Mini-review

InCVAX A novel strategy for treatment of late-stage, metastaticcancers through photoimmunotherapy induced tumor-specicimmunityFeifan Zhou a,1, Xiaosong Li b,1, Mark F. Naylor c, Tomas Hode d, Robert E. Nordquist d,Luciano Alleruzzo d, Joseph Raker d, Samuel S.K. Lam d, Nan Du b, Lei Shi e, Xiuli Wang e,Wei R. Chen a,d,*a Biophotonics Research Laboratory, Center for Interdisciplinary Biomedical Education and Research, University of Central Oklahoma, Edmond, Oklahoma73034, USAb Department of Oncology, the First Aliated Hospital of Chinese PLA General Hospital, Beijing 100048, Chinac Dermatology Associates of San Antonio Texas, San Antonio, Texas 78233, USAd Immunophotonics Inc., 4320 Forest Park Avenue #303, St. Louis, Missouri 63108, USAe Department of Dermatology and Venereology, Shanghai Skin Diseases Hospital, Shanghai 200050, China

A R T I C L E I N F O

Article history:Received 20 November 2014Received in revised form 20 January 2015Accepted 21 January 2015

Keywords:inCVAXLaser immunotherapy (LIT)N-dihydro-galacto-chitosan (GC)Metastatic cancersAntitumor immune responses

A B S T R A C T

A novel, promising potential cancer vaccine strategy was proposed to use a two-injection procedure forsolid tumors to prompt the immune system to identify and systemically eliminate primary and meta-static cancers. The two-injection procedure consists of local photothermal application on a selected tumorintended to liberate whole cell tumor antigens, followed by a local injection of an immunoadjuvant thatconsists of a semi-synthetic functionalized glucosamine polymer, N-dihydro-galacto-chitosan (GC), whichis intended to activate antigen presenting cells and facilitate an increased uptake of tumor antigens. Thisstrategy is thus proposed as an in situ autologous cancer vaccine (inCVAX) that may activate antigen pre-senting cells and expose them to tumor antigens in situ, with the intention of inducing a systemic tumorspecic T-cell response. Here, the development of inCVAX for the treatment of metastatic cancers in thepast decades is systematically reviewed. The antitumor immune responses of local photothermal treat-ment and immunological stimulationwith GC are also discussed. This treatment approach is also commonlyreferred to as laser immunotherapy (LIT).

2015 Elsevier Ireland Ltd. All rights reserved.

Introduction

Metastases are the major cause of treatment failure and cancer-related deaths. Unfortunately, tumor metastases to multiple sitesoften occur so early that by the time primary tumors are detected,a large portion of the patientswith solid tumors already have eitherclinically apparentmetastases or clinically occultmicro-metastases[13]. Many approaches such as chemotherapy, radiation therapy,hormonal therapy, and other targeted therapies have been appliedto treatmetastatic cancers. Although some typesofmetastatic cancerscan be treated effectively with current treatments, many cannot.

The ultimate control of cancer has been suggested to lie in thehost immune surveillance and defense system [46]. Immuno-therapy has been considered a promising treatment approach [7],and various strategies have been proposed, including cytokine

therapy [8,9], dendritic cell-based vaccines [10,11], and check-point inhibitors and immune-activating antibodies, many of whichhave begun to be used in clinical studies, either alone or in variouscombinations with other therapies [1214]. However, immuno-therapy is only accepted as a treatment option for a few indications,andmost cancers still avoid or escape immune control [1517]. Deathfrom metastatic cancer remains the most likely prognosis.

It is theorized that a metastasis originates from a primary cancer.Moreover, metastatic cancer cells and cells of the original cancerusually have some molecular features in common, such as the ex-pression of certain proteins or the presence of specic chromosomechanges [1]. The ideal cancer therapy should not only destroy theprimary tumors, but also at the same time trigger the immunesystem to recognize, track down, and destroy any remaining tumorcells, be they at or near the site of the primary tumor or at distantsites. In view of these desirable properties, some local therapy mo-dalities that are combined with immunotherapy have beendeveloped for metastatic cancers [12,18,19]. One such strategy isto combine photothermal therapy with an injection of animmunoadjuvant that consists of a semi-synthetic functionalized

* Corresponding author. Tel.: +405 974 5147; fax: +405 974 3812.E-mail address: wchen@uco.edu (W.R. Chen).

1 Both authors contributed equally to this work.

http://dx.doi.org/10.1016/j.canlet.2015.01.0290304-3835/ 2015 Elsevier Ireland Ltd. All rights reserved.

Cancer Letters 359 (2015) 169177

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glucosamine polymer,N-dihydro-galacto-chitosan (GC), which in turninduces an in situ autologous cancer vaccine (inCVAX). This treat-ment approach is also commonly referred to as laser immunotherapy(LIT) [18,2023].

It is well known that one of themajor reasons that tumors escapethe host immune system is their insucient expression of mol-ecules necessary for antigen processing and presentation [17]. Laser-induced tumor cell death, on the other hand, can release tumorantigens into the surrounding milieu [24]. Concomitantly,immunoadjuvants for cancer immunotherapy can promote antigenuptake and presentation by antigen-presenting cells (APCs), thustriggering specic antitumor immunity [20,22]. The pre-clinicalstudies of inCVAX showed that the combination of the selective pho-tothermal interaction and GC, both applied locally, could not onlydestroy the treated primary tumors but could also eradicate un-treated metastases at distant sites [20,21,2527]. First-in-humanclinical studies on late-stage, metastatic melanoma and breast cancerpatients were also conducted, and the preliminary data indicate thatinCVAX shows promise [28,29].

It is noticeable that the photothermally induced tumor cell deathby inCVAX provided a source of tumor antigens released from thehosts own tumor, which contrasts other current vaccine strate-gies (peptide or cDNA), most of which concentrate on a single andidentied tumor-specic antigen (TSA) [30]. Since TSAs vary fromtumor to tumor and from patient to patient, inCVAX, in lieu ofcurrent vaccine strategies, may have the potential to vaccinate pa-tients against multiple TSAs that originate from the individualpatients own tumors.

Components of inCVAX

inCVAXutilizes twomajor interactions: (1) a selective local tumorcell death and tumor antigen release bydirect delivery of laser energyinto the tumor tissue; (2) the local administration of GC as animmunoadjuvant, enhancing antigen uptake and presentation, elic-iting a specic antitumor immunity [21,31]. The two componentsof inCVAX play a unique role on the induced antitumor immuneresponse.

