A Phase I Single-Arm Study of Biweekly NHS-IL12 in Patients With Metastatic Solid Tumors.

BACKGROUND: NHS-IL12 is a first-in-class, recombinant fusion protein composed of the human monoclonal antibody NHS76 (binds exposed DNA/histones at sites of intratumoral necrosis) fused to 2 IL-12 heterodimers. The maximum tolerated dose (MTD) and recommended phase II dose (RP2D) of NHS-IL12 monotherapy given subcutaneously (SC) every 4 weeks was previously reported. The study was expanded to include a high-exposure cohort with NHS-IL12 SC every 2 weeks (q2w). METHODS: This single-arm, phase I trial evaluated NHS-IL12 12 µg/kg SC q2w or 16.8µg/kg SC q2w in patients with metastatic solid tumors. The primary endpoint was safety. RESULTS: Using a 3+3 design, 13 patients with advanced cancer were enrolled and 12 were dose-limiting toxicity (DLT) evaluable. There was 1 DLT (Grade 3 aspartate transaminase/alanine transaminase [AST/ALT] elevation). Other grade 3 toxicities included: flu-like symptoms 1/13 (8%), decreased absolute lymphocyte count (ALC) 1/13 (8%), decreased white blood cell count (WBC) 1/13 (8%), but most adverse events reported were low grade and self-limiting grade. Fifty percent of evaluable patients (6/12) experienced stable disease (SD) with 42% (5/12) developing progressive disease (PD) at the first restaging. CONCLUSION: Biweekly NHS-IL12 was well tolerated in this small phase I study. Additional studies incorporating NHS-IL12 with other immunomodulating agents are underway. (ClinicalTrials.gov Identifier: NCT01417546).

A Randomized Phase II Trial of mFOLFOX6 + Bevacizumab Alone or with AdCEA Vaccine + Avelumab Immunotherapy for Untreated Metastatic Colorectal Cancer.

BACKGROUND: FOLFOX plus bevacizumab is a standard of care (SOC) for first-line treatment of microsatellite-stable metastatic colorectal cancer (MSS mCRC). This study randomized patients to SOC or SOC plus avelumab (anti-PD-L1) plus CEA-targeted vaccine. METHODS: Patients with untreated MSS mCRC enrolled to a lead-in arm assessing safety of SOC + immuno-oncology agents (IO). Next, patients were randomized to SOC or SOC + IO. The primary endpoint was progression-free survival (PFS). Multiple immune parameters were analyzed. RESULTS: Six patients enrolled to safety lead-in, 10 randomized to SOC, and 10 to SOC + IO. There was no difference in median PFS comparing SOC versus SOC + IO (8.8 months (95% CI: 3.3-17.0 months) versus 10.1 months (95% CI: 3.6-16.1 months), respectively; hazard ratio 1.061 [P = .91; 95% CI: 0.380-2.966]). The objective response rate was 50% in both arms. Of patients analyzed, most (8/11) who received SOC + IO developed multifunctional CD4+/CD8+ T-cell responses to cascade antigens MUC1 and/or brachyury, compared to 1/8 who received SOC alone (P = .020). We detected post-treatment changes in immune parameters that were distinct to the SOC and SOC + IO treatment arms. Accrual closed after an unplanned analysis predicted a low likelihood of meeting the primary endpoint. CONCLUSIONS: SOC + IO generated multifunctional MUC1- and brachyury-specific CD4+/CD8+ T cells despite concurrent chemotherapy. Although a tumor-directed immune response is necessary for T-cell-mediated antitumor activity, it was not sufficient to improve PFS. Adding agents that increase the number and function of effector cells may be required for clinical benefit.

An IL-15 superagonist/IL-15Rα fusion complex protects and rescues NK cell-cytotoxic function from TGF-ß1-mediated immunosuppression.

