-dose MTX, cisplatin, and doxorubicin (MAP regimen), which improved survival rates when combined with surgical resection [12]. Subsequent studies, such as the EURAMOS-1 trial, have focused on understanding prognostic factors and disease progression, demonstrating the importance of tailored monitoring and treatment strategies based on patient characteristics and tumor location [13]. Recent investigations have explored novel therapies, including immunotherapy and targeted treatments, which show promise in improving outcomes for both human and canine osteosarcoma, reflecting advancements in veterinary and human oncology [14, 15]. Furthermore, observational studies have indicated that adding agents, such as mifamurtide, to conventional chemotherapy can enhance progression-free survival (PFS) in localized osteosarcoma, underscoring the ongoing evolution of therapeutic approaches [16].

Early chemotherapy trials

The introduction of chemotherapy in the 1970s marked a revolutionary era in the treatment of osteosarcoma. Prior to this period, the prognosis of osteosarcoma patients was poor, with long-term survival rates of approximately 20% following surgical resection alone. The advent of chemotherapy has brought new hope and significantly altered the clinical approach to aggressive cancer, transforming patient outcomes [12, 17].

Early clinical trials in the 1970s and 1980s were pivotal for demonstrating the efficacy of several chemotherapeutic agents. MAP regimens have emerged as cornerstone treatments for osteosarcoma [18]. MTX, a folate antagonist, inhibits DNA synthesis and cell replication, making it highly effective against rapidly dividing cancer cells. Doxorubicin, an anthracycline antibiotic, intercalates DNA strands, prevents replication, and ultimately leads to cell death. Cisplatin, a platinum-containing compound, forms DNA crosslinks and induces apoptosis of cancer cells. Each of these drugs has a unique mechanism of action in the burgeoning field of osteosarcoma chemotherapy [12, 13, 19].

Initially, the effectiveness of these drugs was explored in isolation; however, subsequent clinical trials have investigated their combined use. The synergistic effects of these drugs, when used together, provide a multifaceted attack on cancer cells, significantly enhancing overall treatment efficacy. This combined approach not only increased survival rates but also allowed for limb-salvage surgeries, greatly improving the quality of life of patients. The success of these chemotherapeutic regimens has laid the foundation for modern osteosarcoma treatment protocols and continues to influence ongoing research and therapeutic strategies.

The multi-institutional osteosarcoma study (MIOS)

One landmark trial that underscored the success of combination chemotherapy in osteosarcoma treatment was the MIOS [20]. Between the late 1970s and the early 1980s, MIOS played a pivotal role in establishing standard chemotherapy regimens still in use today. This extensive study involved numerous institutions and a large patient cohort and provided robust and comprehensive data on the efficacy of combination chemotherapy [20, 21].

The MIOS trial specifically compared the outcomes of patients receiving a combination chemotherapy drug MAP regimen along with surgical resection to those receiving surgery alone. The results were groundbreaking; patients in the combination therapy group had significantly improved survival rates. This study not only validated the use of chemotherapy in treating osteosarcoma but also emphasized the critical importance of a multidisciplinary treatment approach [20, 22].

The findings from MIOS and other early trials have laid the foundation for contemporary osteosarcoma treatment protocols. Currently, the standard treatment involves neoadjuvant chemotherapy (administered before surgery) to shrink the tumor, followed by surgical resection, and adjuvant chemotherapy (administered after surgery) to eradicate any remaining cancer cells [23, 24]. This multimodal approach has become the cornerstone of osteosarcoma management and substantially improved patient outcomes [20–24]. Such advancements have transformed osteosarcoma from a nearly uniformly fatal disease to one with a considerably improved prognosis, highlighting the ongoing need for research and innovation in cancer treatment.

Evolution of chemotherapy regimens

Chemotherapy regimens for osteosarcoma have evolved significantly over the decades, driven by relentless research and clinical trials aimed at enhancing patient outcomes. Early chemotherapy trials focused on single agents such as high-dose MAP regimens, which showed promise in treating osteosarcoma. However, the combination regimens truly transformed treatment outcomes. For example, a study that reviewed the history of osteosarcoma therapy highlighted the importance of combining effective systemic chemotherapy with surgical resection at all detectable disease sites [12]. A notable study from India demonstrated the success of sequential chemotherapy protocols, which significantly improved event-free survival and overall survival (OS) rates over two decades [25]. Additionally, the clonal evolution of metastatic osteosarcoma shaped by cisplatin treatment underscores the need for the continuous adaptation of chemotherapy regimens to address resistance [26].

Chemotherapy resistance in osteosarcoma is a complex issue driven by a combination of genetic, epigenetic, and microenvironmental factors that hinder treatment effectiveness. A significant portion of mutations in osteosarcoma tumors (up to 63%) are heterogeneous, leading to varied responses to chemotherapy [27]. Genetic changes like point mutations and gene amplifications can alter drug targets, reducing their efficacy [28]. Furthermore, chemotherapy-induced transcriptional and post-transcriptional changes can modify metabolic pathways and contribute to resistance [29]. Cancer stem cells, with their enhanced DNA repair mechanisms and inherent resilience, play a significant role in resistance. The tumor microenvironment (TME), supported by cancer-associated fibroblasts and immune cells, provides a niche that promotes resistance [27]. Epithelial-mesenchymal transition (EMT) further complicates treatment by increasing cell survival and invasiveness [28].

Effectively managing chemotherapy resistance in osteosarcoma requires a multifaceted approach that targets the diverse mechanisms involved. For instance, disrupting the CXCR4 signaling pathway can sensitize osteosarcoma cells to doxorubicin by altering glycolytic activity and promoting apoptosis [30]. Metformin has been shown to inhibit the YY1/MDR1 pathway, increasing the cytotoxic effects of Adriamycin in resistant osteosarcoma cells [31]. Dual inhibition of anti-apoptotic proteins like Bcl-2 and Bcl-xL, using selective inhibitors such as navitoclax, significantly reduces the viability of doxorubicin-resistant cells [32]. Combination therapies, such as using tyrosine kinase inhibitors with standard chemotherapy, have also shown promise in addressing the TME and improving drug delivery [33]. While these strategies offer potential, the complex nature of osteosarcoma resistance necessitates a comprehensive approach that includes personalized medicine and combination therapies to overcome it.

Introduction of ifosfamide and etoposide (IE)

Ifosfamide and etoposide (IE) are often used in combination chemotherapy regimens for osteosarcoma, especially in cases that are high-risk, relapsed, or refractory to standard treatment. Ifosfamide, an alkylating agent, and etoposide, a topoisomerase inhibitor, have distinct mechanisms of action that can target cancer cells in multiple ways, potentially leading to improved treatment outcomes [34–36].

Studies have investigated the use of ifosfamide as a continuous infusion for 14 days in patients with relapsed or refractory osteosarcoma. Some patients experienced tumor shrinkage and improvements in progression-free survival (PFS) and overall survival (OS) [37]. However, this treatment approach was also associated with significant toxicity, including neurotoxicity (e.g., confusion and lethargy), renal toxicity, and hematologic toxicity (low blood cell counts) [37]. While continuous infusion ifosfamide may be a promising alternative for treating relapsed osteosarcoma, further research is needed to confirm its efficacy, minimize risks, and improve patient outcomes, particularly in the refractory osteosarcoma population where a significant unmet clinical need remains.

While ifosfamide and etoposide (IE) have been used as a salvage regimen for soft tissue sarcomas, their efficacy as a single agent is limited, and they are associated with significant toxicity [34]. However, combining IE with other agents has shown some promise. One study investigated apatinib, an anti-angiogenesis tyrosine kinase inhibitor (TKI), in combination with IE and demonstrated some antitumor activity in relapsed or refractory osteosarcoma [35, 36]. It’s important to note that osteosarcoma is a bone cancer, not a soft tissue sarcoma. Another study explored the combination of lenvatinib, another TKI, with IE in a multicenter phase 1/2 trial for refractory or relapsed osteosarcoma, showing some encouraging results [38].

The combination of ifosfamide, carboplatin, and etoposide (ICE) has also been studied in sarcomas, demonstrating improved outcomes compared to IE alone, particularly in patients with fewer prior lines of systemic chemotherapy [39]. However, these combination regimens are associated with increased toxicity. Ifosfamide can cause encephalopathy, nephrotoxicity, and hemorrhagic cystitis, while etoposide can lead to myelosuppression and an increased risk of secondary malignancies. These adverse effects necessitate careful consideration of dosages and schedules to balance efficacy with patient safety. For example, a study examining the combination of apatinib with IE found clinically meaningful antitumor activity but emphasized the importance of managing associated toxicities (Table 1) [40].

Table 1.

Clinical trials and studies ongoing for treatment of osteosarcoma with targeted therapies