Photothermal therapy (PTT)

PTT is a therapeutic strategy for local treatment of cancer thatuses heat generated from absorbed light energy to destroy tumorcells, which could be highly specic, much less invasive, and rathereffective due to the intensive light directed to the target tumor[32,33]. Laser ablation can generate a thermal gradient inside thetarget tissue, which may induce different biological responses. Itshould be noted that at temperatures above 105 C, carbonizationand evaporation of tissue are induced, which are generally consid-ered undesirable in laser ablation, as it changes the optical propertiesof the tissue and reduces the light penetration into the deeper targettissue [34]. To increase its ecacy, the parameters of the laser shouldbe controlled according to the interactions with tumor tissue.

To study the thermal effect on tumor tissue, selective photo-thermal interaction using laser, and in situ light-absorbing dye, wasinitially developed. Specically, an 805-nm laser was used in con-junctionwith indocyanine green (ICG), since biological tissues exhibita deep penetrability with low absorption of near-infrared (NIR)photons in the wavelength of 805 nm, and ICG solution has a highabsorption peak around 800 nm, which makes it an ideal candi-date for selective photothermal therapy [32,35,36].

In the past decade, PTT using various types of NIR-absorbingagents has also been explored to enhance the local thermal effects,including gold nanoparticles, carbon nanotubes and othernanomaterials [3742]. Moreover, direct application of PTT throughinterstitial bers, without the use of NIR-absorbing agents, has been

explored, yielding positive results. However, although PTT has beendemonstrated to be a powerful approach to destroy or interrupttumor cells, the control of metastatic tumors using PTT alone hasnot been achieved thus far.

Immunoadjuvant

Clinical application of adjuvantsAdjuvants are immunological agents thatmodify or augment an

immune response, usually to a vaccine, without having any specif-ic antigenic effect on their own [43,44]. Althoughhundredsof adjuvantcandidates have been developed by preclinical and clinical testing,most of them failed to be approved for routine application [45].Muchprogress has been made during the past decades and several noveladjuvants have been licensed for human use in different countries,including aluminum salts, squalene-oil-water emulsion (MF-59),monophosphory lipid A (MPL) and virosomes [46,47]. The mostcommon adjuvants for human use today are still aluminum hy-droxide and aluminum phosphate. However, the application ofaluminumhydroxidehasnot beenwidelyusedbecauseof aluminum-related macrophage myofascitis. Novel adjuvants still need to bedeveloped for clinical practice. An ideal adjuvant would elicit apersistent, high anity immune response to an antigenwhile beingnon-toxic, biodegradable, non-immunogenic and chemically denedfor reproducible manufacture [43,4851].

Immunoadjuvants are often used tomodify or augment the effectsof cancer vaccine by stimulating the immune system to respond tothe vaccine more vigorously, and thus providing increased immu-nity against the cancer [52,53]. Adjuvants exert their effects throughdifferent mechanisms [54,55]: (1) elevate the presence of anti-gens in the blood; (2) help absorb the antigen by antigen presentingcells; (3) activate macrophages, lymphocytes, and other immunecells; (4) support the production of cytokines.

N-dihydro-galacto-chitosan (GC)Chitin, sometimes referred to as Poly-N-acetylglucosamine

(PNAG), is a naturally occurring biopolymer that is an important com-ponent in, for example, many biolms, exoskeletons of crustaceansand insects, and cell walls in fungi. While only soluble in acidic so-lutions [56,57], it has been found that chitin and its N-deacetylatedform (dPNAG), referred to as chitosan if the degree of deacetylationis >50%, have certain immune stimulating properties, and they havethus been proposed as potential adjuvants for clinical practice [58,59].The immune response induced by PNAG and dPNAG is inuencedby the presence of antigens [60], and it is believed that chitosan inparticular could enhance both humoral and cell-mediated immuneresponses when vaccinated with antigens [6164].

To enhance the immunological functions of chitosan, and toimprove its bioavailability and usability in biomedical applica-tions by increasing its water solubility, N-dihydro-galacto-chitosan(GC) was synthesized by attaching galactose molecules to the freeamino groups on chitosan [31,65], as shown in Fig. 1. As afunctionalized, soluble new chemical entity, GC apparently pos-sesses improved properties for immunological stimulation in non-clinical and clinical studies, indicating its effect as immunoadjuvantin the treatment of metastatic cancers with the inCVAX approach[18,20,21,2527,31].

Effects of inCVAX

As a novel technique, inCVAX was performed with a consistentdemonstration of a long-term response of the immunological defensesystem against residual primary andmetastatic tumor cells. Throughlocal intervention, inCVAX could destroy treated primary tumorswhile also eradicating untreated metastases at distant sites[18,20,26,31].

170 F. Zhou et al./Cancer Letters 359 (2015) 169177

Effects of inCVAX on animal tumor model

The initially proposed inCVAX approach combined three majorcomponents: a near-infrared laser (805 nm), a light-absorbing agent(ICG), and an immunoadjuvant (GC). For the inCVAX implementa-tion, ICG was injected intratumorally, followed by topical laserirradiation to enhance light absorption in the tumor tissue [18,27].The procedure has since then been improved by removing the ICGcomponent, and the optical ber is now injected intratumorally fordirect photothermal application. GC, as an immunological stimu-lant, is injected into the same location (in the tumor) where the laserirradiationwas performed to further enhance immune response afterthe photothermal treatment [21,66]. Fig. 2 shows the non-invasivelaser irradiation of rat tumors.

With a poorly immunogenic rat mammary DMBA-4 tumormodel[27], inCVAX, referred to as LIT in early literatures, was per-formed; the experimental designs and the outcomes are shown inTable 1. In our previous studies, inCVAX treatment resulted in a 38%survival rate until the end of the study at 120 days, with metasta-ses completely destroyed in 40 days. However, in the untreatedgroup, all rats died by 40 days with multiple metastases in the in-guinal and axillary areas. Only rats administered with the effectivephotothermal application and GC achieved long-term survival [18,21].In addition, the lack of tumor resistance observed in rats following

freezethaw lysate immunization and surgical resection revealeda low level of immunogenicity in the DMBA-4 tumor model [27].This implies that there may be strongly augmented antitumorimmune responses induced in this tumor model.

Fig. 1. Reaction of primary amino group of N-deacetylated poly-N-acetylglucosamine (dPNAG), or chitosan, with the aldehyde group of galactose yields a Schiff base whichis in equilibriumwith its Amadori product. Subsequent treatment with sodium borohydride leads to the reduction of the double bonds and the formation of stable functionalizedchitosan, or glycated chitosan (GC). Reproduced with permission from Ref. 31. Copyright 2003, Humana Press Inc.

