Different Behavioral Patterns of SARS-COV-2 in Patients with Various Types of Cancers: A Role for Chronic In�ammation Induced by Macrophages

SARS-CoV-2 is a single strand RNA virus that causes ongoing pneumonia-like and severe respiratory disease. This disease is very important in immunosuppressed patients. Since not many studies have been done in people with cancer as high risk group, in this study we examined the different behaviors of this virus in different types of cancer. In this study, patients with a variety of solid and hematologic cancers were screened for SARS-CoV-2. The rates of infection, the rate of mortality, in patients with various cancers, as well as in the families of these patients have been studied. In this study, it was found that the prevalence of SARS-CoV-2 in patients with solid cancers was signi�cantly lower than hematological cancers. Also, the mortality rate in hematological cancers was higher than in solid cancers (P<0.05). Assessments the families of these patients, it was found that patients with solid cancers were also less infected than their family members (P<0.05). Researchers of this study attributed these results to the chronic in�ammation and M1 macrophages in these patients. It is essential that additional studies are conducted, especially in terms of biological function.


Introduction
Coronavirus disease of 2019  is an infectious disease caused by a member of the coronavirus family (single-strand RNA viruses) named SARS-COV-2 that infects humans.It was reported for the rst time in Wuhan, China in December 2019 (Surveillances, 2020;Wang et al., 2020b;Zhou et al., 2020) and is currently spreading over more than 150 countries around the world (Tavakoli et al., 2020;Youse et al., 2020).SARS-COV-2 causes a broad spectrum of clinical manifestations and most infected people show mild to moderate respiratory symptoms and recover without any special treatment (Bai et al., 2020;Cascella et al., 2020;Recalcati, 2020;Xu et al., 2020;Zhou et al., 2020).But, in high-risk patients with underlying diseases such as diabetes (Gupta et al., 2020), cardiovascular diseases (Chen et al., 2020;Zheng et al., 2020b), cancer (Tian et al., 2020;Wang and Zhang, 2020;Xia et al., 2020), and chronic respiratory diseases (Halpin et al., 2020;Jordan et al., 2020) serious forms of the disease and even in some cases death are observed.The increased human-to-human transmission rate is the main characteristic of the new coronavirus that consequently led to the pandemic spread of COVID-19 (Bai et al., 2020;Kucharski et al., 2020).Transmission of the disease occurs through infected droplets or discharges generated while speaking, sneezing, or coughing by the infected individuals (Qian et al., 2020).Prevention practice is applicable by washing hands, using an alcohol-based rub regularly and not touching the face (Hellewell et al., 2020;Wang et al., 2020a).Although countless clinical trials are ongoing to evaluate the potential e cacy of different treatments in COIVID-19 cases, there is no speci c therapeutics or vaccines available yet (Cascella et al., 2020;Youse et al., 2020).The knowledge regarding lymphocyte subsets and the consequence of immune responses in patients with this disease are very limited.Cancer is considered as a very important risk factor for COVID-19 and based on many previous works; the rate of COVID-19 in patients with cancer has been signi cantly higher than healthy individuals (Burki, 2020;Cortiula et al., 2020;Liang et al., 2020).Also, most studies suggest that patients with cancer are more prone to develop the new coronavirus disease than others (Wang and Zhang, 2020).
However, our information regarding the incidence and severity of COVID-19 disease in various types of cancer is presently very little and more studies are needed to show the relationship between different types of cancer and incidence of COVID-19.Therefore, this study was designed and conducted to evaluate the incidence of COVID-19 in patients with different types of cancer.

Sampling and patients
This retrospective study with the approval number of 99-0213a was carried out on patients with cancer who were referred to the different cancer centers in Tehran, Iran, from November 2019 to May 2020.
Hospitalized cancer patients were previously diagnosed by laboratory and imaging techniques.was clinically diagnosed at the rst step according to the World Health Organization (WHO) protocols with different manifestations including more common symptoms such as dry cough, fever, tiredness; less common symptoms such as sore throat, diarrhea, conjunctivitis, loss of taste or smell, headache, discoloration of ngers, toes, or skin rashes, aches and pains as less common; and serious symptoms such as chest pain or pressure, di culties in breathing or shortness of breath and loss of speech or movement.Then, patients with symptoms detected through the questionnaire results were checked by lung CT scan and Real-Time PCR tests.

