2021 marks the centenary of the bacillus Calmette-Guérin (BCG) vaccine. In our review ‘100 Years of BCG Immunotherapy: From Cattle to COVID-19’, we highlight key milestones in the history of BCG, beginning from its discovery by Alfred Calmette and Camille Guérin to its present day use in tuberculosis (TB) prevention and as intravesical therapy for non-muscle-invasive bladder cancer [1]. Introduced into clinical practice by Alvaro Morales in 1976 [2], and championed by Donald Lamm leading to FDA approval [3], intravesical BCG has become the gold standard treatment for intermediate- and high-risk disease due to its ability to reduce disease recurrence and progression. Yet despite high initial efficacy, we know that BCG does not work for all patients [4,5]. So, what does the future hold for intravesical BCG immunotherapy?

Current research efforts are focused on developing ways to enhance the clinical efficacy of BCG, and it is anticipated that these efforts will reach maturity within the next decade. One strategy to augment treatment response that is being revisited from the 1980s is priming. The concept of priming, while not new, received renewed attention from pre-clinical studies showing that the administration of subcutaneous BCG prior to intravesical BCG instillations accelerates entry of T-cells into the bladder and improves response to BCG [6]. A randomized phase III trial (SWOG Trial S1602; NCT03091660) is underway to determine whether priming with intradermal BCG vaccine prior to standard intravesical BCG might enhance treatment efficacy in patients with high-risk NMIBC. Additionally, this trial will also evaluate the Tokyo strain of BCG against the currently available Tice strain. Evidence from animal studies suggests that certain BCG strains may be more immunogenic than others but, to date, no adequately powered head-to-head trials have been performed to investigate the impact of BCG strain on oncological outcomes [7]. Should the Tokyo strain prove non-inferior, this may ultimately facilitate entry of the strain into the United States market which is currently facing a BCG shortage.

Recombinant BCG (rBCG) techniques may also offer an effective approach to enhance the clinical efficacy of BCG. Most of these rBCGs have been developed as vaccine candidates for improved protection against TB and, in our review, we highlight the promising role of VPM1002 which has been shown to be more immunogenic than conventional BCG [8]. However, these recombinant agents may also be used in the treatment of NMIBC. VPM1002BC is a live recombinant BCG that has been genetically modified to express the listeria toxin listeriolysin. This pore-forming toxin enhances the induction of CD4+ and CD8+ T-cell mediated immune responses [9]. The intravesical application of VP1002BC has already demonstrated safety and tolerability in a phase I trial of patients with BCG failure [10] and the results of a phase II trial are currently pending (NCT023714470). In the muscle-invasive disease setting, a phase II trial is underway to investigate the combination of intravesical VPM1002BC with atezolizumab, cisplatin and gemcitabine to increase the rate of pathologic complete remission at the time of radical cystectomy (NCT04630730). Other rBCGs are still in the pre-clinical phase: rBCG-S1PT, which expresses the detoxified S1 pertussis toxin, has demonstrated enhanced immune activation compared to wild type BC as evidenced by increased cytokine production and tumor cytotoxicity in vitro [11]. Additionally, Cho et al. developed two rBCG strains with resistance to anti-microbial peptides [12]. Bladder cancer cells treated in vitro with these rBCGs demonstrated higher internalization and increased release of anti-tumor cytokines.

Yet another strategy to bolster the anti-tumor activity of BCG may be through combination with checkpoint inhibitors. Upregulation of the PD-1 and PD-L1 signaling pathways has been implicated in BCG treatment failure [13,14]. In an immunohistochemical study of NMIBC samples collected before BCG treatment, PD-L1 expression was detected in 25-28% of non-responders versus 0-4% of responders (p<0.01), suggesting that the combination of BCG with checkpoint inhibitors could yield a durable response in a proportion of patients with BCG resistance [14]. In contrast, a recent RNA-based profiling study of NMIBC specimens was unable to demonstrate any association between high gene expression of PD-L1 and PD-1 and worse BCG responses in three independent cohorts [15]. In any case, a number of phase III trials investigating the synergistic effect of BCG with atezolizumab (ALBAN; NCT03799835), durvalumab (POTOMAC; NCT03528694), pembrolizumab (KEYNOTE-676; NCT03711032) and sasanlimab (CREST; NCT04165317) in BCG-naïve patients are underway and will soon answer the question of whether combination therapy is able to improve outcomes. Additional trials of combination therapy are also ongoing in the setting of BCG-unresponsive disease and include the combination of BCG with nivolumab (CheckMate 7G8; NCT04149574, CheckMate 9UT; NCT03519256) and pembrolizumab (NCT03711032). Other trials of combination therapy in this domain include BCG with gemcitabine (NCT04179162) and the IL-15 superagonist ALT-803 (QUILT 3.032; NCT03022825).

Finally, given that the response to BCG is unpredictable and that we are currently in a period of worldwide BCG shortage, the identification of predictive biomarkers is extremely important. Due to the complexity of the immune pathways induced by BCG, it is unlikely that any single biomarker will be able to reliably predict response in individual patients. Indeed, the search for clinically useful predictive biomarkers is still in its infancy; most candidate biomarkers thus far reported from single institutional series lack subsequent validation [16,17] or possess characteristics rendering them unsuitable for individual patient management [18]. Even so, in time, it is anticipated that as we continue to gain clarity regarding the mechanisms underlying BCG-mediated tumor immunity, the identification of predictive biomarkers will shortly follow.

In conclusion, the next decade of BCG immunotherapy brings much to look forward to. Emerging insights into the mechanisms of action provide new opportunities to improve the anti-tumor activity of BCG be it through priming, recombinant techniques or combination therapies, in addition to developing biomarkers to accurately predict response to BCG. We anticipate that these research endeavors will come to fruition in the next century, thus improving outcomes for patients with non-muscle invasive bladder cancer.

This work was supported by the Wayne B. Duddlesten Professorship in Cancer Research and the Raymond and Maria Floyd Bladder Cancer Research Foundation to Ashish M. Kamat and the Urology Foundation-Fulbright Scholar Award to Niyati Lobo.