Fig. 2. Non-invasive laser irradiation of rat tumors after the injection of inCVAXsolution.

171F. Zhou et al./Cancer Letters 359 (2015) 169177

The function of GC is to enhance the host immune response afterdirect cancer cell destruction by a selective laser photothermal in-teraction. To further test its effects, Chen et al. evaluated severaldifferent adjuvants for immunological stimulation, in combina-tion with a selective photothermal interaction, including completeFreund (CF) adjuvant, incomplete Freund (IF) adjuvant and Cory-nebacterium parvum (CP). In the treatment of DMBA-4 rat tumors,CF, IF and CP raised the cure rates from 0% to 18%, 7% and 9%, re-spectively. In comparison, GC resulted in an average of 29% long-term survival [67,68]. The results indicate that GCwasmore effectivethan other common adjuvants in inducing antitumor immunity incombination with laser treatment, leading to long-term survival oftumor-bearing rats.

Long-term antitumor effect of inCVAX

Tumor rechallenge was performed as shown in Fig. 3, to conrmthe long-term antitumor immunity induced by inCVAX [20]. Theprotective ability of induced immunity was evaluated in severalgroups that were challenged repeatedly with increased inoculation

doses of viable tumor cells. In tumor rechallenge experiments, fteenrats that had been successfully treated with inCVAX wererechallenged with 106 viable tumor cells 120 days after initial in-oculation (105 viable tumor cells); these rats showed total resistanceto the challenge, with neither primary tumors nor metastases ob-served. Eighteen naive age-matched rats were inoculated with thesame number of tumor cells for comparative purposes; they de-veloped primary and metastatic tumors and all died within 30 daysafter inoculation. After the rst rechallenge, the rats from severalexperimental groups were followed by two subsequent chal-lenges in a time interval from 1 to 5 months, with 106 viable tumorcells. The rats successfully treated by inCVAX were totally refrac-tory to all three tumor challenges [20].

Following the rechallenge study, an adoptive immunity trans-fer experiment was performed as demonstrated in Fig. 4, to furtherevaluate the induced long-term antitumor immunity in the suc-cessfully treated rats [20]. The spleen cells harvested fromsuccessfully treated tumor-bearing rats provided 100% immunityin the naive recipients. The passively protected rst cohort rats wereimmune to tumor challenge with an increased tumor dose; theirsplenocytes also prevented the establishment of tumors in the secondcohort of naive recipient rats.

After tumor rechallenge, sera were obtained and analysed fortumor selective antibodies production with histochemical assays[25,27]. Compared to sera from untreated tumor bearing rats, thesera from cured tumor-bearing rats by inCVAX contained antibod-ies that strongly bound to the plasma membrane of both living andpreserved tumor cells. In the western blot assay, two distinct bandswere observed at approximately 45 and 35 kDa using sera from suc-cessfully treated tumor-bearing rats; however, the band intensitywas much weaker using serum from the control tumor bearing rat[25,27]. These results indicate that inCVAX has produced certain an-tibodies in rats that bind or intensify the binding of specic tumorproteins.

Combination of GC with other cancer treatment modalities

As the immunoadjuvant in inCVAX, the application of GC wastested with other local tumor destruction modalities. In the treat-ment of EMT6 mammary tumor in mice, GC of 0.5% and 1.5%concentrations increased the cure rates of Photofrin-based photo-dynamic therapy (PDT) treatment from 38% to 63% and 75%,respectively [67], in this highly immunogenic tumor model. GC of0.5% concentrations increased the cure rates of carbon nanotube-based PTT treatment of EMT6-bearing mice from 44% to 100% andall the cured mice showed resistance to tumor rechallenge [22]. Inthe treatment of line 1 lung adenocarcinoma in mice, a poorly im-munogenic tumor model, a 1.67% GC solution enabled a noncurativemesosubstituted tetra (meta-hydroxy-phenyl) chlorin-based PDT(mTHPC-PDT) to cure 37% of the tumor-bearing mice [67]. In thetreatment of 4T1 mammary sarcoma in mice, another poorly im-munogenic tumor model, 1% GC solution decreased the sizes oftumor metastases when combined with high-intensity focused ul-trasound (HIFU) treatment (instead of photothermal application),and the serum from treatedmice showed signicant cytotoxic effectson 4T1 cells [69]. These results conrmed our previous studies,showing that GC is a critical component in the application of inCVAXfor the treatment of metastatic cancer.

Mechanism of inCVAX

Selective photothermal effect induced by inCVAX

Thermal effect on tumor tissueEffects of tissue temperature elevation have been well estab-

lished in terms of blood ow increases in tumor and normal tissues

Table 1Permutations and treatment parameters of different components of inCVAX fortreatment of DMBA-4 metastatic breast tumors in female rats.

Group Parameters # of rats

Laser Dye/adjuvant

Control 35*ICG injection only 0.25% ICG** 12GC injection only 1% GC** 12Laser only 2 watts; 10 min. 12Laser + ICG 2 watts; 10 min. 0.25% ICG 12Laser + GC 2 watts; 10 min. 1% GC 12ICG + GC injection 0.25% ICG/1% GC** 12Laser + ICG + GC 2 watts; 10 min. 0.25% ICG/1% GC 31*

* The data were collected from two separate experiments.** The ICG, GC, or ICG-GC solution (200 l) was injected directly to the center ofthe primary tumor.Reproduced with permission from Ref. 21. Copyright 2002American Association for Cancer Research.

Fig. 3. Schematic of anti-tumor immunity induced by inCVAX in tumor rechallenge.

172 F. Zhou et al./Cancer Letters 359 (2015) 169177

(T > 40C) [70], cellular cytotoxicity (T > 41.5 C) [71] and vasculardestruction within tumor tissue (T > 42.5 C) [72]. The rate of cellkill doubles for every 1 C increase beyond 43C [73,74]. In addi-tion, tumor tissue is more sensitive to temperature increase thannormal tissue [75].

Raising temperature in tumor tissue can cause cell death or cellstress, leading to the release of tumor antigens and the develop-ment of anti-tumor immunity [24,76]. These antigens include tumor-associated antigens, thermally induced heat shock proteins (HSPs),and a large amount of self-antigens. Antigen presenting cells (APCs),particularly dendritic cells (DCs), can capture these antigens, migrateto lymph nodes, and present the antigens to T cells to induce anti-tumor immune responses [77,78]. Specically, thermal treatmentof primary tumors can induce the release of unique tumor anti-genic peptides that are bound to HSPs [79,80], which in turn canenhance immune responses by facilitating peptide presentation toCD8+ T cells [81].

