Immunotherapy as a potential treatment approach for currently incurable bone metastasis

Abstract

Once cancer metastasizes to the bone, the prognosis of cancer patients becomes extremely poor. Unfortunately, the current most successful treatment for bone metastasis can extend their survival by only a few months. Although recent studies have revealed promising impacts of cancer immunotherapies, their treatment efficacy on bone metastatic diseases remains controversial. Therefore, in this review, we discussed (i) preclinical and clinical evidence of the immunotherapeutic strategies for cancer bone metastasis, mainly focusing on cell-based immunotherapy, cytokine-based immunotherapy, and immune checkpoint blockade, and (ii) current shortcomings of immunotherapy for bone metastasis and their potential future directions. Although the evidence on treatment efficacy and safety, as well as long-term effects, is limited, immunotherapies could induce partial or complete remissions in a few cancer patients with bone metastasis. However, there are still hurdles, such as the immunosuppressive nature of the bone marrow microenvironment and poor distribution of cell-based immunotherapies to bone, that need to be overcome to enhance the treatment efficacy of immunotherapies on bone metastasis. While it is apparent that further investigation is needed regarding immunotherapeutic treatment efficacy in patients with bone metastasis, this therapy may prove to be clinically novel in this subset of cancer patients.

Introduction

Bone metastasis is one of the major causes of death among cancer patients. Indeed, once cancer cells metastasize to the bone, the 5-year survival rate of cancer patients decreases to 1–13% (e.g., 1–2%: lung cancer, rectum cancer, and cervix cancer; 3–10%: colon cancer and prostate cancer; and 13%: breast cancer) [1]. Additionally, the median survival time of patients with bone metastasis is approximately 12–33 months [1]. Although several combinations of systemic treatments (e.g., hormone therapies, chemotherapies, bone-targeted therapies, radiopharmaceuticals, etc.) are known to enhance the overall survival (OS) of patients with bone metastases, they only increase the OS by several months [2]. Therefore, bone metastasis is still considered incurable, and there is a clear need to develop new treatment strategies to eradicate bone metastasis.

Recently, cancer immunotherapies have been viewed as the major breakthrough in cancer treatments. For example, pembrolizumab, a humanized antibody targeting programmed cell death protein 1 (PD-1) on T-cells, has significantly improved the OS of cancer patients, compared to those treated with vehicle [e.g., non-small cell lung cancer (NCT02578680, 22 vs. 10.6 months, p < 0.0001); head and neck squamous cell carcinoma (NCT02358031, 13 vs. 10.7 months, p = 0.0067); and urothelial carcinoma (NCT02256436, 10.3 vs. 7.4 months, p = 0.004)] [3]. Although immune cells are produced in the bone marrow and primarily reside there, the efficacy of cancer immunotherapies is not as high as that of the previously stated conventional treatment strategies for bone metastasis. It has recently been suggested that cancer immunotherapies can be potential alternative treatments for patients with bone metastasis as they can induce a complete remission in those patients [4]. However, little is known as to the mechanisms where by bone metastatic cancer cells and/or the bone marrow microenvironment limit the efficacy of cancer immunotherapy on bone metastasis. We must know how to overcome the limitation of its efficacy before applying cancer immunotherapies for bone metastatic disease.

Therefore, in this review, we will (i) highlight the immunotherapeutic strategies for cancer bone metastasis from previous clinical trials or preclinical studies and (ii) discuss the limitations of these therapeutic strategies to direct future studies.

Immunotherapies for bone metastasis

Cancer immunotherapy is a treatment strategy that uses the patients’ own immune system to eradicate cancer cells. There are several types of immunotherapies, including cell-based immunotherapy, antibody-based immunotherapy, cancer vaccine, oncolytic virus treatment, cytokine-based immunotherapy, and immune checkpoint blockade (ICB). Although many of these treatments have been tested as therapeutic strategies for bone metastasis, we will focus mainly on the effects of cell-based immunotherapy, cytokine-based immunotherapy, and ICB on bone metastasis in this review.