Natural killer (NK) cells are innate cytotoxic lymphocytes that play a fundamental role in the immunosurveillance of cancers. NK cells of cancer patients exhibit impaired function mediated by immunosuppressive factors released from the tumor microenvironment (TME), such as transforming growth factor (TGF)-ß1. An interleukin (IL)-15 superagonist/IL-15 receptor α fusion complex (IL-15SA/IL-15RA; ALT-803) activates the IL-15 receptor on CD8 T cells and NK cells, and has shown significant anti-tumor activity in several in vivo studies. This in vitro study investigated the efficacy of IL-15SA/IL-15RA on TGF-ß1-induced suppression of NK cell-cytotoxic function. IL-15SA/IL-15RA inhibited TGF-ß1 from decreasing NK cell lysis of four of four tumor cell lines (H460, LNCap, MCF7, MDA-MB-231). IL-15SA/IL-15RA rescued healthy donor and cancer patient NK cell-cytotoxicity, which had previously been suppressed by culture with TGF-ß1. TGF-ß1 downregulated expression of NK cell-activating markers and cytotoxic granules, such as CD226, NKG2D, NKp30, granzyme B, and perforin. Smad2/3 signaling was responsible for this TGF-ß1-induced downregulation of NK cell-activating markers and cytotoxic granules. IL-15SA/IL-15RA blocked Smad2/3-induced transcription, resulting in the rescue of NK cell-cytotoxic function from TGF-ß1-induced suppression. These findings suggest that in addition to increasing NK cell function via promoting the IL-15 signaling pathway, IL-15SA/IL-15RA can function as an inhibitor of TGF-ß1 signaling, providing a potential remedy for NK cell dysfunction in the immunosuppressive tumor microenvironment.

ADCC employing an NK cell line (haNK) expressing the high affinity CD16 allele with avelumab, an anti-PD-L1 antibody.

NK-92 cells, and their derivative, designated aNK, were obtained from a patient with non-Hodgkin lymphoma. Prior clinical studies employing adoptively transferred irradiated aNK cells have provided evidence of clinical benefit and an acceptable safety profile. aNK cells have now been engineered to express IL-2 and the high affinity (ha) CD16 allele (designated haNK). Avelumab is a human IgG1 anti-PD-L1 monoclonal antibody, which has shown evidence of clinical activity in a range of human tumors. Prior in vitro studies have shown that avelumab has the ability to mediate antibody-dependent cell-mediated cytotoxicity (ADCC) of human tumor cells when combined with NK cells. In the studies reported here, the ability of avelumab to enhance the lysis of a range of human carcinoma cells by irradiated haNK cells via the ADCC mechanism is demonstrated; this ADCC is shown to be inhibited by anti-CD16 blocking antibody and by concanamycin A, indicating the use of the granzyme/perforin pathway in tumor cell lysis. Studies also show that while NK cells have the ability to lyse aNK or haNK cells, the addition of NK cells to irradiated haNK cells does not inhibit haNK-mediated lysis of human tumor cells, with or without the addition of avelumab. Avelumab-mediated lysis of tumor cells by irradiated haNK cells is also shown to be similar to that of NK cells bearing the V/V Fc receptor high affinity allele. These studies thus provide the rationale for the clinical evaluation of the combined use of avelumab with that of irradiated adoptively transferred haNK cells.

A phase I study of recombinant (r) vaccinia-CEA(6D)-TRICOM and rFowlpox-CEA(6D)-TRICOM vaccines with GM-CSF and IFN-α-2b in patients with CEA-expressing carcinomas.