Photothermal interaction with tumor tissueAn important aspect of photothermal interaction through laser

ablation is that a thermal gradient is generated inside the tissue.Temperature-dependent biological reactions are crucial in thermaltherapy for cancer treatment. When the temperature rises from 43to 100 C, tumor cells undergo coagulative necrosis, but reversiblechanges in tissue occurs only with temperature below 60C [82].In addition, tumor cells swell and break into fragments allowingantigen release, which can induce tumor-specic immune re-sponse in the host.

The selective photothermal effect of inCVAX can thus induce ahigh temperature increase in the target tissue [35,36]. Chen et al.applied magnetic resonance thermometry (MRT) to monitor laser-induced high thermal gradient temperature distribution of targettissue with high spatial resolution. Fig. 5 shows temperature gra-dients in liver tissue acquired by MRT. The temperature mappingresults showed that the selective laser photothermal effect could

Fig. 4. Schematic of anti-tumor immunity induced by inCVAX demonstrated by adoptive immunity transfer after treatment.

Fig. 5. Two-dimensional temperature distributions in liver tissue immediately after interstitial laser irradiation (1.5 W for 10 min) at different locations from the center ofthe tumor (left panel) to the outer edge of the tumor (right panel). Reproduced with permission from Ref. 83. Copyright 2011 Society of Photo Optical Instrumentation Engineers.

173F. Zhou et al./Cancer Letters 359 (2015) 169177

increase tumor temperatures in a range of 1025 C [84]. In a studyby Le et al., different power settings (1.0, 1.25, and 1.5 W) wereapplied for 10min in a rat tumor, demonstrating an increase in tem-perature that ranged from 8 C to 15 C based on different laser powerand distance from the injected optical ber [83].

Different tissue temperatures can induce various immune re-sponses in the host. Within the temperature range of 4060 C,photothermal treatment may induce a high level of cell apoptosisin the tumor tissue, which can create a large antigen load for thegeneration of antitumor immunity [22,85]. In therapeutic terms,the immunogenicity of apoptosis is preferable for immunologicalstimulation compared to necrosis [86]. The released antigens includetumor associated antigens and a large amount of self-antigens,which can be recognized and captured by APCs. Carrying the an-tigens, matured APCs migrate to lymph nodes, where they presentthese antigens to T cells, thus activating cytotoxic T-lymphocytes(CTLs). This process is thus intended to induce a specic cell-mediated antitumor immune response that is effective againsttumor cell antigens [22].

As a chaperone, HSPs work as an endogenous danger signal inthe immune surveillance system; extracellular HSPpeptidecomplexes released from damaged cells can promote thecross-presentation of HSP-bound peptide antigens to major histo-compatibility complex (MHC) class I molecules in DCs, leading toecient induction of antigen-specic CTL [8789]. Treatment byinCVAX could enhance HSP expression in the tumor cells, andexternalization of the apoptotic cells, which could be recognizedbyAPCs through toll like receptor (TLRs), followedby cytokine releasevia Myd88 and NF-kB signaling pathways [85,86,90]. HSPs on thesurface of apoptotic cells could connect the APCs and the tumor cellsas well as enhance the antigen acquisition and activation of APCs,which could initiate a specic antitumor immunity. The roles of HSPsin stimulating both innate immunity and adaptive immunity canexplain at least in part themolecularmechanism bywhich thermalstress bolsters the host immune system [91,92].

Properties of GC

GC works as an immunological stimulant in inCVAX that is in-tended to further enhance the antitumor immune response [18,21].With the direct immunostimulation properties, GC could stimu-late macrophage activation, as measured by TNF- secretion andNO production [85], and enhance CD80 expression that indicatesthe DCmaturation [22]. Studies on the direct interaction with tumorcells showed that GC has no cytotoxicity on tumor cells, but coulddecrease the expression of Twist-1 and Slug, proto-oncogenes com-monly implicated in metastasis [69]. When GC alone is used as atreatment agent on tumor bearing mice, the metastases can be de-creased, and survival time can be extended; however, the survivalrate of animals has not been improved [21,22,69], demonstratingthe adjuvant nature of GC.

Although photothermal application could release a large amountof different antigens for the generation of antitumor immunity, theresponse of the host immune system is limited [22,85]. When com-bined with GC, TNF- secretion of macrophage and CD80 expressionon DCs could be signicantly enhanced [22,85], so could be theantigen acquisition and antigen presentation by APCs [22], thus ac-tivating CTLs [22,85]. These results showed that tumor antigenrelease by laser thermal therapy and the GC enhancement for thegeneration of antitumor immunity were both crucial.

It is not fully understoodhowGC functions as an immunoadjuvant,but it is possible that the carbohydrate structure [31] plays an im-portant role. It has been reported that certain receptors on theimmune cells can preferentially bind to the carbohydrate struc-ture, such as TLRs and the C-type lectin family [93]. These TLRs andC-type lectins have been associated with antigen uptake and may

be important for the migration of DCs and their interactions withlymphocytes [94].

Synergistic effect of inCVAX

Thermal interactionmay induce a local immune-stimulating effectand released tumor antigen could lead to the development ofthermal-based autologous vaccines [95,96]. However, as men-tioned above, tumor debris is apparently not sucient in inducinga potent anti-tumor response [97]. Therefore, additionalimmunological intervention is required to invoke the immune systemto achieve an effective and protective immune response against re-sidual tumor cells.

The principal idea behind inCVAX is an induced systemic, tumor-specic immune response through 1) laser irradiation to selectivelyinterrupt tumor cells, releasing tumor antigens and 2) local admin-istration of GC to activate immune cells, targeting specic tumorcells. In fact, its mechanism, using the treated tumor cells as sourcesof tumor antigens, can be considered as in situ autologous whole-cell cancer vaccination. The schematic of antitumor immune responseinduced by inCVAX is shown in Fig. 6. Inducing systemic anti-tumor immune responses through local, selective photothermalinteraction and active immunological stimulation with the uniqueproperties of GC, inCVAXmay be a promising approach against meta-static cancers [28,29].