Cell-based immunotherapies for bone metastasis

Lymphokine-activated killer (LAK) cell therapy

Cell-based immunotherapies (or adoptive immunotherapies) are a type of therapy that uses patients’ own immune cells (activated ex vivo and then reintroduced into the patient’s body) to target cancer cells [5]. In the early development of cell-based immunotherapies, ex vivo activated peripheral blood leukocytes derived from cancer patients, referred to as lymphokine-activated killer (LAK) cells, were used. Interestingly, LAK cells were also used to treat bone metastatic patients. LAK cells have been tested to treat advanced renal cell carcinoma (RCC) patients with bone metastases in three clinical trials with greatly differing efficacy: (trial 1) some patients presented with partial remission (> 50% decrease in bone metastatic lesions), although recurrence occurred within 6 months of follow-up [6, 7]; (trial 2) IL-2-activated LAK cells induced a partial response in patients; however, they died 10 months and 28 months after LAK treatment [8]; and (trial 3) when IL-2-activated LAK cell therapy was given with interferon-alpha (IFN-α) [9], patients’ bone metastatic lesions progressed [10]. Overall, LAK therapy presents limited efficacy on bone metastasis in addition to renal toxicity.

Autologous cytotoxic T cell therapy

Although LAK cell treatment was somewhat effective, it has been found to induce adverse side effects, such as renal failure, in some patients; therefore, cytotoxic T cells may present as a better option for bone metastasis treatment as they possess specific tumor-killing abilities. It has been demonstrated that when the resting cluster of naïve T cells is treated with anti-CD3 antibodies together with IL-2 ex vivo, they differentiate into cytotoxic CD8⁺ T cells [11]. In a preclinical animal study of melanoma bone metastasis, when mice were intracardiacally inoculated with melanoma cells (B16), all the mice developed bone metastatic lesions (n = 5) [12, 13]. However, when the mice were given the donor T cells (obtained from tumor bearing mice), which were activated with anti-CD3 antibodies and IL-2 ex vivo, the treatment hindered bone metastatic progression and prevented the occurrence of signs of distress, paralysis, or other disturbances in motor activity in recipient mice (n = 5) [12, 13]. Moreover, the therapeutic benefit of the activated donor T cells was reversed when mice were treated with anti-CD8 antibodies (n = 5) [12, 13]. Cytotoxic T cells were also used to treat a breast cancer patient with bone metastasis, in which the patient’s own peripheral blood mononuclear cells (PBMCs) were activated ex vivo using a combination of anti-CD3 antibodies, IL-2, and human leukocyte antigen (HLA)-A2-matched GC022588 gastric cancer cell line [14]. In this clinical case report, the patient experienced a return in serum carcinoembryonic antigen levels (which are sometimes increased in breast cancer patients) to normal range and importantly, bone pain relief [14]. However, when RCC patients with bone metastases (n = 4) were treated with ex vivo activated cytotoxic T cells derived from the patient’s PBMCs using the combination of anti-CD3 antibodies and anti-CD28 antibodies, none of them achieved remission, although two patients showed some regression of bone metastatic lesions, and the only adverse events observed were mild to moderate influenza-like symptoms [15].