NCT number	Study title	Conditions	Interventions	Phase	Enrolment number
Chemotherapy and targeted therapy
05057130	Neoadjuvant combination of doxorubicin, cisplatin and methotrexate in patients aged 24–40 years with primary bone tumors	Osteosarcoma Sarcoma of bone And others	Methotrexate Cisplastin Doxorubicin	P2 P3	50
02986503	European study in bone sarcoma patients over 40 years	Spindle cell sarcoma of +bone Osteosarcoma	Ifosfamide Cisplastin Doxorubicin	NA	100
01258634	A study of pre-operative treatment of newly-diagnosed, surgically-resectable osteosarcoma with doxorubicin, ifosfamide, etoposide, and cisplatin with early metabolic assessment of response	Osteosarcoma Lung metastases	Dexrazoxane Cisplastin Doxorubicin G-CSF PEG-filgrastim Etoposide Ifosfamide Mesna Leucovorin	P1	2
00645632	Combination chemotherapy in treating patients undergoing surgery for newly diagnosed high-grade osteosarcoma	Sarcoma	Cisplastin Doxorubicin HCl Ifosfamide Methotrexate	NA	60
01176981	Outpatient administration of high dose methotrexate (HD MTX) in patients with osteosarcoma	Osteosarcoma	High-dose methotrexate	P3	6
00187109	Recombinant human thrombopoietin in children receiving ifosfamide, carboplatin, and etoposide chemotherapy	Osteosarcoma and others	Carboplatin ifosfamide Etoposide	P1 P2	40
04824352	Apatinib plus chemotherapy (ifosfamide and etoposide) for relapsed or refractory osteosarcoma	Effect of drug Toxicity Osteosarcoma	Apatinib, ifosfamide & etoposide	P2	44
04690231	Apatinib + Ifosfamide and Etoposide for Relapsed or Refractory Osteosarcoma	Effect of Drug Toxicity Secondary Resistance	Apatinib Mesylate Ifosfamide& Etoposide	NA	79
05277480	Apatinib with ifosfamide plus etoposide for relapsed or refractory osteosarcoma	Effect of drug Drug toxicity Osteosarcoma	Apatinib Etoposide	P2	81
04154189	A study to compare the efficacy and safety of ifosfamide and etoposide with or without lenvatinib in children, adolescents and young adults with relapsed and refractory osteosarcoma	Osteosarcoma	Lenvatinib Ifosfamide Etoposide	P2	81
05926492	A clinical study of surufatinib combined with chemotherapy as neoadjuvant Treatment in Osteosarcoma	Osteosarcoma	Surufatinib + chemotherapy Chemotherapy	P2	160
03163381	Apatinib second line treatment for advanced osteosarcoma and soft tissue sarcomas	Osteosarcoma, advanced	Apatinib	P2	40
02711007	Apatinib for advanced osteosarcoma after failure of standard multimodal therapy	Osteosarcoma Metastasis	Apatinib	P2 P3	37
03539172	Radiotherapy concurrent with apatinib in advanced soft tissue and bone sarcomas of head and neck-RASS study	Soft tissue Bone tumor Head and neck cancer Sarcoma	Apatinib mesylate	P2	61
03742193	Pulmonary resectable metastases of osteosarcoma with anti-angiogenics and chemotherapy	Osteosarcoma Pulmonary metastases	Apatinib GD regimen	P2	43
03359018	Apatinib plus anti-PD1 therapy for advanced osteosarcoma	Osteosarcoma	Apatinib SHR-1210	P2	43
04351308	Comparison of MAPI + camrelizumab versus API + apatinib versus MAPI in patients with a poor response to preoperative chemotherapy for newly diagnosed high-grade osteosarcoma	Osteosarcoma	MAPI chemotherapy Apatinib Mesylate Camrelizumab	P2	60
03491371	Apatinib for advanced sarcoma: results from multiple institutions’ off-label use	Advanced sarcoma	Methylsulfonic apatinib	NA	56
06125171	Tucidinostat plus apatinib for advanced osteosarcoma	Osteosarcoma	Tucidinostat, apatinib	P2	46
01216826	Phase II study of everolimus in children and adolescents with refractory or relapsed osteosarcoma	Refractory or relapsed osteosarcoma	Everolimus	P2	20
04044378	Famitinib plus camrelizumab & famitinib alone & famitinib plus ifosfamide in advanced osteosarcoma	Effect of drugs Progression Pediatric cancer	Famitinib Ifosfamide Camrelizumab	P1 P2	0
00977561	A study of cisplatin (or carboplatin) and etoposide with or without figitumumab (CP-751,871) in patients with extensive-stage small cell lung cancer	Solid tumor	Biological: figitumumab Cisplatin (carboplatin) Etoposide	P2	9
00147537	Combination study of CP-751,871 with paclitaxel and carboplatin in advanced lung cancer	Carcinoma, non-small-cell lung	Biological:CP-751,871 Paclitaxel Carboplatin Erlotinib	P1 P2	282
00668148	A five-tier, open-label study of IMC-A12 in advanced sarcoma	Ewing’s sarcoma rhabdomyosarcoma And others	Biological: IMC-A12 (cixutumumab)	P2	113
01016015	Temsirolimus and cixutumumab in treating patients with locally advanced, metastatic, or recurrent soft tissue sarcoma or bone sarcoma	Metastatic osteosarcoma And others	Biological: cixutumumab Temsirolimus	P2	178
00831844	Cixutumumab in treating patients with relapsed or refractory solid tumors	Osteosarcoma And others	Biological: cixutumumab	P2	116
01614795	Cixutumumab and temsirolimus in treating younger patients with recurrent or refractory sarcoma	Childhood sarcomas recurrent osteosarcoma And others	Biological: cixutumumab Temsirolimus	P2	46
02546544	Eurosarc trial of linsitinib in advanced ewing sarcoma	Relapsed and refractory ewing sarcoma	Linsitinib	P2	16
02057380	A rollover study for subjects that have participated in an astellas sponsored linsitinib trial	Advanced solid tumors	Linsitinib Erlotinib Paclitaxel Bortezomib Dexamethasone	P2	13
00889382	A study evaluating intermittent and continuous OSI-906 and weekly paclitaxel in patients with recurrent epithelial ovarian cancer (and other solid tumors)	Ovarian cancer solid tumors	OSI-906 Paclitaxel	P1 P1	152
01759303	Study of pazopanib in the treatment of osteosarcoma metastatic to the lung	Osteosarcoma Metastatic osteosarcoma	Pazopanib	P1	12
01130623	A phase I study of pazopanib as a single agent for children with refractory solid tumors	Osteosarcoma And others	Pazopanib (GW786034)	P1	0
02357810	Pazopanib hydrochloride and topotecan hydrochloride in treating patients with metastatic soft tissue and bone sarcomas	Osteosarcoma And others	Pazopanib HCl Oral Topotecan Hydrochloride	P2	178
01532687	Gemcitabine with or without pazopanib in treating patients with refractory soft tissue sarcoma	Osteosarcoma And others	Gemcitabine Gemcitabine HCl Pazopanib Pazopanib hydrochloride Placebo administration	P2	54
02180867	Radiation therapy with or without combination of chemotherapy or pazopanib before surgery in treating patients with newly diagnosed non-rhabdomyosarcoma soft tissue sarcomas that can be removed by surgery	Osteosarcoma And others	Doxorubicin Doxorubicin Hydrochloride Ifosfamide Pazopanib Pazopanib hydrochloride Radiation therapy	P2 P3	140
00889057	Sorafenib in relapsed high grade osteosarcoma	Osteosarcoma	Sorafenib	P2	35
00330421	Sorafenib in treating patients with soft tissue sarcomas	Osteosarcoma And others	Sorafenib tosylate	P2	15
01804374	P II open label, non-randomized study of sorafenib and everolimus in relapsed and non-resectable osteosarcoma	Metastatic osteosarcoma Relapsed osteosarcoma	Sorafenib Everolimus	P2	38
00880542	Sorafenib and ifosfamide in treating patients with high-grade soft tissue sarcoma or bone sarcoma that can be removed by surgery	Osteosarcoma	Sorafenib Ifosfamide	P2	7
01518413	Dose escalation study of sorafenib and irinotecan combination therapy in pediatric patients with solid tumors	And others	Sorafenib Irinotecan	P1	17
01946529	Therapeutic trial for patients with ewing sarcoma family of tumor and desmoplastic small round cell tumors	Osteosarcoma And others	vincristine doxorubicin cyclophosphamide Ifosfamide etoposide temozolomide temsirolimus bevacizumab sorafenib	P2	24
05395741	Regorafenib in Patients with refractory primary bone tumors	Osteosarcoma Ewing sarcoma of bone	Regorafenib	P1 P2	30
01900743	Phase II study of regorafenib in metastatic soft tissue sarcoma	Sarcoma	Regorafenib Placebo	P2	219
04698785	Efficacy of regorafenib combined with best supportive care as maintenance treatment in high grade bone sarcomas patients	Bone sarcoma Osteosarcoma	Regorafenib Placebo	P2	60
04803877	SARC038: Phase 2 study of regorafenib and nivolumab in osteosarcoma	Osteosarcoma	Regorafenib 40 MG Regorafenib 20MG Nivolumab	P2	48
05830084	P Ib/regorafenib with conventional chemotherapy, in newly diagnosed patients with multimetastatic ewing sarcoma	Bone cancer	Regorafenib tablet	P1	24
02389244	A P II study evaluating efficacy and safety of regorafenib in patients with metastatic bone sarcomas	Osteosarcoma And others	Regorafenib Placebo	P2	132
02048371	SARC024: a blanket protocol to study oral regorafenib in patients with selected sarcoma subtypes	Osteosarcoma And others	Regorafenib Placebo	P2	131
04698785	Efficacy of regorafenib combined with best supportive care as maintenance treatment in high grade bone sarcomas patients	Bone sarcoma Osteosarcoma	Regorafenib and BSC Treatment by placebo and best supportive care	P2	60
04055220	Efficacy and safety of regorafenib as maintenance therapy after first-line treatment in patients with bone sarcomas	Osteosarcoma And others	Regorafenib Placebo	NA	168

Fine-tuning of chemotherapy regimens

Subsequent clinical trials have focused on refining chemotherapy regimens for osteosarcoma to maximize efficacy while minimizing side effects. Dose adjustments, timing of administration, and supportive care measures were evaluated. For instance, hydration protocols and protective agents such as mesna have been introduced to reduce the risk of hemorrhagic cystitis caused by ifosfamide. Additionally, population pharmacokinetic studies have been conducted to optimize individual dosing for high-dose methotrexate, demonstrating the potential for tailored treatment approaches to reduce toxicity and improve outcomes [41]. Supportive care strategies, such as standardized documentation for high-dose MTX treatment, have significantly improved patient safety by reducing the incidence of adverse events such as acute kidney injury and delayed MTX clearance [42]. These measures highlight the importance of balancing the efficacy and safety of chemotherapy protocols.

Clinical trials have played a crucial role in testing new chemotherapy regimens involving IE, particularly for cancers such as osteosarcoma, small-cell lung cancer, and testicular cancer. These studies have often compared new regimens to standard regimens, meticulously recording response rates, PFS, and OS. For example, a study on the combination of ICE showed significant survival benefits in patients with relapsed or refractory bone and soft-tissue sarcomas (Table 1) [39]. Another study demonstrated the effectiveness of lenvatinib combined with IE in improving the outcomes of refractory or relapsed osteosarcoma [40]. Currently, there are many clinical studies recruiting patients with osteosarcoma and other bone sarcomas, as reported in Table 1.

