Gutting it Out: Developing Effective Immunotherapies for Patients With Colorectal Cancer.

Risk factors for colorectal cancer (CRC) include proinflammatory diets, sedentary habits, and obesity, in addition to genetic syndromes that predispose individuals to this disease. Current treatment relies on surgical excision and cytotoxic chemotherapies. There has been a renewed interest in immunotherapy as a treatment option for CRC given the success in melanoma and microsatellite instable (MSI) CRC. Immunotherapy with checkpoint inhibitors only plays a role in the 4%-6% of patients with MSIhigh tumors and even within this subpopulation, response rates can vary from 30% to 50%. Most patients with CRC do not respond to this modality of treatment, even though colorectal tumors are frequently infiltrated with T cells. Tumor cells limit apoptosis and survive following intensive chemotherapy leading to drug resistance and induction of autophagy. Pharmacological or molecular inhibition of autophagy improves the efficacy of cytotoxic chemotherapy in murine models. The microbiome clearly plays an etiologic role, in some or most colon tumors, realized by elegant findings in murine models and now investigated in human clinical trials. Recent results have suggested that cancer vaccines may be beneficial, perhaps best as preventive strategies. The search for therapies that can be combined with current approaches to increase their efficacy, and new knowledge of the biology of CRC are pivotal to improve the care of patients suffering from this disease. Here, we review the basic immunobiology of CRC, current "state-of-the-art" immunotherapies and define those areas with greatest therapeutic promise for the future.

The consensus immunoscore: toward a new classification of colorectal cancer.

In its latest edition, the WHO classification of the Digestive System Tumors introduced for the first time the immune response as essential and desirable diagnostic criteria for colorectal cancer. The immune response within the tumor microenvironment is therefore clinically relevant. The consensus Immunoscore has a prognostic value that has been confirmed in a meta-analysis on more than 10,000 patients, and it provides a reliable estimate of the recurrence risk in colon cancer. The international validation of the prognostic value of the consensus Immunoscore for time to recurrence, disease-free survival and overall survival in colon cancer together with its predictive value of response to chemotherapy provides valuable information for patient care management.

The consensus Immunoscore in phase 3 clinical trial (N0147) and impact on patient management decisions.

The consensus Immunoscore is a routine assay quantifying the adaptive immune response within the tumor microenvironment. It has a prognostic value that has been confirmed in a phase 3 clinical trial (NCCTG N0147) in stage III colon cancers. Moreover, results from another phase 3 randomized trial revealed the predictive value of Immunoscore for response to adjuvant chemotherapy duration. These results highlight the clinical utility of Immunoscore. In its latest edition, the World Health Organization classification of Digestive System Tumors introduced for the first time the immune response as an essential and desirable diagnostic criterion for colorectal cancer. Within the tumor microenvironment, the immune response provides an important estimate of the risk of recurrence and death in colon cancer. The international validation of the prognostic value of the consensus Immunoscore together with its prognostic value in the N0147 trial and its predictive utility for response to chemotherapy in stage III patients provide valuable information for patient management.

Immunoscore and its introduction in clinical practice.

Current cancer classification is based on intrinsic tumor parameters such as characteristic of tumor cells, tumor progression, tumor invasion and the presence of distant metastases, as summarized by the AJCC/UICC-TNM stratification system. There is currently no immune-based classification of cancer. We provided evidence that the immune contexture, as defined by the type, density, functional immune orientation and the location of immune cells within tumor, strongly influences the outcome of colorectal cancer (CRC) patients. To translate this information to the clinic, we developed an immunohistochemistry and digital pathology-based assay named Immunoscore that allows the quantification of two T cell subsets (CD3 and CD8) in two tumor regions (core and invasive margin of tumors). We have demonstrated that the standardized consensus Immunoscore has a prognostic value superior and independent to the one of the traditional TNM system in an international study in stage I-III colon cancer patients, therefore validating the Immunoscore as the first immune-based scoring system. In additional analyses, we showed the prognostic value of Immunoscore for stage II-IV CRC patients, as well as the predictive value of Immunoscore in stage III colon cancer patients. The Immunoscore may help predict and stratify patients who will benefit from adjuvant chemotherapy. Moreover, the Immunoscore allows the classification of tumors from hot to cold categories, and could consequently guide clinical decision for the choice of treatment.

Engineered Bispecific Antibodies with Exquisite HIV-1-Neutralizing Activity.