An alternative method to activate cytotoxic T cells in patients is through infusion of activated dendritic cells (DCs), an antigen-presenting cell type [16]. Briefly, in this process DCs are first stimulated with granulocyte–macrophage colony-stimulating factor (GM-CSF) and prostatic acid phosphatase (PAP, known to be specifically expressed on the surface of prostate cancer cells [17]), and the resulting DCs are able to activate PAP-targeting cytotoxic T cells [17]. This concept has been used to treat prostate cancer through products, such as Sipuleucel-T (APC8015) [17]. Sipuleucel-T therapy has improved the survival of metastatic castration-resistant prostate cancer (CRPC) patients. In two phase III clinical trials of Sipuleucel-T in patients with metastatic CRPC, it was revealed that Sipuleucel-T extended the median survival of treated patients [Trial I (n = 127), Sipuleucel-T survival: 25.9 months vs. placebo: 21.4 months, p < 0.01 [18]; Trial II (n = 512), Sipuleucel-T survival: 25.8 months vs. placebo: 21.7 months, p = 0.03 [19]]. In addition, adverse events, including anorexia, anxiety, depression, flank pain, and contusion, as well as hydronephrosis, in Sipuleucel-T group were less frequent than those in placebo group (7.1% in placebo group vs. 3.8% in Sipuleucel-T group) [19]. Moreover, APC8015F, a version of Sipuleucel-T salvage treatment that is initiated using cryopreserved cells from patients, has also shown to enhance patient survival. A historical control trial of APC8015F in metastatic CRPC patients selected from the control arm of previous studies (NCT00005947, NCT01133704, and NCT00065442) demonstrated that APC8015F improved the median survival of those patients [APC8015F: 20.0 months vs. historical controls (NCT00005947, NCT01133704, and NCT00065442): 9.8 months, p < 0.001] [20]. Although these trials were not intended to determine the treatment effects of DC-based autologous cytotoxic T cell therapy on bone metastatic patients, these findings suggest that DC-based autologous cytotoxic T cell therapy can be an effective treatment for prostate cancer patients with bone metastases, since 72.8% of metastatic CRPC patients develop bone metastases [21].

Tumor-infiltrating lymphocyte (TIL) therapy

While LAK cells and autologous cytotoxic T cells used for cell-based immunotherapies are obtained from patients’ peripheral blood, the use of T cells existing in the tumor as cell-based immunotherapies has recently been appreciated. This treatment strategy is known as tumor-infiltrating lymphocytes (TIL) therapy. Intriguingly, compared to LAK cell and autologous cytotoxic T cell therapies, TIL therapy appears less toxic and has more specific tumor-killing abilities [22, 23]. When mice inoculated intravenously with MC-38 colon adenocarcinoma were treated with LAK cell therapy and cyclophosphamide, all the mice developed liver metastases [24]. However, when the MC-38-bearing mice were treated with TIL therapy and cyclophosphamide (n = 12), none of mice developed liver metastases [24]. In a phase I clinical trial of TIL therapy in metastatic malignant melanoma (n = 20, including one bone metastatic patient), TIL therapy with a single dose of cyclophosphamide induced either a complete or partial remission in 11 patients (the patient with bone metastasis achieved a partial remission for 4 months) [25]. The adverse effects observed in this study were mainly chills (10/20) and gastrointestinal symptoms, including nausea and vomiting (10/20) and diarrhea (9/20) [25]. Importantly, a randomized comparative clinical trial in cancer patients (lung cancer, breast cancer, or gastric cancer) with bone metastases (n = 20) using TIL therapy revealed that the patients’ median survival time was significantly extended by the combination of TIL therapy and standard of care therapy (183 days vs. 82.3 days, p < 0.05), compared to that of patients treated with standard of care therapy alone [23]. In addition, TIL therapy produced an analgesic effect on patients with bone metastasis, which ultimately improved quality of life; however, adverse events were not reported [23].

Cytokine-based immunotherapy for bone metastasis

As mentioned above, cytokines have been used to stimulate immune cells ex vivo to induce their tumor-killing abilities. However, cytokines themselves have also been known as immune inducers (or immune modulators). IL-2 is well known to enhance tumor-killing abilities of T cells, natural killing (NK) cells, and B cells [8]. When kidney cancer patients with bone metastases (n = 6) were co-treated with famotidine (antihistamine, known to increase TIL) and high-dose IL-2, three of these patients showed response to this treatment strategy (for 3, 7, and 14 months) and one patient achieved complete remission in two bone sites for 14 months [26]. IFN-α was known to promote the differentiation and activity of host immune cells [9]. Conversely, a phase III clinical trial of RCC patients with metastases (N = 750) demonstrated IFN-α did not improve OS in patients with or without bone metastases compared to Sunitinib [27].

