Introduction

In the past decade, for the first time, there has been a decrease in the incidence of lung cancer, with the incidence in women decreasing at only about half the rate of men (1.1% vs. 2.6% annually).[1] However, with an estimated 1.8 million deaths each year worldwide, lung cancer remains the leading cause of cancer death, accounting for 18% of cancer deaths.[2] As of 2023, lung cancer is predicted to account for 238,340 new cases and 127,070 deaths in the United States alone.[1] Contributing to 85% of the histological types of lung cancer, non-small cell lung carcinoma (NSCLC) encompasses lung adenocarcinoma (40% of all lung cancers), squamous cell carcinoma (30% of all lung cancers), large cell (undifferentiated) carcinoma (15% of all lung cancers), and other rare mixed histotypes such as adenosquamous carcinoma and sarcomatoid carcinoma.[3] On the other hand, ~15% of all lung cancers are classified as small cell lung cancer (SCLC), also referred to as oat cell cancer, which grows and spreads faster than NSCLC.

Different kinds of cancer treatment approaches have been developed to fight lung cancer with the most common being systemic chemotherapy, especially platinum-based agents, in combination with radiotherapy and surgery for tumor resection.[4,5] Over the past few decades, rapid diagnosis and standard treatment progression have not yielded significant survival benefits; the 5-year survival rate for lung cancer in most countries is less than 30%.[6] Considerable improvements have been made in the targeted therapies against molecular alterations during malignancy. Clinical trials that target oncogenic mutations of anaplastic lymphoma kinase (ALK) and epidermal growth factor receptor (EGFR) via tyrosine kinase inhibitors (TKIs) have shown improved quality of life and response rate in patients with lung cancer.[7,8]

Immunotherapies alone or in combination with targeted therapies may be the ultimate curative option. The representative immunotherapy for lung cancer is monoclonal antibodies targeting immune checkpoints. Cytotoxic T-lymphocyte-associated antigen 4 (CTLA-4) and programmed cell death-1 (PD-1) are two of the most widely studied immune-checkpoint receptors for lung cancer therapy. However, a vast majority of patients are refractory to checkpoint blockade and its efficacy depends critically on the presence of sufficient tumor-specific lymphocytes.[9] Adoptive cell therapies (ACTs) are in clinical trials with promising results, but many hurdles still exist, especially for chimeric antigen receptor (CAR)-T cell therapy.[10] Other groups have focused on reviewing the molecular[11] and tumor heterogeneity[12], epidemiology,[13] biomarkers as diagnostic tools,[14] targeted treatments,[15] CAR-T cell therapy challenges,[16] and more recently first-line immunotherapies for lung cancer.[17–19] In this review, we provide a comprehensive overview of the current therapies with an emphasis on immunotherapies against lung cancer, specifically the most recent progress of adoptive cell therapies and challenges for future research and cell therapy applications.

Current Standards of Treatment for Lung Cancer

The first line of treatment for different lung cancer histotypes varies according to age, regions affected, extent of disease, tolerance to medication, and patient preference. Treatment options include surgery, radiation, chemotherapy, targeted therapy, immunotherapy, or a combination of treatments. For NSCLC, the first line of treatment is often surgery to remove the tumor. For larger tumors or those that have spread to nearby lymph nodes, a combination of surgery, chemotherapy, and radiation therapy may be recommended. As for SCLC, the first line of treatment is typically chemotherapy and radiation therapy. Surgery may be an option for early-stage SCLC, but it is not usually recommended for advanced cases.