However, these enhanced regimens have been associated with increased toxicity. Ifosfamide is associated with side effects such as encephalopathy, nephrotoxicity, and hemorrhagic cystitis, whereas etoposide can cause myelosuppression and increase the risk of secondary malignancies. These adverse effects necessitate careful reassessment of dosages and schedules to ensure that increased efficacy does not result in unsustainable costs to patient health. For instance, a study investigating the combination of apatinib with IE found clinically meaningful antitumor activity but also noted the importance of managing the associated toxicities (Table 1) [36].

Current standard of care for osteosarcoma

The current standard of care for osteosarcoma involves a multidisciplinary approach that combines surgery, chemotherapy, and radiotherapy to effectively manage the disease. Standard chemotherapy regimens typically include a high-dose MAP regimen with the possible addition of ifosfamide to high-risk or metastatic cases. Complete surgical resection of the primary tumor and any metastatic site is crucial for improving the survival outcomes. Advanced radiotherapy techniques, such as carbon-ion radiotherapy and stereotactic radiosurgery, are also being explored for patients who are medically inoperable or refuse to undergo surgery. This comprehensive approach has significantly improved survival rates, particularly in patients with localized disease, although challenges remain for those with metastatic or recurrent osteosarcoma [6, 43].

The importance of understanding the genetics and biology of osteosarcoma to develop patient-specific therapies has been emphasized. Despite the long-standing use of the MAP regimen as the standard of care, ongoing research aims to develop novel therapeutics targeting the complex heterogeneity of osteosarcoma tumors [44]. For instance, systemic chemotherapy has significantly changed the natural history of osteosarcoma, with adjuvant and neoadjuvant chemotherapy combined with surgery being the current standard [45]. However, the high rates of treatment failure and morbidity present significant challenges, highlighting the need for continued research and clinical trials [46].

Neoadjuvant and adjuvant chemotherapy

The treatment paradigm for osteosarcoma involves a multimodal approach that includes neoadjuvant chemotherapy, surgical resection, and adjuvant chemotherapy. Neoadjuvant chemotherapy is administered preoperatively to shrink the primary tumor, making it more manageable for surgical resection and targeting micrometastatic diseases that may not be detectable through imaging. This approach also provides an early assessment of tumor responsiveness to chemotherapy, guiding postoperative treatment decisions. Adjuvant chemotherapy follows surgical resection to eliminate any remaining microscopic disease and to reduce the risk of recurrence.

Key clinical trials, such as those conducted by the European Osteosarcoma Intergroup, have provided robust evidence to support this multimodal approach. These studies have shown that incorporating neoadjuvant chemotherapy enhances the feasibility of limb-salvage surgeries and improves OS rates compared to historical controls, where surgery alone or surgery with postoperative chemotherapy was employed. The high-dose MAP regimen, which has been proven effective in improving survival rates and reducing the likelihood of metastasis. Patients receiving neoadjuvant chemotherapy followed by surgery have higher survival rates and better functional outcomes than those who undergo surgery alone [47]. Another study highlighted the significant benefits of combining neoadjuvant chemotherapy with limb-sparing surgery, with improved postoperative limb function and long-term quality of life [48].

The role of neoadjuvant chemotherapy in the treatment of osteosarcoma is further supported by studies that focus on its impact on long-term outcomes and quality of life. Research has shown that patients treated with neoadjuvant chemotherapy followed by surgery and adjuvant chemotherapy experience better OS and lower recurrence rates than those treated with surgery alone [49].

Salvage surgery vs. amputation

Current research underscores the evolving preference for limb-salvage surgery (LSS) over amputation in managing osteosarcoma, highlighting the significant advancements in surgical techniques and chemotherapy protocols. Studies have shown that LSS offers oncologic outcomes comparable to those of amputation, with improved functional and psychosocial benefits for patients. LSS has been reported to significantly improve disease-free and OS rates compared to amputation, especially with neoadjuvant chemotherapy enhancing 5-year survival rates [50]. It has also been reported that some patients initially treated with LSS opt for revision amputation because of chronic pain and functional limitations, suggesting the need for thorough preoperative discussions about potential outcomes [51]. Another study focusing on population-based evidence showed that LSS leads to better OS and disease-specific survival rates than amputation for early-stage primary bone tumors of the extremities [52].

Targeted therapies

Recent research on targeted therapies for osteosarcoma, has focused on exploiting the unique molecular and genetic characteristics of the disease. The role of major signaling pathways, such as phosphatidylinositol 3-kinase/protein kinase B (PI3K/Akt)/mTOR, janus kinase/signal transducers and activators of transcription (JAK/STAT), and Wnt/β-catenin, in osteosarcoma pathogenesis has been highlighted in several studies, proposing that novel targeted therapies could be developed by focusing on these pathways [53, 54]. These studies will aid in the development of various targeted therapies and molecular markers [55, 56]. The modulatory roles of signaling pathways and their potential as treatment targets for osteosarcoma have been discussed, along with therapies developed over the past decade [57]. Molecular markers and signaling pathways such as PD-1/PD-L1 and nuclear factor kappa light-chain-enhancer of activated B cells (NF-κB) have been identified as novel targets, offering new avenues for treatment [56]. Recently, the use of alpha particle-emitting isotopes like Radium-223 for targeted radiation therapy, particularly for bone-forming tumors and metastases expressing IGF-1R or human epidermal growth factor receptor 2 (HER2), has opened a new perspective for combination therapy [58].

Targeted therapies: IGF-1R inhibitors

The role of IGF-1R in osteosarcoma and its potential as a therapeutic target are focal points of recent research, underscoring its critical role in tumor growth, survival, and metastasis. Several studies have investigated the mechanisms by which IGF-1R influences osteosarcoma progression and explored targeted therapeutic approaches. A study identified recurrent mutations in the IGF signaling pathway in osteosarcoma in a small subset of cases (7%), and IGF-1R amplification was observed in 14% of tumors [59]. This mutational mechanism is age-independent, contributing to the development of osteosarcoma in children and adults alike, emphasizing its crucial role in the development and progression of the disease. These mutations disrupt normal IGF signaling, which is vital for regulating cell growth and survival, and are associated with tumor aggressiveness and treatment resistance. Additionally, the study uncovered distinct genomic rearrangements in osteosarcoma that further complicate the cancer’s biology. These alterations are linked to tumor progression, suggesting that IGF signaling could be a promising therapeutic target. The findings provide valuable insights into the molecular mechanisms of osteosarcoma and propose that targeting IGF signaling may offer new opportunities for improving treatment outcomes in affected patients [59].

Some studies have demonstrated that miR-939-5p inhibits IGF-1R, leading to decreased cell proliferation, migration, and invasion in osteosarcoma by deactivating the PI3K/AKT pathway [60, 61]. A quinazoline derivative, HMJ-30, was reported to disrupt IGF-1R signaling, thereby inhibiting osteosarcoma cell invasiveness and reversing the epithelial-mesenchymal transition (EMT) [62]. IGF-1R overexpression driven by copy number gain could serve as a potential therapeutic target in osteosarcoma [63]. It was also evaluated that NT157, an IRS-1/2 inhibitor, showed significant inhibition of osteosarcoma cell growth and migration, providing a promising therapeutic approach [64]. Metformin enhanced osteosarcoma sensitivity to chemotherapy via the IGF-1R/miR-610/FEN1 pathway, suggesting its potential as an adjuvant therapy [65].

The development of IGF-1R inhibitors has marked a significant advancement in the treatment of osteosarcoma, with both monoclonal antibodies and small-molecule tyrosine kinase inhibitors (TKIs) showing potential in preclinical and clinical settings [63, 65]. The use of soluble IGF-1R mutants to block the IGF-1R signaling axis significantly inhibited tumor growth and metastasis in preclinical models [66]. A correlation between nuclear IGF-1R and poor survival outcomes revealed the potential of targeting nuclear IGF-1R for better clinical outcomes [67]. Still, there are some limitations, such as in a phase 2 trial of R1507, a monoclonal antibody targeting IGF-1R, which showed limited activity in recurrent or refractory sarcomas but highlighted the importance of identifying predictive factors for clinical benefit (Table 1) [68]. As discussed, these inhibitors have shown promising results but have yielded mixed clinical results, underscoring the complexity of osteosarcoma [10].

Small-molecule inhibitors targeting the tyrosine kinase domain of IGF-1R have emerged as promising agents for the treatment of osteosarcoma. These inhibitors penetrate cellular membranes and inhibit IGF-1R kinase activity, effectively blocking downstream signaling pathways critical for tumor growth and metastasis. A computational study identified two potential natural inhibitors, ZINC000014946303 and ZINC000006003042, which showed high binding affinity to IGF-1R and favorable safety profiles, highlighting their potential as IGF-1R inhibitors [69]. Additionally, ABSK021, a potent and selective colony-stimulating factor-1 receptor (CSF-1R) inhibitor, demonstrated strong efficacy in preclinical osteosarcoma models by inhibiting CSF-1R activity and reducing macrophage infiltration [70]. TKIs for osteosarcoma and Ewing’s sarcoma have highlighted their potential when combined with conventional chemotherapy or other targeted agents [71].

IGF-1R-targeted therapies for osteosarcoma have highlighted the potential and challenges of using monoclonal antibodies, such as figitumumab and cixutumumab. A study involving cixutumumab in women with breast cancer that progressed on endocrine therapy found that while the antibody was well-tolerated, it did not significantly improve clinical outcomes (Table 1) [72]. Interestingly, lower expression levels of insulin receptor isoforms correlate with better progression-free survival (PFS) and overall survival (OS), suggesting a potential target for therapeutic efficacy [73, 74]. Another study on ganitumab, another IGF-1R antibody, in combination with dasatinib, showed a 22% disease control rate in rhabdomyosarcoma, indicating some effectiveness in co-targeting pathways to overcome resistance [75]. Additionally, research on targeted alpha-particle therapies, such as radium-223 and actinium-225, linked to anti-IGF1R antibodies, presents a novel approach for treating osteosarcoma by leveraging the near-universal expression of IGF1R in these tumors [58].

Figitumumab

The monoclonal antibody figitumumab, targeting IGF-1R, has been evaluated in phase II clinical trials for osteosarcoma, highlighting both its potential and limitations (Table 1). A key study of figitumumab revealed that while the antibody was well tolerated, its overall clinical benefit was limited, with modest response rates observed in patients with refractory or relapsed osteosarcoma. This outcome underscores the complexity of osteosarcoma and suggests that IGF-1R inhibition alone may not suffice for significant therapeutic effects, indicating the need for combination therapies to enhance efficacy [76].

