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The past number of years have seen much progress for immune-based approaches in solid tumours, with FDA approvals for various strategies, including dendritic cell vaccination as well as the much-vaunted and so-called ‘immune checkpoint inhibitors’. These advances have engendered great optimism (and no small amount of hype), largely on account of the higher ceiling for immunotherapy compared with traditional treatment modalities, in particular their potential for long-term control of metastatic disease.
The principle of immunotherapy — manipulating by whatever means a patient’s immune system to treat cancer, as opposed to directly assaulting the dividing malignancy with cytotoxic drugs or signal transduction inhibitors — is well and truly proven at this stage, but we must remember that outside of some banner indications (MSI-hi cancers, melanoma), the hard reality right now is that the majority of patients will not derive benefit from an immune-based approach. The next frontier, therefore, is for immunotherapy to prove itself in patients who have either developed resistance or have diseases which as a group are not particularly immunogenic. So far, with notable exceptions, this has included cancers of the gastrointestinal tract.
Immunotherapy has a long and colourful history, stretching back to Coley’s toxins in the late 1800s, when William Coley, a surgeon in New York, began administering — with mixed results — direct intratumoural injections of bacteria with the aim of producing an inflammatory reaction against the cancer. Much of the subsequent century was spent elucidating a basic understanding of the immune system itself and it is perhaps surprising to see how recently this began to come together. (The T-cell receptor was not identified until the early 1980s; it was 1992 before the role of CD28 co-stimulation was clarified). The current field has been built on the back of these and many other basic science discoveries, mapped out in increments by scientists working in largely neglected fields, leading to our current understanding of what is perhaps the central relationship in tumour immunology: How a T-cell interacts with an antigen that is presented to it in the context of major histocompatibility complex (MHC) proteins by antigen-presenting cells (APCs).
In the meantime, cancer research proceeded apace, largely ignoring any role for the immune system, with most researchers regarding cancer as solely a disease of genetic aberrancy. The immune system was simply a barrier to giving higher doses of chemotherapy. The power of the immune system to effect remissions in metastatic cancer was first demonstrated for cytokines in renal malignancies and adoptive cellular therapies in melanoma, but these treatments had limited export potential. It was James Allison’s insight that by focusing on endogenously-activated T-cell responses in cancer (rather than the cancer itself) — blocking their inhibitory mechanism, thereby amplifying these responses by drugs with off-the-shelf potential — which led to the paradigm shift we are currently experiencing.
Immune checkpoint inhibitors in GI cancers: Current status
The most high-profile immunotherapy currently in use, and the primary driver of the current enthusiasm for immune approaches, is the family of drugs known as ‘checkpoint inhibitors’. This is a broad and semi-accurate term, which generally refers to drugs targeting inducibly expressed proteins on the surface of a T-cell following activation.
The first to enter the clinic was anti-CTLA4 therapy, inhibiting a protein (cytotoxic T-lymphocyte-associated protein 4), which appears on the surface of a T-cell approximately 24 hours after it is activated and whose purpose is to out-compete the co-stimulation protein CD28 — with which it is homologous — thereby attenuating the immune response and protecting the host from excessive T-cell autoimmune damage. Blocking this inhibitory pathway by a monoclonal antibody, such as ipilimumab, enhances the T-cell response, inhibiting the inhibition or — as is often explained colloquially to patients — ‘releasing the brakes’. (CTLA-4 is also expressed constitutively on T-regulatory cells and the relative contribution of this to therapeutic efficacy in humans is somewhat controversial). The other major ‘checkpoint’ in current use is programmed death (PD)-1, which is likewise a marker of T-cell activation, although high levels of PD-1 — as can occur in chronic viral infections — leads to dysfunctionality. (PD-1hi T-cells are not very proliferative or cytotoxic and secrete less interferon).
Inhibiting PD-1 (by monoclonal antibodies such as pembrolizumab or nivolumab) can reverse this and reinvigorate the T-cell.
The ligand for PD-1 (mainly PD-L1) is expressed by tumour cells, protecting them against T-cell attack, as well as on various other immune cells, which contribute to the immunosuppressed microenvironment in which tumours thrive. Therefore, inhibiting the PD-1/PD-L1 axis has several effects that are likely non-redundant and not fully understood.