Novel features of inCVAX

Individualized therapy

As an autologous vaccine-like approach using patients ownwholetumor cells as the source of antigens, the vaccine has access to thewhole spectrum of unique and shared antigens expressed by thetargeted tumor in each individual patient. Furthermore, inCVAX re-quires no ex vivo selection and processing of patient antigens, and,with the unique properties of GC, may overcome other obstacles,including poor immunogenicity or different antigen expressions.

Minimally invasive

inCVAX is a local, minimally invasive modality that, based onpreliminary studies [29], mainly appears to induce local discom-fort; the most common adverse effects have been shown to be localpain, mild swelling, and blisters at the treatment sites. While ad-ditional clinical trials are necessary, to date, no grade 3 or 4 adverseevents were reported in the initial clinical trials [29], which sug-gests that inCVAX may have the potential to improve the qualityof patients lives.

The local injection of GC is a simple procedure. The techniqueof laser irradiation is intended to be straightforward for use by phy-sicians. The equipment for inCVAX is small and can be stored andtransported in a briefcase, making it suitable for professional clin-ical centers as well as small clinics in rural areas and in developingcountries.

Prospect

The effect of inCVAX has been investigated through cellular andanimal studies. Furthermore, inCVAX has been administered in initialclinical trials for the treatment of patients with metastatic breastcancer, many of whom have failed other available modalities, withpromising outcomes [28,29]. Specically, the data indicate thatinCVAX may be capable of reducing or eliminating treated primarybreast tumors and untreated metastases, as well as prolongingpatient survival [29]. We intend to further develop inCVAX so thatit can be administered to other solid tumors endoscopically, such

174 F. Zhou et al./Cancer Letters 359 (2015) 169177

as cervical cancer and colon cancers. As a new technology underdevelopment, much remains to be understood, but with furtherstudies of the fundamental mechanism of inCVAX, and with con-tinued clinical trials, it is believed that inCVAX may benet cancerpatients in the future, particularly those who havemetastatic tumorsand have failed conventional therapies.

The concept of inCVAX, using the synergistic effect of acute, localtumor cell destruction and prolonged, systemic immunological stim-ulation, may potentially be used with many cancer treatmentmodalities. For example, radiation therapy, a well-established localtreatment method, may be an excellent candidate for concomi-tant use with inCVAX or GC. Radiation, through its direct, precise,and effective interaction, interrupts DNAs in tumor cells, leading totumor cell death, mainly through apoptotic pathways. These deador dying cells could be ideal sources of tumor antigens. Therefore,ionizing radiation, in combination with the in situ application of GC,holds promise for the treatment of metastatic tumors. Further in-vestigation on the effects of the combination of radiation and GCis ongoing.

Acknowledgments

This study was supported in part by a grant from the US Na-tional Institutes of Health (R21 EB0155091-01), as well as by grantsfrom National Natural Science Foundation of China (no. 81472796;81000994), Beijing Natural Science Foundation (4153064), andBeijing Nova Program (Z131107000413104).

Conict of interest

The authors declare no conict of interest.

References

[1] L.A. Liotta, W.G. Stetler-Stevenson, Principles of molecular cell biology of cancer:cancer metastasis, in: V.T. DeVita, S. Hellman, S.A. Rosenberg (Eds.), Cancer:Principles & Practice of Oncology, J.B. Lippincott Co., Philadelphia, PA., 1993,pp. 134149.

[2] G.P. Gupta, J. Massagu, Cancer metastasis: building a framework, Cell 127 (4)(2006) 679695.

[3] S. Kraljevic Pavelic, M. Sedic, H. Bosnjak, S. Spaventi, K. Pavelic, Metastasis: newperspectives on an old problem, Mol. Cancer 10 (2011) 22.

[4] J.N. Blattman, P.D. Greenberg, Cancer immunotherapy: a treatment for themasses, Science 305 (5681) (2004) 200205.

[5] O.J. Finn, Cancer immunology, N. Engl. J. Med. 358 (25) (2008) 27042715.[6] S.A. Rosenberg, Progress in human tumour immunology and immunotherapy,

Nature 411 (6835) (2001) 380384.[7] I. Mellman, G. Coukos, G. Dranoff, Cancer immunotherapy comes of age, Nature

480 (7378) (2011) 480489.[8] A. Carpi, A. Nicolini, A. Antonelli, P. Ferrari, G. Rossi, Cytokines in the

management of high risk or advanced breast cancer: an update and expectation,Curr. Cancer Drug Targets 9 (8) (2009) 888903.

[9] S. Kim-Schulze, B. Taback, H.L. Kaufman, Cytokine therapy for cancer, Surg.Oncol. Clin. N. Am. 16 (4) (2007) 793818.

[10] E.A. Eksioglu, S. Eisen, V. Reddy, Dendritic cells as therapeutic agents againstcancer, Front. Biosci. 15 (2010) 321347.

[11] P. Kalinski, J. Urban, R. Narang, E. Berk, E. Wieckowski, R. Muthuswamy, Dendriticcell-based therapeutic cancer vaccines: what we have andwhat we need, FutureOncol. 5 (3) (2009) 379390.

[12] A.P. Castano, P. Mroz, M.R. Hamblin, Photodynamic therapy and anti-tumourimmunity, Nat. Rev. Cancer 6 (7) (2006) 535545.

[13] H.G. Zhang, K. Mehta, P. Cohen, C. Guha, Hyperthermia on immune regulation:a temperatures story, Cancer Lett. 271 (2) (2008) 191204.

[14] M. Vanneman, G. Dranoff, Combining immunotherapy and targeted therapiesin cancer treatment, Nat. Rev. Cancer 12 (4) (2012) 237251.

[15] M. Ahmad, R.C. Rees, S.A. Ali, Escape from immunotherapy: possiblemechanismsthat inuence tumor regression/progression, Cancer Immunol. Immunother.53 (10) (2004) 844854.

[16] W. Zou, Immunosuppressive networks in the tumour environment and theirtherapeutic relevance, Nat. Rev. Cancer 5 (4) (2005) 263274.

[17] C.L. Zindl, D.D. Chaplin, Immunology: tumor immune evasion, Science 328(5979) (2010) 697698.

Fig. 6. Schematic of the systemic antitumor immune response induced by inCVAX.

175F. Zhou et al./Cancer Letters 359 (2015) 169177

[18] W.R. Chen, R.L. Adams, R. Carubelli, R.E. Nordquist, Laser-photosensitizer assistedimmunotherapy: a novel modality in cancer treatment, Cancer Lett. 115 (1)(1997) 2530.