Furthermore, studies on IGF-1R-targeting antibodies similar to ganitumab in other sarcomas, such as Ewing sarcoma, showed that while IGF-1R antibodies can achieve partial responses, the rapid development of resistance and modest OS benefits highlight the necessity of co-targeting strategies or combinations with other treatments [77]. In a meta-analysis of IGF-1R-targeted therapies, the combination of IGF-1R and mTOR inhibitors demonstrated improved PFS, suggesting that combination approaches might provide more sustained clinical benefits [78].

Cixutumumab

Cixutumumab, combined with the mTOR inhibitor temsirolimus, was evaluated in a phase I study of children with recurrent solid tumors, including osteosarcoma (Table 1). The rationale behind this combination therapy is based on preclinical evidence suggesting that dual inhibition of the IGF-1R and mTOR pathways synergistically enhances the antitumor activity. This study determined the maximum tolerated doses for the combination and noted that while the treatment was feasible and showed some biological activity, its clinical efficacy remained limited. Predominant dose-limiting toxicities included mucositis, hypercholesterolemia, fatigue, thrombocytopenia, and increased alanine aminotransferase levels. Despite some clinical activity, especially in patients with fewer prior treatments, the overall response rates are modest, highlighting the need to optimize treatment regimens and explore additional therapeutic targets [79, 80].

Linsitinib (OSI-906)

Linsitinib (OSI-906) is a dual inhibitor targeting both IGF-1R and the insulin receptor, showing significant antitumor activity in preclinical osteosarcoma models. In a phase I clinical trial involving patients with advanced solid tumors, including osteosarcoma, linsitinib was well-tolerated and exhibited preliminary antitumor effects (Table 1). The study determined the maximum tolerated dose to be 400 mg once daily and 150 mg twice daily, with dose-limiting toxicities including hyperglycemia and elevated liver enzymes. Although the observed antitumor activity was promising, these initial findings necessitate further clinical trials specifically focusing on osteosarcoma to confirm its efficacy and potential as a standard treatment option. Moreover, understanding the molecular mechanisms and identifying therapeutic target predictive of the response to linsitinib is crucial for optimizing its therapeutic application in patients with osteosarcoma [81–83].

TKIs

TKIs have emerged as promising targeted therapies for osteosarcoma. These inhibitors interfere with specific signaling pathways crucial for cancer cell survival, proliferation, and metastasis by inhibiting tyrosine kinase activity [84, 85]. Tyrosine kinases are enzymes that catalyze the transfer of phosphate groups to tyrosine residues on proteins, a process vital for the activation of many signaling pathways involved in cell division, survival, and migration. Dysregulation of tyrosine kinases due to mutations or overexpression is common in various cancers, including osteosarcoma [86]. By blocking these enzymes, TKIs can disrupt aberrant signaling pathways, potentially leading to reduced tumor growth and metastasis [87, 88].

Several tyrosine kinases have been implicated in the pathogenesis of osteosarcoma, including vascular endothelial growth factor receptors (VEGFR), Platelet-derived growth factor receptor (PDGFR), receptor tyrosine kinase (KIT), and Mesenchymal Epithelial Transition (MET). Recent studies have shown that multi-target TKIs such as apatinib, cabozantinib, lenvatinib, regorafenib, and sorafenib have significant therapeutic effects in osteosarcoma (Table 1). However, a key challenge is to determine the most critical targets for effective treatment. To achieve a breakthrough in osteosarcoma therapy, it is necessary to simultaneously inhibit several relevant receptor tyrosine kinases [89]. Additionally, a review highlighted the need to overcome drug resistance to TKIs, which is a major barrier to improving OS in patients with osteosarcoma. Strategies to overcome TKI resistance include combining TKIs with other therapeutic agents and using biomimetic nanoparticles to enhance drug targeting and reduce toxicity [85].

Clinical trials and studies

Clinical trials have explored the efficacy of TKIs in treating osteosarcoma, highlighting both their potential and challenges. A meta-analysis comparing the clinical value of PD-1 inhibitors and TKIs in advanced osteosarcoma found that TKIs provide better OS and PFS than PD-1 inhibitors. However, the combination of TKIs and PD-1 inhibitors may offer enhanced benefits, despite significant adverse effects [85]. A phase II study of surufatinib, a multi-targeted TKI, in patients with osteosarcoma and soft tissue sarcoma who failed standard chemotherapy showed promising results, with a progression-free rate at 12 weeks of 60% and a disease control rate of 80%, although treatment-related toxicities were notable [90].

Recent advances have highlighted the efficacy of anti-angiogenic multi-receptor TKIs such as pazopanib, sorafenib, and regorafenib in osteosarcoma and Ewing sarcoma [36, 38, 91, 92]. Despite positive clinical data, these agents have yet to gain regulatory approval for these indications, and resistance remains a significant hurdle [93]. An overview of the progress and challenges in applying TKIs to osteosarcoma emphasizes the issue of acquired drug resistance, suggesting the need for combination therapies and innovative strategies to overcome this barrier [86].

Pazopanib

Pazopanib, a multi-targeted TKI that inhibits VEGFR, PDGFR, and KIT, has been evaluated in various clinical trials for its efficacy in osteosarcoma treatment (Table 1). A phase II study demonstrated that pazopanib provided modest improvement in PFS in patients with advanced osteosarcoma who had exhausted other treatment options [94]. This trial highlighted manageable toxicity profiles, suggesting the potential of pazopanib as a treatment for these patients [91]. Other studies have investigated the role of pazopanib in sarcomas. Additionally, a retrospective cohort study on pediatric sarcoma patients treated with pazopanib indicated its effectiveness and well-tolerated profile, with a 64% OS rate in the entire cohort [95, 96].

Sorafenib

Sorafenib, which targets RAF kinases VEGFR, PDGFR, and KIT, has shown promise when combined with everolimus, an mTOR inhibitor, in treating osteosarcoma (Table 1). A phase II trial combining these two drugs demonstrated improved PFS in patients with refractory or relapsed osteosarcoma compared with historical controls, suggesting potential synergistic effects [97, 98]. Preclinical models support these findings, showing that the combination abrogates mTORC1 and mTORC2 upregulation, enhances antiproliferative and proapoptotic effects, impairs tumor growth, and reduces migratory and metastatic potential [99, 100].

A patient-derived orthotopic xenograft mouse model study revealed that a combination of sorafenib and everolimus regressed doxorubicin-resistant osteosarcoma, indicating the potential of this combination therapy to overcome drug resistance in osteosarcoma [101]. However, further clinical trials are necessary to validate these findings and optimize dosing regimens to minimize the adverse effects.

Regorafenib

Regorafenib, a multikinase inhibitor that targets VEGFR, PDGFR, FGFR, and KIT, has shown significant promise in the treatment of metastatic osteosarcoma [101, 102]. In a phase II trial, regorafenib significantly improved PFS in patients with metastatic osteosarcoma who had progressed after standard chemotherapy (Table 1) [103, 104]. These trial results have spurred increased interest in regorafenib as a viable treatment option for advanced osteosarcoma, emphasizing the need for additional research to fully understand its benefits and risks. The REGOSARC study also found that regorafenib significantly prolonged PFS in pretreated patients with advanced non-adipocytic sarcoma, indicating its broad potential for sarcoma treatment [103]. Additionally, another phase II trial, REGOMAIN, is ongoing to study the efficacy of regorafenib as a maintenance treatment in patients with high-grade bone sarcomas, with the aim of reducing the risk of progression after the initial treatment [105, 106]. Further investigation in the SARC024 trial also confirmed the activity of regorafenib in advanced Ewing’s sarcoma and related tumors, underscoring its potential across different sarcoma subtypes [107].

Challenges and future directions

TKIs have shown promise in the treatment of osteosarcoma; however, several challenges remain. A significant concern is the development of resistance to TKIs because cancer cells can activate alternative pathways or acquire mutations that render these drugs ineffective [108]. This resistance often leads to limited long-term benefits, highlighting the need for novel strategies to enhance the treatment’s durability. Additionally, toxicity profiles require careful management to ensure patient safety and adherence to treatment protocols [109]. Combining TKIs with other therapeutic modalities such as immunotherapy or traditional chemotherapy could also enhance their efficacy and help overcome resistance mechanisms [110, 111]. For instance, combining TKIs with immune checkpoint inhibitors or mTOR inhibitors has shown potential in preclinical models and early phase clinical trials, suggesting synergistic effects that warrant further exploration. Both IGF-1R and TKIs represent promising avenues for osteosarcoma therapy, although several challenges and opportunities for improvement remain [112, 113].

Immunotherapy

Immunotherapy is indeed a promising strategy for treating osteosarcoma, harnessing the power of the immune system to target and destroy cancer cells. Various approaches are under investigation, including immune checkpoint inhibitors, adoptive T cell therapy, and cancer vaccines. The tumor microenvironment (TME) of osteosarcoma presents unique challenges and opportunities for immunotherapy. Promising strategies include immune checkpoint inhibitors and adoptive cell therapies, which have shown potential to improve patient outcomes [11, 114]. Previous studies have explored various immunotherapeutic strategies like immunomodulation, vaccine therapy, and adoptive T-cell therapy, noting their potential to enhance outcomes for osteosarcoma patients [115, 116]. The current landscape and future directions of immunotherapy for osteosarcoma underscore the potential of these novel therapies [117]. Several strategies, including immunological checkpoint blockade, highlight the promise of immunotherapy for managing osteosarcoma [118].

However, osteosarcoma can develop resistance to immunotherapy due to the complex interplay between the TME and the intrinsic characteristics of tumor cells. The TME, comprised of immune and non-immune cells, often fosters an immunosuppressive environment in osteosarcoma, hindering the effectiveness of immune checkpoint inhibitors and other immunotherapies [114, 119, 120]. Specific TME subtypes, such as immune-dominant or fibro-dominant, influence treatment responses, with some subtypes exhibiting greater resistance due to their cellular composition and signaling pathways [119]. Intratumoral heterogeneity further complicates treatment by leading to variable immune cell infiltration and allowing immunosuppressive tumor cell populations to proliferate [121]. Unlike hematologic malignancies, solid tumors like osteosarcoma often lack uniform expression of target antigens, making targeted immunotherapy less effective. Additionally, tumor cells employ strategies to evade immune detection, such as altering antigen presentation and creating an immunosuppressive environment. The rapid division and complex metabolism of tumor cells further impede immune responses. These factors contribute to both primary and acquired resistance to immunotherapy [122]. To address these challenges, research is focusing on combination therapies, targeting immunosuppressive molecules, and enhancing cancer cell immunosensitization to improve treatment efficacy and overcome resistance.