Perhaps the most important point to be made about immune checkpoint inhibitors is that their efficacy is dependent upon the presence of an already existing endogenous anti-tumour immune response. If the initial engagement of the T-cell receptor by tumour-derived antigen as presented by the APC does not occur, the immune checkpoint inhibitor will simply not work, not because it is a ‘bad drug’, but because there is no ongoing process of T-cell activation for it to act upon and amplify. What we have seen thus far in the clinic, therefore, has been the relatively ‘low-lying fruit’ of efficacy for monotherapy (ie, single-agent checkpoint inhibition) demonstrated in those tumours that are ‘immunogenic’, meaning that the immune system has already recognised the tumour and is mounting an immune response against it. The checkpoint inhibitor amplifies this anti-tumour immune response by relieving the countermanding inhibition (CTLA4 or PD1 upregulation on the surface of the T-cell or ligation of PD1 by PD-L1 expressed on the tumour cell membrane).
Those tumours in which an immune response is ongoing, and which respond or are likely to respond to checkpoint inhibition, have been categorised descriptively as ‘hot’ tumours. The difficulty we face is that the majority of tumours are either ‘cold’ — ie, the tumour is immunologically inert — or have evoked either a weak immune response or one that is smothered by the heavily immune-suppressive microenvironment.
Immune checkpoint inhibitors — clinical experience to date
The clinical experience of immunotherapy in GI cancers has been generally disappointing. One of the first studies evaluating and showing impressive results for PD1/PDL1-directed therapy was disappointing from a GI cancer viewpoint. There were no responses in any of the cohorts containing patients with colorectal (N=18) and pancreatic (N=14) cancer, and this trend has largely continued in subsequent studies for these diseases. Recent evidence has highlighted the link between the immunogenicity of a tumour and its mutagenic burden, ie, the number of non-synonymous somatic mutations within it, and it is unlikely to be coincidental that the tumours that respond best to checkpoint inhibition (melanoma, non-small lung cancer) are the ones most associated with known mutagens (UV light, cigarette smoke). GI cancers are not highly mutated, which perhaps explains the poor general efficacy of checkpoint inhibition.
Isolated pockets of activity, however, have been seen in GI oncology, suggesting that a proportion are immunogenic, and providing a foothold for further development. The highlights have been as follows:
MSI-hi disease: Cancers which have defects in their mismatch-repair apparatus (either by inherited mutations or somatically via epigenetic silencing) bear as a surrogate marker increasing length of microsatellites leading to instability (MSI). These tumours acquire high numbers of mutations, increasing the chances of generating a neoantigen recognisable by T-cells. It was long appreciated that these tumours were heavily infiltrated by lymphocytes, triggering an adaptive response and PD-L1 upregulation as a result of interferon-gamma secretion. Approximately 5-to-20 per cent (depending on stage) of colorectal cancers and as many as 20-to-30 per cent of gastric or duodenal cancers may be MSI-hi. PD-1 blockade has dramatically changed the outlook for these tumours, to the extent that MSI-hi status should be actively sought after, arguably in all patients.
Hepatocellular carcinoma (HCC): Data from Check- Mate 040 was recently published in The Lancet evaluating nivolumab in patients with different aetiologies of HCC. The results were encouraging, with an overall response rate of about 20 per cent and a median duration of response of 9.9 months and median survival not reached. Interestingly, the efficacy was similar in viral hepatitis-associated versus non-associated tumours. The advantages of an immune approach in HCC are particularly attractive due to the frequent underlying hepatic dysfunction in this disease, which renders conventional drug development difficult.
Gastro-oesophageal: According to a landmark TCGA (The Cancer Genome Atlas) analysis, approximately one-third of gastric cancer cases may be immunogenic, based on gene expression in immune-signalling pathways. The largest study to date evaluating checkpoint inhibition in gastric cancer was recently presented. In this phase 3 trial, patients with gastric cancer (N=493) were randomised to nivolumab or placebo. Whilst the study was statistically positive for median overall survival — a difficult end-point in what was a heavily-pretreated population in whom monotherapy was likely to be effective for only a minority — the real story here was the tail on the curve, with a 12-month survival rate of 26 per cent and a median duration of response of 9.5 months. Recent data also shows a role for PD-1 therapy in squamous cancers of the oesophagus.