Prime-boost vaccination with recombinant (r) vaccinia(V)-CEA(6D)-TRICOM (triad of co-stimulatory molecules B7.1, ICAM-1 and LFA-3) and rFowlpox(F)-CEA(6D)-TRICOM infect antigen-presenting cells and direct expression of co-stimulatory molecules. We hypothesized that co-administration of vaccine with GM-CSF and interferon alpha (IFN-α) would have efficacy in CEA-expressing cancers. Patients with CEA-expressing cancers received the rV-CEA(6D)-TRICOM vaccine subcutaneously (s.c.) on day 1 followed by GM-CSF s.c. to the injection site on days 1-4. In Cycle 1, patients received thrice weekly s.c. injections of IFN-α-2b the week after rV-CEA(6D)-TRICOM. In Cycles 2-4, patients received thrice weekly s.c. injections of IFN-α-2b the same week that rF-CEA(6D)-TRICOM was given. The first cohort received no IFN followed by dose escalation of IFN-α in subsequent cohorts. Thirty-three patients were accrued (mean 59.8 years). Grade 3 toxicities included fatigue and hyperglycemia. Grade 4-5 adverse events (unrelated to treatment) were confusion (1), elevated aspartate transaminase (AST)/alanine transaminase (ALT) (1), and sudden death (1). No patients had a partial response, and eight patients exhibited stable disease of ≥3 months. Median progression-free survival and overall survival (OS) were 1.8 and 6.3 months, respectively. Significantly higher serum CD27 levels were observed after vaccine therapy (p = 0.006 post 1-2 cycles, p = 0.003 post 3 cycles, p = 0.03 post 4-7 cycles) and 42 % of patients assayed developed CEA-specific T cell responses. Pre-treatment levels of myeloid-derived suppressor cells correlated with overall survival (p = 0.04). Administration of IFN-α led to significantly increased OS (p = 0.02) compared to vaccine alone. While the vaccine regimen produced no clinical responses, IFN-α administration was associated with improved survival.

Pan-Bcl-2 inhibitor, GX15-070 (obatoclax), decreases human T regulatory lymphocytes while preserving effector T lymphocytes: a rationale for its use in combination immunotherapy.

Bcl-2 inhibitors are currently being evaluated in clinical studies for treatment of patients with solid tumors and hematopoietic malignancies. In this study we explored the potential for combining the pan-Bcl-2 inhibitor GX15-070 (GX15; obatoclax) with immunotherapeutic modalities. We evaluated the in vitro effects of GX15 on human T cell subsets obtained from PBMCs in terms of activation, memory, and suppressive function. Our results indicated that in healthy-donor PBMCs, mature-activated T cells were more resistant to GX15 than early-activated T cells, and that GX15 preserved memory but not non-memory T cell populations. Furthermore, GX15 increased the apoptosis of regulatory T cells (Tregs), profoundly downregulated FOXP3 and CTLA-4 in a dose-dependent manner, and decreased their suppressive function. Treating PBMCs obtained from ovarian cancer patients with GX15 also resulted in increased CD8(+):Treg and CD4(+):Treg ratios. These results support preclinical studies in which mice vaccinated before treatment with GX15 showed the greatest reduction in metastatic lung tumors as a result of increased apoptotic resistance of mature CD8(+) T cells and decreased Treg function brought about by GX15. Taken together, these findings suggest that when a Bcl-2 inhibitor is combined with active immunotherapy in humans, such as the use of a vaccine or immune checkpoint inhibitor, immunotherapy should precede administration of the Bcl-2 inhibitor to allow T cells to become mature, and thus resistant to the cytotoxic effects of the Bcl-2 inhibitor.

Soluble CD27-pool in humans may contribute to T cell activation and tumor immunity.

The interaction between CD27 and its ligand, CD70, has been implicated in regulating cellular immune responses to cancer. In this article, we report on the role of soluble CD27 (sCD27) in T cell activation and its elevation in the serum of cancer patients after immunotherapy. In vitro, sCD27 is preferentially derived from activated CD4(+) T cells. Adding sCD27 to stimulated PBMCs increases T cell activation and proliferation, and is associated with the immunologic synapse-related proteins myosin IIA, high mobility group box 1, and the TCR Vß-chain. The pool of serum sCD27 is shown to be greater in healthy donors than in cancer patients. However, metastatic cancer patients treated with immunotherapy showed a significant increase in the serum sCD27-pool posttherapy (p < 0.0005); there was also an increased trend toward an association between enhanced sCD27-pool posttherapy and overall survival (p = 0.022). The identification of sCD27 as an immune modulator associated with enhanced human T cell activation in vitro and in vivo provides a rationale for developing new immunotherapeutic strategies aimed at enhancing sCD27 for treating cancer and potentially other diseases.