Immune checkpoint inhibitors in osteosarcoma

The advent of immune checkpoint inhibitors has revolutionized cancer treatment, offering new hope for patients with various malignancies, including osteosarcoma. These therapies target regulatory pathways in T cells to enhance the immune system's ability to fight cancer. In osteosarcoma, the potential of immune checkpoint inhibitors is being actively explored, although the journey is fraught with challenges and mixed results. Shi et al. conducted a meta-analysis revealing that TKIs may offer more benefits than PD-1 inhibitors for advanced osteosarcoma, although combining these therapies shows promise despite the significant side effects [85]. Han et al. developed a novel approach by combining IDO inhibitors with platinum (IV) prodrugs to enhance both chemotherapy and immunotherapy in osteosarcoma [123]. Peng et al. analyzed combination therapies involving immune checkpoint inhibitors, emphasizing the importance of understanding the TME and immune cell infiltration for better therapeutic responses [124]. Muñoz-García et al. highlighted recent advancements in osteosarcoma treatment, including immune checkpoint inhibitors, although clinical trials have shown only slight improvements over traditional therapies [125]. Finally, Zhang et al. reviewed the current status and challenges of immune checkpoint inhibitors in osteosarcoma, noting the potential of these therapies despite issues such as drug resistance and severe side effects [126]. Such as pembrolizumab and nivolumab, both anti-PD-1 antibodies, function by blocking the PD-1 pathway, which tumors exploit to evade the immune response. These therapies aim to restore the capacity of the immune system to recognize and attack cancer cells. The potential of these drugs in treating osteosarcoma is being actively explored.

Pembrolizumab trials

Early phase trials involving pembrolizumab in osteosarcoma have provided critical insights but also underscored the complexities involved in treating this disease with immunotherapy (Table 2). For instance, Boye et al. conducted a phase II trial of pembrolizumab in patients with relapsed or refractory osteosarcoma and demonstrated limited efficacy. Only a small subset of patients experienced durable responses, indicating that, while some patients may benefit significantly from pembrolizumab, the majority do not respond favorably [127]. The SARC028 trial highlighted the association between sarcoma-associated immune infiltrate and the response to pembrolizumab, indicating that higher densities of activated T cells and tumor-associated macrophages expressing PD-L1 correlate with better outcomes [128]. The comparative study of pembrolizumab and nivolumab in real-life settings showed different adverse event profiles, emphasizing the need for careful management of these therapies [129]. Livingston et al. combined pembrolizumab with doxorubicin in advanced soft-tissue sarcoma, showing manageable toxicity and promising activity, but similar trials in osteosarcoma are needed to further explore combination strategies (Table 2) [130]. Pollack et al. assessed pembrolizumab with doxorubicin in a phase I/II trial and noted a partial response rate of 13% in sarcoma patients, suggesting some potential for combination therapies to enhance outcomes [131]. Groisberg et al. highlighted that checkpoint inhibitors, including pembrolizumab, show varied responses across different sarcoma subtypes, stressing the need for more tailored approaches [132]. Lastly, Kelly et al. explored the combination of epacadostat (an IDO1 inhibitor) with pembrolizumab but found limited antitumor activity, indicating that achieving adequate inhibition of immunosuppressive pathways remains a challenge [133].

Table 2.

Summarizes the selected ongoing clinical trials of immunotherapy for osteosarcoma

NCT number	Study title	Conditions	Interventions	Phase	Enrolment number
Immunotherapy
02301039	SARC028: A P II study of the anti-PD1 antibody pembrolizumab (MK-3475) in patients with advanced sarcomas	Soft tissue sarcoma Bone sarcoma	Pembrolizumab	P2	144
03013127	A study of pembrolizumab in patients with relapsed or metastatic osteosarcoma not eligible for curative surgery	Osteosarcoma	Pembrolizumab	P2	12
05182164	Combination of pembrolizumab and cabozantinib in patients with advanced sarcomas	Osteosarcoma And others	Association of pembrolizumab + cabozantinib	P2	119
03056001	Safety, tolerability, and efficacy of doxorubicin and pembrolizumab for sarcoma	Soft tissue sarcoma adult Soft tissue sarcoma, child	Pembrolizumab Doxorubicin	P2	30
02888665	Pembrolizumab and doxorubicin hydrochloride in treating patients with sarcoma that is metastatic or cannot be removed by surgery	Sarcoma	Pembrolizumab Doxorubicin HCl	P1 P2	37
03414229	A study of epacadostat, an IDO1 inhibitor, in combination with pembrolizumab in patients with metastatic and/or locally advanced sarcoma	Sarcoma	Pembrolizumab Epacadostat	P2	30
03628209	Nivolumab or nivolumab and azacitidine in patients with recurrent, resectable osteosarcoma	Osteosarcoma Osteosarcoma in children Osteosarcoma recurrent| Sarcoma	Nivolumab Azacitidine	P1 P2	21
02982486	A P II of nivolumab plus ipilimumab in non-resectable sarcoma and endometrial carcinoma	Osteosarcoma And others	Ipilimumab Nivolumab	P2	60
03277924	Trial of sunitinib and/or nivolumab plus chemotherapy in advanced soft tissue and bone sarcomas	Soft tissue sarcoma Bone sarcoma	Sunitinib Nivolumab Epirubicin Ifosfamide Doxorubicin Dacarbazine Cisplatin Methotrexate	P1 P2	270
03190174	Nivolumab (Opdivo®) Plus ABI-009 (Nab-rapamycin) for advanced sarcoma and certain cancers	Osteosarcoma And others	NazRapamycin Biological: Nivolumab	P1 P2	34
03628209	Nivolumab or nivolumab and azacitidine in patients with recurrent, resectable osteosarcoma	Osteosarcoma	Nivolumab Vorinostat	P1 P2	21
04897321	B7-H3-specific chimeric antigen receptor (CAR-T) Autologous T-cell therapy for pediatric patients with solid tumors (3CAR)	Osteosarcoma And others	Fludarabine Cyclophosphamide MESNA B7-H3 CAR T cells	P1	32
04864821	Clinical study of CD276 targeted autologous CAR-T cell infusion in patients with CD276 positive advanced solid tumor	Osteosarcoma Neuroblastoma Gastric cancer Lung cancer	Targeting CD276 CAR T cells	EARLY P1	24
02107963	A Phase I trial of T cells expressing an anti-GD2 CAR-T in children and young adults with GD2+ solid tumors	Sarcoma Osteosarcoma and others	Biological: anti-GD2-CAR engineered T cells AP1903 Cyclophosphamide	P1	15
03373097	Anti-GD2 CAR-T cells in pediatric patients affected by high risk and/or relapsed/refractory neuroblastoma or other GD2-positive solid tumors	Osteosarcoma and others	Biological:GD2-CART01	P1	42
06412458	IM83 clinical study of CAR-T cell therapy in patients with relapsed or refractory osteosarcoma	Refractory osteosarcoma|recurrent osteosarcoma	Biological:IM83 CAR-T cells	P1	9
01953900	iC9-GD2-CAR-VZV-CTLs/refractory or metastatic GD2-positive sarcoma and neuroblastoma	Osteosarcoma Neuroblastoma	Genetic: GD2 T cells Biological:VZV vaccine Fludarabine Cyclophosphamide	P1	26
05312411	A phase I feasibility and safety study of fluorescein-specific (FITC-E2) CAR T cells in combination with parenterally administered folate-fluorescein (UB-TT170) for osteogenic sarcoma	Osteosarcoma	Biological: SCRI-E2CAR_EGFRtv1 UB_TT170	P1	21
04433221	Combination immunotherapy targeting sarcomas	Sarcoma Osteoid sarcoma Ewing sarcoma	Biological: multiple sarcoma-specific CAR-T cells and sarcoma vaccines	P1 P2	20

Nivolumab trials

Nivolumab, another PD-1 inhibitor, has been tested in early-phase trials for osteosarcoma with results similar to pembrolizumab (Table 2). These studies highlight the complexities and challenges of treating aggressive cancers with immunotherapy. Zheng et al. demonstrated that nivolumab effectively inhibited metastasis in a humanized mouse model of osteosarcoma by increasing CD4+ and CD8+ lymphocytes, although it did not affect primary tumor growth [134]. Reichardt et al. explored the efficacy and safety of combining nivolumab with trabectedin in pretreated patients with advanced soft tissue sarcomas, no8ting a significant difference in PFS between the different cohorts, although these findings may need further investigation in osteosarcoma [135]. Zhou et al. studied the anti-PD-L1 antibody ZKAB001 as a maintenance therapy in patients with localized high-grade osteosarcoma, showing manageable safety and preliminary evidence of clinical activity [136]. These trials underscore the modest response rates and the need for a deeper understanding of tumor biology and the immune landscape to improve treatment outcomes with PD-1 inhibitors in osteosarcoma.

Adoptive T-cell therapy

Adoptive T-cell therapy is a form of cancer immunotherapy that utilizes modified T-cells to target and destroy cancer cells. This approach encompasses two main types: chimeric antigen receptor (CAR) T cells and T-cell receptor (TCR)-engineered T-cells. CAR T-cell therapy involves genetically engineering a patient’s T cells to express a synthetic receptor (CAR) that can recognize specific antigens on the surface of cancer cells [137–139]. This CAR combines the antigen-binding domain of an antibody with the signaling domains of a TCR, allowing the modified T-cells to effectively target and eliminate tumor cells [140]. While CAR-T cell therapies have demonstrated remarkable success in treating hematological malignancies, their application in solid tumors like osteosarcoma presents challenges due to factors like poor T-cell infiltration and an immunosuppressive tumor microenvironment (TME) [137, 141].