Anal cancer: Though curable in the localised setting, metastatic anal cancer is generally poorly-responsive to chemotherapy. Encouraging data was initially published for pembrolizumab and subsequently — in the phase 2 setting — for nivolumab, in which nine of 37 patients (24 per cent [95 per cent, CI 15-33]) had an objective response, with a median depth of response of 70 per cent (IQR, 57-90). Certain tumour characteristics were associated with a higher odds of response (PD-L1 expression, presence of intratumoural lymphocytes). These results suggest that across GI cancers as a group, there are subsets of patients who seem to benefit from single-agent checkpoint inhibition. For the field as a whole these preliminary results are really just a starting point and clearly need to be built upon.
Translational efforts: Making ‘cold’ tumours ‘hot’
The central question facing us today is how to make immunologically-inert tumours amenable to immunotherapy. In other words, how to cause a tumour to become inflamed, how to turn a ‘cold’ tumour into a ‘hot’ one. This can be accomplished — at least pre-clinically — in a dizzying variety of ways. Some of these experimental efforts are highly preliminary, awaiting translational proof of principle, whilst others are employing modalities such as radiation or ablative methods, for example, which have been around for a long time but are being repurposed with an immune intent in mind. It is impossible to comprehensibly list all of these efforts and what follows is a selective (and highly subjective) summary of these efforts.
Experimental immune strategies
Agonistic co-stimulatory antibodies: Rather than ‘inhibiting the inhibition’, T-cell agonism can directly stimulate immune cells and several of these proteins (OX40, CD40, CD137) are the target of much translational research. OX40, for example, is a member of the tumour necrosis factor (TNF) superfamily that is expressed transiently on CD4 and CD8 cells during antigen-specific priming, inducing their expansion and differentiation and promoting long-lived memory T-cells. There is a strong rationale for combining T-cell agonists with immune checkpoint inhibitors, perhaps in combination with vaccination.
This approach for OX40 agonism recently proved capable — preclinically in a poorly-immunogenic mouse model — of reversing CD8 T-cell anergy to a tumour-associated antigen and promoting robust expansion and function of these cells.
Vaccines: Many different vaccine approaches — peptide vaccines, allogenic or even autologous tumour-derived vaccines —have been tested down the years with very limited success, due to, for whatever reason, an inability to induce potent anti-tumour immunity that was clinically relevant, rather than serologically so. Checkpoint inhibitors can be a natural ally for amplifying vaccine-induced T-cell responses. But in addition to this, technological advances have also led to the development of better vaccines. For example, it is now possible to create personalised vaccines that induce immunity to tumour-associated neoantigens, and several companies are attempting to bring this to market. Microbial organisms can also be availed of, making use of their visibility to the innate immune system, thereby acting as a ‘Trojan horse’ to be taken up by dendritic cells and containing a genome that can be modified to enhance immune function (and limit pathogenicity of the infectious agent). An example of the latter are drugs such as CRS-207, which use the bacterium Listeria monocytogenes as a platform for activation of immunity directed against mesothelin-expressing cancer cells, such as pancreatic cancer. Viruses can also be used as potential vaccines, an example of which is the adenovirus serotype five currently in development for colorectal cancer.
Oncolytic viruses: Viruses can also be used as properly infective agents with the aim of spreading to and lysing cancer cells directly. In a throwback to Coley’s toxins, the intratumoural injection of genetically-modified viruses has been shown to invoke potent immune responses, and herein lies their relevance for immunotherapy. These so-called oncolytic viruses are native or engineered viruses that preferentially replicate in cancer cells. Selective tumour cell replication is thought to depend on infection of neoplastic cells, which harbour low levels of protein kinase R (PKR) and dysfunctional type I IFN-signalling elements. The FDA approved the first oncolytic virus for cancer [Talimogene laherparepvec (T-VEC)], for melanoma on October 15, 2015. The concept of combining oncolytic viruses with checkpoint inhibition has two enormous theoretical advantages: Tumour selectivity, and the ability to trigger adaptive immunity. Direct tumour cell lysis causes the release of soluble tumour antigens that can prime and promote tumour-specific immunity, which of course can be amplified by combination with either CTLA-4 and PD-1 blockade. In the first published experience of this approach, a phase 1b trial of T-VEC plus ipilimumab, 18 patients with unresectable melanoma were treated, with an objective response seen in 56 per cent of patients (33 per cent had a complete response). Most of the oncolytic virus clinical trials to date have involved intratumoural injection. Pexa-Vec (JX-594) — a vaccinia virus engineered for the expression of transgenes encoding human granulocyte-macrophage colony-stimulating factor (GM-CSF) — can be administered systemically by intravenous administration. In a study published in Nature, patients with HCC had a dose-dependent, almost doubling in median survival, and evidence of an abscopal effect — reductions in tumours distant from the area injected.