Identification by digital immunohistochemistry of intratumoral changes of immune infiltrates after vaccine in the absence of modifications of PBMC immune cell subsets.

Preclinical studies have demonstrated that the combination of systemic subcutaneous (s.c.) vaccination with intratumoral (i.t.) vaccination was superior in the induction of antitumor activity vs. vaccination with either route alone. A subsequent phase I study employing i.t.-s.c. vaccination was carried out in men with locally recurrent or progressive prostate cancer. rF-PSA-TRICOM (PROSTVAC) vaccine was administered intraprostatically and rV-PSA-TRICOM followed by rF-PSA-TRICOM vaccine was administered systemically. In that study no dose limiting toxicities were observed, 19/21 patients had stable or improved prostate-specific antigen (PSA) values and tumor-infiltrating lymphocytes (TILs) increased in post- vs. pre-treatment tumor biopsies, analyzed employing conventional immunohistochemistry (IHC). In the studies reported here, 31 phenotypes of peripheral blood mononuclear cells (PBMCs) were analyzed prevaccination and postvaccination as well as the functions of PBMC regulatory T cells (Tregs) and natural killer cells. A trend was observed in decreases in serum PSA with the reduction of circulating Tregs postvaccination. Digital IHC was employed prevaccination and postvaccination to measure CD4 and CD8 TILs, as well as Treg TILs by conventional IHC. Few correlations were observed with CD4, CD8 or Treg in TILs vs. PBMCs. However, patients with lower levels of CD4 TILs prevaccination showed the greatest increases in CD4 TILs postvaccine, while Treg TILs decreased postvaccine. There was also a strong correlation between decreases in serum PSA and increases in CD8 TILs postvaccine. These studies provide additional rationale for the use of i.t.-s.c. vaccinations and demonstrate a noncoordinate expression of specific immune subsets in PBMCs vs. tumor.

Identification and characterization of agonist epitopes of the MUC1-C oncoprotein.

The MUC1 tumor-associated antigen is overexpressed in the majority of human carcinomas and several hematologic malignancies. Much attention has been paid to the hypoglycosylated variable number of tandem repeats (VNTR) region of the N-terminus of MUC1 as a vaccine target, and recombinant viral vector vaccines are also being evaluated that express the entire MUC1 transgene. While previous studies have described MUC1 as a tumor-associated tissue differentiation antigen, studies have now determined that the C-terminus of MUC1 (MUC1-C) is an oncoprotein, and its expression is an indication of poor prognosis in numerous tumor types. We report here the identification of nine potential CD8⁺ cytotoxic T lymphocyte epitopes of MUC1, seven in the C-terminus and two in the VNTR region, and have identified enhancer agonist peptides for each of these epitopes. These epitopes span HLA-A2, HLA-A3, and HLA-A24 major histocompatibility complex (MHC) class I alleles, which encompass the majority of the population. The agonist peptides, compared to the native peptides, more efficiently (a) generate T-cell lines from the peripheral blood mononuclear cells of cancer patients, (b) enhance the production of IFN-γ by peptide-activated human T cells, and (c) lyse human tumor cell targets in an MHC-restricted manner. The agonist epitopes described here can be incorporated into various vaccine platforms and for the ex vivo generation of human T cells. These studies provide the rationale for the T-cell-mediated targeting of the oncogenic MUC1-C, which has been shown to be an important factor in both drug resistance and poor prognosis for numerous tumor types.

Identification and characterization of a cytotoxic T-lymphocyte agonist epitope of brachyury, a transcription factor involved in epithelial to mesenchymal transition and metastasis.