Targeted therapy with TKIs is another current standard of lung cancer treatment. A well-established molecular target for lung cancer is EGFR, a receptor tyrosine kinase also known as ErbB1/human epidermal growth factor receptor 1 (HER1) included in the ErbB family of tyrosine kinase receptors which also comprise ErbB2/human epidermal growth factor receptor 2 (HER2)/Neu, ErbB3/human epidermal growth factor receptor 3 (HER3), and ErbB4/human epidermal growth factor receptor 4 (HER4).[20] EGFR activates many cellular signaling pathways that regulate growth, proliferation, and survival, and as the ERBB1 (EGFR) gene mutation was the first discovered mutation in NSCLC patients, it is the most common target of targeted therapy, which is characterized by 20% of patients with lung adenocarcinoma. First-generation TKIs, including gefitinib, icotinib, and erlotinib, which cause a reversible blockade of ATP-binding sites thereby stopping downstream signaling, have shown better responses and improved survival compared to cytotoxic therapy in previously untreated patients with EGFR mutations. Second-generation EGFR TKIs include afatinib and dacomitinib, which are irreversible inhibitors that also target HER2 and HER4 and provide an alternative for patients who acquired resistance to first-generation TKIs. Both afatinib and dacomitinib showed improved survival compared to gefitinib. The third-generation TKIs, such as osimertinib, rociletinib, olmutinib, and lazertinib, presented a new treatment option for patients with T790M EGFR-mutant tumors, which is the most common mechanism of resistance in patients who receive first- and second-generation TKIs. The ALK gene encodes a tyrosine kinase receptor that shares a high level of similarity with the insulin receptor but with a unique glycine-rich extracellular domain. Crizotinib is an oral competitive ATP inhibitor of ALK, MET, and ROS1 tyrosine kinases with activity against ALK fusion-positive NSCLC. Second-line and third-line treatment for NSCLC depends on the gene mutations found in the tumor resulting from the treatments that patients have already received.[21] More therapeutic options will be available for NSCLC patients with rare genetic aberrations and molecular alterations, mutations, and rearrangements including BRAF, KRAS, NTRK, and RET. New therapeutic targets in NSCLC are being evaluated in ongoing clinical trials and their efficacy has been recently reviewed.[22]

Immunotherapy for Lung Cancer

Among the strategies against lung cancer, the most promising are immunotherapies which include (1) non-specific immunotherapies such as cytokines, (2) monoclonal antibodies (mAbs) and immune checkpoint inhibitors, (3) oncolytic virus therapy, (4) cancer vaccines, and (5) adoptive cell therapy (e.g., T cells and natural killer [NK] cells), alone or in combination with other therapeutic agents [Figure 1].

Figure 1:

Immunotherapy types for lung cancer. ADCC: Antibody-dependent cell-mediated cytotoxicity; ADCP: Antibody-dependent cell mediated phagocytosis; CTLA-4: Cytotoxic T-lymphocyte-associated antigen 4; GzmB: Granzyme B; IL: Interleukin; MHC: Major histocompatibility complex; NF-κB: Nuclear factor-κB; NK: Natural killer; PD-1: Programmed cell death-1; PD-L1: Programmed cell death ligand-1; PRF: Platelet-rich fibrin; TCR: T cell receptor.

Non-specific immunotherapies

Toll-like receptor (TLR), a pattern recognition receptor (PRR) that recognizes pathogen-associated molecular patterns (PAMPs) to induce antigen-specific innate immunity, agonists have been investigated for their ability to enhance anti-tumor immune responses.[23] TLR9 is expressed by human B cells and plasmacytoid dendritic cells (pDCs). Synthetic unmethylated 5′–C–phosphate–G–3′ (CpG) oligodeoxynucleotides (CpG ODN) can activate TLR9 to reduce immune tolerance and promote anti-tumor response. Two Phase III trials evaluated TLR9 agonist PF-3512676 in combination with first-line paclitaxel/carboplatin[24] and gemcitabine/cisplatin[25], respectively, in advanced NSCLC; however, both studies were terminated due to lack of efficacy and increased toxicity. Other TLR9 agonists, such as IMO-2055[26] and MGN1703[27], are still in early development. The anti-tumor potential for TLR7 agonists has also been shown in pre-clinical studies via activation of pDCs, whereas TLR7 expression on lung cancer cells and its stimulation by TLR7 agonists promote tumor progression and resistance to chemotherapy.[28] Finally, TLR3 agonists can enhance tumor-specific T-cell responses and are currently being used as adjuvants for cancer vaccines in preclinical models.[29]

The use of cytokines is another emerging form of immunotherapy against lung cancer. Mammalian actin-binding protein-1 (MABP1) is a human monoclonal antibody that targets interleukin (IL)-1α. In a Phase I study, 16 patients with NSCLC who were refractory to the standard chemotherapy were given MABP1 in a dose-escalating manner.[30] The median overall survival (OS) was 7.6 months, and the median progression-free survival (PFS) was 57 days. Interestingly, there was a correlation between the patients with anti-EGFR pretreatment prior to MABP1 treatment and a better treatment outcome.[30] Median OS in the anti-EGFR pretreated patients was 9.4 months, versus non-pretreated patients with a median OS of 4.8 months. Median PFS was also higher in the anti-EGFR pretreated group than non-pretreated group (97 days and 78 days, respectively). The sample size was small, and the study was underpowered to be conclusive. Nevertheless, given the relative success in the anti-EGFR pretreatment group, the relationship of EGFR, IL-1α, and NSCLC should be an area of continued study, especially considering the encouraging results from MABP1 in the treatment of colorectal cancer.