STING pathway agonists: The stimulator of IFN genes (STING) pathway senses cytosolic tumour-derived DNA within dendritic cells that have managed to infiltrate tumours. Following activation, the STING pathway triggers an influx of anti-tumour T-cells. Pharmacologic agonists of the STING pathway have demonstrated proof of principle in multiple poorly-immunogenic syngeneic murine models and are currently in early-phase clinical testing. Combination with checkpoint inhibition is the obvious next step.
Cellular therapies (TIL, CAR, engineered TCR): The potential of cellular approaches in solid tumours was recently illustrated in spectacular fashion by Rosenberg and colleagues in two high-profile case reports. Both cases utilised the recent invention of tandem minigene (TMG) constructs to characterise the immunogenicity of a patient’s individual mutations (and corresponding neoantigens) in order to identify the relevant reactive population of T-cells. Via adoptive transfer of enriched populations of these cells, the investigators were able to produce durable tumour regression in metastatic chemo-refractory cholangiocarcinoma and KRAS-mutated colorectal cancer. Other possible cell-based approaches — in the case of a pre-identified antigen — are T-cells with genetically engineered receptors (restricted therefore by HLA context) or chimeric antigen receptor (CAR) T-cell therapy against extracellular targets and are non-MHC restricted. CARs are hybrid receptors that link the tumour antigen specificity and affinity of an antibody-derived, single-chain, variable fragment with the signalling endodomains associated with T-cell activation. CAR therapy targeting CD19 has yielded extraordinary clinical responses against some haematological tumours. Solid tumours in general remain an important challenge to cell transfer approaches but the case reports mentioned above provide huge impetus to the field.
Combination with standard therapies: It is now appreciated that standard treatment modalities (chemotherapeutics, radiation, interventional radiologic procedures) that have been used for decades can also activate the immune system, either indirectly — by depleting immune suppressive cells — or directly, via release of tumour antigen or provocation of immunogenic cell death. In GI cancer, this applies to some of the most commonly-employed chemotherapeutics, such as gemcitabine and oxaliplatin. The obvious implication is that these previously unappreciated or purely academic effects can now be taken advantage of by combination with checkpoint inhibition. Traditional therapies are, therefore, being re-evaluated — their dose, their schedule, their presumed activity — and administered now with an immune effect in mind, leading to a repurposing. We recently showed that ablative therapies — performed in a subtotal fashion in metastatic disease — can be effective when used in combination with anti-CTLA-4 therapy (NCT01853618) in patients with liver cancers.
The advantages of immune-based treatments are clear, such that the initial results of the checkpoint inhibitors, in particular, have captured the imaginations of physicians and the general public alike. Whilst the benefit seen thus far is real, we must acknowledge that it has been confined to a minority of patients. Those with GI cancers, in particular, could be forgiven for feeling left behind in the midst of all this hype and chatter. One hopes this will change in time.
There certainly are grounds for optimism, and the foundation for future development seems strong. Technological advances have led to superior pre-clinical models, as exemplified in pancreatic cancer by the autochthonous KPC mice for example, which better approximate the human situation, including the suppressive microenvironment and allow for better drug development and clinical trials.
Trial design itself has become more flexible, moving slightly away from the traditional and more rigid phase 1/2 pathway of cytotoxic development, with less emphasis on the initial dose-escalation portion, in favour of studies allowing multi-histology cohorts.
Co-operation across institutions and, more importantly, pharmaceutical companies has greatly improved, underpinned by government endeavours such as the former US Vice President Joe Biden’s Cancer Moonshot initiative, or philanthropic ones such as The Parker Institute for Cancer Immunotherapy.
It is a time of great optimism and energy, which has come, we must never forget, from incremental basic science discoveries, to say nothing of the effort and willingness of patients and their families to take part in clinical trials.