The transcription factor brachyury is a major driver of epithelial to mesenchymal transition in human carcinoma cells. It is overexpressed in several human tumor types versus normal adult tissues, except for testes and thyroid. Overexpression is associated with drug resistance and poor prognosis. Previous studies identified a brachyury HLA-A2 cytotoxic T-lymphocyte epitope. The studies reported here describe an enhancer epitope of brachyury. Compared to the native epitope, the agonist epitope: (a) has enhanced binding to MHC class I, (b) increased the IFN-γ production from brachyury-specific T cells, (c) generated brachyury-specific T cells with greater levels of perforin and increased proliferation, (d) generated T cells more proficient at lysing human carcinoma cells endogenously expressing the native epitope, and (e) achieved greater brachyury-specific T-cell responses in vivo in HLA-A2 transgenic mice. These studies also report the generation of a heat-killed recombinant Saccharomyces cerevisiae (yeast) vector expressing the full-length brachyury gene encoding the agonist epitope. Compared to yeast-brachyury (native) devoid of the agonist epitope, the yeast-brachyury (agonist) enhanced the activation of brachyury-specific T cells, which efficiently lysed human carcinoma cells. In addition to providing the rationale for the recombinant yeast-brachyury (agonist) as a potential vaccine in cancer therapy, these studies also provide the rationale for the use of the agonist in (a) dendritic cell (DC) vaccines, (b) adjuvant or liposomal vaccines, (c) recombinant viral and/or bacterial vaccines, (d) protein/polypeptide vaccines, (e) activation of T cells ex vivo in adoptive therapy protocols, and (f) generation of genetically engineered targeted T cells.

A combination trial of vaccine plus ipilimumab in metastatic castration-resistant prostate cancer patients: immune correlates.

We recently reported the clinical results of a Phase I trial combining ipilimumab with a vaccine containing transgenes for prostate-specific antigen (PSA) and for a triad of costimulatory molecules (PROSTVAC) in patients with metastatic castration-resistant prostate cancer. Thirty patients were treated with escalating ipilimumab and a fixed dose of vaccine. Of 24 chemotherapy-naïve patients, 58 % had a PSA decline. Combination therapy did not exacerbate the immune-related adverse events associated with ipilimumab. Here, we present updated survival data and an evaluation of 36 immune cell subsets pre- and post-therapy. Peripheral blood mononuclear cells were collected before therapy, at 13 days and at 70 days post-initiation of therapy, and phenotyped by flow cytometry for the subsets of T cells, regulatory T cells, natural killer cells, and myeloid-derived suppressor cells. Associations between overall survival (OS) and immune cell subsets prior to treatment, and the change in a given immune cell subset 70 days post-initiation of therapy, were evaluated. The median OS was 2.63 years (1.77-3.45). There were trends toward associations for longer OS and certain immune cell subsets before immunotherapy: lower PD-1(+)Tim-3(NEG)CD4EM (P = 0.005, adjusted P = 0.010), higher PD-1(NEG)Tim-3(+)CD8 (P = 0.002, adjusted P = 0.004), and a higher number of CTLA-4(NEG) Tregs (P = 0.005, adjusted P = 0.010). We also found that an increase in Tim-3(+) natural killer cells post- versus pre-vaccination associated with longer OS (P = 0.0074, adjusted P = 0.015). These results should be considered as hypothesis generating and should be further evaluated in larger immunotherapy trials.

Phase I trial of a recombinant yeast-CEA vaccine (GI-6207) in adults with metastatic CEA-expressing carcinoma.