CAR T-cells

Chimeric antigen receptor (CAR) T-cell therapy utilizes genetically modified T-cells expressing receptors that target specific antigens on cancer cells. These CAR-T cells combine the antigen-binding domain of an antibody with the signaling domains of a T-cell receptor (TCR), enabling the modified T-cells to recognize and attack tumor cells with enhanced specificity and potency. This approach holds promise for osteosarcoma treatment, although challenges such as tumor immune evasion and limited CAR T-cell persistence in the tumor microenvironment need to be addressed.

Several studies have explored CAR T-cell therapies for osteosarcoma. For example, Adeshakin et al. demonstrated that Regnase-1 knockout can improve the antitumor activity of B7-H3-CAR T cells [142]. Robbins et al. proposed CAR-engineered NK cells as an alternative, showing enhanced therapeutic efficacy and safety [138]. Zhang et al. highlighted the efficacy of B7-H3 targeted CAR T cells in preclinical models [143]. Hidalgo et al. discussed a switchable CAR T-cell strategy targeting B7-H3 for a controllable antitumor response [139]. Other studies have investigated complementary approaches, such as membrane-anchored and tumor-targeted IL-12 therapy [144], and strategies to overcome challenges in solid tumors [145]. Beyond osteosarcoma, research has focused on optimizing CAR T-cell production and quality control [140], developing versatile CAR T-cell systems with broader antigen recognition [146], and providing comprehensive overviews of the engineering process, mechanisms, challenges, and future prospects of CAR T-cell therapy [147].

TCR engineered T-cells

T-cell receptor (TCR) engineering enhances the natural specificity of T cells by modifying their TCRs to recognize tumor-associated antigens presented by major histocompatibility complex (MHC) molecules on cancer cells [148]. This approach expands the range of targetable cancer epitopes by leveraging the inherent ability of T cells to detect and respond to intracellular antigens. Greenbaum et al. discuss the challenges and advancements in engineering T cells to express specific TCRs for targeting cancer, focusing on strategies to enhance TCR T-cell efficacy and overcome obstacles in solid tumor treatments [148]. Tsuji et al. describe a method for rapid construction of TCR-expression libraries from tumor specimens, facilitating the identification of TCR genes for various tumor antigens [149]. Kok et al. demonstrate the potential of TCR-edited T cells targeting shared neoantigens derived from common cancer mutations, utilizing CRISPR technology to enhance T-cell functionality [150]. Sun et al. review advancements in TCR-T cell therapies, focusing on deriving tumor antigen-specific TCRs and clinical achievements in treating solid tumors, while addressing off-target toxicities and antigen loss [151]. Delfanti et al. explore the use of invariant NK-T (iNKT) cells engineered with tumor-specific TCRs, highlighting strong antitumor activity through dual targeting [152]. Further research includes studies on novel TCR knockout reporter assays [153], coexpression of CD8α in TCR-engineered CD4+ T cells [154], and the development of AFNT-111 targeting the KRAS G12V mutation [155]. Finally, Baulu et al. provide an overview of clinical results, challenges, and future directions of TCR-engineered T cell therapies in solid tumors, emphasizing the need for improved toxicity assessment and understanding of resistance mechanisms [156].

Early-phase clinical trials of CAR T-cell therapy in patients with osteosarcoma have yielded important insights into the feasibility, safety, and potential efficacy of this therapeutic approach. These trials are critical for identifying optimal conditions to maximize the effectiveness of adoptive T-cell therapies against osteosarcoma. Several studies have explored different CAR T-cell constructs targeting various antigens. [157] demonstrated the robust antitumor activity of FOLR1-CAR T cells targeting the folate receptor alpha in preclinical models of osteosarcoma, observing complete resolution of both local and pulmonary disease [156]. Hsu et al. [158] investigated EphA2-directed CAR T cells for osteosarcoma and Ewing sarcoma, reporting potent antitumor efficacy in vitro and in vivo. Yang et al. [144] proposed an alternative strategy using IL-12-based attIL12-PBMCs, which exhibited significant antitumor efficacy in osteosarcoma models.

GD2-targeted CAR T-cells

GD2-targeted CAR T-cell therapy has also shown promise in early-phase trials. GD2, a disialoganglioside expressed on the surface of various tumors including osteosarcoma, has been investigated as a target antigen [159]. [160] assessed the safety and feasibility of GD2-CAR T-cell therapy in a phase I trial, finding the therapy to be well-tolerated and achieving stable disease in 76.9% of patients on day 28 post-infusion, although all patients eventually progressed [160]. This study also identified immune determinants of CAR T-cell expansion, suggesting the potential for pre-treatment immune profiling to predict treatment response. Ramakrishna et al. [159] evaluated a third-generation GD2-CAR T-cell construct in a phase I trial, demonstrating feasibility, safety, and manageable side effects, with 76.9% of patients achieving stable disease at day 28 post-infusion. Further research has explored factors influencing GD2 expression and alternative CAR targets. A 2022 study found that GD2 expression in osteosarcoma cell lines was higher in confluent cell cultures and associated with increased sensitivity to GD2-specific CAR T-cell cytolysis [160]. Another study highlighted the potential of NKG2D-CAR T cells in targeting osteosarcoma cells [161, 162].

CAR-T cell therapy for osteosarcoma: addressing challenges and enhancing efficacy

Adoptive T cell therapy, including CAR T-cell and TCR-engineered T-cell therapies, holds immense potential for treating osteosarcoma. However, key challenges remain. Identifying suitable target antigens with high specificity to osteosarcoma cells is crucial to minimize off-tumor toxicity [143, 163]. While GD2 is a promising target, its expression on healthy cells necessitates exploring other candidates, such as B7-H3, though consistent expression remains challenging. Additionally, the immunosuppressive tumor microenvironment (TME) presents a significant hurdle. Osteosarcoma tumors evade immune surveillance through various mechanisms, including immune checkpoint molecule expression (e.g., PD-L1), immunosuppressive cytokine secretion, and recruitment of regulatory T cells and myeloid-derived suppressor cells [159, 164]. These factors can hinder the efficacy of adoptive T cell therapies by inhibiting their activity and persistence. To address these challenges, combination strategies are being investigated. Combining CAR T-cell therapy with immune checkpoint inhibitors, like pembrolizumab, may enhance antitumor immune responses by mitigating inhibitory signals within the TME. Furthermore, engineering CAR T-cells to express cytokines like IL-15 or co-stimulatory molecules could improve their persistence and functionality within the hostile TME [160]. While early-phase trials have demonstrated the feasibility and safety of adoptive T cell therapies in osteosarcoma, with some patients experiencing disease stabilization or partial responses, overcoming these challenges is essential for achieving durable and effective treatments. Ongoing research and innovative strategies are crucial for realizing the full potential of adoptive T cell therapy for osteosarcoma patients.

Antibody–drug conjugates (ADCs)

Antibody–drug conjugates (ADCs) are emerging as a powerful weapon in the fight against osteosarcoma, a devastating bone cancer that primarily affects children and adolescents. Traditional treatments like chemotherapy often come with severe side effects and limited efficacy. ADCs offer a more targeted approach by linking a potent cytotoxic drug to an antibody that specifically recognizes and binds to a protein found on the surface of cancer cells. This targeted delivery system acts like a guided missile, delivering the toxic payload directly to the tumor while sparing healthy tissues. One promising target for ADCs in osteosarcoma is LRRC15, a protein that is highly expressed on the surface of tumor cells. Preclinical studies have demonstrated the remarkable efficacy of LRRC15-targeting ADCs, such as ABBV-085. These ADCs have shown the ability to induce significant tumor regression and hinder tumor growth in experimental models of osteosarcoma, all while minimizing damage to surrounding healthy cells [165, 166]. Beyond their direct cancer-killing effects, LRRC15-targeting ADCs may also stimulate the patient’s own immune system to attack the tumor by altering the tumor microenvironment [167]. While challenges such as variable LRRC15 expression levels and the potential for drug resistance need to be addressed, these ADCs represent a significant step towards safer and more effective treatments for osteosarcoma. Early clinical trials, including a phase I study of ABBV-085 (NCT02565758), have provided encouraging results, demonstrating the safety and potential efficacy of this approach [168].

In addition to LRRC15, another promising target for ADCs in osteosarcoma is B7-H3, a protein found on the surface of various pediatric cancers. Several B7-H3-targeting ADCs have shown remarkable anti-tumor activity in preclinical studies. Ifinatamab deruxtecan (I-DXd; DS-7300) has generated considerable excitement due to its potent effects against osteosarcoma and other pediatric tumors, warranting further clinical investigation [169]. Similarly, DS7300a and m276-SL-PBD have demonstrated impressive results in preclinical osteosarcoma models, inducing substantial tumor shrinkage, slowing tumor growth, and improving survival rates [170]. Another promising B7-H3-targeting ADC, MGC018, has not only shown efficacy in shrinking tumors but has also demonstrated a favorable safety profile with minimal off-target toxicity [171]. These findings collectively suggest that B7-H3-targeting ADCs hold the potential to provide more effective and less toxic treatment options for young patients battling osteosarcoma and other solid tumors [172]. The development of both LRRC15- and B7-H3-targeting ADCs represents a major advancement in the treatment of pediatric osteosarcoma. These targeted therapies offer a ray of hope for improved outcomes and reduced side effects in young patients facing this challenging disease. Moving forward, research efforts should focus on exploring combination therapies to enhance efficacy, identifying biomarkers to predict patient response, and optimizing the drug component of the ADCs to maximize their cancer-killing power. These efforts will be crucial in translating the promising preclinical findings into tangible clinical benefits for patients with osteosarcoma.

Epigenetic modification therapy in osteosarcoma

Recent studies have highlighted novel therapeutic approaches for osteosarcoma, focusing on epigenetic modifications, including DNA methylation, histone modifications, and HDAC inhibition [173–176]. These approaches target the underlying epigenetic mechanisms driving tumorigenesis and progression. Research has focused on DNA methylation patterns in osteosarcoma, identifying differential methylation in numerous CpGs and loci, including key tumor suppressor and oncogene regions [173]. This suggests the involvement of DNMT3B and TET1 in osteosarcoma development. Further research has reviewed epigenetic mechanisms in chondrosarcoma, noting similar regulatory roles of DNA and histone modifications, indicating broader applicability to other sarcomas, including osteosarcoma [174]. These findings provide comprehensive insights into epigenetic changes in osteosarcoma, highlighting therapeutic targets and pathways that could inform treatment and prognosis [175, 176]. Additionally, a prognostic model based on epigenetic modification-related genes has been developed, offering potential tools for better clinical decision-making in osteosarcoma therapy [177–180].