COVID-19 real-time RT-PCR assays
All samples were handled according to laboratory biosafety guidelines of WHO for COVID-19 and RNAs were extracted using a QIAamp viral RNA extraction mini kit (QIAGEN, Hilden, Germany) according to the manufacturer's instructions.250 µl of each respiratory tract specimen was used for RNA extraction and the extracted RNAs were kept at -80°C until use.

Data analysis
The Shapiro-Wilk test was used to assess the normality of the data distribution.All data were analyzed with GraphPad Prism software (GraphPad Software, Inc.) and Fisher's exact test was used to compare the results.P-value of < 0.05 was considered statistically signi cant.
The Demographic and baseline clinical characteristics of COVID-19 patients in solid cancers are described in Table 1.Among cancer patients, Breast, Gastric, and Lung cancers were the most prevalent forms of cancer (P < 0.05).Also, most of the patients were in stage I or II of their disease.Based on clinical manifestations, 86.74% (314/362) of patients with solid tumors showed no clinical signs and only 5 (1.38%) patients indicated severe clinical manifestations of COVID-19 that these differences were statically signi cant (P < 0.001).Results of the con rmatory tests indicated that 9 (2.48%) patients with different types of solid tumors were positive for COVID-19, whereas, 46 (12.70%) patients had close contact with COVID-19 patients in their family and were closely exposed (Fig. 1).Among different types of malignancies, DLBL (Diffuse large B-cell lymphoma) was the most frequent type (31.10%), followed by AML (25.35%) and ALL cancers (19.61%).The morbidity rate of COVID-19 in hematologic malignancies (54.96%) was signi cantly higher than the solid tumors (P = 0.004).Also, in patients with hematologic malignancies, the rates of clinical manifestations (mild (26.13%) and severe (19.61%)) were higher in comparison to the solid type.
Most of the patients in both groups of malignancies (hematologic and solid) had a history of antitumor chemotherapy and were divided into two main clusters according to the received treatments: 1) patients with chemotherapy during the last 1-4 weeks, 2) patients with chemotherapy during 1-6 months.Clinical manifestations of these two clusters are shown in Table 3. Patients with clinical manifestations of COVID-19 who had received chemotherapy during the last 4 weeks were examined using CT-scan and Real-Time PCR tests and our results showed that the sensitivity of the real-time PCR test was signi cantly higher than the CT-scan.