The efficacy of IL-2 treatment in patients with NSCLC remains questionable. In a Phase III study, 239 patients with stage IIIb or IV NSCLC were randomly assigned to receive either gemcitabine + cisplatin (chemotherapy only), or gemcitabine + cisplatin + IL-2 (chemotherapy + IL-2). The median OS and median PFS were 10.5 and 6.6 months, respectively, in the chemotherapy + IL-2 arm and 12 and 6.9 months, respectively, in the chemotherapy only arm. The statistical differences of these two groups were nonsignificant in both OS and PFS.[31] Yet, in a separate Phase I/II trial, 45 patients with stage III NSCLC were studied. Thirteen patients received gemcitabine, cisplatin, and recombinant IL-2 (rIL-2), followed by surgery (chemotherapy + IL-2 + surgery). The remaining 32 patients received gemcitabine, cisplatin, and surgery (chemotherapy + surgery). The 5-year OS rate in the chemotherapy + IL-2 + surgery group was 59% compared to the group without rIL-2 therapy at 32%.[32] A striking 66% reduction in the hazard of death was observed in patients receiving rIL-2 immunotherapy. Despite this, no significant difference was found between the two groups in OS and event-free survival (EFS). Overall, further research is warranted to determine the efficacy of IL-2 immunotherapy treatment in patients with NSCLC.

In a first-in-Human Phase I study of subcutaneous outpatient recombinant human IL15 (rhIL15), E. coli-derived rhIL-15 was subcutaneously administered to patients with melanoma, renal cell carcinoma, squamous cell head and neck carcinoma, and NSCLC, in which the maximum tolerated dose of IL-15 administered subcutaneously was significantly higher than the dose that was feasible by intravenous bolus injection, and reached 3.0 μg/kg per day.[33] In another study, the number of circulating NK cells dramatically increased with IL-15 administration in a dose-dependent fashion. ALT-803, a superagonist complex of an IL-15 mutein bound to the sushi domain of IL-15Rα fused to the immunoglobulin G1 Fc, in combination with nivolumab (anti-PD-1 mAb) may repeatedly elicit objective responses to anti-PD-1 immunotherapy after relapse or treatment failure in patients with NSCLC.[34] Finally, the roles of IL-12, IL-21, as well as interferons in the treatment of lung cancer have not been fully explored.

Talactoferrin alfa, a recombinant human lactoferrin, is an orally active dendritic cell-mediated immunotherapy thought to interact with DCs in the gut wall leading to the production of an immunostimulatory cytokine cascade, thereby stimulating the migration and maturation of tumor antigen-presenting DCs.[35] Despite demonstrating anti-tumor activity in a variety of different pre-clinical models, these findings were not validated in an international randomized Phase III trial referred to as the FORTIS-M study in patients with advanced NSCLC, and therefore further testing came to a halt.[36]

Monoclonal antibodies

mAbs are proteins that are designed to target specific proteins on the surface of cancer cells. These mAbs can be used to either block the signals that cancer cells use to grow and divide or to mark the cancer cells for recognition and destruction by the immune system. A critical part of the immune system is to keep itself from attacking normal cells, and therefore the body uses “checkpoint” proteins on immune cells to turn on/off an immune response. Cancer cells can develop strategies to use these checkpoints to evade the immune system, which can then be targeted by drugs called immune checkpoint inhibitors (ICIs). ICIs have been largely used for NSCLC treatment.