Research on DNMT inhibitors for osteosarcoma treatment has revealed promising avenues for therapy by targeting epigenetic modifications. MC3343, a novel non-nucleoside DNMT inhibitor, has shown potential as an adjuvant treatment. Unlike traditional nucleoside inhibitors, MC3343 effectively induces osteoblastic differentiation and synergizes with conventional chemotherapeutics, enhancing DNA damage and cell death [181–183].

Histone modifications also play a crucial role in regulating chromatin structure and gene expression. Modifications like acetylation and methylation affect chromatin configuration, influencing transcriptional activity. Histone acetylation typically leads to transcriptional activation, while deacetylation is associated with repression [184, 185].

These modifications form a cell type-specific chromosomal bar code, ensuring consistent gene expression patterns and cell identity. Additionally, modifications like histone sumoylation play diverse roles in co-transcriptional processes, chromatin remodeling, and DNA damage response. Even in archaeal organisms, histone-based chromatin structures regulate global gene expression, highlighting the evolutionary conservation and significance of epigenetic mechanisms [184, 185].

HDAC inhibitors

Histone deacetylase (HDAC) inhibitors have shown promise in osteosarcoma therapy by modulating histone acetylation to restore normal gene expression patterns [186–188]. These agents induce cell cycle arrest, apoptosis, and differentiation of osteosarcoma cells. One example is vorinostat, which targets multiple pathways driving cancer progression and metastasis by inhibiting class I, II, and IV HDACs [186]. This leads to the accumulation of acetylated histones and proteins, impacting gene expression. Vorinostat reduces the proliferation and metastatic capacity of osteosarcoma cells, while also promoting apoptosis [186]. Another HDAC inhibitor, romidepsin, inhibits class I and II HDACs, leading to restored tumor suppressor gene expression [187, 188]. It induces cell cycle arrest and apoptosis, particularly in rapidly growing osteosarcoma cells, and targets survival signaling pathways [189]. Preclinical studies show romidepsin compromises osteosarcoma growth in vitro and in vivo, including established lung metastases [190].

Beyond vorinostat and romidepsin, other HDAC inhibitors like AR-42 and WT161 have shown greater apoptotic responses and synergistic effects with conventional chemotherapies [191, 192]. Depletion of HDAC2, a therapeutic target, has been linked to increased stemness and tumorigenic potential. Panobinostat and romidepsin, which target HDAC1/2, have proven effective in inhibiting both primary and metastatic osteosarcoma growth [191, 192].

Clinical trials investigating HDAC inhibitors for osteosarcoma treatment have shown promising results (Table 3). A phase I trial combining vorinostat with etoposide and cisplatin demonstrated tolerability and preliminary activity in patients with advanced solid tumors, including osteosarcoma. Another study focused on valproic acid in combination with other chemotherapy agents, which showed enhanced anticancer effects in osteosarcoma models. Entinostat has shown potential to upregulate FAS expression and reduce pulmonary metastasis. Panobinostat and romidepsin have also shown efficacy in both primary and metastatic osteosarcoma, with preclinical studies highlighting their ability to prevent tumor growth and metastasis [190, 193].

Table 3.

Ongoing clinical trials of epigenetic modifications for osteosarcoma treatment

NCT number	Study title	Conditions	Interventions	Phase	Enrolment number
Epigenetic therapy
02959164	Decitabine and gemcitabine for pancreatic cancer and sarcoma	Pancreatic ductal adenocarcinoma Sarcoma	Decitabine Gemcitabine	P1	36
01241162	Decitabine followed by a cancer antigen vaccine for patients with neuroblastoma and sarcoma	Osteogenic sarcoma neuroblastoma Rhabdomyosarcoma Ewings sarcoma Synovial sarcoma	Decitabine and vaccine therapy	P1	19
04833582	A study of ZN-c3 in combination with gemcitabine in subjects with osteosarcoma	Osteosarcoma	Gemcitabine	P1 P2	84
03598595	Gemcitabine, docetaxel, and hydroxychloroquine in treating participants with recurrent or refractory osteosarcoma	Recurrent osteosarcoma Refractory osteosarcoma	Docetaxel Gemcitabine Hydroxychloroquine	P1 P2	31
02429973	Trial with gemcitabine and rapamycin in second line of metastatic osteosarcoma	Osteosarcoma	Gemcitabine rapamycin	P2	33
01204450	Temsirolimus and valproic acid in treating young patients with relapsed neuroblastoma, bone sarcoma, or soft tissue sarcoma	Brain and central nervous system tumors Neuroblastoma Sarcoma Unspecified childhood solid tumor, protocol specific	Temsirolimus Valproic acid	P1	7
04897880	A study of panobinostat in pediatric patients with solid tumors including MRT/ATRT	Rhabdoid tumor And others	Panobinostat	P2	25
00112463	Depsipeptide (Romidepsin) in treating patients with metastatic or unresectable soft tissue sarcoma	Osteosarcoma and others	Romidepsin	P2	40
04308330	Vorinostat in combination with chemotherapy in relapsed/refractory solid tumors and CNS malignancies	Ewing sarcoma Rhabdomyosarcoma Wilms tumor Neuroblastoma Hepatoblastoma Germ cell tumor	Vorinostat	P1	30
01294670	Clinical study of vorinostat in combination with etoposide in pediatric patients < 21 years at diagnosis with refractory solid tumors	Solid tumors Relapsed/refractory sarcomas	Vorinostat etoposide	P1	27
00106626	Suberoylanilide hydroxamic acid in advanced solid tumors	Advanced cancer	Vorinostat Suberoylanilide hydroxamic acid (SAHA) Pemetrexed and cisplatin	P1	52

Despite their promise, the clinical use of HDAC inhibitors is limited by significant side effects, including toxicity to normal cells [194, 195], hematological issues, fatigue [196], and specific adverse effects like renal dysfunction, enhanced liver enzymes [187], and gastrointestinal disturbances [197]. The impact on normal proliferating cells and amplified toxicity in combination therapies pose challenges for patient tolerance and adherence [190, 196]. These off-target effects and systemic toxicity highlight the need for more selective compounds, targeted delivery, and careful therapeutic strategies to improve safety and efficacy [194, 195].

Combination therapies

Epigenetic therapies are emerging as a promising avenue for treating osteosarcoma by targeting the epigenetic modifications that drive the disease. These modifications, which include DNA methylation and histone modifications, can silence crucial genes and disrupt cellular processes, leading to cancer development. One promising strategy involves combining DNMT inhibitors, which block DNA methylation, with HDAC inhibitors, which prevent the removal of acetyl groups from histones. This combination therapy has demonstrated synergistic effects in reactivating silenced tumor suppressor genes and improving therapeutic outcomes [190, 198–200]. Studies have shown that combining DNMT and HDAC inhibitors can effectively reduce cancer stem-like cell populations, which are believed to contribute to tumor growth and recurrence. For example, in a mammary tumor model, the combination of 5-azacytidine (a DNMT inhibitor) and butyrate (an HDAC inhibitor) significantly reduced cancer stem cell abundance and improved survival. This synergistic effect is attributed to the disruption of key signaling pathways, such as DNA damage repair mechanisms, which enhances the overall antitumor efficacy [190, 198–200].

Furthermore, these epigenetic therapies can modulate the tumor microenvironment (TME) and increase the expression of immune-related genes, thereby enhancing the efficacy of immunotherapies. The combined action of DNMT and HDAC inhibitors leads to a more open chromatin structure, facilitating gene transcription and reactivation of epigenetically silenced tumor-suppressor and osteo/chondrogenesis-related genes [201]. This combination therapy also reactivates p53-independent apoptotic pathways, which are often silenced in multi-drug-resistant osteosarcoma cells. Consequently, this approach not only promotes growth arrest and apoptosis but also directs osteosarcoma cells towards osteoblast differentiation, counteracting drug resistance [201]. The combination of DNMT and HDAC inhibitors has shown promising results in overcoming multi-drug resistance in osteosarcoma, offering a potential solution for patients with non-responsive disease. While not specific to osteosarcoma, similar combinations have been found to enhance immune responses in other cancers, suggesting potential benefits for osteosarcoma treatment [202]. This dual approach provides a more comprehensive strategy by targeting both DNA methylation and histone modification, which enhances the reactivation of tumor-suppressor genes and improves treatment outcomes for osteosarcoma.

DNA demethylation therapy has proven effective in osteosarcomas, reactivating multiple tumor suppressors and osteo/chondrogenesis-related genes, contributing to the epigenetic reprogramming that underlies its therapeutic efficacy. Additionally, the regulation of CXCL12 via DNMT1 affects tumor progression and immune response, highlighting the potential of targeting this pathway in osteosarcoma treatment [203–205]. Ongoing research is focused on identifying specific epigenetic alterations to predict the response to these therapies, which is crucial for their successful clinical implementation (Table 3). Thus, epigenetic therapies, particularly the combination of DNMT and HDAC inhibitors, hold significant promise for treating osteosarcoma. By targeting the epigenetic modifications that drive the pathogenesis of the disease, these therapies can reactivate silenced tumor suppressor genes, modulate the TME, and enhance the efficacy of immunotherapy. Continued research in this area is crucial to optimize treatment strategies and improve outcomes for patients with osteosarcoma.

Viral therapy: a novel therapeutic strategy

Viral therapy using oncolytic viruses has emerged as a promising and innovative therapeutic approach for the treatment of osteosarcoma (Table 4). Oncolytic viruses (OVs) represent a promising therapeutic strategy for osteosarcoma by leveraging their ability to selectively infect and lyse cancer cells while sparing normal cells. Several studies have demonstrated the efficacy of OVs in osteosarcoma models. For example, the oncolytic adenovirus Delta-24-ACT has shown significant antitumor effects in osteosarcoma, promoting immune responses and extending survival in mouse models. This therapy led to both primary tumor reduction and the suppression of spontaneous metastases. Another study highlighted the potential of oncolytic virotherapy to reverse chemoresistance in osteosarcoma by suppressing MDR1 expression, thereby enhancing the efficacy of chemotherapeutic agents such as doxorubicin. Additionally, OVs have been explored in combination with immunotherapies, showing the potential to enhance immune-mediated tumor clearance and establish immune memory against osteosarcoma cells [206, 207].