Yeast-CEA (GI-6207) is a therapeutic cancer vaccine genetically modified to express recombinant carcinoembryonic antigen (CEA) protein, using heat-killed yeast (Saccharomyces cerevisiae) as a vector. In preclinical studies, yeast-CEA induced a strong immune response to CEA and antitumor responses. Patients received subcutaneous vaccines every 2 weeks for 3 months and then monthly. Patients were enrolled at 3 sequential dose levels: 4, 16, and 40 yeast units (10(7) yeast particles/unit). Eligible patients were required to have serum CEA > 5 ng/mL or > 20 % CEA(+) tumor block, ECOG PS 0-2, and no history of autoimmunity. Restaging scans were performed at 3 months and then bimonthly. Peripheral blood was collected for the analysis of immune response (e.g., by ELISPOT assay). Twenty-five patients with metastatic CEA-expressing carcinomas were enrolled. Median patient age was 52 (range 39-81). A total of 135 vaccines were administered. The vaccine was well tolerated, and the most common adverse event was grade 1/2 injection-site reaction. Five patients had stable disease beyond 3 months (range 3.5-18 months), and each had CEA stabilization while on-study. Some patients showed evidence post-vaccination of increases in antigen-specific CD8(+) T cells and CD4(+) T lymphocytes and decreases in regulatory T cells. Of note, a patient with medullary thyroid cancer had substantial T cell responses and a vigorous inflammatory reaction at sites of metastatic disease. Yeast-CEA vaccination had minimal toxicity and induced some antigen-specific T cell responses and CEA stabilization in a heterogeneous, heavily pre-treated patient population. Further studies are required to determine the clinical benefit of yeast-CEA vaccination.

Elevated serum soluble CD40 ligand in cancer patients may play an immunosuppressive role.

Tumor cells can induce certain cytokines and soluble receptors that have a suppressive effect on the immune system. In this study, we showed that an extracellular portion of a membrane-bound ligand of CD40 (soluble CD40 ligand; sCD40L) was significantly elevated in the serum of cancer patients compared with healthy donors. In addition, PBMCs from cancer patients had a relatively larger population of myeloid-derived suppressor cells (MDSCs), defined as CD33(+)HLA-DR(-) cells, and these cells expressed higher levels of CD40. T-cell proliferation and IFN-γ production decreased when stimulated T cells were cocultured with an increased amount of autologous MDSCs. The addition of recombinant monomeric sCD40L enriched MDSCs and had an additive inhibitory effect on T-cell proliferation. PBMCs cultured in vitro with sCD40L also showed an expansion of regulatory T cells (CD4(+)CD25(high)Foxp3(+)), as well as induction of cytokines, such as IL-10 and IL-6. Moreover, sCD40L-induced enrichment of programmed death-1-expressing T cells was greater in cancer patients than in healthy donors. Preexisting sCD40L also inhibited IL-12 production from monocytes on activation. These data suggest that the higher levels of sCD40L seen in cancer patients may have an immunosuppressive effect.

Adjuvant nivolumab, capecitabine or the combination in patients with residual triple-negative breast cancer: the OXEL randomized phase II study.

Chemotherapy and immune checkpoint inhibitors have a role in the post-neoadjuvant setting in patients with triple-negative breast cancer (TNBC). However, the effects of nivolumab, a checkpoint inhibitor, capecitabine, or the combination in changing peripheral immunoscore (PIS) remains unclear. This open-label randomized phase II OXEL study (NCT03487666) aimed to assess the immunologic effects of nivolumab, capecitabine, or the combination in terms of the change in PIS (primary endpoint). Secondary endpoints included the presence of ctDNA, toxicity, clinical outcomes at 2-years and association of ctDNA and PIS with clinical outcomes. Forty-five women with TNBC and residual invasive disease after standard neoadjuvant chemotherapy were randomized to nivolumab, capecitabine, or the combination. Here we show that a combination of nivolumab plus capecitabine leads to a greater increase in PIS from baseline to week 6 (91%) compared with nivolumab (47%) or capecitabine (53%) alone (log-rank p = 0.08), meeting the pre-specified primary endpoint. In addition, the presence of circulating tumor DNA (ctDNA) is associated with disease recurrence, with no new safety signals in the combination arm. Our results provide efficacy and safety data on this combination in TNBC and support further development of PIS and ctDNA analyses to identify patients at high risk of recurrence.

Up-regulation of proliferative and migratory genes in regulatory T cells from patients with metastatic castration-resistant prostate cancer.