While the search for an efficacious HIV-1 vaccine remains elusive, emergence of a new generation of virus-neutralizing monoclonal antibodies (mAbs) has re-ignited the field of passive immunization for HIV-1 prevention. However, the plasticity of HIV-1 demands additional improvements to these mAbs to better ensure their clinical utility. Here, we report engineered bispecific antibodies that are the most potent and broad HIV-neutralizing antibodies to date. One bispecific antibody, 10E8V2.0/iMab, neutralized 118 HIV-1 pseudotyped viruses tested with a mean 50% inhibitory concentration (IC50) of 0.002 µg/mL. 10E8V2.0/iMab also potently neutralized 99% of viruses in a second panel of 200 HIV-1 isolates belonging to clade C, the dominant subtype accounting for ∼50% of new infections worldwide. Importantly, 10E8V2.0/iMab reduced virus load substantially in HIV-1-infected humanized mice and also provided complete protection when administered prior to virus challenge. These bispecific antibodies hold promise as novel prophylactic and/or therapeutic agents in the fight against HIV-1.

Effects of antigen-expressing immunostimulatory liposomes on chemotaxis and maturation of dendritic cells in vitro and in human skin explants.

PURPOSE: Antigen-Expressing Immunostimulatory Liposomes (AnExILs) represent a novel DNA vaccination platform based on the production of protein antigens from DNA templates inside liposomes mediated by an in vitro transcription and translation (IVTT) mix. The aim of this study was to analyze the effects of AnExILs on different dendritic cells (DCs) models and to better understand the role of the different components of this formulation on its adjuvanticity. METHODS: The effect of ß-galactosidase-expressing AnExILs on maturation and particle uptake by murine DC cell line, fresh human monocyte-derived DCs or human dermal DCs in skin explants was investigated and compared to the effects of either plain liposomes or IVTT mix alone. RESULTS: AnExILs induced efficient DC chemotaxis and promoted up-regulation of maturation markers on murine DCs, due to the presence of IVTT in the formulation. Furthermore, the amount of active ßGal associated with DCs was higher for AnExILs than for free ßGal expressed in IVTT or ßGal encapsulated into non-adjuvanted liposomes. Most interestingly, the same trend was observed with human DCs. CONCLUSIONS: Both IVTT mix and liposomal vehicles were shown to be key components of the AnExIL formulation responsible for its adjuvanticity. AnExILs combine antigen production, adjuvanticity and delivery in one system, and can efficiently activate both murine and human DCs.

Induction of humoral and cellular immune responses by antigen-expressing immunostimulatory liposomes.

Recently we have shown that liposomes can be used as artificial microbes for the production and delivery of DNA-encoded antigens. These so-called antigen-expressing immunostimulatory liposomes (AnExILs) were superior in inducing antigen-specific antibodies compared to conventional liposomal protein or DNA vaccines when tested in mice after i.m. immunization. In this study, we investigated the capacity of AnExILs to induce T-cell responses. By using a plasmid vector encoding a model antigen under control of both the prokaryotic T7 and the eukaryotic CMV promoter we hypothesized that antigen production could lead to CTL activation via two distinct routes: i. production of antigens inside the AnExILs with subsequent cross-presentation after processing by APCs and ii. endogenous production of antigens after AnExIL-mediated transfection of the pDNA. Although we were not able to demonstrate transfection-mediated expression of luc-NP in mice, i.m. injection of AnExILs producing luc-NP resulted in T-cell responses against the encoded NP epitope, as determined by tetramer staining. T-cell responses were comparable to the responses obtained after i.m. injection of naked pDNA. In order to find out whether CTL activation was caused by cross-presentation of the exogenous antigens produced inside AnExILs or by endogenous antigen production from transfection with the same pDNA source a second study was initiated in which the contribution of each of these effects could be separately determined. These results demonstrate that the observed T-cell responses were not exclusively caused by cross-presentation of the AnExIL-produced antigens alone, but were rather a combination of dose-dependent antigen cross-presentation and low levels of endogenous antigen production.

Anti-adenovirus humoral responses influence on the efficacy of vaccines based on epitope display on adenovirus capsid.

The efficacy of recombinant adenoviruses (Ads) vaccine vectors is diminished by the high prevalence of anti-Ad antibodies (Abs) that hampers gene transfer. Epitope display on Ad capsid constitutes an alternative approach to bypass anti-Ad Ab capacity from blocking antigen expression. To understand the role of the epitope insertion site, an ovalbumin-derived epitope was genetically inserted into either Ad hexon or fiber proteins. Hexon-modified Ads triggered higher anti-ovalbumin Ab responses after one injection but surprisingly fiber-modified Ads were by far more potent after two or several administrations. Our data unravel a role for anti-Ad humoral immunity in controlling anti-epitope humoral responses.