Table 4.

Ongoing clinical trials of virotherapy for osteosarcoma treatment

NCT number	Study title	Conditions	Interventions	Phase	Enrolment number
Viral therapy
06171282	A clinical study on oncolytic virus injection (R130) for the treatment of advanced bone and soft tissue tumors	Osteosarcoma Sarcoma Soft tissue sarcoma Bone tumor	Recombinant oncolytic herpes simplex virus type I (R130)	Early Phase 1	9
05851456	A clinical study on oncolytic virus injection (R130) for the treatment of relapsed/refractory bone and soft tissue tumors	Osteosarcoma Sarcoma Soft tissue Sarcoma Bone tumor	Recombinant oncolytic herpes simplex virus type I (R130)	Early Phase 1	20
00503295	Safety and efficacy study of REOLYSIN® in the treatment of bone and soft tissue sarcomas metastatic to the lung	Osteosarcoma Ewing sarcoma Malignant fibrous histiocytoma Sarcoma, synovial Fibrosarcoma	REOLYSIN®	P2	53
05860374	A clinical study on oncolytic virus injection (R130) for the treatment of advanced solid tumors	Sarcoma Carcinoma Breast cancer Pancreatic Cancer Colorectal Cancer Gastric cancer Liver cancer Lung cancer Gynecologic cancer	Recombinant oncolytic herpes simplex virus type 1 (R130)	Early Phase 1	20
05644509	Study on the treatment of advanced malignant solid tumor with revottack and PD-1 inhibitor	Advanced	Oncolytic virus injection (Revottack) PD-1 inhibitor	P1	10
03647163	Ph I/II trial of systemic VSV-IFNβ-NIS in combination with checkpoint inhibitor therapy in patients with select solid tumors	Solid tumor Non-small cell lung cancer Neuroendocrine carcinoma Renal cell carcinoma	VSV-IFNβ-NIS Pembrolizumab Ipilimumab + nivolumab	P1 P2	86
02923466	Ph1 administration of VSV-IFNβ-NIS monotherapy and in combination with avelumab in Pts with refractory solid tumors	Malignant solid tumour	VSV-IFNβ-NIS VSV-IFNβ-NIS + avelumab	P1	76
01628640	Viral therapy in treating patient with refractory liver cancer or advanced solid tumors	Advanced malignant solid neoplasm Hepatocellular carinoma	Vesicular stomatitis virus	P1	17
06080984	The application of novel oncolytic virus in late stage solid tumors	Late stage solid tumors	Oncolytic virus SDJ001 Oncolytic virus YD06-1	P1	24
05205408	A study of intratumoral administration of oncolytic virus injection (RT-01) in patients with advanced solid tumors	Advanced solid tumors	Oncolytic virus injection (RT-01)	P1	7

OVs directly lyse cancer cells by replicating within them, causing cell rupture and subsequent cell death. This leads to the release of viral progeny and tumor antigens, which in turn stimulate a robust antitumor immune response, effectively turning the tumor into an in situ vaccine. Moreover, OVs can be genetically engineered to express therapeutic genes such as cytokines or immune checkpoint inhibitors, further enhancing their antitumor activity. The combination of these mechanisms results in a multifaceted attack on the tumor, promoting both direct oncolysis and immune-mediated tumor clearance [208, 209].

Recent research on oncolytic viruses in osteosarcoma treatment has demonstrated promising advancements in utilizing these viruses to selectively target and kill cancer cells. One study focused on Delta-24-ACT, an oncolytic adenovirus engineered to target cancer cells and potentiate immune responses, which showed significant antitumor effects in both primary tumors and metastases in a pediatric osteosarcoma model [206, 210, 211]. Another study explored the potential of OBP-702, an oncolytic adenovirus expressing the tumor suppressor p53, to reverse chemoresistance in osteosarcoma by suppressing MDR1 expression, highlighting the enhanced efficacy of combination therapy [207, 212, 213]. Additionally, a comprehensive review emphasized the safety and potential of various oncolytic viruses in treating osteosarcoma, noting the need for combination strategies to maximize therapeutic benefits [214].

Oncolytic viral therapy (OVT) offers a promising approach for osteosarcoma treatment, harnessing the ability of viruses to selectively target and destroy cancer cells while stimulating the immune system to fight the tumor (Table 4). However, translating this promising approach into effective human therapies faces several challenges.

Differences between animal models and humans

One significant hurdle is the notable differences observed between human and animal models of osteosarcoma. These differences, stemming from variations in immune activation, tumor microenvironment (TME), and genetic factors, can complicate the translation of findings from preclinical studies to human applications. For instance, canine models, while valuable, exhibit a median survival of 10–12 months, contrasting with the 70% 5-year survival rate in humans [215]. This disparity highlights the challenges in extrapolating results from animal studies to human patients. While oncolytic viruses (OVs) like Delta-24-ACT have shown promising tumor regression, immune activation, and enhanced survival in murine models, and oncolytic vesicular stomatitis virus (VSV-IFNβ-NIS) elicits robust immune responses in canines [216], the complexity of the human immune landscape and tumor heterogeneity often leads to unpredictable responses. Furthermore, human and canine immune systems exhibit distinct characteristics. Human T-cells show a stronger Th1 bias, producing higher levels of IFN-γ compared to canine T-cells, influencing the efficacy of oncolytic therapies [217]. Conversely, canine macrophages are more responsive to IFN-γ, leading to distinct immune activation profiles [217]. The TME also differs significantly between species. In humans, immunosuppressive monocytes and macrophages can limit OVs efficacy, necessitating combination therapies like immune checkpoint inhibitors. In contrast, canine models often show heightened inflammation and micronecrosis in response to OVs, reflecting differences in immune and TME dynamics [218, 219]. These findings underscore the need for tailored approaches in OVs research and clinical applications, accounting for species-specific immune and tumor responses to optimize outcomes in treating osteosarcoma.

Limitations and challenges of OVT

Beyond species-specific variations, OVT faces inherent limitations and potential side effects. Key challenges include difficulties in viral delivery due to the TME and intratumoral heterogeneity, which restrict viral spread and reduce therapeutic efficacy [220]. The complex vascular structure of osteosarcoma tumors further complicates effective distribution [220]. The immunosuppressive TME, often dominated by M2-polarized tumor-associated macrophages, can inhibit immune responses against tumor cells, limiting the therapeutic impact [221]. Additionally, the direct lytic activity of OVs may be insufficient, leading to suboptimal tumor cell death [221]. While generally safe, OVT carries a risk of adverse effects, particularly in immunocompromised patients or those undergoing concurrent therapies [214, 222]. The potential for immune evasion and the frequent need for combination therapies further underscore the complexity of effectively using OVs for osteosarcoma.

Overcoming challenges and future directions

Despite these challenges, ongoing research is focused on overcoming the obstacles hindering OVT. Efforts are underway to develop viruses with improved target fidelity and safety, leveraging genetic engineering to enhance conditional viral replication within tumor cells. Combining OVs with other cancer therapies, such as chemotherapy and immunotherapy, is also being explored to improve patient outcomes. For instance, studies have focused on the synergy of OVs with various cancer therapeutics to overcome viral suppression and stromal barriers [223], while others have discussed advancements in OV-modification strategies and the promising clinical outcomes of OV combinations with other therapies [224]. The challenges in OV delivery and the potential of combination therapies have also been reviewed, underscoring the need for further optimization of the delivery routes [225]. Continued research and clinical trials are essential to address these challenges and fully realize the therapeutic potential of oncolytic viruses for osteosarcoma treatment. By improving viral delivery, enhancing tumor cell killing, and stimulating robust anti-tumor immunity, OVT holds the promise of becoming a valuable weapon in the fight against osteosarcoma.

Conclusion

In conclusion, advancements in targeted therapies and immunotherapies have opened new avenues for osteosarcoma treatment. IGF-1R inhibitors and TKIs show significant promise, but their efficacy is often hampered by tumor heterogeneity and compensatory signaling pathways. Combination therapies with oncolytic viruses offer potential solutions to these challenges. Immunotherapeutic strategies, particularly CAR T-cell therapies targeting GD2, have shown encouraging results in early phase clinical trials. Continued research and clinical trials are essential to optimize these treatments, overcome resistance mechanisms, and improve patient outcomes.

Abbreviations

CAR-T

Chimeric antigen receptor-t cells

DNMT

DNA methyl transferase

HDAC

Histone deacetylase

HER2

Human epidermal growth factor receptor 2

IGF-1R

Insulin like growth factor-1 receptor

ICE

Ifosfamide, carboplatin, and etoposide

IE

Ifosfamide and Etoposide

JAK/STAT

Janus kinase/signal transducers and activators of transcription

KIT

Receptor tyrosine kinase

LSS

Limb-salvage surgery

MET

Mesenchymal epithelial transition

mTOR

Mammalian target of rapamycin

MTX

Methotrexate

MIOS

Multi-Institutional Osteosarcoma Study

NF-κB

Nuclear factor kappa light-chain-enhancer of activated B cells

NK cells

Natural killer cells

OVs

Oncolytic viruses

OS

Overall survival

PD-1

Programmed death-1

PD-L1

Programmed death ligand-1

PFS

Progression-free survival

PI3K/Akt

Phosphatidylinositol 3-kinase/protein kinase B

PDGFR

Platelet-derived growth factor receptor

TCR

T-cell receptor

TME

Tumor microenvironment

VEGFR

Vascular endothelial growth factor receptors

Author contributions

All authors contributed to the manuscript. All authors read and approved the final version of the manuscript. VKM—conceptualization of the paper, design of the paper, data collection and analysis, a major contributor in writing the manuscript, supervising, approval of the final version of the manuscript. AGM—design of the paper, data collection and analysis, creation of the figures. SHP- a major contributor in review and editing of the manuscript. KCN—a major contributor in writing the manuscript, supervising, Funding acquisition, approval of the final version of the manuscript.

Funding

This research was supported by the Hallym University Medical Center Research Fund (MH 4.0).

Data availability

No datasets were generated or analysed during the current study.

Declarations

Ethics approval and consent to participate

Not applicable (review paper).

Competing interests

The authors declare no competing interests.