A higher frequency of regulatory T cells (Tregs) has been observed in peripheral blood mononuclear cells (PBMC) of patients with different types of solid tumors and hematological malignancies as compared to healthy donors. In prostate cancer patients, Tregs in PBMC have been shown to have increased suppressive function. Tumor-induced biological changes in Tregs may enable tumor cells to escape immunosurveillance. We performed genome-wide expression analyses comparing the expression levels of more than 38,500 genes in Tregs with similar suppressive activity, isolated from the peripheral blood of healthy donors and patients with metastatic castration-resistant prostate cancer (mCRPC). The differentially expressed genes in mCRPC Tregs are involved in cell cycle processes, cellular growth and proliferation, immune responses, hematological system development and function and the interleukin-2 (IL-2) and transforming growth factor-ß (TGF-ß) pathways. Studies revealed that the levels of expression of genes responsible for T-cell proliferation (C-FOS, C-JUN and DUSP1) and cellular migration (RGS1) were greater in Tregs from mCRPC patients as compared to values observed in healthy donors. Increased RGS1 expression in Tregs from mCRPC patients suggests a decrease in these Tregs' migratory ability. In addition, the higher frequency of CD4(+) CD25(high) CD127(-) Tregs in the peripheral blood of mCRPC patients may be the result of an increase in Treg proliferation capacity. Results also suggest that the alterations observed in gene expression profiles of Tregs in mCRPC patients may be part of the mechanism of tumor escape from host immune surveillance.

Effect of talactoferrin alfa on the immune system in adults with non-small cell lung cancer.

BACKGROUND: Talactoferrin alfa (talactoferrin), an agent with immune-stimulating properties, has demonstrated safety and preliminary efficacy in clinical trials. METHODS: Ten patients (five males and five females) with stage IV non-small cell lung cancer (NSCLC) in a single-arm pilot study received orally administered talactoferrin (1.5 g, b.i.d.) for up to 24 weeks. Radiographic and immunologic studies were performed at baseline and at weeks 6 and 12. Circulating immune cells (natural killer cells [NKCs], CD4+, CD8+, and regulatory T cells) and systemic cytokine levels were measured to assess immune response. RESULTS: Patients enrolled in the study had received a median of four prior chemotherapy regimens, and all patients were symptomatic. Talactoferrin was well tolerated, with no grade 3 or 4 toxicities. Median time to progression (TTP) and overall survival were 6 weeks and 14.5 weeks, respectively. The four patients with ≥9 weeks TTP had evidence of immunologic activity (three with increased NKC activity). CONCLUSIONS: The median of four previous chemotherapy regimens, with elevated levels of interleukin (IL) 6 and tumor necrosis factor-alfa in most patients, suggests these patients were poor candidates for immunotherapy.

Adjuvant nivolumab, capecitabine or the combination in patients with residual triple-negative breast cancer: the OXEL randomized phase II study.

Chemotherapy and immune checkpoint inhibitors have a role in the post-neoadjuvant setting in patients with triple-negative breast cancer (TNBC). However, the effects of nivolumab, a checkpoint inhibitor, capecitabine, or the combination in changing peripheral immunoscore (PIS) remains unclear. This open-label randomized phase II OXEL study (NCT03487666) aimed to assess the immunologic effects of nivolumab, capecitabine, or the combination in terms of the change in PIS (primary endpoint). Secondary endpoints include the presence of ctDNA, toxicity, clinical outcomes at 2-years and association of ctDNA and PIS with clinical outcomes. Forty-five women with TNBC and residual invasive disease after standard neoadjuvant chemotherapy were randomized to nivolumab, capecitabine, or the combination. Here we show that a combination of nivolumab plus capecitabine leads to a greater increase in PIS from baseline to week 6 (91%) compared with nivolumab (47%) or capecitabine (53%) alone (log-rank p = 0.08), meeting the pre-specified primary endpoint. In addition, the presence of circulating tumor DNA (ctDNA) was associated with disease recurrence, with no new safety signals in the combination arm. Our results provide efficacy and safety data on this combination in TNBC and support further development of PIS and ctDNA analyses to identify patients at high risk of recurrence.

Immune correlates of clinical parameters in patients with HPV-associated malignancies treated with bintrafusp alfa.