Targeting ICIs, such as CTLA-4, PD-1, and PD-L1, has demonstrated promising clinical efficacy and durable responses in a broad spectrum of cancers, including lung cancer. PD-1 and PD-L1 targeted immunotherapy drugs that have been approved to treat NSCLC include atezolizumab (Tecentriq) and durvalumab (Imfinzi) which target PD-L1, and nivolumab (Opdivo), pembrolizumab (Keytruda), and cemiplimab (Libtayo) which target PD-1.[37] CTLA-4 (or CD152) is expressed on T lymphocytes, and competes with CD28, which is a co-stimulatory receptor on T lymphocytes, by binding with B7 (CD80 or CD86) on antigen-presenting cells (APCs) with higher affinity and avidity. An inhibitory signal is generated when CTLA-4 binds with B7 on activated T cells. Drugs that block the CTLA-4 pathway include ipilimumab (Yervoy), a human IgG1 mAb, which is given in combination with nivolumab (Opdivo), and sometimes in combination with chemotherapy. Tremelimumab is another human IgG2 mAb directed at CTLA-4. Durvalumab is also frequently combined with tremelimumab for dual checkpoint inhibitor studies in NSCLC.

The combination of immune checkpoint blockade and chemotherapy produces a synergized effect to confer better survival outcomes. Chemotherapy is the first choice for patients who lack targetable driver mutations. Pembrolizumab targeting PD-1 was combined with chemotherapy in several clinical trials (KEYNOTE-021,[38] KEYNOTE-189,[39] and KEYNOTE 407[40]). In 2018, the combination of pembrolizumab with pemetrexed and carboplatin was approved as a first-line treatment for metastatic NSCLC patients with no driver mutation, irrespective of PD-L1 expression based on the results shown by KEYNOTE-021. A subsequent Phase III trial concluded that the addition of pembrolizumab to chemotherapy resulted in a significantly longer OS and PFS than chemotherapy alone (KEYNOTE-189).[39] Later, an expanded approval was obtained for the combination of pembrolizumab with carboplatin and paclitaxel/nab-paclitaxel (Abraxane) for metastatic squamous NSCLC, irrespective of PD-L1 expression (Keynote-407). In 2018, the efficacy of nivolumab in combination with ipilimumab, an ICI targeting CTLA-4, was first demonstrated in a Phase I trial (CheckMate 012) as a first-line treatment of advanced NSCLC with a tolerable safety profile and a high and durable response.[41] Durvalumab was also combined with osimertinib (a third-generation EGFR-TKI) in a Phase III trial (CAURAL); however, the clinical trial was terminated early because of an increased incidence of interstitial lung disease-like events. In the CheckMate 012 trial, nivolumab combined with erlotinib was tolerable in patients with EGFR-mutated advanced NSCLC.[42]

Antibody-drug conjugates (ADCs) are a novel class of biopharmaceutical drugs in which a monoclonal antibody is linked to a cytotoxic drug. Traztuzumab deruxtecan (T-DXd), which consists of trastuzumab and the payload deruxtecan, a topoisomerase I inhibitor, is the first ADC approved for patients with HER2 mutation-positive NSCLC.[43] Two other ADCs (patritumab deruxtecan,[44] and telisotuzumab vedotin[45]) have been granted U.S. Food and Drug Administration (FDA) Breakthrough Therapy Designation and are currently under evaluation. Moreover, novel ADCs targeting HER2,[46] HER3,[47] Trop-2,[48,49] and c-Met,[50] either as monotherapy or in combination strategies are being clinically evaluated in lung cancer.

Considering many patients experience lung tumor recurrence within five years following surgery, studies have focused on perioperative regimens that combine immunotherapy with chemotherapy. The AEGEAN clinical trial assessed the activity of durvalumab, a human IgG1 mAb targeting PD-L1, alone or in combination with platinum-based chemotherapy prior to surgery. Findings revealed perioperative immunotherapy in resectable NSCLC demonstrated improved pathologic complete response and event-free survival in patients without prior treatment, and perioperative durvalumab plus neoadjuvant chemotherapy was associated with significantly greater event-free survival and pathological complete response than neoadjuvant chemotherapy alone.[51] The same study was repeated but not as effective when pembrolizumab was used.[52] Further studies are required to find appropriate target patients that may benefit from mAb combinational therapies.