Immune checkpoint blockade for bone metastasis

Immune checkpoint molecules consist of a receptor and ligand complex, which regulates immune activities [28]. There are two types of immune checkpoint molecules: stimulatory checkpoint molecules [e.g., CD27 (receptor)/CD70 (ligand) and CD28/CD86 (a.k.a. B7–2) or CD80 (a.k.a. B7–1)] and inhibitory checkpoint molecules [e.g., CD279 (a.k.a. PD-1)/CD274 [a.k.a. programmed cell death protein ligand 1 (PD-L1)] and CD152 [a.k.a. cytotoxic T-lymphocyte associated protein 4 (CTLA-4)]/CD86 (a.k.a. B7–2) or CD80 (a.k.a. B7–1)] [28, 29]. The receptors, which are commonly expressed on the surface of T and B cells, are responsible for exerting stimulatory or inhibitory immune responses after binding with their respective ligands.

In the tumor microenvironment, tumor cells and tumor-infiltrating immune cells highly express PD-L1, which upon binding to PD-1 on T cells directly inhibits T cell-mediated immune response to cancer. In addition, since CD28 binding with CD86 or CD80 results in T cell activation, high levels of CTLA-4 on tumor-infiltrating T regulatory cells (Treg) competing with CD28 for CD86 or CD80 binding results in the inactivation of T cells [30, 31]. Both mechanisms ultimately contribute to cancer cell evasion of immune surveillance [28]. As stated above, blocking inhibitory checkpoint molecule pathways has been somewhat successful to treat cancer and has been appreciated as a breakthrough cancer treatment [32]. Antibodies, which target either the PD-1/PD-L1 pathway or the CTLA-4 pathway, were developed in the clinic for cancer treatment. These antibodies, such as ipilimumab (anti-CTLA-4), nivolumab (anti-PD-1), and atezolizumab (anti-PD-L1), have been FDA approved to treat various types of cancers including melanoma, non-small cell lung cancer (NSCLC), and RCC [32]. However, currently, these treatment strategies have negative treatment effects on patients with bone metastases.

In a two-center retrospective study of the combination of anti-PD-1 therapy and anti-angiogenic therapy in metastatic NSCLC patients (n = 57), bone metastasis was considered a negative predictor of prognosis (p < 0.01) [33]. Moreover, none of the bone metastatic patients responded to this combination strategy, although its objective response rate was 19.3% [33]. Similarly, a prospective, single-institution study of the second line anti-PD-1/PD-L1 therapy in metastatic NSCLC patients (n = 66) revealed that bone metastasis was an independent predictor of shorter survival [hazard ratio = 1.89 (confidence interval (CI) 1.02–3.51, p = 0.049)] [34]. For advanced RCC, a phase III clinical trial of nivolumab, an anti-PD-1 antibody, demonstrated that nivolumab (n = 76) expanded median OS in bone metastatic patients [18.5 months, 95% CI 10.2—not reached (NR)] compared to everolimus (mTOR inhibitor) chemotherapy (n = 70) (13.8 months, 95% CI 7.0–16.4) [35]. No obvious difference in adverse event rates was observed in bone metastatic patients from the nivolumab group or the everolimus group [35]. Another phase III clinical trial compared ipilimumab (n = 399), an anti-CTLA-4 antibody, with placebo (n = 199) in patients with metastatic (80% bone metastatic) chemotherapy-naïve, CRPC. Unfortunately, no significant difference in median OS was observed between the two groups, though ipilimumab increased median progression-free survival (PFS) for 1.8 months [hazard ratio (HR), 0.67; 95.87% CI 0.55 to 0.81] [36]. Bone pain response was not evaluated due to the small number of patients experiencing pain [36]. Despite limited efficacy of ICB on bone metastasis, ICB may help control bone remodeling. Nivolumab was found to prevent bone destruction by inhibiting TRAP + osteoclast differentiation in a bone metastatic mouse model, although there was no effect on tumor growth [37].