[19] M. Mitsunaga, M. Ogawa, N. Kosaka, L.T. Rosenblum, P.L. Choyke, H. Kobayashi,Cancer cell-selective in vivo near infrared photoimmunotherapy targetingspecic membrane molecules, Nat. Med. 17 (12) (2012) 16851691.

[20] W.R. Chen, A.K. Singhal, H. Liu, R.E. Nordquist, Antitumor immunity inducedby laser immunotherapy and its adoptive transfer, Cancer Res. 61 (2) (2001)459461.

[21] W.R. Chen, H. Liu, J.W. Ritchey, K.E. Bartels, M.D. Lucroy, R.E. Nordquist, Effectof different components of laser immunotherapy in treatment of metastatictumors in rats, Cancer Res. 62 (15) (2002) 42954299.

[22] F. Zhou, S. Wu, S. Song, W.R. Chen, D.E. Resasco, D. Xing, Antitumorimmunologically modied carbon nanotubes for photothermal therapy,Biomaterals 33 (11) (2012) 32353242.

[23] T. Hode, X. Li, M.F. Naylor, L. Hode, P. Jenkins, G. Ferrel, et al., Laserimmunotherapy, in: M.R. Hamblin, S.K. Sharma (Eds.), Handbook ofPhotomedicine, Taylor & Francis, New York, 2013.

[24] M.H. den Brok, R.P. Sutmuller, R. van der Voort, E.J. Bennink, C.G. Figdor, T.J.Ruers, et al., In situ tumor ablation creates an antigen source for the generationof antitumor immunity, Cancer Res. 64 (11) (2004) 40244029.

[25] W.R. Chen, W.-G. Zhu, J.R. Dynlacht, H. Liu, R.E. Nordquist, Long-term tumorresistance induced by laser photo-immunotherapy, Int. J. Cancer 81 (5) (1999)808812.

[26] W.R. Chen, H. Liu, J.A. Nordquist, R.E. Nordquist, Tumor cell damage andleukocyte inltration after laser immunotherapy treatment, Lasers Med. Sci.15 (1) (2000) 4348.

[27] W.R. Chen, S.W. Jeong, M.D. Lucroy, R.F. Wolf, E.W. Howard, H. Liu, et al., Inducedanti-tumor immunity against DMBA-4 metastatic mammary tumors in ratsusing a novel approach, Int. J. Cancer 107 (6) (2003) 10531057.

[28] X. Li, M.F. Naylor, R.E. Nordquist, T.T. Kent, H.C. Anthony, M. Cynthia, et al., Insitu photoimmunotherapy for late-stage melanoma patients: a preliminarystudy, Cancer Biol. Ther. 10 (11) (2010) 10771214.

[29] X. Li, G.L. Ferrel, M.C. Guerra, T. Hode, J.A. Lunn, O. Adalsteinsson, et al.,Preliminary safety and ecacy results of laser immunotherapy for the treatmentof metastatic breast cancer patients, Photochem. Photobiol. Sci. 10 (5) (2011)817821.

[30] J. Copier, A. Dalgleish, Overview of tumor cell-based vaccines, Int. Rev. Immunol.25 (56) (2006) 297319.

[31] W.R. Chen, R. Carubelli, H. Liu, R.E. Nordquist, Laser immunotherapy: a noveltreatment modality for metastatic tumors, Mol. Biotechnol. 25 (1) (2003) 3743.

[32] W.R. Chen, R.L. Adams, S. Heaton, D.T. Dickey, K.E. Bartels, R.E. Nordquist,Chromophore-enhanced laser-tumor tissue photothermal interaction using808 nm diode laser, Cancer Lett. 88 (1) (1995) 1519.

[33] C. Liang, S. Diao, C. Wang, H. Gong, T. Liu, G. Hong, et al., Tumor metastasisinhibition by imaging-guided photothermal therapy with single-walled carbonnanotubes, Adv. Mater. 26 (32) (2014) 56465652.

[34] M.N. Iizuka, I.A. Vitkin, M.C. Kolios, M.D. Sherar, The effects of dynamic opticalproperties during interstitial laser photocoagulation, Phys. Med. Biol. 45 (5)(2000) 13351357.

[35] W.R. Chen, R.L. Adams, K.E. Bartels, R.E. Nordquist, Chromophore-enhanced invivo tumor cell destruction using an 808-nm diode laser, Cancer Lett. 94 (2)(1995) 125131.

[36] W.R. Chen, R.L. Adams, A.K. Higgins, K.E. Bartels, R.E. Nordquist, Photothermaleffects on murine mammary tumors using indocyanine green and an 808-nmdiode laser: an in vivo ecacy study, Cancer Lett. 98 (2) (1996) 169173.

[37] X.H. Huang, I.H. El-Sayed, W. Qian, M.A. El-Sayed, Cancer cell imaging andphotothermal therapy in the near-infrared region by using gold nanorods, J.Am. Chem. Soc. 128 (6) (2006) 21152120.

[38] F. Zhou, D. Xing, Z. Ou, B. Wu, D.E. Resasco, W.R. Chen, Cancer photothermaltherapy in the near-infrared region by using single-walled carbon nanotubes,J. Biomed. Opt. 14 (2009) 021009.

[39] K. Yang, S. Zhang, G. Zhang, X. Sun, S.T. Lee, Z. Liu, Graphene in mice: ultrahighin vivo tumor uptake and ecient photothermal therapy, Nano Lett. 10 (9)(2010) 33183323.

[40] L. Cheng, K. Yang, Y. Li, J. Chen, C. Wang, M. Shao, et al., Facile preparation ofmultifunctional upconversion nanoprobes for multimodal imaging and dual-targeted photothermal therapy, Angew Chem Int Edit 50 (32) (2011) 73857390.

[41] F. Zhou, S. Wu, B. Wu, W.R. Chen, D. Xing, Mitochondria-targeting single-walledcarbon nanotubes for cancer photothermal therapy, Small 7 (19) (2011)27272735.

[42] D. Jaque, L. Martnez Maestro, B. del Rosal, P. Haro-Gonzalez, A. Benayas, J.L.Plaza, et al., Nanoparticles for photothermal therapies, Nanoscale 6 (2014)94949530.

[43] X. Li, M. Min, N. Du, Y. Gu, T. Hode, M. Naylor, et al., Chitin, chitosan, and glycatedchitosan regulate immune responses: the novel adjuvants for cancer vaccine,Clin. Dev. Immunol. 2013 (2013) 387023.