Oncolytic virus therapy

There are currently 14 clinical trials of lung cancer-associated oncolytic viruses.[53] Oncolytic viruses are designed to preferentially infect and replicate in tumors, which not only promotes cell death but also enhances immune mediators entering the tumor microenvironment and induces systemic anti-tumor immunity. Phase 1 study of intravenous administration of the chimeric adenovirus enadenotucirev in patients undergoing primary tumor resection (previously known as ColoAd1), a tumor-selective chimeric adenovirus, stimulated local high anti-tumor immune response and the infiltration of CD8+ T cells in NSCLC.[54] Furthermore, using oncolytic adenovirus or vaccinia virus-infected reprogrammed somatic-derived tumor cell vaccine (VIReST) regimen to vaccinate high-risk populations can prevent tumor progression and initiate long-term anti-tumor immune response, which can be used in the treatment and prevention of lung cancer. Myxoma virus (MYXV) is a distinct Leporipoxvirus initially identified as the causative agent for myxomatosis, a lethal disease specific to European rabbit strains (Oryctolagus cuniculus) but nonpathogenic for other mammals. A rabbit-specific poxvirus, MYXV, is able to infect and replicate in tumor cells derived from diverse species, and oncolytic activity has been demonstrated for several human tumor lineages. Kellish et al[55] selected human SCLC as a solid tumor model for preclinical testing for MYXV therapy. “Oncolytic therapeutic vaccine” is a combination strategy utilizing oncolytic virus encoding one or more tumor-associated antigens or neoantigens.

Cancer vaccines

The normal lung environment is constantly exposed to foreign antigens, including inanimate dust, viruses, bacteria, and fungi. Imbalances in immune activation and immune suppression can lead to autoimmune diseases such as asthma or interstitial lung disease. Generating vaccines to target lung cancer requires fine-tuning immune activation. There are protein and peptide vaccines (MAGE-A3, L-BLP25) as well as cell-based vaccines (GVAX, Belagenpumatucel-L) targeting lung cancer.

NSCLC patients whose tumors express the tumor-associated antigen (TAA) melanoma associated antigen-A3 (MAGE-A3) are often associated with poorer prognosis. The MAGE-A3 vaccine is a recombinant MAGE-A3 protein combined with the immunostimulant AS02B that was assessed in a large, double-blind, randomized Phase III trial (MAGRIT) in patients with stage IB, II, or IIIA MAGE-A3-positive NSCLC. Unfortunately, this study found no overt increase in disease-free survival in the vaccine group compared with placebo in either the overall population or in the patients who did not receive chemotherapy, and further development of the MAGE-A3 immunotherapy for NSCLC patient use has been stopped.

TG4010 is a vaccine targeted against Mucin1 (MUC1) antigen composed of a recombinant vaccinia virus (modified virus of Ankara or MVA) that encodes human MUC1 and IL-2 (MVA-MUC1-IL-2).[56] Belagenpumatucel-L is an allogeneic tumor cell vaccine containing four irradiated NSCLC cell lines and an antisense plasmid of tranforming growth factor-β (TGF-β).[57] The vaccine aims to increase the immune response to NSCLC through downregulation of TGF-β expression. Like many other solid tumors, lung cancers utilize a variety of mechanisms to evade the host immune response, including downregulation of human leukocyte antigen (HLA)/major histocompatibility complex (MHC) molecules, loss or modulation of tumor antigen expression or secretion of immunosuppressive cytokines. These can at least partially explain why vaccines have not worked in lung cancer. Sequencing the “tumor mutanome” allows for in silico prediction of the immunogenicity of mutated peptides, which can then be used for tracking tumor-specific T-cell responses to develop a personalized vaccine.[58] The peptide neoantigen vaccine may not be enough to induce an effective tumor-specific immune response, and a combination therapy approach may be needed. For example, the combination of checkpoint blockade and vaccine is a logical next step in cancer immunotherapy. Despite the advances with checkpoint blockade, not all patients respond, and the addition of vaccine therapy may further enhance T-cell proliferation.

Adoptive Cellular Therapy

There are currently three ACT techniques: tumor-infiltrating lymphocytes (TILs), T cell receptor-engineered T (TCR-T) cells, and chimeric antigen receptor T (CAR-T) cells [Figure 2]. TIL therapy involves the expansion of a heterogeneous population of endogenous T cells found in a harvested tumor, while CAR-T cells and TCR-Ts involve the expansion of a genetically engineered T-cell directed toward specific antigen targets.[59] While successful application of ACT has been seen in hematologic malignancies, ACT in solid tumors, especially in lung cancer, is still in its early stages.

Figure 2:

ACT for lung cancer. ACT: Adoptive cell therapy; MHC: Major histocompatibility complex; TAA: Tumor-associated antigen.