Current shortcomings of immunotherapy for bone metastasis

As we have discussed, some types of immunotherapies (including cell-based and cytokine-based) have demonstrated moderate treatment efficacy in bone metastatic cancer patients (Table 1); however, bone metastasis overall remains a disease that does not respond well to immunotherapies [20, 33–36, 38]. Although their efficacy has been limited at this point, immunotherapies have potential for the treatment of bone metastasis, since bone marrow harbors various immune cells. To enhance the treatment efficacy of immunotherapies on bone metastasis, we must first identify the key barriers limiting their efficacy. Like other organs, bone marrow contains not only immune-supportive cells, but also immunosuppressive cells. However, bone marrow contains more immunosuppressive cells than other organs [39] in order to protect hematopoietic stem cells, which are crucial for maintaining blood formation from autoimmune insults [40]. For example, mesenchymal stem cells (MSCs) are known as one of the major components of the bone marrow microenvironment, or the “niche,” for hematopoietic stem cells. MSCs were also reported to inhibit T cells’ production of interferon-gamma (IFN-γ) in vitro, which is known as an anti-tumor molecule [41]. This implies MSCs may play a role in the inhibition of immune response to bone metastasis. Moreover, a recent bone marrow single cell sequencing analysis revealed additional contributors to the immunosuppressive environment in bone metastasis. This study compared four groups of patient samples: (i) solid bone marrow prostate cancer metastatic tissue, (ii) liquid bone marrow from the vertebral level of spinal cord compression, (iii) liquid bone marrow from vertebral body distant from the tumor site, and (iv) non-cancer patient bone marrow [42]. After analyzing the sequencing data, they found differences in monocyte and macrophage cell populations that were present. In group (i) with solid bone marrow metastatic tissue, tumor inflammatory monocytes (TIMs) and tumor associated macrophages (TAMs) were observed, while in groups (ii-iv) such populations were not observed [42]. TIMs and TAMs may ultimately contribute to an immunosuppressive microenvironment through support of tumor growth by expansion of the suppressive Treg cells [42]. In the bone marrow, bone metastatic cancer cells escape from immunosurveillance by indirectly inducing anti-inflammatory cytokine transforming growth factor-beta (TGF-β), which is released from the bone resorbed by osteoclasts [43]. TGF-β inhibits T-helper-1 (Th1) development [43], which is known to promote CD8 T cell activity by cytokines such as IL-2 [44]. Overall, current findings suggest that bone metastasis has been difficult to treat with immunotherapies (Fig. 1, left). Additionally, poor distribution of cell-based immunotherapies to bone has been considered as one of their shortcomings. Indeed, a rodent study revealed that LAK cells distribute thirty times higher to lung, liver, and spleen than femur, muscle, and prostate [45]. Moreover, a study of adoptive cellular therapy for acute myeloid leukemia also unveiled that double negative T cells infused into mice lowly distributed to the bone marrow compared with the heart, liver, kidney, and blood [46]. The distribution of therapeutic chimeric antigen receptor-modified (CAR)-T cells was also found to be much lower in the bone marrow than in the spleens, lungs, livers, and kidneys of mice with acute lymphoblastic leukemia [47]. This phenomenon may be caused by the way of therapeutic cell injection since intravenous administration mainly passes through lung, kidney, and liver.

Table 1.