[44] E. Montomoli, S. Piccirella, B. Khadang, E. Mennitto, R. Camerini, A. De Rosa,Current adjuvants and new perspectives in vaccine formulation, Expert Rev.Vaccines 10 (7) (2011) 10531061.

[45] F.R. Vogel, M.F. Powell, A compendium of vaccine adjuvants and excipients,Pharm. Biotechnol. 6 (1995) 141228.

[46] M. Yang, Y. Yan, M. Fang, M. Wan, X. Wu, X. Zhang, et al., MF59 formulated withCpG ODN as a potent adjuvant of recombinant HSP65-MUC1 for inducinganti-MUC1(+) tumor immunity in mice, Int. Immunopharmacol. 13 (4) (2012)408416.

[47] C.W. Cluff, Monophosphoryl lipid A (MPL) as an adjuvant for anti-cancervaccines: clinical results, Adv. Exp. Med. Biol. 667 (2010) 111123.

[48] R.K. Gupta, G.R. Siber, Adjuvants for human vaccines-current status, problemsand future prospects, Vaccine 13 (14) (1995) 12631276.

[49] D.F. Hoft, V. Brusic, I.G. Sakala, Optimizing vaccine development, Cell. Microbiol.13 (7) (2011) 934942.

[50] J. Cebon, Cancer vaccines: where are we going?, Asia Pac J Clin Oncol. S1 (2010)S9S15.

[51] J. Schlom, Therapeutic cancer vaccines: current status and moving forward, J.Natl Cancer Inst. 104 (8) (2012) 599613.

[52] G. Leroux-Roels, Unmet needs in modern vaccinology: adjuvants to improvethe immune response, Vaccine 28 (S3) (2010) C25C36.

[53] C. Mesa, L.E. Fernndez, Challenges facing adjuvants for cancer immunotherapy,Immunol. Cell Biol. 82 (2004) 644650.

[54] V.E. Schijns, Mechanisms of vaccine adjuvant activity: initiation and regulationof immune responses by vaccine adjuvants, Vaccine 21 (910) (2003) 829831.

[55] A.S. McKee, M.K.L. MacLeod, J.W. Kappler, P. Marrack, Immune mechanisms ofprotection: can adjuvants rise to the challenge?, BMC Biol. 8 (2010) 37.

[56] M. Sugano, S. Watanabe, A. Kishi, M. Izume, A. Ohtakara, Hypocholesterolemicaction of chitosans with different viscosity in rats, Lipids 23 (3) (1988) 187191.

[57] Y.C. Chung, J.Y. Yeh, C.F. Tsai, Antibacterial characteristics and activity ofwater-soluble chitosan derivatives prepared by the Maillard reaction, Molecules16 (10) (2011) 85048514.

[58] K. Suzuki, Y. Okawa, K. Hashimoto, S. Suzuki, M. Suzuki, Protecting effect of chitinand chitosan on experimentally induced murine candidiasis, Microbiol.Immunol. 28 (8) (1984) 903912.

[59] V. Jarmila, E. Vavrkov, Chitosan derivatives with antimicrobial, antitumourand antioxidant activities a review, Curr. Pharm. Des. 17 (32) (2011) 35963607.

[60] K. Nishimura, S. Nishimura, N. Nishi, I. Saiki, S. Tokura, I. Azuma, Immunologicalactivity of chitin and its derivatives, Vaccine 2 (1) (1984) 9399.

[61] I.M. van der Lubben, J.C. Verhoef, G. Borchard, H.E. Junginger, Chitosan formucosal vaccination, Adv. Drug Deliv. Rev. 52 (2) (2001) 139144.

[62] R.C. Read, S.C. Naylor, C.W. Potter, J. Bond, I. Jabbal-Gill, A. Fisher, et al., Effectivenasal inuenza vaccine delivery using chitosan, Vaccine 23 (35) (2005)43674374.

[63] E.A. McNeela, I. Jabbal-Gill, L. Illum, M. Pizza, R. Rappuoli, A. Podda, et al.,Intranasal immunization with genetically detoxied diphtheria toxin inducesT cell responses in humans: enhancement of Th2 responses and toxin-neutralizing antibodies by formulation with chitosan, Vaccine 22 (8) (2004)909914.

[64] D.A. Zaharoff, C.J. Rogers, K.W. Hance, J. Schlom, J.W. Greiner, Chitosan solutionenhances both humoral and cell-mediated immune responses to subcutaneousvaccination, Vaccine 25 (11) (2007) 20852094.

[65] F. Xu, H. Liu, X. Wu, H. Jiang, R.E. Nordquist, W.R. Chen, Measurement of x-rayattenuation coecients of aqueous solutions of indocyanine green and glycatedchitosan, Med. Phys. 26 (7) (1999) 13711374.

[66] W.R. Chen, H. Liu, W. Yue, J. Wang, R.E. Nordquist, Dynamically observingintratumor injection of laser-absorbing dye and immunoadjuvant using digitalx-ray imaging technique, Opt. Eng. 40 (7) (2001) 12491254.

[67] W.R. Chen, M. Korbelik, K.E. Bartels, H. Liu, J. Sun, R.E. Nordquist, Enhancementof laser cancer treatment by a chitosan-derived immunoadjuvant, Photochem.Photobiol. 81 (1) (2005) 190195.

[68] W.R. Chen, Z. Huang, M. Korbelik, R.E. Nordquist, H. Liu, Photoimmunotherapyfor cancer treatment, J. Environ. Pathol. Toxicol. Oncol. 25 (12) (2006) 281291.

[69] Y.L. Chen, C.Y. Wang, F.Y. Yang, B.S. Wang, J.Y. Chen, L.T. Lin, et al., Synergisticeffects of glycated chitosan with high-intensity focused ultrasound onsuppression of metastases in a syngeneic breast tumor model, Cell Death Dis.5 (2014) e1178.

[70] H.S. Reinhold, B. Endrich, Tumor microcirculation as a target for hyperthermia,Int. J. Hyperthermia 2 (2) (1986) 111137.

[71] A.J. Thistlethwaite, D.B. Leeper, D.J. Moylan, R.E. Nerlinger, pH distribution inhuman tumors, Int. J. Radiat. Oncol. Biol. Phys. 11 (9) (1985) 16471652.

[72] W.C. Dewey, L.E. Hopwood, S.A. Sapareto, L.E. Gerweck, Cellular responses tocombinations of hyperthermia and radiation, Radiology 123 (2) (1977) 463474.