Tumor-infiltrating lymphocyte (TIL) therapy

TILs are a heterogeneous population of cells that are polyclonal and have the capability to recognize various tumor antigens. The utility of autologous TIL therapy as an effective anti-cancer treatment was first reported in metastatic melanoma patients by Rosenberg and colleagues in 1988.[60] The most encouraging clinical trials to date were conducted in patients with malignant melanoma, a tumor that shows a good response to immune therapy.[61–65] For example, patients with metastatic melanoma who received autologous TILs can yield durable, complete responses.[62] The overall response (OR) rates and complete response (CR) rates for these patients were 40–50% and 20%, respectively.[62] Specifically, patients who achieve CR have demonstrated long-term disease-free survival.[62] In addition, for patients with advanced metastatic melanoma who progressed on multiple prior therapies, including anti-PD-1, autologous TIL therapy also yielded a 38% response rate.[66] There is growing evidence showing that TIL therapy has led to durable remissions in subsets of patients with cervical cancer,[67] colorectal cancer,[68] and breast cancer.[69]

The success of TIL therapy is driven by neoantigen-directed T cells and the number of neoantigens that are targeted. The tumor mutational load and predicted neoantigen load have been shown to correlate with clinical outcomes for TIL therapies as well as PD-1 inhibitors.[70] The fact that metastatic lung cancer is responsive to PD-1/PD-L1 immunotherapy and possesses a high mutational burden makes it an ideal candidate for ACT with TILs. However, there have been limited published studies of TIL therapies in the treatment of NSCLC. The first randomized clinical trial assessing the efficacy of TIL therapy for NCSLC was reported in 1994.[71] Patients who received TILs had significantly longer survival than those who underwent standard treatment (median survival 22.4 months vs. 14.1 months).[71] This study was the initial proof that TIL therapy can be applied successfully to patients with late-stage lung cancer. In a recent Phase I single-arm clinical trial, Creelan et al[72] tested the safety and efficacy of TILs in anti-PD-1 resistant metastatic lung cancer patients. Twenty patients with metastatic NSCLC were enrolled and underwent harvesting of TILs before the initiation of ICB treatment; 16 patients subsequently received a TIL infusion and a high dose of IL-2, followed by maintenance nivolumab.[72] The patients received a lymphocyte-depleting chemotherapy regimen before the TIL infusion. Of 13 evaluable patients, 3 had confirmed responses, and 11 had a reduction in tumor burden, with a median best change of 35%. Two patients achieved complete responses ongoing 1.5 years later.[72] Although this study enrolled a small group of patients, the work highlights that TIL therapy can be clinically beneficial in patients with ICB-resistant NSCLC, inducing durable complete responses in some patients. More recently, a prospective, open-label, multi-cohort, non-randomized, multicenter Phase II study was started to evaluate autologous TIL therapy in patients with solid tumors (NCT03645928). Cohort 3B investigated TIL monotherapy in patients with advanced or metastatic NSCLC, who had been treated with 1–3 prior lines of systemic therapy, including either ICI or oncogene-directed therapy. Of 28 patients who received TIL therapy, the objective response rate (ORR) was 21.4% (6/28).[73] Among them, one patient had a complete metabolic response (defined as the visual absence of pathological fluorodeoxyglucose uptake in all baseline lesions identified on the baseline positron emission tomography [PET] scan), and 2 responses occurred in patients who were PD-L1 negative.[73] All responders received ≥2 prior lines of systemic therapy.[73] This study demonstrated that TILs could be a treatment option in NSCLC patients who had previously received ICIs. Other ongoing clinical trials of TILs in NSCLC are summarized in Table 1.