Summary of immunotherapy trials for cancer patients with bone metastasis

Immunotherapy	Study type	Treatment	Cancer	Outcome	Adverse events	Metastatic sites	PMID
Adoptive immunotherapies	Clinical Trial (Phase Unspecified, 1987, USA)	LAK	RCC	5 Bone metastatic patients: 2/5 had PR [Patient 1: < 6 months and Patient 2: no recurrence within 6 months]	Malaise, fever, renal failure	Bone, lung, brain, skin, liver, subcutaneous tissue, inferior vena cava	PMID: 2845776
Adoptive immunotherapies	Clinical Trial (Phase Unspecified, 1994, Japan)	LAK	RCC	2 Bone metastatic patients: 2/2 had PR [patient 1: < 10 months and patient 2: < 28 months]	Hypotension, heart failure, artery occlusion	Bone, muscle, brain, liver, lung, pleura, peritoneum	PMID: 7645136
Adoptive immunotherapies	Clinical Trial (Phase I, 1995, Japan)	LAK + IFN-#x3B1;	RCC	PD	Fever, anemia, gastric ulcers	Bone, lung, brain	PMID: 7609359
Adoptive immunotherapies	Case Report (2006, Japan)	Autologous T cell	BCa	CEA reduction, bone pain relief	NA	Bone	PMID: 16786141
Adoptive immunotherapies	Clinical Trial (Phase I, 2003, USA)	Autologous T cell	RCC	2 Bone metastatic patients: 2/2 had PR	Influenza-like symptoms	Bone, lung, brain, liver	PMID: 14506142
Adoptive immunotherapies	Clinical Trial (Phase III, 2006, USA)	Sipuleucel-T	CRPC	Extend median survival [25.9 months vs. placebo 21.4 months]	Anorexia, anxiety, depression, flank pain, hydronephrosis	Bone	PMID: 16809734
Adoptive immunotherapies	Clinical Trial (Phase III, 2010, USA)	Sipuleucel-T	CRPC	Extend median survival [25.8 months vs. placebo 21.7 months]	Anorexia, anxiety, depression, flank pain, hydronephrosis	Bone	PMID: 20818862
Adoptive immunotherapies	Clinical Trial (Phase III, 2015, USA)	Sipuleucel-T salvage	CRPC	Extend median survival [20.0 months vs. historical controls 9.8 months]	Chills, nausea, fever	Bone	PMID: 25943532
Adoptive immunotherapies	Clinical Trial (Phase I, 1988, USA)	TIL	Melanoma	CR or PR [10 patients without bone metastasis] and PR [1 patient with bone metastasis: 4 months]	Chills, nausea, vomiting, diarrhea	Bone, lung, brain, subcutaneous tissue, spleen	PMID: 3264384
Adoptive immunotherapies	Clinical Trial (Phase Unspecified, 1994, China)	TIL	Lung cancer, BCa, Gastric cancer	Extend median survival [183 days vs. standard of care therapy 82.3 days]	NA	Bone, lung, brain, liver	PMID: 7956700
Cytokine-based immunotherapy	Clinical Trial (Phase Unspecified, 2006, USA)	Famotidine + IL-2	Kidney cancer	6 Bone metastatic patients: 1/6 had CR [14 months] and 2/6 had PR [Patient 1: 3 months, Patient 2: 7 months]	Hypertension, elevated creatinine, pulmonary edema	Bone, lung, liver	PMID: 17105423
Cytokine-based immunotherapy	Clinical Trial (Phase III, 2008, USA)	IFN-α	RCC	Did not improve OS compared to Sunitinib	Fatigue, anemia, lymphopenia, nausea	Bone, lung, liver	PMID: 19487381
Immune checkpoint blockade	Retrospective Study (2020, China)	anti-PD-1 + antiangiogenic agent	NSCLC	ORR 19.3%	Proteinuria, Hypotension, Fatigue	Bone, liver, brain, adrenal gland	PMID: 33039958
Immune checkpoint blockade	Clinical Trial (Phase III, 2017, USA, France, Italy, Spain, Australia, Denmark, Finland, UK, Germany, Japan)	anti-PD-1	RCC	Extend median survival [18.5 months vs. chemotherapy 13.8 months]	Adverse events incidence reduced compared to chemotherapy	Bone, lung, liver	PMID: 28262413
Immune checkpoint blockade	Clinical Trial (Phase III, 2017, USA, Portland, France, Australia, Canada, Poland, Spain, Germany, Brazil, Mexico, Denmark, the Netherlands)	anti-CTLA-4	CRPC	Increased median PFS by 1.8 months compared to placebo	Diarrhea, rash, pruritus	Bone	PMID: 28034081