[73] S.A. Sapareto, W.C. Dewey, Thermal dose determination in cancer therapy, Int.J. Radiat. Oncol. Biol. Phys. 10 (6) (1984) 787800.

[74] S.B. Field, C.C. Morris, The relationship between heating time and temperature:its relevance to clinical hyperthermia, Radiother. Oncol. 1 (2) (1983) 179186.

[75] L.J. Anghileri, J. Robert, Hyperthermia in Cancer Treatment, CRC Press, BocaRaton, FL, 1986.

[76] T.J. Yoon, J.Y. Kim, H. Kim, C. Hong, H. Lee, C.K. Lee, et al., Anti-tumorimmunostimulatory effect of heat-killed tumor cells, Exp. Mol. Med. 40 (1)(2008) 130144.

[77] E. Jager, D. Jager, A. Knuth, Antigen-specic immunotherapy and cancer vaccines,Int. J. Cancer 106 (6) (2003) 817820.

[78] A. Mukhopadhaya, J. Mendecki, X. Dong, L. Liu, S. Kalnicki, M. Garg, et al.,Localized hyperthermia combined with intratumoral dendritic cells inducessystemic antitumor immunity, Cancer Res. 67 (16) (2007) 77987806.

[79] T. Chen, J. Guo, C. Han, M. Yang, X. Cao, Heat shock protein 70, released fromheat-stressed tumor cells, initiates antitumor immunity by inducing tumor cellchemokine production and activating dendritic cells via TLR4 pathway, J.Immunol. 182 (3) (2009) 14491459.

[80] M.N. Rylander, Y. Feng, J. Bass, K.R. Diller, Heat shock protein expression andinjury optimization for laser therapy design, Lasers Surg. Med. 39 (9) (2007)731746.

176 F. Zhou et al./Cancer Letters 359 (2015) 169177

[81] Z. Prohaszka, Chaperones as part of immune networks, Adv. Exp. Med. Biol. 594(2007) 159166.

[82] M.R. Hamblin, Y.Y. Huang, Handbook of Photomedicine, CRC Press, New York,2013.

[83] K. Le, X. Li, D. Figueroa, R.A. Towner, P. Garteiser, D. Saunders, et al., Assessmentof thermal effects of interstitial laser phototherapy on mammary tumors usingproton resonance frequency method, J. Biomed. Opt. 16 (12) (2011) 128001.

[84] Y. Chen, S.C. Gnyawali, F. Wu, H. Liu, Y.A. Tesiram, A. Abbott, et al., Magneticresonance imaging guidance for laser photothermal therapy, J. Biomed. Opt.13 (4) (2008) 044033.

[85] F. Zhou, S. Song, W.R. Chen, D. Xing, Immunostimulatory properties of glycatedchitosan, J Xray Sci Technol. 19 (2) (2011) 285292.

[86] F. Zhou, D. Xing, W.R. Chen, Regulation of HSP70 on activating macrophagesusing PDT-induced apoptotic cells, Int. J. Cancer 125 (6) (2009) 13801389.

[87] J. Gong, Y. Zhang, J. Durfee, D. Weng, C. Liu, S. Koido, et al., A heat shock protein70-based vaccine with enhanced immunogenicity for clinical use, J. Immunol.184 (1) (2010) 488496.

[88] B.H. Segal, X.Y. Wang, C.G. Dennis, R. Youn, E.A. Repasky, M.H. Manjili, et al.,Heat shock proteins as vaccine adjuvants in infections and cancer, Drug Discov.Today 11 (1112) (2006) 534540.

[89] M. Nishikawa, S. Takemoto, Y. Takakura, Heat shock protein derivatives fordelivery of antigens to antigen presenting cells, Int. J. Pharm. 354 (12) (2008)2327.

[90] S. Song, F. Zhou, W.R. Chen, D. Xing, PDT-induced HSP70 externalizationup-regulates NO production via TLR2 signal pathway in macrophages, FEBS Lett.587 (2) (2013) 128135.

[91] T. Torigoe, Y. Tamura, N. Sato, Heat shock proteins and immunity: applicationof hyperthermia for immunomodulation, Int. J. Hyperthermia 25 (8) (2009)610616.

[92] W.L. Yang, D.G. Nair, R. Makizumi, G. Gallos, X. Ye, R.R. Sharma, et al., Heat shockprotein 70 is induced in mouse human colon tumor xenografts after sublethalradiofrequency ablation, Ann. Surg. Oncol. 11 (4) (2004) 399406.

[93] F. Sallusto, M. Cella, C. Danieli, A. Lanzavecchia, Dendritic cells usemacropinocytosis and the mannose receptor to concentrate macromoleculesin the major histocompatibility complex class II compartment: downregulationby cytokines and bacterial products, J. Exp. Med. 182 (2) (1995) 389400.

[94] C.G. Figdor, Y. van Kooyk, G.J. Adema, C-type lectin receptors on dendritic cellsand Langerhan cells, Nat. Rev. Immunol. 2 (2) (2002) 7784.

[95] P.H. Mller, K. Ivarsson, U. Stenram, M. Radnell, K.-G. Tranberg, Comparisonbetween interstitial laser thermotherapy and excision of an adenocarcinomatransplanted into rat liver, Br. J. Cancer 77 (11) (1998) 18841892.

[96] R.F. Sanchez-Ortiz, N. Tannir, K. Ahrar, C.G. Wood, Spontaneous regression ofpulmonary metastases from renal cell carcinoma after radio frequency ablationof primary tumor: an in situ tumor vaccine?, J. Urol. 170 (1) (2003) 178179.

[97] E. Jager, D. Jager, A. Knuth, Antigen-specic immunotherapy and cancer vaccines,Int. J. Cancer 106 (6) (2003) 817820.

177F. Zhou et al./Cancer Letters 359 (2015) 169177

InCVAX A novel strategy for treatment of late-stage, metastatic cancers through photoimmunotherapy induced tumor-specific immunity Introduction Components of inCVAX Photothermal therapy (PTT) Immunoadjuvant Clinical application of adjuvants N-dihydro-galacto-chitosan (GC) Effects of inCVAX Effects of inCVAX on animal tumor model Long-term antitumor effect of inCVAX Combination of GC with other cancer treatment modalities Mechanism of inCVAX Selective photothermal effect induced by inCVAX Thermal effect on tumor tissue Photothermal interaction with tumor tissue Properties of GC Synergistic effect of inCVAX Novel features of inCVAX Individualized therapy Minimally invasive Prospect Acknowledgments Conflict of interest References