Table 1 - Selected clinical trials of TIL therapy in lung cancer. NCT No. Title Phase Types of cancer Status NCT03215810 Nivolumab and tumor infiltrating lymphocytes (TIL) in advanced non-small cell lung cancer I Stage IV or recurrent NSCLC Completed NCT05681780 Clinical trial of CD40L-augmented TIL for patients with EGFR, ALK, ROS1 or HER2-driven NSCLC I/II Stage IV or recurrent NSCLC Recruiting NCT05878028 L-TIL plus tislelizumab for PD1 antibody resistant aNSCLC II Stage IV NSCLC Recruiting NCT04614103 Autologous LN-145 in patients with metastatic non-small-cell lung cancer II Metastatic stage IV NSCLC Recruiting NCT03419559 Study of autologous tumor infiltrating lymphocytes (LN-145) in combination with durvalumab in non-small cell lung cancer II Stage III or stage IV NSCLC Withdrawn NCT03645928 Study of autologous tumor infiltrating lymphocytes in patients with solid tumors II Unresectable or metastatic melanoma, advanced, recurrent or metastatic HNSCC, stage III or stage IV NSCLC Recruiting NCT05566223 CISH inactivated TILs in the treatment of NSCLC (CheckCell-2) I/II PD-L1 negative or positive metastatic NSCLC Not yet recruiting NCT05573035 A study to investigate LYL845 in adults with solid tumors I Melanoma, NSCLC, CRC Recruiting NCT05576077 A study of TBio-4101 (TIL) and pembrolizumab in patients with advanced solid tumors (STARLING) I Advanced or metastatic breast carcinoma, colorectal adenocarcinoma, uveal melanoma, cutaneous melanoma, NSCLC, HNSCC Recruiting NCT02133196 T cell receptor immunotherapy for patients with metastatic non-small cell lung cancer II stage IV or unresectable NSCLC Recruiting NCT03903887 A study of anti-PD1 antibody-activated TILs in non-small cell lung cancer I/II Stages II–IIIA NSCLC Unknown status NCT05361174 A Study to investigate the efficacy and safety of an infusion of IOV-4001 in adult participants with unresectable or metastatic melanoma or stage III or IV non-small-cell lung cancer I/II Unresectable or metastatic melanoma or stage III or IV NSCLC Recruiting NCT01820754 Evaluation of circulating T cells and tumor infiltrating lymphocytes (TILs) during/after pre-surgery chemotherapy in NSCLC II Stages 1B, 2, or 3 NSCLC Completed

ALK: Anaplastic lymphoma kinase; aNSCLC: Advanced non-small cell lung cancer (NSCLC); CISH: Cytokine inducible SH2 containing protein; CRC: Colorectal cancer; EGFR: Epidermal growth factor receptor; HER2: Human epidermal growth factor receptor 2; HNSCC: Head and neck squamous cell carcinoma; L-TIL: Liquid tumor infiltrating lymphocytes; PD1: Programmed cell death 1; ROS1: ROS proto-oncogene 1, receptor tyrosine kinase; TIL: Tumor infiltrating lymphocyte.

TIL immunotherapy involves several laboratory and clinical steps, which start with surgical resection of tumor tissue from the patients and continue with tumor processing to establish T cell cultures, which are then expanded in IL-2-containing media. In 2017, Ben-Avi et al[74] optimized the expansion method of lung cancer-derived TILs under a good manufacturing practice (GMP) environment. After isolation from the tumor tissue, the TILs were expanded with anti-CD3 antibody, IL-2, and irradiated peripheral blood mononuclear cells from non-related donor cells in G-Rex flasks. This method can yield up to 5 × 1010 functional TILs for infusion starting from 1 × 106 live nucleated cells isolated from a single lung tumor piece. This study indicates that TILs from lung cancer patients can be expanded to treatment levels under GMP conditions.

TIL may hold some distinguishing advantages for treating solid tumors. Firstly, TILs have diverse TCR clonality, are capable of recognizing an array of tumor antigens, and therefore may be superior in tackling tumor heterogeneity compared to other ACT approaches, such as CAR-T and TCR-T cell therapy. In line with this, TIL has demonstrated better clinical efficacy than CAR-T in solid tumors containing high mutation load, such as melanoma.[75] In addition, off-target toxicity has seldom been reported in TIL therapy, probably due to the negative selection of TCRs of TIL during the early development of T cell immunity. On the contrary, the engineered tumor-targeting single-chain variable fragments (scFv) in CAR-T or affinity-enhanced TCR in TCR-T products may lead to toxicity if they bear cross-reactivity with antigens on normal tissues. However, TIL therapy also has some disadvantages. Currently, the most widely used TIL production method is to isolate infiltrating lymphocytes from tumor tissues and then culture and expand these cells in vitro. The administration of high-dose IL-2 used as a standard of care to support the growth and activity of infused TIL, however, may restrain the clinical application of TIL therapy. High-dose IL-2 could induce systemic toxicity,[76] and could also promote regulatory T cells that suppress the anti-tumor response of TIL.[77] Furthermore, the short-term persistence of autologous mature TILs in vivo is also a challenge to achieve the maximal outcome of TIL therapy.