LAK lymphokine-activated killer cell therapy, TIL tumor-infiltrating lymphocytes therapy, RCC advanced renal cell carcinoma, BCa breast cancer, CRPC castration-resistant prostate cancer, NSCLC non-small cell lung cancer, CR complete response, PR partial response, PD progressed disease; OS overall survival, PFS progression-free survival, ORR objective response rate

Fig. 1.

Models of specific immune evasion in bone metastasis and the potential therapeutic strategies. As shown in the left panel, when bone metastatic cancer cells interact with osteoclasts in the bone marrow, these cancer-associated osteoclasts mediate bone resorption, in which the resorbed bone releases transforming growth factor-beta (TGF-β). TGF-β released from bone is known to inhibit the differentiation of CD4 T cells (CD4 T) into T helper 1 cells (Th1). Additionally, bone marrow mesenchymal stem cells (MSCs) prevent the secretion of interferon-gamma (IFN-γ, a cytokine known to suppress tumor growth) from CD8 T cells (CD8 T) (based on in vitro studies). These bone marrow-specific mechanisms allow bone metastatic cancer cells to evade the immune system, which contributes to the limited treatment efficacy of immunotherapy for bone metastasis. Conversely, as shown in the right panel, TGF-β that was released from the bone can be blocked with anti-TGF-β antibodies, which allows CD4 T cells to differentiate into Th1 cells. By releasing cytokines, Th1 cells are then able to support CD8 T cells to kill tumor cells. Further, treatment with an indoleamine 2,3-dioxygenase (IDO1) inhibitor, 1-methyl-l-tryptophan (1-l-MT) can prevent the MSC-mediated inhibition of IFN-γ release from T cells (based on in vitro studies). Through activation of CD8 T cells and IFN-γ release, cytotoxicity to bone metastatic cancer cells can be increased, thus enhancing immune response in the bone marrow microenvironment. We propose that by enhancing CD8 T cell tumor-killing ability through anti-TGF-β antibody and 1-l-MT treatment, efficacy of immunotherapies for bone metastatic disease may be improved