PURPOSE: Bintrafusp alfa is a bifunctional agent consisting of an anti-human PD-L1 antibody linked to two TGFßRII. It is designed to act both as a checkpoint inhibitor and to 'trap' TGFß in the tumor microenvironment. Phase I and II clinical studies demonstrated clinical activity in patients with a range of human papillomavirus (HPV)-associated cancers. The purpose of the studies reported here was the interrogation of various aspects of the peripheral immunome in patients with HPV-associated cancers, both prior to and early in the treatment regimen of bintrafusp alfa to better understand the mode of action of the agent and to help define which patients are more likely to benefit from bintrafusp alfa treatment. PATIENTS AND METHODS: The peripheral immunome of patients (n=65) with HPV+ malignancies was analyzed both prior to treatment with bintrafusp alfa and day 14 post-treatment for levels and changes in (1) 158 different immune cell subsets, (2) multiple plasma soluble factors including analytes reflecting immune stimulatory and inhibitory status, (3) complete blood counts, and in a subset of patients (4) TCR diversity and (5) HPV-specific T-cell responses. RESULTS: Interrogation of the peripheral immunome prior to bintrafusp alfa treatment revealed several factors that associated with clinical response, including (1) higher levels of sCD27:sCD40L ratios, (2) lower levels of TGFß1 and 12 additional factors associated with tumor mesenchymalization, and (3) higher CD8+ T cell:MDSC ratios. Analysis at 2 weeks post bintrafusp alfa revealed that eventual clinical responders had fewer increases in IL-8 levels and the neutrophil to lymphocyte ratio, and higher levels of HPV-16 specific CD8+ T cells. This study also provided information concerning differences in the peripheral immunome for patients who were naïve versus refractory to prior checkpoint inhibition therapy. While preliminary, two multivariate models developed predicted clinical benefit with 76%-91% accuracy. CONCLUSIONS: These studies add insight into the mechanism of action of bintrafusp alfa and provide evidence that the interrogation of both cellular and soluble components of the peripheral immunome of patients with HPV-associated malignancies, either prior to or early in the therapeutic regimen, can provide information as to which patients are more likely to benefit with bintrafusp alfa therapy.

Combination Therapies for HPV-Associated Malignancies.

Human papillomavirus (HPV)-associated malignancies cause almost all cases of cervical cancer in women, and a significant percentage of head and neck cancer, together totaling almost 5% of the global cancer burden, and representing an important public health issue. The approval and use of two prophylactic HPV vaccines, Gardasil® and Cervarix®, have significantly decreased infections with HPV, but unfortunately, prophylactic vaccination does not treat established infections or malignancies resulting from HPV. Therefore, therapies for HPV-associated malignancies are necessary to improve the quality of life and survival in patients with these diseases. This review will detail new combinations of therapies in clinical development for HPV-associated malignancies.

Preclinical study of a novel therapeutic vaccine for recurrent respiratory papillomatosis.

Activation of antigen-specific T-lymphocyte responses may be needed to cure disorders caused by chronic infection with low-risk human papillomavirus (lrHPV). Safe and effective adjuvant therapies for such disorders are needed. The safety and efficacy of a novel gorilla adenovirus vaccine expressing a protein designed to elicit immune responses directed against HPV6 and HPV11, PRGN-2012, was studied using in vitro stimulation of T lymphocytes from patients with recurrent respiratory papillomatosis, in vivo vaccination studies, and therapeutic studies in mice bearing tumors expressing lrHPV antigen. PRGN-2012 treatment induces lrHPV antigen-specific responses in patient T lymphocytes. Vaccination of wild-type mice induces E6-specific T-lymphocyte responses without toxicity. In vivo therapeutic vaccination of mice bearing established HPV6 E6 expressing tumors results in HPV6 E6-specific CD8+ T-lymphocyte immunity of sufficient magnitude to induce tumor growth delay. The clinical study of PRGN-2012 in patients with disorders caused by chronic infection with lrHPV is warranted.