Discussion
In the current study, the clinical characteristics of 571 cancer patients (with solid or hematologic cancer) were described.Cancer patients who were referred to the cancer research centers during the past 8 months were primarily examined using a questionnaire.Patients were divided into different clinical categories based on the clinical signs (mild and severe) recorded in the Iranian Coronavirus Control Organization.Patients with clinical symptoms of the new coronavirus disease and chemotherapy in the last 4 weeks were re-examined using con rmatory tests for COVID-19 (CT-scan and Real-Time PCR).Also, all family members of patients with cancer were examined for COVID-19 to determine if the patients were at risk for the disease.The results of our study revealed that the rates of mortality, manifestations, and especially severe manifestations of COVID-19 in hematological malignancies were signi cantly higher than solid cancers and patients with solid cancers showed more resistance to the disease.Interestingly, patients with solid tumors who had close contact with SARS-COV-2 infected individuals in their families were resistant and not infected.This resistance existed even after chemotherapy.Patients with different types of hematological malignancies showed severe symptoms of the new coronavirus disease after chemotherapy, and some even died.The mortality rate of COVID-19 in the group of solid cancers was much lower and no death was reported.Different behavioral patterns observed in patients with various types of cancers in the face of COVID-19 disease are very important and the researchers of the current study noticed this difference during the last 3 months.
In ammatory cells play important roles in cancer (Fulop et al., 2020;Grizzi et al., 2020;Tseng et al., 2020).Cancer expansion and development are multifaceted processes that are also regulated by the surrounding microenvironment generated by the innate and adaptive immune cells such as dendritic cells, T cells, and macrophages (Katz and Rabinovich, 2020;Rezaei et al., 2020).Macrophages are present in all tissues and play imperative roles in the preservation of homeostasis in tissues (Nouwen and Everts, 2020;Wu and Lu, 2020).Macrophages are regularly differentiated from peripheral blood monocytes and categorized into three types, namely, tissue-resident macrophages (TRM), myeloid-derived suppressor cells (MDSC), and tumor-associated macrophages (TAMs) (Beatson et al., 2020;Etzerodt et al., 2020;Gross-Vered et al., 2020;Heideveld et al., 2020;Liu et al., 2020b).
In terms of their function Macrophages are divided into two types; classically activated M1 or good macrophages and alternatively activated M2 subtypes or bad macrophages (Klichinsky et al., 2020).M1 macrophages are presented in T-helper type 1 (Th1) responses that are promoted by Th1 cytokines such as interleukin-12 (IL-12) and IL-18.This type of macrophages generate reactive oxygen/nitrogen species and pro-in ammatory cytokines such as IL-1β, IL-6, tumor necrosis factor α (TNF-α), and are important in innate host defense and killing tumor cells (Zhang et al., 2014;Genin et al., 2015;Chunying Xie, 2020).
Cytokine release syndrome (CRS) is a complete response to the in ammation caused by usual infections.Many drugs and other factors have been designed to induce the production of a large amount of proin ammatory cytokines and increase the strength of the immune system (Hirano and Murakami, 2020).CRS is commonly happening during immune system-related diseases or immune-related therapies and also in infectious diseases such as viral infections (Hirano and Murakami, 2020;Liu et al., 2020a).IFN-γ is one of the most important functional cytokines which plays an important role in defense against intracellular pathogens and also cancers (Liu et al., 2020a;Mehta et al., 2020;Zheng et al., 2020a).The results of various studies have shown that the levels of pre-in ammatory cytokines and IFN-γ in patients with solid cancers are naturally elevated (Gaire et al., 2020;Zhang et al., 2020).The study conducted by Qing et al (2020) reported that the levels of IL-1B, IFN-γ, IP-10, and monocyte chemoattractant protein 1 (MCP-1) in patients with COVID-19 was signi cantly high.These cytokines are Th1 activators and are key factors for the activation of speci c immunity and unfortunately are positively correlated with the severity of COVID-19 (Ye et al., 2020).
CRS and cytokine storms have shown a direct association with COVID-19 in acute in ammations, but the behavior of the disease in chronic in ammations is completely different.In chronic in ammations, the interaction between macrophages and solid tumors leads to the production of IFN-γ by M1 TAMs.The high levels of IFN-γ in chronic in ammations result in the resistance of the patients with solid tumors to COVID-19 in comparison to the patients with hematologic malignancies.In a comparable study, many studies revealed that vaccinations with BCG vaccine cause chronic and non-speci c immunity that surprisingly decreases the incidence and mortality of COVID-19 (Bodova et al., 2020;O'Neill and Netea, 2020).
One of the important aspects of this study, which is different from other investigations, is the survey of family and people who are in contact with the patient.Our study and other similar studies have shown that many people with no clinical symptoms were molecularly positive.These people are carriers and can transmit the disease.
The importance of this group (asymptomatic carriers) will be much greater if they are in contact with cancer patients.Chemotherapy weakens the immune system and increases the risk of developing the disease.Researchers are concerned about the presence of asymptomatic carriers in contact with candidates for chemotherapy.Therefore, it is recommended that either all patients await chemotherapy and families and individuals in contact with them undergo a molecular analysis prior to chemotherapy, or chemotherapy is performed under hygienic and supervised measures.
In this study, the incidence rate of COVID-19 in patients with cancer was investigated.Two groups of patients were studied, including patients with solid cancer and patients with hematological malignancies.
The results showed that COVID-19 was much less common in patients with solid cancers in comparison to patients with hematological malignancies.Even in cases that family members of patients were infected with COVID-19, patients with solid tumors were not infected.Researchers of this study attributed these results to the chronic in ammation and M1 macrophages in these patients.Due to the importance of COVID-19 and cancer in the world and based on the results of this study, it is essential that additional studies are conducted, especially in terms of biological function.Finally, researchers of this study invite other researchers from all research centers around the world to cooperate and participate in further supplementary studies.

Figures
Figure 1 Results of the con rmatory tests indicated that 9 (2.48%) patients with different types of solid tumors were positive for COVID-19, whereas, 46 (12.70%) patients had close contact with COVID-19 patients in their family and were closely exposed

Table 1
Demographic and baseline clinical characteristics of COVID-19 patients in solid tumors

Table 3
The clinical manifestations of patients with solid and hematologic malignancies and different regimes of chemotherapy.