Future directions and conclusions

The current mainstream treatment strategies for bone metastasis are the combinations of bone-targeted treatments [e.g., bisphosphonates and denosumab (a human monoclonal anti-receptor activator of nuclear factor κB ligand (RANKL) antibody), and radium-223 dichloride] and systemic treatments (e.g., hormone therapies, chemotherapies). While these strategies have been somewhat effective (improving OS by a few months), they are mainly palliative [48]. Therefore, new treatment strategies other than combination therapies to eradicate bone metastasis are urgently needed. Although, to date, bone metastasis has been considered as an “immune-cold” disease, immunotherapies have potential to become paradigm shift therapies, since bone marrow contains a huge number of immune cells. However, as stated above, there are still several difficulties that need to be overcome before applying immunotherapies for bone metastasis. One example is the immunosuppressive effects of bone marrow. If these effects can be reversed, the efficacy of immunotherapies can be enhanced. Indeed, anti-TGF-β antibody treatment enhanced the efficacy of ICB immunotherapy. In a metastatic CRPC mouse model, treatment with anti-CTLA4 antibody combined with anti-TGF-β antibody significantly expanded intra-tumoral CD8⁺ T cells, whereas single treatment with either one of the two antibodies could not achieve obvious CD8⁺ T cell expansion [43]. Further, anti-TGF-β antibody treatment improved the OS (p = 0.06) and reduced the burden of bone metastases by approximately 70–80% (p = 0.001) in a metastatic breast cancer mouse model compared to the control group [49]. Considering that osteoclasts mediate TGF-β release from the bone, combining inhibition of osteoclasts by anti-RANKL antibody with ICBs has induced a dramatic partial response of bone metastasis and complete bone pain relief in a melanoma patient with widespread bone metastases [50]. Currently, a phase Ib/II clinical trial (ACTRN12618001121257) comparing combination therapy of RANKL inhibitor and ICB with ICB alone in resectable NSCLC is ongoing [51]. Moreover, the indoleamine-pyrrole 2,3-dioxygenase (IDO1) inhibitor, known as 1-methyl-l-tryptophan (1-l-MT), treatment may be a potential way to inhibit the immunosuppressive effects mediated by MSCs [52]. In a co-culture experiment of MSCs with T cells (exposed to immunostimulants, such as lipopolysaccharides (LPS) or poly(cytidylic-inosinic) acid), 1-l-MT significantly increased T cell proliferation compared to nontreatment control [52]. Although further in vivo experiments are needed, these findings suggest that by combining these anti-immunosuppressive strategies, the efficacy of immunotherapies for bone metastasis can be improved (Fig. 1, right).

To enhance the treatment efficacy and reduce potential off-target effects of cell-based immunotherapies, their bone tropism needs to be improved. Recent gene editing techniques enable us to engineer T cells to recognize specific tumor antigens; these strategies include (i) T-cell receptor (TCR)-T cell technology: stimulates TCR to recognize tumor antigen, and (ii) CAR-T cell technology: artificial antigen receptors to recognize tumor antigen [53]. To engineer TCR-T cells, modified TCRs that specifically recognize tumor antigens are integrated onto the T cell membrane [53]. Compared to TCR-T cells, CAR-T cells use chimeric antigen receptors that include scFv domains derived from antibody, CD3ζ, and transmembrane domain, which enable CAR-T cells to recognize cell surface antigens directly without the presentation of major histocompatibility complex (MHC) [53]. Therefore, these technologies may be used to enhance bone tropism of cell-based immunotherapies if TCR or CAR-T cells can be effectively designed to target cells in the bone marrow (e.g., osteoblasts, osteoclasts). By doing so, these T cell engineering technologies have a potential to enhance the treatment efficacy of cell-based immunotherapies on bone metastatic cancer cells. Further studies are warranted prior to applying this idea as a means to improve their bone tropism.

Although recent clinical trials have demonstrated that ICB has limited effects on cancer patients with bone metastases, several case reports have revealed that ICB could induce complete remission in cancer patients with bone metastases (pembrolizumab, an anti-PD-1 antibody, in three bone metastatic lung cancer patients [54, 55]; and ipilimumab in two bone metastatic prostate cancer patients [56]). Additionally, as discussed, therapies using cytotoxic T cells have been somewhat effective for bone metastasis. Therefore, the ICB can be considered as a potential treatment for bone metastasis if the functions of cytotoxic T cells are enhanced. Intriguingly, recent studies revealed that alteration of the gut microbiota of cancer patients can improve the treatment efficacy of ICB by enhancing tumor-killing activities of cytotoxic T cells [32, 57–60]. Thus, strengthening cytotoxic T cell activities by manipulating the gut microbiota may improve the treatment efficacy of ICB on bone metastatic cancer patients. Additional studies are needed to confirm this hypothesis.

Although further clinical evaluation and improvement of its treatment efficacy are clearly warranted, immunotherapy could be a breakthrough for cancer patients with bone metastases and help alleviate many of the challenges and hurdles faced by bone metastatic patients and their families.